Clinical Immunology_ Principles and Practice ( PDFDrive )

1116 Part NINE Transplantation



TABLE 82.1 Sources of Hematopoietic GvHD. Several methods are available to attain T-cell
Stem Cells for transplantation depletion.
In the past, the method of soybean lectin agglutination and
Hematopoietic Stem Cell transplantation (HSCt) From a E-rosetting was frequently used. With this method, soybean lectin
related Donor allowed agglutination of the majority of mature marrow cells,
Bone marrow from a human leukocyte antigen (HLA)–genotypically which were removed by sedimentation. Further depletion of T
identical sibling
Bone marrow from an HLA-phenotypically identical family member lymphocytes was achieved by rosetting with sheep erythrocytes
Bone marrow from a haploidentical parent (E-rosetting technique) and density gradient centrifugation.
T cell depleted by negative selection with soybean lectin Importantly, T-cell depletion by soybean lectin agglutination
agglutination and rosetting with sheep red blood cells and E-rosetting maintains all immature marrow cells in the final
(E-rosetting) preparation.
T cell depleted by negative selection with monoclonal antibodies T-cell depletion can also be achieved by incubation of bone
Positive selection of CD34 cells
+
marrow with monoclonal antibodies (mAbs) to T lymphocytes
HSCt From a Matched Unrelated Donor plus complement. Campath-1 G, Leu 1, and other mAbs have
Unmanipulated bone marrow been used for this purpose, but the degree of T-cell depletion
+
T cell–depleted bone marrow by means of positive selection of CD34 that is achieved with these agents is less effective than with the
cells soybean lectin and E-rosetting, and therefore a higher incidence
+
Positively selected peripheral blood CD34 cells of GvHD has been reported. More recently, use of mAbs directed
against the αβ form of the T-cell receptor (TCR) and against
HSCt From an Unmanipulated related or Unrelated CD19 has entered clinical practice, with excellent results. 4
Cord Blood
However, the most common method to obtain transplantable
+
HSCs is represented by positive selection of CD34 cells using
mAbs. This method allows very robust depletion of mature T
−
KEY CONCEPtS cells; however, it also removes immature CD34 cells and other
Sources of Stem Cells and Selection of Donors cells (especially stromal marrow cells) that can facilitate stem
cell engraftment.
for Hematopoietic Stem Cell Transplantation in Selection of the best donor is another important aspect of T
Primary Immunodeficiencies cell–depleted haploidentical HSCT for SCID. In general, the
donor is represented by one of the parents, since the volume of
Sources of hematopoietic stem cells (HSCs) for transplantation include
bone marrow, peripheral blood, and cord blood. bone marrow that can be collected is much higher than it would
If the donor is a genotypically human leukocyte antigen (HLA)–identical be if a haploidentical sibling were to serve as donor. Maternal
sibling, unmanipulated bone marrow is used as source of stem cells. T-cell engraftment in utero is a common finding in infants with
Whenever the donor is HLA-mismatched to the recipient, T-cell depletion SCID and has been observed in almost 40% of patients with
must be performed to eliminate mature T lymphocytes from the graft. SCID. In such cases, T cell–depleted HSCT should be performed
Methods for T-cell depletion of the bone marrow include use of soybean using the mother as donor, if possible, since transplantation
lectin agglutination and E-rosetting, depletion with monoclonal antibodies from the father might cause a graft-versus-graft reaction.
(mAbs), and positive selection of stem cells.
Cord blood is a rich source of stem cells. However, the volume of cord In Utero Haploidentical HSCT
blood is limited, so its use is mainly restricted to young patients.
The number of volunteers included in Bone Marrow Donor Registries is The identification of a growing number of immunodeficiency-
expanding. Consequently, there is a continuous increase in the number causing genes has resulted in continuous improvement in prenatal
of matched unrelated donor (MUD) transplantations performed for diagnosis, which, in most cases of severe immunodeficiency, can
patients with primary immunodeficiencies. now be accomplished on chorionic villi DNA at 10–11 weeks
Whenever cord blood or MUD stem cells are used, conditioning regimen
must be given to the recipient before transplantation to facilitate of gestation. This has prompted prenatal transplantation of
+
engraftment of donor stem cells. parental positively selected CD34 stem cells into the peritoneum
Therapeutic options for patients with severe combined immunodeficiency of fetuses prenatally diagnosed with SCID, under ultrasound
(SCID) and other severe forms of primary immunodeficiency (PID) guidance.
include transplantation from HLA-genotypically identical donor, mis- The rationale underlying in utero HSCT is based on a
matched related donors (MMRD), and MUDs.
lower risk of graft rejection resulting from decreased fetal
immunocompetence (although this consideration is not relevant
in the case of fetuses with SCID), a presumed induction of
HSCT From a Haploidentical Donor tolerance to paternal antigens (which might favor successful
Unfortunately, the option of related HLA-identical HSCT is engraftment after postnatal transplantation from the same donor),
limited only to a minority of patients. When no such donor is the predicted competition between donor and autologous stem
available, stem cell transplantation from a haploidentical parent cells at a time when several empty niches should be available
should be considered, particularly in infants with SCID. for stem cell engraftment, the potential ability to provide preemp-
The rationale for haploidentical HSCT is based on the ability tive treatment (thus reducing the risk of postnatal infection),
of donor-derived stem cells to repopulate the recipient’s vestigial and the lower cost of the procedure that does not require pro-
thymus and give rise to fully mature T lymphocytes. Indeed, longed hospitalization. However, in utero HSCT is associated
this is a life-saving procedure that has been successfully applied with the potential risks of fetal loss and of GvHD. Finally, if
to several hundreds of infants with SCID. 1,2 maternal T cells had engrafted into the fetus with SCID, trans-
+
However, this procedure requires careful removal of T lym- plantation of paternal CD34 cells might cause graft-versus-graft
phocytes from the graft, as these would otherwise cause severe reaction.

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1117


In a series of five in utero HSCT performed at the University recipient’s cells. Furthermore, conditioning regimens can cause
+
of Brescia, Italy, all three cases with B SCID survived, with toxicity of several organs. Myeloablative regimens cause anemia,
evidence of T-cell reconstitution. However, transient or no thrombocytopenia, and leukopenia. Consequently, supportive
−
immune reconstitution was observed in two fetuses with B SCID. treatment with RBC and platelet transfusions is necessary during
The use of in utero HSCT for SCID is now largely unjustified, the aplastic phase. Finally, the leukopenia predisposes the patient
also in consideration of the fact that newborn screening allows to an increased risk of life-threatening bacterial or fungal
rapid identification of infants with SCID and permits rapid infections.
referral to HSCT, with long-term survival that exceeds 90%. 5 The frequency and severity of these complications depend
on the type of transplant, the possible use of a conditioning
HSCT From Matched Unrelated Donors regimen, and specific considerations related to the underlying
Since the first successful HSCT performed in 1977 in an infant disorder and to the clinical status of the recipient before
with SCID, HSCT from MUD has been increasingly used to transplantation.
treat severe PID. MUD HSCT has been shown to be more effective
than T cell–depleted HSCT in patients with immunodeficiencies KEY CONCEPtS
1
other than SCID, and it has been successfully used also in infants Rejection of Donor Stem Cells by the Host
with SCID. 6 Immune System and Reaction of Donor’s T
Transplantation from MUD has been facilitated by the increas-
ing number of volunteer donors included in registries worldwide. Lymphocytes to the Host
In addition, advances in the quality of the techniques used for The host T cells are responsible for the elimination of donor stem cells,
HLA typing permits this identification of an optimal MUD and regardless of the degree of human leukocyte antigen (HLA) compatibility.
reduction in the risk of GvHD. As of April 2016, more than 28 Under these circumstances, other than in patients with severe combined
million donor volunteers and UCB units were included in the immunodeficiency (SCID), a chemotherapy-based conditioning regimen
Bone Marrow Donors Worldwide (BMDW) registry. At present, must be used before transplantation to allow for donor stem cell
engraftment.
it takes only a few weeks to identify a MUD. However, the prob- If T cells are present in the graft, and especially if there is HLA-
ability of finding a suitable donor is lower for selected ethnic or incompatibility between the donor and the recipient, the donor’s T
racial groups that are poorly represented among volunteer donors. lymphocytes may react to host alloantigens and cause graft-versus-host
Importantly, MUD HSCT requires use of a preparative disease (GvHD).
chemotherapy regimen in the recipient (even in the case of SCID) Risk factors for GvHD include HLA-mismatch between donor and recipient,
and GvHD prophylaxis (because of likely disparity between donor older age at transplantation, gender mismatch, and previous viral
and recipient at minor histocompatibility loci), whereas neither infections.
one is necessary for related HLA-identical HSCT in SCID infants. GvHD may develop early after transplantation (acute GvHD), or at 100
days or more after transplant (chronic GvHD). GvHD is one of the
major causes of death and long-term disability after stem cell
HSCT Using Unmanipulated Cord Blood transplantation.
As opposed to MUD HSCT, which requires identification, Prevention is the best approach to the management of GvHD. Prevention
willingness, and medical clearance of an adult volunteer, stored of GvHD is based on selection of optimal donor and vigorous T-cell
cord blood is readily available as a source of stem cells for depletion in the case of HLA mismatch between the donor and the
transplantation. In addition, the risk of GvHD at any given degree recipient. Immunosuppressive drugs, such as cyclosporine, are another
potent form of GvHD prevention.
of HLA matching is lower when using cord blood compared
with MUD HSCT so that greater HLA disparity with the recipient
can be tolerated. However, the number of cells contained in any
defined unit is still a major limitation of cord blood. Low Graft Rejection
cell dose is not usually a problem for transplants performed in Graft rejection reflects the presence of immunocompetent cells
infants with SCID or other severe forms of immune deficiency in the host that specifically recognize and react to donor-derived
because of the low weight of the recipient. Indeed, unrelated stem cells. Several factors influence the likelihood of graft rejec-
umbilical cord stem cell transplantation has been successfully tion, in particular (i) the degree of immunocompetence of the
7
used in dozens of patients with severe PID. In practice, an host; (ii) the degree of HLA disparity between donor and recipient;
unrelated HSC donor should be simultaneously searched for in (iii) the number and source of stem cells infused; (iv) the type
cord blood banks and in bone marrow donor registries for patients of conditioning regimen used; and (v) the possible presensitization
lacking an HLA-identical sibling HSC donor. The option of of the host to donor histocompatibility antigens.
performing cord blood transplants should be based on urgency In the case of infants with SCID, graft rejection is an unlikely
of the transplantation, the cell dose required, and the number event because of the virtual lack of T cells that characterizes
of HLA disparities. these conditions. However, natural killer (NK) cells are present
As for HSCT from MUD, transplantation using cord blood in several forms of SCID and may contribute to graft rejection.
usually requires pretransplantation conditioning and GvHD Indeed, among infants with SCID who received HSCT without
prophylaxis, irrespective of the underlying disease. conditioning chemotherapy, reduced overall survival and increased
requirement of additional procedures have been observed in
Complications of Hematopoietic Stem those with NK SCID compared with NK SCID (Fig. 82.1). In
−
+
8
Cell Transplantation other forms of PID, there is sufficient immune function in the
A variety of complications can compromise the success of HSCT. host to allow for rejection of donor-derived stem cells, unless
Among these, incompatibility between donor and recipient can an appropriate conditioning regimen is used before HSCT.
lead to graft rejection by the host immune system or to GvHD In children with nonmalignant disease, the most commonly
caused by alloreactivity of donor-derived lymphocytes to the used myeloablative conditioning regimen consists of busulfan

1118 Part NINE Transplantation


100 TABLE 82.2 Staging and Grading of acute
NK-ve 87%
80 Graft-Versus-Host Disease (aGvHD)
NK+ve 62% Stage Skin Liver Gastrointestinal system
(Bilirubin) (GI) (Stool output)
% surviving 0 None <2 mg/dL Adult: <500 mL/day
60
40
1 Rash <25% BSA 2–3 mg/dL Child: <10 mL/kg/day
Adult: 500–999 mL/day
20 Child: 10–19.9 mL/kg/day or
P<0.01 persistent nausea,
vomiting, or anorexia, with
0 a positive upper GI biopsy
0 2000 4000 6000 2 Rash 25–50% 3–6 mg/dL Adult: 1000–1500 mL/day
Days BSA Child: 20–30 mL/kg/day
FIG 82.1 Kaplan-Meyer survival curve after nonconditioned 3 Rash >50% BSA 6.1–15 mg/ Adult: >1500 mL/day
transplantation for infants with severe combined immunodefi- (generalized dL Child: >30 mL/kg/day
erythroderma)
+
ciency (SCID) with normal natural killer (NK) cell count (NK ) or 4 Generalized >15 mg/dL Severe abdominal pain with
−
with lack or markedly reduced number of NK cells (NK ). (With erythroderma or without ileus
permission from Hassan A, Lee P, Maggina P, et al. Host natural with bullous
killer immunity is a key indicator of permissiveness for donor formation
cell engraftment in patients with severe combined immunode- Grade
ficiency. J Allergy Clin Immunol 2014;133:1660–6.) Grade 0 No stage 1–4 of any organ
Grade 1: Stage 1 or 2 skin involvement; no liver or gut involvement
Grade 2: Stage 1–3 skin involvement; Grade 1 liver or gut
and fludarabine, with or without the addition of antithymocyte involvement
globulin (ATG) or anti-CD52 (alemtuzumab) mAb. However, Grade 3: Stage 2 or 3 skin involvement
infants and children with preexisting organ damage are highly Grade 4: Stage 1–4 skin involvement; stage 2–4 liver or gut
sensitive to the toxic effects of drugs. In these cases, reduced involvement
intensity regimens are preferred. To this purpose, lower doses
of busulfan are used. Treosulfan is an alternative to busulfan
9
and has reduced toxicity. Finally, the combination of fludarabine
and melphalan is another common form of reduced-intensity of transplantation. As the types of donor, stem cell sources, and
conditioning. conditioning regimens have changed, it is now recognized that
symptoms of aGvHD can present later than 100 days and that
Acute Graft-Versus-Host Disease cGvHD is better defined by its distinct clinical manifestations
Acute GvHD (aGvHD) is the result of alloreactivity of donor- rather than the time of onset alone. These clinical manifestations
derived T lymphocytes versus the recipient’s antigens and is one include skin changes (scleroderma-like lesions, hyperpigmenta-
of the most severe complications of HSCT. It may occur as early tion, hyperkeratosis, skin atrophy, ulcerations), tissue fibrosis
as 1 week after HSCT and is potentially fatal. Clinical manifesta- and limitation of joint motility, fibrosis of exocrine glands (“sicca
tions of aGvHD include maculopapular skin rash (that tends to syndrome”), fibrosis of lungs and liver, increased susceptibility
10
be confluent), diarrhea, and liver abnormalities (hepatomegaly, to infections, immune dysregulation, and autoimmunity.
10
elevated liver enzymes, increased levels of conjugated bilirubin). Consequently, cGvHD poses a major burden on the patients’
The disease may progress to severe skin manifestations, with quality of life and can be fatal.
exfoliative dermatitis, and significant liver and gut damage (with Although the incidence of cGvHD is lower in children than
intractable watery or bloody diarrhea, protein-losing enteropathy, in adults treated by allogeneic HSCT, the risk factors and the
and abdominal pain). In the most severe cases, leakage of spectrum of clinical manifestations are similar.
intravascular fluids into the interstitium (so-called third space Acute GvHD represents a major risk factor for cGvHD, and
filling) leads to generalized edema. Bone marrow aplasia and a yet cGvHD can be observed even without preceding aGvHD,
high susceptibility to infections (including reactivation of her- and when present, it does not represent merely the continuation
pesvirus infections) are also often observed in severe aGvHD. of aGvHD. Older age of the recipient at HSCT, transplantation
The severity of aGvHD is evaluated according to grading from a multiparous female donor into a male recipient (with
(Table 82.2). Major risk factors for aGvHD include HLA-mismatch reactivity to Y chromosome-associated antigens), and incompat-
between donor and recipient, older age of the recipient or donor, ibility at minor histocompatibility loci represent additional risk
10
10
gender mismatch, and stem cell source. However, aGvHD may factors for cGvHD. Furthermore, use of peripheral blood stem
also be observed following related HLA-identical HSCT, par- cells carries an increased risk of cGvHD compared with use of
ticularly when a conditioning regimen is used. bone marrow stem cells.
Finally, transfusion-associated aGvHD is a very severe com-
plication after HSCT, which can be effectively prevented by using Prevention of GvHD
irradiated (1500–3000 rad) and filtered blood derivatives. Prevention is the most effective approach to GvHD, and use of
a fully matched donor remains the best method of prevention.
Chronic Graft-Versus-Host Disease Alternatively, if a related HLA-mismatched donor is used for
Chronic GvHD (cGvHD) has traditionally been defined as transplantation, it is essential that the graft is vigorously T-cell
symptoms that persist or appear after 100 days since the time depleted.

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1119


3
Whenever a conditioning regimen is used in the transplantation HLA-mismatched transplantation for SCID. In spite of this,
protocol, pharmacological prophylaxis of GvHD must also be infections remain the major cause of death. Challenging
included, even in the case of HSCT from a related HLA-identical viruses in infants with SCID include adenovirus, cytomegalovirus
donor. (CMV), parainfluenza type III virus, and Epstein-Barr virus
Long-standing approaches to prevention of GvHD include (EBV). In particular, CMV infection after HSCT can cause
2
cyclosporine daily for 6 months or methotrexate (15 mg/m on interstitial pneumonia, enteritis, hepatitis, and encephalitis.
2
the first day after HSCT, and then 10 mg/m at days +3, +6, and EBV can also cause lymphoproliferative disease. Filtering of
+11 after transplantation), or a combination of the two. Newer blood derivatives removes leukocytes and thus reduces the
regimens substitute another calcineurin inhibitor, tacrolimus, risk of transfusion-associated infections. Several antiviral
for cyclosporine. Serotherapy with ATG or alemtuzumab is often drugs (acyclovir, ganciclovir, foscarnet, cidofovir) are now available
13
included in the preparatory regimen, with the aim of reducing and have good results, especially against CMV, and preemptive
the risk of GvHD. The timing of administration of alemtuzumab administration of anti-CD20 mAb is efficacious in the preven-
as part of the preparatory regimen has opposed effects on the tion of EBV-driven lymphoproliferative disease. Furthermore,
prevention of aGvHD and the speed of T-cell reconstitution. In transfusion of virus-specific cytotoxic T lymphocytes (CTLs)
particular, use of alemtuzumab in the days that immediately represents another important resource to fight severe viral
precede transplantation has more potent effects on aGvHD infections. 14
prevention but is associated with delayed T-cell reconstitution, Pneumocystis jiroveci is a common cause of pneumonia in
whereas the opposite is observed when the drug is administered severely immunocompromised patients. Treatment is based on
a few weeks before HSCT. Finally, mycophenolate mofetil and intravenous cotrimoxazole (20 mg/kg/day).
methotrexate can also be used to prevent GvHD. Aspergillus infection is a severe complication in patients with
Attempts to prevent cGvHD have been less satisfactory. In chronic granulomatous disease (CGD) and in patients with
particular, use of a prolonged immune suppression after transplant profound neutropenia. Voriconazole offers some advantage
does not decrease the incidence of cGvHD. compared with liposomal amphotericin B for treatment of invasive
aspergillosis, whereas prophylactic itraconazole reduces the
Treatment of GvHD incidence of fungal infections in patients with CGD before
Once GvHD has developed, treatment is mainly based upon the transplantation.
use of immunosuppressive drugs. Corticosteroids remain the Bacterial infections are usually amenable to successful treat-
first-line therapy and are usually effective, especially for mild ment, if the pathogen is identified, and appropriate and aggressive
and moderate forms of aGvHD. Second-line therapy includes use of antibiotics is initiated. Prophylactic administration of
ATG, mycophenolate mofetil, cyclosporine, or tacrolimus. mAbs, immunoglobulins (Igs) following HSCT also reduces the frequency
such as anti-CD25 (daclizumab), are also used. However, the and severity of infections. Administration of Bacille Calmette-
efficacy of second-line agents in severe or steroid unresponsive Guérin (BCG) vaccine at birth to prevent tuberculosis is still
aGvHD is limited. practiced in many countries worldwide. In severely immuno-
Treatment of cGvHD is also based on immunosuppression, compromised infants who undergo HSCT, this live vaccine may
but with limited efficacy. 11,12 Topical steroids and calcineurin cause disease, which often manifests at the time of engraftment
inhibitors may alleviate mucosal and skin symptoms. Systemic with local and systemic immune reconstitution inflammatory
steroids have been shown to improve survival, but at the risk of syndrome (IRIS).
significant adverse effects. Ursodeoxycholic acid may be useful
in cGvHD with significant liver involvement. Extracorporeal Toxicity Related to Conditioning Regimen
photopheresis can be used with the goal to induce tolerance; Chemotherapeutic agents that are used in the conditioning
typically, its benefits, if present, are delayed until 2–3 months regimen of HSCT often cause significant short-term and long-
after initiation of treatment. Use of hydroxychloroquine, myco- term toxicity. Chemotherapeutic drugs that damage the liver
phenolate mofetil, anti-TNF-α mAb, etanercept (a recombinant vascular endothelium, particularly busulfan and cyclophospha-
form of soluble tumor necrosis factor [TNF]-receptor), and mide, can cause veno-occlusive disease (VOD), which is clinically
anti-CD20 antibody (rituximab) remains investigational. marked by painful hepatomegaly, jaundice, ascites, fluid retention,
and weight gain and can ultimately result in fatal multiorgan
Infections failure (MOF). Defibrotide is the most effective agent studied
Infections represent one of the major complications following to date in the treatment of VOD. Busulfan can also cause seizures
HSCT. Patients with severe PID are intrinsically highly susceptible and lung damage. Cyclophosphamide can cause hemorrhagic
to infections. In infants with SCID and with other forms of cystitis, a syndrome of inappropriate antidiuretic hormone
combined immune deficiency, viral and opportunistic infections secretion, or more rarely, cardiac disturbances.
can develop before transplantation and are one of the factors Long-term hormonal complications are more common
2
that adversely affect the outcome of HSCT itself. Regardless of when total body irradiation (TBI) is used. However, the busulfan
the type of underlying PID, T cell–depleted HSCT carries a high and cyclophosphamide regimen can cause delayed puberty or
risk of infections because of the longer time required to achieve sterility, and thyroid dysfunction is frequently observed, even in
immune reconstitution. Furthermore, use of a pretransplant patients who have not received TBI. Delayed or incomplete tooth
conditioning regimen with myeloablative and immunosuppressive eruption is also a possible consequence of conditioning regimens.
drugs, and GvHD prophylaxis, contribute to the increased Effects on final height and growth, as well as long-term neuro-
susceptibility to infections after HSCT. cognitive effects, are emerging as more children are treated
Strict isolation of the patients during and after HSCT, and followed up. Patients with defects of DNA repair (e.g.,
and prophylactic administration of antibiotics, have been some forms of SCID) are particularly at risk for drug-related
associated with a better survival rate, particularly after related toxicities.

1120 Part NINE Transplantation



CLINICaL PEarLS 1.0 YEAR OF TRANSPLANTATION
Considerations of Stem Cell Transplantation
Unique to Severe Combined 0.8
Immunodeficiency (SCID)

Patients with SCID are strongly impaired in their ability to reject allogeneic 0.6 2000–2005 1995–1999
cells, including stem cells. Therefore no chemotherapy is required in
these patients in order to achieve T-cell reconstitution following stem Proportion surviving 1968–1994
cell transplantation.
The quality and the kinetics of T-cell reconstitution following stem cell 0.4
transplantation depend on the type of transplant.
If unmanipulated bone marrow from genotypically human leukocyte
antigen (HLA)-identical related donor is used, mature T cells contained 0.2 P =.0003
in the graft expand as early as 2 weeks after transplantation and
provide a rapid source of immune competence.
A similar phenomenon is also observed following unmanipulated matched 0
unrelated donor (MUD) transplantation. In this case, however, drugs
used in conditioning regimen and for prevention of graft-versus-host 0 12 24 36 60 120
disease (GvHD) decrease the degree of early expansion of donor-derived Time after transplantation (months)
T cells. Months 0 6 12 24 36 60 120
In contrast, appearance of naïve T cells occurs only at 3 months or more
after transplantation, regardless of degree of HLA matching between Number at risk
donor and recipient. Consequently, following haploidentical transplanta- 1968–1994 361 245 187 172 166 151 88
tion, there is a prolonged period during which the recipient remains 1995–1999 157 113 95 78 66 40 3
lymphopenic and at high risk of infections. 2000–2005 181 111 74 49 26 4 0
In the absence of pretransplant conditioning, following haploidentical
transplantation for SCID, engraftment is usually restricted to T lym- FIG 82.2 Cumulative probability of survival in patients with severe
phocytes. This may cause persistent B-cell dysfunction. Furthermore, combined immunodeficiency (SCID) after hematopoietic stem
+
patients with natural killer (NK) SCID may show some ability to reject cell transplantation (HSCT) according to the period in which
stem cells from HLA-mismatched donors. For these reasons, several transplanted. (With permission from Gennery AR, Slatter MA,
centers include use of conditioning regimen for the hematopoietic Grandin L, et al. Transplantation of hematopoietic stem cells
stem cell transplantation (HSCT) in babies with SCID who do not have and long-term survival for primary immunodeficiencies in Europe:
a matched related donor available.
entering a new century, do we do better? J Allergy Clin Immunol
2010;126:602-10.e1–11.)

HSCT for SCID the use of two parents as sequential donors was associated with
General Considerations immune reconstitution.
SCID is a medical emergency and is uniformly fatal unless In contrast, chemotherapy is typically used for HSCT from
promptly diagnosed and successfully treated. With a few excep- unrelated donors and—irrespective of donor type—for “leaky”
tions in which alternative strategies (gene therapy, enzyme forms of SCID with presence a significant number of autologous,
replacement therapy) can be used, allogeneic HSCT represents partially functioning T lymphocytes.
the most effective form of treatment for these disorders.
SCID is also a unique situation in which the virtual lack of Survival Following HSCT for SCID
T lymphocytes strongly impairs the ability of the recipient to The two largest series of patients with SCID treated by HSCT
reject the graft. Furthermore, donor-derived lymphoid progenitor in Europe and in North America included 699 and 240 cases,
cells have a striking advantage for in vivo T-cell differentiation. respectively. 1,2
Consequently, use of pretransplantation chemotherapy and In the European study, which included transplantations
immune suppression is not required to attain T-cell reconstitution. performed between 1968 and 2005, 203 patients received HSCT
However, although there is a general consensus that no condition- from genotypically (n = 135) or phenotypically (n = 68) identical
ing is needed for HSCT from an HLA-identical related donor, related donors, with 10-year overall survival rates of 84% and
there is controversy as to whether chemotherapy should be used 64%, respectively. This figure is clearly superior to the 54% survival
for T cell–depleted mismatched HSCT. Although use of condition- rate observed among 415 patients who had received related
ing chemotherapy carries the risk of drug-related toxicity, it HLA-mismatched HSCT. Finally, in this study, 81 patients received
1
favors the engraftment of donor-derived stem cells and may lead HSCT from MUDs, and 66% survived. In the North American
to a better recovery of humoral immunity and of improved study, which included transplantations performed between 2000
2
thymopoiesis. In approximately 30% of the cases, unconditioned, and 2009, the overall 5-year survival rate was 74%. In particular,
T cell–depleted MMRD transplantation may fail to induce T-cell the survival rate was highest among recipients of HSCT from
reconstitution in infants with SCID. In this case, booster trans- HLA-matched sibling donors (MSDs) (97%), followed by
plants without chemotherapy may be helpful. A recent report unconditioned HSCT from MMRDs (79%), HSCT from URDs
15
analyzed the outcome of such a procedure. Among 49 patients (74%), HSCT from MMRDs with conditioning (66%), and HSCT
with SCID who received such booster transplants at Duke from UCB (58%).
University Medical Center, 31 (63%) survived for up to 28 years. Survival after HSCT for SCID has improved over the years
1
Older age at HSCT and persistence of preexisting infections (Fig. 82.2), reflecting more effective treatment and prevention
represented significant risk factors for death. In several patients, of disease-related and transplantation-associated complications,

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1121


such as infections and GvHD. In particular, according to the Factors that may contribute to such poor outcome may include
1
European study, the current survival rate after HSCT from the presence of autologous NK cells that may mediate graft
HLA-identical related donors is 90%, whereas 3-year survival rejection. Consistent with this, in a UK series of 77 infants with
after HSCT from related HLA-mismatched donors has improved SCID who received HSCT, survival rate was higher among patients
+
−
from 49% in transplantations performed up to 1994 to 69% in with NK SCID than among recipients with NK SCID (see Fig.
17
those performed between 1995 and 1999. However, no further 82.1). Furthermore, RAG and Artemis deficiencies cause a later
improvement in survival has been observed since that time. block in T-cell development compared with the majority of
+
Several factors influence survival after HSCT for SCID. In genetic defects associated with B SCID. This may lead to a
particular, younger age at transplantation leads to superior stronger competition between donor-derived and autologous
−
survival. In the North American study, among 68 infants who T-cell progenitors in patients with B SCID who receive uncon-
2
were treated before 3.5 months of life, 64 (94%) survived. ditioned HSCT.
Similarly high survival rates were observed also among infants Finally, in a series of 106 patients with SCID caused by adenos-
−
older than 3.5 months with no history of infections or whose ine deaminase deficiency (ADA SCID), unconditioned trans-
infections had resolved before HSCT (90% and 84% 5-year plantation from MMRDs was associated with a high risk of graft
2
survival rates, respectively). Collectively, these data emphasize failure. 18
the importance of early diagnosis to prevent infections and
provide strong support to newborn screening for SCID. For CLINICaL PEarLS
patients of any age who do not have MSDs and have an active Outcome of Hematopoietic Stem Cell
infection at the time of HSCT, a higher survival rate (65%) is
observed with unconditioned transplantation from MMRDs. 2 Transplantation (HSCT) for Primary
The SCID genotype and phenotype may also affect the outcome Immunodeficiencies
+
of HSCT. In the European series, infants with B SCID had For severe combined immunodeficiency (SCID), the 5-year survival rate
superior 10-year survival after HSCT compared with infants of patients transplanted between 2000 and 2009 is over 74% and is
−
1
with B SCID (70% vs 51%, respectively; Fig. 82.3), although higher (97%) for patients who receive transplant from matched sibling
no significant difference in survival between these two groups donors.
2
of infants with SCID was observed in the North American study. Improvements in clinical care (both in critical care and in prevention and
Furthermore, haploidentical transplantation without conditioning treatment of infections) have also contributed to the improved outcome
of HSCT for SCID, including transplantation from matched unrelated
or with immunosuppression only in patients with recombinase donors (MUDs). However, the decision to attempt a MUD transplanta-
activating gene (RAG) or Artemis deficiency is associated with tion for SCID must be weighed against the risks associated with the
16
a high risk of graft failure or of incomplete T-cell reconstitution. time interval required to identify such a donor.
Factors influencing survival after transplantation for SCID include younger
age at transplant and lack of active infections at the time of
1.0 transplantation.
The decline of T-cell function that is observed at 10 years or more after
transplantation remains a concern and may cause clinical problems.
0.8 Therefore there is a need for improvements in the procedures used
to facilitate and sustain stem cell engraftment and/or to boost donor-
Other For primary immunodeficiencies other than SCID, there has been a
derived immunity.
Proportion surviving 0.6 B– progressive improvement in the outcome following stem cell trans-
B+
plantation. In particular, results are good both for human leukocyte
antigen–identical transplants (with a survival rate of 70% or more,
0.4
depending on the disease) and for MUD or cord blood transplantations.
Reduced-intensity conditioning regimens have been often used in
these patients, in the attempt to reduce the risks of drug-related
0.2 P <.0001
toxicity.
0
Complications Following HSCT for SCID
0 12 24 36 60 120 Despite advances in prophylaxis and treatment, infections
Time after transplantation (months) (especially those caused by viruses) remain a significant cause
of death after HSCT for SCID. In a report of 166 transplantations
Months 0 6 12 24 36 60 120 performed at their center, Buckley et al. indicated that viral
18a
Number at risk infections accounted for 30 of the 40 deaths observed. CMV
B+ 345 247 195 168 147 111 55 and adenovirus were responsible for nine deaths each, and 18
B– 300 184 135 109 91 69 33 additional deaths resulted from infections caused by EBV,
Other 54 38 26 22 20 15 3 enteroviruses, parainfluenza virus type 3, varicella virus, herpes
FIG 82.3 Cumulative probability of survival in patients with T B or simplex virus, and respiratory syncytial virus. In a series of 240
−
−
T B severe combined immunodeficiency (SCID) after hemato- patients with SCID who received HSCT in North America between
−
+
poietic stem cell transplantation (HSCT). (With permission from 2000 and 2009, infections accounted for 24 of 62 deaths. 2
Gennery AR, Slatter MA, Grandin L, et al. Transplantation of Viral and opportunistic infections are more common early
hematopoietic stem cells and long-term survival for primary after HSCT, especially in recipients of T cell–depleted haploidenti-
immunodeficiencies in Europe: entering a new century, do we cal HSCT, because of the delay in achieving immune reconstitu-
do better? J Allergy Clin Immunol 2010;126:602-10.e1–11.) tion. Incomplete recovery of immune function at 1 year after

1122 Part NINE Transplantation


HSCT is associated with a higher risk of late infections. In a Finally, prolonged nutritional support has been reported after
single-center study of 90 patients with SCID treated with HSCT, HSCT for SCID. This complication is more frequent among
11 (12%) developed significant infectious complications 2–17 patients treated by mismatched related or unrelated donor HSCT,
19
years after transplantation. Among late infections, chronic skin especially if cGvHD, immune dysregulation, and poor immune
warts caused by papilloma virus have been observed in a sig- reconstitution are also present. Infants with Artemis deficiency
19
nificant fraction of infants with γc or JAK3 deficiency after HSCT. (leading to impaired DNA repair) are at particularly high risk
This complication may occur also in patients who attain robust for late complications, including growth retardation, requirement
immune function and probably results from signaling defects for nutritional support, and dental abnormalities. 16,19
that involve extrahematopoietic cells, such as keratinocytes.
GvHD is another major complication of HSCT for SCID. In Quality and Kinetics of T-Cell Immune Reconstitution
a series of 240 patients with SCID who received HSCT in North The effectiveness of HSCT in SCID is well illustrated by the
America between 2000 and 2009, the cumulative incidence of normalization of the number and function of T lymphocytes that
aGvHD of grade 2–4 at 100 days after transplant was 20%, with is achieved after transplantation (Figs. 82.4 and 82.5). The efficacy
2
no significant difference based on donor type. By contrast, in of the procedure has been demonstrated in all forms of SCID,
their article describing the experiences of HSCT for SCID at although the T-lymphocyte count after HSCT tends to be lower
two centers (Brescia, Italy; and Toronto, Canada), Grunebaum in patients with adenosine deaminase (ADA) deficiency, possibly
et al. reported that aGvHD developed in four (31%) of 13 patients reflecting irreversible thymic damage. Moreover, normalization
who received related HLA-identical HSCT, 18 (45%) of 40 patients of T-lymphocyte count after HSCT demonstrates the ability of
treated with T cell–depleted haploidentical transplantation, and stem cells to seed and differentiate in a vestigial thymus.
6
30 (73%) of 41 patients receiving MUD HSCT. Buckley et al. The kinetics of T-lymphocyte reconstitution differs substan-
reported that GvHD occurred in 45 (30.2%) of 149 patients tially, depending on the type of transplant. The unmanipulated
given T cell–depleted mismatched parental bone marrow, eight graft from a related HLA-identical donor contains mature T
(47%) of 17 patients given unfractionated HLA-identical marrow, lymphocytes. Homeostatic as well as antigen-driven expansion
18a
and four (80%) of five patients given placental blood. In most of these mature T cells occurs as early as 2 weeks after transplanta-
1
cases, GvHD occurred when there was presence of transplacentally tion (Fig. 82.6). These T cells have a memory (CD45RO)
acquired T lymphocytes. In most cases in this study, the GvHD phenotype, are fully competent, and, in fact, provide the recipient
observed was mild (grade I or II) and required no treatment; with functional immunity.
however, 11 patients developed GvHD grade III or IV and required Mature T cells are present also in bone marrow grafts collected
treatment with steroids, cyclosporine, and/or tacrolimus. None from MUDs. However, the use of conditioning in MUD HSCT
of these patients has died since then, but one has developed impairs, at least in part, immune development in such transplants,
cGvHD. Although continuous improvement in HLA typing has compared with unconditioned HSCT from related HLA-identical
resulted in a progressively lower incidence of aGvHD in the last donors (see Fig. 82.6).
+
+
decades, these data illustrate the need for careful monitoring of In contrast, newly generated, naïve (CD45RA CD31 ) T
infants treated with other than related HLA-identical HSCT and lymphocytes do not appear in circulation until 3–6 months after
call for adherence to guidelines on the use of immunosuppression HSCT, irrespective of the type of transplant (HLA-identical or
for GvHD prophylaxis after MUD HSCT or after conditioned mismatched), and their number tends to peak at approximately
T cell–depleted haploidentical transplantation. 1 year after HSCT, when a fully polyclonal T-cell repertoire is
Chronic GvHD disease has been reported in 10 (11%) of 90 usually observed. These naïve T lymphocytes are the product of
patients who have survived for at least 2 years after receiving ongoing active thymopoiesis, as shown by the fact that they
HSCT for SCID in Paris. Six of them developed disseminated contain T-cell receptor excision circles (TRECs). TRECs are
cGvHD, and three died of cGvHD and related infectious complica- extrachromosomal DNA episomes, which are generated during
19
tions. A similar rate of cGvHD (15% at 2 years post-HSCT) V(D)J recombination (Chapter 4) and are not duplicated during
has been observed in the North American series of 240 infants mitosis. Therefore TRECs identify newly generated naïve T
with SCID. 2 lymphocytes, and their enumeration in peripheral blood is used
Immune dysregulation and autoimmunity represent additional as a method to identify babies with SCID at birth. 20
complications of HSCT for SCID. In a joint series of 94 infants The kinetics of T-cell reconstitution is influenced by the
with SCID who received transplantations in Brescia (Italy) and recipient’s age. Transplantations performed early in life (at <3.5
2
Toronto (Canada), we have reported that six of 41 patients who months of age) lead to superior thymic output. This may reflect
received MUD HSCT, and five of 40 children treated with T lack of thymic damage (which is often observed in older infants
cell–depleted haploidentical transplantation developed auto- after infections); alternatively, it is possible that a younger thymus
6
immune cytopenias. These complications may develop a few has intrinsic superior ability to support active thymopoiesis.
months after HSCT (when skewing of the T-cell repertoire may Quantitation of TRECs sequentially after HSCT is an accepted
be observed) or may persist, particularly in infants with delayed approach to assess engraftment of bona fide stem cells and to
and incomplete immune reconstitution. In particular, Neven monitor persistence of immunity. Although an earlier study had
et al. have reported that among 90 long-term survivors after shown that levels of TRECs tended to decline at 10 years after
HSCT for SCID, 12 patients suffered from autoimmune and HSCT in recipients of unconditioned mismatched-related
inflammatory complications at more than 2 years after HSCT transplants, more recent observations from the same group
for SCID, and in six of them, the onset of such complications indicate that robust thymopoiesis and generation of a diversified
19
was within the first 2 years after transplantation. These late repertoire of T lymphocytes were maintained over the long term
manifestations of immune dysregulation are often associated after HSCT. 21
with incomplete immune reconstitution and may lead to a poor Vigor of immune reconstitution at >2 years after HSCT is
outcome. influenced by the donor type, use of conditioning, and SCID

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1123


10000

8000 T lymph B lymph NK lymph
Lymphocytes/µL 6000


4000

4000


γc def JAK3 def IL7R def RAG def Art. def ADA def Unknown Normal
N=15 N=15 N=4 N=6 N=7 N=4 N=13 range

10000

8000
Lymphocytes/µL 6000


4000

4000


γc def JAK3 def IL7R def RAG def Art. def ADA def Unknown Normal
N=11 N=11 N=4 N=3 N=3 N=4 N=6 range
+
+
FIG 82.4 Mean (±SE) numbers of CD3 T cells, CD19 B cells, and CD16 natural killer cells before
+
transplantation (A) and at the most recent evaluation after transplantation (B) performed at the
University of Brescia, Italy, according to the type of severe combined immunodeficiency (SCID).
300 Pre-HSCT Post-HSCT
TRECs (per 10 3 lymphocytes) 200
250


150

100



A 50 γc def JAK3 def IL7R def RAG def Art. def ADA def Unknown Normal
range
Proliferative response to PHA (cpm x 10 -3 ) 200
300
250


150

100





range
B 50 γc def JAK3 def IL7R def RAG def Art. def ADA def Unknown Normal
FIG 82.5 Number of T-cell receptor excision circles (TRECs) (A) and in vitro proliferative response
to phytohemagglutinin (PHA) (B) before transplantation and at the last follow-up after transplantation
in a series of 42 infants with severe combined immunodeficiency (SCID) treated at the University
of Brescia, Italy, according to the type of SCID.

1124 Part NINE Transplantation


More limited data are available about reconstitution of NK
−
3000 cell function. In patients with NK SCID, NK cells are often the
first cells to appear after haploidentical HSCT. Lower NK-cell
counts are observed at long-term follow-up after HSCT in patients
CD3 + lymphocytes/µl 2000 with γc or JAK3 defects.
2500
HSCT for Combined Immunodeficiencies Other Than SCID
1500
The Primary Immune Deficiency Treatment Consortium (PIDTC)
related HLA-identical
1000
MUD
forms of combined immunodeficiency characterized by residual
Related HLA-mismatched has established criteria to distinguish typical SCID from other
500 (although lower than normal) T cell–mediated immunity.
23
Omenn syndrome is a fatal disorder, unless treated with HSCT.
0
0 1 3 6 12 15 24 Satisfactory results have been obtained with transplantation from
related HLA-identical donors, but less so from haploidentical
Months since HSCT donors. However, an improved outcome has been observed in
+
FIG 82.6 Kinetics of CD3 T lymphocytes reconstitution in 48 the last 15 years. Mazzolari et al. have reported that 9 of 11
infants with severe combined immunodeficiency (SCID) following patients with Omenn syndrome were alive after HSCT; importantly,
human leukocyte antigen (HLA)-identical (n = 12), matched of these, only one had a matched sibling, whereas two infants
unrelated donor (MUD) (n = 15) or haploidentical (n = 21) hema- had a phenotypically identical related donor, three were treated
topoietic stem cell transplantation (HSCT), performed at the with MUD HSCT, and five received a haploidentical HSCT. 22
+
University of Brescia, Italy. Geometrical mean CD3 T-cell counts HSCT has been attempted also in other predominant T-cell
are shown. immunodeficiencies, such as purine nucleoside phosphorylase
deficiency, cartilage hair hypoplasia, and other forms of T-cell
phenotype. In particular, in a large series of patients with SCID activation deficiency. Overall, survival after HSCT for these
who received HSCT in North America between 2000 and 2009, disorders is worse than in typical SCID (approximately 50%). 1
higher counts of CD3 cells at 2–5 years after transplantation Major histocompatibility complex (MHC) class II deficiency
were associated with HSCT from MSDs, use of conditioning, remains a very difficult disease to treat with transplantation. In
+
−
2
and B or NK SCID phenotype. Use of conditioning resulted the European series, cumulative survival after HSCT performed
1
also in a higher count of naïve CD4 cells. 2 in 1995–2005 was around 40%. Many patients with this deficiency
fail to reconstitute the number of circulating CD4 T lymphocytes,
Reconstitution of B- and NK-Cell Immunity probably because the lack of expression of HLA class II molecules
In contrast to what is observed for T lymphocytes, the engraftment on thymic epithelial cells prevents positive selection of CD4
of B cells after HSCT for SCID is often problematic and delayed. lymphocytes.
In their series, Buckley et al. reported that only five of 17 The mainstay of treatment for patients with complete DiGeorge
24
survivors of HLA-identical HSCT and 33 of 109 survivors of syndrome is represented by thymic transplantation. However,
unconditioned haploidentical HSCT had evidence of donor- HSCT or even transplantation of unmobilized peripheral blood
derived B lymphocytes; overall, 63 of 126 survivors required mononuclear cells may be attempted if an HLA-identical donor
18a
intravenous immunoglobulins. Booster transplants have been is available; in such cases, immune reconstitution is provided
used to overcome these problems; 33 of 49 patients who received by mature T lymphocytes contained in the graft.
such booster transplants at Buckley et al.’s institution were Patients with CD40 ligand (CD40L) deficiency suffer from
reported to be alive with improved immune function. 18a recurrent bacterial and opportunistic (Pneumocystis jiroveci,
In Europe, use of pretransplantation conditioning has been Cryptosporidium parvum) infections, resulting in increased
advocated in haploidentical HSCT with the goal of facilitating mortality in childhood and young adulthood. This has prompted
engraftment of stem cells and thus also of B lymphocytes. Maz- use of HSCT in the treatment of this disease. In a series of 38
25
zolari et al. have reported that donor B-cell engraftment was patients transplanted in Europe, 26 (68.4%) survived. Early
achieved in only one of 11 survivors after unconditioned HSCT age at transplantation and lack of preexisting pulmonary disease
compared with 26 of 29 patients who received a myeloablative were associated with a more favorable outcome. Successful outcome
conditioning regimen. 22 has been also reported after HSCT for CD40 deficiency.
Recently, higher B-cell counts and a better chance of attaining X-linked immunodeficiency with ectodermal dystrophy, caused
independence from Ig replacement therapy in recipients of condi- by mutations of the IKBKG (NEMO) gene that impair nuclear
tioned HSCT for SCID have been confirmed also in a large North factor (NF)-κB signaling, may also present with features of
2
American study. Finally, attainment of normal B-cell function combined immunodeficiency. Colitis is also frequently seen and
may also depend on the nature of the genetic defect, as shown may be caused by immune dysregulation as well as abnormalities
+
by the fact that among infants with B SCID, those who have of NF-κB signaling in the gut epithelial cells. Initial observations
an IL-7RA gene defect usually develop normal B-cell immunity supported the notion that use of myeloablative regimens for
after HSCT, even if no donor-derived B cells are present, whereas HSCT is poorly tolerated in this condition, also because of
patients with γc or JAK3 deficiency (both of which compromise increased mucosal toxicity. However, graft failure has been
B-cell function) often remain dependent on Ig substitution therapy observed when using reduced-intensity conditioning, and in
if engraftment of donor-derived B cells is not achieved. This contrast, several patients have tolerated myeloablative regimens
26
reflects the importance of interleukin-21 (IL-21)–mediated, γc/ well and have attained immune reconstitution. Yet, persistence
JAK3-dependent signaling for germinal-center reaction, plasma- of the colitis after HSCT has been reported in a proportion of
27
cell differentiation, and antibody production. the patients. Conflicting results have also been reported after

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1125


HSCT in patients with hypermorphic mutations of the IKBA 1
gene, another cause of combined immune deficiency with 0.9
ectodermal dystrophy. 28 0.8
Wiskott-Aldrich Syndrome 0.7
Bone marrow transplantation for correction of WAS was 0.6
attempted as early as 1968, with partial success. Full correction Survival 0.5
following HSCT was first reported in 1978 when a more robust 0.4
conditioning regimen was used. Since then, results of related 0.3
HLA-identical HSCT in WAS have been consistently good, with
continuous improvement in recent years, and excellent results 0.2
have been also achieved with HSCT from MUDs. A multiinsti- 0.1
tutional study of 194 patients with WAS treated with HSCT 0
showed an overall survival of 84%, and a 5-year survival as high 0 500 1000 1500 2000 2500 3000
29
as 89.1% for transplantations performed since year 2000. For Days following HCT
patients treated with MUD HSCT, younger age (<5 years) at FIG 82.7 Long-term survival of patients after reduced intensity
transplantation was associated with an improved outcome. conditioning hematopoietic stem cell transplantation (HSCT) for
Patients with preexisting autoimmunity or recurrent/severe X-linked lymphoproliferative disease type 1 (XLP1). (With permis-
infections had a higher rate of complications after transplantation. sion from Moratto D, Giliani S, Bonfim C, et al. Long-term
Mixed chimerism was associated with the presence of immu- outcome and lineage-specific chimerism in 194 Wiskott–Aldrich
nological abnormalities and increased risk of autoimmunity, Syndrome patients treated by hematopoietic cell transplantation
and myeloid chimerism <50% was associated with persistent between 1980–2009: an international collaborative study. Blood
29
thrombocytopenia. Use of a conditioning regimen other than 2011;118:1675–84.)
fully myeloablative conditioning was associated with reduced
disease-free survival in another study of 14 patients. 30
was 80% at 1 year and 71% in the long term; mixed chimerism
Cytotoxicity Defects was common but, in most cases, was sufficient to control the
Familial hemophagocytic lymphohistiocytosis (FHL) comprises disease. Infectious complications (especially viral infections) were
a genetically heterogeneous group of disorders of T cell– and recorded in the majority of the patients. 33
NK cell–mediated cytotoxicity. Although chemotherapy may XLP type 2 (XLP2), caused by mutations in the X-linked
induce remission, patients with FHL tend to relapse and ultimately inhibitor of apoptosis (XIAP) gene, is associated with various
die, mostly as a result of multiorgan failure observed in the phenotypes, including XLP, HLH, and severe colitis. An inter-
accelerated phase of the disease. At present, HSCT is the only national survey of transplantation for this condition identified
curative approach to FHL. However, patients with FHL often 19 patients who underwent HSCT, in which fully myeloablative
are critically ill, with extensive organ involvement and/or active (n = 7), reduced-intensity (n = 11), or intermediate-level (n =
34
infections, and may suffer from refractory disease. For these 1) conditioning was used. Myeloablative conditioning was
reasons, patients are unusually prone to developing transplantation- associated with poor survival (14%) and a high rate of serious,
related toxicities and complications. Recently, significantly transplantation-related toxicities. Survival was better (55%) in
improved outcomes with increased survival have been reported patients who received reduced-intensity conditioning and was
after HSCT with the use of a reduced-intensity conditioning especially good (86%) in those in remission from HLH. 34
31
regimen. However, use of reduced-intensity conditioning is The hematological and immune complications of Chediak-
associated with a higher risk of mixed and recipient chimerism Higashi syndrome (CHS) can be cured with bone marrow
31
(65%), even for HSCT from matched donors. Selection of transplantation. In a series of 35 patients, 5-year survival after
35
optimal family donors is an important issue for HSCT in FHL. HSCT was 62%. Mortality was higher in patients who were in
Functional and genetic studies should be used to screen potential the life-threatening accelerated phase of the disease at the time
family donors to avoid obtaining transplants from genetically of transplantation and in those who received HSCT from an
35
affected but as-yet asymptomatic subjects. alternative related donor. Use of MUDs represents a valid option
Excellent results have been reported also after HSCT for for HSCT in patients with CHS. However, the long-term outcome
X-linked lymphoproliferative disease type 1 (XLP1). In an of patients with CHS treated with HSCT remains unclear,
international series of 91 patients with XLP caused by a SH2D1A especially since neurological deterioration has been consistently
gene defect, survival was 81.4% in 43 patients treated with HSCT observed several years after transplantation.
32
compared with 62.5% in 48 untransplanted patients. Further- Griscelli syndrome type 2 (GS2) is a genetic disease character-
more, the majority of untransplanted survivors required Ig ized by hemophagocytic lymphohistiocytosis and a high risk of
replacement therapy, whereas good immune reconstitution was neurological complications. Correction of the cytotoxicity defect
achieved in most transplanted patients. However, survival was after HSCT has been reported in a small series of patients with
poorer (50%) among patients with a previous history of hemo- GS2, but neurological sequelae remain a challenge and may develop
phagocytic lymphohistiocytosis (HLH), suggesting that ideally even in patients without preexisting neurological problems. 36
the transplantation should be performed before onset of EBV
32
infection. More recently, a single-center experience involving Phagocytic Cell Disorders
16 patients with XLP1 showed that reduced-intensity conditioning Although regular administration of prophylactic antibiotics and
with alemtuzumab, fludarabine, and melphalan is effective even antifungal agents (with the possible addition of interferon [IFN]-
33
in patients with XLP1 with a history of HLH (Fig. 82.7). Survival γ) has clearly improved the outcome in patients with CGD, this

1126 Part NINE Transplantation


remains a severe disorder with a rather high risk of complications However, a subgroup of these patients fail to respond to G-CSF,
37
and death, especially in patients with oxidase-null phenotype. and some of them are at high risk for development of myelogenous
Therefore there has been a renewed interest in HSCT for CGD. leukemia. A retrospective multicenter study of 136 patients with
In a survey of the European experience in 1985–2000, 27 patients SCN who underwent allogeneic HSCT between 1990 and 2012
with CGD were treated with HSCT; most of them received reported a 3-year overall survival rate of 82%, with 17%
41
unmodified bone marrow from an HLA-identical sibling after transplantation-related mortality. Transplantation at a younger
myeloablative conditioning. Overall, 23 of the 27 patients were age (<10 years) and use of matched related or unrelated donors
reported to have survived, and the disease had been cured in 22 were associated with an improved outcome.
38
of them. In this study, all 18 patients who were infection-free
at the time of transplantation survived. However, a real break- Other Primary Immune Deficiencies
through in the treatment of CGD was provided by a prospective IFN-γ receptor 1 deficiency leads to severe mycobacterial
multicenter study in 56 patients with CGD, 42 of whom had infections, with a high rate of mortality early in life. Although
high-risk features, including treatment-refractory infections and HSCT should theoretically correct the disease, results have been
39
severe inflammatory complications. In this study, reduced- disappointing, with few exceptions. An international survey of
intensity conditioning (with low-dose or targeted busulfan eight transplanted patients showed that only two were in full
administration, high-dose fludarabine, and serotherapy) and remission 5 years after transplantation. The high level of IFN-γ
transplantation from matched related (n = 21) or unrelated (n in these patients inhibits engraftment of donor-derived HSCs,
= 35) donors resulted in a 2-year overall survival rate of 96% accounting for the high rejection rate. Correction of the disease
and an event-free survival rate of 91% (Fig. 82.8). Stable (≥90%) has been reported with HSCT following control of mycobacterial
myeloid chimerism was documented in 93% of surviving patients. infection, normalization of IFN-γ levels, and use of myeloablative
A low rate of aGvHD of grade III/IV (4%) and of cGvHD (7%) conditioning.
was observed. Immunodysregulation/polyendocrinopathy/enteropathy/X-
HSCT is a successful and lifesaving procedure also in patients linked (IPEX) syndrome is a severe disorder with immune
with the complete form of leukocyte adhesion defect type 1 dysregulation caused by mutations of the FOXP3 gene, which
(LAD-1). A multicenter study of 36 such patients who underwent plays a critical role in the development and function of regulatory
HSCT between 1993 and 2007 reported an overall survival of T cells (Tregs). Patients with IPEX often die early in infancy.
42
75%, with similar results when matched related or unrelated HSCT is the only curative approach to treat this disease. Selective
donors were used. Mortality was higher (four of eight cases) after advantage for donor-derived Tregs has been observed in patients
43
haploidentical HSCT. Stable mixed multilineage chimerism is with mixed chimerism after HSCT. However, other studies have
40
sufficient to cure the disease. More recently, employing blockade reported a higher risk of graft failure and increased occurrence
of the P40 subunit of IL-23 and IL-12 with ustekinumab has of posttransplantation autoimmune cytopenias following HSCT
been reported resolve inflammatory symptoms in a patient with with reduced-intensity conditioning. 44
LAD-1 after 1 year of therapy. 40a Dedicator of cytokinesis 8 (DOCK8) deficiency is a recently
Administration of recombinant G-CSF is the treatment of described form of combined immunodeficiency with elevated
choice for patients with severe congenital neutropenia (SCN). serum IgE, cutaneous viral infections, and a high risk of malig-
nancy. HSCT represents an effective form of treatment for this
disease. In a series of 11 patients, seven of whom received HSCT
100 from related donors and four from unrelated donors, survival
was excellent (91%), and selective advantage for donor-derived
45
80 T cells and switched memory cells was noticed. In other studies,
successful outcomes were reported with either myeloablative or
reduced-intensity conditioning. HSCT has been also successfully
EFS survival (%) 60 used in patients with combined immune deficiency resulting
46
from DOCK2 deficiency.
40
FUTURE TRANSLATIONAL RESEARCH FOR HCST IN
THE TREATMENT OF PID
20
MUD Although HSCT has clearly shown its efficacy in patients with
MRD severe forms of PID, several goals remain to be met. In particular,
0 the main areas of interest for future development include (i)
0 24 48 72 96 120 methods to improve and sustain engraftment of stem cells; (ii)
Months after HSCT strategies to facilitate engraftment and to reduce the incidence
35 12 3 2 1 0 of GvHD in recipients of mismatched or matched unrelated
21 12 3 3 3 0 transplants; (iii) attempts to improve thymopoiesis, with the
FIG 82.8 Event-free survival (EFS) in patients with chronic goal to accelerate immune reconstitution and to avoid or postpone
granulomatous disease (CGD) after reduced-intensity conditioning long-term decline of immunity; and (iv) design strategies to
hematopoietic stem cell transplantation (HSCT) from matched reduce the burden of infections after HSCT.
related donors (MRD) or matched unrelated donors (MUD). (With Generation of a diversified pool of naïve T lymphocytes after
permission from Eapen M, DeLaat CA, Baker KS, et al. Hema- HSCT depends on thymic function. Use of cytotoxic drugs and
topoietic cell transplantation for Chediak-Higashi syndrome. Bone GvHD are significant risk factors for posttransplantation T-cell
47
Marrow Transplant 2007;39:411–5.) deficiency as they interfere with normal thymic function. Use

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1127


of mAbs against CD117 (c-kit) has been proposed to displace 5. Kwan A, Abraham RS, Currier R, et al. Newborn screening for severe
autologous stem cells and favor engraftment of donor-derived combined immunodeficiency in 11 screening programs in the United
cells, while avoiding the drug-related toxicity of chemotherapy States. JAMA 2014;312:729–38.
48
and radiotherapy. Recognition that B-cell engraftment after 6. Grunebaum E, Mazzolari E, Porta F, et al. Bone marrow transplantation
for severe combined immunodeficiency. JAMA 2006;295:508–18.
HSCT is hampered by competition between host and donor 7. Gennery AR, Cant AJ. Cord blood stem cell transplantation in primary
49
early B-cell precursors may prompt novel forms of conditioning immune deficiencies. Curr Opin Allergy Clin Immunol 2007;7:528–34.
regimens targeted to host pro-B lymphocytes, in an attempt to 8. Hassan A, Lee P, Maggina P, et al. Host natural killer immunity is a key
facilitate development of donor-derived B cells, even in the indicator of permissiveness for donor cell engraftment in patients with
unconditioned HLA-identical setting. severe combined immunodeficiency. J Allergy Clin Immunol
Strategies aimed at improving thymic function after HSCT 2014;133:1660–6.
may include (i) protection of thymic stroma that supports 9. Slatter MA, Rao K, Amrolia P, et al. Treosulfan-based conditioning
thymopoiesis and (ii) direct stimulation of early T-cell progenitors. regimens for hematopoietic stem cell transplantation in children with
Keratinocyte growth factor (KGF) is potentially attractive because primary immunodeficiency: United Kingdom experience. Blood
it protects the thymic stroma. Administration of KGF before 2011;117:4367–75.
HSCT has been shown to enhance thymopoiesis and peripheral 10. Ferrara JL, Levine JE, Reddy P, et al. Graft-versus-host disease. Lancet
2009;373:1550–61.
T-cell numbers and to reduce the incidence and severity of GvHD 11. Wolff D, Gerbitz A, Ayuk F, et al. Consensus conference on clinical
50
in murine models of HSCT. However, more must be learned practice in chronic graft-versus-host disease (GVHD): first-line and
about the long-term outcome of this treatment before clinical topical treatment of chronic GVHD. Biol Blood Marrow Transplant
trials can be started. 2010;16:1611–28.
Attempts to accelerate immune reconstitution might be based 12. Wolff D, Schleuning M, von Harsdorf S, et al. Consensus conference on
on the use of cytokines, such as IL-7, which promote T-cell clinical practice in chronic GVHD: second-line treatment of chronic
development and maturation in the thymus. However, experience graft-versus-host disease. Biol Blood Marrow Transplant 2011;17:1–17.
in severely immunodeficient mice has shown that infusion of 13. Tomblyn M, Chiller T, Einsele H, et al. Guidelines for preventing
IL-7 provides limited benefit in the MHC-compatible or partially infectious complications among hematopoietic cell transplantation
compatible setting, whereas it has facilitated T-cell development recipients: a global perspective. Biol Blood Marrow Transplant
2009;15:1143–238. Erratum in: Biol Blood Marrow Transplant
in a fully mismatched setting. 2010;16:294.
14
Finally, besides use of virus-specific CTLs, infusion of donor 14. Naik S, Nicholas SK, Martinez CA, et al. Adoptive immunotherapy for
T cells that have antiinfective activity but no GvHD activity primary immunodeficiency disorders with virus-specific T lymphocytes. J
might help reduce infection-related mortality after HSCT. Such Allergy Clin Immunol 2016;137:1498–505.
strategy could be based on removal of alloreactive cells by negative 15. Teigland CL, Parrott RE, Buckley RH. Long-term outcome of
selection using anti-CD25 or anti-CD69 antibodies. non-ablative BMT in patients with SCID. Bone Marrow Transplant
2013;48:1050–5.
ON tHE HOrIZON 16. Schuetz C, Neven B, Dvorak CC, et al. SCID patients with ARTEMIS vs
RAG deficiencies following HCT: increased risk of late toxicity in
The following strategies may help improve even further the outcome of ARTEMIS-deficient SCID. Blood 2014;123:281–9.
hematopoietic stem cell transplantation (HSCT) for primary immunode- 17. Hassan A, Lee P, Maggina P, et al. Host natural killer immunity is a key
ficiency (PID) in the future: indicator of permissiveness for donor cell engraftment in patients with
Methods to improve and sustain engraftment of stem cells severe combined immunodeficiency. J Allergy Clin Immunol
Strategies to facilitate engraftment and reduce the occurrence of graft- 2014;133:1660–6.
versus-host disease (GvHD) in patients with mismatched or matched 18. Hassan A, Booth C, Brightwell A, et al. Outcome of hematopoietic stem
unrelated donor (MUD) transplant cell transplantation for adenosine deaminase-deficient severe combined
Approaches to increase thymopoiesis and accelerate immune immunodeficiency. Blood 2012;120:3615–24.
reconstitution 18a. Buckley RH. Transplantation of hematopoietic stem cells in human
Design of therapeutic strategies to reduce the infectious complications severe combined immunodeficiency: longterm outcomes. Immunol Res
of HSCT 2011;49(1-3):25–43.
19. Neven B, Leroy S, Decaluwe H, et al. Long-term outcome after
hematopoietic stem cell transplantation of a single-center cohort of 90
Please check your eBook at https://expertconsult.inkling.com/ patients with severe combined immunodeficiency. Blood
for self-assessment questions. See inside cover for registration 2009;113:4114–24.
details. 20. Kwan A, Abraham RS, Currier R, et al. Newborn screening for severe
combined immunodeficiency in 11 screening programs in the United
REFERENCES States. JAMA 2014;312:729–38.
21. Sarzotti-Kelsoe M, Win CM, Parrott RE, et al. Thymic output, T-cell
1. Gennery AR, Slatter MA, Grandin L, et al. Transplantation of diversity, and T-cell function in long-term human SCID chimeras. Blood
hematopoietic stem cells and long-term survival for primary 2009;114:1445–53.
immunodeficiencies in Europe: entering a new century, do we do better? J 22. Mazzolari E, Forino C, Guerci S, et al. Long-term immune reconstitution
Allergy Clin Immunol 2010;126:602–10.e1–11. and clinical outcome after stem cell transplantation for severe T-cell
2. Pai SY, Logan BR, Griffith LM, et al. Transplantation outcomes for severe immunodeficiency. J Allergy Clin Immunol 2007;120:892–9.
combined immunodeficiency, 2000-2009. N Engl J Med 2014;371:434–46. 23. Shearer WT, Dunn E, Notarangelo LD, et al. Establishing diagnostic
3. Klein OR, Chen AR, Gamper C, et al. Alternative-donor hematopoietic criteria for severe combined immunodeficiency disease (SCID), leaky
stem cell transplantation with post-transplantation cyclophosphamide SCID, and Omenn syndrome: the Primary Immune Deficiency
for nonmalignant disorders. Biol Blood Marrow Transplant Treatment Consortium experience. J Allergy Clin Immunol 2014;133:
2016;22:895–901. 1092–8.
+
+
4. Handgretinger R. Negative depletion of CD3 and TcRαββ T cells. Curr 24. Markert ML, Devlin BH, Chinn IK, et al. Thymus transplantation in
Opin Hematol 2012;19:434–9. complete DiGeorge anomaly. Immunol Res 2009;44:61–70.

1128 Part NINE Transplantation


25. Gennery AR, Khawaja K, Veys P, et al. Treatment of CD40 ligand 38. Seger RA, Gungor T, Belohradsky BH, et al. Treatment of chronic
deficiency by hematopoietic stem cell transplantation: a survey of the granulomatous disease with myeloablative conditioning and an
European experience, 1993–2002. Blood 2004;103:1152–7. unmodified allograft: a survey of the European experience, 1985–2000.
26. Permaul P, Narla A, Hornick JL, et al. Allogeneic hematopoietic stem cell Blood 2002;100:4344–50.
transplantation for X-linked ectodermal dysplasia and immunodeficiency: 39. Güngör T, Teira P, Slatter M, et al. Reduced-intensity conditioning and
case report and review of outcomes. Immunol Res 2009;44:89–98. HLA-matched haemopoietic stem-cell transplantation in patients with
27. Pai SY, Levy O, Jabara HH, et al. Allogeneic transplantation successfully chronic granulomatous disease: a prospective multicentre study. Lancet
corrects immune defects, but not susceptibility to colitis, in a patient with 2014;383:436–48.
nuclear factor-κB essential modulator deficiency. J Allergy Clin Immunol 40. Qasim W, Cavazzana-Calvo M, Davies EG, et al. Allogeneic hematopoietic
2008;122:1113–18. stem-cell transplantation for leukocyte adhesion deficiency. Pediatrics
28. Fish JD, Duerst RE, Gelfand EW, et al. Challenges in the use of allogeneic 2009;123:836–40. Erratum in: Pediatrics 2009;123:1436.
hematopoietic SCT for ectodermal dysplasia with immune deficiency. 40a. Moutsopoulos NM, Zerbe CS, Wild T, et al. Interleukin-12 and
Bone Marrow Transplant 2009;43:217–21. interleukin-23 blockade in leukocyte adhesion deficiency type 1. N Engl J
29. Moratto D, Giliani S, Bonfim C, et al. Long-term outcome and Med 2017;376:1141–6.
lineage-specific chimerism in 194 Wiskott–Aldrich Syndrome patients 41. Fioredda F, Iacobelli S, van Biezen A, et al. Stem cell transplantation in
treated by hematopoietic cell transplantation between 1980–2009: an severe congenital neutropenia: an analysis from the European Society for
international collaborative study. Blood 2011;118:1675–84. Blood and Marrow Transplantation. Blood 2015;126:1885–92.
30. Stepensky P, Krauss A, Goldstein G, et al. Impact of conditioning on 42. Rao A, Kamani N, Filipovich A, et al. Successful bone marrow
outcome of hematopoietic stem cell transplantation for Wiskott-Aldrich transplantation for IPEX syndrome after reduced-intensity conditioning.
syndrome. J Pediatr Hematol Oncol 2013;35:e234–8. Blood 2007;109:383–5.
31. Marsh RA, Jordan MB, Filipovich AH. Reduced-intensity conditioning 43. Horino S, Sasahara Y, Sato M, et al. Selective expansion of donor-derived
haematopoietic cell transplantation for haemophagocytic regulatory T cells after allogeneic bone marrow transplantation in a
lymphohistiocytosis: an important step forward. Br J Haematol patient with IPEX syndrome. Pediatr Transplant 2014;18:E25–30.
2011;154:556–63. 44. Kucuk ZY, Bleesing JJ, Marsh R, et al. A challenging undertaking: Stem
32. Booth C, Gilmour KC, Veys P, et al. X-linked lymphoproliferative disease cell transplantation for immune dysregulation, polyendocrinopathy,
due to SAP/SH2D1A deficiency: a multicenter study on the enteropathy, X-linked (IPEX) syndrome. J Allergy Clin Immunol
manifestations, management and outcome of the disease. Blood 2016;137:953–955.e4.
2011;117:53–62. 45. Al-Herz W, Chu JI, van der Spek J, et al. Hematopoietic stem cell
33. Marsh RA, Bleesing JJ, Chandrakasan S, et al. Reduced-intensity transplantation outcomes for 11 patients with dedicator of cytokinesis 8
conditioning hematopoietic cell transplantation is an effective treatment deficiency. J Allergy Clin Immunol 2016;138:852–9.
for patients with SLAM-associated protein deficiency/X-linked 46. Dobbs K, Domínguez Conde C, Zhang SY, et al. Inherited DOCK2
lymphoproliferative disease type 1. Biol Blood Marrow Transplant Deficiency in Patients with Early-Onset Invasive Infections. N Engl J Med
2014;20:1641–5. 2015;372:2409–22.
34. Marsh RA, Rao K, Satwani P, et al. Allogeneic hematopoietic cell 47. Krenger W, Blazar BR, Holländer GA. Thymic T-cell development in
transplantation for XIAP deficiency: an international survey reveals poor allogeneic stem cell transplantation. Blood 2011;117:6768–76.
outcomes. Blood 2013;121:877–83. 48. Czechowicz A, Kraft D, Weissman IL, et al. Efficient transplantation via
35. Eapen M, DeLaat CA, Baker KS, et al. Hematopoietic cell transplantation antibody-based clearance of hematopoietic stem cell niches. Science
for Chediak-Higashi syndrome. Bone Marrow Transplant 2007;39: 2007;318:1296–9.
411–15. 49. Liu A, Vosshenrich CA, Lagresle-Peyrou C, et al. Competition within the
36. Pachlopnik Schmid J, Moshous D, Boddaert N, et al. Hematopoietic stem early B cell compartment conditions B cell reconstitution after
cell transplantation in Griscelli syndrome type 2: a single-center report hematopoietic stem cell transplantation in non-irradiated recipients.
on 10 patients. Blood 2009;114:211–18. Blood 2006;108:1123–8.
37. Kuhns DB, Alvord WG, Heller T, et al. Residual NADPH oxidase and 50. Wils EJ, Cornelissen JJ. Thymopoiesis following allogenic stem cell
survival in chronic granulomatous disease. N Engl J Med transplantation: new possibilities for improvement. Blood Rev
2010;363:2600–1. 2005;19:89–98.

CHaPtEr 82 Immune Reconstitution Therapy for Immunodeficiency 1128.e1


MUL t IPLE-CHOICE QUES t IONS

1. What distinguishes the kinetics of immune reconstitution C. Presence of active infections at the time of transplantation
following hematopoietic stem cell transplantation (HSCT) D. Age younger than 2 months of life
with unmanipulated bone marrow from a matched sibling E. Transplantation from a haploidentical donor
donor versus a T cell–depleted transplantation from a hap- 3. Use of reduced intensity conditioning regimens may reduce
loidentical donor in an infant with severe combined immu- drug-related toxicity but often results in mixed chimerism
nodeficiency (SCID)? after HSCT for primary immune deficiency (PID). In many
A. Transplantation from a matched sibling donor will provide cases, mixed chimerism is sufficient to control the disease
rapid engraftment of B lymphocytes. phenotype, but in some forms of PID, it is associated with
B. Transplantation from an human leukocyte antigen (HLA)– an increased risk of complications. Can you identify in which
matched donor allows rapid T-cell engraftment. of the following immunodeficiency this is the case?
C. Only transplantation from a matched sibling donor permits A. Familial hemophagocytic lymphohistiocytosis (FHL)
durable T-cell reconstitution. B. Chronic granulomatous disease (CGD)
D. Transplantation from an HLA-matched donor allows rapid C. Immune dysfunction/polyendocrinopathy/enteropathy/X-
multilineage hematopoietic engraftment. linked (IPEX) syndrome
E. There are no differences in the kinetics of immune D. Wiskott-Aldrich syndrome (WAS)
reconstitution.
E. Leukocyte adhesion defect type 1 (LAD-1)
2. Which of the following factors negatively affect outcome of
HSCT for SCID?
+
A. B SCID phenotype
B. History of previous infections

83






Hematopoietic Stem Cell Transplantation for

Malignant Diseases



Pashna N. Munshi, Scott D. Rowley, Robert Korngold








Hematopoietic stem cell transplantation (HSCT) is effective treat- tumor vaccines) to induce an effective immune responsiveness
ment for most hematological malignancies, including leukemia, to the residual disease after transplantation.
lymphoma, multiple myeloma (MM), and clonal myelodysplastic
and myeloproliferative diseases (MPDs), as well as nonmalignant
diseases, such as autoimmune disorders and hemoglobinopathies. THERAPEUTIC PRINCIPLES
Autologous HSCT (auto-HSCT) is commonly used as therapy Autologous and Allogenic Transplantation
for patients with malignancies sensitive to chemotherapy or
radiotherapy in a dose-responsive manner. These patients Autologous Transplantation
receive intensive cytoreductive regimens designed to eliminate • Based on chemotherapy or radiotherapy dose-sensitivity of disease
all tumor cells but which, in so doing, also destroy the patient’s being treated
hematopoietic function needed for blood formation. Infusion of • Requires collection and storage of adequate hematopoietic stem cells
(HSCs), preferably before extensive alkylating agent or purine analogue
previously collected hematopoietic stem cells (HSCs) will rescue therapy
the patient from the marrow-ablative effects of this treatment. • Lower risk of graft failure (no immunological rejection)
Allogeneic HSCT (allo-HSCT), in addition to reconstitution of • No routine posttransplantation immunosuppression
bone marrow function, achieves an immunotherapeutic benefit • Minimal risk of graft-versus-host disease (GvHD)
from the donor natural killer (NK) and T cells infused into • No graft-versus-tumor (GvT) effect
the graft attacking residual tumor cells that persist after the • More rapid posttransplantation immune reconstitution
• Risk of tumor cell contamination in HSC product
conditioning regimen, thereby greatly reducing the risk of later • Not useful for diseases in which normal HSCs cannot be collected
relapse of the disease. Thus allo-HSCT, in contrast to auto-HSCT, (e.g., chronic myelogenous leukemia, myelodysplasia)
does not require administration of dose-intensive regimens to
achieve complete tumor cell kill, and nonmyeloablative regimens Allogeneic Transplantation
may be used to “condition” the host for transplantation. • Rescues bone marrow function after dose-intensive therapy
Auto-HSCT (including syngeneic twins) is justified by the • Effective with reduced-intensity conditioning regimens
dose-sensitivity of most hematological malignancies. Although • Achieves a GvT effect in many malignancies
there is some evidence that a more robust immunological recovery • Risk of GvHD distinct from the beneficial GvT effect
• Higher risk of transplantation-related complications that may offset
after auto-HSCT predicts for a lower risk of relapse, possibly the benefit of the GvT effect
opening an area of research in graft (or host) modification to • Risk of immunological graft rejection
enhance such recovery, treatment of the disease is primarily a • Slow posttransplantation immune reconstitution
result of the dose-intensive, myeloablative chemotherapy or • No risk of tumor-cell contamination with healthy donor
radiotherapy administered. Infusion of cells is only required to
recover hematopoiesis, and the stem cell infusion is, therefore,
intended to treat the deleterious effect of chemotherapy on bone Allo-HSCT has a much lower relapse risk compared with
marrow function and not the disease itself. The primary complica- auto-HSCT as a result of a beneficial immunological graft-versus-
tions of auto-HSCT result from the administration of a dose- tumor (GvT) effect achieved by engraftment of the donor immune
intensive regimen and include a period of marrow hypoplasia, system. Allograft recipients, however, face a much higher risk of
possibly requiring blood transfusions and antibiotics. Nonhe- treatment-related mortality (TRM) from the detrimental immu-
matological toxicities, including mucositis resulting in inanition nological GvH response against healthy tissues of the patient. The
and diarrhea, and damage to other organs, such as the lung, principal complication of allo-HSCT is graft-versus-host disease
liver, and kidney, limit the amount of chemotherapy that can (GvHD), which can occur early (acute GvHD, within the first
be administered. Currently, the treatment-related mortality risk several weeks) (Table 83.1) or late (chronic GvHD, months to
for most treatment regimens is ≤5%. Relapse of disease, particu- several years) after transplantation. The rate of overall incidence
larly for patients who come to transplantation with chemotherapy- of moderate to severe acute GvHD (aGvHD) is 35%–80% for all
refractory disease, is the primary cause of failure of auto-HSCT. patients receiving a human leukocyte antigen (HLA)–matched
Improvements in the outcome of auto-HSCT will require new related or unrelated donor stem cell transplant, and aGvHD is
conditioning regimens with greater tumor cell kill, effective a primary cause of death in 10–20% of these patients. Chronic
posttransplantation consolidation therapies, or strategies (e.g., GvHD (cGvHD), a clinicopathologically distinctive form of this

1129

1130 PART NINE Transplantation



TABLE 83.1 Acute Graft-Versus-Host (e.g., A*02:01 vs A*02:02). The risks of aGvHD, cGvHD, and
Disease (GvHD) transplantation-related mortality increase with the number of
HLA mismatches, and ideally, genotypically matched unrelated
Clinical Staging donors are sought for patients lacking an HLA-identical sibling.
Stage Skin Liver Gut Disparity at HLA-A, -B, -C, and DRB1 alleles are definite risk
factors for survival after unrelated donor transplantation, whereas
1 Maculopapular Bilirubin Diarrhea 500–1000 mL/ single HLA-DQ or -DP mismatches appear to be better tolerated
rash <25% 2–3 mg/dL day, or
BSA Persistent nausea and/or permissive, meaning they do not have a deleterious impact
2
2 Maculopapular Bilirubin Diarrhea 1000–1500 mL/ on clinical outcome.
rash 25–50% 3–6 mg/dL day Not all mismatches result in a deleterious clinical outcome,
BSA and identification of permissive mismatches will improve the
3 Generalized Bilirubin Diarrhea >1500 mL/day number of donors available to the patient. Algorithms accounting
erythroderma 6–15 mg/dL for possible permissive mismatching in the selection of unrelated
4 Desquamation Bilirubin Pain ± ileus donors are currently being developed and tested. 3
and bullae >15 mg/dL
formation GvHD and GvT occurring after HLA-compatible sibling
transplantation demonstrate the importance of minor histo-
Clinical Grading
compatibility antigens (miHAs) on the outcome of allo-HSCT.
Functional MiHAs are derived from polymorphic sites in normal proteins
Overall Grade Skin Liver Gut Impairment between individuals and are constantly processed by proteasome
0 (none) 0 0 0 0 activity and presented on the cell surface by MHC molecules;
1 (mild) 1–2 0 0 0 they can thus be recognized by T cells from a HLA-matched
2 (moderate) 1–3 1 1 1 donor. As a result, hundreds of miHAs may be variably expressed
3 (severe) 2–3 2–3 2–3 2 on host tissues and can trigger an alloresponse from donor T
4 (life-threatening) 1–4 1–4 1–4 3 cells, thereby causing GvHD. Unfortunately, only a few miHAs

Adapted from: Harris AC, Young R, Devine S, et al. International, Multicenter have been identified, such as those on the Y chromosome leading
Standardization of Acute Graft-versus-Host Disease Clinical Data collection: A Report to a higher risk of GvHD in male recipients of cells from female
from the Mount Sinai Acute GVHD International Consortium. Biol Blood Marrow donors; miHA matching of a male donor for a male recipient
Transplant 2016;22:4–10.
is not a component of current algorithms for donor selection.
Research in the field of additional criteria for donor selection
is ongoing. Human NK cells possess clonally distributed, inhibi-
alloreaction, occurs in up to 80% of recipients, may involve aspects tory receptors termed “killer cell immunoglobulin-like receptors”
of regulatory T cell (Treg) dysfunction and autoimmune-like (KIRs) that recognize epitopes shared by groups of HLA class I
responses, and may require years of therapy before tolerance is alleles (KIR ligands). KIR–ligand mismatching in the GvH
achieved allowing withdrawal of immunosuppressant medications. direction appears to result in lower risk of relapse- and nonrelapse-
Other significant complications of allo-HSCT relate to problems related mortality after allo-HSCT. Activating KIRs transduce
of inadequate reconstitution of the immune system of the patient signals to activate NK cells, and the presence of these cells is
and the concomitant risk of opportunistic infections, including associated with a lower risk of leukemia relapse after unrelated
viral and fungal infections that are rarely observed in healthy hosts. and haploidentical transplantation as well as protection against
Despite the apparent risks associated with allo-HSCT, this treat- certain viral infections, such as human immunodeficiency/
ment holds great promise as a curative therapy for several tumor acquired immunodeficiency syndrome (HIV/AIDS) or hepatitis
types, particularly with strategies either to enhance the efficacy C virus (HCV) infection. This effect appears to be more evident
of the GvT effect or decrease the toxicity of the graft-versus-host in haploidentical transplantation and in umbilical cord blood
(GvH) response. Efforts have been made in recent years to make (UCB) transplantation. The benefit of KIR–ligand mismatching
HSCT less toxic by development of low-dose, nonmyeloablative is less obvious (and more controversial) in unrelated transplanta-
conditioning regimens that allow the treatment of older patients tions and after transplantation with T cell–replete grafts in which
or those with comorbid health issues that otherwise preclude the antitumor effects of NK cells may be obscured by GvH
treatment with high doses of chemotherapy. 1 reactions mediated by T cells.
IMMUNE MECHANISMS RELATED TO ALLO-HSCT Graft-Versus-Host Disease
GvHD is caused by mature donor T cells contaminating the
Histocompatibility HSC inoculum, which recognize HLA or miHA differences
The HLA major histocompatibility complex (MHC; Chapter 5) expressed by host antigen-presenting cells (APCs) and tissues
4
is the primary consideration in the selection of a donor for allo- (Fig. 83.1). Cytokines released from host cells after a patient
HSCT, since its loci contribute significantly to host-versus-graft has received dose-intensive tumor cytoreductive chemotherapy
(HvG), leading to immunological rejection of donor HSC, and or radiotherapy conditioning create an inflammatory environment
to GvH (leading to GvHD and GvT) reactions. HLA antigens that enables the generation of a response of infused donor T
are classified as class I (HLA-A, -B, -C) and class II (HLA-DR, cells against host antigens. This initiates a cascade of T-cell
-DQ, -DP) molecules, and typing of donors and patients can activation events, which results in proliferation, release of
be performed using low- or high-resolution techniques. The additional inflammatory cytokines, and the generation of effector
low-resolution serological techniques can determine a phe- T cells that can infiltrate target tissue, particularly the lymphoid
notypic mismatch (e.g., A02 vs A03), whereas high-resolution system, intestinal tract, skin, and liver and mediate the destruction
molecular techniques can identify allelic genotypic differences of host cells in those organs. Both CD4 and CD8 T cells can be

CHAPTER 83 Hematopoietic Stem Cell Transplantation for Malignant Diseases 1131



KEY CONCEPTS
Irradiation Chemotherapy DC Donor cells
Graft-Versus-Host Disease (GvHD)
DC/Mø
• Caused by donor–recipient differences in:
IL-12 • Major histocompatibility complex (MHC) molecules
• Minor histocompatibility antigens (miHAs)
• Mediated by mature donor CD4 and/or CD8 T cells
• Requires inflammatory cytokines
TNFα, IL-1
CD4 NK • Primary target organs include lymphoid system, skin, gastrointestinal
tract, and liver
Upregulates adhesion • Acute and chronic forms
molecules on endothelium Activation and CD8
and chemokines proliferation
IFNγ, TNFα
IL-2 may also be additional tumor-specific or hematopoietic tissue
5
Effector T cells Mø LPS specific T cells. Thus the overriding goal is to be able to manipu-
late the donor HSC inoculum in such a way as to avoid GvHD
but to still be able to mediate a GvT effect. 6

GVHD tissue damage TNFα Clinical Aspects of aGvHD
lymphoid, skin, gut, liver Target cell apoptosis induction Usually developing within the first 3 months after transplantation,
FIG 83.1 Development of Graft-Versus-Host Disease (GvHD). aGvHD is a clinical diagnosis with characteristic but nondiag-
Cytoreductive preconditioning treatment of patients with nostic pathological findings, with the most common presenting
hematological malignancies with total body irradiation (TBI) or manifestations including skin rash; nausea, anorexia, and diarrhea;
7,8
chemotherapeutic drugs causes damage to epithelium in the skin and jaundice, depending on the target organ(s) most affected.
and gastrointestinal tract and activates the release of inflammatory In addition to the increased risks of developing aGvHD related to
cytokines by dendritic cells (DCs) and macrophages (Mϕ) in those the extent of HLA and miHA disparity, additional factors include
tissues. These cytokines include tumor necrosis factor-α(TNF-α) the advanced age of either the donor or the recipient, gender
and interleukin (IL)-1, which upregulate adhesion molecules and disparity (female donor–male recipient), donor parity (female
chemokine release in the vascular endothelium of the tissues. donors), and infusion of T cell–replete HSC products. Condition-
Activated DCs also migrate to the lymphoid system, where they ing with reduced-intensity regimens, with lower regimen-related
can present recipient histocompatibility antigens to infused donor toxicities to nonhematological tissues, also results in a lower risk
T cells that are in the hematopoietic stem cell (HSC) graft. DCs of aGvHD and may delay the onset of its initial manifestation.
release IL-12, which helps activate CD4 and CD8 T cells, as well Interactions between microbial-associated molecules and innate
as natural killer (NK) cells. These responding cells proliferate and immune receptors (e.g., Toll-like receptors [TLRs]) appear to
produce additional inflammatory cytokines, including interferon-γ be involved in GvHD pathogenesis, as demonstrated in both
(IFN-γ), TNF-α, and IL-2. Mϕ are activated by both IFN-γ and murine models and human transplantation. Research has shown
lipopolysaccharide (LPS) produced by bacteria found in the an interaction between the host’s gut microbiota and immune
9
intestinal tract, and these cells then produce high levels of more system. On the basis of this knowledge, experiments to decrease
TNF-α. TNF-α has many properties, including direct induction of transplantation-related GvHD by altering the gut microbiome
apoptosis on cells in the tissues of target organs of GvHD, but are being actively pursued.
it also helps effector T cells to home to and enter tissue sites Pharmacological agents are the mainstay of aGvHD prophy-
through the vascular endothelium. Effector T cells specific for laxis. Most patients receive a combination of a calcineurin
host histocompatibility antigens then get reactivated and perform inhibitor (tacrolimus or cyclosporine) along with an antime-
their effector functions, including release of inflammatory and tabolite, such as methotrexate or mycophenolate mofetil (MMF).
cytolytic cytokines and direct killing of recipient-type cells in the Methotrexate is associated with delayed engraftment, mucositis,
lymphoid compartment and in the skin, gut, and liver. idiopathic pneumonia syndrome, and other transplantation-
related complications, which has prompted the development of
other combination regimens, such as a calcineurin inhibitor in
combination with sirolimus or MMF, or reduced doses of
involved in GvHD, depending on the specific class I or class II methotrexate. The addition of antithymocyte globulin (ATG)
HLA or miHA disparities involved. to the conditioning regimen lowers the incidence of both GvH
The simplest way to avoid the development of GvHD is to and HvG reactions because of its persistence for several days
deplete the donor HSC graft of T cells before infusion to a cell after HSC infusion, effectively depleting T cells from the graft
5
dose below 10 cells/kg body weight. This approach has succeeded as well as the host. However, patients treated with ATG may face
in significantly diminishing the incidence of GvHD, but other higher risks of infectious complications, including Epstein-Barr
complications related to the ensuing delay in immune reconstitu- virus (EBV)–associated posttransplantation lymphoproliferative
tion of recipients, an increased risk of relapse from loss of the disorder as a result of the greater immunosuppression achieved.
GvT effect, and a higher rate of engraftment failure (also from Novel combinations of agents continue to be explored to reduce
loss of the GvH effect) have resulted in inconsistent improvement the cost and morbidity of aGvHD and its treatments.
in long-term survival, compared with T cell–replete products. The administration of high-dose cyclophosphamide, an
The conundrum is that the same alloreactive donor T cells that alkylating agent of the nitrogen mustard family, after transplanta-
mediate GvHD can also cause a GvT response, although there tion reduces the risks of acute and/or chronic GvHD and is a

1132 PART NINE Transplantation


prime example of drug-induced immunological tolerance, a bortezomib, or high doses of potent glucocorticoids before
concept first demonstrated by Schwartz and Dameshek in their collection and storage of autologous HSCs, resulting in a period
10
10
experiments using 6-mercaptopurine. Subsequently, in 1963, of immune dysregulation occurring early after autologous HSCT.
Berenbaum and Brown demonstrated immune tolerance to skin Patients with “engraftment syndrome” may require intensive
11
allografts in adult mice by using cyclophosphamide. Post- treatment with corticosteroids and a calcineurin inhibitor, with
transplantation cyclophosphamide given on days 3 and 4 after courses more similar to those experienced by patients undergoing
transplantation is now a standard regimen used in haploidentical allo-HSCT.
allogeneic transplantations in combination with other immu-
nosuppressive drugs and as a single agent in matched unrelated Clinical Aspects of cGvHD
and related transplantations, reducing the complications of graft cGvHD is the leading cause of late transplant-related mortality
failure and life-threatening aGvHD associated with transplantation (TRM) among those undergoing allo-HSCT and resembles
of HLA disparate grafts. 12 autoimmune disorders, such as scleroderma, Sjögren syndrome,
Glucocorticoids with a calcineurin inhibitor remain the and primary biliary cirrhosis. Diagnosis is, as with aGvHD, based
standard approach to initial systemic management of clinically on clinical observations and secondary confirmation with labora-
significant aGvHD. About 30–50% of patients will respond to tory or pathology tests (Table 83.2). A falling performance status,
initial therapy, and patients who fail to respond have a poor progressive weight loss, or recurrent infections are usually signs
prognosis, as additional agents added for control greatly increase of severe cGvHD. About 50% of long-term survivors will develop
the risk of opportunistic infections and other treatment-related cGvHD at a median of 9 months after transplantation, and
complications. The use of higher doses of corticosteroids, or the patients must be monitored closely for this complication for at
addition of ATG, for example, in the initial treatment of aGvHD least 3 years so that appropriate treatment can be initiated before
do not improve patient outcomes and should be reserved for extensive end-organ damage ensues. Permanent ocular or oral
patients who fail initial therapy. A number of drugs, including sicca syndrome and pulmonary dysfunction can result if cGvHD
ATG, pentostatin, switching to tacrolimus from cyclosporine, and is not appropriately treated, leading to a marked deterioration
newer monoclonal antibodies (mAbs) have shown limited activity in quality of life.
in the salvage treatment of patients with steroid-refractory aGvHD. Factors predictive of the development of cGvHD include the
Extracorporeal exposure of peripheral blood mononuclear cells degree of HLA and miHA disparity, as well as prior aGvHD,
(PBMCs) to the photosensitizing agent 8-methoxypsoralen and older patient age, the source of HSCs (greater risk after peripheral
ultraviolet A (UVA) radiation (photopheresis) is effective in the blood stem cell [PBSC] transplantation than after bone marrow
treatment of selected diseases mediated by T cells, including transplantation), gender (female donor–male recipient), and
both aGvHD and cGvHD, although the mechanism of this effect donor lymphocyte infusion (DLI) after transplantation. Inflam-
remains to be elucidated. matory events, such as sunburn or surgical procedures, can
precipitate cGvHD. Patients who develop cGvHD have a higher
Autologous GvHD risk of TRM but a lower risk of relapse as a result of the immu-
15
A form of aGvHD can also occur after auto-HSCT, probably as a nological GvT effect. T-cell depletion of the graft or treatment
manifestation of immune system dysregulation during reconstitu- with ATG may decrease the risk of cGvHD, although this has
tion of the immune system after dose-intensive therapy. Initially not been demonstrated in all studies. Most patients require at
studied in murine models of auto-HSCT, it was demonstrated least two drugs for effective treatment of cGvHD, with the
that the abrupt withdrawal of cyclosporine could induce clinical standard initial treatment being glucocorticoids and a calcineurin
features nondistinguishable from that observed after allo-HSCT. inhibitor. About half the patients do not achieve a complete
A mechanism that involves the depletion of central memory cells remission with first-line therapy, although the manifold signs
has been proposed, although depletion of Tregs with inhibition and symptoms of cGvHD complicate the definition of response
of peripheral tolerance may also be involved. Autologous GvHD to treatment. There are no clear recommendations regarding
is also reported in patients not receiving posttransplantation second-line treatments and various pharmacological and immu-
immunomanipulations (spontaneous GvHD). Based on the clini- nological techniques have been used. Photopheresis has an overall
cal benefits of GvT observed in allo-HSCT, several clinical trials response rate of 50–60%, with many patients achieving complete
with administration of cyclosporine with or without interferon remission. At least transient responses can be achieved with
(IFN) were performed in patients undergoing auto-HSCT without rituximab treatment, an anti-CD20 chimeric antibody, which
13
obvious GvT effect. Both induced and spontaneous autologous illustrates the humoral immunity contribution to cGvHD. The
aGvHD are usually self-limiting complications, and are usually demonstration of activation of the Janus kinase (JAK) pathways
easily managed with a course of corticosteroids in patients with in activated T lymphocytes has led to studies demonstrating a
more extensive involvement, in contrast to the more extensive potential benefit of Jak2 inhibitors in the management of steroid-
and difficult-to-manage aGvHD that occurs after allo-HSCT. refractory cGvHD.
Furthermore, cGvHD does not occur after auto-HSCT.
A spontaneously occurring severe, and sometimes fatal, reac- GvT Responses
tion involving the gastrointestinal (GI) system with severe diarrhea Although recognized in mouse models of transplantation in the
during the periengraftment period is reported in patients 1950s, the first clinical report of a relationship between GvHD
16
undergoing auto-HSCT for MM and other malignancies (“engraft- and GvT was published in 1979. This relationship between the
14
ment syndrome”). The presentation is clearly distinct from the incidence (but not the severity) of aGvHD or cGvHD and the
spontaneous or induced GvHD previously observed in patients relapse rate of chronic myeloid leukemia (CML) and, to a lesser
undergoing auto-HSCT. “Engraftment syndrome” may be a extent, acute myeloid leukemia (AML), acute lymphocytic
consequence of prior treatments with immunomodulatory drugs, leukemia (ALL), and MM has been observed in patients who
such as lenalidomide or thalidomide, the proteasome inhibitor received allo-HSCT. Relapses after HSCT are thought to occur

CHAPTER 83 Hematopoietic Stem Cell Transplantation for Malignant Diseases 1133



TABLE 83.2 Staging of Chronic Graft-Versus-Host Disease (GvHD)
Target Organ Score 0 Score 1 Score 2 Score 3
Performance score KPS 100% KPS 80–90% KPS 60–70% KPS <60%
Skin No symptoms <18% BSA 19–50% or sclerotic, still able to >50% or “Hidebound”
pinch
Mouth No symptoms Mild symptoms, no Moderate symptoms, decreased Severe symptoms with
limitations oral intake major decrease in intake
Eyes No symptoms Mild dry eyes Moderate dry eyes, drops >3×/day Severe dry eyes affecting
daily activities
Gastrointestinal tract No symptoms Symptoms without Symptoms with moderate weight Symptoms with weight loss
weight loss loss (5–15%) >15%
Liver Normal LFTs LFTs elevated <2× upper LFTs elevated 2–5× upper limits LFTs elevated >5× upper
limits of normal of normal limits of normal
Lungs No symptoms Mild symptoms Moderate symptoms Severe symptoms
FEV 60–79% FEV 40–59% FEV <40%
Joints and fascia No symptoms Mild tightness not Tightness affecting daily activities Contractures with significant
affecting daily activities loss of range of motion
Female genital tract No symptoms Symptomatic with middle Symptomatic with dyspareunia Symptomatic with strictures
signs on examination
At least one diagnostic and one distinctive sign is necessary to make a diagnosis of cGvHD.
BSA, Body Surface Area; FEV, Forced Expiratory Volume; KPS, Karnofsky Performance Status; LFT, Liver Function Test.
Adapted from: Filipovitch AH, Weisdorf D, Pavletic S, et al. National Institutes of Health Consensus Development Project on Criteria for Clinical Trials in Chronic Graft-versus-Host
Disease: 1. Diagnosis and Staging Working Group Report. Biol Blood Marrow Transplant 2005; 11: 945–56.


CLINICAL PEARLS GvHD GvT
Factors Predicting Chronic Graft-Versus-Host
Disease (GvHD)
• Degree of human leukocyte antigen (HLA) incompatibility Target tissue- Shared alloantigens: Bcr-abl
• Major histocompatibility complex (MHC) restricted miHA MHC, miHA hematopoietic-restricted
• Minor histocompatibility antigens (miHAs) disparity miHA proteinase-3 c-akt
• Presence of prior acute GvHD
• Source of stem cells (higher risk in peripheral blood versus bone
marrow)
• Donor gender (female donor → male recipient)
• Use of donor lymphocyte infusion (DLI) following hematopoietic stem
cell transplantation (HSCT)
• Inflammatory events; surgeries, phototoxicity, alcohol consumption FIG 83.2 Graft-Versus-Host Disease (GvHD) and Graft-Versus-
Tumor (GvT) Responses. Donor T cell responses to recipient
antigens can cause GvHD but can also target residual leukemia
because of survival of malignant cells harbored in bone marrow cells. T cells causing GvHD may recognize ubiquitous or tissue-
after administration of the pretransplantation conditioning restricted antigens (either major histocompatibility complex [MHC]
regimen and their outgrowth several months later. Relapse remains or minor histocompatibility antigens [miHAs]). Many of these
the major cause of treatment failure after allo-HSCT despite the recipient antigens may also be expressed by the leukemia cells
intended GvT effect of this treatment modality. The ability to and allows for a GvT response. Additional leukemia-specific (e.g.,
mediate an effective GvT response likely depends on several bcr-abl, proteinase 3, or c-akt) or tissue-restricted antigens (some
factors, including the presentation of appropriate antigens by miHAs, as those expressed only by certain lineages of hematologi-
MHC class I and/or class II molecules on the tumor cells that cal cells) may be dominantly expressed by the tumor cells and
can be recognized by effector CD4 or CD8 T cells; lack of strong can be targeted by donor T cells without causing GvHD.
Treg activity that may be induced by cytokines from the tumor
cells; tumor cell susceptibility to lysis by effector T cells (e.g.,
the level of B-cell lymphoma 2 (BCL-2) expression and the ability Immunotherapy remains the major strategy to combat relapse
to resist apoptosis induction); ability of T cells to home to sites occurring after allo-HSCT. This can be achieved by reducing
of tumor growth; and the direct effect of immunosuppressive immunosuppression, offering a second allo-HSCT, or infusing
cytokines, such as transforming growth factor-β (TGF-β), additional lymphocytes from the HSC donor (DLI). As one can
17
produced by the tumor cells. Many types of tumor cells expect, all these modalities pose the challenge of associated
6
downregulate expression of MHC on their surface, and perhaps morbidities, particularly GvHD. More defined immunothera-
CML and AML are most susceptible to GvT responses because peutic approaches using chimeric antigen receptors (CARs)
the myeloid lineage is adapted for antigen presentation and high expressed on T cells (CAR T cells; Chapter 77) also show some
MHC expression. A number of novel immunotherapeutic promise to target residual tumor cells that remain after allo-HSCT,
approaches are being developed to overcome these obstacles and with a possible low risk of GvHD. However, recent studies in
enhance GvT responses, keeping in mind that GvHD has to also murine models have raised a cautionary note concerning the
be avoided or minimized to improve outcomes (Fig. 83.2). capacity of CAR T cells to inflame GvHD. 18

1134 PART NINE Transplantation



KEY CONCEPTS with aggressive lymphomas, ALL, MM, and CLL, resulting in long-
25
Major Issues Related to Success of Allogeneic lasting remissions following heavy pretreatment. Developed in
the late 1980s and requiring decades of fine-tuning and extensive
Hematopoietic Stem Cell Transplantation clinical trials, CAR T cells have the ability to recognize and target
• Graft-versus-host disease (GvHD) tumor cells via their reprogrammed T-cell receptor (TCR) toward
• Graft-versus-tumor (GvT) responses the malignant cell and costimulatory molecules engineered into
• Kinetics and completeness of immune reconstitution the patient’s own T cells. This form of treatment engaging a
• Opportunistic infections from delayed immune reconstitution combination of gene therapy, cell therapy, and immunotherapy has
• Patient comorbid conditions increasing risk of toxicities led to unprecedented results in patients without robust treatment
• Chemotherapy sensitivity of disease being treated options for their aggressive malignancies or following relapse
after allo-HSCT. The primary limitation of CAR T–cell therapy is
Immunomodulation is frequently the first treatment option the need to identify tumor-specific antigens to avoid the serious
for patients in relapse after allo-HSCT, with rapid withdrawal toxicity of “on target” but “off tumor” response.
of immunosuppressive medications and infusion of donor T Another potential approach to augment GvT responses fol-
6
cells. This treatment approach requires that donor T-cell chi- lowing HSCT to prevent or treat relapse is the use of checkpoint
merism be sustained to avoid rejection of effector cells through inhibitors, currently accomplished by mAb blockade of negative
an HvG mechanism. Patients with low-grade lymphoid malignan- regulatory signals from either cytotoxic T lymphocyte antigen-4
cies, such as CLL, indolent non-Hodgkin lymphoma (NHL), (CTLA4) or programmed death (PD)-1 proteins on the surface
mantle cell NHL, and CML have the highest likelihood of response. of activated T cells upon interaction with their respective ligands,
A survey of 25 transplantation programs identified 140 patients B7-2 (CD86) and PD-L1 (B7-H1), on APCs and tumor cells
who had received DLI and reported a complete response rate (Chapter 77). Checkpoint inhibitors show strong clinical efficacy
of 60% in patients with CML. Response rates were higher in in the treatment of relapsed hematological malignancies, such
patients with cytogenetic and chronic-phase relapse (75.7%) as CLL and Hodgkin lymphoma. 26,27 In a phase I dose-escalation
than in patients with accelerated-phase (33.3%) or blastic-phase study using the anti-CTLA4 mAb ipilimumab for relapse after
(16.7%) relapse, and almost 90% of these patients remained in allo-HSCT, Bashey et al. concluded that ipilimumab did not
19
remission at 2 years after treatment. The activity of DLI in induce or exacerbate clinical GvHD but could cause organ-specific
AML and ALL is not as robust in comparison to that of in CML. immune adverse events and regression of malignancy following
However, it remains the main treatment strategy to treat relapse allo-HSCT. 28
following allo-HSCT. DLI can be used in combination with
chemotherapy to maintain control of disease control during the Adjuvant Therapy With HSCT
time required for the development of the GvT effect. The relatively higher incidence of relapse after high-dose therapy
The number of lymphocytes infused is important in achieving leads to the hypothesis that chemoablation therapy with auto-
the DLI effect, although it may be possible to induce GvT using HSCT, curative for some patients, could be viewed as a platform
doses of lymphocytes that are less likely to result in GvHD. A for other approaches effective in eliminating the minimal residual
large retrospective analysis demonstrated that using an initially disease (MRD) in patients destined to experience relapse.
20
lower cell dose reduced GvHD and improved survival. One Additional or yet higher-dose chemotherapy or radiotherapy,
method to reduce GvHD is to genetically insert suicide genes unless directly targeted to the tumor, increases the risk of
into the T cells being given for DLI, allowing specific ablation nonhematopoietic toxicity and TRM from causes other than
21
of these cells if this complication occurs. Another method to relapse. The correlation of more rapid lymphocyte recovery with
lower GvHD rates complicating DLI is to deplete the DLI product a decreased risk of relapse, although likely a reflection of host
of GvHD-inducing cells. Studies in mice have shown that naïve factors and not direct evidence of a GvT effect after auto-HSCT,
subset of CD8 T cells lead to more GvHD, whereas the effector supports attempts to use HSCT as a tumor-debulking platform
memory subsets (T EM ) of CD8 and CD4 T cells moderate the for posttransplantation immunotherapies. Immunotherapies are
graft-versus-leukemia (GvL) effect without causing GvHD. 22,23 of interest in this regard and include administration of post-
The delay in response between DLI and the development of a transplantation cytokines; the addition of tumor-specific antibod-
GvT effect suggests that only a minority of the cells infused ies, used before and/or after HSCT as an “in vivo purge”; and
recognize the tumor cell antigens and must undergo in vivo the development of tumor-specific vaccines, such as with tumor
expansion before the therapeutic effect is achieved. It may be antigen–pulsed dendritic cells (DCs). The immunoglobulin (Ig)
possible to develop leukemia-specific cytotoxic T cells in the idiotype found in most patients with MM and indolent NHL,
laboratory, decreasing the delay in effect and possibly increasing which is unique to the malignant clone, may serve as a unique
the GvT potential. Donor immunity can be transferred, at least tumor-associated antigen for DC pulsing. The treatment appeared
29
transiently, to the host, as demonstrated by delayed-type transfu- to be well tolerated, and after vaccination, idiotype-specific
sion reactions to host red blood cells (RBCs), mediated by donor responses have been observed, characterized by T cell–proliferative
24
lymphocytes transfused along with the HSC product. Adoptive responses with cytokine release and the production of antiidiotype
transfer of donor immunity against specific targets can be antibodies, although, again, a clinical benefit remains elusive.
achieved, but its persistence requires immunization of both the
donor and the recipient. A clinical utility for such manipulation CLINICAL HSCT
of the immune system has not yet been demonstrated, but it is
clearly of interest as a technique to prevent posttransplantation Sources of HSCs
infections and/or disease relapse. Bone marrow, peripheral blood stem cells (PBSCs), or UCB
A rapid and major leap in the field of adoptive transfer of are all appropriate sources of HSCs for transplantation, with
immunity is the development of CAR T–cell therapies for patients the primary differences being the quantities of HSCs in each

CHAPTER 83 Hematopoietic Stem Cell Transplantation for Malignant Diseases 1135


product and the quantities and functions of other blood cells Influence of graft, donor, and host factors on allogeneic HSC engraftment
comprising most of the cellular content of the HSC product that
may influence the immunological outcomes of transplantation. Greater Lesser
PBSCs have virtually replaced bone marrow as the HSC source for HSC dose
auto-HSCT and are widely used for allo-HSCT. Transplantation
of PBSCs achieves more rapid hematological recovery in both the
auto- and allo-HSCT settings, resulting in lower complication T-cell dose
rates and lower costs of treatment. PBSCs appear to improve
overall survival and disease-free survival of patients with advanced
hematological malignancies who undergo allo-HSCT, although Engraftment HLA-compatibility Rejection
30
at the cost of an increased risk of extensive cGvHD. Use of
PBSCs results in a decreased CD4 T-helper cell-1 (Th1)/Th2 ratio, Pretransplant
which may adversely affect the ability to counteract infections conditioning
and may favor cGvHD development. It is also interesting to note Posttransplant
that PBSC products tend to contain about 10 times more T cells immunosuppression
than bone marrow products, yet those T cells are less functional,
possibly because of inhibition by granulocytes activated by the FIG 83.3 Effect of Donor and Graft Characteristics on Allo-
granulocyte–colony-stimulating factor (G-CSF) used for HSC geneic Hematopoietic Stem Cell Transplantation (Allo-HSCT).
mobilization. 31 Hematological engraftment requires that the host-versus-graft
UCB is a rich source of HSCs, with the major limitation for (HvG) reaction be overwhelmed, either by administration of a
clinical use being the small quantity of cells collected, resulting more intensive conditioning regimen before transplantation, use
in slower recovery of hematological function and increased risk of more immunosuppressive medications after transplantation,
of failure of sustained engraftment. A very significant advantage closer matching of donor with host, and infusion of donor
is the lower risk of GvHD because of the relative immaturity lymphocytes that can affect development of GvHD.
of the donor immune system, allowing for the use of HLA-
mismatched units without a prohibitory increase in the risk of KEY CONCEPTS
GvHD. In general, as with other sources of HSCs, outcomes of
UCB transplantation reflect patient characteristics, with lower Selection of Hematopoietic Stem Cell (HSC)
survival probabilities for patients with advanced diseases or poorer Products for Transplantation
performance status at time of transplantation. A number of Bone Marrow
investigators proposed combining units of UCB to increase the • Adequate quantities of cells can be obtained from most patients and
cell dose infused into the patient, possibly increasing the speed donors
of hematological recovery, but also (in theory at least, but not • Cytokine administration before harvesting may increase the quantities
shown yet in clinical studies) potentiating the GvT effect and of HSCs collected and the number and function of accessory cells
reducing the risk of relapse. Ex vivo expansion of one or more affecting transplant outcomes
UCB units is also being studied as a mechanism to achieve more • Lower risk of chronic graft-versus-host disease (GvHD) than peripheral
rapid hematological recovery. blood stem cells (PBSCs)
Cell dose is an important predictor of outcome for both PBSCs
auto-HSCT and allo-HSCT, and HSCs comprise a very small • Faster hematological recovery than with bone marrow or umbilical
portion (generally <1%) of the marrow, PBSC, or UCB product. cord blood (UCB)
It is now recognized that successful establishment of donor cell • Better survival after related-donor hematopoietic stem cell transplanta-
chimerism after allo-HSCT is a complex interplay of pre- and tion (HSCT) for patients with advanced malignancy
posttransplantation suppression of the host immune system, • Lower tumor cell contamination than with bone marrow collected
dose of HSCs and accessory cells (including donor lymphocytes) from autologous patients
contained in the graft, and donor HLA compatibility (Fig. 83.3). • Cytokine administration before harvesting may increase the quantities
of hematopoietic stem cells (HSCs) collected and the number and
HLA-mismatching, T-cell depletion, and less intensive pretrans- function of accessory cells affecting transplantation outcomes
plantation conditioning regimens all raise the risk of graft failure.
Importantly, the duration of aplasia predicts the incidence of UCB
32
TRM after auto-HSCT or allo-HSCT. Auto-HSCT has a neg- • Relative immaturity of donor immune system permits multiple-antigen
ligible risk of engraftment failure if the viability of HSCs is mismatched transplantation
maintained during processing and storage. The speed of hema- • Transplantation outcomes similar to mismatched unrelated bone marrow
tological recovery is related to the quantity of HSCs reinfused transplantation
+
33
in an exponential relationship. Increasingly higher CD34 cell • Slower hematological recovery than with either PBSC or bone marrow
• Availability of stored units facilitates transplantation for patients with
(a surrogate marker of immature HSCs) doses result in greater immediate need of treatment
likelihood of rapid recovery of PBSC counts. At lower doses,
there is considerable heterogeneity in engraftment speed, especially
for platelet recovery, with some patients experiencing quick and disease-free survival significantly favored patients receiving
34
engraftment despite low doses of PBSCs. Products containing marrow or PBSC grafts with higher cell doses, but exceeding
+
+
6
6
≥2-3 × 10 CD34 cells/kg recipient weight have more consistent a CD34 cell dose of about 10 × 10 /kg results in lower survival,
rapid granulocyte and platelet engraftment. likely as a result of higher risk of chronic GvHD. 35
Similarly, cell dose is a predictor of outcome for patients HSC products are not uniform in their characteristics, and
+
undergoing allo-HSCT. The actuarial 5-year TRM, overall survival, the relative contributions of CD34 cell dose and the doses of

1136 PART NINE Transplantation


accessory cells, including CD4, CD8, and NK-cell populations, demonstrated through evaluation of single nucleotide tandem
on the outcome of transplantation remain unconfirmed. Whether repeats (short tandem repeats [STRs]) by using molecular analysis.
or not this is clinically significant, the ratio and quantity of cell Obviously, such studies are of no value in assessing engraftment
populations collected will be affected by changes in the cytokines after auto-(syngeneic)HSCT.
and chemokines, used singly or in combination, to mobilize Much of the emphasis in transplantation has been on myeloid
HSCs into peripheral blood for collection or to alter bone marrow engraftment because initial patient survival depends on recovery
products collected from patients and donors. of phagocytes and, to a lesser extent, platelets. Immune reconstitu-
tion, and in particular donor T-cell reconstitution, in patients
Purging of Cell Populations receiving HSCT is often hampered by older recipient age, dimin-
For auto-HSCT, postcollection attempts to purge tumor cells ished functional status of the thymus, cytokine milieu at the time
potentially contaminating the graft have not demonstrated a of transplantation, and posttransplantation immunosuppressive
survival advantage for the transplant recipient. T-cell depletion treatments. The thymus involutes rapidly after childhood, and in
of donor HSCs by various methods is an effective means of the older adult, it is only able to contribute a very small portion
reducing the incidence of aGvHD or cGvHD; but it increases to the mature T-cell compartment. The thymic tissue may be
the risk of graft failure, opportunistic infections, and relapse, damaged as a result of a myeloablative conditioning regimen,
effectively nullifying any advantage. Higher doses of HSCs, which or it can also be a target of alloreactive donor T cells mediating
can be achieved by the use of large quantities of PBSCs or by GvHD. As a result, restoration of the T-cell compartment in
combining marrow and PBSC components, will offset this risk patients is often slow, particularly for CD4 T cells, and may
of graft failure. Partial T-cell depletion, as opposed to maximal be at suboptimal levels for many months to over a year. This
T-cell depletion, may also reduce the risk of graft failure. Some situation, of course, endangers the ability of the patient to stave
centers are exploring posttransplantation T-cell add-back after off opportunistic infections, such as from the herpes family of
the acute inflammatory effects of the conditioning regimen have viruses and fungal pathogens. If donor T cells are provided in
resolved in an effort to maintain the GvT effect while still reducing the HSC inoculum, some reconstitution of the T-cell repertoire,
GvHD morbidity and mortality. mostly CD8 T cells, is provided by the mechanism of nonthymic
homeostatic expansion, although the level of diversity may be
Expansion of HSC Products limited. Experimentally, administration of cytokines, such as
Ex vivo expansion of UCB HSCs is one potential mechanism to interleukin-7 (IL-7), after HSCT, can enhance thymic function
offset the low cell dose and delayed hematological recovery after and help donor T-cell reconstitution. B-cell reconstitution, in
transplantation. However, no expansion technique has yet achieved contrast, is not that problematic in terms of the regeneration of
this goal, and such endeavors are complicated by the difficulty the immune repertoire, although the ability to actually respond
in identifying the pluripotent HSCs in comparison with lineage- effectively to an infection with antibody production may still
committed progenitor cells. Furthermore, it is likely that expansion depend on the availability of antigen-specific CD4 T cells.
techniques will result in T-cell depletion, resulting in the other Administration of Ig to patients with low IgG levels can prevent
complications described above. some of the infectious complications. Patients receiving HSCT
who are conditioned with myeloablative regimens usually attain
Hematological Recovery high levels of donor chimerism in their lymphoid compartment
HSC engraftment encompasses two concepts: (i) recovery of within a few months of transplantation. This often correlates
hematopoietic and immunological function and (ii) the rate at with the ability of alloreactive donor T cells, capable of mediating
which this recovery occurs. Delay in or failure of sustained GvHD, to target residual recipient HSC elements so that the
engraftment after myeloablative-conditioning regimen administra- primary source of de novo lymphoid reconstitution will be from
tion greatly increases treatment morbidity and cost. Engraftment the donor origin. By the same token, high donor chimerism is
failure can occur as a result of inadequate HSC quantity from also associated with a lower incidence of relapse of malignancy.
poor collection or loss in postcollection processing, inadequate
host support of the infused cells, posttransplantation events or CONDITIONING REGIMENS
medications, or HvG rejection (see Fig. 83.3). Engraftment failure
is a very rare complication of auto-HSCT and is most likely a Dose-Intensive and Reduced Intensity Chemotherapy
consequence of poor preservation of HSCs after collection. In The pretransplantation regimen is intended to accomplish two
allo-HSCT, the risk of engraftment failure is proportional to the goals: ablate the tumor and achieve adequate immunosuppression
donor HLA–miHA disparity, occurring more commonly in to allow donor engraftment. For auto-HSCT, only the dose-
unrelated donor transplants than in sibling donor transplants, sensitivity of the tumor being treated need be considered. Lower
and with HLA-mismatched transplants. Engraftment failure is dose, nonmyeloablative regimens are not used in auto-HSCT
also increased by T-cell depletion of marrow inoculum because because with such regimens, infusion of HSCs to reconstitute
of the loss of the GvH effect against residual host immune cells. marrow function would not be needed. Total body irradiation
Chimerism assessment is important in evaluating graft function (TBI) was initially used for conditioning of transplant recipients.
after allo-HSCT. A fall in PBSC counts could indicate HvG This modality achieves tumor cytotoxicity; treatment of sanctuary
rejection of the graft or early relapse after transplantation or sites of disease, such as the central nervous system (CNS) and
could result from GvHD or viral infection. Documentation of testes; and profound immunosuppression. TBI is usually combined
stable persistence of donor T cells in the recipient’s blood will in sequence with chemotherapy agents, such as cyclophosphamide
help discriminate between these possibilities. It is also important or etoposide. Busulfan-based regimens were developed as alterna-
that sustained lymphoid chimerism be demonstrated if DLI is tives to TBI for patients who had received prior dose-limiting
to be used in the treatment of disease relapse after transplantation. radiotherapy and to avoid the effects of TBI on growth and
The level of donor–host chimerism after allo-HSCT is best development in children. A review of several studies that compared

CHAPTER 83 Hematopoietic Stem Cell Transplantation for Malignant Diseases 1137


the use of busulfan and TBI found no statistically significant in the initial treatment of CML, will reduce the numbers of
difference in survival or disease-free survival for patients with patients requiring HSCT in the management of their disease.
36
CML or AML. Specific regimens are commonly used in the Organizations, such as American Society of Blood and Marrow
treatment of certain malignancies, such as dose-intensive mel- Transplantation, have published guidelines for treatment and
phalan in the treatment of MM and carmustine (BCNU)–contain- reviews of the efficacy of treatment, identifying areas requiring
ing regimens used in the treatment of lymphoma. additional research. 39,40
CLINICAL PEARLS Acute Myelogenous Leukemia
Conditioning Regimens for Hematopoietic Stem The primary clinical questions in HSCT in the treatment of
Cell Transplantation (HSCT) Transplantation AML concern patient selection and timing of treatment. Most
patients with AML will achieve remission with initial chemo-
Autologous HSCT therapy, but even with appropriate postremission consolidation,
• Allows dose-intensive therapy the majority of patients (≈65%) will relapse within 1–2 years.
• Allows use of marrow-toxic agents Older age, the presence of defined cytogenetic abnormalities,
• Regimens designed for optimal tumor cytotoxicity inability to achieve a complete remission (CR) with the initial
course of therapy, and history of a preceding marrow disorder
Allogeneic HSCT
• Must achieve adequate patient immunosuppression (reduce host- or receipt of prior chemotherapy (“secondary AML”) are predic-
versus-graft [HvG] reaction) to achieve engraftment tors for failure of nontransplantation therapy. Patients with these
• Allows but does not require dose-intensive therapy adverse risk factors may be offered HSCT in first remission in
• Allows but does not require marrow-toxic agents place of nontransplantation consolidation chemotherapy. Numer-
• Need not be tumor specific ous studies of patients entering their first remission compared
standard consolidation therapy with dose intensification with
The myeloablative conditioning regimens currently used have auto- or allo-HSCT. In general, auto-HSCT was not shown to
been tested in dose-escalation studies to achieve the maximal toler- be more effective than nontransplantation consolidation che-
ated doses in otherwise healthy patients. Nonmarrow toxicities, motherapy, whereas allo-HSCT had the lowest risk of relapse.
such as pneumonitis, mucositis, and hepatic venoocclusive disease, Despite the higher transplantation-related complications, in at
limit further dose escalation of standard TBI- or chemotherapy- least two major studies, patients assigned to allo-HSCT achieved
based regimens. New approaches include the addition of targeted a significantly better disease-free survival (DFS) compared with
therapies to the conditioning regimen, such as tumor-directed patients assigned to either chemotherapy or auto-HSCT. Allo-
mAbs or radioimmunoconjugates that will not increase the toxicity HSCT in first remission is particularly beneficial for patients
to other organs. Tandem transplantation with the combination of with adverse risk features, achieving about a 50–70% DFS, and
a dose-intensive regimen with auto-HSCT followed, after recovery is the treatment of choice for patients with adverse risk cytogenet-
from the immediate regimen-related toxicities, by allo-HSCT ics or leukemia that arises from prior chemotherapy or other
using a reduced-intensity regimen is a novel approach to combine marrow diseases.
the benefits of each transplantation modality.
Hematological malignancies are predominantly diseases Myelodysplastic Syndromes
of older adults. The potent GvT effect that is observed after Myelodysplastic syndrome (MDS) comprises a heterogeneous
allo-HSCT allows allograft recipients to be treated with lower- group of clonal hematological disorders characterized by expan-
dose nonmyeloablative regimens with the immunosuppressive sion of abnormal HSCs engendering variable degrees of cytopenia
properties of the regimen to reduce HvG reactions and facilitate and frequent evolution to AML. Currently, allo-HSCT is the
engraftment becoming more important than direct cancer cytotox- only modality of treatment that can achieve long-term control
icity. The primary requirement in developing a reduced-intensity of disease; auto-HSCT is not feasible because of the inability to
regimen is the need to achieve adequate immunosuppression collect normal HSCs from these patients. The best results are
to permit the development of hematopoietic chimerism, which seen in patients with earlier-stage disease, although for patients
became feasible with the development of the purine analogue with earlier stage MDS, a “watchful waiting” approach, with
family of drugs. A variety of regimens are available, including transplantation performed at the time of disease progression,
combinations of fludarabine with melphalan and fludarabine may be appropriate.
with busulfan. Among the least toxic are regimens that involve
37
a single fraction of TBI, based on the work by Storb et al., who Chronic Myelogenous Leukemia
proposed that the HvG reaction leading to HSC rejection and Allo-HSCT is an appropriate treatment for CML with long-term
the GvH reaction could both be modified by an appropriate survival rates >80% for younger patients undergoing related donor
immunosuppressive regimen administered after transplantation, transplantation within the first year after diagnosis. However,
allowing a reduction in the intensity of the pretransplantation inhibitors of the tyrosine kinase (TK) encoded by the Philadelphia
conditioning regimen. 37,38 chromosome have relegated transplantation to the treatment of
patients with advanced disease or the rare patient with CML not
HSCT FOR INDIVIDUAL DISEASES responsive to that targeted therapy. The probability of long-term
survival is lower for patients who undergo allo-HSCT more than 1
The treatment of diseases by auto- or allo-HSCT continues to year after diagnosis and for patients with more advanced diseases.
evolve as the understanding of the biology of these diseases The effect of prior therapy with a TK inhibitor on the outcome
becomes more clearly understood. Improvements in nontrans- of transplantation is not known. CML is highly responsive to
plantation treatments available to patients, such as the develop- the immunological GvT effect, and many patients in relapse
ment of targeted tyrosine kinase inhibitors, which are very effective after transplantation can be salvaged by the administration of

1138 PART NINE Transplantation


DLI. CML is a stem cell disease, and the collection of normal melphalan with auto-HSCT can achieve about a 40% complete
HSCs is unlikely for most patients, precluding auto-HSCT. Of response rate. Tandem auto-HSCT with two cycles of dose-
speculative interest is the collection of autologous cells after intensive therapy at 2- to 6-month intervals can improve progres-
cytogenetic remission is achieved with the use of a TK inhibitor, sive free survival and in some studies, overall survival. This
for subsequent use at the time of disease relapse. approach to treatment is still debated. Ultimately, most patients
experience relapse and succumb to the disease. Although auto-
Myeloproliferative Diseases HSCT is not considered curative for this disease, the median
MPDs are clonal stem cell diseases and include CML (discussed duration of time without treatment is 2–3 years, and ≈15% of
above), polycythemia vera, primary myelofibrosis, essential patients may not need treatment for progressive disease for 5
thrombocytosis, chronic myelomonocytic leukemia, and MPD years or longer, which greatly improves their quality of life.
not otherwise characterized. These disorders display overlapping Allo-HSCT with use of a reduced-intensity conditioning regimen
clinical features and may also show features more characteristic decreases the risk of TRM to <10%, but patients with advanced
of MDS, but they exhibit different natural histories and different disease are unlikely to respond to this treatment. The lower risk
therapeutic requirements. Allo-HSCT is effective in ablating the of complications with nonmyeloablative conditioning regimens
abnormal clone in these disorders. The timing of treatment is has led to studies of tandem auto-HSCT, using an intensive
important because of the frequently long natural history of these chemotherapy regimen to achieve tumor debulking, followed
diseases. Anemia, older age, and cytogenetic abnormalities all 2–6 months later by allo-HSCT using a reduced-intensity regimen
predict poor survival in primary myelofibrosis, suggesting that to achieve the GvT effect. This approach to the treatment of
patients with anemia and an abnormal karyotype should proceed MM is being studied in large multicenter trials, with some studies
to allo-HSCT. showing a survival advantage for recipients of allo-HSCT.
Acute Lymphoblastic Leukemia Non-Hodgkin Lymphoma
In contrast to treatment of the pediatric patient with ALL The lymphomas (Chapter 79) represent a diverse group of
(Chapter 78), only very few adults with ALL are cured with malignant diseases of B and T lymphocytes, comprising some
nontransplantation induction and consolidation regimens. As of the slowest- to the fastest-growing human malignancies, with
with AML, adverse risk features include certain cytogenetic a range of curability achieved by nontransplantation therapies.
changes and the inability to achieve remission with initial induc- As a group, the lymphomas exhibit a strong dose–response
tion chemotherapy. Allo-HSCT in first remission is clearly effective relationship to chemotherapy or radiotherapy, and the benefit
in the management of patients with defined adverse risk cyto- of dose-intensive treatment with auto-HSCT has been well
genetics, such as translocation involving chromosome 4 and 11, established. Auto-HSCT avoids the morbidity of allo-HSCT and
or 9 and 22 (Philadelphia chromosome). Recent randomized is the preferred approach for the majority of patients, with some
studies also demonstrate a survival advantage for patients who prominent exceptions discussed below. Popular chemotherapy
have intermediate risk of disease and are undergoing allo-HSCT, regimens include cyclophosphamide, BCNU, and etoposide (CBV)
especially since current salvage regimens for patients in relapse or BEAM (BCNU, etoposide, cytarabine, and melphalan).
have a very low likelihood of achieving a second remission. There However, no single chemotherapeutic or radiation-based regimen
is little evidence to support the effectiveness of auto-HSCT despite has emerged as a superior treatment.
its theoretical potential.
Low-Grade NHL
Chronic Lymphocytic Leukemia Low-grade NHL, in general, exhibits a variable and prolonged
Many patients and physicians are reluctant to support aggressive natural course, with many patients not requiring treatment until
treatment of CLL with allo-HSCT because of the frequently symptoms or organ toxicity appear. Therefore most of the experi-
indolent nature and long natural history of this disease, as well ence with HSCT has been in patients after initial relapse rather
as the advanced age of most patients at time of diagnosis. Yet than at the time of initial diagnosis. Few randomized studies
disease progression is inevitable, and the aggressiveness of this comparing auto-HSCT with nontransplantation therapies for
disease can be predicted by various features, including chromo- patients have been reported. A number of phase II and registry
some 17p and 11q deletions and detection of ZAP-70 mutation data have been published for auto-HSCT, and although response
and CD38 expression. Therefore HSCT should be considered a rates are high, a continuing pattern of relapse has been observed.
treatment choice, especially for younger patients or those with In contrast, patients who undergo allo-HSCT experience a higher
adverse risk features. probability of TRM but a lower risk of relapse after transplanta-
The exquisite sensitivity of the CLL cells to the GvT effect tion. Indolent B-cell NHL, similar to other low-grade diseases,
allows for the use of reduced-intensity regimens with lower risks such as CLL, appears to be very sensitive to the GvT effect of
of regimen-related mortality in these, generally, older patients. allo-HSCT, and using reduced-intensity regimens will result in
Durable control of disease can be achieved for about 50% of lower transplantation-related complications. The difference in
patients. The extensive infiltration of bone marrow by malignant relapse rates between auto-HSCT and allo-HSCT could result
lymphocytes and the lack of the GvT effect preclude auto-HSCT. from the possible reintroduction of lymphoma cells in the HSC
In the few clinical studies that have been reported, high CR product but is more likely from the lack of a GvT effect of
responses have been obtained with auto-HSCT, but relapses were auto-HSCT.
the frequent cause of treatment failure.
Aggressive NHL
Multiple Myeloma Auto-HSCT is the standard of care for patients with B-cell NHL
MM is a malignancy of plasma cells (Chapter 80), with a median in first “chemotherapy-sensitive” relapse. The success of this
patient age of 65 years at time of diagnosis. In MM, dose-intensive therapy reflects the extent and the responsiveness of the disease

CHAPTER 83 Hematopoietic Stem Cell Transplantation for Malignant Diseases 1139


to chemotherapy at the time of transplantation, with relapse HSCT or who are not candidates for auto-HSCT because of
the major cause of treatment failure. The Parma randomized aggressive, chemotherapy-refractory disease.
trial for high-dose therapy followed by auto-HSCT reported
a significantly higher event-free survival for patients treated Solid Tumors
with auto-HSCT than for the group receiving standard-dose Skin and colonic mucosa are primary targets of both acute and
treatment (46% vs 12%). Furthermore, it is notable that no cGvHD, and this would suggest that allo-HSCT would be effective
patients assigned to the conventional-dose salvage therapy therapy in the treatment of cancers of these organs. Yet allo-HSCT
could be rescued at the time of the second relapse with delayed is not effective in the control of these cancers, illustrating the
transplantation. The efficacy of auto-HSCT is diminished discrimination between target antigens of normal tissues of GvH,
for patients previously treated with rituximab, an anti–B-cell such as the colonic crypt cells, compared with antigens expressed
chimeric antibody, and who relapse within 12 months of by tumors derived from these tissues targeted by GvT. With the
41
initial therapy. Other circumstances, in which HSCT may be exception of allo-HSCT in the treatment of renal cell cancer, in
indicated, include HSCT for patients who respond slowly to which a GvT effect was seen to be clinically evident in some but
initial therapy, have high-risk disease, are resistant to initial not all studies, transplantation in the treatment of solid tumors,
40
therapy, or are in relapse with chemotherapy-insensitive disease. such as neuroblastoma, Wilm tumor, and germ cell tumors, is
Results for patients with truly chemotherapy-refractory disease limited to auto-HSCT with one or more cycles of dose-intensive
are, however, poor, and such patients should be considered for chemotherapy.
allo-HSCT.
Mantle cell lymphoma is known for its unremitting clinical FUTURE DIRECTIONS
course when treated conventionally and has proven relatively
resistant to dose-intensive treatment, especially when used for Advances in HLA typing, chemotherapy conditioning regimens,
management of relapsed disease. Mantle cell NHL appears to supportive care, and our understanding of the biology of the
be very sensitive to the GvT effect of allo-HSCT allowing treat- HvG and GvH reactions have greatly reduced the toxicity of
ment with reduced-intensity regimens. both auto-HSCT and allo-HSCT. Although the diseases treat-
Burkitt lymphoma, Burkitt-like lymphomas, and lymphoblastic able with HSCT are those characterized by dose sensitivity,
lymphomas are high-grade NHLs associated with relatively poor currently available dose-intensive conditioning regimens have
long-term survival rates. The role of both auto-HSCT and allo- been pushed to maximal tolerable doses. Newer approaches
HSCT in these disorders remains unclear. There are no convincing to the treatment of these malignancies will likely include the
risk models that identify clear indications for transplantation as addition of therapies that target the malignancy, such as radioim-
part of the initial therapy. The lack of an obvious GvT effect in munoconjugates that will add minimal toxicity to other organs.
these disorders suggests that auto-HSCT can provide a reasonable The availability of reduced-intensity regimens permits tandem
treatment. Transplantation registry data suggest that disease status transplantation, using dose-intensive therapy with auto-HSCT to
at the time of transplantation is the most important predictor achieve maximal tumor-cell debulking, followed, after recovery
42
of outcome in patients with high-grade disease. T-cell lym- from transplantation-related toxicities, by Allo-HSCT, using a
phomas are less common than B-cell lymphomas. There is no reduced-intensity regimen to achieve an immunological GvT
well-defined management strategy for these disorders, and effect. Advancements in the field of haploidentical HSCT have
treatment is based on the disease stage and immunopathological facilitated immediate availability of donors for patients with
grade. Disease control with T cell–type treatment regimens, high-risk malignancies when selection of unrelated donors is
compared with the more common B cell–type treatment regimens, limited by ethnicity and time constraints. Finally, the ability to
is lower, although the comparison may not be appropriate, as use HSCT as a platform for protein- or cellular-based vaccination
T-cell diseases present at more advanced stages compared with strategies is the subject of considerable interest.
B-cell diseases. The GvT effect may be more evident in T-cell
NHL, particularly for virus-associated NHL, and allo-HSCT is ON THE HORIZON
the preferred treatment for some patients.
Allogeneic Hematopoietic Stem Cell Transplantation (HSCT)
Hodgkin Lymphoma Development of donor selection algorithms
Many patients with Hodgkin lymphoma will achieve durable Strategies to maximize graft-versus-tumor (GvT) effect
Advent of haploidentical HSCT allows for easy donor availability
remissions with nontransplantation chemotherapy and/or radia- Refining haploidentical HSCT techniques for different diseases and use
tion therapy, and algorithms for staging and treatment of this of KIR testing for donor selection
disease are well defined. Dose-intensive therapy with auto-HSCT Combination conditioning regimens
is available to those patients who do not achieve a remission or Developing regimens with lower toxicities
who relapse after initial therapy. The outcome for patients whose Posttransplantation maintenance/treatment
remission lasted <1 year is dismal with standard dose second-line Posttransplantation maintenance/treatment for high risk of relapse after
HSCT
treatments, and approximately 40–50% of patients with Hodgkin Immune effector cell therapies
lymphoma who suffer a relapse within 1 year will achieve durable Platform for other treatments; chimeric antigen receptor (CAR) T–cell
remissions after auto-HSCT. This approach can also overcome therapy, checkpoint inhibitor blockade
drug resistance and lead to an overall survival rate of 34–50%
for patients with chemotherapy-refractory disease. Autologous HSCT
Allo-HSCT is not the first choice for the treatment of relapsed Tumor-specific conditioning regimens
Hodgkin lymphoma because of the higher transplantation-related Improve ratio of toxicity to tumor-cell cytotoxicity
complications despite the evidence of an effective GvT effect. Posttransplantation vaccination strategies
Checkpoint inhibitor blockade in clinical trials for high-risk malignancies
Allo-HSCT is offered to patients who suffer relapse after auto-

1140 PART NINE Transplantation


Please check your eBook at https://expertconsult.inkling.com/ 22. Anderson BE, McNiff J, Yan J, et al. Memory CD4+ T cells do not induce
for self-assessment questions. See inside cover for registration graft-versus-host disease. J Clin Invest 2003;112:101–8.
details. 23. Chen BJ, Deoliveira D, Cui X, et al. Inability of memory T cells to induce
graft-versus-host disease is a result of an abortive alloresponse. Blood
2007;109:3115–23.
REFERENCES 24. Rowley SD, Donato ML, Bhattacharyya P. Red blood cell incompatible
allogeneic hematopoietic progenitor cell transplantation. Bone Marrow
1. Sorror ML, Maris MB, Storer B, et al. Comparing morbidity and Transplant 2011;46:1167–85.
mortality of HLA-matched unrelated donor hematopoietic cell 25. Jackson HJ, Rafiq S, Brentjens RJ. Driving CAR T-cells forward. Nat Rev
transplantation after nonmyeloablative and myeloablative conditioning: Clin Oncol 2016;13:370–83.
influence of pretransplantation comorbidities. Blood 2004;104:961–8. 26. Bachireddy P, Burkhardt UE, Rajasagi M. Haematological malignancies: at
2. Pidala J, Lee SJ, Ahn KW, et al. Nonpermissive HLA-DPB1 mismatch the forefront of immunotherapeutic innovation. Nat Rev Cancer
increases mortality after myeloablative unrelated allogeneic hematopoietic 2015;15:201–15.
cell transplantation. Blood 2014;124:2596–606. 27. Ansell SM, Lesokhin AM, Borrello I, et al. PD-1 Blockade with nivolumab
3. Confer DL, Abress LK, Navarro W, et al. Selection of adult unrelated in relapsed or refractory Hodgkin’s lymphoma. N Engl J Med
hematopoietic stem cell donors: beyond HLA. Biol Blood Marrow 2015;372:311–19.
Transplant 2010;16:S8–S11. 28. Bashey A, Medina B, Corringham S, et al. CTLA4 blockade with
4. Socie G, Blazar BR. Acute graft-versus-host disease: from the bench to the ipilimumab to treat relapse of malignancy after allogeneic hematopoietic
bedside. Blood 2009;114:4327–36. cell transplantation. Blood 2009;113:1581–8.
5. Bleakley M, Riddel SR. Molecules and mechanisms of the graft-versus- 29. Leitch HA, Connors JM. Vaccine therapy for non-Hodgkin’s lymphoma
leukaemia effect. Nat Rev Cancer 2004;4:371–80. and other B-cell malignancies. Curr Opin Investig Drugs 2005;6:
6. Kolb H-J. Graft-versus-leukemia effects of transplantation and donor 597–604.
lymphocytes. Blood 2008;112:4371–83. 30. Anasetti C, Logan BR, Lee SJ, et al. Peripheral-blood stem cells versus
7. Glucksberg H, Storb R, Fefer A, et al. Clinical manifestations of bone marrow from unrelated donors. N Engl J Med 2012;367:1487–96.
graft-versus-host disease in human recipients of marrow from 31. Vasconcelos ZF, Santos BM, Costa ES, et al. T-lymphocyte function from
HLA-matched sibling donors. Transplantation 1974;18:295–304. peripheral blood stem-cell donors is inhibited by activated granulocytes.
8. Przepiorka D, Weisdorf D, Martin P, et al. 1994 Consensus conference on Cytotherapy 2003;5:336–45.
acute GvHD grading. Bone Marrow Transplant 1995;15:825–8. 32. Offner F, Schoch G, Fisher LD, et al. Mortality hazard functions as related
9. Mathewson ND, Jenq R, Mathew AV, et al. Gut microbiome-derived to neutropenia at different times after marrow transplantation. Blood
metabolites modulate intestinal epithelial cell damage and mitigate 1998;88:4058–62.
graft-versus-host disease. Nat Immunol 2016;17:505–13. 33. Rowley SD, Zuehlsdorf M, Braine HG, et al. CFU-GM content of bone
10. Schwartz R, Dameshek W. Drug-induced immunological tolerance. marrow graft correlates with time to hematologic reconstitution
Nature 1959;183:1682–3. following autologous bone marrow transplantation with
11. Berenbaum MC, Brown IN. Prolongation of homograft survival in mice 4-hydroperoxycyclophosphamide purged bone marrow. Blood
with single doses of cyclophosphamide. Nature 1963;200:84. 1987;70:271–5.
12. Robinson TM, O’Donnell PV, Fuchs EJ, et al. Haploidentical 34. Ringden O, Barrett AJ, Zhang MJ, et al. Decreased treatment failure
bone marrow and stem cell transplantation: experience with in recipients of HLA-identical bone marrow or peripheral blood
post-transplantation cyclophosphamide. Semin Hematol 2016;53:90–7. stem cell transplants with high CD34 cell doses. Br J Haematol
13. Yeager AM, Vogelsang GB, Jones RJ, et al. Induction of cutaneous 2003;121:874–85.
graft-versus-host disease by administration of cyclosporine to patients 35. Heimfeld S. HLA identical stem cell transplant: is there an optimal CD34
undergoing autologous bone marrow transplantation for acute myeloid cell dose? Bone Marrow Transplant 2003;31:839–45.
leukemia. Blood 1992;79:3031–5. 36. Socie G, Clift RA, Blaise D, et al. Busulfan plus cyclophosphamide
14. Keung YK, Beaty MW, Pettenati M, et al. Possible role of engraftment compared with total-body irradiation plus cyclophosphamide before
syndrome and autologous graft-versus-host disease in myelodysplastic marrow transplantation for myeloid leukemia: long-term follow-up of 4
syndrome after autologous stem cell transplantations: retrospective randomized studies. Blood 2001;98:3569–74.
analysis and review of the literature. Clin Lymphoma Myeloma Leuk 37. Storb R, Yu C, Wagner JL, et al. Stable mixed hematopoietic chimerism in
2010;10:129–33. DLA-identical littermate dogs given sublethal total body irradiation
15. Pidala J, Kim J, Anasetti C, et al. The global severity of chronic before and pharmacological immunosuppression after marrow
graft-versus-host disease, determined by National Institutes of Health transplantation. Blood 1997;89:3048–54.
consensus criteria, is associated with overall survival and non-relapse 38. McSweeney PA, Niederwieser D, Shizuru JA, et al. Hematopoietic cell
mortality. Haematologica 2011;96:1678–84. transplantation in older patients with hematologic malignancies:
16. Weiden PL, Flournoy N, Thomas ED, et al. Antileukemia effect of replacing high-dose cytotoxic therapy with graft-versus-tumor effects.
graft-versus-host disease in human recipients of allogeneic-marrow grafts. Blood 2001;97:3390–400.
N Engl J Med 1979;300:1068–73. 39. Oliansky DM, Gordon LI, King J, et al. The role of cytotoxic therapy with
17. van den Brink MRM, Porter DL, Giralt S, et al. Relapse after allogeneic hematopoietic stem cell transplantation in the treatment of follicular
hematopoietic cell therapy. Biol Blood Marrow Transplant lymphoma: an evidence-based review. Biol Blood Marrow Transplant
2010;16:S138–S145. 2010;16:443–68.
18. Jacoby E, Yang Y, Qin H, et al. Murine allogeneic CD19 CAR T cells 40. Oliansky DM, Antin JH, Bennett JM, et al. The role of cytotoxic therapy
harbor potent antileukemic activity but have the potential to mediate with hematopoietic stem cell transplantation in the therapy of
lethal GVHD. Blood 2016;127:1361–70. myelodysplastic syndromes: an evidence-based review. Biol Blood
19. Collins RH, Shpilberg O, Drobyski WR, et al. Donor leukocyte infusions Marrow Transplant 2009;15:137–72.
in 140 patients with relapsed malignancy after allogeneic bone marrow 41. Gisselbrecht C, Glass B, Mounier N, et al. Salvage regimens with
transplantation. J Clin Oncol 1997;15:433–44. autologous transplantation for relapsed large B-cell lymphoma in the
20. Guglielmi C, Arcese W, Dazzi F, et al. Donor lymphocyte infusion for rituximab era. J Clin Oncol 2010;28:4184–90.
relapsed chronic myelogenous leukemia: prognostic relevance of the 42. Sweetenham JW, Pearce R, Taghipour G, et al. Adult Burkitt’s and
initial cell dose. Blood 2002;100:397–405. Burkitt-like non-Hodgkin’s lymphoma – outcome for patients treated
21. Ciceri F, Bonini C, Stanghellini MT, et al. Infusion of suicide-gene- with high-dose therapy and autologous stem-cell transplantation in first
engineered donor lymphocytes after family haploidentical haematopoietic remission or at relapse: results from the European Group for Blood and
stem-cell transplantation for leukaemia (the TK007 trial): a Marrow Transplantation. J Clin Oncol 1996;14:2465–72.
non-randomised phase I-II study. Lancet Oncol 2009;10:489–500.

CHAPTER 83 Hematopoietic Stem Cell Transplantation for Malignant Diseases 1140.e1


MULTIPLE-CHOICE QUESTIONS

1. What is the objective of using a conditioning regimen for 3. Which of the following increases the risk of relapse after an
hematopoietic stem cell transplantation (HSCT)? allogeneic stem cell transplantation?
A. To reduce the bulk of malignant cells A. Human leukocyte antigen (HLA) mismatched donor
+
6
B. To decrease the risk of graft rejection B. CD34 cell dose less than 2 × 10 /kg patient weight
C. To improve the engraftment of donor hematopoietic stem C. Presence of acute or chronic GvHD
cells (HSCs) D. Disease burden prior to stem cell transplantation
D. All the above
4. Use of which strategy decreases the risk for acute and/or
2. What is the main mechanism of graft-versus-leukemia (GvL) chronic GvHD?
effect using donor lymphocyte infusion? A. Unrelated donor
A. Release of cytokine mediators generated by the graft-versus- B. Female donor for male recipient
host disease (GvHD) reaction initiated by donor lymphocyte C. Total body irradiation as part of the conditioning regimen
infusion (DLI) D. T-cell depletion of graft
B. Provide CD4 and CD8T cells that recognize target antigens
on tumor cells
C. Enhance inhibitory receptors (killer cell immunoglobulin-
like receptors [(KIRs]) recognized by donor natural killer
(NK) cells
D. Increase the T-cell receptor (TCR) repertoire diversity

84






Immunoglobulin Therapy:

Replacement and Immunomodulation



Mark C. Ballow







Over 70 years ago, a cold ethanol fraction of plasma that contained KEY CONCEPTS 1
an enriched fraction of gammaglobulin was used intramuscularly
as passive immunotherapy for the treatment and protection of Properties of Intravenous Immunoglobulin (IVIG)
infectious pathogens and subsequently as antibody replacement and Subcutaneous Immunoglobulin (SCIG)
therapy for patients with primary immune deficiency diseases • Plasma fractionation (first step) by cold ethanol / Cohn-Oncley modifica-
(PIDDs). This Cohn ethanol plasma fraction of immunoglobulin tion (fraction II)
G (IgG) remained the principal form of therapy until 1981 when • >98% IgG; >90% monomeric IgG
an intravenous (IV) preparation (i.e., intravenous immunoglobu- • Traces of other immunoglobulins (i.e., IgA and IgM) and serum
lin [IVIG]) became available. Subsequently, Imbach observed proteins
that thrombocytopenia resolved when patients with immune • Addition of sugar, and amino acids, stabilizes IgG from aggregation
deficiency were treated with IVIG. This observation led to the • Multiple viral inactivation/removal steps
• Cold ethanol fraction
use of IVIG in patients with autoimmune idiopathic thrombo- • Chromatography
cytopenic purpura, and an expansion on the use of IVIG as • Solvent/detergent treatment
immunomodulatory therapy in several FDA approved autoimmune • Caprylate fractionation
disorders (Table 84.1). Most IVIG preparations are derived from • Nanofiltration
plasma by the Cohn ethanol fractionation method or the Cohn- • Depth filtration
Oncley modification. IVIG products are modified to prevent the • Pasteurization
formation of IgG aggregates and to make IVIG suitable for the • Intact Fc receptor important for biological function
• Opsonization and phagocytosis
IV route. Excipients, such as sugars (maltose or D-sorbitol) or • Complement activation
amino acids (glycine or L-proline), stabilize the IgG molecules • Antibody dependent cytotoxicity
from aggregation. Cold ethanol fractionation as the first step in • Normal half-life comparable to serum IgG
plasma processing inactivates human immunodeficiency virus • Normal proportion of IgG subclasses
(HIV). Treatment with low pH, solvent and detergent, pasteuriza- • Broad spectrum of antibodies to bacterial and viral agents
tion or nanofiltration/depth filtration in combination, depending
on the Ig product, is used as further 5–7 steps for viral inactivation
and removal. Commercial lots of IVIG are derived from ≈15 000
donors (not to exceed 60 000 donors according to the U.S. Food and a comparison of low-dose with high-dose IVIG therapy.
3
and Drug Administration [FDA] and the Plasma Protein Thera- Quartier et al. performed a retrospective study of the clinical
peutics Association). Each lot must contain adequate levels of features and outcomes of 31 patients with X-linked agamma-
antibody to measles. These products may vary slightly among globulinemia receiving replacement IVIG therapy between 1982
manufacturers and from lot to lot, but they are generally com- and 1997. Although early treatment with IVIG and achieving a
parable with regard to clinical efficacy, but perhaps not tolerability. trough serum IgG level of >500 mg/dL was effective in preventing
The newer Ig formulations are isoosmolar, low-sodium, and severe acute bacterial infections, these levels did not prevent
low-IgA liquid products. The characteristics of IVIG preparations pulmonary disease, sinusitis, or meningoencephalitis. The authors
available in the United States are reviewed in Table 84.2. suggested that more intensive therapy to maintain a higher serum
In this chapter, the application of IVIG as replacement therapy IgG trough level (e.g., >800 mg/dL) may improve the pulmonary
in patients with primary immune deficiency (PID) and the outcome.
potential mechanisms of action of Ig therapy in the treatment Several studies have shown that pulmonary abnormalities
of autoimmune and inflammatory diseases are reviewed. For are the most important factors associated with morbidity and
more information on the use of Ig therapy in specific diseases, mortality in patients with PIDs. The number of infections, days
the reader is referred to an evidence-based review of the topic on antibiotic therapy, days missed from school or at work, and
elsewhere. 1,2 hospitalized days may not be sufficient indicators of adequate
treatment. Therefore the improvement or maintenance of
REPLACEMENT THERAPY WITH IVIG pulmonary function is an important measure of the success of
4
therapy. Orange et al. examined the impact of serum IgG trough
Several early trials in the 1980s have been conducted to examine levels on pneumonia incidence in patients with PIDDs receiving
the efficacy of IVIG with intramuscular (IM) gammaglobulin, replacement IVIG therapy in a meta-analysis of published clinical

1143

1144 ParT TEN Prevention and Therapy of Immunological Diseases



TABLE 84.1 FDa-approved Indications KEY CONCEPTS 2
for IVIG Treatment of Patients With Primary Immune
• Primary humoral immunodeficiency disease—as replacement Deficiency With Intravenous Immunoglobulin
therapy
• Immune thrombocytopenic purpura—to prevent severe bleeding • Starting dose—400–600 mg/kg every 4 weeks.
• Children with HIV/AIDS and recurrent infections—to prevent • Maintain trough level of >500 mg/dL or a serum immunoglobulin G
serious bacterial infections (IgG) level 450 mg/dL over baseline.
• B-cell chronic lymphocytic leukemia with recurrent infections and • Recent studies indicate the most patients may require a trough or
humoral immune deficiency—to prevent bacterial infection steady-state level of 750–850 mg/dL.
• Kawasaki disease—to prevent coronary artery aneurysms • The most important dose of intravenous immunoglobulin (IVIG) is
• Bone marrow transplantation—to decrease risk of infection, the dose that keeps the patient free of infections (i.e., biological
interstitial pneumonia, and graft-versus-host disease in the first 100 trough level).
days after transplantation • Higher trough levels (>800 mg/dL) may be necessary to prevent chronic
• Chronic inflammatory demyelinating polyneuropathy pulmonary changes and enteroviral meningoencephalitis.
• Multifocal motor neuropathy • Depending on the IgG catabolism and/or control over infections,
decrease infusion interval to every 3 weeks.
FDA, U.S. Food and Drug Administration; IVIG, intravenous immunoglobulin; • Subcutaneous immunoglobulin (SCIG) offers an advantage in achieving
HIV, human immunodeficiency virus; AIDS, acquired immunodeficiency syndrome. higher steady-state plasma IgG levels.
• It may take ≥3 months to achieve a steady state after a change in
dose.
studies. The results of their analysis showed that trough IgG • Monitor serum blood urea nitrogen (BUN) and creatinine and perform
levels increased 121 mg/dL for every 100 mg/kg increase in dose, liver function tests every 6–12 months.
which resulted in a decrease in pneumonia incidence by 27%. • Keep a log of dose, manufacturer, lot number, and reactions for each
There was a strong correlation between increasing trough IgG infusion.
5
levels and a decrease in pneumonia. Bonagura et al. discussed • For patients with rate-related adverse side effects. Consider pretreat-
ment with:
the value of “biological” trough levels for the serum IgG related • Acetaminophen
to the patient’s clinical course (i.e., frequency of infections) instead • Diphenhydramine
of arbitrarily choosing a trough value of 500 mg/dL. A study of • Nonsteroidal antiinflammatory drugs (NSAIDs)
90 patients with common variable immune deficiency (CVID) • Corticosteroids
with follow-up of 22 years showed that the dose of IVIG needed • Consider alternate administration of SCIG in patients with frequent
to keep a patient free of infection varied among patients, systemic adverse reactions.
emphasizing that the goal of replacement therapy should be
improvement of clinical outcome, not a specific IgG trough level. 6
The overall consensus among clinical immunologists is that cost and inconvenience, self-administration and home treatment
an IVIG dose of 400–600 mg/kg/month is a good starting point. have been used successfully. For home therapy, patients need to
(see Key Concepts 2) Catabolism of infused Ig varies among be selected carefully. Infusions should be done only in the presence
individuals. The rate of elimination of IgG may be higher during of a responsible adult who is ready to provide assistance. Home
a period of active infection. For persons with a higher catabolism treatment has been reported to be as effective as hospital treatment
of infused IgG or more frequent infections, infusions every in terms of the frequency of infections, days missed from school
3 weeks with smaller doses may be more efficacious. In the final or work, antibiotics used, and Ig level achieved. Patients receiving
analysis, trough levels are only a general guide, and the clinical home treatment should be seen regularly (i.e., approximately
well-being of the patient is a more important parameter. Clearly, every 6 months) to monitor clinical status, liver function, and
with higher trough levels, the incidence of pneumonia and serum IgG level.
comorbid conditions (e.g., bronchiectasis and meningoencepha- The first description on the use of the SC route for Ig replace-
litis) is reduced. 3-6 ment therapy was published in 1980. It was reported to be safe,
For replacement therapy in patients with PID all brands of well tolerated, and effective in achieving adequate serum IgG
IVIG are probably equivalent, although there are differences in levels. In a multicenter study of 165 patients with hypogam-
viral inactivation processes (see Table 84.2). The choice of brands maglobulinemia receiving subcutaneous infusions (27 030 at
may be dependent on the hospital, payor, or home care formulary; home), a significant reduction in adverse systemic reactions with
the distributor availability and cost; and, most importantly, patient SC administration, compared with the IM or IV administration,
tolerability. The dose, manufacturer, and lot number should be was observed. Anaphylactoid reactions did not occur. Several
recorded for each infusion to facilitate review procedures for studies have shown enhancements in quality-of-life measure-
7,8
adverse events or other consequences. It is crucial to record all ments. Local tissue reactions occur with subcutaneous
side effects that occur during the infusion. It is also recommended immunoglobulin (SCIG) infusions, including swelling, soreness,
that liver and renal functions be monitored periodically, approxi- redness, induration, local heat, itching, and bruising. There are
mately once or twice a year. Changes in weight or growth in advantages and disadvantages for each of the routes of administra-
children or changes in clinical response or adverse events may tion of replacement Ig therapy. (Clinical Pearls)
necessitate a more frequent laboratory evaluation. More frequent SCIG is a suitable alternative to IVIG and may present certain
serum IgG levels may be necessary for patients on home therapy, opportunities for optimizing care for patients with PIDD at
9
especially for those receiving subcutaneous (SC) products to home. SCIG is an excellent alternative to IVIG for those patients
evaluate compliance. unable to tolerate the IV route of Ig replacement due to systemic
In patients with active infection, especially patients with CVID, side effects with IVIG administration. SCIG also leads to higher
10
the initial (first) dose can be halved (i.e., 200–300 mg/kg) and serum IgG steady-state levels and may eliminate the wear-off
the dose repeated 2 weeks later to achieve a full dose. To minimize effects of IVIG replacement therapy. 11,12

CHaPTEr 84 Immunoglobulin Therapy: Replacement and Immunomodulation 1145



TABLE 84.2 Characteristics of Immunoglobulin (Ig) Preparations
Parenteral Iga
Brand Stabilizer/ Form and Final Content
(Manufacturer) Manufacturing Process pH Sodium Concentrations (µg/mL) antiviral Steps
Intravenous IgG
Gammagard S/D Cohn-Oncley cold ethanol 6.4–7.2 Albumin, glycine, Lyophilized <2.2 µg/ PEG precipitation, solvent
(Shire) fractionation, followed by glucose; powder mL detergent treated, low
ultracentrafiltration and ion sodium 8.5 mg/ 5%, 10% (5% pH, depth filtration
exchange chromatography mL / 17 mg/mL solution)
Gammagard Liquid Cohn-Oncley cold ethanol 4.6–5.1 Glycine; 10% liquid 37 µg/mL S/D treatment,
(Shire) fractionation, followed by trace sodium solution nanofiltration, low pH
ultracentrafiltration and ion incubation
exchange chromatography
Gamunex -C Cohn-Oncley cold ethanol 4.0–4.5 Glycine; 10% liquid 46 µg/mL Caprylate precipitation,
(Instituto Grifols, fractionation, diafiltration and trace sodium solution depth filtration, low pH
SA) depth filtration; caprylate/ incubation
anion-exchange chromatography
Privigen Cold alcohol fractionation, octanoic 4.6–5.0 L-Proline 10% solution ≤25 µg/ Depth filtration, pH4
(CSL Behring AG) fractionation, chromatography trace sodium mL incubation, nanofiltration,
and depth filtration TSE reduction steps
Gammaked Cohn-Oncley cold ethanol 4.6–4.5 Glycine 10% liquid 46 µg/mL Caprylate precipitation,
(Grifols -Distributed fractionation, diafiltration and solution depth filtration, low pH
by: Kedrion depth filtration; caprylate/ incubation
Biopharma) anion-exchange chromatography
Flebogamma DF Cold alcohol fractionation, ion 5.0–6.0 D-Sorbitol 5% and 10% <50 µg/ PEG precipitation,
(Instituto Grifols, exchange chromatography, liquid solution mL for pasteurization, S/D
SA) polyethylene glycol precipitation, the 5% treatment, nanofiltration,
pH4 treatment
Octagam Cohn-Oncley cold ethanol 5.1–6.0 Maltose 5% and 10% <100 µg/ Solvent/detergent, pH4
(Octapharma) fractionation, chromatography solution mL incubation, ultrafiltration
Gammaplex Kistler & Nitschmann fractionation, 4.8–5.1 D-sorbitol, 5% solution <10 µg/ S/D treatment,
(Bio Products chromatography, DEAE glycine and mL nanofiltration, low pH
Laboratory) Sephadex chromatography, polysorbate 80 incubation
CM-Sepharose chromatography Sodium
30–50 mmol/L
Bivigam Cohn-Oncley cold ethanol 4.0–4.6 Glycine 10% ≥ 200 µg/ S/D, nanofiltration, fraction
(Biotest fractionation, ion-exchange 0.1–0.14 M mL precipitation
Pharmaceuticals chromatography sodium
Corporation)
Carimmune NF Kistler & Nitschmann fractionation, 6.4–6.8 Sucrose Lyophilized 720 µg/ pH4, trace pepsin, depth
(CSL Behring AG) Cold alcohol fractionation, <20 mg sodium/g preparation—3%, mL filtration, nanofiltration,
Chromatography, nanofiltration protein 6%, 9%, or TSE reduction steps
12%
Subcutaneous IgG
Hizentra Cold alcohol fractionation, octanoic 4.6–5.2 L-Proline 20% ≤50 µg/ Depth filtration, pH4
(CSL Behring AG) acid fractionation, anion Trace sodium mL incubation, depth
exchange chromatography filtration, nanofiltration,
TSE reduction steps
Gamunex-C Cold ethanol fractionation, 4.0–4.5 Glycine; 10% 46 µg/mL Caprylate precipitation,
(Instituto Grifols, diafiltration and depth filtration; trace sodium depth filtration, low pH
SA) caprylate/anion exchange incubation
chromatography
Gammaked Cold ethanol fractionation, 4.6–4.5 Glycine 10% 46 µg/mL Caprylate precipitation,
(Grifols -Distributed Depth filtration, caprylate/ depth filtration, low pH
by: Kedrion anion-exchange chromatography incubation
Biopharma)
Gammagard Liquid Cohn-Oncley cold ethanol 4.6–5.1 Glycine; 10% solution 37 µg/mL S/D treatment,
(Shire) fractionation, followed by trace sodium nanofiltration, low pH
ultracentrafiltration and ion incubation
exchange chromatography
HyQvia Cohn-Oncley cold ethanol 4.6–5.1 Glycine 10% plus human 37 µg/mL S/D treatment,
(Shire) fractionation, followed by recombinant nanofiltration, low pH
ultracentrafiltration and ion hyaluronidase incubation
exchange chromatography
CUVITRU Cohn-Oncley cold ethanol 4.6–5.1 Glycine 20% 80 µg/mL S/D treatment, ultra/
(Shire) fractionation, followed by diafiltration, low pH
ultracentrafiltration and ion incubation
exchange chromatography
Contents of table summarized from manufacturers’ literature.
PEG, polyethylene glycol; S/D, solvent detergent; TSE, transmissible spongiform encephalopathy.

1146 ParT TEN Prevention and Therapy of Immunological Diseases



CLINICaL PEarLS 1 need to be instructed on the correct technique, made to apply it
Comparison of Routes of Administration of under close supervision, and educated about the recognition of
possible side effects. SCIG infusion is safer, better tolerated, and
Immunoglobulin Therapy in Patients With preferred by some patients. Several studies in Europe and Canada
Primary Immune Deficiency have shown that health care costs of SCIG therapy are lower
compared with IVIG. 14-16 It should be considered as an alternative,
Intravenous (IV) route
Advantages especially in those patients with systemic adverse reactions from
• Achieve rapid plasma levels IV administration. Recently, an alternative method for the admin-
• Can use this route in patients with bleeding disorders istration of Ig has been approved and has been referred to as
17,18
• 3–4 week intervals facilitated SC infusion. In this approach, human recombinant
hyaluronidase is used to enhance the volume of Ig (10% liquid
Disadvantages Ig) that can be infused in the SC space, allowing for monthly
• Need IV access doses of Ig to be administered in one or two sites. The hyaluronidase
• Interrupt patient’s schedule for 3–5-hour period depolymerizes the hyaluronan temporarily allowing the Ig greater
• Often needs to come to a hospital or infusion center access to the lymphatics of the SC space, facilitating the absorption
• System side effects may be more frequent in some patients
of Ig. This method of administration of 10% Ig has been shown
Subcutaneous (SC) route in pharmacokinetic studies to result in bioavailability of 93% of
Advantages the IVIG dose and thus does not require an upward adjustment
• IV access not needed for those patients with poor venous access factor in calculating the dose of Ig replacement. More details about
11,19,20
• Eliminate trough levels SC infusions can be found in several reviews.
• Achieve a stable serum level of immunoglobulin G (IgG)
• May eliminate 3rd–4th week fatigue prior to next infusion
(wear off) ADVERSE EVENTS ASSOCIATED WITH
• Less systemic adverse effects than IV route IVIG THERAPY
• More flexibility for patient’s (parent’s) schedule
• Distance from infusion center or hospital Rate-Related Adverse Events
• Young adults going to college
Typical rate-related adverse reactions with IVIG include tachy-
Disadvantages cardia, dyspnea, chest tightness, back pain, arthralgia, myalgia,
• Minor local reactions at the site of infusion hypertension or hypotension, headache, pruritus, rash, and
• Patient reliability low-grade fever. Mild to moderate reactions occur in 5–15% of
• Need for a pump infusions; severe reactions occur in <1% of patients. Of course,
in patients with autoimmune disorders, reaction rates are higher
with higher doses. Patients with more profound immunodeficiency
SCIG products are 10% or 20% formulations, the former being or patients with active infections also tend to have more severe
similar in composition to IVIG products. The calculated dose for reactions. Other factors that contribute to adverse reactions
SC administration is generally 100–150 mg/kg weekly. Depending include change of IVIG products, concomitant infections, higher
on the weight or body mass of the patient and the concentration concentrations or lyophilized products, and rapid infusion rates
21
of the SCIG (i.e., 10% or 20%), infusions may have to be given (reviewed in the report by Stiehm ). The cause of the reactions
more frequently than every 7 days. Pharmacokinetics studies in is thought to be related to the anticomplementary activity of
clinical trials have suggested that upward adjustments in the dose IgG aggregates in the IVIG in which immune complexes form
of 37% of the IVIG dose may be needed to achieve comparable between infused antibodies and antigens of infectious agents in
bioavailability, defined as the area under the serum concentration the patient. The other possible mechanism is that the formation
13
curve (AUC). This adjustment in dosing between IVIG and SCIG of oligomeric or polymeric IgG complexes interacts with Fc
has not been mandated by European regulatory agencies, and in receptors and triggers the release of inflammatory mediators.
the United States some clinical immunologists select dosing based These rate-related reactions occur less frequently with the newer
on optimization of prevention of infections, as noted above for IVIG products that are liquid and isoosmolar. Headaches are
5,6
the IVIG dosing. Each subcutaneous infusion requires a small the most frequent symptom associated with IVIG infusions
portable syringe driver–type pump together with a 10–20-mL occurring in 5–20% of infusions and one-third of patients.
syringe and an infusion set with a specialized SC 25–27-gauge Slowing the infusion rate or discontinuing therapy until symptoms
needle. The length of the needle may have to be adjusted for the subside may diminish the reaction. Pretreatment with nonsteroidal
thickness of the subcutaneous tissue of each patient. Before infusion, antiinflammatory drugs (NSAIDs), acetaminophen (15 mg/kg/
the line needs to be checked to ensure that there is no blood dose), diphenhydramine (1 mg/kg/dose), and/or hydrocortisone
return. Infusions can be given weekly at multiple sites or more (6 mg/kg/dose, maximum 100 mg) 1 hour before the infusion
often (e.g., daily), if needed, to maintain adequate serum IgG may prevent adverse reactions. Oral hydration prior to the infusion
levels. The 20% SCIG product can be given every 2 weeks. Infusion is often helpful. Switching products may also lead to adverse
sites are usually on the abdominal wall and lateral thigh. In adults, events in 15–18% of patients and should be discouraged.
20–35 mL can be infused into a single site, depending on the
amount of subcutaneous tissue. A general guideline for infusion Central Nervous System–Related Adverse Events
rate is 15–35 mL/hr per site depending on the weight and sub- Aseptic meningitis has been reported as one of the complications
cutaneous tissue of the patient. The SCIG schedule should be of IVIG, especially with large doses and rapid infusions and in
21
started 1 week after the last dose of IVIG, or in a new patient the treatment of patients with autoimmune disease. Interestingly,
loaded with 4 or 6 doses of SCIG. Before home treatment, patients this rarely occurs in subjects with immunodeficiencies. Symptoms,

CHaPTEr 84 Immunoglobulin Therapy: Replacement and Immunomodulation 1147



CLINICaL PEarLS 2 successfully with conservative therapy, but deaths were reported
Adverse Events Associated With in 17 patients who had serious underlying conditions. Most
cases of this adverse event were associated with IVIG
Immunoglobulin Therapy products containing sucrose as a stabilizer. Risk factors for this
rate-related adverse reaction include preexisting renal insufficiency, diabetes
• Infusion site erythema, swelling, pain, itching mellitus, dehydration, patient age >65 years, sepsis, paraprotein-
• Headache emia, and concomitant use of nephrotoxic agents. Newer IVIG
• Myalgia, back pain, arthralgia products are using alternative stabilizers (e.g., amino acids) instead
• Malaise, fatigue of sucrose.
• Chills, Fever
• Pruritus
• Rash—urticaria
• Nausea, vomiting CLINICaL PEarLS 3
• Tachycardia Risk Factors for Adverse Events
• Dyspnea, chest pain, or tightness
• Hypotension/hypertension
• Infusion issues
• Prior history of an infusion reaction with an immunoglobulin (Ig)
Central Nervous System product
• Severe headaches • First infusion in a patient with active infection or inflammation
• Trigger migraine headaches • Changing immunoglobulin products
• Aseptic meningitis* • Rapid infusion and/or large dose
• Patient factors
renal • Preexisting renal insufficiency
• Azotemia • Prior history of thrombotic event
• Renal failure • Autoimmune disorder
• Diabetes mellitus
Thromboembolic Events* • Age—older age
• Thrombosis/cerebral infarction • Hyperlipidemia or elevated cholesterol
• Myocardial infarction • Dehydration with volume depletion
• Pulmonary embolism • Hypercoagulable state
• Indwelling catheters
• Paraproteinemia or other causes of hyperviscosity
anaphylaxis From anti-Immunoglobulin E (IgE) antibodies to Iga
• Cardiac or peripheral vascular disorders
• Estrogen use
Other (Isolated reports) • Smoking
• Cardiac rhythm abnormalities
• Coagulopathy
• Serum sickness
• Hemolysis—alloantibodies to blood type A/B
• Cryoglobulinemia Thromboembolic Events
• Neutropenia This adverse effect was observed mainly in patients receiving
• Alopecia large doses of IVIG for autoimmune diseases. Patients with
• Uveitis elevated serum viscosity (e.g., cryoglobulinemia, hypergam-
• Noninfectious hepatitis
• Progressive neurodegeneration maglobulinemia, and hypercholesterolemia) are at risk for
developing a critical increase in serum viscosity with IVIG,
*See text for predisposing risk factors. especially high doses that predispose them to thromboembolic
events, such as myocardial infarction, stroke, deep vein thrombosis,
or pulmonary embolism. (Clinical Pearls 3) Recently, a contami-
nating procoagulant factor (e.g., factor XIa) has been implicated
22
including headache, stiff neck, and photophobia, usually develop in these thromboembolic events. Patients at risk are older (>65
within 24 hours after completion of the infusion and may last years), on multiple drugs, and have comorbidities, such as diabetes,
3–5 days. Spinal fluid pleocytosis occurs in most patients. Long- hypertension, and so on. Patients at risk should be well hydrated,
term complications are minimal. The etiology of aseptic meningitis IVIG should be administered at lower rates, and products with
is unclear, but migraine has been reported as a risk factor low sodium and an osmolality in the physiological range should
and may be associated with recurrence despite the use of be selected. Ig manufacturers have taken steps to remove or
different IVIG preparations and slower rates of infusion. (Clinical reduce this procoagulant activity from their products. 23
Pearls 2)
Transfusion Reaction Caused by Antibodies Against IgA
Renal Adverse Events True anaphylaxis is rare in patients with selective IgA deficiency
Acute renal failure is a rare but significant complication of IVIG and CVID who develop IgE antibodies to IgA after treatment
treatment and has been associated in the past with products with Ig; this adverse event appears to be much less frequent than
containing sucrose. Histopathological findings of acute tubular originally thought. Patients with CVID have IgG antibodies
necrosis, vacuolar degeneration, and osmotic nephrosis were (10–22% in various studies) to IgA, but there is no correlation
24
suggestive of osmotic injury to the proximal renal tubules. Most to the presence of these antibodies and adverse reactions. Patients
patients (95%) had received large doses for treatment of auto- with anti-IgA antibodies who have had reactions to IVIG have
immune diseases. The majority of the cases were treated tolerated SCIG. 25

1148 ParT TEN Prevention and Therapy of Immunological Diseases



Other Adverse Reactions KEY CONCEPTS 3
A number of other adverse reactions have been reported in Mechanisms of Action of Intravenous
association with IVIG infusions. (Clinical Pearls 2) These side Immunoglobulin (IVIG) in Autoimmune and
effects are usually less common and are discussed in more detail Inflammatory Diseases
elsewhere. 21
• Blockade of Fc receptors on macrophages of the reticuloendothelial
system of liver and spleen
Summary: Ig Replacement in Treatment of • Restoration of idiotypic/antiidiotypic network
Immune Deficiency • Suppression or neutralization of cytokines by specific antibodies in
Ig replacement is the mainstay of treatment for primary humoral the IVIG
immune deficiency. The goal of treatment is to provide a broad • Block binding of adhesion molecules on leukocytes to vascular
endothelium
spectrum of antibodies to prevent infections and chronic • Inhibition of complement uptake on target tissues
long-term complications. The usual dose is 400–600 mg/kg/ • Neutralization of microbial toxins
month, but this may vary among individuals, and some patients • Blockade of Fas ligand mediated apoptosis by anti-Fas antibodies in
may require higher doses. A serum trough level above 500 mg/ the IVIG
dL has been shown to be effective in the prevention of • Induction of apoptosis with anti-Fas antibodies at high concentrations
serious bacterial infections. However, recent studies have sug- of IVIG
gested that even higher doses (e.g., a “biological” trough level • Neutrophil apoptosis by anti-Siglec-9 antibodies in IVIG
and achieving trough levels of IgG in the range of 750–900 mg/ • Saturation of FcRn receptors to enhance the clearance of
autoantibodies
6
dL) may be desirable. Ig can be given intramuscularly, • Induction of inhibitory FcγInd receptors on effector macrophages
intravenously, or subcutaneously. SC administration has been • Neutralization of growth factors for B cells (e.g., B cell–activating
proven to be safe and is a good alternative in some patients, factor ([BAFF])
especially those experiencing side effects of administration by • Inhibition of T-cell proliferative responses
the IV route. Generally, IVIG replacement therapy is considered • Expansion and/or activation of a population of regulatory T cells (Tregs)
safe in the majority of patients. Side effects are usually mild and • Enhancement of the cyclooxygenase 2 (COX-2) pathway by increasing
prostaglandin E2 (PGE 2 ) from dendritic cells (DCs)
treatable with premedication. Good manufacturing practices, • Downregulation of the T-helper 17 (Th17) pathway
improved screening of plasma donors, testing of the source • Modulation of DC function through C-type lectin receptors (DCLRs)
plasma, and additional viral inactivation and removal steps have DC-specific intercellular adhesion molecule-grabbing nonintegrin
made IVIG a better and safer plasma-derived product. To (DC-SIGN; spleen) and DCLR (lung and lymph nodes)
optimize the care of patients with PIDs, there are eight guiding
principles for the use of Ig replacement therapy. (see Therapeutic
Principles) have shown that IVIG leads to the rapid reversal of the throm-
bocytopenia. Autoantibody-opsonized platelets are destroyed in
the spleen and the liver by FcγR-mediated phagocytic clearance.
MECHANISMS OF ACTION OF IG (Key Concepts 4)
THERAPY IN AUTOIMMUNE AND
INFLAMMATORY DISEASES Interactions of Idiotype and Antiidiotype as
Immune Modulation
Although we still do not understand all the mechanisms by which Idiotype–antiidiotype interactions between antiplatelet glyco-
IVIG has immunomodulatory effects, knowledge of the actions protein IIb (GPIIb)/IIIa autoantibodies and IVIG may be another
of IVIG in these diseases will allow for definition of appropriate mechanism by which IVIG could affect autoantibody production
indications and schedules of administration of IVIG and the and effector function in ITP and may be playing a role in the
design of new-generation IVIG products that are better able to long-term immune effects in ITP. IVIG contains antiidiotypic
target the immune perturbations in autoimmune and inflam- antibodies to anti-DNA, anti–factor VIII, antineutrophil cyto-
matory processes. (Key Concepts 3) Furthermore, additional plasmic antibody (ANCA) autoantibodies, antithyroid autoan-
multicenter placebo-controlled clinical trials are needed to confirm tibodies, and others. A proposed mechanism of action of IVIG
clinical efficacy. A more detailed discussion of the evidence-based in ANCA-positive vasculitis is binding or neutralization of the
treatment of autoimmune and inflammatory disorders can be ANCA autoantibodies by antiidiotypic antibodies in IVIG. Studies
1,2
found elsewhere. This chapter will not review the indications have shown that IVIG contains antibodies with idiotypic specifici-
for IVIG therapy; the reader is directed to other works, including ties that can bind and neutralize potentially pathogenic auto-
comprehensive reviews and practice-based guidelines, for more antibodies in autoimmune neurological diseases, such as
details about the various indications for IVIG therapy in Guillain-Barré syndrome (GBS), chronic inflammatory demy-
26
autoimmune and inflammatory disorders. This section will address elinating polyneuropathy (CIDP), and myasthenia gravis (MG).
the possible mechanisms of actions of Ig therapy; more than Antiidiotypic antibodies in IVIG directed against idiotopes on
one mechanism(s) may be important in the immune modulation the anti-GM 1 Ig molecule block the binding of the anti-GM 1
of these autoimmune disorders. antibodies to its target antigen. Support for a similar mechanism
in MG comes from the fact that IgG or F(ab’) 2 fragments in
Blockade of Fc Receptors of the IVIG preparations are capable of binding to anti–acetylcholine
Reticuloendothelial System receptor antibodies in vitro. Additionally, using in vitro nerve-
27
Idiopathic thrombocytopenic purpura (ITP) results from acceler- muscle preparations, Buchwald et al. showed that the F(ab’) 2
ated platelet destruction attributable to an immunological process portion of IVIG neutralized the “blocking” effect of serum from
resulting in bleeding that may be life threatening. Studies in ITP patients with acute GBS. (Key Concepts 5)

CHaPTEr 84 Immunoglobulin Therapy: Replacement and Immunomodulation 1149


These antiidiotypic antibodies may also act in concert with to the sialylation of the glycan component of the Fc fragment.
their effects on the FcγRIIB receptor on B cells to produce a The important glycan moiety in the IgG molecule is located at
297
negative “off signal” to the B cells synthesizing these autoimmune the asparagine (Asn ) site in the second domain of the constant
antibodies. Thus the antiidiotypic antibodies in IVIG may be region. Using a K/BxN serum-induced arthritis model in mice,
32
beneficial by restoring the idiotypic control network in these Kaneko et al. showed that IVIG at 1 g/kg inhibited the inflam-
autoimmune diseases. matory arthritic process. Deglycosylated or neuraminidase-treated
IVIGs were unable to inhibit this inflammation. IVIG enriched
The Role of the FcRn Receptor on Immune Modulation for the sialylated glycan moiety had comparable inhibitory effects
The FcRn receptor (neonatal Fc receptor) was identified as the on the inflammatory process at only one-10th of the dosage
receptor responsible for protecting IgG from catabolism in the used with intact IVIG. This inhibitory activity was dependent
33
endocytic vesicles of the endosome, and this explains the relatively on FcγRIIB expression on effector macrophages. Anthony et al.
long half-life of this plasma protein. IVIG may accelerate the engineered a recombinant/sialylated human IgG1 Fc protein that
catabolism of IgG autoantibodies by saturating these protective had 35-fold enhanced immune-modulating activity compared
34
receptors in direct proportion to the relative concentration of with native IVIG. Anthony et al. performed studies to examine
exogenous plasma levels of IgG from IVIG. In mouse models the mechanism by which the sialylated Fc fragment could mediate
of bullous pemphigoid and arthritis, IVIG treatment resulted its antiinflammatory activity and to identify the target cell that
in a reduction in pathogenic antibodies to levels below the initiates this antiinflammatory pathway. A splenic marginal zone
disease-causing threshold, and this effect was attenuated in macrophage expressing the C-type lectin receptor (i.e., SIGN-R1)
28
29
FcRn-deficient mice. Hansen and Balthasar showed in a rat was required for the antiinflammatory activity of IVIG in concert
model of immune thrombocytopenia that IVIG enhanced the with its ability to bind to sialylated Fc domains. These authors
clearance of antiplatelet antibodies in a dose-dependent manner proposed that the interaction between sialylated IgG Fc and
30
by saturation of the FcRn receptor. These same investigators SIGN-R1 leads to the upregulation of the inhibitory FcγRIIB
estimated that approximately 50% of the overall effect of IVIG receptor on effector cells. They suggested that DC-SIGN (dendritic
in ITP may be attributed to IVIG-mediated acceleration of the cell–specific intercellular adhesion molecule-grabbing noninte-
elimination of antiplatelet antibodies by the FcRn saturation grin), the human orthologue of SIGN-R1, has a comparable role
mechanism and showed that in FcRn knock-out mice, IVIG in the antiinflammatory effects of IgG Fc fragment on human
35
failed to increase the clearance of antiplatelet antibodies. However, macrophages and dendritic cells (DCs). Anthony et al. presented
very little data, if any, are available in human autoimmune disease data in their murine model that this immune-modulating pathway
to support this mechanism. may be mediated by the production of interleukin-33 (IL-33)
by DCs, which, in turn, produce IL-4 from basophils, leading
KEY CONCEPTS 4 to the increased expression of the FcγRIIB receptor on effector
macrophages. Of note, observations in patients with chronic
Proposed Mechanisms of Action of intravenous inflammatory demyelinating polyneuropathy (CIDP) treated
immunoglobulin (IVIG) in Autoimmune Idiopathic with high-dose IVIG demonstrated upregulation of the FcγRIIB
Thrombocytopenic Purpura receptor on peripheral blood monocytes. Others, however, have
36
questioned whether this pathway of immune modulation is
• Fc receptor blockade of reticuloendothelial system important in humans. 37-39 Nevertheless, these exciting studies
• Fcγ receptor downregulation
• Idiotype–antiidiotype interaction between antiplatelet GPIIb/IIa auto- define, at least in mice, an important mechanism by which IVIG
antibodies and the antiidiotypic antibodies in IVIG modulates immune processes mediated through sialylated Fc
• Activation of inhibitory receptor FcγRIIB on the IgG molecule and the receptors on DCs and effector
• Saturation of FcRn receptor to accelerate the catabolism of antiplatelet macrophages (i.e., SIGN-R1 and FcγRIIB) involved in this
autoantibodies antiinflammatory process.


Modulation of Immunoregulatory Function Through KEY CONCEPTS 5
the Fc Receptor Proposed Mechanisms of Action of IVIG in
The FcγRIIB receptor provides an inhibitory signal to cells through Neuromuscular Diseases
a pathway mediated by an immunoregulatory tyrosine-based • Modulation of proinflammatory cytokines, e.g., TNF-GR α, IL-1GR β
inhibition motif (ITIM). Studies have shown that IVIG stimulates in GBS
these inhibitory FcγRIIB receptors found on a variety of cell • Reduction in muscle levels of ICAM-1, TGF-GR β, and TGF-GR β
types, including macrophages, B cells, and a subpopulation of mRNA in dermatomyositis
31
T cells. Samuelsson et al., using a murine model of ITP, showed • Neutralization by antiidiotypic antibodies in IVIG of potentially pathogenic
autoantibodies, e.g., anti-Gm1 antibodies in GBS and CIDP, and anti-
that the administration of IVIG prevented platelet destruction AchR in MG
by a pathogenic monoclonal autoantibody. Protection was • IVIG saturation of FcRn receptors to accelerate degradation of IgG
associated with the induction of the expression of FcγRIIB autoantibodies
receptors on splenic macrophages. This inhibitory FcγRIIB • Inhibition of complement deposition and formation of MAC on
receptor was required for the protection of the animals against endomysial capillaries in dermatomyositis
the monoclonal autoantibody, since disruption of the receptor AchR, acetylcholine nicotinic receptor; CIDP, chronic inflammatory
by either genetic deletion or a blocking monoclonal antibody demyelinating polyneuropathy; FcRn, neonatal Fc receptor;
32
(mAb) reversed the therapeutic effects of IVIG. Kaneko et al. GBS, Guillain-Barré syndrome; MAC, membranolytic attack complex;
showed that that the inhibitory properties of IVIG were linked MG, myasthenia gravis.

1150 ParT TEN Prevention and Therapy of Immunological Diseases



Neutralizing Antibody Activity in IVIG Against superantigens, which could be used in the treatment of streptococ-
cal toxic shock syndrome. The neutralizing capacity of IVIG
Bacterial Toxins against these bacterial superantigens is important because of
Kawasaki syndrome (KS), an acute multisystem disease of their potential to stimulate the production of proinflammatory
unknown etiology, primarily affects infants and young children. cytokines that lead to clinical disease. A number of in vitro studies
Although KS occurs worldwide in children of all racial groups, have shown that IVIG can inhibit the production of, or bind to
it is most prevalent in Japan and in children of Japanese ancestry. and neutralize, a number of cytokines and growth factors from
Although the acute illness is generally self-limiting, coronary various cell types. 42-44 Thus IVIG may exert its antiinflammatory
artery abnormalities related to a generalized inflammation and effects in many different types of inflammatory diseases by
immune activation of small- and medium-sized blood vessels interrupting or modifying a number of different steps in the
develop in up to 25% of untreated patients. Although the etiology inflammatory cascade, from the inhibition of effector cell function
remains unknown, the clinical features and laboratory findings to reduction in cytokine-induced endothelial cell activation, or
suggest an infectious or postinfectious process. The administration the “neutralization” of proinflammatory cytokines.
of high-dose IVIG, together with aspirin, is the standard of care
in the treatment of KS.
KS is associated with marked activation of T cells and Modulation of Adhesion Molecules on
monocytes–macrophages. On the basis of immunological and Endothelial Cells and Antibodies in IVIG
clinical features of KS overlapping with those of bacterial toxic to Cell Surface Receptors
shock–like syndrome, studies were carried out to determine if IVIG contains a number of natural autoantibodies that may
KS is associated with exposure to a superantigen, such as a have immune-modulating activities (e.g., antibodies to CD4,
bacterial toxin. Acute KS is associated with marked immune major histocompatibility complex [MHC] class I molecules,
activation and increased circulating cytokine levels. Some of cytokines, adhesion molecules, and Siglec molecules on leukocytes
these cytokines elicit proinflammatory and prothrombotic and other cell surface molecules). The “natural” antibodies in
responses by inducing the expression of leukocyte adhesion IVIG have also been shown to bind to a number of plasma and
molecules, which localize inflammatory cells to vascular endo- tissue proteins, including B cell–activating factor (BAFF),
thelial cells. The expression of endothelial–leukocyte adhesion granulocyte macrophage–colony-stimulating factor (GM-CSF),
molecules has been demonstrated in acute KS, and its downregula- liver antigens, and beta-amyloid peptide. 42,45 The binding of native
tion correlates with favorable response to IVIG treatment. The IgG can be significantly increased by mild denaturing conditions
magnitude and persistence of proinflammatory cytokine synthesis (e.g., mild pH treatment and cold ethanol precipitation used
46
have been reported to constitute a risk for the development of during the manufacturing of IVIG). St-Amour et al. suggested
coronary artery abnormalities. that these commercial fractionation processes for IVIG could
IVIG has been shown to contain high titers of specific antibod- contribute to its therapeutic antiinflammatory effects.
47
ies inhibitory to the activation of T cells by staphylococcal and Vassilev showed that IVIG contains specific antibodies
streptococcal superantigens (Chapter 6). These findings may to a 10-peptide sequence, including the RGD (Arg-Gly-ASP)
account for the observation that treatment of acute KS with motif that is expressed in adhesion molecules on a variety
IVIG results in a marked reduction of macrophage and T-cell of cell surfaces. Most integrins bind to this RGD sequence.
48
40
activation. In this regard, the efficacy of IVIG in suppressing Gill et al. reported that IVIG inhibits leukocyte recruitment
the immune activation associated with KS and, more importantly, into inflammatory tissues by inhibiting selectin and integrin
its ability to prevent the development of coronary artery abnor- binding. In a mouse model of sickle cell vaso-occlusive crisis,
malities in this illness may relate to the neutralizing antibody IVIG was shown to inhibit neutrophil adhesion to the vascular
activity of IVIG against these bacterial toxins. Toxin neutralization endothelium, resulting in an increase in capillary blood flow
49
is not likely the only beneficial effect of IVIG in KS. Blocking and reversal of the vessel occlusion. IVIG could modulate this
or neutralizing cytokines by the anticytokine antibodies in cytokine-mediated endothelial cell activation by neutralizing the
IVIG may modulate the local inflammatory responses of effects of the cytokines, inhibiting endothelial cell responses to the
blood vessels in KS by modifying leukocyte adhesion after cytokines, or inhibiting the production of cytokines and growth
increasing the expression of cell-surface determinants on vascular factors. These mechanisms of IVIG may be playing an important
endothelial cells. role in preventing coronary artery abnormalities in patients
IVIG may have therapeutic value in the treatment of patients with KS.
with toxic shock syndrome secondary to Staphylococcus aureus Toxic epidermal necrolysis (TEN) and Stevens-Johnson
or Streptococcus pyogenes exotoxins. In an open study by the syndrome (SJS) are severe drug-induced skin diseases (reviewed
41
50
Canadian Streptococcal Study Group , IVIG appeared to be in Letko et al. ). TEN results in apoptotic epidermal cell death,
beneficial in patients with streptococcal toxic shock syndrome. in which there is separation of large areas of the skin at the
In a meta-analysis of IVIG treatment of neonatal sepsis, there epidermal junction, producing the appearance of scalded skin.
was a sixfold decrease in mortality. IVIG inhibits Staphylococcus Keratinocyte apoptosis that precedes epidermal detachment is
exotoxin–induced T-cell activation and contains antibodies against an early event in TEN. A number of drugs, including sulfonamides,
exotoxins responsible for toxic shock syndrome. Great variations anticonvulsants, and NSAIDs, can cause TEN and SJS. The
51
in neutralizing activity against streptococcal pyrogenic exotoxins mortality rate can be as high as 30%. Viard et al. studied serum
can be found in different brands and even among different lots samples from patients with TEN and found that the sera of these
of IVIG. However, these findings suggest that it is possible to patients had very high levels of soluble Fas ligand (sFasL).
select one IVIG preparation that contains high levels of neutral- Keratinocytes normally express the death receptor Fas. The
izing activity against a wide variety of Group A streptococcal keratinocytes of patients with TEN also express very high levels

CHaPTEr 84 Immunoglobulin Therapy: Replacement and Immunomodulation 1151


62
51
of active Fas ligand. In their small pilot study, Viard et al. model downregulated the Th-17 pathway. In patients with KS
administered IVIG (0.2–0.75 g/kg for 4 consecutive days) to 10 and GBS, clinical improvement with IVIG therapy correlated
patients with TEN. In all 10 patients, the progression of skin with increased number and function of Tregs. 63,64
disease was rapidly arrested within 24–48 hours, with rapid skin
healing and no adverse effects. In in vitro studies, IVIG completely Summary: IVIG in Treatment of Autoimmune and
inhibited Fas-mediated keratinocyte apoptosis. This effect was Inflammatory Diseases
related to the presence of naturally occurring Fas-blocking Clearly, IVIG (IgG) has a number of immune-modulating effects
antibodies in IVIG, which inhibit Fas-mediated keratinocyte cell and has been found to be an effective treatment for a wide
death. spectrum of autoimmune and inflammatory diseases. 44,65 At
present, IVIG is FDA approved for only a few autoimmune and
Modulation of Complement Effector Function inflammatory diseases. In autoantibody-mediated disease, the
The principal inflammatory mechanism in dermatomyositis Fc domain appears to be the important IgG moiety that leads
(DM) is complement (C)-dependent microangiopathy with to immune modulation. The importance of sialylation of the
activation of C3 and deposition of the complement C5b-9 Fc fragment remains controversial, as do the mediators involved
membrane attack complex (MAC) on the endomysial capillar- (e.g., IL-33 and IL-4). Differences in animal models, IVIG source,
ies. 26,52 Basta et al. showed that IVIG can inhibit the uptake of route and timing of the administration of the IVIG, mouse strain,
53
C components on target tissues. In patients with DM who were and other variables may account for the differences in the
treated with IVIG, C3 deposition was reduced with corresponding experimental observations of laboratories. In T-cell mediated
decreases in complement expression on endomysial capillaries. 26,52 animal models of disease, such as EAE, there is strong evidence
IVIG prevents the uptake of complement components and that there is upregulation of Tregs and inhibition of the Th17
formation of the MAC on the endomysial capillaries in the muscle pathway. These effects may be mediated by the F(ab’)2 portion
tissues of patients with DM. Consequently, IVIG allowed neo- of the IgG molecule, and not the Fc domain. Furthermore, there
vascularization to occur with reversal of the ischemic process, is also controversy over the receptor on antigen-presenting cells
resulting in muscle tissue healing. This effect of IVIG on comple- (APCs), such as macrophages and DCs, which are involved in
ment deposition may be relevant to other autoimmune neurologi- the immune-modulating process mediated by IVIG. In certain
cal diseases, such MG, GBS, and CIDP, in which complement animal models, the SIGN-R1 (or in humans DC-SIGN) is
54
may be playing a role in the tissue damage. 26,52 Arumugam et al. important, and in other models (e.g., murine asthma), a novel
showed that IVIG protects the brain against ischemic injury C-type lectin receptor (DCIR) appears to be important. These
mediated by complement in a mouse model of experimental differences undoubtedly relate to the disease model. Two studies
stroke. have demonstrated the importance of the PGE 2 pathway on
IVIG-induced immune modulation mediated by the F(ab’)2
Effects of Ig on the Regulatory T-Cell Pathways portion of the IgG molecule. These observations may point to
In a mouse model of multiple sclerosis (MS), IVIG has been the possibility of alternative treatment regimens that employ
+
shown to expand and enhance the function of FoxP3 regulatory the selective increase in PGE 2 in certain autoimmune or inflam-
55
T cells (Tregs) while inhibiting the differentiation of T-helper matory disorders to increase Tregs and inhibit the production
17 (Th17) and Th1 cells. This protective effect of IVIG was of certain cytokines. Further clarification in human disease models
56
lost in mice that were depleted of Tregs. These changes were using in vitro human cells is important. These mechanisms are
independent of FcγRIIB and the Fc domain, since F(ab’) 2 frag- not mutually exclusive, and probably more than one mechanism
ments led to similar changes in Th17 cells, Tregs, and clinical is playing a role in the efficacy of Ig therapy in an autoimmune
efficacy in this experimental allergic encephalomyelitis (EAE) disease process. A better understanding of the pathogenic
model. Desialylated IVIG had the same immune-modulating mechanisms involved in these diseases will undoubtedly lead to
58
57
effects as “native” IVIG. Investigations by Trinath et al. sug- a more effective therapy with IVIG and more specific, modified
gested that the mechanism by which IVIG induces Tregs was forms of this biological product.
the enhancement of the cyclooxygenase 2 (COX-2) pathway
via increased expression of prostaglandin E2 (PGE 2 ) from
human DCs.
Kaufman et al. have extensively studied the effects of IVIG ON THE HOrIZON
therapy in an ovalbumin-sensitized mouse model of asthma. Translational Research Opportunities Related to
They reported that IVIG markedly attenuated lung inflammation, Immunoglobulin Therapy
decreased bronchial hyperresponsiveness to methacholine, and
59
suppressed the Th2 pathway. The draining pulmonary lymph • Elucidation of mechanism(s) of action will lead to more precise bio-
nodes of IVIG-treated mice showed a significant increase in engineered molecular products to treat autoimmune and inflammatory
diseases.
+
+
+
CD4 CD25 FoxP3 regulatory cells. IVIG-primed DCs on adoptive • Clarification of the mechanisms of action of intravenous immunoglobulin
transfer to ovalbumin (OVA)–sensitized and challenged mice (IVIG) in autoimmune and inflammatory diseases should lead to better
60
abrogated airway hyperresponsiveness and induced Tregs. In understanding of the pathobiology of these diseases.
61
their model system, Massoud et al. reported that sialylated IgG • Enhanced product purification—identification of the minor components
bound to a novel C-type lectin receptor (i.e., dendritic cell in Ig products that may contribute to adverse reactions should lead
immunoreceptor [DCIR]) induced Tregs. Thus a number of to improved manufacturing processes and improved product tolerability
studies have demonstrated the importance of the induction for patients (e.g., procoagulant factors, isohemagglutinins)
+
of FoxP3 Tregs by IVIG in modulating the autoimmune/ • Ig delivery—development of innovative approaches to the delivery of
Ig products to patients to enhance safety and compliance.
antiinflammatory process. In contrast, Ig therapy in this murine

1152 ParT TEN Prevention and Therapy of Immunological Diseases


Please check your eBook at https://expertconsult.inkling.com/ immunoglobulins for primary immunodeficiency. J Allergy Clin
for self-assessment questions. See inside cover for registration Immunol 2012;130:951–7.
details. 19. Wasserman RL. Progress in gammaglobulin therapy for
immunodeficiency: from subcutaneous to intravenous infusions and back
again. J Clin Immunol 2012;32(6):1153–64.
REFERENCES 20. Bonilla FA. Pharmacokinetics of immunoglobulin administered via
intravenous or subcutaneous routes. Immunol Allergy Clin North Am
1. Orange JS, Hossny EM, Weiler CR, et al. Use of intravenous 2008;28(4):803–19, ix.
immunoglobulin in human disease: a review of evidence by members of 21. Stiehm ER. Adverse effects of human immunoglobulin therapy. Transfus
the Primary Immunodeficiency Committee of the American Academy of Med Rev 2013;27:171–8.
Allergy, Asthma and Immunology. J Allergy Clin Immunol 22. Daniel GW, Menis M, Sridhar G, et al. Immune globulins and thrombotic
2006;117:S525–53. adverse events as recorded in a large administrative database in 2008
2. Perez E, Bonilla T, Chinen J, et al. Update on the use of Immune Globulin through 2010. Transfusion 2012;52:2113–21.
(IG) in human disease: a review of evidence by members of the primary 23. Farrugia A, Quinti I. Manufacture of immunoglobulin products for
immunodeficiency committee of the American Academy of Allergy, patients with primary antibody deficiencies- the effect of processing
Asthma and Immunology. J Allergy Clin Immunol 2017;139(3):S1–46. conditions on product safety and efficacy. Front Immunol 2014;5:
3. Quartier P, Debre M, DeBlie J, et al. Early and prolonged 1–8.
intravenous immunoglobulin replacement therapy in childhood 24. Horn J, Thon V, Bartonkova D, et al. Anti-IgA antibodies in common
agammaglobulinemia: a retrospective survey of 31 patients. J Pediatrics variable immunodeficiency (CVID): diagnostic workup and therapeutic
1999;134:589–96. strategy. Clin immunol 2007;122(2):156–62.
4. Orange JS, Grossman WJ, Navickis RJ, et al. Impact of trough IgG on 25. Rachid R, Bonilla FA. The role of anti-IgA antibodies in causing adverse
pneumonia incidence in primary immunodeficiency: a meta-analysis of reactions to gamma globulin infusion in immunodeficient patients: a
clinical studies. Clin immunol 2010;137(1):21–30. comprehensive review of the literature. J Allergy Clin Immunol
5. Bonagura VR, Marchlewski R, Cox A, et al. Biologic IgG level in primary 2012;129(3):628–34.
immunodeficiency disease: the IgG level that protects against recurrent 26. Dalakas M. Mechanistics effects of IVIG in neuroinflammatory diseases:
infection. J Allergy Clin Immunol 2008;122(1):210–12. conclusions based on clinicopathologic correlations. J Clin Immunol
6. Lucas M, Lee M, Lortan J, et al. Infection outcomes in patients with 2014;34(Suppl1):S120–6.
common variable immunodeficiency disorders: relationship to 27. Buchwald B, Ahangari R, Weishaupt A, et al. Intravenous
immunoglobulin therapy over 22 years. J Allergy Clin Immunol immunoglobulins neutralize blocking antibodies in Guillain-Barre
2010;125(6):1354–60 e4. syndrome. Ann Neurol 2002;51:673–80.
7. Jiang F, Torgerson TR, Ayars AG. Health-related quality of life in patients 28. Li N, Zhao M, Hilario-Vargas J, et al. Complete FcRn dependence for
with primary immunodeficiency disease. Allergy Asthma Clin Immunol intravenous Ig therapy in autoimmune skin blistering diseases. J Clin
2015;11:27–38. Invest 2005;115:3440–50.
8. Routes J, Costa-Carvalho B, Grimbacher B, et al. Health-related quality of 29. Hansen RJ, Balthasar JP. Effects of intravenous immunoglobulin on
life and health resource utilization in patients with primary platelet count and antiplatelet antibody disposition in a rat model of
immunodeficiency disease prior to and following 12 months of immune thrombocytopenia. Blood 2002;100:2087–93.
immunoglobulin G treatment. J Clin Immunol 2016;36(5):450–61. 30. Hansen RJ, Balthasar JP. Mechanisms of IVIG action in immune
9. Jolles S, Orange JS, Gardulf A, et al. Current treatment options with thrombocytopenic purpura. Clin Lab 2004;50:133–40.
immunoglobulin G for the individualization of care in patients with 31. Samuelsson A, Towers TL, Ravetch JV. Anti-inflammatory activity of IVIG
primary immunodeficiency disease. Clin Exp Immunol 2015;179: mediated through the inhibitory Fc receptor. Science 2001;291:484–6.
146–60. 32. Kaneko Y, Nimmerjahn F, Ravetch JV. Anti-inflammatory activity of
10. Berger M, Rojavin M, Kiessling P, et al. Pharmacokinetics of immunoglobulin G resulting from Fc sialylation. Science 2006;313:670–3.
subcutaneous immunoglobulin and their use in dosing of replacement 33. Anthony RM, Nimmerjahn F, Ashline D, et al. Recapitulation of IVIG
therapy in patients with primary immunodeficiencies. Clin immunol anti-inflammatory activity with a recombinant IgG Fc. Science
2011;139(2):133–41. PubMed PMID: 21353644. 2008;320(373-376).
11. Berger M. Subcutaneous administration of IgG. Immunol Allergy Clin 34. Anthony RM, Wermeling F, Karlsson M, et al. Identification of a receptor
North Am 2008;28(4):779–802, viii. required for the anti-inflammatory activity of IVIG. Proc Natl Acad Sci
12. Lawo J-P, Hubsch A, Rojavin M. Quantification of the wear-off effect USA 2008;105:19571–8.
towards the end of the IVIG infusion interval:Pooled data analysis. 35. Anthony RM, Kobayashi T, Wermeling F, et al. Intravenous
J Allergy Clin Immunol 2014;133(2):AB179. gammaglobulin suppresses inflammation through a novel T(H)2 pathway.
13. Berger M, Jolles S, Orange JS, et al. Bioavailability of IgG administered by Nature 2011;475(7354):110–13.
the subcutaneous route. J Clin Immunol 2013;33(5):984–90. 36. Tackenberg B, Jelcic I, Baerenwaldt A, et al. Impaired inhibitory Fcgamma
14. Ducruet T, Levasseur M-C, Roches A, et al. Pharmacoeconomic receptor IIB expression on B cells in chronic inflammatory demyelinating
advantages of subcutaneous versus intravenous immunoglobulin polyneuropathy. Proc Natl Acad Sci USA 2009;106(12):4788–92.
treatment in a Canadian pediatric center. J Allergy Clin Immunol 37. Campbell IK, Miescher S, Branch DR, et al. Therapeutic effect of IVIG on
2013;131(2):585–7. inflammatory arthritis in mice is dependent on the Fc portion and
15. Fasth A, Nystrom J. Quality of life and health-care resource utilization independent of sialylation or basophils. J Immunol 2014;192(11):
among children with primary immunodeficiency receiving home 5031–8.
treatment with subcutaneous human immunoglobulin. J Clin Immunol 38. Bayry J, Bansal K, Kazatchkine M, et al. DC-SIGN and a2,6-sialylated IgG
2008;28(4):370–8. Fc interaction is dispensable for the anti-inflammatory activity of IVIG.
16. Misbah S, Sturzenegger MH, Borte M, et al. Subcutaneous Proc Natl Acad Sci USA 2009;106:E24.
immunoglobulin: opportunities and outlook. Clin Exp Immunol 39. Yu X, Vasiljevic S, Mitchell DA, et al. Dissecting the molecular mechanism
2009;158(Suppl. 1):51–9. of IVIg therapy: the interaction between serum IgG and DC-SIGN is
17. Ponsford M, Carne E, Kingdon C, et al. Facilitated subcutaneous independent of antibody glycoform or Fc domain. J Mol Biol
immunoglobulin (fSCIg) therapy- practical considerations. Clin Exp 2013;425(8):1253–8.
Immunol 2015;182:302–13. 40. Burns J, Franco A. Ihe immunomodulatory effects off intravenous
18. Wasserman RL, Melamed I, Stein MR, et al. Recombinant human immunoglobulin therapy in Kawasaki disease. Expert Rev Clin Immunol
hyaluronidase-facilitated subcutaneous infusion of human 2015;11(I7):819–25.

CHaPTEr 84 Immunoglobulin Therapy: Replacement and Immunomodulation 1153


41. Kaul R, McGeer A, Norrby-Teglund A, et al. Intravenous immunoglobulin complement-mediated neuronal cell death. Proc Nat Acad Sci
therapy for streptococcal toxic shock syndrome—a comparative 2007;104:14104–9.
observational study. Clin Infect Dis 1999;28:800–7. 55. Ephrem A, Chamat S, Miquel C, et al. Expansion of CD4+CD25+
42. Bayry J, Negi VS, Kaveri SV. Intravenous immunoglobulin therapy in regulatory T cells by intravenous immunoglobulin: a critical factor in
rheumatic diseases. Nat Rev Rheumatol 2011;7:349–59. controlling experimental autoimmune encephalomyelitis. Blood 2008;111:
43. Tha-In T, Bayry J, Metselaar HJ, et al. Modulation of the cellular immune 715–22.
system by intravenous immunoglobulin. Trends Immunol 2008;29: 56. Othy S, Hegde P, Topcu S, et al. Intravenous gammaglobulin inhibits
608–15. encephalitogenic potential of pathogenic T cells and interferes with their
44. Ballow M. The IgG molecule as a biological immune response trafficking to the central nervous system, implicating sphingosine-1
modifier:mechanisms of action of intravenous immune serum globulin phosphate receptor 1-mammalian target of rapamycin axis. J Immunol
in autoimmuen and inflammatory disorders. J Allergy Clin Immunol 2013;190(9):4535–41.
2011;127:315–23. 57. Othy S, Topcu S, Saha C, et al. Sialylation may be dispensable for
45. Kaveri SV, Lacroix-Desmazes S, Bayry J. The antiinflammatory IgG. reciprocal modulation of helper T cells by intravenous immunoglobulin.
N Engl J Med 2008;359:307–9. Eur J Immunol 2014.
46. St-Amour I, Laroche A, Bazin R, et al. Activation of cryptic IgG reactive 58. Trinath J, Hegde P, Sharma M, et al. Intravenous immunoglobulin
with BAFF, amyloid beta peptide and GM-CSF during the industrial expands regulatory T cells via induction of cyclo-oxygenase-2-dependent
fractionation of human plasma into therapeutic intravenous prostaglandin E2 in human dendritic cells. Blood 2013;122(8):1419–27.
immunoglobulins. Clin immunol 2009;133:52–60. 59. Kaufman GN, Massoud AH, Audusseau S, et al. Intravenous
47. Vassilev TL, Kazatchkine MD, Van Huyen J-PD, et al. Inhibition of immunoglobulin attenuates airway hyperresponsiveness in a murine
cell adhesion by antibodies to Arg-Gly-Asp (RGD) in normal model of allergic asthma. Clin Exp Immunol 2011;41(5):718–28.
immunoglobulin for therapeutic use. Blood 1999;11:3624–31. 60. Massoud AH, Guay J, Shalaby KH, et al. Intravenous immunoglobulin
48. Gill V, Doig C, Knight D, et al. Targeting adhesion molecules as a attenuates airway inflammation through induction of forkhead box
potential mechanism of action for intravenous immunoglobulin. protein 3-positive regulatory T cells. J Allergy Clin Immunol 2012;129(6):
Circulation 2005;112:2031–9. 1656–65 e3.
49. Chang J, Shi PA, Chiang EY, et al. Intravenous immunoglobulins reverse 61. Massoud AH, Yona M, Xue D, et al. Dendritic cell immunoreceptor: a
acute vaso-occlusive crises in sickle cell mice through rapid inhibition of novel receptor for intravenous immunoglobulin mediates induction of
neutrophil adhesion. Blood 2008;111:915–23. regulatory T cells. J Allergy Clin Immunol 2014;133(3):853–63 e5.
50. Letko E, Papaliodis DN, Papaliodis GN, et al. Stevens-Johnson syndrome 62. Maddur MS, Vani J, Hegde P, et al. Inhibition of differentiation,
and toxic epidermal necrolysis: a review of the literature. Ann Allergy amplification, and function of human TH17 cells by intravenous
Asthma Immunol 2005;94:419–36. immunoglobulin. J Allergy Clin Immunol 2011;127(3):823–30 e1-7.
51. Viard I, Wehrli P, Bullanim R, et al. Inhibition of toxic epidermal 63. Olivito B, Taddio A, Simonini G, et al. Defective FoxP3 expression in
necrolysis by blockade of CD95 with human intravenous patients with acute Kawasaki disease and restoration by intravenous
immunoglobulin. Science 1998;282:490–3. immunoglobulin therapy. Clin Exp Rheumatol 2010;28(Suppl. 57):
52. Lunemann J, Nimmerjahn F, Dalakas MC. Intravenous imunoglobulin 93–7.
in neurology-mode of action and clinical efficacy. Nat Rev Neurol 64. Tsurikisawa N, Saito H, Oshikata C, et al. High dose intravenous
2015;11(2):80–9. immunoglobulin treatment increases regulatory T cells in patients with
53. Basta M. Modulation of complement-mediated immune damage by eosinophilic granulomatosis with polyangiitis. J Rheumatol 2012;39:
intravenous immune globulin. Clin Exp Immunol 1996;104(Suppl. 1): 1019–25.
21–5. 65. Ballow M. Mechanisms of immune regulation by IVIG. Curr Opin
54. Arumugam TV, Tang S, Lathia JD, et al. Intravenous immunoglobulin Allergy Clin Immunol 2014;14:509–15.
protects the brain against experimental stroke by preventing

CHaPTEr 84 Immunoglobulin Therapy: Replacement and Immunomodulation 1153.e1


MULTIPLE-CHOICE QUESTIONS

1. The BEST strategy for dosing of immunoglobulin (Ig) replace- 3. Which of the following receptors during IVIG treatment of
ment therapy in patients with antibody deficiency diseases autoimmune disease is enhanced on effector macrophages?
is by achieving a serum IgG level: A. CD25
A. Of 500 mg/dL B. FcγRI
B. Of 1000 mg/dL C. FcγRIIB
C. Of 200 mg/dL over baseline D. TACI
D. That minimizes the number of infections
4. Which mechanism of action of IVIG is important in the rapid
2. Thromboembolic events associated with intravenous immu- increase in platelet counts in patients with autoimmune
noglobulin (IVIG) therapy in autoimmune diseases is thought thrombocytopenic purpura (ITP)?
to be caused by: A. Saturation of the FcRn receptors
A. Osmolality of the product B. Neutralization of cytokines
B. Sodium content C. Blocking the uptake of complement on platelets
C. Factor XIa D. Blocking the Fc receptors on macrophages
D. IgA in the product

85







Gene Therapy for Primary Immune
Deficiency Diseases



Caroline Y. Kuo, Donald B. Kohn







Gene therapy as it is being applied for primary immune deficien- and post-transplant immune suppression; essentially, there should
cies (PIDs) represents an autologous hematopoietic stem cell be no risk of graft-versus host-disease (GvHD) from the autolo-
transplant (HSCT), in which a patient’s own stem cells are gous graft (Table 85.1). Additionally, gene therapy could have
genetically corrected and transplanted back (Fig. 85.1). Thus, increased efficacy compared to allogeneic HSCT in some condi-
gene therapy for PID builds upon decades of experience using tions, due to the potential to over-express the relevant gene
allogeneic HSCT from a healthy donor, where replacement of product (e.g. adenosine deaminase enzyme) and lead to a
some or all of a PID patient’s bone marrow hematopoietic stem supra-physiological effect from the engineered graft.
cells (HSCs) with HSC from a healthy donor can be curative of However, gene therapy may have unique risks: the potential
the disease. Although initial efforts in gene therapy were not for the genetic manipulation of the genome of the stem cells by
beneficial for the patients involved, there has been steady progress either gene addition or gene correction causing malignant
with the methods, and there is now clear-cut therapeutic efficacy transformation and leukoproliferative complications. Also, gene
using gene therapy/autologous HSCT for the three most com- therapy could have decreased efficacy if the percentage of
monly transplanted PIDs: severe combined immune deficiency transplanted stem cells that are successfully gene-corrected is
(SCID), Wiskott-Aldrich syndrome (WAS), and chronic granu- low, if the level of expression of the inserted transgene is sub-
lomatous disease (CGD)(Chapters 22, 34). Most gene therapy optimal, or if the process of ex vivo gene manipulation impairs
efforts to date have used gene addition methods in which a the stem cell’s capacity for long-term hematopoiesis. Additionally,
normal copy of the relevant gene is added to the patient’s cells, developing gene therapy for PID will require a separate research
usually using a viral vector. More recently, methods are under endeavor for each genetic etiology (e.g., ADA SCID, XSCID, Rag1
development to perform gene correction, using homologous SCID, Rag2 SCID, etc.), whereas allogeneic HSCT is a more “one
recombination (HR) DNA repair of double-stranded breaks size fits all” approach requiring less individualization for similar
induced near the site of genome mutations by site-specific classes of disorders.
endonucleases.
GENE TRANSFER TO HEMATOPOIETIC STEM CELLS
KEY CONCEPTS
For gene therapy to have an enduring effect in PID, the gene
• Some severe primary immune deficiencies (PIDs) can be treated by addition or correction must occur in the long-term pluripotent
transplantation of hematopoietic stem cells (HSCs), but this is optimal HSCs by some method that will lead to the corrective gene being
with a well-matched immune-compatible donor and may entail immune passed on to the billions of progeny blood cells made from each
complications.
• Gene therapy using autologous HSCs that have been gene-corrected HSC. Gene modification of the far more numerous, but short-
(either added or endogenously-corrected) may avoid the immune lived, progenitor cells would only lead to transient presence of
complications of allogeneic transplant and confer similar benefits. gene-corrected blood cells. The technical challenge is for the
• Stable gene addition to HSC can be done using integrating viral vectors, gene delivery to HSCs to be efficient (high percentage of the
from retrovirus and lentivirus. cells modified), to yield persistent expression, and to have minimal
• Early-phase clinical trials have been performed by applying gene therapy immediate cytotoxicity or long-term genotoxicity.
to PIDs.
• Gene therapy using gamma-retroviral vectors led to immune reconstitu- Many of the gene transfer methods used for research purposes,
tion for several forms of severe combined immune deficiency, Wiskott- such as transfection, electroporation, and nano-particle delivery
Aldrich syndrome and chronic granulomatous disease, but some patients are themselves transient, and the inserted gene would be lost by
developed leukoproliferative complications. dilution as the stem cells proliferate. Thus, most of the studies
• More recent trials using safer vectors are continuing to yield clinical to date have used viral vectors derived from the Retroviridae
efficacy with good safety profiles. family that integrate their genomes into the chromosomes of
• New techniques are being developed for precise gene editing in HSCs, the target cell to achieve persistence of the normal gene (Fig.
which may allow application to a wider spectrum of PIDs.
85.2). Genes delivered by murine gamma-retroviral, human
lentiviral (HIV), spumaviral (foamy) or alpha-retroviral vectors
The key potential advantage of autologous transplant with remain permanently covalently linked to the cellular chromosomal
gene therapy, compared to allogeneic HSCT, is that it may have DNA for stable transmission to progeny cells. The steps involved
reduced risks and a better safety profile, because it eliminates in using these types of vectors to produce a gene-modified HSC
the need for pre-transplant immune-suppressive conditioning graft are described in the Therapeutic Principles box.

1155

1156 ParT TEN Prevention and Therapy of Immunological Diseases








34
34 34





Collect cells from Enrich for stem cells by Transduce cells to Transplant corrected
bone marrow CD34 selection correct gene defect cells into patient
FIG 85.1 Gene Therapy to Treat Primary Immune Deficiencies (PIDs) by Autologous Trans-
plantation of Gene-Corrected Hematopoietic Stem Cells (HSCs). HSC may be obtained from
the bone marrow, by granulocyte–colony-stimulating factor (G-SCF) mobilization and leukapheresis
(peripheral blood stem cells [PBSC]), or from umbilical cord blood of a PID patient. The HSC may
+
be enriched by isolating the CD34 fraction of cells using immunomagnetic separation methods.
+
The CD34 cell population is then cultured for gene manipulation (gene addition or gene correction).
The gene-corrected autologous HSCs are then transplanted back to the patient.


TABLE 85.1 advantages and Disadvantages of allogeneic vs. autologous (Gene Therapy)
HSCT for PIDs

Transplant Type advantage Disadvantage
Allogeneic HSCT Normal function of relevant gene assured; benefit expected if Need suitable matching donor
sufficient donor chimerism
Well-established and long-term experience with benefits/risks Immune: GvHD and rejection risks.
Excellent outcomes with matched sibling donor Requires immune modulation: immune ablative
conditioning; graft manipulation; and GvHD prophylaxis
and treatment.
May contribute to morbidity.
Autologous Gene Patient is donor Potential genotoxicity from gene addition or correction
Therapy HSCT causing cell loss, dysfunction or transformation.
No risks of GvHD. Need to gene-modify high percentage of primary HSC
with minimal toxicity.
May not require immune suppression before (e.g., flu/ATG) Low fractional correction may blunt efficacy.
and after (e.g., corticosteroids, calcineurin inhibitor) HSCT.
Risks of rejection may be less than with allogeneic cells. Prior chemotherapy or marrow dysfunction may preclude
use of autologous stem cells
Transgene over-expression may have benefits (e.g., Immunogenicity of transgene products is not well defined.
adenosine deaminase).
Gene Addition (e.g., Currently showing efficacy for SCID, WAS, and CGD (and for Risks for insertional mutagenesis (gene disruption, gene
lentiviral vector) non-PIDs; e.g., X-ALD, MLD, β-thalassemia). activation) from semi-random insertions into target cell
genomes.
Current generation of SIN lentiviral vectors has significantly Transgene not under normal transcriptional control.
reduced genotoxicity, compared to earlier-used gamma- Function of transgene (level, lineage, longevity) may vary
retroviral vectors. and be variegated.
Gene Correction (e.g., Corrected endogenous gene should have normal function. Risks of local or off-target gene disruption (insertion/
nuclease/HDR) deletion).
Risk of translocations.
CGD, chronic granulomatous disease; GvHD, graft-versus-host disease; HSC, hematopoietic stem cells; HSCT, hematopoietic stem cell transplantation; PID, primary immune
deficiencies; SCID, severe combined immune deficiency; WAS, Wiskott-Aldrich syndrome.


clinical approaches advanced in the past three decades (Table
CLINICAL TRIALS OF GENE THERAPY FOR 85.3). Because each genetic form of PID requires a separate
PRIMARY IMMUNE DEFICIENCIES development path from pre-clinical studies to define efficacy
and safety through clinical trials, each individual PID will be
To date, clinical trials of autologous transplant/gene therapy discussed separately; although they share a common progression
have been performed for five PID disorders — multiple trials through the different eras.
for ADA-deficient SCID, XSCID, CGD, and WAS, and one trial Over these eras, the preferred vector for gene delivery to HSCs
for leukocyte adhesion defect (LAD) (Table 85.2). These have has progressed from murine gamma-retroviral vectors (γ-RV)
occurred over roughly demarcated eras as the technology and to HIV-1–based lentiviral vectors in which the enhancers in the

CHaPTEr 85 Gene Therapy for Primary Immune Deficiency Diseases 1157


γ-Retroviral long-terminal repeats are self-inactivated (SIN vectors). Lentiviral
Lentiviral ψ Gene vectors have several potential advantages over γ-RV, including
Foamy viral the ability to more effectively transduce human HSCs in a shorter
α-Retroviral Package as period of ex vivo culture, the capacity to carry longer and more
pseudotyped vector
Administer Gene addition to complex genetic sequences (such as cellular gene enhancers and
marrow stem cells with vector promoters), and less tendency to insert near the 5′ ends of genes
conditioning which may decrease risks for trans-activating expression of the
Collect, isolate adjacent cellular genes. The combination of recombinant cytokines
stem cells used to activate the HSCs to facilitate transduction was advanced
PID ψ Gene as factors were identified that acted on the earliest HSCs, including
patient Transplant flt-3 ligand and thrombopoietin, combined with c-kit ligand.
gene-modified
stem cells Role of Cytoreductive Conditioning to
Gene correction of stem cells
with site-specific nuclease Facilitate Engraftment
and homologous donor Initial trials of gene therapy for PID did not administer pre-
FIG 85.2 Autologous Transplantation of Gene-Corrected transplant cytoreductive conditioning, due to potential risks of
Hematopoietic Stem Cells (HSC). Gene addition may be per- chemotherapy or radiation with unknown prospects of benefit
formed using Retroviridae-derived vectors (from gamma-retro- from the gene transfer procedure. It is well-known from multiple
viruses, lentiviruses, foamy viruses or alpha-retroviruses) to transplant studies in mice and large animal models that there
transfer a normal copy of a relevant gene into HSCs collected is minimal if any engraftment of autologous HSCs when given
and isolated from a primary immune deficiency (PID) patient. without prior conditioning, unless extraordinary numbers of
The gene-containing vector is packaged in a suitable envelope cells (e.g., 30–50x higher than standard cell dose/kg) are given;
(pseudotyped) for gene addition to human HSC. Alternatively, even mega-dose transplants without conditioning lead to only
the HSC may be gene-corrected using site-specific endonucleases low levels of engraftment (e.g., 1%), although this may be
1
to augment homologous recombination–directed gene correction. persistent. While there was initial reluctance to use conditioning
The patient may receive marrow cytoreductive conditioning to in gene therapy trials when there had not previously been efficacy,
enhance engraftment after transplant of the gene-modified stem now with the clear-cut benefits that may be obtained from gene
cells. therapy, the necessity to use conditioning for autologous gene
therapy HSCT (which is less than that needed for allogeneic
HSCT) is becoming recognized as the standard. The amount
and types of chemotherapy drugs used for conditioning has
varied, depending on the disease setting. For SCID, where a
relatively low level of gene-corrected HSCs may support immune
reconstitution, reduced-intensity conditioning (RIC), such as
THEraPEUTIC PrINCIPLES relatively low dose busulfan alone (e.g. 4–6 mg/kg), may be
Steps for Gene Transfer to Hematopoietic Stem sufficient. For other disorders where less of a selective advantage
Cells for Clinical Transplantation may exist for the gene-corrected cells, a higher level of engraftment
of modified HSC may be needed, and thus stronger conditioning
Package as Pseudotyped Vector regimens have been used, reaching levels for myeloablative
Transfect packaging cell with vector plasmid and virion/envelope conditioning (MAC) (e.g., busulfan, 12–16 mg/kg). For the WAS
plasmids. where it may be necessary to ablate the pre-transplant immunity
Collect released vector from cell culture medium (DNAse). to reduce risks for post-transplant autoimmunity, immune-
Purify and concentrate vector (e.g., ion-exchange chromatography, suppressive drugs (e.g., fludarabine, rituximab) have been added
tangential flow filtration).
Aliquot. Certify. Release. to conditioning regimens. These combined iterative approaches
to improving gene therapy have led to the current state, where
Transduce Stem Cells With Vector clinical benefits are being routinely achieved, as detailed below
+
Isolate CD34 stem/progenitor cells from clinical source (bone marrow, for each disorder.
mobilized peripheral blood, cord blood). New methods are being explored to “make space” in the
+
Grow CD34 cells in serum-free medium plus recombinant cytokines marrow using alternatives to chemotherapy and radiation, such
(e.g., ckit ligand, flt-3 ligand, thrombopoietin). as monoclonal antibodies to markers present on HSCs. These
Add vector to cells and culture for transduction. are moving toward clinical assessments and may in the future
Link vector sequences covalently into stem cell’s chromosomal DNA.
Formulate and characterize cell product for release. facilitate engraftment with lower short-term and long-term risks
than with chemotherapeutic agents.
administer Marrow Cytoreductive Conditioning
May deliver single or combination chemotherapeutic agents or monoclonal Adenosine Deaminase (ADA)-Deficient Severe
antibodies to “make space” in marrow. Combined Immune Deficiency (SCID; Chapter 35)
The first clinical trial of gene therapy for an inherited disorder
Transplant Gene-Modified Stem Cells (other than a premature attempt at gene therapy for beta-
Infuse gene-modified cell product intravenously. thalassemia) was directed against adenosine deaminase (ADA)-
Stem cells engraft and transmit transgene to all progeny blood cells. deficient Severe Combined Immune Deficiency (SCID).
Transgene produces necessary gene product to correct genetic ADA-SCID was the first form of human SCID for which the
deficiency.
responsible gene was identified and cloned, allowing gene therapy

1158 ParT TEN Prevention and Therapy of Immunological Diseases



TABLE 85.2 Clinical Trials of Gene Therapy for Primary Immune Deficiencies
PID Investigator(s) Year Vector/Target Conditioning NCT #
ADA SCID Blaese, Anderson, Culver 1990 LASN (MLV LTR) None —-
PBMC
Bordignon, Mavilio 1992 G1ADA None —-
(MLV LTR)
PBMC and BM
Hoogerbrugge, Valerio 1993 MLV-ADA None —-
BM CD34 +
Kohn, Parkman 1993 G1NA-ADA (MLV LTR) None —-
UCB CD34 +
Onodera/Sakiyama 1995 LASN None —-
(MLV LTR)
PBMC
Aiuti/Roncarolo 1998 G1ADA Busulfan 4 mg/kg 00599781
(MLV-LTR)
BM CD34 +
Gaspar/Thrasher 1999 SFFV-ADA-wpre Melphalan; busulfan —-
BM CD34 +
Otsu/Ariga 2003 GCsap-M-ADA BM CD34 + None —-
Kohn/Candotti 2001 MND-ADA and GCsap-M-ADA None (4). —-
BM CD34 + Busulfan (6)
GlaxoSmithKlein 2008 G1ADA Busulfan 00598481
BM CD34 +
Kohn/Candotti 2009 MND-ADA Busulfan 90 mg/m2 00794508
BM CD34 +
Gaspar 2012 EFS-ADA Busulfan 01380990
+
CD34 BM/PBSC
Kohn 2013 EFS-ADA Busulfan 01852071
BM CD34 +
XSCID Cavazzana-Calvo/ Fischer 1998 MFG-IL2Rg None —-
BM CD34 +
Thrasher/Gaspar 1999 MLV-IL2Rg None —-
BM CD34 +
Thrasher/Fischer/ 2011 SIN γRV EFS-IL2Rg None Fr: 01410019
Cavazzana/Pai/ Williams BM CD34 + UK: 01175239
US: 01129544
Sorrentino 2012 EFS-IL2Rg-Ins None/ Busulfan 01512888
BM CD34 +
Malech/DeRavin 2011 EFS-IL2Rg-Ins Busulfan 01306019
CD34 PBSC
+
LAD Hickstein, Bauer 1999 MLV-CD18 None —-
BM
CGD Malech 1995 MLV-p47 None —-
+
CD34 PBSC
Malech 1998 MLV-gp91phox None —-
Ott/Grez 2009 SFFV-gp91phox 8 mg/kg 00927134
CD34 PBSC
+
Kang/Malech 2010 MT-gp91phox 10 mg/kg —-
+
CD34 PBSC
Thrasher/ Serve/ 2013 pChim-gp91phox (G1XCGD) pK adjusted, ~12 mg/kg 01855685
Reichenbach CD34 PBSC
+
Kohn/Williams/Kang 2015 pChim-gp91phox (G1XCGD) pK adjusted, ~12 mg/kg 02234934
+
CD34 PBSC
WAS Klein 2008 SFFV-WAS gRV Busulfan —-
Aiuti 2011 1.6hWASP-WPRE LV Bu/Flu/Rituximab 01515462
Hacein-Bey Abina… 2011 Bu/flu (rituximab or alemtuzimab 02333760
Cavazzana for auto-immunity)
Thrasher 2011 Bu/Flu 01347242
Pai 2011 Bu/Flu 01410825
CGD, chronic granulomatous disease; LAD, leukocyte adhesion defect; PID, primary immune deficiencies; SCID, severe combined immune deficiency; WAS, Wiskott-Aldrich
syndrome; XSCID, X-linked SCID.

CHaPTEr 85 Gene Therapy for Primary Immune Deficiency Diseases 1159



TABLE 85.3 Eras in the advancement of Clinical Gene Therapy for PIDs
Predominant Conditioning Typical
Era Vector Growth Factors regimen PIDs Treated Outcomes
Early γ-RV IL3/IL6/SCF None ADA XSCID CGD LAD Insufficient engraftment.
(~1990–1999) No efficacy.
Middle γ-RV SCF/FLT3L/TPO RIC-MAC ADA XSCID CGD WAS Efficacy.
(~1998–2006) Genotoxicity in some.
Current SIN LV SCF/FLT3L/TPO RIC-MAC ADA XSCID CGD WAS Efficacy.
(~2007–present) Good safety profile.



2
to be approached. As a ubiquitously-expressed housekeeping lymphopenic SCID patients. Indeed robust reconstitution of T
enzyme, the non-regulated expression of an introduced ADA cell immunity was achieved, although there were variable
gene was expected to be tolerated and potentially beneficial. responses of B cells. However, after 2-3 years from treatment,
Also, the ability to treat ADA-SCID with a bone marrow transplant several of the patients developed a severe complication from the
from a matched sibling donor without the use of cytoreductive retroviral vector of leukemia-like leukoproliferation. New vectors
or immune suppressive conditioning was taken to imply that were developed that lacked the strong enhancer elements of the
there is a selective advantage for ADA-replete T lymphocytes retroviral vectors that caused the insertional oncogenesis (self-
over ADA-deficient cells and that only a modest number of inactivating or “SIN” vectors). One published study using a SIN
engrafted gene-corrected HSC may provide clinical benefit. gamma-retroviral vector demonstrated immune restoration
A series of clinical trials were performed using gamma- without any evidence of leukoproliferation. New studies are in
14
retroviral vectors and targeting either peripheral blood T cells progress using lentiviral vectors. The first of these studies treated
or bone marrow hematopoietic stem cells. While initial studies a group of XSCID patients who had undergone allogeneic
in the early 1990’s did not achieve clinical efficacy, subsequent transplant some years earlier, but who had not achieved complete
trials that applied reduced-intensity condition prior to transplant immune reconstitution and had significant morbidity from poor
15
to increase engraftment of gene-corrected HSCs have led to growth and development. Following gene therapy using a
immune restoration in the majority of more than 40 treated lentiviral vector and non-myeloablative conditioning, all have
3-6
patients, without vector-related complications. The relatively improved general well-being, with development of sufficient
low dose chemotherapy and absence of graft versus host disease antibody production to be able to stop immunoglobulin replace-
makes these autologous transplants well-tolerated with essentially ment therapy. Studies treating more typical XSCID infants are
none of the clinical side-effects seen in allogeneic HSCT with now underway at several centers in the US and Europe. Thus,
conditioning. One of the retroviral vectors used in studies at the ability of gene therapy to provide clinical benefit to these
Telethon Gene Therapy Program for Genetic Diseases, of the difficult patients is indeed gratifying.
Hospital San Raffaele, in Milan Italy was licensed by GSK and
has received licensure approval in the European Union, a major Leukocyte Adhesion Defect (LAD; Chapter 22)
advance for gene therapeutics which is available for patients. Bauer and Hickstein performed gene therapy for two LAD patients
Other ongoing clinical trials in the US and the UK are using a using a gamma-retroviral vector expressing human CD18 to
16
lentiviral vector for ADA SCID with excellent initial clinical results transduce bone marrow cells without conditioning. There were
in terms of efficacy and safety. Thus, gene therapy for ADA SCID no beneficial effects of gene therapy and both later succumbed
has become a major treatment option for patients, as the efficacy to complications of their immune deficiency. These investigators
and safety profile have been favorable and may exceed those of have developed a highly effective vector from the spumavirus
matched unrelated or haplo-identical transplants, although there called Human Foamy Virus and a clinical trial is under
have not been randomized controlled trials. development. 17

X-linked SCID (Chapter 35) Chronic Granulomatous Disease (CGD; Chapter 22)
The second genetic form of human SCID for which the relevant Chronic Granulomatous Disease (CGD) is a rare primary
7,8
gene was identified and cloned is the X-linked form (XSCID). immunodeficiency (1 in 200,000 live births in the US), first
The responsible gene, IL2R γ , encodes the common cytokine termed Fatal Granulomatous Disease of Childhood in 1959 to
receptor chain (or γ c ), a component of several multimeric recep- describe individuals affected by recurrent, invasive bacterial and
18
tors for a family of lymphopoietic cytokines: IL-2, IL-4, IL-7, fungal infections complicated by granuloma formation. Clinical
IL-9, IL-15 and IL-21. Defects of γ c severely impair the develop- presentation is explained by defects in any of the components
ment of multiple components of the immune system and patients that comprise the nicotinamide adenine dinucleotide phosphate
typically have severely reduced numbers of T and NK cells, and (NADPH) oxidase complex leading to phagocyte dysfunction.
may have immature but non-functional B cells. Retroviral- In its fully assembled form, the NADPH oxidase complex is
mediated gene transfer of a normal human IL2R γ cDNA was composed of five proteins, two of which are membrane-bound
shown to restore cytokine-induced signaling activity and lym- (gp91phox and p22phox) and three of which are cytosolic
phocyte function in patients’ cells and in murine models. 9-12 (p47phox, p67phox, and p40phox). All five components are
XSCID patients were first treated by gene therapy using necessary for proper NADPH oxidase function without which
5,13
gamma-retroviral vectors. No conditioning was given, relying affected individuals are particularly predisposed to infections
on the potent selective survival and proliferation advantage with Aspergillus species, Staphylococcus aureus, Burkholderia
gene-corrected lymphoid cells were expected to have in cepacia, Serratia marcescens, Nocardia species, and less commonly,

1160 ParT TEN Prevention and Therapy of Immunological Diseases


Salmonella and BCG infections outside of North America. Defects was obtained, gamma-retroviral vectors were constructed and
in gp91phox encoded by CYBB account for the most common shown to correct several manifestations of the disorder in patient
X-linked form of CGD while the remaining defects are inherited derived cells and in murine models. However, it is not fully
19
in an autosomal recessive (AR) pattern. Since its initial descrip- known what levels of WAS transgene expression are needed to
tion almost six decades ago, CGD has evolved from a disease safely and effectively correct the major manifestations of
with early mortality to one with relatively good outcomes and lymphocyte and platelet dysfunction. A concern has been raised
multiple treatment options. that sub-optimal levels or low frequency of expression of WAS
While prophylactic antimicrobial and immunomodulatory protein (WASP) could allow auto-immunity to develop. If
agents have dramatically decreased infection rates in CGD patients, correction is only partial, (e.g., some proportion of B cells are
curative therapy can only be achieved with HSCT. Transplant not gene corrected and have defective auto-regulatory function),
outcomes have continued to improve over the years, with good auto-immunity may occur. 16
results reported even in patients with high-risk features such as A first trial of gene therapy for WAS used a gamma-retroviral
intractable infections and auto-inflammation using reduced- vector with a very potent long terminal repeat promoter enhancer
13
intensity conditioning protocols. Nevertheless, allogeneic HSCT (from the Spleen Focus Forming Virus). G-CSF-mobilized
can still be complicated by graft-versus-host disease and pre- peripheral blood stem cells were used as the HSC source and
existing infections and is not preferred for those without an conditioning with myeloablative dosages of busulfan was given
HLA-matched stem cell donor. For these patients, autologous prior to transplant. There were excellent initial results in terms
HSCT with gene-modified cells is becoming a more viable and of immune reconstitution and platelet counts, demonstrating
realistic option. that gene therapy can be therapeutic for this disorder. However,
The first gene therapy trials for CGD began in the mid-1990’s there was subsequently a very high incidence of acute leukemia
again using gamma-retroviral vectors. As with ADA SCID, initial among these patients, developing within a few years from treat-
trials in which pre-transplant conditioning was not used did ment, with ALL, AML or both occurring in a shocking 7 of 9
17
not lead to efficacy as there was minimal if any engraftment subjects. The mechanism of insertional oncogenesis from the
of gene-corrected stem cells. In subsequent studies where gamma-retroviral vector was clearly the cause, with the leukemias
non-myeloablative conditioning with busulfan was used, showing clonal vector integrations adjacent to known proto-
increasingly higher levels of engraftment of corrected stem oncogenes, such as LMO2 and MDS1.
cells was achieved. The best and worst of these studies was one While that trial was being initiated, work was ongoing through
using a gammaretroviral vector with a potent transcriptional a multi-center collaboration in the EU to develop a SIN lentiviral
control element to drive high level expression of the gp91phox vector for WAS that uses the promoter from the WAS gene per
gene. The three treated subjects had initial development of se to drive expression of the WAS cDNA. Extensive pre-clinical
neutrophils with restored oxidase function and cleared severe studies showed the efficacy of this vector to improve immunologic
resistant infections. However, insertional oncogenesis occurred and hematologic parameters in murine and human disease
in this study also, and myelodysplasia or frank myeloid leukemia models. 18,19 And, it displayed significantly lower risks for geno-
developed. More recently, trials are using a lentiviral vector with a toxicity than the type of gamma-retroviral vector used in the
myeloid-specific transcriptional control element intended to drive first trial.
gp91phox expression in mature myeloid cells, where it is needed, The lentiviral vector with the WAS promoter has now been
but to not have activity in HSCs that are the likely targets for used in parallel but independent phase I/II clinical trials per-
transformation. Initial results are showing safety and evidence of formed in several countries. 20,21 Relatively high intensity condition-
efficacy. 14 ing was administered to attempt to obtain high-level engraftment
Recently, the group at NIH has also investigated the use of of gene-corrected HSCs, with some differences among the centers
site-specific endonucleases (a zinc-finger nuclease, discussed in the precise conditioning regimen used. All used moderate
below) to target integration of a normal gp91phox cDNA expres- doses of busulfan (pharmacokinetically [pK]-adjusted) and
15
sion cassette to the AAVS1 gene “safe harbor site”. Good levels fludarabine, and variably, rituximab or other serotherapy agents.
of engraftment were demonstrated in mouse models, with up These trials have uniformly demonstrated efficacy and safety.
to 11% of human cell expressing gp91phox in vivo. This work Findings have included clinical improvement in general health,
represents the first step towards targeted gene editing for X-CGD bleeding incidents, and eczema, and as expected for an autologous
and suggests that it can also be possible to integrate the transgene transplant, no problems from graft versus host disease. Most
in its natural location in the genome under control of its have had good recovery of T, B, NK cell numbers and functions.
endogenous promoter. However, there has been only modest and variable improvements
for platelet levels (e.g. 20-60,000/uL), with evidence that higher
WISKOTT-ALDRICH SYNDROME (WAS; transplanted dose of transduced cells led to higher platelet
20
Chapter 35) counts. No new onset of auto-immunity has been reported,
although in some cases pre-existing problems have persisted. In
The Wiskott Aldrich Syndrome (WAS), initially described as fact, decreases of indices of auto-immunity and improved B cell
22
an X-linked syndrome in kindreds in Germany and the US, tolerance have been documented after gene therapy for WAS.
presents with multiple clinical manifestations, including the Polyclonal vector integration patterns were seen, with no reported
classical triad of immune deficiency, eczema and thrombocy- development of clonal expansions or frank leukoproliferative
topenia. The complex immune deficiency involves defects of T, complications. Integration site patterns resemble those seen in
B, NK and antigen-presenting cells. The identified WAS gene other trials using lentiviral vectors into human HSCs, with highly
encodes a 501 amino acid proline-rich protein that has multiple diverse vector integration sites and no predilection for cancer-
identified functional domains, placing it at the nexus of intracel- related genes, significantly different than the pattern seen in the
lular signaling and cytoskeleton control. Once the WAS cDNA gamma-retroviral vector trials.

CHaPTEr 85 Gene Therapy for Primary Immune Deficiency Diseases 1161


These results are promising and of significant clinical benefit deficiency caused by hypogammaglobulinemia, the high rate of
and safety, but would be better if more normal levels of platelets other clinical complications that CVID patients may experience
could be achieved. It is possible that the level of expression of necessitate new treatments. However, gene therapy requires
WASP from the WAS gene promoter in the lentiviral vector is knowing the responsible pathogenic gene, and has been limited
inadequate to support normal platelet production or survival. to monogenic disorders. To date, a single gene defect has not
If so, introducing more than one copy of the vector per cell been identified in the majority of patients with CVID. There are
could lead to higher levels of WASP and higher platelet counts. some known CVID genes, including TNFRSF13B (encoding TACI
The WAS gene promoter that has been used as a regulatory 8–10%), TNFRSF13C (encoding BAFF-R), COS, CD19, and
element (a 1.6 kb fragment from the 5′ end of the gene) may MSH5, which in total may be responsible for 10–15% of all
be sub-optimal to drive sufficient expression; there may be other CVID patients. A separate gene therapy project would be needed
regulatory elements from the WAS genomic locus not included to develop treatment for each causal gene, through the full
in the vectors that are needed for some essential aspect of expres- spectrum from pre-clinical activities to clinical trial performance.
23
sion (lineage, level, longevity). Astrakhan et al reported that a And, because most of these known CVID-causing genes are
lentiviral vector using a stronger internal promoter (a retroviral involved in cell stimulation and signaling, they may require
LTR) to drive the WASP cDNA led to better restoration of T regulated, rather than ubiquitous, constitutive expression for
and B cell activity in the WAS gene knock-out mouse model. safety. Because of these constraints, it is not currently possible
Alternatively, the absolute number of gene-corrected HSCs to apply gene therapy for the majority of patients with CVID.
engrafted may mediate the platelet levels and a higher percentage There has been growing recognition of numerous immune
of transduced cells or higher numbers of gene-corrected cells dysregulation and auto-inflammatory syndromes due to auto-
may support higher platelet levels. Relative risks of multiple somal dominant gain-of-function mutations (e.g. STAT3, MEFV,
26
integrants versus a stronger promoter are not known. As for the IL-1, NFkB, interferon pathways). The pathology attributed to
other disorders discussed here, direct gene correction should blood cells (as opposed to those effects on other tissues) may
yield normal, physiologic expression of the WAS gene. be corrected by HSCT, and here too autologous transplant/gene
therapy may have advantages over allogeneic. Gene modification
GENE THERAPY CONSIDERATIONS FOR OTHER PID of a patient’s HSC by addition of a shRNA cassette or gene disrup-
tion using site-specific endonucleases may suppress expression
Currently, a new development project is needed to bring gene of the dominant gene. And, direct correction of the pathogenic
therapy to clinical application for each individual PID-causing mutation could also be beneficial, as discussed below. It is likely
locus, such as the more than 20 human genes that may cause that the gene modification would need to be efficient to yield a
SCID, 5 or more CGD loci, several for Hemophagocytic Lym- high fractional correction of the engrafting stem cells. In this
phoistiocytosis (HLP), X-linked lymphoproliferative disease setting, some pre-transplant immune suppression may be needed
(XLP), etc. Each gene and disease setting poses different challenges to ablate pre-existing auto-immunity. And, perhaps most chal-
in terms of necessary gene transfer efficiency, level of expression, lenging for gene therapy are the PIDs that also have major somatic
need for regulation of the transferred gene, safety considerations, or developmental problems, such as chromosomal abnormalities,
and measurable end-points. ataxia-telangiectasia, and others. Here the gene therapy with the
X-linked agammaglobulinemia (XLA; Chapter 34) is another HSC may benefit that component of the disorder but would not
logical disease to consider treating by autologous transplant with address the others; systemic delivery of genes or delivery to the
gene therapy, since normal B cell development from HSC with CNS is under study, but it is not yet sufficiently efficient for
a normal Bruton tyrosine kinase (BTK) gene should correct the most clinical needs.
immune deficiency. Because of the good clinical effects from
immunoglobulin replacement therapy for XLA and the toxicities GENE CORRECTION (EDITING) FOR GENE
from HSCT, especially from chemotherapy and GVHD, HSCT THERAPY OF PID
is rarely done for XLA patients. A few XLA patients have had
allogeneic transplants from healthy donors and have developed A major paradigm shift in gene therapy is under way as methods
B cell reconstitution. to perform precise edits of cells’ genomes are being developed.
Gene therapy studies in BTK gene knock-out mice have shown As an alternative to the semi-random insertion of normal copies
that it can lead to immune reconstitution, using lentiviral vectors of the relevant gene delivered by a viral vector, as discussed for
with B lymphoid-specific promoters. 24,25 While no adverse effects all of the studies above, techniques are being established to either
were seen from constitutive expression of the BTK gene in these correct specific bases in the DNA, or to insert (or remove) gene
murine studies, these do not constitute formal toxicology studies sequences at specific sites by harnessing cellular DNA repair
which would be needed before clinical applications. It is likely pathways. These DNA repair mechanisms normally correct the
that regulated expression of BTK, rather than constitutive, many double-stranded DNA breaks that occur during DNA
ubiquitous expression, is needed for highest efficacy and safety. replication or from environmental genotoxic agents (ionizing
In theory, lentiviral viral vectors using components of the BTK radiation, chemicals). To simplify a highly complex topic, there
gene transcriptional regulatory sequences could yield vectors are two major DNA repair pathways for rejoining the sequences
with the desired expression specificity. Alternatively, gene cor- that flank a double-stranded DNA break: non-homologous end
rection of the BTK gene, using the methods discussed below, joining (NHEJ) and homologous recombination (HR). NHEJ
could restore precise BTK expression regulation and may have reconnects the broken ends of the chromosomes in a way that
the greatest safety profile. often leads to insertion or deletion of DNA bases (indels) at the
Common Variable Immune deficiency (CVID) comprises the junctional site. This is, in essence, a mutagenic process and
most common severe human PID (Chapter 34). While immu- may be used to disrupt genes to knock-out their activity,
noglobulin replacement therapy can ameliorate the immune examples being the HIV-1 co-receptor CCR5, dominant-active

1162 ParT TEN Prevention and Therapy of Immunological Diseases


Codon#: 1 2 3 4 5 6 7 8 9 Gene Sequence-specific
(1) Wild-type 5’-ATG CCTTGA AATTCG GGGCGA TTGACC-3’ endonuclease Mutation
Gene 3’-TAC GGAACT TTAAGC CCCGCT AACTGG-5’ Promoter
1 2 3 4 5
(2) Mutant 5’-ATG CCTTGA AAT ACG GGGCGA TTGACC-3’
Gene 3’-TAC GGAACT TTA TGC CCC GCTAAC TGG-5’
1 2 34 5 pA Homology arm flanked,
(3) Repair template 5’-GA AAT TCG GGGC-3’ codon-optimized cDNA/pA
Donor
(4) Corrected 5’-ATG CCTTGA AAT TCG GGG CGATTG ACC-3’
Gene 3’-TAC GGAACT TTA AGC CCC GCTAAC TGG-5’
FIG 85.3 Site-Specific Gene Repair by Homologous Recom- Inserted cDNA transcribed
from endogenous promoter
bination (HR). In this example, instead of the wild-type gene Mutation
sequence (1), a patient’s gene (2) has a mutation from a base-pair Promoter 1 2 3 45 pA 1 2 3
substitution of an A for a T at the start of the 5th codon (red).
An artificial donor template (3), here as a single-stranded AAAAA
deoxyoligonucleotide of 12 bases in length (green), is provided FIG 85.4 Site-Specific Insertion of a cDNA to Override Down-
with the correct base present at the site of the patient’s mutation Stream Gene Mutations. A prototypical gene is shown with 5
(blue). If the donor template is used to repair a double-stranded exons (yellow boxes 1–5) and an upstream promoter (red box).
DNA break induced near the mutation by a site-specific endo- A mutation in exon 2 (black X) inactivates the gene. A sequence-
nuclease (red arrow), sequences from the donor (green) will be specific endonuclease (red arrow) is designed to introduce a
incorporated into the patient’s gene (4), introducing the normal double-stranded DNA break, in this case in the 5′ untranslated
corrective base pair (blue). region of the gene (blue and green lines). A cDNA molecule
contains the contiguous exons of the gene (orange 1–5), codon-
optimized to increase expression and to eliminate homology
with the endogenous exons to eliminate illegitimate recombination
transcriptional factors such as CTLA-4, or STAT3 Gain of Function events, and a polyadenylation signal (pA) to terminate transcrip-
alleles, a transcriptional repressor of fetal hemoglobin, etc. There tion. The cDNA is flanked by the sequences from the endonucle-
have been clinical trials for patients with HIV infection in which ase cleavage site (blue and green lines). The donor can be inserted
the gene for CCR5 HIV co-receptor was disrupted using a zinc into the nuclease target site by homologous recombination,
finger nuclease (ZFN) to make a double stranded DNA break placing the cDNA under transcriptional control of the endogenous
while NHEJ was allowed to repair the break, leading to indels gene promoter; the cDNA transcript (red arrow) would override
that inactivated the gene and co-receptor expression. 27 any downstream mutations in the gene.
HR is a more precise repair mechanism that normally uses a
copied sister chromosome or the other homologue as a template
to repair the break; sequences of the template are copied into
the repair site and if there are differences, the template acts as a Clustered, Regularly Interspaced, Short Palindromic Repeats
donor for the new sequences. For gene correction, an artificial (CRISPRs) that allow the introduction of a double-stranded
donor template is provided to the cells to instruct the introduc- DNA break at unique sites in the mammalian genome with high
tion of the intended sequence changes (Fig. 85.3). Beyond its specificity. Current methods introduce the nuclease into HSCs
use for modification of single base pairs, as illustrated here, in a method where it will only be present transiently, such as
HR can be used to introduce whole gene sequences into the electroporation of in vitro transcribed messenger RNA encoding
target site by flanking a gene cassette with “homology arms” the nuclease proteins. The homologous donor template is
that consist of the DNA sequences homologous to the target site introduced as either a short (e.g. 50–100 bases) single-stranded
(Fig. 85.4). deoxy-oligonucleotide that is co-electroporated with the nuclease,
Although HR has been used to introduce genes into cells, or as a longer sequence carried by a viral vector that does not
such as murine embryonic stem cells to make gene knockout integrate into the target cell chromosome, including adeno-
and knockin mice, it is generally a low-frequency event (occurring associated virus (AAV) or an integrase-defective lentiviral vector
6
4
in 1/10 –1/10 cells) and requires the use of selectable markers (IDLV). The development and application of this technology
to isolate the rare desired recombinant. Whereas cloning and has evolved at an incredibly rapid pace and is approaching the
expansion of murine (and human) embryonic stem cells can be levels of efficiency needed for clinical applications.
done to produce populations of the rare recombinant cells, this One caveat is that potential off-target activity of the nucleases
is not possible with primary HSCs, which cannot be expanded could cause genotoxicity from either disruption of unintended
to any great degree from single cells. Methods achieving high genes or even introduction of chromosomal translocations
efficiency of gene modification with low cytotoxicity in large between two cut sites produced simultaneously, e.g., one on-target
numbers of primary stem cells are needed for clinical applications and one off-target. Current studies are assessing the consequences
to autologous HSCT. The major breakthrough in this area comes of this potential genotoxicity in human HSCs, while ongoing
from the observation that HR is vastly more frequent when a work is seeking to improve the specificity of the nucleases to
double-stranded DNA break is introduced close to the target eliminate or greatly minimize off-target activity.
28
site; then, the repair donor molecule can be used at efficiencies Gene correction will have several advantages over gene
in the range ~1–50% of treated cells. addition. It would avoid the potential problems from randomly
Several classes of designer site-specific endonucleases have inserting vectors that may disrupt or over-stimulate adjacent
been derived, including ZFN, introduced above; Transcription cellular genes, as discussed above for the retroviral vectors.
Activator-Like Effector Nucleases (TALENs); and more recently Crucially, correcting the endogenous gene keeps its expression

CHaPTEr 85 Gene Therapy for Primary Immune Deficiency Diseases 1163


under normal physiological control. This will be essential for gene using an AAV vector to express the SERPING1 gene encoding
therapy of many PIDs, where it could be problematic if the gene the C1 inhibitor. 30
is expressed constitutively from a viral vector. For example, it was
shown that retroviral vector delivery of a normal CD40 ligand ADVANCING GENE THERAPY FOR PID FROM
gene corrected murine models of X-linked hyper-IgM syndrome EXPERIMENTAL TO STANDARD OF CARE
(XHIM), but it subsequently led to the development of lympho-
mas in the mice from constitutive rather than regulated CD40 Initial trials of gene therapy for PIDs were all performed at
29
ligand expression. Bruton tyrosine kinase (BTK), defective in tertiary-level academic medical centers, often with federal or
X-linked agammaglobulinemia, similarly may need to be expressed disease-foundation research grant funding, and they tested initial
specifically at certain stages of B-lymphocyte development for hypotheses concerning safety and efficacy. These centers may
safety and efficacy. A long list of other loci involved in PIDs continue to perform early phase clinical trials for specific dis-
may similarly be best approached by gene correction, including orders, especially where there is local expertise on the disease
SLAM, XIAP, JAK3, FOXP3, IL-10, IL7Ralpha, TACI, CTLA4, being studied. However, there is an ongoing transition to
etc. While current gene correction techniques are likely below industry-sponsored trials focused on drug product development,
the frequency of efficiency needed to yield clinical benefits for as the effective vectors and the gene-modified stem cell products
most disorders, this field is moving at a lightning pace with new they compose are advanced to licensure and marketing as
advances being reported weekly in the activity and specificity pharmaceutical agents. A common model used by these new
of a whole host of nucleases and other genome editing tools. gene therapy companies is one of central processing, at one or
a few high-grade commercial GMP facilities. The autologous
USE OF PLURIPOTENT STEM CELLS AS A SOURCE patient cells are procured at their home institution (by leuko-
OF HSC FOR GENE THERAPY OF PID pheresis or bone marrow harvest), shipped to the central process-
ing site for genetic manipulation and cryopreservation, and then
The establishment of human pluripotent stem cells (hPSCs), returned to be administered locally. Currently, gene therapy
initially as human embryonic stem cells (hESCs) and subsequently transplants have been performed at a limited number of clinical
as induced pluripotent stem cells (iPSCs), has brought the promise trial sites due to the cell processing expertise needed; presumably
of novel models to study human diseases (“disease in a dish”) under the commercial model, it may be done at any suitable
and to provide renewable sources of patient-compatible cells for hematopoietic stem cell transplant center, with the company
cellular therapies. The essentially unlimited ability to expand selling the processed cell product, like a medical device or an
hPSCs and their capacity to produce any of the cell-types in the unrelated stem cell product. Alternatively, self-contained cell
body has led to investigations to harness them for regenerative processing and manipulation devices are being developed that
medicines. In the treatment of PIDs, hPSC could provide an could allow gene modification of stem cells to be done at
ideal target to produce autologous HSCs with precise gene cor- individual institutions, without requiring highly trained staff or
rection. While techniques have been developed to perform the high-grade GMP facilities.
gene modification strategies that may be sufficiently robust for Major issues remain to be determined about cost and
clinical applications, the major roadblock is the current inability reimbursement for gene therapy. Effective gene therapy for the
to produce clinically-relevant numbers of transplantable HSCs severe diseases being approached would be expected to lead to
from hPSCs. The HSC is a relatively evanescent state and it large lifetime savings in medical costs. The one-time price can
has not been possible to “freeze” differentiation from hPSC be compared to the costs faced by the patient encumbered by
at that state, although it has been possible to proceed right the progressive nature of the underlying disease, to the costs
through the HSC stage to produce relatively pure populations for long-term protein-based therapies and possibly even to the
of individual mature blood cells. As with gene correction, the costs for allogeneic transplantation. However, those one-time
pace of scientific progress with these phenomenal cells is proceed- costs will be expensive for clinical gene therapy transplants
ing rapidly, and it is likely that new sources of gene-corrected using pharmaceutical-grade vectors and commercial-level cell
autologous HSCs will be available for clinical transplants in the processing with the attendant quality control. Thus, a single large
near future. expenditure for gene therapy may eventually be cost-effective
compared to ongoing medical costs; however, the method
GENE THERAPY FOR PID INVOLVING SERUM of financing the large up-front charge, at least in the United
PROTEIN DEFICIENCIES States with its multiple insurance companies, remains to be
determined.
Whereas many of the severe PIDs are due to blood cell defects
and thus responsive to HSCT, others result from deficiencies of CONCLUSION
serum proteins, such as complement components. Here, gene
therapy may be approached, as is being done for hemophilia, In the past decades, gene therapy for PIDs has gone from a
by direct in vivo administration of a gene-containing vector that dream for the future to a clinical reality. Gene therapy for ADA-
can permanently insert the gene into target tissues–such as the deficiency SCID has been safe and effective in most treated
liver, skeletal muscle, or endothelium–that can serve as protein patients. Although initial trials of gene therapy for XSCID, CGD
sources. In vivo gene delivery is being studied using retroviral and WAS showed efficacy, they were marred by an unacceptably
and lentiviral vectors or adeno-associated virus (AAV) vectors, high rate of complications from genotoxicity associated with
and methods for in vivo gene correction using site-specific gamma-retroviral vectors. New vectors are now in clinical trials
nucleases and homologous donor sequences are also under and are showing excellent safety profiles and higher efficacy in
development. To this end, Crystal and co-workers have recently clinical trials for ADA-deficient SCID, XSCID, WAS, and CGD
reported studies in a murine model of hereditary angioedema as well as other non-PID disorders. Approaches to direct gene

1164 ParT TEN Prevention and Therapy of Immunological Diseases


correction are being developed that may broaden indications 12. Bauer TR Jr, Hickstein DD. Gene therapy for leukocyte adhesion
for PIDs that may be treated by gene therapy (e.g., XHIM, XLA, deficiency. Curr Opin Mol Ther 2000;2:383–8.
etc.). Thus, the continued efforts of scientists and physicians to 13. Güngör T, Teira P, Slatter M, et al. Reduced-intensity conditioning and
develop gene therapy are leading to a new therapeutic modality, HLA-matched haemopoietic stem-cell transplantation in patients with
ideally to permanently and safely cure these diseases. chronic granulomatous disease: a prospective multicentre study. Lancet
2014;383:436–48. Available from: http://www.ncbi.nlm.nih.gov/
pubmed/24161820.
ON THE HOrIZON 14. Santilli G, Almarza E, Brendel C, et al. Biochemical correction of X-CGD
• Continued expansion of the genetic types of PID being treated by by a novel chimeric promoter regulating high levels of transgene
gene addition using integrating vectors (e.g. other forms of SCID, expression in myeloid cells. Mol Ther 2011;19:122–32.
other forms of CGD, LAD, HLH, etc.). 15. De Ravin SS, Reik A, Liu P-Q, et al. Targeted gene addition in human
• Development of safe and effective marrow conditioning regimens CD34+ hematopoietic cells for correction of X-linked chronic
that are not chemotherapy-based (e.g. monoclonal antibodies) granulomatous disease. Nat Biotechnol. Nature Publishing Group
• Application of gene editing to broader spectrum of PIDs: e.g. X-linked 2016;(February):1–8. Available from: http://www.nature.com/doifinder/
agammaglobulinemia, X-linked Hyper-IgM syndrome (CD40 ligand 10.1038/nbt.3513.
deficiency), RAG1-deficient SCID, Gain of Function STAT1, IPEX). 16. Becker-Herman S, Meyer-Bahlburg A, Schwartz MA, et al. WASp-deficient
• Understand the molecular pathogenesis of more PID (e.g. CVID, auto- B cells play a critical, cell-intrinsic role in triggering autoimmunity. J Exp
immune) and develop effective gene therapy approaches. (Some may Med 2011;208:2033–42.
require more than just HSC correction, e.g. ataxia telangiectasia). 17. Boztug K, Schmidt M, Schwarzer A, et al. Stem-cell gene therapy for the
• Production of autologous HSC by cellular reprogramming coupled to Wiskott-Aldrich syndrome. N Engl J Med 2010;363:1918–27.
effective expansion of HSC 18. Charrier S, Dupré L, Scaramuzza S, et al. Lentiviral vectors targeting
WASp expression to hematopoietic cells, efficiently transduce and correct
Please check your eBook at https://expertconsult.inkling.com/ cells from WAS patients. Gene Ther 2007;14:415–28. Available from:
for self-assessment questions. See inside cover for registration http://www.nature.com/doifinder/10.1038/sj.gt.3302863%5Cnpapers3://
details. publication/doi/10.1038/sj.gt.3302863.
19. Scaramuzza S, Biasco L, Ripamonti A, et al. Preclinical safety and
REFERENCES efficacy of human CD34+ Cells transduced with lentiviral vector for the
treatment of Wiskott-Aldrich syndrome. Mol Ther 2013;21:
1. Stewart FM, Crittenden RB, Lowry PA, et al. Long-term engraftment of 175–84. Available from: http://dx.doi.org/10.1038/mt.2012.23/
normal and post-5-fluorouracil murine marrow into normal nature06264.
nonmyeloablated mice. Blood 1993;81:2566–71. 20. Hacein-Bey Abina S, Gaspar HB, Blondeau J, et al. Outcomes following
2. Blaese RM, Culver KW, Miller AD, et al. T lymphocyte-directed gene gene therapy in patients with severe Wiskott-Aldrich syndrome. JAMA
therapy for ADA- SCID: initial trial results after 4 years. Science 2015;313:1550–63. Available from: http://jama.jamanetwork.com/
1995;270:475–80. article.aspx?doi=10.1001/jama.2015.3253.
3. Candotti F, Shaw KL, Muul L, et al. Gene therapy for adenosine 21. Aiuti A, Biasco L, Scaramuzza S, et al. Lentiviral hematopoietic stem cell
deaminase – deficient severe combined immune deficiency : clinical gene therapy in patients with Wiskott-Aldrich syndrome. Science
comparison of retroviral vectors and treatment plans. Blood 2013;341:1233151. Available from: http://www.ncbi.nlm.nih.gov/
2012;120:3635–46. pubmed/23845947.
4. Cicalese MP, Ferrua F, Castagnaro L, et al. Update on the safety and 22. Pala F, Morbach H, Castiello MC, et al. Lentiviral-mediated gene therapy
efficacy of retroviral gene therapy for immunodeficiency due to restores B cell tolerance in Wiskott-Aldrich syndrome patients. J Clin
adenosine deaminase deficiency. Blood 2016;128:45–54. Invest 2015;125:3941–51. Available from: http://www.pubmedcentral
5. Shaw KL, Sokolic R, Davila A, et al. Phase II clinical trial of gene therapy .nih.gov/articlerender.fcgi?artid=4607131&tool=pmcentrez&rendertype
for adenosine deaminase-deficient severe combined immune deficiency =abstract.
(ADA-SCID). Mol Ther 2014;22(Suppl. 1):S107. 23. Astrakhan A, Sather BD, Ryu BY, et al. Ubiquitous high-level gene
6. Candotti F, Johnston JA, Puck JM, et al. Retroviral-mediated gene expression in hematopoietic lineages provides effective lentiviral gene
correction for X-linked severe combined immunodeficiency. Blood therapy of murine Wiskott-Aldrich syndrome. Blood 2012;119:
1996;87:3097–102. Available from: http://www.ncbi.nlm.nih.gov/ 4395–407.
pubmed/8605322. 24. Ng YY, Baert MRM, Pike-Overzet K, et al. Correction of B-cell
7. Cavazzana-Calvo M, Hacein-Bey S, de Saint Basile G, et al. Role of development in Btk-deficient mice using lentiviral vectors with
interleukin-2 (IL-2), IL-7, and IL-15 in natural killer cell differentiation codon-optimized human BTK. Leukemia 2010;24:1617–30.
from cord blood hematopoietic progenitor cells and from gamma c 25. Kerns HM, Ryu BY, Stirling BV, et al. B cell-specific lentiviral
transduced severe combined immunodeficiency X1 bone marrow cells. gene therapy leads to sustained B-cell functional recovery in a
Blood 1996;88:3901–9. murine model of X-linked agammaglobulinemia. Blood 2010;115:
8. Hacein-Bey-Abina S, Hauer J, Lim A, et al. Efficacy of gene therapy for 2146–55.
X-linked severe combined immunodeficiency. N Engl J Med 26. Hagin D, Burroughs L, Torgerson TR. Hematopoietic Stem Cell
2010;363:355–64. Available from: http://www.pubmedcentral.nih.gov/ Transplant for Immune Deficiency and Immune Dysregulation Disorders.
articlerender.fcgi?artid=2957288&tool=pmcentrez&rendertype=abstract. Immunol Allergy Clin North Am 2015;35:695–711.
9. Gaspar HB, Cooray S, Gilmour KC, et al. Long-term persistence of a 27. Tebas P, Stein D, Tang WW, et al. Gene editing of CCR5 in autologous
polyclonal T Cell pepertoire after gene therapy for X-linked severe CD4 T cells of persons infected with HIV. N Engl J Med 2014;370:
combined immunodeficiency. Sci Transl Med 2011;3:97ra79. 901–10. Available from: http://www.pubmedcentral.nih.gov/articlerender.
10. Hacein-Bey-Abina S, Pai S-Y, Gaspar HB, et al. A modified γ-retrovirus fcgi?artid=4084652&tool=pmcentrez&rendertype=abstract.
vector for X-linked severe combined immunodeficiency. N Engl J Med 28. Porteus MH, Baltimore D. Gene Targeting in Human Cells. Science
2014;371:1407–17. Available from: http://www.nejm.org/doi/abs/ 2003;300(May):75390.
10.1056/NEJMoa1404588. 29. Brown MP, Topham DJ, Sangster MY, et al. Thymic lymphoproliferative
11. De Ravin SS, Wu X, Moir S, et al. Lentiviral hematopoietic stem cell gene disease after successful correction of CD40 ligand deficiency by gene
therapy for X-linked severe combined immunodeficiency. Sci Transl Med transfer in mice. Nat Med 1998;4:1253–60.
2016;8:335ra57. Available from: http://stm.sciencemag.org/ 30. Pagovich OE, Qui T, Whaley AS, et al. One-time gene therapy for
content/8/335/335ra57.figures-only. hereditary angioedema. Mol Ther 2016;24(Suppl. 1):S298.

CHaPTEr 85 Gene Therapy for Primary Immune Deficiency Diseases 1164.e1


MULTIPLE-CHOICE QUESTIONS

3-5 multiple choice review questions - these questions should 3. Which are true about gene editing:
have explanations for the correct answer. 1. Gene editing may be preferred to gene addition for primary
immune deficiencies where the relevant gene needs to be
1. Which of the following vector types is effective for high
efficiency permanent gene addition to hematopoietic stem expressed under precise regulation.
cells: 2. Cells can repair double-stranded DNA breaks by two
A. Adeno-associated virus (AAV) predominate methods: Non Homologous End Joining
B. Gamma-retroviral (NHEJ), which tends to introduce insertions and deletions
C. Plasmid – by transfection or electroporation (indels) at the chromosome repair junction, and Homolo-
D. Lentiviral gous Recombination (HR), which can copy genetic
E. Adenovirus information from a homologous donor sequence into the
1. A, B, C repair junction.
2. B and D 3. The first human clinical trial using gene editing disrupted
3. A-D the gene encoding an essential HIV-1 co-receptor protein
4. All are correct in autologous T cells.
4. The introduction of a double-stranded DNA break increases
2. Which of the following primary immune deficiencies have the frequency of homologous recombination for site-specific
shown good clinical responses to gene therapy: gene editing.
A. Wiskott-Aldrich Syndrome (WAS) 5. Gene editing methods can site-specifically disrupt, correct
B. X-linked Severe Combined Immune Deficiency (XSCID) or introduce a gene.
C. X-linked Chronic Granulomatous Disease (XCGD) 1. A, B, C
D. X-linked Agammaglobulinemia (XLA) 2. B and D
E. Common Variable Immune Deficiency (CVID) 3. A-D
1. A, B, C 4. All are correct
2. B and D
3. A-D
4. All are correct