Clinical Immunology_ Principles and Practice ( PDFDrive )

79









Lymphomas



Stefania Pittaluga, Tapan Bhavsar, Elaine S. Jaffe






The classification of malignant lymphomas continues to undergo KEY CONCEPTS
revision based on insights gained through the application of Lymphoma
immunological and molecular techniques and the application of
these discoveries to individualized therapeutic approaches. Early • Classification consists of a list of individual disease entities defined
classifications were based on morphological characteristics of the by morphological, immunophenotypic, genetic, and clinical features.
neoplastic elements; however, with increasing knowledge of the • Neoplastic cells are related to the postulated normal counterpart,
complexity of the immune system, a more functional approach was when possible.
sought. Differentiation schemes provided a useful starting point for • Histological grade should be applied within individual diseases.
understanding lymphomas (Fig. 79.1). High-throughput genomic • Clinical factors for individual patients, as measured by the International
Prognostic Index (IPI) and gene expression profiling, are useful in
studies of both tumor RNA and tumor DNA have been applied to predicting clinical outcome.
lymphomas to define their molecular signatures with the aim of
improving the understanding of oncogenic pathways and their clinical
implications. These studies have led to new prognostic and diagnostic
tools, and as a result, more targeted therapies have emerged. 1

Splenic marginal Classic Hodgkin DLCBL (ABC-type) CLL
zone lymphoma lymphoma PMBCL mutated V -gene


Acute lymphoblastic
leukemia Marginal zone
Memory B cell

Mantle Crippled
zone B-cell

MALT lymphoma
Germinal
NaÏve B cell center
Lymphoplasmacytic
lymphoma

CLL Plasma cell
unmutated V-gene
Plasmablast

Mantle cell Follicular lymphoma NLPHL DLCBL (GCB-type) Multiple myeloma
lymphoma Burkitt lymphoma

V-gene recombination Clonal expansion, somatic mutation Class switching Differentiation
FIG 79.1 Normal B-Cell Differentiation in Relation to a Secondary B Follicle, Mutational
Stages of the Immunoglobulin Genes, and Cellular Counterparts for B-Cell Lymphomas.
Simplified version of B-cell development indicates points at which V-gene recombination, clonal
expansion, and somatic mutations occur in relation to a secondary B follicle. B-cell lymphomas
are related to different stages of B-cell differentiation and function. ABC-type, activated B-cell-like
type; CLL, chronic lymphocytic leukemia/lymphoma; DLBCL, diffuse large B-cell lymphoma;
GCB-type, germinal-center B-cell-like type; MALT lymphoma, marginal zone B-cell lymphoma of
mucosa-associated lymphoid tissue (MALT) type; NLPHL, nodular lymphocyte-predominant Hodgkin
lymphoma; PMBCL, primary mediastinal B-cell lymphoma.

1065

1066 ParT EIGhT Immunology of Neoplasia


The guiding principles of the World Health Organization (GEP) in lymphomas has generated distinct molecular “signatures”
(WHO) classification of neoplasms of the hematopoietic and for a variety of disease entities, in some cases corresponding
lymphoid tissues, published in 2001 and updated in 2008 and more closely to different stages of lymphoid differentiation
2
2016, were based on the Revised European American Classification and in other instances offering insights into mechanisms of
of Lymphoid Neoplasms (REAL), which was published by the neoplastic transformation with delineation of alterations in
International Lymphoma Study group in 1994. The focus was specific pathways. The advent of whole-genome sequencing
on the identification of individual diseases based on an integration contributes additional new insights into pathogenetic mechanisms.
of morphological, immunophenotypic, genetic, and clinical Recently, a revision of the nearly 8-year-old WHO classification
features. The recent application of gene expression profiling of lymphoid neoplasms (Table 79.1) was published clarifying

TABLE 79.1 WhO Classification of Tumors of hematopoietic and Lymphoid Tissues (2016)
B-Cell Neoplasm Primary effusion lymphoma
Precursor B-cell lymphoblastic leukemia/lymphoma HHV8 positive DLBCL, NOS*
B-lymphoblastic leukemia/lymphoma, not otherwise specified Burkitt lymphoma
B-lymphoblastic leukemia/lymphoma with recurrent genetic Burkitt-like lymphoma with 11q aberration*
abnormalities High grade B-cell lymphoma, with MYC and BCL2 and/or BCL6
rearrangements*
Mature B-Cell Neoplasm High grade B-cell lymphoma, NOS*
B-cell lymphoma unclassifiable, with features intermediate between
Chronic lymphocytic leukemia/small lymphocytic lymphoma diffuse large B-cell lymphoma and classic Hodgkin lymphoma
Monoclonal B-cell lymphocytosis*
B-cell prolymphocytic leukemia T-Cell Neoplasm
Splenic B-cell marginal zone lymphoma
Hairy cell leukemia Precursor T-cell lymphoblastic leukemia/lymphoma
Splenic B-cell lymphoma/leukemia, unclassifiable
Splenic diffuse red pulp small B-cell lymphoma Mature T-Cell and NK-Cell Neoplasms
Hairy cell leukemia-variant T-cell prolymphocytic leukemia
Lymphoplasmacytic lymphoma T-cell large granular lymphocytic leukemia
Waldenstrom macroglobulinemia Chronic lymphoproliferative disorder of NK-cells
Monoclonal gammopathy of undetermined significance (MGUS), IgM* Aggressive NK leukemia
+
µ heavy-chain disease Systemic EBV T-cell lymphoma of childhood*
γ heavy-chain disease Hydroa vacciniforme-like lymphoproliferative disorder*
α heavy-chain disease Adult T-cell leukemia/lymphoma
Monoclonal gammopathy of undetermined significance (MGUS), IgG/A* Extranodal NK-/T-cell lymphoma, nasal type
Plasma cell myeloma Enteropathy-associated T-cell lymphoma
Solitary plasmacytoma of bone Monomorphic epitheliotropic intestinal T-cell lymphoma*
Extraosseous plasmacytoma Indolent T-cell lymphoproliferative disorder of the GI tract*
Monoclonal immunoglobulin deposition diseases* Hepatosplenic T-cell lymphoma
Extranodal marginal zone lymphoma of mucosa-associated Subcutaneous panniculitis-like T-cell lymphoma
lymphoreticular tissue (MALT) lymphoma Mycosis fungoides
Nodal marginal zone lymphoma Sézary syndrome
+
Pediatric nodal marginal zone lymphoma Primary cutaneous CD30 positive T-cell lymphoproliferative disorders
Follicular lymphoma Primary cutaneous anaplastic large-cell lymphoma
In situ follicular neoplasia* Lymphomatoid papulosis
Duodenal-type follicular lymphoma* Primary cutaneous γ/δ T-cell lymphoma
Pediatric-type follicular lymphoma* Primary cutaneous CD8 positive aggressive epidermotropic cytotoxic
Large B-cell lymphoma with IRF4 rearrangements* T-cell lymphoma
+
Primary cutaneous follicle-center lymphoma Primary cutaneous acral CD8 T-cell lymphoma*
Mantle cell lymphoma Primary cutaneous CD4 positive small/medium T-cell lymphoproliferative
In situ mantle cell neoplasia * disorder*
Diffuse large B-cell lymphoma (DLBCL), NOS Peripheral T-cell lymphoma, NOS
Germinal-center B-cell type* Angioimmunoblastic T-cell lymphoma
Activated B-cell type* Follicular T-cell lymphoma*
T-cell/histiocyte rich large B-cell lymphoma Nodal peripheral T-cell lymphoma with TFH phenotype*
Primary DLBCL of the central nervous system Anaplastic large-cell lymphoma, ALK positive
Primary cutaneous DLBCL, leg type Anaplastic large-cell lymphoma, ALK negative*
EBV DLBCL, NOS* Breast implant-associated anaplastic large-cell lymphoma*
+
+
EBV mucocutaneous ulcer*
DLBCL associated with chronic inflammation hodgkin Lymphoma
Lymphomatoid granulomatosis Nodular lymphocyte predominant Hodgkin lymphoma
Primary mediastinal (thymic) large B-cell lymphoma Classic Hodgkin lymphoma
Intravascular large B-cell lymphoma Nodular sclerosis Hodgkin lymphoma
ALK positive large B-cell lymphoma Lymphocyte-rich classic Hodgkin lymphoma
Plasmablastic lymphoma Mixed cellularity classic Hodgkin lymphoma
Large B-cell lymphoma arising in HHV8-associated multicentric Lymphocyte-depleted classic Hodgkin lymphoma
Castleman disease
*Refers to changes from the 2008 classification.
Note: More common entities are underlined. Provisional entities in italics.
From Swerdlow SH, Campo E, Pileri SA, et al. The 2016 revision of the World Health Organization (WHO) classification of lymphoid neoplasm. Blood 2016;127:2375–90.

ChaPTEr 79 Lymphomas 1067


the diagnosis and management of lesions at the very early stages which occasionally can manifest plasmacytoid differentiation.
of lymphomagenesis, refining diagnostic criteria for some entities, Many but not all patients with LPL may have clinical evidence
and detailing the expanding genetic/molecular landscape of of Waldenström macroglobulinemia (WM), which is based on
numerous lymphoid neoplasms and their clinical correlates. 3 the detection of an IgM monoclonal gammopathy of any con-
This chapter focuses on the classification of neoplasms derived centration and is associated with bone marrow involvement by
from mature B cells, T cells, and natural killer (NK) cells, with LPL (Chapter 80).
emphasis on malignant lymphomas. Other chapters in this volume In LPL, cells have surface and cytoplasmic Ig, usually IgM
cover lymphoid leukemias (Chapter 78) and immunosecretory (and usually lacking IgD), and express B cell–associated antigens
disorders, including plasma cell neoplasms (Chapter 80). Special (CD19, CD20, CD22, CD79a). They are generally negative for
attention is devoted to the impact of clinical features (e.g., age CD5 and cyclinD1, distinguishing LPL from CLL and MCL,
and anatomical site) on disease definition and a greater apprecia- respectively. CD25, CD10, or CD11c may be weakly expressed
tion of early events in neoplastic transformation. These early in some cases. The postulated normal counterpart is thought to
lesions can sometimes be detected in otherwise healthy individuals, be a postfollicular medullary cord B cell, based, in part, on the
and these lesions may or may not progress to overt lymphoma presence of somatic mutations in the Ig heavy-chain and light-
or leukemia. These entities appear to carry fewer genetic aber- chain variable region genes.
rations compared with the conventional forms of the disease, The recent identification of MYD88 L265P mutation (found
which perhaps explains their indolent clinical behavior. 4 in approximately 90% of WM cases) has become a reliable marker
5
supporting a diagnosis of LPL. This mutation is also found in
a significant proportion of IgM, but not IgG or IgA, monoclonal
gammopathy of undermined significance (MGUS) cases.
CLINICaL PEarLS
Indolent Lymphomas Mantle-Cell Lymphoma
MCL usually presents in adults (median age 62; variable male
• Natural history: survival measured in years predominance) with advanced-stage disease—involving lymph
• Least sensitive to therapy nodes, Waldeyer ring lymphoid tissue, spleen, bone marrow, and
• Good response to low-dose oral alkylating agents, radiotherapy, and
steroids, but no curability peripheral blood. Gastrointestinal (GI) tract involvement is
• Higher response rate and complete remission with combination of common and is associated with a lymphomatous polyposis picture.
standard chemotherapy and anti-CD20 monoclonal antibody Retrospective studies have shown a poor prognosis (median
• Gene expression profiling can help identify patients who might benefit survival 3–5 years), with a high relapse rate following initial
from high-dose chemotherapy and autologous stem-cell transplantation, remission. MCL is composed of small lymphoid cells with slightly
which is a potentially curative modality
irregular nuclear contours, finely clumped chromatin, and scant
cytoplasm. Specifically, blastoid and pleomorphic variants have
been associated with a more aggressive clinical course. The
+
+
+
MATURE B-CELL NEOPLASMS postulated normal counterpart is a CD5 “naïve” sIgM and sIgD
B cell, found in peripheral blood and in the mantles of reactive
KEY CONCEPTS follicles.
MCL is characterized by a molecular hallmark, t(11;14)
Somatic Mutation in Relation to Normal (q13;q32)—involving cyclin D1 (CCND1) and the IGH genes;
B-Cell Development an overexpression of CCND1 is believed to be essential in the
+
• Premutational stage: circulating naïve B cells (immunoglobulin (Ig)M / pathogenesis. Rare variants negative for CCND1 but with similar
6
+
D ) before antigen exposure immunomorphology and GEP have also been identified. Half
• Stage of somatic mutation, clonal expansion, and isotype switch: at of the CCND1 expression/rearrangement-negative forms have
the germinal center CCND2 translocations, often with IGK or IGL as a partner
7
• Postmutational stage: selected B cells move to the periphery (post– locus, which could be of diagnostic utility. SOX11 is overex-
germinal center) or to the recirculating pool (memory B cells), or pressed in most CCND1-positive and CCND1-negative cases.
8
undergo terminal differentiation (plasma cells)
Besides the CCND1/IGH translocation, additional alterations
involving other cell cycle regulatory proteins (RB, p53, CDK
inhibitors) have been described in the more aggressive forms
Lymphoplasmacytic Lymphoma of MCL.
Lymphoplasmacytic lymphoma (LPL) is a disease of adult life In-situ mantle cell neoplasia (ISMCN) (according to new
(median age in the 60s), usually presenting with generalized 2016 classification; previously “in-situ” MCL) describes clonal
lymphadenopathy, constitutional symptoms, and splenomegaly. proliferation of cyclin D1-positive cells restricted to the inner
Histologically, there is a diffuse interfollicular proliferation mantle cuffs in an otherwise reactive lymph node/lymphoid
of small lymphocytes (many with plasmacytoid features) and tissue and usually represents an incidental finding. Some cases
2
plasma cells, with or without immunoglobulin (Ig)–filled will eventually progress to overt MCL ; however, the risk of
intranuclear inclusions (Dutcher bodies) and sparing of the progression is difficult to ascertain, as the number of reported
sinuses. An increased number of mast cells and iron-laden cases is few. Another newly identified nonnodal variant is
macrophages can be seen. Although many B-cell neoplasms characterized by a leukemic phase without nodal disease but
occasionally show maturation to plasmacytoid or plasma cells often long-standing splenomegaly. These cases develop from
with cytoplasmic Ig, the term LPL should be restricted to tumors IgHV-mutated SOX11-negative B cells, carry t(11;14) with few
lacking features of other well-defined entities, such as chronic additional chromosomal abnormalities but lack expression
lymphocytic leukemia (CLL) or mantle-cell lymphoma (MCL), of SOX11. 9

1068 ParT EIGhT Immunology of Neoplasia


The proliferation rate based on Ki67-positivity has been Surface Ig is positive, most commonly with IgM expression, but
considered of prognostic relevance. More recently, GEP using IgG or IgA can also be seen in many cases. CD10 and BCL6 are
genes involved in cell cycle progression and DNA synthesis has positive, but CD5 is usually negative.
identified a proliferation signature—defining different prognostic The 2008 WHO classification had recognized variants of
groups and showing correlation with cytological subtype to some FL, namely, pediatric FL, GI tract FL, and other extranodal FL.
extent. For example, a high proliferation rate has been shown Pediatric-type FL (now a definite entity in the 2016 classification)
in the blastoid variant. is more common in males and presents as localized nodal disease.
The treatment approach for newly diagnosed patients with It typically has an expansile serpiginous pattern, with relatively
MCL depends on their eligibility for stem cell transplantation monotonous, medium-sized, blastoid cellular composition. BCL2,
(SCT). However, purging of the neoplastic cells is difficult to BCL6, and MYC rearrangements are lacking. Complete remissions
achieve because of frequent bone marrow involvement, and may be obtained with either surgical excision or local radiation
autologous SCT has not proven be curative. The incorporation therapy. Some studies have raised the possibility that pediatric-
of rituximab into chemotherapeutic regimens has become an type FL might be a “benign clonal proliferation with low malignant
evidence-based standard of care. Of the current drugs used for potential.” 11,12
MCL, the application of the BTK inhibitor ibrutinib might change The WHO classification recognizes “FL in situ” (now termed
treatment paradigms by obviating the need for transplantation in-situ follicular neoplasia [ISFN]) as a distinctive lesion. It should
10
in younger patients and chemotherapy in older patients. The be distinguished from partial involvement of FL. ISFN shows
patients with the nonnodal variant as described above do not involvement of germinal centers by CD10 and BCL2-positive
appear to require aggressive chemotherapy. 9 cells carrying t(14;18) in an otherwise reactive lymph node, which
is typically an incidental finding. Patients have a very low risk
Follicular Lymphoma of progression, but ISFN may be detected in association with
Follicular lymphoma (FL) is the most common subtype of other forms of B-cell lymphoma, necessitating additional clinical
13
non-Hodgkin lymphoma (non-HL) in the United States and assessment. Fewer chromosomal abnormalities are noted in
14
accounts for approximately 45% of all newly diagnosed cases. IFSN in comparison with partial or overt FL. Patients lacking
It has a peak incidence in the fifth and sixth decades and is rare evidence of FL at staging have a low risk of developing the disease;
under the age of 20 years. Both sexes are equally affected. Most this phenomenon appears to represent the tissue counterpart of
patients have stage 3 or 4 disease at diagnosis, with generalized circulating clonal B-cells carrying t(14;18) as detected in healthy
lymphadenopathy and bone marrow involvement. Approximately individuals. A higher level of circulating t(14;18) positive lym-
–4
10% of patients have circulating malignant cells; appropriate phocytes (>10 of total cells) indicates a higher risk for FL.
immunophenotypic or molecular analyses may reveal involvement The new 2016 classification addresses GI tract FL as duodenal-
of peripheral blood in a higher proportion of patients. type FL, as these can also be seen elsewhere in the GI tract and
FL is composed of varying proportions of follicle center–type have features overlapping with ISFN and extranodal mucosa-
14
cells, centrocytes, and centroblasts, with centroblasts representing associated lymphoreticular tissue (MALT) lymphoma. These
the proliferative component. According to the WHO classification, present as small mucosal polyps or nodules, and most cases
all low-grade FL are combined into a single category, grade are discovered incidentally on endoscopy. The lesions are
1/2—with an overall predominance of centrocytes and fewer usually low-grade and have an indolent clinical course, and most
than 15 centroblasts per high-power field (hpf). Grade 3 (with patients have been managed without therapy. Local recurrences
>15 centroblasts/hpf) is further subdivided into 3A and 3B, based in the intestine may occur, but spread beyond the small intestine
on the presence or absence of background centrocytes. FL is rare.
represents the neoplastic counterpart of the reactive germinal- Primary cutaneous follicle center lymphoma frequently lacks
center cells; intraclonal heterogeneity with high numbers of the BCL2 translocation and BCL2 expression. However, when
somatic and ongoing Ig mutations can be detected in the neo- BCL2 expression is detected, the possibility that this may represent
plastic cells, as in the normal counterparts. Biologically, grade a secondary site of involvement should be considered. These
3B is more closely related to diffuse large B-cell lymphoma lesions are generally managed locally with conservative approaches
(DLBCL) compared with FL. and have a low risk of spread beyond the skin. The scalp is a
The vast majority of FL (approximately 90%) is associated common site of presentation.
with a t(14; 18) involving rearrangement of the BCL2 gene. This Two recent variants or subtypes of FL have been described;
translocation appears to result in constitutive expression of BCL2 one subtype described by Karube et al. occurring in older
−
+
protein, which is capable of inhibiting apoptosis in lymphoid individuals showed a CD10 /MUM1 immunophenotype that
cells. The cells of FL accumulate and are at risk for secondary tends to show grade 3 morphology, usually lacks BCL2 transloca-
15
mutations, which may be associated with histological progression. tions, and often shows BCL6 abnormalities. This is in contrast
It is thought that the BCL2 translocation occurs at a very early to another recently described distinct new provisional entity of
stage of B-cell development, during immunoglobulin gene a low-stage large B-cell lymphoma with IRF4 rearrangement
rearrangement. Mutations in BCL2 can lead to loss of BCL2 common in children or young adults, typically involving the
protein in the presence of the translocation; fluorescence in situ Waldeyer ring and/or the cervical lymph nodes. These cases show
hybridization (FISH) studies for t(14; 18) in these cases are strong IRF4/MUM1 expression, usually with BCL6 and a high
informative. Of note, t(14; 18) is not specific to FL and can be proliferative fraction, and are considered more aggressive than
seen in CLL and other low-grade B-cell lymphomas as well. other pediatric-type FL; however, when treated, they show good
16
Grade 3B FL is less commonly associated with BCL2 translocation response. Another distinctive subtype described by Katzenberger
17
and carries genetic aberrations more commonly seen in DLBCL. et al. is characterized by t(14;18)–negative nodal grade 1/2 FL
The neoplastic cells in FL have a mature B-cell phenotype with with a high proliferation rate that predominantly shows diffuse
expression of the B-cell antigens (CD19, CD20, and CD22). growth pattern and deletions in the chromosomal region 1p36.

ChaPTEr 79 Lymphomas 1069


This entity usually presents with low clinical stage and large or (q32;q21), t(3;14)(q27;q32), and t(3;14)(p14.1;q32); these
localized inguinal tumors. 17 abnormalities are observed with variable frequency, often depend-
19
FL is indolent but is still incurable with available therapeutic ing on the anatomical site. Although several genes are involved
modalities. The occurrence of BCL2 translocation at a very in these translocations, at least three of them—t(11;18), t(1;14),
early stage of B-cell development might be contributory and t(14;18)—share a common pathway, leading to the activation
20
to the difficulty of eradicating the neoplastic clone with chemo- of NFκB and its downstream targets. By genome-wide DNA
therapy. Clinical parameters have been used to develop prognostic profiling integrated with GEP, differences were detected among
indexes (e.g., the Follicular Lymphoma International Prognostic the three main types of marginal zone lymphomas, lending
Index, adjusted from the International Prognostic Index for support to the current WHO classification that separates these
18
aggressive B-cell lymphoma). Recently, much has also been learned three entities. The translocation t(11;18)(q21;q21) is associated
about the mutational landscape of FL. Mutations in CREBBP, exclusively with low-grade extranodal MALT and is not detected
MLL2, and EZH2 have been shown as extremely common in cases with concurrent low-grade and high-grade tumors, or
18
early events and may be potential therapeutic targets. The in primary extranodal large-cell lymphomas—raising doubts
natural history of the disease is associated with histological whether these primary extranodal lymphomas are, in fact, related
progression at both cellular composition and overall pattern to low-grade MALT. The term extranodal marginal zone lymphoma
levels (Fig. 79.2). (MZL) of MALT type applies only to low-grade MALT; the term
high-grade MALT should not be used for extranodal large B-cell
Mucosa-Associated Lymphoid Tissue Lymphomas lymphomas in a MALT site.
Most lymphomas of marginal zone derivation present in extra- There is a strong association between chronic infection
nodal sites and have histopathological and clinical features that with Helicobacter pylori and gastric MALT lymphoma. Other
are part of the spectrum of MALT lymphomas. MALT lymphomas infectious agents have been described in MALT lymphomas
occur most frequently in the stomach, lung, thyroid gland, salivary involving skin (Borrelia burgdorferi), ocular adnexae (Chlamydia
gland, and lacrimal gland. Other less common sites of involvement psittaci), and the small intestine (Campylobacter jejuni);
include the orbit, breast, conjunctiva, bladder, kidney, and thymus. however, a causal relationship has not yet been demonstrated.
MALT lymphomas are characterized by a heterogeneous cellular Chronic antigen stimulation is critical to both the development
composition, including centrocyte-like cells, monocytoid B cells, of a MALT lymphoma and the maintenance of the neoplastic
small lymphocytes, and plasma cells. In most cases, large trans- state. Indeed, in some cases lacking aforementioned genetic
formed cells are uncommon, but reactive germinal centers are aberrations, eradication of H. pylori by antibiotic therapy
nearly always present. Historically, the distinction from reactive has led to the spontaneous remission of gastric MALT
lesions has been problematic. Clonality can be established on lymphoma. MALT lymphomas are positive for B cell–associated
the basis of light-chain restriction or molecular studies. Follicular antigens but are usually negative for CD5 and CD10. BCL6 and
colonization by the neoplastic cells can simulate FL. The clinical CD10 are helpful markers to identify residual reactive germinal-
course is usually quite indolent. MALT lymphomas tend to relapse center cells, especially in cases of follicular colonization. The
in other MALT-associated sites. MALT lymphomas of the salivary putative cell of origin of MZL is a post–germinal-center memory
gland, thyroid gland, and mediastinum (thymus) are usually B cell.
associated with a history of autoimmune diseases, predominantly
Sjögren syndrome. Nodal Marginal Zone Lymphoma
MALT lymphomas have several recurring cytogenetic abnor- Nodal marginal zone lymphoma (NMZL), a primary nodal
malities, including t(11;18)(q21;q21), t(1;14)(p22;q32), t(14;18) disease, resembles other marginal zone lymphomas, extranodal
or splenic types. Adult patients often present with bone marrow
involvement and tend to have a more aggressive clinical course
compared with those with extranodal MALT. In contrast, when
NMZL occurs in children, it shows a striking male predominance,
presence of disrupted follicles with progressive transformation
of germinal centers, presents with localized (head and neck)
disease, and can be managed with local therapies. 19
The neoplastic proliferation is mostly composed of small- to
medium-sized B cells, often with pale cytoplasm. The immuno-
phenotype is similar to that of other marginal zone B-cell
−
−
+
lymphomas, CD20 , CD5 , CD10 , with variable IgD expression.
Some NMZLs have a morphology and immunophenotype similar
to those of splenic marginal zone lymphomas (SMZLs) (see
+
below), and in these cases, the tumor cells are IgD .
Splenic Marginal Zone Lymphomas
SMZLs present in adults and have a slight female gender predilec-
tion, usually with splenomegaly but without peripheral lymph-
adenopathy. Most patients have bone marrow involvement with
FIG 79.2 Follicular Lymphoma. The neoplastic follicles are modest lymphocytosis. Some evidence of plasmacytoid differentia-
similar in size and are partially surrounded by lymphoid cuffs. tion and the presence of a low level M-protein may also be seen.
In contrast to reactive germinal centers, they lack polarization The course is reportedly indolent, and splenectomy may be
and tingible body macrophages (“starry-sky pattern”). followed by a prolonged remission.

1070 ParT EIGhT Immunology of Neoplasia


Histologically, the spleen shows expansion of the white pulp In terms of morphology and phenotype, DLBCL is one of
with a characteristic biphasic pattern—central zone of small the most heterogeneous categories in the WHO classification;
lymphocytes surrounded by a peripheral zone of larger cells currently, several morphological variants as well as specific
2
resembling marginal zone cells. The residual polytypic mantle subtypes are recognized. There has been a great interest in
is not present in this instance. The abundant pale cytoplasm identifying DLBCL features that might be prognostically relevant.
evident in tissue sections may also be seen in peripheral blood To address issues traditionally not resolved by morphological
smears, and the cytological features may be mistaken for those or immunophenotypic features, DLBCL was among the first
of hairy-cell leukemia. The phenotype resembles other marginal lymphomas to be analyzed by complementary DNA (cDNA)
zone B-cell lymphomas with a more frequent sIgD expression. microarray. Based on the differential expression of a large set of
Deletion of 7q31 has been described in SMZL; although it genes by GEP, germinal-center B cell–like (GCB) group and
22
can also be seen in isolated cases of splenic B-cell lymphoma or activated B cell–like (ABC) group have been identified, in
leukemia unclassifiable, it has not been detected in other splenic addition to the previously recognized primary mediastinal (thymic)
21
lymphomas. Studies of the Ig variable genes and IgD expression large B-cell lymphoma (PMBCL). GCB DLBCLs express a set of
have revealed two groups of naïve and memory B-cell origin. genes that are associated with normal germinal-center B cells,
whereas ABC DLBCLs show downregulation of these genes and
CLINICaL PEarLS share similarities with post–germinal-center B cells.
The t(14;18) translocation involving BCL2 and immunoglobu-
Aggressive Lymphomas lin heavy-chain gene has been detected in the GCB subtype, but
not in the other subtypes. Previous studies have shown reduced
• Natural history: survival measured in months. disease-free survival in DLBCL cases with BCL2 overexpression,
• Successful therapy can be achieved with combination chemotherapy.
• Relapses from chemotherapy-induced remission may be cured with irrespective of the translocation. Although no absolute correlation
high-dose chemotherapy with hematopoietic support. between morphology and GEP has been established, interestingly,
• In addition to the International Prognostic Index (IPI), gene expression the majority of DLBCLs with centroblastic morphology falls
profiling can be useful in predicting prognosis and survival of individual into the GCB subtype, whereas those with immunoblastic
patients. morphology usually correlate with the ABC subtype.
DLBCL has an aggressive natural history but generally responds
well to chemotherapy. The complete remission rate with modern
Diffuse Large B-Cell Lymphoma, Not regimens is 75–80%. Currently, with R-CHOP (rituximab–
Otherwise Specified cyclophosphamide, doxorubicin, vincristine, and prednisone)
Diffuse large B-cell lymphoma, not otherwise specified (DLBCL, regimen, the 10-year progression-free and overall survival rates
NOS) is one of the more common subtypes of non-HL, represent- for older patients with advanced-stage DLBCL are 36.5% and
ing up to 40% of cases. This diagnosis is used for both primary 43.5%, respectively. 23
DLBCL as well as for cases that transformed from a low-grade Immunophenotype-based algorithms using CD10/BCL6
lymphoma. It may be nodal or involve extranodal sites, including positivity for GCB and MUM1/IRF4 expression for the
bone, skin, the thyroid gland, the GI tract, and lungs. ABC subtype have been proposed as surrogates for cDNA
DLBCL, NOS is composed of large transformed lymphoid microarray; additionally, BCL2 and IPI are also informative to
cells with nuclei at least twice the size of a small lymphocyte stratify the DLBCL cases. Because of the therapeutic impact, in
(Fig. 79.3). The nuclei generally have vesicular chromatin, the recent publication of the proposed changes in the WHO
prominent nucleoli, and basophilic cytoplasm—resembling either 2016 classification, it has been recommended that these two
centroblasts or immunoblasts, albeit with an overall greater subtypes be identified, on the basis of either commonly used
cellular pleomorphism. immunophenotype-based algorithms or GEP.
T cell/histiocyte–rich large B-cell lymphoma (THRLBCL) is a
distinct clinicopathological entity, rather than a morphological
variant. THRLBCL tends to occur in younger patients compared
with other DLBCL, NOS, and often presents with advanced stage
and bone marrow involvement with an aggressive clinical
behavior.
The WHO classification recognizes that lymphomas arising
from certain anatomical sites may have distinctive clinical and
biological features. Among these are primary DLBCL of the central
nervous system (CNS) and primary cutaneous DLBCL, leg type.
Primary DLBCL of the CNS with some distinctive GEP features
shares some similarities with DLBCL arising in other immune-
privileged sites, such as the testis. Primary cutaneous DLBCL,
leg type, has a GEP resembling the ABC subtype of DLBCL,
presents most often in older females, and generally has an aggres-
sive clinical course.
Several Epstein-Barr virus (EBV)–positive B-cell lymphopro-
liferations are often grouped with DLBCL. In the revised WHO
3
FIG 79.3 Diffuse Large-Cell Lymphoma. The neoplastic cells 2016 classification, EBV-positive DLBCL, NOS has now replaced
have large round to oval nuclei with vesicular chromatin and a previous provisional entity “EBV-positive DLBCL of the elderly”
multiple eosinophilic nucleoli. Numerous mitoses are also present. in the 2008 WHO classification, since it may occur in younger

ChaPTEr 79 Lymphomas 1071


28
patients with a broader morphological spectrum and better by GEP. Tumor necrosis factor receptor–associated factor 1
survival 24,25 as a consequence of decreased immune surveillance. expression and c-REL amplification also are seen in both types
These should be distinguished from EBV-associated atypical of neoplasms and could also be detected with appropriate
hyperplasia and lesions associated with iatrogenic or age-related immunohistochemical studies.
immunosuppression, as well as those typified by an isolated and Gray-zone lymphomas are more common in males than
circumscribed cutaneous or mucosal presentation, with an indolent females, present with bulky mediastinal masses, and appear to
behavior and a self-limiting clinical course as EBV-positive have a more aggressive clinical course compared with either
26
mucocutaneous ulcer. Lymphomatoid granulomatosis is an EBV- PMBCL or CHL. Previously, methylation profiling showed a
30
positive B-cell lymphoproliferative disorder (LPD) associated with signature distinct from both CHL and PMBCL. However, as
an inflammatory background rich in T cells. The lung is nearly shown by FISH studies, gray-zone lymphomas, PBMCL, and
always involved; additionally, skin, kidneys, the liver, and the brain CHL all share a number of common cytogenetic aberrations,
are frequently affected as well. DLBCL associated with chronic including gains at 2p16.1 (REL/BCL11A locus), 9p24.1 (JAK2/
inflammation are EBV-driven large B-cell proliferations encountered PDL2), and rearrangements of 16p13.13 (CIITA). DA-EPOCH
in diverse clinical settings, usually associated with a confined R has been effective in both PMBL and DLBCL; however, it is
anatomical space and a background of chronic inflammation. not clear how these patients should be managed clinically, but
These cases appear to have a good prognosis if successfully resected. they appear to benefit from combined modality therapy. 31,32
Human herpes virus 6/Kaposi sarcoma herpes virus (HHV-8/
KSHV)–associated LPDs are also documented. These include CLINICaL PEarLS
primary effusion lymphoma (PEL), multicentric Castleman disease
(MCD), and lymphomas arising in the context of MCD. The Highly Aggressive Lymphomas
cells of PEL are usually coinfected with EBV, and the disease is • More common in children
most often diagnosed in the setting of HIV infection and • Natural history similar to acute leukemia
immunosuppression. Although pleural or peritoneal effusions • Successful treatment includes high-dose chemotherapy (induction,
are most common, extracavitary PEL can present as a tumor consolidation, and maintenance phases) with central nervous system
mass, usually in extranodal sites. PEL has a phenotype resembling prophylaxis
terminally differentiated B cells (i.e., plasmablastic).
Two other lymphomas with a plasmablastic phenotype include
plasmablastic lymphoma (PBL) and ALK-positive large B-cell Burkitt Lymphoma
lymphoma. PBL is usually positive for EBV, most often extranodal, Burkitt lymphoma (BL) occurs most commonly in children and
and is associated with immunosuppression from either human accounts for up to one-third of all pediatric lymphomas in the
immunodeficiency virus (HIV) infection or advanced age. Recent United States. Three clinical variants of BL are recognized—
studies have identified a high incidence of MYC translocation endemic, sporadic, and immunodeficiency-associated. The
27
in PBL. ALK-positive large B-cell lymphomas show overexpres- endemic cases are seen in equatorial Africa and mostly involve
sion of ALK, usually as a consequence of translocation. They the jaw and other facial bones. African BL occurs in malaria-
mainly affect older individuals but can occur at any age. endemic regions. In non–malaria-endemic regions, such as the
United States, extranodal sites are frequent, including the ileocecal
Primary Mediastinal Large B-Cell Lymphoma region, ovaries, kidneys, or breasts. Bone marrow involvement
PMBCL has a distinct constellation of clinical and morphological is a sign of poor prognosis.
features. PMBCL shows marked female gender predominance EBV has been shown to be a cofactor for the development
in adolescents and young adults. The clinical presentation is that of BL and shows varying degrees of positivity in the variant
of a rapidly growing anterior mediastinal mass with frequent subtypes. BL is one of the more common tumors associated
superior vena cava syndrome and/or airway obstruction. Nodal with HIV. It can present at any time during the clinical course
involvement is uncommon at presentation and also at relapse. and also can be the initial acquired immunodeficiency syndrome
Frequent extranodal sites of involvement, particularly at relapse, (AIDS)–defining illness. Recent GEP data have supported a
include the liver, kidneys, adrenal glands, ovaries, the GI tract, common pathogenetic mechanism in cases of HIV infection
and the CNS. The treatment approach includes aggressive systemic and endemic malaria–related immunosuppression.
chemotherapy plus rituximab, along with radiation therapy used Cytologically, BL shows monomorphic medium-sized cells
in some centers. A relatively abundant pale cytoplasm with distinct with round nuclei, multiple (2–5) basophilic nucleoli, and
cytoplasmic membrane and fine compartmentalizing sclerosis moderately abundant deeply basophilic cytoplasm. Cytoplasmic
are characteristic. The tumor appears to be derived from medul- lipid vacuoles reflecting a high proliferation rate and apoptosis
lary B cells within the thymus gland. These cells express CD20 are frequent. It is the most rapidly growing of all lymphomas,
and CD79a, but not surface Ig. CD23 is frequently positive, and with 100% of the cells in cell cycle at any time. The characteristic
MUM-1/IRF4 coexpression is also common. A unique signature “starry sky” pattern of BL is a manifestation of the numerous
was identified by GEP in PMBCL that shared similarities with benign macrophages that have ingested karyorrhectic or apoptotic
HL cell lines, including constitutive activation of the nuclear tumor cells. BL has a mature B-cell phenotype, expressing CD19,
28
factor-κB and recurrent gains and amplification of c-Rel. The CD20, CD22, CD79a, and monoclonal surface Ig, nearly always
expression of the MAL gene has been detected in PMBCL but IgM. CD10 is positive in almost all cases, whereas CD5, CD23,
not in other DLBCLs. and BCL2 are consistently absent.
The 2008 WHO classification included borderline categories, The pathogenesis of BL is related to the MYC oncogene
one of which manifests features intermediate between DLBCL, translocations seen in virtually 100% of cases. The MYC transloca-
29
especially PMBCL and classic HL (CHL) “gray-zone lymphomas.” tion is considered a primary event and often is the sole karyotypic
A close relationship between PMBCL and CHL has been supported abnormality detected. This is in contrast to other aggressive

1072 ParT EIGhT Immunology of Neoplasia


lymphomas, in which the MYC translocation occurs as a secondary Aggressive NK-cell leukemia is a closely related entity with a
33
event in a more complex karyotype. Most of the translocations similar phenotype, EBV association, and epidemiology. It presents
involve the IGH gene on chromosome 14 and, less commonly, at a younger age than extranodal NK/T-cell lymphoma, is
the light-chain genes on chromosomes 2 and 22. associated with systemic disease, and has a fulminant clinical
Recently, a subset with chromosome 11q alterations that course.
+
morphologically, largely phenotypically, and by GEP resemble There are other EBV T-cell and NK-cell proliferations seen
34
BL but lack MYC translocations has been described as a new mainly in Asian children and in indigenous populations from
provisional entity “Burkitt-like lymphoma with 11q aberration” Central and South Americas and Mexico. These show a broad
in the revised classification. Mutations in the transcription factor range of clinical manifestations from indolent, localized forms
TCF3 or its negative regulator ID3 are present in approximately involving skin, such as hydroa vacciniforme-like lymphoproliferative
70% of sporadic and immunodeficiency-related BL and 40% of disorder (name changed from lymphoma) and mosquito bite
endemic cases. allergy, the latter usually being derived from NK cells, to systemic
+
Few cases show GEP of BL but carry additional BCL2 or EBV T-cell lymphoma (name changed from lymphoproliferative
3
BCL6 cytogenetic abnormalities. These “double-hit” or “triple-hit” disease), a more systemic form characterized by fever, hepato-
lymphomas have an aggressive clinical course and poor prognosis splenomegaly and lymphadenopathy with or without cutaneous
36
and are now reclassified from the previous borderline category manifestations and a fulminant clinical course, and hemophago-
of the 2008 WHO classification now as HGBL with and without cytic lymphohistiocytosis.
MYC and BCL2 or BCL6 translocations (except for those fulfilling
3
the criteria for follicular or lymphoblastic lymphoma). Of note, Nodal T-Cell Lymphoma With T FH Phenotype:
the “double-hit” lymphoma are usually of GCB subtype, whereas Angioimmunoblastic T-Cell Lymphoma
“double expressors” are of the ABC subtype. However, this Recently angioimmunoblastic T-cell lymphoma (AITL) has been
category should not be utilized for otherwise typical DLBCLs grouped under an umbrella category “nodal T-cell lymphoma
with a MYC translocation. with T follicular helper cell (T FH ) phenotype” to highlight a
spectrum of nodal lymphomas with T FH phenotype, including
T-CELL AND NK-CELL NEOPLASMS follicular T-cell lymphoma and other nodal PTCLs with a T FH
phenotype, in addition to AITL. 3
Overview of the Classification of T-Cell Neoplasms AITL presents in adults with generalized lymphadenopathy
Peripheral T-cell lymphomas (PTCLs) are uncommon, representing and prominent systemic symptoms, including fever, weight loss,
fewer than 10% of all non-HLs. The classification of PTCL has and skin rash. Polyclonal hypergammaglobulinemia is usually
always been controversial. The genetic landscape of many entities seen.
has only recently been defined, and immunophenotypic markers Histologically, the nodal architecture is generally effaced,
are less specific for apparently distinct disease entities. For these but peripheral sinuses are often open or dilated. Proliferation
reasons, the WHO classification relied, to a considerable extent, of high endothelial venules (HEVs) is often prominent. Fol-
on clinical presentation to subdivide these tumors. 2 licles are regressed, but a proliferation of dendritic cells (DCs)
often entrapping HEVs is typically seen. The atypical lym-
Extranodal NK/T-Cell Lymphoma, Nasal Type phoid cells have clear cytoplasm and are admixed with small
Extranodal NK/T-cell lymphoma, nasal type, is a distinct clini- lymphocytes, immunoblasts, plasma cells, and histiocytes, with or
copathological entity highly associated with EBV. It affects adults without eosinophils. A relationship to T FH has been confirmed by
(median age 50 years), and the most common clinical presentation GEP. The atypical T cells are usually positive for CD4, CD10, and
is a destructive nasal or midline facial lesion. Palatal destruction, PD-1, a phenotype characteristic of T FH . Chemokine ligand-13
orbital swelling, and edema can be prominent. NK/T-cell lym- (CXCL-13), a chemokine involved in B-cell trafficking into
phomas have been reported in other extranodal sites, including the germinal centers, is also expressed. In keeping with an
skin, soft tissue, testes, the upper respiratory tract, and the GI established derivation from T FH, B-cell proliferation, including
tract. It is much more common in Asians and indigenous popula- marked polyclonal plasmacytosis, is often seen. In some cases, the
+
tions of the Americas than in those of European background, plasma cells may be monoclonal. Background EBV B cells are
+
and this indicates that genetic factors play a role in the patho- almost constant, and progression to EBV B-cell lymphoma may
genesis of these lymphomas. occur. The exact role of EBV in AILT is uncertain; however,
Extranodal NK/T-cell lymphoma, nasal type, is characterized plausible theories include expansion resulting from decreased
by a broad cytological spectrum. Although the cells express some immune surveillance. The majority of cases show clonal rear-
T cell–associated antigens, most commonly CD2, other T-cell rangements of T-cell receptor (TCR) genes, but proliferation of
markers, such as surface CD3, are usually absent. Cytoplasmic B cells also contributes to the histological and clinical picture.
CD3 is positive, but most cases lack clonal T-cell gene rearrange- Patients may initially respond to steroids or mild cytotoxic
ment. In favor of an NK-cell origin, the cells are nearly always chemotherapy, but progression usually occurs. More aggressive
+
CD56 , although CD16 and CD57 are usually negative. EBV is combination chemotherapeutic regimens have led to a higher
invariably positive as shown by in situ hybridization. remission rate, but patients are prone to secondary infec-
The clinical features and treatment response of nonnasal tious complications. The median survival is usually less than
extranodal NK/T-cell lymphoma are different from of those of 5 years.
nasal presentation of this lymphoma in that the addition of New genetic insights have shown recurrent mutations, includ-
radiotherapy in early-stage nasal cases may offer a survival ing TET2, IDH2, DNMT3A, RHOA, and CD28, as well as gene
35
benefit. A hemophagocytic syndrome is a common clinical fusions, such as ITK-SYK or CTLA4-CD28, which affect a sig-
complication and adversely affects survival. Emerging oncogenic nificant proportion of PTCL, NOS cases with a T FH phenotype,
pathways have been identified by GEP. and AITL cases.

ChaPTEr 79 Lymphomas 1073


Peripheral T-Cell Lymphomas, Not Otherwise Specified of implant to lymphoma of about 10 years. Confinement of the
Peripheral T-cell lymphomas, not otherwise specified (PTCL, neoplastic cells to the seroma fluid without capsule invasion
NOS) most often present in adults with generalized lymphade- portends a favorable prognosis.
nopathy, hepatosplenomegaly, and frequent bone marrow Recently GEP studies have shown that ALK-negative ALCLs
involvement. It is a diagnosis of exclusion with most cases being have a signature close to that of ALK-positive counterparts, but
nodal in origin. PTCLs are characterized by cytological and distinct from other NK/T-cell lymphomas. Genetic studies have
phenotypical heterogeneity. Most cases have a mature T-cell shown convergent mutations and kinase fusions leading to
phenotype and express one of the major subset antigens, with constitutive activation of the JAK/STAT3 pathway. The overall
CD4 expression seen more frequently than CD8. These are not survival and disease-free survival are significantly better in
clonal markers, and antigen expression can change over time. ALK-positive cases than in ALK-negative cases. Clinical or
Loss of one of the pan-T-cell antigens (CD3, CD5, CD2, or CD7) prognostic variations exist in ALK-negative ALCLs—a subset
is seen in two-thirds of cases, with loss of CD7 being most with rearrangements at the DUSP22 and IFR4 locus on 6p25
frequent. has a superior prognosis, whereas a small subset with TP63
Recently, GEP has shown a global signature close to one of rearrangements is very aggressive.
the activated T lymphocytes and has identified at least three
subtypes characterized by overexpression of GATA3, TBX21, and Primary Cutaneous ALCL
cytotoxic genes associated with a different clinical behavior and Primary cutaneous ALCL is closely related to lymphomatoid
response to therapy. The clinical course is generally aggressive, papulosis (LyP) and differs at the clinical, immunophenotypic,
especially with a high proliferation signature, with a lower and molecular levels from the systemic form. Indeed, LyP and
response rate than that seen for aggressive B-cell lymphomas. cutaneous ALCL represent a histological or clinical continuum
+
of CD30 cutaneous lymphoproliferative diseases. Small lesions
Anaplastic Large-Cell Lymphoma are likely to regress, whereas patients with large tumor masses
Anaplastic large-cell lymphoma (ALCL) is most common in may develop disseminated disease with lymph node involvement—a
children and young adults, and more common in males than in period of observation is usually warranted before the institution
females. Nodal presentations are most common; however, a variety of any chemotherapy for isolated lesions. Most patients with
of extranodal sites can be involved. primary cutaneous ALCL have multiple skin lesions, but it is a
ALCL is characterized by pleomorphic or monomorphic cells more indolent disease compared with other T-cell lymphomas
+
that have a propensity to invade lymphoid sinuses. The cells of of the skin. Cutaneous ALCL is CD30 but is usually ALK-1– and
classic ALCL have large, often lobulated nuclei with small EMA-negative, and it also lacks the t(2;5) translocation. More
basophilic nucleoli. In some cases, the nuclei may be round. The recently, LyP variants D-E and LyP with 6p25 have been described;
cytoplasm is usually abundant and amphophilic and has distinct appreciation of these variants is important as they can mimic
cytoplasmic borders, and a prominent Golgi region is generally aggressive T-cell lymphoma histologically but clinically are similar
visible. to other forms of LyP A-C.
The expression of the CD30 antigen is a hallmark of this
disease. However, CD30 expression is not specific for ALCL and Subcutaneous Panniculitis-Like T-Cell Lymphoma
may also be seen in other forms of malignant lymphoma, Subcutaneous panniculitis-like T-cell lymphoma (SPTCL) usually
+
including CHL. ALCL-ALK is associated with a characteristic presents with subcutaneous nodules primarily affecting the
chromosomal translocation, t(2;5)(p23;q35), involving the ALK extremities. In its early stages, the infiltrate may appear deceptively
and NPM genes, respectively. A variety of other ALK partners benign, and lesions are often misdiagnosed as panniculitis.
have been identified and monoclonal antibodies to the ALK However, histological progression usually occurs, and subsequent
protein have been able to identify tumor cells regardless of the biopsies show more pronounced cytological atypia.
+
underlying translocation. The neoplastic cells show both nuclear The neoplastic cells are CD8 cytotoxic α/β T cells. They are
and cytoplasmic staining in the majority of cases; the presence also positive for the cytotoxic proteins perforin, granzyme B,
of cytoplasmic staining for ALK is suggestive of a variant and TIA-1 that may be responsible for the cellular destruction
translocation. seen in these tumors. EBV is negative. Some PTCL of γ/δ T-cell
Immunohistochemistry is indispensable in the correct diagnosis derivation may show similar features but differ from SPTCL in
of ALCL. The prominent Golgi region usually shows intense their clinical behavior (more aggressive) and histological pattern,
staining for CD30 and epithelial membrane antigen (EMA). The as they are often not confined to the subcutis.
cells exhibit an aberrant phenotype with loss of many of the T Some patients have a history of autoimmune disease, and in
cell–associated antigens. Both CD3 and CD45RO, the most widely particular, the differential diagnosis of SPTCL with lupus pro-
used pan-T-cell markers, are negative in >50% of cases. CD2 and fundus panniculitis can be challenging. Unlike SPTCL, lupus
CD4 are positive in the majority, whereas CD8 is usually negative. panniculitis usually contains abundant plasma cells, a mixture
−
+
+
+
ALCL cells, despite the CD4 /CD8 phenotype, often express the of CD4 and CD8 cells, and a relative increase of γ/δ T cells
cytotoxic-associated antigens (TIA-1, granzyme B, and perforin). and plasmacytoid DCs.
In addition, clusterin is generally present in ALCL and represents A hemophagocytic syndrome can be a complication of SPTCL
another useful diagnostic marker. Molecular studies in most cases but is more often associated with γ/δ T-cell lymphomas involving
demonstrate TCR rearrangement, confirming a T-cell origin. the subcutaneous tissue. Patients present with fever, pancytopenia,
Improved criteria now exist for the recognition of ALK- and hepatosplenomegaly. It is most readily diagnosed in bone
negative ALCL as a separate entity. It occurs in an older age marrow aspirate smears, where histiocytes containing erythrocytes,
group compared with the ALK-positive cases. Recently, a unique platelets, and other blood elements may be observed. The
form of ALK-negative ALCL arising in association with breast hemophagocytic syndrome usually heralds a fulminant downhill
37
implants has been identified, with a medial interval from time clinical course.

1074 ParT EIGhT Immunology of Neoplasia


Primary Cutaneous γ/δ T-Cell Lymphomas The homing pattern manifested by the malignant cells is similar
Primary cutaneous γ/δ T-cell lymphomas are clinically aggressive to that of normal γ/δ T cells, which also populate the sinusoidal
tumors that can present with involvement of the subcutis and areas of the spleen. The neoplastic cells have a phenotype that
the dermis or with epidermal infiltration. Skin is the most resembles normal γ/δ T cells, usually expressing neither CD4
common presenting site, however, similar lymphomas of γ/δ nor CD8. CD56 is often positive. The cells are positive for the
T-cell origin can present in other extranodal sites, including the cytotoxic protein TIA-1, but are not activated and generally lack
GI tract and lungs. The cells have a cytotoxic phenotype and granzyme B and perforin. In situ hybridization for EBV is negative.
express cytotoxic molecules, and like normal γ/δ T cells, lack The recognition of the atypical cells in bone marrow is greatly
+
CD5. They may be CD8 or, more often, double-negative for aided by immunohistochemical stains showing a sinusoidal
CD4 and CD8. Few cutaneous γ/δ T-cell lymphomas appear to pattern of infiltration.
be TCR silent (negative for both TCR-β and TCR-γ) but share Isochromosome 7 is a consistent cytogenetic abnormality,
the same aggressive clinical behavior. Recurrent mutations of usually seen in conjunction with trisomy 8. Rare cases derived
STAT5B can be seen in T-cell lymphomas of γ/δ origin. 38 from α/β T cells but with similar morphological and biological
features can be seen. Recurrent mutations of STAT5B and less
Mycosis Fungoides and Sézary Syndrome often STAT3 are seen in HSTCL of γ/δ origin. 38
Mycosis fungoides (MF) and Sézary syndrome (SS) are closely Clinically, HSTCL is aggressive. Although patients may respond
related and often considered together from a clinical and biological initially to chemotherapy, relapse occurs in the vast majority of
standpoint but are now regarded as separate diseases. Both are cases. The median survival is less than 3 years. Allogeneic bone
primary cutaneous T-cell malignancies derived from mature skin marrow transplantation is required for sustained remission.
homing CD4 T cells. Epidermotropism is the hallmark of MF;
infiltration of the epidermis produces characteristic Pautrier Adult T-Cell Leukemia/Lymphoma
microabscesses. The cutaneous lesions are categorized as patches, Adult T-cell leukemia/lymphoma (ATLL) is a distinct form of
plaques, and tumors, based on the extent of the infiltrate. A γ/δ T-cell lymphoma associated with the retrovirus human
indolent variant of MF has also been described. SS presents with T-lymphotropic virus-1 (HTLV-1). The highest number of cases
exfoliative erythroderma and circulating cerebriform lymphocytes is seen in Southwestern Japan and the Caribbean basin. The
known as Sézary cells. Clinically, SS is more aggressive than MF. disease has a long latency, and affected individuals are usually
exposed to the virus very early in life. The virus may be transmit-
Enteropathy-Associated T-Cell Lymphoma ted in breast milk and through exposure to blood or blood
Enteropathy-associated T-cell lymphoma (EATL) is highly products. The cumulative incidence of ATLL is estimated to be
associated with celiac disease on a worldwide basis. This disease 2.5% among HTLV-1 carriers. The virus is monoclonally inte-
occurs in adults; the majority has either overt or clinically silent grated into tumor DNA. Different clinical variants of ATLL,
gluten-sensitive enteropathy (Chapter 75). Ulcerative jejunitis may including the acute, lymphomatous, chronic, and smoldering
precede the development of overt EATL and may share a common types, have been recognized. Cutaneous involvement is seen in
clonal T-cell population as with the subsequent lymphoma. The the majority of patients.
small bowel usually shows ulceration with frequent perforation, Peripheral blood involvement is very common, often without
with or without a mass. EATL shows a cytological composition bone marrow disease. The atypical cells are often markedly
of variably sized or polymorphic atypical lymphoid cells. The polylobated and have been referred to as “flower” cells. The cells
adjacent small bowel usually shows villous atrophy associated have a characteristic phenotype that resembles T-regulatory cells
+
+
+
+
+
with celiac disease. The neoplastic cells are CD3 , CD7 T cells (Tregs)—CD3 , CD4 , and CD25 . FoxP3 is expressed in a
that also express the homing receptor CD103. Anaplastic cells minority of tumor cells. The function of the tumor cells as Tregs
strongly positive for CD30 can be present. The cells express may correlate with the associated immunodeficiency.
cytotoxic molecules, a feature shared by nearly all extranodal Somatic gain of function CCR4 mutations have been implicated
40
T-cell lymphomas. The majority belong to the α/β TCR subset, in the pathogenesis of ATLL. Recent integrated molecular analysis
whereas only a minority expresses the γ/δ TCR. Other PTCLs has shown frequent intragenic deletions and mutations of key
+
can present with intestinal disease, including EBV extranodal T-cell signaling molecules in ATLL. 41
T/NK-cell lymphomas and γ/δ T-cell lymphomas, and should
be distinguished from EATL. The clinical course is aggressive. CLINICaL PEarLS
The disease previously known as EATL type II has been now been Hodgkin Lymphoma
formally designated as monomorphic epitheliotropic intestinal T-cell
lymphoma (MEITL); it shows no association with celiac disease • B-cell lineage established in nearly all cases
and appears to have an increased incidence in Asian and Hispanic • Reed-Sternberg cell, the hallmark of the disease, represents a “crippled”
populations. Unlike classic EATL, MEITL is monomorphic, usually germinal-center B cell
39
positive for CD8, CD56, and MAPK. STAT5B mutations are • Nodular lymphocyte-predominant Hodgkin lymphoma considered a
common in MEITL cases, many of which are of γ/δ T-cell origin. related but distinct entity
• Eighty percent of patients are curable with current therapy
Hepatosplenic T-Cell Lymphoma • Stage of disease guides the choice of therapy; even patients with
advanced-stage disease may be cured
Hepatosplenic T-cell lymphoma (HSTCL) shows a marked male • Late complications from treatment include acute leukemia (alkylating
gender predominance, and most patients are young adults. The agents with extended-field radiation therapy), second solid tumors
clinical presentation is that of marked hepatosplenomegaly in (radiation therapy), and premature atherosclerotic coronary artery
the absence of lymphadenopathy. disease (radiation therapy)
The liver and the spleen show marked sinusoidal infiltration, • Cause of death in the first 5–10 years is mainly Hodgkin lymphoma;
after 10 years, mainly secondary malignant tumors
with sparing of both portal triads and white pulp, respectively.

ChaPTEr 79 Lymphomas 1075



HODGKIN LYMPHOMAS Survival for localized cases with or without treatment is long.
However, cases with variant histology are more likely to have
43
HL and non-HL have long been regarded as distinct disease advanced stage disease with a higher relapse rate. Moreover,
entities on the basis of their differences in pathology, phenotype, patients with advanced-stage disease respond poorly to HL
clinical features, and response to therapy. It is now accepted that regimens, such as adriamycin, bleomycin, vinblastine, dacarbazine
the malignant cell of HL is an altered B cell. Therefore it is not (ABVD), and benefit from treatment for aggressive B-cell
44
surprising that biological and clinical overlap occurs between lymphoma. Although considered two separate entities with
these two lymphoma groups, as also shown by GEP in PMBCL major differences, NLPHL shares immunomorphological similari-
and classic Hodgkin cell lines. In spite of the close histogenetic ties with THRLBCL; these similarities have also been shown by
relationship (hence the name Hodgkin lymphoma), these disorders GEP and array comparative genomic hybridization. Two forms
are still treated with different modalities. of histological progression can be seen, either to DLBCL or to
The diagnosis of classic HL (CHL) depends on the identifica- a process resembling THRLBCL.
tion of Hodgkin/Reed–Sternberg (HRS) cells in an appropriate
inflammatory background composed of small T lymphocytes, Classic Hodgkin Lymphoma, Nodular Sclerosis
plasma cells, histiocytes, and granulocytes (often eosinophils). This variant of HL, classic Hodgkin lymphoma, nodular sclerosis
All cases of CHL share certain immunophenotypic and genotypic (CHLNS) is most commonly seen in adolescents and young
+/−
+
−
−
features. The phenotype is CD30 , CD15 , CD45 , and EMA . adults but can occur at any age. Female cases equal or exceed
Expression of B cell–associated antigens is seen in up to 75% of those in males. The mediastinum is commonly involved; stage
cases; however, when present, CD20 staining is usually weaker and bulk of disease have prognostic importance.
or more variable than that seen in normal B cells. CD79a is The tumor has at least a partially nodular pattern, with fibrous
usually negative. Ig and TCR genes are usually germline because bands separating the nodules in most cases. Diffuse areas with
of the paucity of tumor cells in the inflammatory background; necrosis may be present. The characteristic cell is the “lacunar-
however, microdissection can enable amplification for clonal type” Reed-Sternberg (RS) cell, which may be very numerous.
rearrangement of the Ig genes by polymerase chain reaction Diagnostic RS cells are usually also present. The background
(PCR). In addition, the presence of somatic mutations indicates contains lymphocytes, histiocytes, plasma cells, eosinophils, and
transit through the germinal center. neutrophils. Grading (I and II) is based on the proportion of
Sufficient evidence has emerged in recent years to warrant the tumor cells and the presence of necrosis but is considered
the recognition of nodular lymphocyte-predominant HL as a optional. The immunophenotype and genotype are characteristic
distinct entity. Although it resembles other types of HL in having of CHL. However, EBV is infrequently positive— less than 15%
a minority of putative neoplastic cells in a background of benign of cases.
inflammatory cells, it differs morphologically, immunophenotypi- CHLNS is often curable; however, in long-term survivors,
cally, and clinically from CHL. The preferred term Hodgkin the risk of secondary malignancies is increased, especially in
lymphoma, versus Hodgkin disease, reflects current knowledge those receiving both chemotherapy and radiation. CHLNS of
concerning the nature of the neoplastic cell as a lymphocyte. the mediastinum is thought to be closely related to PMBCL, and
both types of tumors can be seen in the same patient, either as
Nodular Lymphocyte-Predominant Hodgkin Lymphoma composite malignancy or sequentially.
Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL)
occurs at all ages but is more common in adult males. It usually Classic Hodgkin Lymphoma, Mixed Cellularity
involves peripheral lymph nodes with sparing of the mediastinum Classic Hodgkin lymphoma, mixed cellularity (CHLMC) is
and is localized at diagnosis, although rarely it may be predominantly seen in male adults. A bimodal age distribution,
disseminated. NLPHL usually has a nodular growth pattern, with peaks in young children and later in older adults can
with or without diffuse areas. The number of infiltrating reactive be present. Both CHLMC and the lymphocyte-depleted form
T cells is variable, and various patterns have been described on (see below) can be associated with underlying HIV infection.
42
the basis of the cellular composition and growth pattern. The The infiltrate is diffuse without band-forming sclerosis, although
atypical cells have vesicular, polylobated nuclei and small fine interstitial fibrosis may be present (Fig. 79.4). HRS
nucleoli. These had been called lymphocytic and/or histiocytic cells are of the classic type. It is often EBV-positive, seen in up
(L&H) cells, or “popcorn” cells, but the term lymphocyte- to 75% of cases. The stage is often advanced at diagnosis.
predominant (LP) cell is now preferred. LP cells differ from classic The clinical course is moderately aggressive, but is often
HRS cells. The background is composed predominantly of curable.
lymphocytes with or without clusters of epithelioid histiocytes.
Plasma cells are infrequent; eosinophils and neutrophils are also Classic Hodgkin Lymphoma, Lymphocyte Depletion
+
rare. The atypical cells are CD45 , positive for B cell–associated Classic Hodgkin lymphoma, lymphocyte depletion (CHLLD) is
+
−
+/−
antigens (CD19, CD20, CD22, CD79a), CDw75 , EMA CD15 , the least common variant of CHL and is most common in older
−/+
−
CD30 and usually sIg , as shown by routine techniques. Small people, in HIV-positive individuals, and in populations of
lymphocytes in the nodules are predominantly B cells with a nonindustrialized countries. It frequently presents with abdominal
mantle zone phenotype. However, numerous T cells are present, lymphadenopathy, and spleen, liver, and bone marrow involve-
+
with CD279 T cells “rosetting” the LP cells. The proportion of ment, but without peripheral adenopathy. The infiltrate is diffuse
T cells tends to increase over time in sequential biopsies. A and often appears hypocellular because of the presence of diffuse
prominent follicular DC meshwork is present within the nodules. fibrosis and necrosis. A large number of HRS cells and occasional
LP cells, when microdissected, have shown to have clonally bizarre “sarcomatous” variants are seen relative to the fewer
rearranged immunoglobulin genes with evidence of somatic number of normal lymphocytes and scarce other inflammatory
hypermutation. cells. The immunophenotype is characteristic of CHL. Since the

1076 ParT EIGhT Immunology of Neoplasia


Please check your eBook at https://expertconsult.inkling.com/
for self-assessment questions. See inside cover for registration
details.


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However, patients usually present with localized disease and tend follicular lymphoma. Blood 2011;118:2976–84.
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similar to other variants of CHL. lymphoma: a genetic perspective. Haematologica 2014;99:481–8.
15. Karube K, Guo Y, Suzumiya J, et al. CD10-MUM1+ follicular lymphoma
lacks BCL2 gene translocation and shows characteristic biologic and
ON ThE hOrIZON clinical features. Blood 2007;109:3076–9.
16. Salaverria I, Philipp C, Oschlies I, et al. Translocations activating IRF4
• In recent years, there has been a greater appreciation of early events identify a subtype of germinal center-derived B-cell lymphoma affecting
in lymphoid neoplasia. predominantly children and young adults. Blood 2011;118:139–47.
• These early lesions in some ways can be considered equivalent to 17. Katzenberger T, Kalla J, Leich E, et al. A distinctive subtype of t(14;18)-
benign neoplasms in the epithelial system. negative nodal follicular non-Hodgkin lymphoma characterized by a
• These are clonal proliferations of B or T cells that carry genetic aber- predominantly diffuse growth pattern and deletions in the chromosomal
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lymphocytic leukemia, multiple myeloma, follicular lymphoma, mantle 18. Loeffler M, Kreuz M, Haake A, et al. Genomic and epigenomic
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• Examples include monoclonal gammopathy of undermined significance, 19. Swerdlow SH. Pediatric follicular lymphomas, marginal zone lymphomas,
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follicular neoplasia, lymphomatoid papulosis, patch stage of mycosis 20. Farinha P, Gascoyne RD. Molecular pathogenesis of mucosa-associated
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• Early lesions appear to lack the secondary and tertiary “hits” seen in lymphoid tissue lymphoma. J Clin Oncol 2005;23:6370–8.
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• Current focus is to define the precise genetic features that distinguish 2010;220:461–74.
early lesions from lymphoma, assess the risk of clinical progression, 22. Rosenwald A, Wright G, Chan WC, et al. The use of molecular profiling
and determine their clinical management. to predict survival after chemotherapy for diffuse large-B-cell lymphoma.
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patients in the LNH-98.5 trial, the first randomized study comparing 2014;123:1187–98.
rituximab-CHOP to standard CHOP chemotherapy in DLBCL patients: a 35. Au WY, Weisenburger DD, Intragumtornchai T, et al. Clinical differences
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27. Taddesse-Heath L, Meloni-Ehrig A, et al. Plasmablastic lymphoma with 39. Deleeuw RJ, Zettl A, Klinker E, et al. Whole-genome analysis and HLA
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2003;198:851–62. 41. Kataoka K, Nagata Y, Kitanaka A, et al. Integrated molecular analysis of
29. Traverse-Glehen A, Pittaluga S, Gaulard P, et al. Mediastinal gray zone adult T cell leukemia/lymphoma. Nat Genet 2015;47:1304–15.
lymphoma: the missing link between classic Hodgkin’s lymphoma and 42. Fan Z, Natkunam Y, Bair E, et al. Characterization of variant patterns of
mediastinal large B-cell lymphoma. Am J Surg Pathol 2005;29:1411–21. nodular lymphocyte predominant Hodgkin lymphoma with
30. Eberle FC, Rodriguez-Canales J, Wei L, et al. Methylation profiling of immunohistologic and clinical correlation. Am J Surg Pathol
mediastinal gray zone lymphoma reveals a distinctive signature with 2003;27:1346–56.
elements shared by classical Hodgkin’s lymphoma and primary 43. Hartmann S, Eichenauer DA, Plutschow A, et al. The prognostic impact
mediastinal large B-cell lymphoma. Haematologica 2011;96:558–66. of variant histology in nodular lymphocyte-predominant Hodgkin
31. Wilson WH, Grant C, et al. Therapy in primary mediastinal B-cell lymphoma: a report from the German Hodgkin Study Group (GHSG).
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32. Dunleavy K, Pittaluga S, Maeda LS, et al. Dose-adjusted 44. Xing KH, Connors JM, Lai A, et al. Advanced-stage nodular lymphocyte
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New Engl J Med 2013;368:1408–16. lymphoma: a matched pair outcome analysis. Blood 2014;123:3567–73.
33. Dave SS, Fu K, Wright GW, et al. Molecular diagnosis of Burkitt’s 45. Nam-Cha SH, Montes-Moreno S, Salcedo MT, et al. Lymphocyte-rich
lymphoma. N Engl J Med 2006;354:2431–42. classical Hodgkin’s lymphoma: distinctive tumor and microenvironment
34. Salaverria I, Martin-Guerrero I, Wagener R, et al. A recurrent 11q markers. Mod Pathol 2009;22:1006–15.
aberration pattern characterizes a subset of MYC-negative high-grade

ChaPTEr 79 Lymphomas 1077.e1


MULTIPLE-C h OICE QUESTIONS

1. In the diagnostic workup of a diffuse large B-cell lymphoma, C. Chronic lymphocytic leukemia
which markers are helpful in the identification of the cell of D. Lymphoplasmacytic lymphoma
origin? E. Multiple myeloma
A. CD10/BCL6/MUM1 3. Which of the following lymphomas are typically associated
B. CD10/P53/MIB-1 with Epstein-Barr virus (EBV) positivity?
C. CD10/BCL2/MUM1 A. Enteropathy-associated T-cell lymphoma
D. CD10/BCL6/MIB1
B. Extranodal natural killer (NK)-/T-cell lymphoma, nasal
2. The presence of MYD88 L265P mutation is associated with type
the diagnosis of: C. Hepatosplenic T-cell lymphoma
A. Mantle cell lymphoma D. Anaplastic large-cell lymphoma, ALK positive
B. Follicular lymphoma E. Anaplastic large-cell lymphoma, ALK negative

80









Monoclonal Gammopathies



Kazunori Murata, Samuel McCash, C. Ola Landgren








Monoclonal gammopathies are a group of disorders characterized Most cases of MGUS are sporadic. However, relatives of persons
by the proliferation of a single clone of plasma cells. These plasma with MGUS (or MM) have an increased risk of developing MGUS
cells produce an immunologically homogeneous immunoglobulin, and related diseases. Acquired angioedema is closely associated
or parts thereof, also known as an M (for monoclonal) protein. with MGUS, being identified in approximately one-third of
This chapter covers 12 recognized types of monoclonal gam- acquired angioedema patients. 1
mopathies: monoclonal gammopathy of undetermined signifi-
cance (MGUS); multiple myeloma (MM); smoldering multiple Clinical Presentation and Laboratory Findings
myeloma (SMM); nonsecretory/oligosecretory myeloma; plasma By definition, all patients with MGUS have no myeloma-associated
cell leukemia (PCL); solitary plasmacytoma of bone (SPB); solitary symptoms attributable to the proliferation of plasma cells or
extramedullary plasmacytoma (SEP); polyneuropathy, organo- the presence of the monoclonal protein. Non-IgM and IgM MGUS
megaly, endocrinopathy, monoclonal protein, skin changes are characterized by the presence of an M-protein produced and
(POEMS) syndrome; Waldenström macroglobulinemia (WM); secreted by clonal plasma cells. These proteins can be detected
heavy chain disease (HCD); light chain amyloidosis (AL); and in serum using either protein electrophoresis or immunofixation
monoclonal immunoglobulin deposition disease. These disorders electrophoresis as well as in a sample from a 24-hour urine
differ in the types of monoclonal proteins that are generated by collection using the same methods.
the diseased cell. The proteins involved include immunoglobulin The quantity of monoclonal protein in all cases of MGUS is
(Ig)-G, IgA, IgM, IgD, IgE, kappa or lambda light chains only, defined as <3 g/dL in the serum. Serum immunofixation elec-
and immunoglobulin heavy chains only. trophoresis (SIF) is used to confirm the presence of an M-protein
and to determine its type. The clonal immunoglobulin distribution
MONOCLONAL GAMMOPATHY OF in MGUS varies according to isotype: IgG 69%, IgM 17%, IgA
UNDETERMINED SIGNIFICANCE 11%, IgD <1%, and biclonal 3%. 2
IgD MGUS is extremely rare with only two cases having been
MGUS is a clinically asymptomatic premalignant clonal plasma reported. Thus finding an IgD M-protein is almost always associ-
cell or lymphoplasmacytic proliferative disorder. It is defined by ated with the diagnosis of MM, light chain amyloid, or PCL.
the presence of the following characteristics: the presence of an Biclonal MGUS is also uncommon, representing only 3% of
M-protein in the serum at a concentration <3 g/dL, bone marrow patients with MGUS. Approximately one-third of MGUS patients
with <10% monoclonal plasma cells, and the absence of end-organ will have a decrease in the concentration of the uninvolved
damage related to the lymphoproliferative process. There are immunoglobulin isotypes (e.g., IgM and IgA in the case of IgG
three distinct clinical types of MGUS: non-IgM MGUS, IgM MGUS), with many demonstrating a reduction in the normal
MGUS, and light chain MGUS (LC-MGUS). polyclonal circulating immunoglobulins. 1
Serum free light chain (sFLC) assays can detect low concentra-
Epidemiology tions of monoclonal free light chain in patient serum. This
MGUS occurs in approximately 1–2% of adults and >2% of the technique can be used to diagnose light chain MGUS and to
general Caucasian population above the age of 50. The mean predict the risk of the progression of MGUS. A monoclonal light
age at diagnosis is 70 years with <2% of patients diagnosed chain can also be found in the urine of approximately 20% of
before the age of 40. The incidence increases with age, and the MGUS patients. 1
prevalence of MGUS in persons ≥50, ≥70, and ≥85 years of age The complete blood count and peripheral smear are typically
is 3.2%, 5.3%, and 7.5%, respectively. 1 normal. Infrequently, rouleaux formation (the phenomenon in
Men are at higher risk of developing MGUS compared with which red cells take on the appearance of stacked coins in diluted
women (4.0% versus 2.7%). The incidence of MGUS in men is suspensions of blood) is seen, and this is typically observed in
estimated to be 120/100 000 at age 50, increasing to 530/100 000 those MGUS patients with elevated serum protein levels. Circulat-
by the age of 90, whereas the incidence rate for women is estimated ing plasma cells of the same isotype can be seen in peripheral
to be 60/100 000 at age 50, increasing to 370/100 000 by the age blood of some patients with MGUS using a slide-based immu-
of 90. Race is also a risk factor, with the incidence of MGUS nofluorescence assay or flow cytometry. 1
being 1.5- to 3-fold higher in Africans and African Americans By definition, a bone marrow aspirate and biopsy must
compared with Caucasians. 1 demonstrate fewer than 10% clonal plasma cells. For IgM

1079

1080 Part eight Immunology of Neoplasia


MGUS, clonal lymphoplasmacytic cells must constitute <10% to idiopathic Bence-Jones proteinuria, light chain MM, AL
infiltration. amyloidosis, or LCDD.
Conventional cytogenetics are normal in patients with MGUS Patients with MGUS progress to a symptomatic plasma cell
due to technical limitations, a low proliferative rate, and the proliferative disorder or lymphoproliferative disorder at a rate
small number of plasma cells within the bone marrow samples. of 1% per year. However, not all persons with MGUS have the
However, studies using more sensitive methods have shown that same risk of disease progression. The risk of progression to a
chromosomal abnormalities are very common in the clonal cells more serious disease ranges widely from 0.6–3.4%/year based
found in MGUS. Analysis by interphase fluorescent in situ on the initial value of serum monoclonal protein as well as from
4
hybridization (FISH) demonstrate chromosomal aneuploidy, in 0.25–2.9%/year according to a risk stratification model. Three
particular hyperdiploidy, in approximately 50% of patients with adverse risk factors have been combined to create a risk stratifica-
MGUS. 1 tion model that is useful in predicting progression of MGUS
5
(non-IgM and IgM) to MM or a related malignancy : The findings
Diagnosis that generate a less favorable outcome include serum monoclonal
Diagnostic criteria for each of the individual types of MGUS protein level ≥1.5 g/dL, non-IgG MGUS, and abnormal serum
are determined by the International Myeloma Working Group free light chain ratio (kappa/lambda free light chain ratio <0.26
(IMWG) and are detailed in Table 80.1. 3 or >1.65).
The stratified absolute chance of disease progression over 20
Clinical Course years is 58% for high-risk MGUS associated with all three risk
The clinical course of MGUS depends largely on the specific factors, 37% for high- or intermediate-risk MGUS based on two
4
type. In non-IgM MGUS a minority of cases will progress to risk factors, 21% for low- or intermediate-risk MGUS associated
the more advanced premalignant stage SMM and to symptomatic with one risk factor, and 5% for low-risk MGUS based on no
5
MM. Less frequently, these patients progress to light chain risk factors. Risk factors for progression of LC-MGUS have yet
amyloidosis, light chain deposition disease (LCDD), or another to be defined.
lymphoproliferative disorder. IgM MGUS can progress to smolder-
ing WM (SWM) and to symptomatic WM. IgM MGUS also KeY CONCePtS
infrequently progresses to IgM MM. LC-MGUS may progress
MGUS
• A premalignant clonal plasma cell/lymphoplasmacytic proliferative
disorder.
TABLE 80.1 iMWg Diagnostic Criteria for • Asymptomatic by definition.
MgUS by type • Defined by the presence of a monoclonal protein (M-protein) in the
serum or bone marrow with <10% monoclonal plasma cells and absence
Non-igM MgUS of end-organ damage related to the proliferative process.
1. The presence of a serum monoclonal protein (M-protein, whether • Three types of MGUS: non-IgM MGUS, IgM MGUS, and light chain
IgA, IgG, or IgD) at a concentration <3 g/dL. MGUS.
2. Fewer than 10% clonal plasma cells in the bone marrow. • Progresses to a symptomatic plasma cell proliferative disorder or
3. The absence of lytic bone lesions, anemia, hypercalcemia, and lymphoproliferative disorder at a rate of 1% per year.
renal insufficiency related to the plasma cell proliferative process. • Patients with MGUS should not be treated but instead monitored for
signs of progressive disease.
igM MgUS
1. The presence of a serum IgM monoclonal protein at a
concentration <3 g/dL. Management and Prognosis
2. Fewer than 10% clonal lymphoplasmacytic/plasma cells in the bone Patients with MGUS should be followed over time with history
marrow. and physical examination for signs and symptoms of progressive
3. The absence of end-organ damage such as anemia, constitutional disease. Treatment of MGUS has not been shown to have any
symptoms, hyperviscosity, lymphadenopathy, or
hepatosplenomegaly related to the plasma cell proliferative effect on mortality, and, as noted above, only a minority of
process. patients will progress to symptomatic disease over a set time
4
interval. All other patients are followed with annual serum and
Light Chain MgUS urinary M-protein, complete blood count, creatinine, and serum
4
1. The presence of an abnormal free light chain ratio (i.e., ratio of calcium. It has been observed that patients with MGUS are at
kappa to lambda free light chains <0.26 or >1.65). increased risk of fracture and thromboembolic disease. Patients
2. Increased level of the appropriate involved light chain (e.g., with MGUS should be evaluated for osteoporosis with a dual-
increased kappa FLC in patients with a ratio >1.65 and increased energy X-ray absorptiometry (DEXA) scan and have their vitamin
lambda FLC in patients with a ratio <0.26). 4
3. No monoclonal immunoglobulin heavy chain (IgG, IgA, IgD, or IgM). D and calcium intake optimized.
4. Fewer than 10% clonal lymphoplasmacytic cells in the bone
marrow. MULTIPLE MYELOMA
5. The absence of lytic bone lesions, anemia, hypercalcemia, and
renal insufficiency related to the plasma cell proliferative process. MM is the neoplastic proliferation of a single clone of plasma
cells producing an M-protein. The difference between MM and
FLC, free light chain; Ig, immunoglobulin; IMWG, International Myeloma Working
Group; MGUS, monoclonal gammopathy of undetermined significance. MGUS lies in the presence of detectable end-organ damage caused
Source: International Myeloma Working Group. Criteria for the classification of by the proliferation of plasma cells and/or the overproduction
monoclonal gammopathies, multiple myeloma and related disorders: a report of the of M-protein. The proliferation of plasma cells in MM causes
International Myeloma Working Group. Br J Haematol 2003;121(5):749–57. PubMed
PMID: 12780789. skeletal bone destruction, primarily in the axial skeleton, and

ChaPter 80 Monoclonal Gammopathies 1081


may also result in anemia. Myeloma patients often present with serum (SPEP) and/or of urine (UPEP) from a 24-hour collection
6
pain and hypercalcemia caused by osteolytic lesions and/or combined with SIF and urine (UIF). SPEP will demonstrate
pathological fractures. Overproduction of M-protein over time a localized band or peak in 82% of patients with myeloma
can contribute to renal failure, hyperviscosity syndrome, and/ (Figs. 80.1 and 80.2).
or recurrent bacterial infections. Addition of serum protein immunofixation electrophoresis
increases the sensitivity to 93%. Adding either the sFLC assay or
Epidemiology urine monoclonal protein studies (UPEP and UIF) increases the
MM accounts for approximately 1% of all cancers and slightly diagnostic sensitivity to 97% or more. The very rare patient who
more than 10% of hematological malignancies in the United lacks detectable M-protein by any of these tests is considered to
States. The annual incidence in the United States is approximately have “nonsecretory myeloma.” Among the 20% with no localized
4 to 5 per 100 000. The incidence of MM in people of African band on SPEP, hypogammaglobulinemia is seen in approximately
background is two to three times that in Caucasians, whereas one-half, with no apparent abnormality in the remainder. 6
the risk is lower in Japanese and in Mexicans. MM is also slightly SIF confirms the presence of an M-protein and determines
more frequent in men than in women with an incidence ratio its type. The malignant plasma cells can produce a complete
of approximately 1.4 : 1. The median age at diagnosis is 66 years; clonal immunoglobulin molecule consisting of heavy and light
only 10% and 2% of patients are diagnosed before 50 and 40 chains, light chains alone, or neither, with the following frequen-
years, respectively. The risk of developing MM is approximately cies on serum immunofixation electrophoresis: IgG 52%, IgA
3.7-fold higher for persons with a first-degree relative diagnosed 21%, IgD 2%, IgM 0.5%, biclonal 2%, kappa or lambda light
with MM. 6 chain only 16%, and negative 17%. 7
Kappa is the predominant light chain compared with lambda
Clinical Presentation by a ratio of 2 to 1, with the exception that lambda light chains
Most patients with MM present with signs and symptoms related are more common in IgD myeloma and myeloma associated
to the infiltration of plasma cells into the bone or other organs with amyloidosis. The level of one of the major uninvolved
as well as kidney damage from excess light chains. Patients often immunoglobulins (i.e., IgM or IgA in the case of IgG myeloma)
complain of fatigue and display pallor upon physical examination. 6 is reduced in 91% of patients overall, and both isotypes are
Bone pain, particularly in the back or chest, and less often reduced in 73%. Up to 20% of myeloma patients have only
in the extremities, is present at the time of diagnosis in approxi- monoclonal light chains in the serum or urine. Approximately
mately 60% of patients. The pain is usually induced by movement 3% of patients with MM have no M-protein in the serum or
and does not occur at night except with change of position. The urine on immunofixation at the time of diagnosis. In approxi-
patient’s height may be reduced by several inches as a result of mately 60% of patients with myeloma who have a normal serum
vertebral collapse. Plasmacytomas of the ribs occur and can and urine immunofixation, monoclonal sFLC can be detected
6
6
present either as expanding costal lesions or soft-tissue masses. in the serum using FLC assays. M-proteins can increase the
Radiculopathy, usually in the thoracic or lumbosacral area, is serum viscosity and erythrocyte sedimentation rate (ESR). The
the most common neurological complication of MM and is due ESR is >20 mm/h in 84% and >100 mm/h in one-third of patients
to compression of the nerve by a paravertebral plasmacytoma with MM.
or, in rare instances, by the bone collapse itself. 6 The sFLC assay measures kappa and lambda light chains that
Extramedullary plasmacytomas (EP) are seen in approximately are unbound to heavy chains in the serum. These assays typically
7% of patients with MM at the time of diagnosis. An additional report out three values: quantitation of free kappa chains,
6% of patients will develop EP later during the course of disease, quantitation of free lambda chains, and the kappa/lambda ratio.
and these can present as large, purplish subcutaneous masses. The kappa/lambda ratio is obtained by dividing the free kappa
Spinal cord compression from an extramedullary plasmacytoma value by the free lambda quantitation (For example, the kappa/
or a bone fragment due to fracture of a vertebral body occurs lambda ratio of a patient with a kappa value of 6.32 mg/dL and
in approximately 5% of patients. Intracranial plasmacytomas a lambda value of 0.51 mg/dL would have a kappa/lambda ratio
are rare and almost always represent extensions of myelomatous of 12.4). The kappa/lambda ratio currently plays a key role in
lesions of the skull or plasmacytomas involving the clivus or the diagnosis and management of patients with monoclonal
6
base of the skull. Patients with MM are at increased risk for gammopathies. The normal kappa/lambda FLC ratio is 0.26 to
infection with Streptococcus pneumoniae and gram-negative 1.65. Abnormal FLC ratios outside this range are usually seen
organisms representing the most frequent pathogens. 6 in clonal plasma cell disorders when there is excess production
of one type of light chain (kappa or lambda). Abnormal FLC
Laboratory Findings ratios are seen in approximately 90% of patients with MM.
Biochemical Tests Patients with otherwise asymptomatic myeloma who have an
The serum creatinine concentration is increased in almost one-half involved/uninvolved FLC ratio of 100 or greater have a risk of
of patients at diagnosis. Hypercalcemia was found in 28% of progression to end-organ damage in the next 2 years of approxi-
one series of patients with MM at the time of diagnosis. Serum mately 80%. For this reason, an involved/uninvolved FLC ratio
calcium greater than 11 mg/dL has been found in 13% of patients of 100 or more is now considered diagnostic of MM. 6
and can require emergent treatment. Elevation of the serum
calcium may be due to binding of the monoclonal protein to Hematology
calcium. A low anion gap may be present due to severe hyper- A normocytic, normochromic anemia (hemoglobin ≤12 g/dL)
calcemia and the presence of a cationic IgG molecule. 6 is present in 73% at diagnosis and in 97% at some time during
The vast majority (97%) of patients with MM will have an the course of MM. Macrocytosis (mean corpuscular volume
M-protein produced and secreted by the malignant plasma cells. >100 fL) has been shown to be present in 9% of patients. The most
M-proteins can be detected by protein electrophoresis of the frequent findings on peripheral smear are rouleaux formation,

1082 Part eight Immunology of Neoplasia


Ref G A M K L

0.305
0.290
0.274
0.259
0.244
0.229
0.214
0.198
0.183
0.168
0.153
0.137
0.122
0.107
0.091
0.078
0.061
0.048
0.030
0.015
0.000 B
A 0 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300
Ref G A M K L
0.40
0.38 1
0.36
0.34
0.32
0.30
0.28
0.26
0.24
0.22
0.20
0.18
0.16
0.14
0.12
0.10
0.08
0.06
0.04
0.02
0.00
0 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300
C D
Fig 80.1 Electropherogram (A) and Immunofixation Electrophoresis (B) of Serum from a
Normal Patient and a Myeloma Patient (C) and (D). Note the M-spike in the gamma region
of the electropherogram and the immunoglobulin G (IgG) kappa band visible in the immunofixation
of the myeloma patient specimen.
leukopenia, and thrombocytopenia. Rouleaux formation is Mature plasma cells are oval with abundant basophilic cytoplasm.
particularly common in patients with elevated serum protein The nucleus is round and eccentrically located with a marked
levels, as noted in the discussion of MGUS. Monoclonal plasma perinuclear cytoplasmic clearing, termed a hof. 80. The nucleus
cells are rarely seen in the peripheral smear in patients with MM; contains “clock face“ or “spoke wheel“ chromatin without nucleoli.
a detectable absolute peripheral blood plasma cell count ≥100 Immature plasma cells have dispersed nuclear chromatin,
cells/µL (≥0.1 × 10/L) is found in approximately 10%. Evaluation prominent nucleoli, and a high nuclear to cytoplasmic ratio (Fig.
6
for circulating monoclonal plasma cells utilizes a slide-based 80.3). The cytoplasm of myeloma cells may contain condensed
immunofluorescence assay, a two-color immunoassay technique or crystallized cytoplasmic immunoglobulin resulting in the
+
−
(ELISPOT), or flow cytometry by gating on CD38 /CD45 cells. 6 following unusual findings, which are not limited to MM: multiple
pale bluish-white, grapelike accumulations, cherry-red refractive
Bone Marrow Evaluation round bodies, vermilion staining glycogen-rich IgA, overstuffed
Bone marrow aspirates and biopsies are key to the diagnosis of fibrils, and crystalline rods.
MM. The bone marrow plasma cell percentage should be estimated Immunohistochemical staining, immunofluorescent studies,
from a core biopsy specimen. However, if the percentage of plasma and flow cytometry detect either kappa or lambda light chains,
cells in the aspirate and core biopsy differ, the higher value should but not both, in the cytoplasm of bone marrow plasma cells
be used. Flow cytometry is not used to determine bone marrow in patients with MM while surface immunoglobulin is absent.
plasma cell percentage for diagnostic purposes. Clonality can The normal kappa/lambda ratio in the bone marrow is 2 : 1. A
be established by demonstrating kappa/lambda light chain ratio of more than 4 : 1 or less than 1 : 2 is considered to meet
6
restriction using flow cytometry, immunohistochemistry, or the definition of kappa or lambda monoclonality, respectively.
immunofluorescence. 6 As with normal plasma cells, myeloma cells express CD79a,
The bone marrow of the vast majority of patients contains p63, CD138, and CD38. In contrast to normal plasma cells,
10% or more clonal plasma cells. However, due to patchy bone myeloma cells infrequently express CD19. Approximately 70%
marrow involvement, a bone marrow aspirate and biopsy may of myeloma cells express CD56, which is typically negative in
6
show less than 10% plasma cells in approximately 4% of patients. normal plasma cells and in PCL. 6

ChaPter 80 Monoclonal Gammopathies 1083


Ref GAM K L fK fL
0.200
0.190
0.180
0.170
0.160
0.150
0.140
0.130
0.120
0.110
0.100
0.090
0.080
0.070
0.060
0.050
0.040
0.030
0.020
0.010
0.000 B
A 0 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300
0.200 Ref GAM K L fK fL
0.190
0.180
0.170
0.160
0.150
0.140
0.130
0.120
0.110
0.100
0.090
0.080
0.070
0.060
0.050
0.040 1
0.030
0.020
0.010
0.000
D
0 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300
C
Fig 80.2 Electropherogram (A) and Immunofixation Electrophoresis (B) of Urine from a
Normal Patient and a Myeloma Patient (C and D). Note the M-spike in the gamma region
seen in the myeloma patient specimen. Both a monoclonal free kappa band as well as a monoclonal
intact immunoglobulin band is visible in the urine immunofixation of the myeloma patient, indicating
the presence of both monoclonal free kappa light chains (Bence-Jones protein) and monoclonal
immunoglobulin M-protein in the patient’s urine.

There is no specific cytogenetic abnormality that is typical of the spine each occur in approximately 20%. The most frequent
or diagnostic of MM. The majority of myeloma tumors have sites of involvement include areas with active hematopoiesis,
genetic abnormalities that can be detected with sensitive molecular such as the vertebral bodies, skull, thoracic cage, pelvis, and
genetic techniques, such as interphase FISH. In contrast, only proximal humeri and femurs. 6
20–30% of patients will have cytogenetic abnormalities detected Low-dose whole-body computed tomography (CT), magnetic
in bone marrow plasma cells by conventional karyotyping; this resonance imaging (MRI), and positron emission tomography
is due to the low number of metaphases in myeloma cells. 6 (PET)/CT scans are helpful in patients who have bone pain
but no abnormalities on routine roentgenograms (Fig. 80.5).
Radiography MRI can detect diffuse and focal bone marrow lesions in
A metastatic bone survey with plain radiographs including the patients with MM without osteopenia or focal osteolytic lesions
humeri and femurs is a key component to the evaluation of a on standard metastatic bone surveys, and bone surveys can
patient suspected of having MM (Fig. 80.4). The skeletal survey detect lesions not found on MRI of the axial skeleton. PET/
for patients with MM includes a posteroanterior view of the CT scanning using fluorine-18-labeled fluorodeoxyglucose
chest; anteroposterior and lateral views of the cervical spine, (FDG) appears to correlate with areas of active lytic bone
thoracic spine, lumbar spine, humeri, and femurs; anteroposterior disease. 6
and lateral views of the skull; and anteroposterior view of the
pelvis. Symptomatic areas are also imaged. Conventional skeletal Diagnosis
surveys reveal punched-out lytic lesions, diffuse osteopenia, or The physical examination, laboratory tests, and imaging studies
fractures in nearly 80% of patients with MM at the time of recommended by the IMWG for the diagnosis of MM are
8
diagnosis. outlined in Table 80.2. The 2015 IMWG criteria for the diagnosis
Focal lytic lesions are found in nearly 60% of patients; of MM require the fulfillment of criteria as outlined in Table
osteoporosis, pathological fractures, or compression fractures 80.3. 8

1084 Part eight Immunology of Neoplasia



KeY CONCePtS Management
Multiple Myeloma Patients who have end-organ damage attributable to the underly-

• A cancer of plasma cells. ing plasma cell disorder have MM and require therapy. The
• Organ damage due to plasma cell proliferation and M-protein following findings, if attributable to the underlying plasma
overproduction. cell disorder, are clear indications for treatment: anemia,
• Pathophysiology of multiple myeloma consists mainly of CRAB: hypercalcemia, renal insufficiency, lytic bone lesions or severe
hyperCalcemia, Renal, Anemia, and Bone. osteopenia, and extramedullary plasmacytoma (excluding
• History and physical examination, laboratory tests, bone marrow testing, SEP). Asymptomatic patients who have one of the follow-
9
and imaging all play key roles in the diagnosis and management of
myeloma. ing biomarkers are also considered to have MM that requires
• Prognosis and treatment vary depending on risk stratification. therapy: ≥60% clonal plasma cells in the bone marrow, involved/
uninvolved FRL ratio of ≥100, more than one focal bone lesion
on MRI. 9
High-dose chemotherapy with autologous hematopoietic
stem cell transplantation (HSCT) is the preferred therapy
for patients with standard- or intermediate-risk myeloma.
Patients eligible for HSCT receive induction therapy for 2–4
months prior to stem cell collection to reduce the number
of tumor cells in the bone marrow and peripheral blood, to
9
lessen symptoms, and to mitigate end-organ damage. The
preferred induction chemotherapy regimen depends on the risk
stratification.
Patients ineligible for HSCT receive induction lenalidomide
plus dexamethasone. In contrast, those receiving alkylator- or
bortezomib-based regimen are treated for approximately 12 to
9
18 months and then observed until progression. Patients with
high-risk MM do poorly with all conventional treatment options.
All patients should be evaluated before each treatment cycle to
determine the degree of disease response.
Patients with one or more lesions on skeletal radiographs
and those with osteopenia should be given bisphosphonate
therapy. Pneumococcal and influenza vaccines should also be
given to all patients. Patients with MM frequently develop
complications related to their disease, including hypercalcemia,
renal insufficiency, infection, and skeletal lesions, which require
Fig 80.3 Photomicrograph of Plasma Cells in a Bone Marrow specific treatment. 9
Biopsy From a Myeloma Patient. Plasma cells show atypical Almost all patients with MM who survive initial treatment
features, including visible nucleoli, multinucleation, high nuclear/ will eventually relapse and require further therapy. Treatment
cytoplasmic ratios, and increased size. (Courtesy Dr. Mikhail options for patients with relapsed or refractory MM include
Roshal, Department of Pathology, Memorial Sloan Kettering HSCT, a reinitiation of the previous chemotherapy regimen, or
Cancer Center.) a trial of a new regimen. 9






















A B
Fig 80.4 Conventional Radiographs Showing Myeloma Osteolytic Lesions in the (A) Skull
and (B) Humerus. (Courtesy Dr. Jonathan Landa, Department of Radiology, Memorial Sloan
Kettering Cancer Center.)

ChaPter 80 Monoclonal Gammopathies 1085





















A B























C
Fig 80.5 Myeloma lytic bone lesions visible in the sacral vertebra by (A) conventional computed
tomography (CT), (B) positron emission tomography (PET)/CT scans, and (C) magnetic resonance
imaging (MRI) demonstrate hypermetabolism in the sacrum. (Courtesy Dr. Jonathan Landa,
Department of Radiology, Memorial Sloan Kettering Cancer Center.)



Prognosis theraPeUtiC PriNCiPLeS
MM is a heterogeneous disease. Some patients progress rapidly Current Treatment of Multiple Myeloma (MM)
despite treatment, and others remain stable without therapy for
a number of years. Myeloma patients are typically staged using • Patients with standard- or intermediate-risk myeloma: High-dose
the Revised International Staging System (R-ISS). This system chemotherapy with autologous hematopoietic stem cell transplantation
stratifies patients with MM into three main risk categories at (HSCT) with induction chemotherapy regimen dependent on risk
time of diagnosis. 10 stratification.
• High-Risk Myeloma: Patients with t(14;16), t(14;20), or • Patients ineligible for HSCT: induction lenalidomide plus dexamethasone.
• Patients with high-risk MM do poorly with all conventional treatment
del17p13 by FISH are considered to have high-risk myeloma. options.
These patients have a median survival of approximately 2–3 • Patients should be evaluated before each treatment cycle to determine
years despite standard treatment. Patients with lactate dehy- degree of disease response.
drogenase (LDH) ≥2 times the institutional laboratory upper • Patients with one or more lesions on skeletal radiographs and those
limit of normal and those with features of primary are also with osteopenia should be given bisphosphonate therapy.
considered to have high-risk MM. • Almost all patients with MM who survive initial treatment will eventually
relapse and require further therapy.
• Intermediate-Risk Myeloma: Patients with t(4;14) or 1q+ by
FISH or deletion 13/hypodiploidy are considered to have
intermediate-risk MM.
• Standard-Risk Myeloma: All other patients with MM who SMOLDERING MULTIPLE MYELOMA
lack any of the high- or intermediate-risk cytogenetic
abnormalities or features are considered to have standard- SMM is defined as follows: M-protein ≥3 g/dL in serum and/or
risk MM; this includes patients with trisomies, t(11;14) and 10–60% bone marrow plasma cells, no end-organ damage (lytic
t(6;14). lesions, anemia, renal disease, or hypercalcemia) attributable to

1086 Part eight Immunology of Neoplasia



TABLE 80.2 exams, tests, and imaging TABLE 80.3 iMWg Criteria for the
Studies recommended by the international Diagnosis of Multiple Myeloma
Myeloma Working group for the Diagnosis Clonal bone marrow plasma cells ≥10% or biopsy-proven bony or
of Multiple Myeloma extramedullary plasmacytoma and any one or more of the following
CRAB (hyperCalcemia, Renal, Anemia, and Bone) features and
1. History and Physical Examination myeloma defining events (MDEs):
2. Routine Testing
• Complete blood count with differential and peripheral blood 1. Evidence of end-organ damage that can be attributed to the
underlying plasma cell proliferative disorder, specifically:
smear review.
• Chemistry panel including calcium and creatinine. • Hypercalcemia: serum calcium >0.25 mmol/L (>1 mg/dL) higher
than the upper limit of normal or >2.75 mmol/L (>11 mg/dL)
• Serum protein electrophoresis, immunofixation electrophoresis.
• Nephelometric quantitation of immunoglobulins. • Renal insufficiency: creatinine clearance <40 mL per minute or
serum creatinine >177 µmol/L (<2 g/dL [20 g/L])
• Routine urinalysis, 24-hour urine collection for proteinuria,
electrophoresis, and immunofixation electrophoresis. • Anemia: hemoglobin value of >20 g/L below the lowest limit of
• Quantification of both urine M-component level and albuminuria. normal, or a hemoglobin value <10 g/dL (100 g/L)
3. Bone Marrow Testing: Obtain an aspirate plus trephine biopsy with • Bone lesions: one or more osteolytic lesions on skeletal
radiography, CT, or PET/CT. If bone marrow has <10% clonal
testing for cytogenetics, fluorescent in situ hybridization (FISH), and
immunophenotyping. plasma cells, more than one bone lesion is required to
4. Imaging distinguish from solitary plasmacytoma with minimal marrow
involvement
• Bone survey including spine, pelvis, skull, humeri, and femurs. 2. Any one or more of the following biomarkers of malignancy
• The IMWG now recommends the use of low-dose whole-body
CT (LDWBCT) or MRI in the workup of smoldering multiple (MDEs):
• 60% or greater clonal plasma cells on bone marrow examination
myeloma (SMM) and solitary plasmacytoma.
• The IMWG now recommends that one of PET/CT, LDWBCT, or • Serum-involved uninvolved free light chain ratio of 100 or
greater, provided the absolute level of the involved light chain is
MRI of the whole body or spine be done in all patients with
suspected smoldering myeloma, with the exact imaging modality at least 100 mg/L (a patient’s “involved” free light chain—either
kappa or lambda—is the one that is above the normal reference
determined by availability and resources.
• Clear evidence of one or more sites of osteolytic bone range; the “uninvolved” free light chain is the one that is
typically in, or below, the normal range)
destruction (≥5 mm in size) seen on CT (including LDWBCT) or
PET/CT does fulfill the criteria for bone disease in multiple • More than one focal lesion on MRI that is at least 5 mm or
greater in size
myeloma and should be regarded as meeting the CRAB
(hyperCalcemia, Renal, Anemia, and Bone) requirement, whether Source: Rajkumar SV, Dimopoulos MA, Palumbo A, Blade J, Merlini G, Mateos MV,
or not the lesions can be visualized on skeletal radiography. et al. International Myeloma Working Group updated criteria for the diagnosis of
• Increased uptake on PET/CT alone is not adequate for the multiple myeloma. Lancet Oncol. 2014;15(12):e538–48. doi: 10.1016/S1470-
diagnosis of multiple myeloma; evidence of underlying osteolytic 2045(14)70442-5. PubMed PMID: 25439696.
bone destruction is needed on the CT portion of the
examination.
• Bone densitometry studies are not sufficient to determine the
presence of multiple myeloma.
• The IMWG no longer recommends the presence of osteoporosis
or vertebral compression fractures in the absence of lytic lesions abnormal plasma cell immunophenotype, and abnormalities on
as being sufficient evidence of bone disease for purposes of the imaging (MRI or PET/CT). 11,12 Patients with SMM should not
diagnostic criteria. be treated but instead closely monitored with follow-up every
3–4 months with treatment initiated upon progression to MM. 11
Source: Rajkumar SV, Dimopoulos MA, Palumbo A, Blade J, Merlini G, Mateos MV,
et al. International Myeloma Working Group updated criteria for the diagnosis of
multiple myeloma. Lancet Oncol 2014;15(12):e538–48. doi: 10.1016/S1470- NONSECRETORY MYELOMA
2045(14)70442-5. PubMed PMID: 25439696.
Approximately 3% of patients with MM have no M-protein in
the serum or urine based on immunofixation electrophoresis at
the time of diagnosis. Patients with myeloma who have normal
serum and urine immunofixation electrophoresis as well as a
the underlying plasma cell disorder, and no amyloidosis. SMM normal serum FLC ratio are considered to have true nonsecretory
is differentiated from MGUS by the quantity of M-protein in myeloma. Of these, approximately 85% will have M-protein that
serum (≥3 g/dL), urine (≥500 mg/24 h), and/or the percentage can be detected in the cytoplasm of the neoplastic plasma cells
of clonal bone marrow plasma cells (10–60%). by immunohistochemistry; however, they have impaired secretion
of this protein. The other 15% do not have immunoglobulin
Prognosis and Management detectable in the plasma cells. 6
Patients with SMM will progress to symptomatic MM or AL Patients with true nonsecretory myeloma need to be monitored
amyloidosis at an approximate rate of 10%/year for the first 5 mainly on the basis of imaging tests and bone marrow studies.
years, 3%/year for the next 5 years, and 1–2%/year for the fol- Patients with nonsecretory MM are not at risk for myeloma
lowing 10 years. The majority of patients with SMM will progress, kidney as long as light chains cannot be detected in the urine,
11
with a median time to progression of 4.8 years. The following but they are at risk for other complications of MM. 6
factors are associated with an increased risk of progression:
abnormal free light chain κ/λ ratio (<0.125 or >8.0), bone marrow OLIGOSECRETORY MYELOMA
clonal plasma cells 50–60%, serum M-protein ≥3 g/dL (or progres-
sive increase in M-protein level), IgA SMM, immunoparesis, Approximately 5–10% of patients with MM have oligosecretory
high-risk genetic features, increased circulating plasma cells, myeloma at the time of diagnosis as defined by the absence of

ChaPter 80 Monoclonal Gammopathies 1087


measurable disease in serum or urine using the following anemia or renal insufficiency attributable to the underlying plasma
parameters: serum M-protein <1 g/dL and urine M-protein cell disorder.
<200 mg/24 hours. 6
Monitoring these patients is difficult using the standard serum Diagnosis
and urine electrophoretic tests. In most of these patients, the The diagnosis of SPB requires the following: biopsy-proven
serum FLC assay can be used to monitor the disease, provided solitary tumor of bone with evidence of clonal plasma cells,
that the serum FLC ratio is abnormal and the involved (affected) metastatic bone survey and either CT (PET/CT) or MRI of the
FLC level is ≥10 mg/dL. Patients with oligosecretory disease spine and pelvis showing no other lytic lesions, bone marrow
may need to be monitored with imaging and bone marrow aspirate, and biopsy that contains no clonal plasma cells as well
studies, particularly if the baseline FLC levels are unmeasurable as no findings attributable to clonal plasma cell proliferative
8
(<10 mg/dL). disorder (anemia, hypercalcemia, or renal insufficiency). The
presence of an M-protein does not exclude the diagnosis of SPB,
PLASMA CELL LEUKEMIA as a low level of M-protein may be present in 30–75% of cases.
This M-protein may or may not disappear with treatment.
PCL is a very rare and aggressive variant of MM. It is characterized
by high levels of plasma cells circulating in peripheral blood. Management and Prognosis
PCL may originate de novo (primary PCL) or as a secondary The primary treatment for patients with SPB is localized radiation
transformation of MM (secondary PCL). The incidence of PCL therapy (RT). Surgery may be required for patients with structural
in Europe has been estimated at 4 cases/10 000 000 persons/year. instability of the bone, retropulsed bone, or rapidly progressive
Secondary PCL occurs as a progression of disease in 1–4% of symptoms from cord compression. Bisphosphonates are not
all cases of MM. 13 recommended for patients with SPB, except in the setting of
underlying osteopenia. 14
Clinical Presentation and Diagnosis The median overall survival of patients with SPB is approxi-
Patients with PCL typically present with signs and symptoms mately 10 years. Overall survival rates at 5 and 10 years are
13
similar to those seen in MM as well as in other leukemias. The approximately 75% and 45%, respectively, with corresponding
diagnosis of PCL is based upon the evaluation of peripheral disease-free survival rates of 45% and 25%. A little more than
blood smear, bone marrow aspirate and biopsy, and protein half of patients with SPB will eventually develop overt MM.
electrophoresis. PCL diagnosis is confirmed when a monoclonal Patients with true SPB who meet the strict criteria listed above
population of plasma cells is present in the peripheral blood have a recurrence/progression rate of approximately 10% within
with an absolute plasma cell count exceeding 2000/µL and 20% 3 years. 14
of the peripheral blood white cells. 13
Management and Prognosis SOLITARY EXTRAMEDULLARY PLASMACYTOMA
Treatment generally consists of induction therapy followed by SEPs are plasma cell tumors that arise outside the bone marrow.
HSCT, if eligible for HSCT, and chemotherapy alone for those SEP refers to a solitary nonosseous plasma cell neoplasm in the
13
ineligible for HSCT. The prognosis of PCL is poor with a median absence of any other sign of MM. The SEP lesions are most
survival of 7–11 months. Median survival for PCL occurring often located in the head and neck region, mainly in the upper
after refractory or relapsing MM is 2–7 months. 13 aerodigestive tract, but may also occur in the gastrointestinal
tract, urinary bladder, central nervous system, thyroid, breast,
SOLITARY PLASMACYTOMA OF BONE testes, parotid gland, lymph nodes, and skin. The median age at
diagnosis for SEP is 55 to 60 years; approximately two-thirds of
SPB is a localized tumor in the bone comprised of a single clone patients are male, and SEP accounts for approximately 3% of
of plasma cells in the absence of other features of MM. Approxi- plasma cell malignancies. 15
mately 5% of all cases of plasma cell disorders are SPB. The
incidence of SPB is approximately 0.15 cases/100 000 person-years Clinical Presentation
with approximately 450 new cases per year in the United States. Most patients present with symptoms related to the location of
The incidence is highest in patients of African background and the mass. Approximately 80% involve the upper respiratory tract
lowest in Asians and Pacific Islanders. Men are diagnosed twice (i.e., oronasopharynx and paranasal sinuses) and cause epistaxis,
as frequently as women. The median age at diagnosis is 55–65 nasal discharge, or nasal obstruction. Less common sites of
years compared with a median age at diagnosis of 71 years for involvement include the gastrointestinal tract, liver, lymph nodes,
patients with MM. 14 testes, skin, and central nervous system. Primary plasmacytoma
of the lung often presents as a pulmonary nodule or hilar mass
Clinical Presentation with or without hemoptysis. 15
Most patients present with skeletal pain or a pathological fracture
of the affected bone. Patients with vertebral involvement may Diagnosis
have severe back pain or neurological compromise. Less com- By definition, patients with SEP do not have anemia, hypercal-
monly, SPB can extend into the surrounding soft tissue, resulting cemia, renal insufficiency, or bone lesions attributable to the
in a palpable mass. The most common bones involved are those underlying plasma cell disorder. The diagnosis of SEP requires
with active hematopoiesis; the axial skeleton is more commonly the following: biopsy-proven extramedullary tumor with evidence
involved than is the appendicular skeleton, and involvement of of clonal plasma cells, metastatic bone survey and either MRI
the distal appendicular skeleton below the knees or elbows is or PET/CT of the spine and pelvis that show no lytic lesions,
14
extremely rare. By definition, patients with SPB do not have bone marrow aspirate and biopsy without clonal plasma cells,

1088 Part eight Immunology of Neoplasia


and absence of anemia, hypercalcemia, or renal insufficiency TABLE 80.4 iMWg Diagnostic Criteria for
attributable to a clonal plasma cell proliferative disorder. 16 the Diagnosis of POeMS Syndrome
Some patients may have a small amount of M-protein, usually
IgA, in the serum or urine, which often disappears after treatment. Both Mandatory Criteria:
The bone marrow of patients with SEP does not have clonal 1. Polyneuropathy
plasma cells, whereas some patients with a solitary extramedullary 2. Monoclonal plasma cell proliferative disorder
lesion may demonstrate up to 10% clonal plasma cells, in which Plus at Least One Major Criterion:
case the diagnosis is SEP with minimal marrow involvement. 15 3. Osteosclerotic or mixed sclerotic/lytic lesion visualized on plain
films or computed tomography (CT)
Management and Prognosis 4. Castleman disease
The treatment of choice for SEP is RT. Small lesions may be 5. Elevated serum or plasma vascular endothelial growth factor
cured with surgery alone; no adjuvant RT is indicated unless (VEGF) levels (at least three to four times the upper limit of
there is suspicion of residual local disease. 15 normal)
Less than 7% of patients with SEP will develop a local recur- Plus at Least One Minor Criterion:
rence after tumoricidal radiation. Approximately 10–15% of
patients ultimately will develop MM; the progression rate is 1. Organomegaly (splenomegaly, hepatomegaly, or
lymphadenopathy)
higher (20%) in patients with SEP with minimal marrow involve- 2. Extravascular volume overload (peripheral edema, ascites, or
ment. Five-year overall survival rates range from 40–85%. 15 pleural effusion)
3. Endocrinopathy (adrenal, thyroid, pituitary, gonadal, parathyroid, or
pancreatic disorder excluding diabetes mellitus or hypothyroidism)
POEMS SYNDROME 4. Skin changes (hyperpigmentation, hypertrichosis, glomeruloid
hemangiomata, plethora, acrocyanosis, flushing, white nails)
POEMS (polyneuropathy, organomegaly, endocrinopathy, 5. Papilledema
monoclonal protein, skin changes) syndrome is characterized 6. Thrombocytosis or polycythemia
by the presence of a monoclonal plasma cell disorder, peripheral
neuropathy, and one or more of the following features: osteo- POEMS, polyneuropathy, organomegaly, endocrinopathy, monoclonal protein, skin
sclerotic myeloma, Castleman disease, increased levels of serum changes.
Source: Rajkumar SV, Dimopoulos MA, Palumbo A, Blade J, Merlini G, Mateos MV,
vascular endothelial growth factor (VEGF), organomegaly, et al. International Myeloma Working Group updated criteria for the diagnosis of
endocrinopathy, edema, typical skin changes, and papilledema. multiple myeloma. Lancet Oncol 2014;15(12):e538–48. doi: 10.1016/S1470-
The cause of POEMS syndrome is unknown, although chronic 2045(14)70442-5. PubMed PMID: 25439696.
overproduction of proinflammatory and other cytokines such
as VEGF appears to be a major feature of this disorder. POEMS
syndrome is a rare disorder, but the exact incidence is unknown.
POEMS syndrome commonly presents in the fifth to sixth macytic cells and a monoclonal IgM gammopathy in the blood.
decade. 17 WM is a rare disorder with an incidence of approximately 3/
million people/year, with 1400 new cases diagnosed in the United
Clinical Presentation States each year. The median age at diagnosis is 64 years; less
By definition, all patients have peripheral neuropathy and a than 1% of patients are diagnosed under 40 years of age, and
monoclonal plasma cell disorder, almost always of the lambda approximately 60% are males. WM is more common in Caucasians
light chain type. In addition, almost all patients have osteosclerotic than in other ethnic groups, and familial predisposition is present
lesions, whereas Castleman disease is present in approximately in up to 20%. 18
15%. 17
Clinical Presentation
Diagnosis Patients usually present in their seventh decade with symptoms
The IMWG has developed a series of criteria required for the related to the infiltration of the hematopoietic tissues or to the
diagnosis of POEMS syndrome; other signs and symptoms that effects of monoclonal IgM in the blood. WM most commonly
do not fall into these criteria may also be present, as outlined presents with pallor, oronasal bleeding, systemic complaints
in Table 80.4. 8 (weakness, fatigue, weight loss, fever, night sweats), and organo-
megaly (enlarged lymph nodes, spleen, and/or liver). Involvement
Management and Prognosis of the bone or kidneys is uncommon. 18
There is no standard treatment for POEMS syndrome. Those An important presentation includes central nervous system
with limited disease may be treated with RT. Patients with signs and symptoms due to the hyperviscosity syndrome (blurring
widespread bone lesions may be treated with therapy similar to or loss of vision, headache, ataxia, dementia, stroke, or coma).
17
that used for MM. POEMS syndrome is chronic, and patients A classic finding in WM associated with hyperviscosity is the
with POEMS syndrome survive three times longer than those presence of dilated, segmented, and tortuous retinal veins. Another
with MM. The natural history is one of progressive peripheral major presentation involves neurological symptoms such as
neuropathy until the patient is bedridden. Death usually occurs paresthesias and weakness as well as cranial nerve palsies and
from inanition or a terminal bronchopneumonia. 17 sudden deafness. Patients with WM who are asymptomatic are
considered to have SWM. 18
WALDENSTRÖM MACROGLOBULINEMIA Laboratory Findings and Diagnosis

WM is a rare clinicopathological entity demonstrating 10% or Patients with WM must have 10% or greater infiltration of the
greater infiltration of the bone marrow by clonal lymphoplas- bone marrow by clonal lymphoplasmacytic cells and a monoclonal

ChaPter 80 Monoclonal Gammopathies 1089


IgM gammopathy in the blood. The diagnosis of WM is made present with symptoms due to hyperviscosity should undergo
when the following two criteria are met: (i) presence of an IgM immediate therapeutic plasmapheresis, followed by chemotherapy
21
monoclonal paraprotein on serum immunofixation and (ii) 10% for control of the malignant clone. Median survival in WM is
or more of the bone marrow biopsy sample demonstrates infiltra- approximately 5–8 years from the time of diagnosis. However,
tion by small lymphocytes that exhibit plasmacytoid or plasma outcomes are highly variable. A staging system for WM has been
cell differentiation (lymphoplasmacytic features or lymphoplas- developed based on a prospective multicenter observational study
macytic lymphoma) with an intertrabecular pattern. This infiltrate outlined in Table 80.6. 22
+
should express a typical immunophenotype (e.g., surface IgM ,
+
−
−
+
+
+
+
+/−
CD5 , CD10 , CD19 , CD20 , CD22 , CD23 , CD25 , CD27 , HEAVY CHAIN DISEASES
−
+
−
19
FMC7 , CD103 , CD138 ). WM must be differentiated from
IgM MGUS, MM, chronic lymphocytic leukemia (CLL), and The HCDs are rare B-cell proliferative disorders characterized
mantle-cell lymphoma. by the production of an M-protein consisting of a portion of
the immunoglobulin heavy chain without a bound light chain.
Management and Prognosis Three types of HCD are recognized, based upon the class of
Many patients with WM are asymptomatic and can be observed immunoglobulin heavy chain produced by the malignant cell:
for months to years after the diagnosis is established before 1. Alpha HCD is a form of mucosa-associated lymphoreticular
requiring treatment. Asymptomatic patients with adequate tissue (MALT) lymphoma that is also called immuno-
hemoglobin and platelet levels are followed every 6 months with proliferative small intestinal disease (IPSID), Mediterranean
20
complete blood counts and monoclonal protein levels. Treatment lymphoma, or Seligmann disease.
is indicated in those who meet one or more of the conditions 2. Gamma HCD (Franklin disease) is typically associated with
presented in Table 80.5. 21 the presence of a systemic lymphoma, often of mixed
Symptomatic WM is treated with a regimen that incorporates lymphoid-plasmacytic character.
rituximab in combination with other agents. Symptomatic patients 3. Mu HCD has clinical features resembling small lymphocytic
with low tumor burden and minimally symptomatic patients lymphoma/CLL, often with distinctive vacuolated lymphocytes/
may be treated with the single agent rituximab. Patients who plasma cells in the bone marrow.
Clinical Presentation
The clinical presentation of the HCDs is that of a patient with
TABLE 80.5 Clinical and Laboratory a low-grade B-cell malignancy. Alpha HCD is a form of MALT
indications for initiation of treatment for lymphoma, with the same histological features of MALT-type
Waldenström Macroglobulinemia (WM) gastrointestinal lymphomas with marked plasma cell differentia-
Clinical indications for therapy initiation: tion. It is the most common form of HCD and occurs in patients
• Recurrent fever, night sweats, weight loss, fatigue from the Mediterranean region or Middle East, usually young
• Hyperviscosity males, and is often associated with relatively poor sanitation.
• Lymphadenopathy that is either symptomatic or bulky (≥5 cm in The gastrointestinal tract is most commonly involved in alpha
maximum diameter) HCD, resulting in abdominal pain, malabsorption with chronic
• Symptomatic hepatomegaly and/or splenomegaly diarrhea, steatorrhea, and loss of weight. Growth retardation,
• Symptomatic organomegaly and/or organ or tissue infiltration digital clubbing, and mesenteric lymphadenopathy may also be
• Peripheral neuropathy due to WM 23
present.
Laboratory indications for therapy initiation: Patients with gamma HCD typically present with systemic
• Symptomatic cryoglobulinemia symptoms, lymphadenopathy, splenomegaly, and/or anemia, and
• Cold agglutinin anemia occasionally with palatal and uvular swelling. The median age
• Immune hemolytic anemia and/or thrombocytopenia of patients with gamma HCD is 60 to 70 years, although the
• Nephropathy related to WM condition has been noted in persons younger than age 20.
• Amyloidosis related to WM Autoimmune manifestations are seen in about one-third of
• Hemoglobin ≤10 g/dL patients. 23
9
• Platelet count <100 × 10 /L
Mu HCD is the least common of the HCDs and has features
Source: Dimopoulos MA, Kastritis E, Owen RG, Kyle RA, Landgren O, Morra E, et al. resembling CLL/small lymphocytic lymphoma, although periph-
Treatment recommendations for patients with Waldenstrom macroglobulinemia (WM) eral adenopathy is less common than in CLL. Osteolytic lesions
and related disorders: IWWM-7 consensus. Blood 2014;124(9):1404–11. doi: 10.1182/ or pathological fractures occasionally have been reported in
blood-2014-03-565135. PubMed PMID: 25027391; PubMed Central PMCID:
PMC4148763. patients. Unlike those with alpha and gamma HCD, some patients
TABLE 80.6 Staging System for Waldenström Macroglobulinemia

Stage risk Characteristics Five-Year Survival Five-Year Progression-Free Survival
A Low B2M <3 mg/L and Hgb ≥12.0 g/dL 87% 83%
B Medium B2M <3 mg/L and Hgb <12.0 g/dL 63% 55%
C Medium B2M ≥3 mg/L and serum IgM ≥4.0 g/dL 53% 33%
D High B2M ≥3 mg/L and IgM <4.0 g/dL 21% 12%
B2M, Beta 2 microglobulin.
Source: Dhodapkar MV, Jacobson JL, Gertz MA, Rivkin SE, Roodman GD, Tuscano JM, et al. Prognostic factors and response to fludarabine therapy in patients with Waldenstrom
macroglobulinemia: results of United States intergroup trial (Southwest Oncology Group S9003). Blood 2001;98(1):41–8. PubMed PMID: 11418461.

1090 Part eight Immunology of Neoplasia


with mu HCD have increased free light chain secretion, and they LIGHT CHAIN AMYLOIDOSIS
may develop cast nephropathy or amyloidosis. 23
Amyloidosis refers to the deposition of fibrils composed of
Diagnosis low-molecular-weight subunits of normal serum proteins. AL
The diagnosis of the HCDs can be made on histopathological amyloidosis is a specific form of amyloidosis caused by the
examination of affected tissues and/or analysis of serum or urine deposition of monoclonal light chains. These light chains undergo
samples by electrophoresis and immunofixation electrophoresis. transformation to a beta-pleated fibrillar configuration. Deposi-
A typical M-spike in the serum or urine may not always be tion of these light chains in various organs results in organ failure.
present. On serum or urine immunofixation electrophoresis or AL amyloidosis is an uncommon disorder of older adults,
immunohistochemical staining of affected tissues, the abnormal and the exact incidence is unknown. In the United States, the
protein consists of a heavy chain without an associated light incidence is approximately 6–10 cases per million person-years.
chain. Immunohistochemical staining of affected tissue shows Age-specific incidence rates increase in each decade of life after
the presence of a clonal population of cells staining positively age 40 years. The median age at diagnosis is 64 years, and less
for a heavy chain but negative for both kappa and lambda light than 5% of patients are under the age of 40. There is a male
chains. 23 predominance, with men accounting for 65–70% of patients. AL
Alpha HCD most commonly involves infiltration of the jejunal amyloidosis occurs in all races and all geographic locations. 24
mucosa with plasmacytoid cells, whereas the duodenum and
ileum are affected less often. The diagnosis depends upon the Clinical Presentation
recognition of a monoclonal alpha heavy chain without an AL amyloidosis is a systemic disorder that can present with a
associated light chain in the serum, urine, intestinal secretions, variety of symptoms, including significant proteinuria, edema,
or the cells infiltrating the intestinal mucosa. The serum protein hepatosplenomegaly, otherwise unexplained heart failure, and
electrophoretic pattern is normal in one-half of cases, and in carpal tunnel syndrome. Although virtually all patients have
the remainder an unimpressive broad band may appear in the multisystem amyloid deposition, it is not uncommon for a patient
alpha-2 or beta mobility regions. The amount of alpha heavy to present with evidence of mainly one organ involvement.
chain in the urine is small. 23 Nonspecific systemic symptoms, including fatigue and
Gamma HCD is typically associated with a polymorphous unintentional weight loss, are common in patients with AL
infiltrate, including admixtures of lymphocytes, plasmacytoid amyloidosis. Renal involvement occurs in approximately 70%
lymphocytes, plasma cells, immunoblasts, and eosinophils. of patients and most often presents as asymptomatic proteinuria
The electrophoretic pattern often shows a broad-based band or nephrotic syndrome. Cardiac involvement is seen in approxi-
more suggestive of a polyclonal increase, but the gamma chains mately 60% of patients and typically is characterized by thickening
are monoclonal. The diagnosis is established by immunofixa- of the interventricular septum and ventricular wall. This can
tion electrophoresis, which shows that the abnormal protein lead to systolic or diastolic dysfunction with symptoms of heart
consists only of gamma heavy chains without associated light failure. Other manifestations that may be seen include sudden
chains. 23 death or syncope due to arrhythmia or heart block and, rarely,
Mu HCD is characterized by vacuolated plasma cells or angina or infarction due to accumulation of amyloid in the
lymphoid cells observed in the bone marrow, together with coronary arteries. 24
panhypogammaglobulinemia. The serum protein electrophoretic Hepatomegaly with or without splenomegaly is seen in as
pattern is usually normal, except for hypogammaglobulinemia. 23 many as 70% of patients. Potential gastrointestinal manifestations
include bleeding, gastroparesis, constipation, bacterial overgrowth,
Management and Prognosis malabsorption, and intestinal pseudo-obstruction resulting from
Initial treatment of alpha HCD consists of eradication of any dysmotility. 24
concurrent infection with appropriate antibiotics. Those with Mixed sensory and motor peripheral neuropathy and/or
symptomatic disease who do not respond adequately to antibiotics autonomic neuropathy is a prominent feature in AL amyloidosis.
may be treated with chemotherapy similar to that used for non- Symptoms of numbness, paresthesia, and pain are frequently
23
Hodgkin lymphoma, such as the CHOP regimen. Treatment noted, as in peripheral neuropathy of many other causes. Com-
of gamma HCD is indicated only for symptomatic patients. pression of peripheral nerves, especially the median nerve within
Appropriate chemotherapy includes melphalan plus prednisone the carpal tunnel, can cause more localized sensory changes.
or regimens used to treat B-cell lymphoma, such as the combina- Symptoms of bowel or bladder dysfunction and findings of
tion of cyclophosphamide, vincristine, and prednisone, with or orthostatic hypotension may be due to autonomic nervous system
without doxorubicin, or rituximab if the abnormal cells express damage. 24
23
CD20. Treatment for symptomatic patients with mu HCD is Amyloid infiltration of skeletal muscles may cause visible
similar to that employed in patients with CLL (e.g., glucocorti- enlargement. Macroglossia, or lateral scalloping of the tongue
coids, alkylating agents, fludarabine). 23 from impingement on the teeth, is characteristic of AL amyloid.
In the absence of therapy, alpha HCD typically is progressive Arthropathy may be due to amyloid deposition in joints and
and fatal. The prognosis of gamma HCD is variable and ranges surrounding structures. The “shoulder pad“ sign is visible
from the asymptomatic presence of a stable monoclonal heavy enlargement of the anterior shoulder due to fluid in the gleno-
chain in the serum or urine (e.g., MGUS) to a rapidly progressive humeral joint and/or amyloid infiltration of the synovial
downhill course over a short duration (e.g., highly aggressive membrane and surrounding structures. 24
lymphoma). Median survival is 7.4 years with an extremely wide Purpura, characteristically elicited in a periorbital distribution
23
range from 1 month to more than 21 years. The course of mu (raccoon eyes) by a Valsalva maneuver or minor trauma, is present
HCD is variable, and survival ranges from a few months to many in only a minority of patients, but it is highly characteristic of
years. 23 AL amyloidosis. Other signs of skin involvement include waxy

ChaPter 80 Monoclonal Gammopathies 1091


thickening, easy bruising, and subcutaneous nodules or plaques. of AL amyloidosis. Kidney or liver biopsy is positive for amyloid
Amyloidosis may also be directly associated with a bleeding deposition in over 90% of cases; however, a high success rate
diathesis. Approximately 10% of patients have coexisting MM. 24 can also be achieved by less invasive procedures, such as abdominal
fat pad aspirate, rectal biopsy, bone marrow biopsy, or skin
KeY CONCePtS biopsy. 24
Amyloid appears as a pink, amorphous, waxy substance with
Light Chain Amyloidosis a characteristic “cracking” artifact on hematoxylin and eosin–
• A specific form of amyloidosis caused by tissue deposition of mono- stained biopsies (Fig. 80.6a). The presence of amyloid fibrils can
clonal light chains. be confirmed by their characteristic appearance on electron
• A systemic disorder with multisystem amyloid deposition causing a microscopy and by their ability to bind Congo red or thioflavin-T
variety of nonspecific symptoms. (Fig. 80.6b and 80.6c). 24
• Demonstration of amyloid deposition in affected organs is key to Staining for kappa and lambda light chains, transthyretin,
diagnosis.
• Poor long-term prognosis due to organ dysfunction caused by amyloid and serum amyloid A can determine the type of amyloidosis.
24
deposition. Positive staining for kappa or lambda indicates AL amyloidosis.
Immunofluorescence microscopy using light chain–specific
antisera and other specialized techniques may show deposition
Laboratory Findings and Diagnosis of a monoclonal (lambda or kappa) light chain in the kidney
Demonstration of amyloid fibrils upon histological evaluation or other affected tissues. Laser microdissection of tissue followed
of an affected organ or a surrogate site is required for the diagnosis by analysis by tandem mass spectrometric proteomic analysis


























A B























C
Fig 80.6 Photomicrograph of Plasmacytoma With AL Amyloid Deposition. (A) Hematoxylin
and eosin stain, (B) Congo red staining without polarization, (C) Congo red staining under polarized
light. (Courtesy Dr. Mikhail Roshal, Department of Pathology, Memorial Sloan Kettering Cancer
Center.)

1092 Part eight Immunology of Neoplasia


may be helpful in identifying the type of amyloid protein (e.g., due to cardiac or hepatic failure as well as infection. There is,
monoclonal light chain versus other forms of amyloid proteins). however, significant variability in median survival, depending
An alternative to mass spectrometry is to use immunoelectron on the nature, number, and extent of organ involvement. Median
microscopy, if available. 24 survival may range from 4 to 6 months in patients diagnosed
Patients with AL amyloidosis frequently have chromosomal at an advanced stage, to in excess of 5 years in patients with
abnormalities, but there is no single chromosomal change that limited organ involvement. Two key prognostic factors include
is diagnostic of this disease. Chromosomal abnormalities reported the presence and severity of cardiac involvement and the presence
with moderate to high frequency in patients with AL amyloidosis of concurrent myeloma. 26
include t(11;14)(q13;q32), del(13q14), and gain of 1q21. 24
Most patients with AL amyloidosis have little or no intact MONOCLONAL IMMUNOGLOBULIN
monoclonal immunoglobulin, but are characterized by the DEPOSITION DISEASES
presence of monoclonal free light chain. The monoclonal light
chain type is lambda in approximately 70% of cases, kappa in Monoclonal immunoglobulin deposition diseases are a non-
25%, and biclonal in 5%. SPEP demonstrates a localized band amyloid monoclonal immunoglobulin chain deposition disease
or peak in less than 50% of patients with AL amyloidosis. that is caused by a clonal plasma cell proliferative disorder. These
Immunofixation electrophoresis detects a serum or urinary diseases are characterized by the deposition of immunoglobulin
monoclonal protein in nearly 90% of cases of AL amyloidosis. light or heavy chain fragments, leading to organ dysfunction.
When serum and urine immunofixation electrophoresis is There are two types of immunoglobulin deposition diseases:
combined with serum free light chain ratio analysis, a monoclonal heavy chain deposition disease (HCDD) and LCDD. They are
protein can be detected in virtually all cases. 24 differentiated from light chain and heavy chain amyloidosis in
Bone marrow biopsy specimens typically demonstrate a slightly that the light chain fragments do not have the necessary biochemi-
increased percentage of plasma cells that may appear morphologi- cal characteristics to form amyloid fibrils.
cally normal. Less commonly, the bone marrow demonstrates
overt myeloma or lymphoplasmacytic lymphoma. A clonal excess Clinical Presentation
of plasma cells (lambda or kappa) can also be demonstrated HCDD is extremely rare, with 37 cases reported in the literature
by immunoperoxidase staining or flow cytometric analysis of from 1992 to 2011. The majority of patients present with hyper-
24
specimens of involved bone marrow. Diagnostic criteria for tension, progressive renal dysfunction, anemia, and nephrotic
27
AL amyloidosis have been developed by the Mayo Clinic and syndrome with microhematuria. Patients with LCDD usually
the IMWG that require the presence of the four criteria outlined present with renal, cardiac, or hepatic involvement and may have
8
in Table 80.7. Approximately 2–3% of patients with AL amy- underlying MM or a lymphoproliferative disorder. 24
loidosis will not meet the requirement for evidence of a mono-
clonal plasma cell disorder listed. Laboratory Findings
In LCDD, a population of clonal plasma cells produces mono-
Management and Prognosis clonal light chain fragments that are deposited as granules in
Amyloidosis patients meeting the criteria in Table 80.8 are the tissues. The deposits in LCDD are almost always composed
25
considered to be eligible for autologous HSCT. Those eligible of kappa light chains; they are granular, not fibrillar, and do not
for HSCT may be treated with melphalan followed by autologous bind Congo red, thioflavin-T, or serum amyloid P component.
HSCT. Those not eligible for HSCT may be treated with Tissue deposits of fragments of monoclonal light chains are
melphalan/dexamethasone or a bortezomib-based regimen. 25 commonly seen and can occur in a variety of organs, including
AL amyloidosis typically has poor long-term prognosis. Death the kidney, heart, liver, and small intestine. Routine electrophoretic
is typically due to organ dysfunction, with major causes of death techniques may not demonstrate a monoclonal protein in the
serum or urine in some patients, but it may be detectable by
serum free light chain analysis. 26
TABLE 80.7 iMWg Diagnostic Criteria for
the Diagnosis of Light Chain amyloidosis

1. Presence of an amyloid-related systemic syndrome (e.g., renal, TABLE 80.8 eligibility Criteria for
liver, heart, gastrointestinal tract, or peripheral nerve involvement). autologous hSCt in Light Chain
To be included as a diagnostic criterion, the organ damage must be amyloidosis Patients
felt to be related to amyloid deposition and not to another common
disease, such as diabetes or hypertension. 1. Physiological age ≤70
2. Positive amyloid staining by Congo red in any tissue (e.g., fat 2. Troponin T <0.06 ng/mL
aspirate, bone marrow, or organ biopsy) or the presence of amyloid 3. N-terminal pro-brain natriuretic peptide (NT proBNP) <5000 ng/L
fibrils on electron microscopy. 4. Creatinine clearance ≥30 mL/min (unless on chronic stable dialysis)
3. Evidence that the amyloid is light chain–related is established by 5. Eastern Cooperative Oncology Group (ECOG) performance status
direct examination of the amyloid using spectrometry-based ≤2
proteomic analysis or immunoelectron microscopy. 6. New York Heart Association functional status Class I or II
4. Evidence of a monoclonal plasma cell proliferative disorder (e.g., 7. No more than two organs significantly involved (liver, heart, kidney,
presence of a serum or urine M-protein, abnormal serum free light or autonomic nerve)
chain ratio, or clonal plasma cells in the bone marrow). 8. No large pleural effusions
9. No dependency on oxygen therapy
Source: Rajkumar SV, Dimopoulos MA, Palumbo A, Blade J, Merlini G, Mateos MV,
et al. International Myeloma Working Group updated criteria for the diagnosis of HSCT, hematopoietic stem cell transplantation.
multiple myeloma. Lancet Oncol 2014;15(12):e538–48. doi: 10.1016/S1470- Source: Gertz MA. How to manage primary amyloidosis. Leukemia. 2012;26(2):191–8.
2045(14)70442-5. PubMed PMID: 25439696. doi: 10.1038/leu.2011.219. PubMed PMID: 21869840.

ChaPter 80 Monoclonal Gammopathies 1093


In HCDD, a population of clonal plasma cells produces both Lancet Oncol 2014;15(12):e538–48. doi:10.1016/S1470-2045(14)70442-5.
light and heavy monoclonal immunoglobulin chains or only PubMed PMID: 25439696.
short (truncated) heavy chains that are unable to form amyloid 9. Rajkumar SV. Overview of the management of multiple myeloma.
fibrils but are deposited as granules in the tissues. HCDD can UpToDate: Wolters Kluwer; 2016.
be identified by immunofluorescence staining of biopsy tissue 10. Palumbo A, Avet-Loiseau H, Oliva S, et al. Revised International Staging
System for Multiple Myeloma: A Report From International Myeloma
with anti–heavy chain antibodies. 26 Working Group. J Clin Oncol 2015;33(26):2863–9. doi:10.1200/
Management and Prognosis JCO.2015.61.2267. PubMed PMID: 26240224, PubMed Central PMCID:
PMC4846284.
The therapeutic approaches for both LCDD and HCDD are similar 11. Rajkumar SV. Smoldering multiple myeloma. UpToDate: Wolters Kluwer;
to those employed for patients with MM. Serial free light chain 2016.
26
assays can be used to monitor response to treatment in LCDD. 12. Kristinsson SY, Holmberg E, Blimark C. Treatment for high-risk
The prognoses of LCDD and HCDD vary considerably, depending smoldering myeloma. N Engl J Med 2013;369(18):1762–3. doi:10.1056/
on the nature, number, and extent of organ involvement. 26 NEJMc1310911#SA1. PubMed PMID: 24171527.
13. Rajkumar SV. Plasma cell leukemia. UpToDate: Wolters Kluwer; 2016.
14. Rajkumar SV. Diagnosis and management of solitary plasmacytoma of
ON the hOriZON bone. UpToDate: Wolters Kluwer; 2016.
15. Rajkumar SV. Diagnosis and management of solitary extramedullary
• Two monoclonal antibody drugs (mAb) have recently been approved plasmacytoma. UpToDate: Wolters Kluwer; 2016.
by the FDA for the treatment of multiple myeloma, with more on the
way. 16. Kyle RA, Rajkumar SV. Criteria for diagnosis, staging, risk stratification
• Caution is warranted when monitoring myeloma patients on mAb and response assessment of multiple myeloma. Leukemia 2009;23(1):3–9.
drugs, as these biologicals may cause false positive results by protein doi:10.1038/leu.2008.291. PubMed PMID: 18971951, PubMed Central
electrophoresis of the serum and immunofixation electrophoresis (IFE). PMCID: PMC2627786.
17. Rajkumar SV. POEMS syndrome. UpToDate: Wolters Kluwer; 2016.
18. Rajkumar SV. Epidemiology, pathogenesis, clinical manifestations and
Please check your eBook at https://expertconsult.inkling.com/ diagnosis of Waldenstrom macroglobulinemia. UpToDate: Wolters
Kluwer; 2016.
for self-assessment questions. See inside cover for registration 19. Owen RG, Treon SP, Al-Katib A, et al. Clinicopathological definition of
details. Waldenstrom’s macroglobulinemia: consensus panel recommendations
from the Second International Workshop on Waldenstrom’s
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sonc.2003.50082. PubMed PMID: 12720118.
1. Rajkumar SV. Diagnosis of monoclonal gammopathy of undetermined 20. Rajkumar SV. Treatment and prognosis of Waldenstrom
significance. UpToDate: Wolters Kluwer; 2016. macroglobulinemia. UpToDate: Wolters Kluwer; 2016.
2. Kyle RA, Therneau TM, Rajkumar SV, et al. Prevalence of monoclonal 21. Dimopoulos MA, Kastritis E, Owen RG, et al. Treatment
gammopathy of undetermined significance. N Engl J Med recommendations for patients with Waldenstrom macroglobulinemia
2006;354(13):1362–9. doi:10.1056/NEJMoa054494. PubMed PMID: (WM) and related disorders: IWWM-7 consensus. Blood
16571879. 2014;124(9):1404–11. doi:10.1182/blood-2014-03-565135. PubMed
3. International Myeloma Working Group. Criteria for the classification of PMID: 25027391, PubMed Central PMCID: PMC4148763.
monoclonal gammopathies, multiple myeloma and related disorders: a 22. Dhodapkar MV, Jacobson JL, Gertz MA, et al. Prognostic factors and
report of the International Myeloma Working Group. Br J Haematol response to fludarabine therapy in patients with Waldenstrom
2003;121(5):749–57. PubMed PMID: 12780789. macroglobulinemia: results of United States intergroup trial (Southwest
4. Rajkumar SV. Clinical course and management of monoclonal Oncology Group S9003). Blood 2001;98(1):41–8. PubMed PMID:
gammopathy of undetermined significance. UpToDate: Wolters Kluwer; 11418461.
2016. 23. Rajkumar SV. The heavy chain diseases. UpToDate: Wolters Kluwer; 2016.
5. Rajkumar SV, Kyle RA, Therneau TM, et al. Serum free light chain ratio is 24. Rajkumar SV. Clinical presentation, laboratory manifestations, and
an independent risk factor for progression in monoclonal gammopathy diagnosis of immunoglobulin light chain (AL) amyloidosis (primary
of undetermined significance. Blood 2005;106(3):812–17. doi:10.1182/ amyloidosis). UpToDate: Wolters Kluwer; 2016.
blood-2005-03-1038. PubMed PMID: 15855274, PubMed Central 25. Gertz MA. How to manage primary amyloidosis. Leukemia
PMCID: PMC1895159. 2012;26(2):191–8. doi:10.1038/leu.2011.219. PubMed PMID: 21869840.
6. Rajkumar SV. Clinical features, laboratory manifestations, and diagnosis 26. Rajkumar SV. Prognosis and treatment of immunoglobulin light chain
of multiple myeloma. UpToDate: Wolters Kluwer; 2016. (AL) amyloidosis and light and heavy chain deposition diseases.
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Working Group updated criteria for the diagnosis of multiple myeloma.

ChaPter 80 Monoclonal Gammopathies 1093.e1


MUL ti PL e -C h O i C e QU e S ti ONS

1. Which of the following is a “myeloma defining event“ according 3. Which one of these differentiates Waldenström macroglobu-
to the 2014 International Myeloma Working Group (IMWG) linemia (WM) from IgM MGUS?
guidelines? A. IgM M-protein ≥1 g/dL
A. One focal 4-mm lesion on magnetic resonance imaging B. ≥10% plasma cells in bone marrow
B. M-protein >0.5 g/dL C. Renal damage
C. Back pain D. Patient age <50
D. Clonal bone marrow plasma cells ≥60%
2. Which one of these is a risk factor for progression from
monoclonal gammopathy of undetermined significance
(MGUS) to multiple myeloma or a related malignancy?
A. Immunoglobulin G (IgG) MGUS
B. Abnormal serum free light chain ratio
C. M-protein ≥0.3 g/dL
D. Patient age >50

81






Concepts and Challenges in Organ

Transplantation: Rejection,

Immunosuppression, and Tolerance



Kathryn J. Wood, Sushma Shankar, Joanna Hester, Fadi Issa







The clinical era of transplantation began on December 23, 1954, rejection, whereas allografts (transplantation of tissues from
when Dr. Joseph Murray and colleagues performed the first genetically different individuals) usually evoke an aggressive
successful renal transplant on the genetically identical Herrick immune response as a result of the antigenic differences between
1
twins. Solid organ transplantation (SOT) has since transformed the donor and the recipient. Tissue transplanted between species,
the landscape of modern medicine, offering dramatic improve- or xenografts, are also possible but not yet in clinical use in cell
ments in patient survival and quality of life in many end-stage and organ transplantations because these usually trigger a more
diseases. The development of powerful immunosuppressive rapid and aggressive immune response compared with an allograft.
regimens and cutting-edge biological agents represents an elegant This section considers the processes involved in allorecognition
proof of concept, translating seminal work from the laboratory and graft destruction.
bench to the patient’s bedside. However, this ground-breaking
field within clinical immunology is not free of setbacks. Rejection Innate Immunity and Ischemia-Reperfusion Injury
of transplanted organs and tissues can result in devastating Trauma to the graft is initiated during, or even before, retrieval
problems for the patient, and potent immunosuppression is because death of the donor brainstem induces hemodynamic
associated with substantial comorbidity, including exposure to and neuroendocrine responses; in addition, organs obtained from
infection, malignancy, and cardiovascular risks that may be fatal. donors after their cardiac death suffer a period of warm ischemia.
Theoretically, immunosuppression withdrawal and allograft- During cold storage, loss of intracellular potassium and cell injury
specific protection against host responses is the ultimate treatment continues. This leads to graft damage through cell swelling
to offer a transplant recipient. Although this is undoubtedly a and a buildup of toxic metabolites, leaving the graft susceptible
challenging goal, it is being realized in defined subgroups of to further ischemia–reperfusion injury on rapid warming fol-
recipients. This chapter examines the rejection response, outlines lowing revascularization. Efforts are made during storage
the mainstay of current immunosuppressive therapy, and discusses and organ implantation surgery to reduce the metabolic rate
the latest advances in the search for transplantation tolerance. through storage on ice and perfusion with specialized fluids.
This approach aims to minimize the accumulation of toxic
REJECTION metabolites and the resultant pH changes to reduce the effect
of cold ischemic time on posttransplantation outcomes, such as
The immune system is complex and has evolved to protect the delayed graft function. 2
individual from harm. This harm may be either in the form of Tissue injury leads to the expression of damage-associated
foreign pathogenic microorganisms or premalignant mutations molecular patterns (DAMPs), such as heparin sulfate, heat shock
in the individual’s own cells. To achieve this safely, the immune proteins (HSPs), nucleic acids, and high-mobility group box-1
system must have the ability to distinguish “self” from “non-self” (HMGB1) protein. These are identified by invariant pattern-
or “altered-self” to avoid damaging the host itself. Any immune recognition receptors (PRRs) of the innate immune system, for
response that is generated must also be proportional to the threat, example, Toll-like receptors (TLRs) (Chapter 3). This process
and thus antigens encountered in the context of inflammation results in the local production of inflammatory mediators, such
will prime T cells and evoke a more aggressive immune response. as interleukin (IL)-1 and -6, chemokines, and the expression of
Although an effective immune response is essential to survival adhesion molecules within the graft. These early innate responses
in the context of infection or malignancy, it can represent sig- are seen in autografts as well as isografts and are generally not
nificant management challenges in the transplantation setting. alloantigen specific, although a subcategory of innate cells does
Some elements of the immune system respond to the general display a limited degree of alloantigen reactivity. In turn, this
trauma associated with organ retrieval, perfusion, and surgery, triggers activation of macrophages and dendritic cells (DCs) via
3
whereas others respond after specific recognition of antigenic cell-surface and internal PRRs, causing both cell types to show
differences between the donor and the recipient. If this process greater antigen-presenting capacity and to enter a cytocidal state.
continues in an uncontrolled manner, it inevitably leads to Endogenous signals, such as stress, can also activate the comple-
destruction of the graft. Thus isografts or syngeneic grafts (grafting ment cascade to generate several products, including complement
of tissue between two genetically identical individuals such that component C3 in the graft, which promotes DC maturation and
there are no antigenic differences) do not typically result in subsequently their ability to activate T cells. DCs are also activated

1097

1098 Part NiNe Transplantation



Trauma of transplantation antigens can also be significant, particularly in bone marrow
Organ retrieval, perfusion, and transplantation expression of transplantation (BMT), where activation of the adaptive immune
proinflammatory cytokines and recruitment of inflammatory cells into the graft system of the donor may result in graft-versus-host disease
(GvHD). These miH antigens may be derived from a wide variety
of polymorphic proteins and are encoded for a vast array of
Presentation of alloantigen to recipient T cells: genes distributed throughout the genome.
Direct, indirect, and semidirect presentation of alloantigen to recipient APC
Signal 1: Recognition of Alloantigen
Class I MHC molecules are cell surface glycoproteins expressed
T-cell activation:
TCR signal on most nucleated cells and are recognized by the T-cell receptors
Costimulation (TCRs) of CD8 T cells. Class II MHC molecules are not expressed
Cytokine generation by every cell in the body; rather they are found on DCs, B
lymphocytes, macrophages, and, in humans, endothelial cells.
MHC class II molecules are recognized by T cells bearing the
Cell-mediated immunity Th17 Humoral immunity CD4 surface glycoprotein. Expression of both class I and II MHC
Th1 Th2 molecules can be increased or, in the latter case, induced during
inflammation, in particular by IFN-γ.
Class I and II MHC proteins contain a highly polymorphic
groove or cleft, which can bind peptides, formed on a platform
of beta strands, flanked by two α helices. Antigen processing
T-cell migration: within antigen-presenting cells (APCs) results in the production
Upregulation of MHC and adhesion molecules attraction of peptides that can bind in these grooves producing a MHC–
of leucocytes into graft through chemotaxis peptide complex that can be recognized by T cells (Chapter 6).
These peptides may be of self origin or derived from foreign
molecules (e.g., from an allograft after transplantation or from
Graft destruction:
Infiltration of macrophages, cytokines, cytotoxic T cells, and a virus after an infection). In general, peptides derived from
antibodies leading to graft injury molecules present inside the cell are processed and loaded into
MHC class I molecules, whereas molecules present outside the
FiG 81.1 Mechanisms Leading to Graft Injury. Interplay of
innate and adaptive immune responses results in eventual graft cell in the extracellular environment are processed into peptides
destruction. that load into class II molecules. However, cross-presentation
can also occur, whereby some APCs, notably DCs, process
extracellular proteins such that they can be loaded onto MHC
class I molecules and presented to CD8 T cells, leading to
by interferon-γ (IFN-γ) produced by natural killer (NK) cells. “cross-priming.” 6
In one or more of these ways the innate immune system, activated The TCR is composed of two chains that confer MHC–peptide
by local tissue injury, promotes the initiation of adaptive immune specificity and is associated with a complex of polypeptides
responses when there are antigenic differences between the donor referred to collectively as CD3 (Chapter 4). On antigen recognition
and the recipient. Activation of the adaptive immune system by a TCR, CD3 delivers intracellular signals to the T cell (Chapter
results in a series of effector mechanisms, both cell and antibody 12). Recognition of antigen by the TCR–CD3 complex is the
mediated, that lead to further graft injury (Fig. 81.1). first step in T-cell activation and commonly referred to as “signal
A severely damaged graft will initiate a more aggressive 1.” This initial meeting between host naive and memory T cells
immune response, and this may explain the superior outcomes and alloantigen from the foreign transplant is believed to take
in living donor grafts compared with deceased donor grafts (as place in secondary lymphoid organs rather than in the transplanted
7
the former are subject to a lower degree of ischemic injury), graft itself. The inflammatory response triggered in the allograft
even in the presence of significant major histocompatibility by retrieval and implantation of the organ or tissue, as outlined
4
complex (MHC) mismatches. The immune response evoked above, not only initiates the migration of donor-derived passenger
by the damaged tissue also accounts for the higher rates of leukocytes but also their maturation into functional APCs
rejection observed in individuals with delayed graft function. 5 expressing high levels of donor MHC molecules.
The presentation of intact allogeneic MHC molecules by
Initiation of the Adaptive Immune System donor-derived passenger leukocytes to T cells is known as the
Recognition of differences between donor and recipient major direct pathway of allorecognition. This is the dominant pathway
and/or minor histocompatibility (miH) molecules or antigens through which the immune reaction to the graft is initiated, and
by T cells is central to the adaptive immune response. The T cells responding to direct antigen presentation constitute a
extensive polymorphism of MHC genes (Chapter 5) makes vast majority of the alloreactive immune repertoire, estimated
8
complete matching of unrelated individuals rare, and therefore, at 10% of T cells. After a time, the donor lymphocytes trans-
almost inevitably, transplantation of cells or organs between planted with the graft are depleted; however, this does not abrogate
genetically unrelated individuals occurs across an MHC mismatch. rejection. The immune response against an allograft is maintained
Knowledge of the MHC and of the structure of MHC molecules by recipient APCs, largely consisting of DCs and B cells, which
has contributed significantly to our understanding of how rejec- process and present peptides derived from allogeneic MHC
tion is triggered and has facilitated the development of novel molecules shed from the graft (soluble MHC molecules or
immunosuppressive drugs. Additionally, transplantation between apoptotic cells) as well as miH antigens. This route of allorecogni-
MHC-identical siblings has shown that differences in miH tion is known as the indirect pathway. Experimental models have

CHaPter 81 Concepts and Challenges in Organ Transplantation 1099


Recipient APC
Direct presentation Semidirect presentation

Donor APC
Donor APC
Class I MHC

CD8 +




Exosome +
Class II MHC Class II MHC CD4
C





CD4 +
A

Indirect presentation


Recipient APC



CD4 +

B
FiG 81.2 Antigen Presentation. (A) Direct presentation: passenger donor antigen-presenting
cells (APCs) present alloantigen to recipient T cells in lymphoid tissue. (B) Indirect presentation:
alloantigen from donor cells is processed and presented by recipient APCs via major histocompat-
ibility complex (MHC) class II to recipient CD4 cells. (C) Semidirect presentation: donor MHC
+
class I and class II may be transferred to the surface of recipient APCs enabling presentation of
alloantigen to recipient T cells.



suggested that indirect presentation of donor antigens may play APC T cell
a greater role in rejection than direct presentation overall, as it
9
continues for as long as the graft remains in situ. It is also now CD80 CTLA-4
known that the direct and indirect pathways of allorecognition
interact as a third pathway referred to as the semidirect pathway CD86 CD28
of antigen presentation. In the semidirect pathway of allorecogni-
tion, donor MHC proteins are transferred intact to recipient CD40 CD154
APCs (through membrane transfer or the exosomal route),
enabling them to present allogeneic MHC–peptide complexes
to recipient T cells. This MHC transfer is temperature and energy FiG 81.3 Costimulation. Costimulation follows binding to major
dependent and requires close cell-to-cell contact. Both MHC histocompatibility complex (MHC) class I and class II on antigen-
class I and class II may be transferred, although class II MHC presenting cells (APCs) and involves signal transduction via
10
appears to be transferred more efficiently. These three modes intracellular proteins and increased interaction affinity via the
of antigen presentation are illustrated in Fig. 81.2. binding of several cell surface proteins. Several costimulatory
molecule pairs have been identified.
Signal 2: Costimulation
T lymphocyte activation relies not only on contact of the membrane of the T cell reorganizes such that the TCR–CD3
TCR–CD3 complex with its specific MHC–peptide complex complex and costimulatory molecules are brought together
(“signal 1”) but also on signals delivered by the interaction of in the cell membrane to form the immunological synapse.
several costimulatory receptors and their ligands, known col- Signal 1 is specific for the antigen involved, whereas costimulation
lectively as “signal 2” (Fig. 81.3). During T-cell activation, the is not.

1100 Part NiNe Transplantation


Costimulatory receptors fall into two major families, the B7 The pattern of cytokine production through the interaction
family (e.g., CD28 and CD152) and the tumor necrosis factor of the combination of signaling processes determines the nature
(TNF) family (e.g., CD154 and CD70) (Chapter 12). Although of the response, in which either cell-mediated or antibody-
several costimulatory molecule pairs have been identified, the mediated immunity is seen to dominate (Chapter 16). The T
CD28 and CD154 pathways have been the most clearly defined cells known to promote a cell-mediated response, and the
to date. CD80 and CD86 are receptor ligands on the surface of cytokines they produce, are termed T-helper 1 (Th1) cells. These
DCs and other cells that can bind to CD28 on T cells, resulting cells are associated with the production of IFN-γ. T cells promot-
in the activation of additional signal–transduction pathways ing a humoral response are referred to as Th2 and are associated
within the T cell. This has a number of effects: lowering the with the generation of IL-4, -5, and -6. Additionally, a Th17
threshold for T-cell activation; increasing glucose metabolism, population that has been identified is characterized by the produc-
12
as well as cytokine and chemokine expression, including IL-2 tion of IL-17 and promoting the infiltration of neutrophils.
production; reducing T-cell death through apoptosis; and expand- Th22 cells, which express IL-13, IL-22, and TNF-α, have also
ing the number of T cells that respond through proliferation. been described. A subset of CD4 cells called regulatory T cells
CD80 and CD86 are also linked to an inhibitory receptor, CD152 (Tregs) can also be induced following antigen exposure in the
13
or cytotoxic T lymphocyte antigen-4 (CTLA-4), which inhibits periphery (pTregs; Chapter 18). These cells secrete IL-10 or
T-cell activation possibly by competing with CD28 to bind with transforming growth factor (TGF) and have suppressive or
CD80 or CD86. CTLA-4 has a 10-fold higher receptor affinity regulatory functions against effector cells and APCs. The balance
compared with CD28 and is rapidly upregulated following T-cell of these responses results in either graft injury or the induction
activation. Additionally, CD152 activates tryptophan catabolism of tolerance (discussed below).
in DCs, which results in the inhibition of proliferation and
promotes apoptosis of the responding T cells. Related to this Memory T Cells
pathway are other pathways involving members of the B7–CD28 Following exposure to an antigen, antigen-specific memory T
family of molecules, including ICOS/B7h, PD1/PD-L1/PD-L2, and B cells are generated. These memory cells are then able to
and B7-H3. Each of these can play a role in T-cell costimulation produce a more rapid and intense immune reaction if the antigen
and later coinhibition, but for the early phase of the response, is encountered on a second occasion because they have a lower
the CD28 pathway dominates. CD40 belongs to the TNF receptor activation threshold and are less dependent on costimulation.
family, expressed on all APCs, and binds to CD154 (CD40L), Transplant recipients, particularly older patients or those who
which is present on activated CD4 cells, a subset of CD8 cells have had previous antigen exposure through previous transplanta-
and NK cells. CD40 stimulation causes triggering signals for tion, blood transfusion, or pregnancy, may therefore have specific
antibody production and induces MHC expression on APCs, anti-donor memory cells. Memory-type responses may also occur
thus amplifying antigen presentation. 11 as a result of antigen receptor cross-reactivity known as heter-
In clinical transplantation, treatment with calcineurin inhibi- ologous immunity.
tors (CNIs) to block signal 1 and/or the use of T cell–depleting
agents has been highly successful in the prevention and reversal Cell Migration
of rejection episodes. More recently, blockade of costimulatory After activation, chemotactic cytokines, or chemokines, generated
pathways has also been shown to be effective. These approaches at the site of inflammation attract leukocytes that have been
are discussed in the “Immunosuppression” section. primed in secondary lymphoid tissues to the graft (Chapter 10).
Once activated, leukocytes upregulate chemokine receptors, thus
Signal 3: Proliferation and Differentiation of enabling their migration along a chemokine gradient. Vessels in
Effector T Cells proximity to the graft become dilated, allowing increased blood
Following alloantigen recognition by the TCR–CD3 complex flow, and the endothelium becomes activated. Activated endo-
and costimulation, a process begins resulting in “signal 3.” This thelial cells externalize Weibel-Palade bodies containing P-selectin,
is the complex process involving the recruitment and phosphoryla- and chemokines generated in the graft, such as IL-8, RANTES,
tion of a series of signaling molecules, which induce intracellular and MCP-1, stick to the vascular endothelium (Chapter 11).
biochemical processes that lead to the activation of three This signals the passing leukocytes in postcapillary venules to
signal–transduction pathways: the calcium–calcineurin pathway, leave laminar flow of blood and move closer to the vessel edge
the Ras- and Rac-mitogen–activated protein kinase pathways, so that they can interact with the activated endothelial cells.
and the protein kinase C nuclear factor κB (NF-κB) pathway. Extravasation then occurs in a multistep process, involving
These pathways interact with inositol triphosphate (IP3) and molecules on the surface of activated leukocytes called Sialyl-Lewis
diacylglycerol (DAG), formed from the hydrolysis of phospha- moieties, which interact with endothelial P-selectin, forming
tidylinositol 4,5-bisphosphonate, to activate transcription factors: temporary bonds that form and break causing the leukocytes to
nuclear factor of activated T cells (NFAT), activating protein-1 “roll.” Chemokines on the endothelial surface then induce a
(AP-1), and NF-κB, respectively. These transcription factors cause conformational change in leukocyte integrin molecules, allowing
the expression of many genes leading to upregulation of growth high-affinity binding to endothelial adhesion molecules, such
factors and cytokines, in particular, IL-2 and CD25 (IL-2Rα). as intercellular adhesion molecule-1 (ICAM-1). When this occurs,
Growth signals are then delivered through the phosphoinositide- the leukocytes stop rolling, and this allows them to extravasate
3-kinase (PI3K) and mechanistic target of rapamycin (mTOR) and follow the chemokine gradient further into the graft, where
pathways to promote cell cycle progression, and initiation of damage then occurs.
clonal expansion and differentiation of activated T cells to express
effector functions. These mechanisms are targeted by various The Effector Response and Graft Destruction
immunosuppressive agents discussed in the “Immunosuppression” The immune system generates many different effector mecha-
section and illustrated in Fig. 81.5. nisms, depending both on the challenge it meets and the

CHaPter 81 Concepts and Challenges in Organ Transplantation 1101


microenvironments that are present. In transplantation, the type mediators, such as TNF-α. Perforins insert into the target cell
of transplanted tissue, graft site, and immune status of the membrane to form pores, allowing granzyme to enter the cell,
recipient at time of transplantation may modify this response. and this causes proteolysis and activates the caspase cascade. Fas
Although initiation of rejection in a nonsensitized recipient is ligand binds to Fas on the target cell and also initiates the caspase
principally T-cell dependent, many components of the immune cascade. These processes induce target cell apoptosis and acute
system contribute to the subsequent destruction of the trans- cellular rejection and typically occur 1 week to 3 months after
planted tissue. Graft destruction may be alloantigen specific, or transplantation.
there may be bystander tissue destruction. The nature of the Additionally a nonspecific delayed-type hypersensitivity
immune cells involved in the effector response is reflected in the response occurs, usually mediated by CD4 cells that are attracted
characteristics of the resulting damage and in the speed of onset to the graft and involving the release of multiple proinflammatory
of tissue destruction. cytokines, including IL-1, IFN-γ, and TNF-α. This leads to the
recruitment and activation of further leukocytes, affects graft
Acute Antibody-Mediated Rejection cell permeability and vascular smooth muscle tone, thus affecting
Alloantigen-specific antibodies, or alloantibodies, are secreted graft physiology and contributing to acute and chronic rejection.
by plasma cells. Alloantibodies are produced after alloantigen- CD4 alloreactive T cells responding to donor-derived peptides
driven B-cell activation in the presence of T-cell help, such as bound to recipient MHC class II molecules have also been cor-
can occur during rejection or following a blood transfusion. related with chronic allograft dysfunction. 15
Antibodies that cross-react with alloantigens can also be generated As acute allograft rejection is initiated by the recognition of
as a result of infections that result in heterologous immunity. polymorphic donor MHC molecules by recipient T cells, it follows
In the former, in addition to DCs, the B cells themselves may that transplantation of MHC incompatible tissues will elicit a
act as APCs. MHC class II molecules are presented to and bind strong, T cell–dependent immune response to donor tissues.
immunoglobulins (Igs) on the surface of B cells, enabling the B In general, mismatches for only class I MHC result in slower
cell to internalize the alloantigen and process it into peptides rates of rejection compared with grafts with differences in
16
that are presented at the cell surface within MHC class II molecules class II MHC or both class I and II. Rejection can still
(Chapter 6). The subsequently activated T cells produce cytokines occur when transplantation has taken place between MHC-
that activate B cells enabling them to differentiate into matched siblings because of T-cell recognition of minor histo-
alloantibody-producing plasma cells. When allospecific antibodies compatibility antigens.
encounter their specific antigen, antibody-mediated rejection
occurs. Antibodies function through a number of effector Delayed Allograft Rejection and Dysfunction
mechanisms. Antibody binding activates endothelial cells within Rejection later after transplantation, which is often referred to
the graft, resulting in the expression of adhesion molecules, as chronic rejection or chronic allograft dysfunction, comprises
cytokines, and chemokines as well as the synthesis of tissue factor. a number of mechanisms. It is characterized by a cellular infiltrate
Antibody binding can trigger complement activation, which can constituted of macrophages, eosinophils, NK cells, and T cells,
17
result in cell lysis and graft damage directly or indirectly by the where the primary target is the vascular endothelium. Damaged
binding of complement components to the graft and the recruit- endothelium allows platelet deposition on the arterial wall and
ment of macrophages and neutrophils. Additionally damage is growth factor production. This, in turn, leads to smooth muscle
caused by a mechanism whereby effector cells, such as NK cells proliferation in the arterial wall media. Proliferating muscle cells
and macrophages, bind to the Fc non–antigen-specific portion invade the intima and contribute to intimal fibrosis. These changes
of the antibody. This encourages NK cells and macrophages to are concentric, affecting all graft arteries. In heart transplants,
kill any target cells with antibody bound to their surface. This this may appear as transplant arteriosclerosis. Chronic rejection
is a nonspecific process termed antibody-dependent cellular therefore manifests in an organ-specific manner. For example,
cytotoxicity (ADCC) and can contribute to graft rejection. 14 within renal glomeruli glomerulosclerosis and interstitial fibrosis
When donor-specific alloantibodies (DSAs) exist in a recipient may develop, whereas in lungs, bronchiolitis obliterans develops.
prior to transplantation (e.g., as a result of previous transplanta- The underlying mechanistic processes remain similar.
tion, blood transfusion, or pregnancy), a dramatic response is
seen upon perfusion of the transplanted graft. Coagulation and Clinical Implications
complement cascades are activated, resulting in extensive To prevent hyperacute rejection, patients on transplant waiting
thrombosis and graft infarction within minutes. This so-called lists are monitored routinely for the development of anti–human
hyperacute rejection is now very rarely seen in clinical practice leukocyte antigen (HLA) antibodies as a marker of sensitization.
because of the advances in screening and cross-matching tech- Immediately before transplantation, a further check is performed
niques as well as desensitization techniques, such as plasmapheresis by mixing recipient serum and donor splenocytes and observing
18
and intravenous immunoglobulin (IVIG). for cell lysis. The extent of the risk of hyperacute rejection is
determined by the antibody target and quantitative titer of the
Acute Cellular Rejection anti-donor antibodies. Highly sensitized recipients can remain
19
As described above, following the non–antigen-specific innate on waiting list for extended periods but may have successful
response to organ retrieval and implantation, an inflammatory transplantations after desensitization, as mentioned previously,
environment is created within the graft, thus promoting an by removal of existing antibodies through plasma exchange or
adaptive cellular response over the ensuing days. Naïve cytotoxic adsorption or by depletion of B cells with immunosuppressive
T cells, activated by CD4 cells clustering with APCs, migrate to agents, such as rituximab or proteasome inhibitors. Despite the
the graft, where they recognize allogeneic class I MHC molecules. extensive screening performed to detect donor HLA antibodies
This causes them to release cytotoxic molecules, such as perforin before transplantation, preformed antibodies against donor
and granzyme B; upregulate surface Fas ligand; and secrete soluble non-MHC antigens may exist and have the ability to induce

1102 Part NiNe Transplantation



1. Normal

2. Antibody-mediated rejection
Acute antibody-mediated rejection
I. ATN-like – C4d+, minimal inflammation
II. Capillary - margination and/or thromboses, C4d+
III. Arterial – v3, C4d+
Chronic active antibody-mediated rejection
Glomerular double contours and/or peritubular capillary basement membrane multilayering
and/or interstitial fibrosis/tubular atrophy and/or fibrous intimal thickening in arteries, C4d

3. Borderline changes: ‘suspicious’ for acute T-cell-mediated rejection
This category is used when no intimal arteritis is present, but there are foci of tubulitis

4. T-cell-mediated rejection
Acute T-cell-mediated rejection
IA. Cases with significant interstitial infiltration (>25% of parenchyma affected, i2 or i3)
and foci of moderate tubulitis (t2)
IB. Cases with significant interstitial infiltration (>25% of parenchyma affected, i2 or i3)
and foci of severe tubulitis (t3)
IIA. Cases with mild to moderate intimal arteritis (v1)
IIB. Cases with severe intimal arteritis comprising >25% of the luminal area (v2)
III. Cases with ‘transmural’ arteritis and/or arterial fibrinoid change and necrosis of medial
smooth muscle cells with accompanying lymphocytic inflammation (v3)

Chronic active T-cell-mediated rejection
‘Chronic allograft arteriopathy’ (arterial intimal fibrosis with mononuclear cell infiltration in
fibrosis, formation of neo-intima)


5. Interstitial fibrosis and tubular atrophy, no evidence of any specific etiology
I. Mild interstitial fibrosis and tubular atrophy (<25% of cortical area)
II. Moderate interstitial fibrosis and tubular atrophy (26–50% of cortical area)
III. Severe interstitial fibrosis and tubular atrophy/loss (>50% of cortical area)
(may include nonspecific vascular and glomerular sclerosis, but severity
graded by tubulointerstitial features)

6. Other: Changes not considered to be due to rejection-acute and/or chronic

FiG 81.4 Banff Diagnostic Criteria for Renal Allografts. Histological criteria have been developed
and are regularly reviewed to aid diagnosis of the cause of chronic allograft dysfunction. Determining
the cause of dysfunction aids decision making with regard to pathology and management. C4d,
complement degradation product C4d.




vascular rejection. There is increasing interest in determining complement fixation in not only late rejection but also early
the impact of such antibodies on graft outcome. rejection. Antibody-mediated rejection (ABMR) is frequently
Acute graft rejection is suspected when there is a sudden seen following heart, lung, and kidney transplantations; however,
20
deterioration in allograft function. Biopsy of the transplanted liver allografts are relatively protected. Nevertheless ABMR is
tissue and histological evaluation are performed, resulting increasingly recognized as a cause of graft damage in liver
in diagnosis of rejection and numerical grading of severity. transplantation. This is explored further in the discussion below
Grading schemes, such as the Banff criteria shown in Fig. 81.4, on tolerance in the liver (Fig. 81.6).
which provide semiquantitative measures for histopathological Although improved immunosuppression regimens have led
assessment of the inflammatory response, have been devel- to a reduction in the occurrence of acute rejection, chronic
oped for specific organs and form the basis on which further rejection has become more evident and remains a significant
clinical management is guided; decisions to treat with high-dose contributor to late graft loss. Nonimmunological factors, including
steroids or other immunosuppressive medications can thus CNI-induced toxicity, advanced donor age, ischemic injury during
be made after structured analysis of a biopsy specimen of the implantation, hypertension, and infection, also contribute to
transplant. chronic allograft dysfunction, and modifying these factors can
21
In addition to the diagnosis of cell-mediated rejection, modern improve graft outcome. However, immunological processes
staining techniques have enabled the identification of complement play a significant role with increased levels of pretransplantation
component 4d (C4d) in biopsy specimens from rejecting tissues, anti-HLA antibodies, de novo posttransplantation donor-specific
thus providing indirect evidence of antibody deposition and antibodies and antibodies against non-HLA antigen MHC class

CHaPter 81 Concepts and Challenges in Organ Transplantation 1103



APC


Rituximab
MHC:peptide
Signal 1 Basiliximab
Belatacept
IL-2

Signal 2
CD28 TCR:CD3 Anti-CD3 IL-2 receptor
mAB
Ca 2+
Cyclosporine Signal 3
Tacrolimus Rapamycin
rATG Calcineurin mTOR

CD52 Cyclin/CDK
NF-AT
Steriods Cell Mycophenolate
IL-2 transcription cycle Azathioprine
Alemtuzumab


Immunodepletion
FiG 81.5 Immunosuppressive Drugs and Their Targets. Signal 1 results from major histocompat-
ibility complex (MHC): antigen recognition through the T-cell receptor (TCR)–CD3 complex, a
process blocked by anti-CD3 mAbs and indirectly by rituximab. Signal 2 results in costimulation,
a process which can be blocked by belatacept. Costimulation results in full activation of the
TCR–CD3 complex, initiating signal transduction, Signal 3: Downstream signaling pathways result
in calcineurin activation, a stage which can be inhibited by tacrolimus and cyclosporine. Activated
calcineurin dephosphorylates nuclear factor of activated T cells (NFAT), allowing interleukin-2
(IL-2) transcription, a process that can be inhibited by steroids. IL-2 receptor stimulation, a step
which can be blocked by basiliximab, activates the mechanistic target of rapamycin (mTOR) signaling
cascade, which can be inhibited by sirolimus. This pathway induces the T cell to enter cell cycle
and proliferate, which, in turn, can be blocked by mycophenolate and azathioprine. Rabbit anti-
thymocyte globulin (rATG) exerts polyclonal effects, and alemtuzumab binds to CD52, both
resulting in immunodepletion,




I polypeptide-related sequence A (MICA) being seen to affect Clinical regimens implemented today are focused on target-
allograft survival. ing the steps involved in the T cell–mediated immune response
The correlation among late acute rejection, chronic allograft to alloantigen. As outlined earlier, there are three main stages
dysfunction, and graft loss has been consistently reported, and in this pathway: recognition of alloantigen or self antigen;
there is increasing evidence that subclinical rejection may represent costimulation; and proliferation/differentiation of effector T
22
an important factor in predicting graft loss. Establishing a clear cells. The current clinical paradigm is based on blockade of
link between subclinical rejection and development of chronic at least one of these stages and/or by total immunodepletion
allograft dysfunction may be difficult, however, because of the therapy as well as by alterations in lymphocyte trafficking (see
interplay of other factors, including CNI toxicity. 21 Fig. 81.5).
The risk of acute graft rejection is greatest during the initial
IMMUNOSUPPRESSION three months following transplantation; therefore current
immunosuppression strategies are primarily based on a potent
In recent years, substantial advances in immunosuppressive induction regimen using a monoclonal antibody (mAb) or poly-
strategies and their translation to routine clinical practice have clonal antibody, followed by “maintenance immunosuppression”
revolutionized management and outcomes in solid organ often consisting of calcineurin inhibitors (CNIs, cyclosporine or
transplantation (SOT)—an option that has become the therapy tacrolimus), an antiproliferative agent (mycophenolate mofetil,
of choice for many end-stage organ diseases. Short-term outcomes, MMF), and low-dose corticosteroids (prednisolone). Provided
such as patient and allograft survival at 1 year, acute rejection there are no episodes of acute rejection, the doses of these agents
rates, and time-course of disease progression and symptom are gradually reduced, and then maintenance immunosuppres-
control have steadily improved since the first successful trans- sion is continued indefinitely (Fig. 81.6). This dose reduction is
plantations performed over 50 years ago. based on the concept that as graft inflammation subsides and

1104 Part NiNe Transplantation



Living donor kidney Simultaneous pancreas/kidney
Heart-beating donor kidney
Non-heart-beating donor kidney
HLA mismatch £ 2 HLA mismatch ³ 2
Alemtuzumab induction on day 0+1
Maintenance with daily mycophenolate and tacrolimus
Low intensity protocol High intensity protocol B No steroids


Basiliximab induction on day 1 +4 Basiliximab induction on day 1 +4
Maintenance with daily Maintenance with daily
azathioprine and tacrolimus mycophenolate and tacrolimus
A reducing dose of steriods over 2 months reducing dose of steriods over 2 months
FiG 81.6 Examples of Immunosuppressive Protocols in Clinical Use. (A) Patients receiving
a renal transplant from a living donor or heart-beating donor are stratified according to human
leukocyte antigen (HLA) mismatch. When HLA mismatch ≤2, the patient is treated with a low
intensity protocol; when HLA mismatch ≥2, the patient is treated with a high intensity protocol.
(B) Pancreas and non–heart-beating donor kidneys have higher immunogenicity, and so induction
is with a potent monoclonal antibody, alemtuzumab; therefore steroids are not required.


TABLE 81.1 immunosuppressive therapies in transplantation: Maintenance and
induction agents
Drugs Mechanisms adverse effects
Maintenance agents
Azathioprine Inhibits purine and DNA synthesis, inhibits cell proliferation Bone marrow depression, opportunistic infection,
macrocytosis, liver toxicity
Cyclosporine Binds to cyclophilin, inhibits calcineurin-phosphatase, blocks Hypertension, hyperlipidemia, nephrotoxicity, hepatotoxicity,
NFAT dephosphorylation, blocks IL-2 transcription and & T-cell pancreatitis, peptic ulcers, thrombotic microangiopathy,
activation opportunistic infection, neurotoxicity, tremor, gingival
hyperplasia, hirsutism
Mycophenolate Inhibits inosine-monophosphate-dehydrogenase, inhibits purine Gastrointestinal symptoms, bone marrow depression,
mofetil synthesis and blocks cell proliferation opportunistic infection in particular CMV and BK nephropathy
Rapamycin Binds to FKBP12, inhibits mTOR and blocks IL-2–driven cell Delayed graft function, delayed wound healing, mouth ulcers,
proliferation pneumonitis, increased proteinuria, peripheral edema,
hyperlipidemia
Steroids Induces phospholipase A 2 inhibitory proteins, inhibits arachidonic Diabetes, delayed wound healing, peptic ulcers, psychosis,
acid synthesis, inhibits prostaglandins and leukotrienes osteoporosis, infection, blurred vision, fluid retention, weight
gain, acne, constipation
Tacrolimus Binds to FKBP12, inhibits calcineurin-phosphatase and blocks Posttransplantation diabetes mellitus, nephrotoxicity,
T-cell activation thrombotic microangiopathy, neurotoxicity
induction agents
Antithymocyte Polyclonal effects not well characterized; immunosuppressive Polyclonal effects: cytokine release syndrome, serum
globulin efficacy attributed to T-cell depletion through apoptosis, sickness, leukopenia, thrombocytopenia. De novo tumors
antibody-dependent cytolysis and complement-dependent lysis and opportunistic infection: CMV and HSV.
Alemtuzumab Binds to CD52 antigen, (expressed on 95% of peripheral blood Opportunistic infection: disseminated Candida and CMV
lymphocytes, NK cells, macrophages, and thymocytes) infection
Results in profound lymphopenia
Basiliximab Binds to IL-2R with similar affinity as IL-2, thereby inhibiting Occasional hypersensitivity reactions, inadequate
IL-2-driven T-cell proliferation immunosuppression in immunologically high-risk recipients
IL-2, interleukin-2; IL-2R, interleukin-2 receptor; NFAT, nuclear factor of activated T cells; DHFR, dihydrofolate reductase; mTOR, mechanistic target of rapamycin;
CMV, cytomegalovirus; HSV, herpes simplex virus.


donor-derived APCs are cleared, the immunogenicity and risk An introduction to the immunosuppressive agents currently in
of rejection decreases. widespread use is provided below.
Chronic immunosuppression is associated with several
undesirable sequelae (Table 81.1), in particular an increased Immunodepletion
relative risk of infections and malignancy, cardiovascular morbid-
ity (the leading cause of death with a functioning graft in The more recent therapeutic strategies are based on induction
transplant recipients), and target organ damage. therapies that concentrate on profound immune cell depletion
Many different immunosuppressive regimens for SOT are in at the time of transplantation, when immune activation is most
clinical use and are dependent on local factors and guidelines. intense.

CHaPter 81 Concepts and Challenges in Organ Transplantation 1105



CLiNiCaL PearLS powered to show overall superiority of one agent over the other
Treatment Options for Rejection and were powered to address safety rather than efficacy parameters
24
or used a limited daclizumab dosage regimen. More robust
• Acute cellular rejection studies that are designed to evaluate the efficacy of rATG relative
• High-dose steroids to IL-2R mAbs specifically, are needed to definitively establish
• Rejection refractory to steroid treatment the use of rATG with respect to these monoclonal agents.
• Antithymocyte globulin
• Alemtuzumab Anti-CD52 Monoclonal Antibody (Alemtuzumab)
• Acute humoral rejection Anti-CD52 mAb (alemtuzumab) is a humanized rat IgG2b
• Plasmapheresis
• Intravenous immunoglobulin directed against the CD52 antigen, which is expressed on 95%
• Rituximab of peripheral blood lymphocytes, NK cells, macrophages, and
27
thymocytes ; thus almost all mononuclear cells are affected.
The profound and long-lasting lymphopenia produced after the
administration of one or two doses of alemtuzumab is likely
Antithymocyte Globulin explained by such abundance on monocyte cell surfaces. Examina-
Rabbit-derived antithymocyte globulin (rATG) is a lymphocyte- tion of the peripheral blood lymphocytes from recipients after
depleting polyclonal IgG preparation with specificity toward alemtuzumab induction has identified a subset of T cells, pre-
human thymocytes. It binds primarily to the peripheral blood dominantly CD4 central memory cells that survive despite
lymphocytes as well as to those present in the lymphoid organs, alemtuzumab induction and appear largely resistant to depletion;
including lymph nodes, spleen, and thymus, as demonstrated these memory T cells express lower CD52 levels compared with
by in vivo studies in monkeys. The agent’s polyclonal nature naïve T cells. 28,29 CD52 is not present on granulocytes, platelets,
enables it to display specificity toward a wide variety of molecules erythrocytes, or hematopoietic stem cells (HSCs).
expressed on the surface of T cells, B cells, DCs, NK cells, and After binding to CD52, alemtuzumab causes cell death through
endothelial cells, including those involved in T-cell activation, several mechanisms: complement-mediated cytolysis, antibody-
proliferation, apoptosis, signal transduction, cell adhesion, and mediated cytotoxicity, and apoptosis. The plasma elimination
trafficking. half-life is approximately 12 days, whereas its clinical effects are
The precise mechanism of action underlying the immunosup- far more persistent. Lymphocyte depletion of >99% can be seen
pressive efficacy of rATG in SOT recipients is unknown at present, after a single dose, with lymph node depletion taking up to 3–5
although has been primarily attributed to T-cell depletion. Data days compared with <1 hour seen in the peripheral lymphocytes.
from in vitro studies have suggested that rATG modulates the Different subpopulations display varying rates of recovery,
expression of various lymphocyte surface antigens resulting depending on the subpopulation of interest: NK cells are almost
−
in apoptosis, antibody-dependent cytolysis, or complement- unaffected and decrease only transiently (a population of CD52
dependent lysis. Lymphocyte depletion with rATG has been NK cells has also been identified); monocyte and B-cell recovery
further demonstrated in adult recipients of renal transplants in can be seen at 3 and 12 months, respectively; T cells levels recover
several randomized, comparative clinical studies, with repopula- to only 50% of baseline at 36 months. 30
23
tion reported to take at least 3 months. More recently, data Alemtuzumab is currently being used for induction in SOT
from preclinical and clinical studies have suggested that rATG with the aim of minimizing maintenance immunosuppression.
therapy may induce the expansion and enrichment of certain It was first used in transplantation as an induction agent in
+
+
+
+
31
Treg subsets, such as CD4 CD25 Forkhead box P3 (FOXP3 ), 1998, in 13 renal transplant recipients who received low-dose
+
+
CTLA-4 , and glucocorticoid-induced TNF receptor (GITR+) cyclosporine alone as maintenance therapy. At 6–12 months
Tregs from human peripheral blood lymphocytes or peripheral follow-up, patient and graft survival were 100%, and there were
24
blood mononuclear cells in vitro ; these Treg subsets are key to two episodes of acute rejection. The 5-year results of the initial
32
immune response modulation and are further discussed in the series, together with another 20 patients who were subsequently
“Tolerance” section. enrolled, showed no significant difference in graft or patient
In terms of clinical use, rATG induction in combination with survival or in acute rejection rates compared with a retrospective,
tacrolimus-based immunosuppressive therapy is more effective contemporaneous control group of 66 renal transplant recipients
in preventing episodes of acute renal graft rejection than with who had received no induction, but triple immunosuppression
tacrolimus-based therapy without induction, as reported by therapy alone (cyclosporine, azathioprine, and prednisolone). The
primary endpoint data from two, randomized, open-label, study did also find, however, more episodes of late rejection in
multicenter trials. 25,26 Moreover, the median time to biopsy-proven the alemtuzumab-treated group. Indeed, severe lymphopenia and
acute rejection (BPAR) was >1 week longer in rATG induction homeostatic cytokines are known to drive the rapid homeostatic
than in noninduction; however, it should be noted that there proliferation of naïve and memory T cells, and lymphocytes
was no significant difference between induction and noninduction generated under such conditions have been previously found
regimens in terms of patient or graft survival. to be potent alloreactive cells, inevitably triggering rejection in
33
Several randomized, multicenter studies have reported that animal models. Investigation of this phenomenon in human
28
the efficacy of rATG induction therapy is generally no different recipients illustrated that the recovery of the immune system
from that of basiliximab or low-dose daclizumab (IL-2 receptor after alemtuzumab induction differed with respect to CD4 and
[IL-2R] mAbs, which are an alternative induction agent; discussed CD8 T cells. CD8 T cells recovered to pretransplantation levels
later in this section) with regard to the incidence of BPAR, graft by 6 months after alemtuzumab induction, whereas the number
loss, or death at 6 and 12 months after transplantation, in the of CD4 T cells remained low, even at 1-year after transplantation.
context of renal transplant recipients receiving triple immunosup- Repopulating CD8 T cells were mainly of the immunosenescent
−
+
pressive maintenance therapy. These trials, however, were not CD28 CD8 phenotype, and the percentage of this population

1106 Part NiNe Transplantation


was lower in those recipients who experienced rejection compared episodes as long ago as 1980. However, severe adverse effects of
with those who did not mount an immune response to their muromonab-CD3, such as cytokine-release syndrome (owing
allograft. A series of in vitro experiments demonstrated that the to the propensity of muromonab-CD3 to initially activate T
−
+
expanded CD28 CD8 T cells compete for immune space with cells, releasing TNF-α, and IL-2), resulted in preference for rATG
CD4 T cells, suppressing their proliferation and thus delaying and anti-CD52 agents, as outlined above. Furthermore, OKT3,
CD4 T-cell recovery. This delay might be associated with the an mAb of mouse origin, was limited by host production of
clinical outcome because CD4 T cells, notably CD4 T effector human antimouse antibodies (HAMAs) that bind to circulating
memory cells, have been shown to be associated with rejection. 34 reagent molecules, neutralizing their activity. Nonactivating,
Despite considerable experience with this agent over many therefore less toxic, humanized Fc-receptor nonbinding anti-CD3
years, largely in renal transplantation, until recently few random- mAbs, such as teplizumab and visilizumab, have been tested in
ized controlled trials had been conducted. Two randomized clinical trials in autoimmune disease settings (type 1 diabetes,
controlled clinical trials, the INTAC and 3C studies, investigating Crohn disease) as well as in renal and pancreatic islet transplanta-
41
alemtuzumab induction in renal transplantation have now tion. Indeed, these studies have suggested a further mechanism
provided further insight. 35,36 The data from these studies support of action with regard to anti-CD3 antibodies, and this could be
the use of alemtuzumab as an induction agent in low-risk patients, helpful in the setting of transplantation: Circulating CD8 T cells
and the incidence of biopsy-confirmed acute rejection at 1 year isolated from patients with type I diabetes, after treatment with
was lower in patients treated with alemtuzumab compared with anti-CD3 mAb, may have regulatory function ex vivo; recent
those treated with basiliximab induction. Similar to ATG, data have demonstrated functional human CD8 Tregs in vivo
42
alemtuzumab cannot be used as a solitary immunosuppressive after administration of anti-CD3 mAb. Further work is needed
therapy and requires long-term maintenance immunosuppression to elucidate the action of such agents on the immune response
to prevent allograft rejection. Importantly, in the 3C study, to both self and alloantigens to realize their full potential.
alemtuzumab induction therapy was found to result in reduced
CNI and mycophenolate exposure and steroid avoidance, reducing Anti-CD20 Monoclonal Antibody (Rituximab)
the risk of biopsy-proven acute rejection in a broad range of Rituximab is an anti-CD20, chimeric mAb that eliminates most
patients receiving a kidney transplant, and this supported earlier B cells from the circulation. Originally approved to treat B-cell
data from smaller studies, including a study in lung transplant lymphoproliferative diseases in patients other than transplant
37
recipients. However, patients with preformed DSAs despite a recipients as well as to treat posttransplantation lympho-
negative cross-match are at high risk of adverse outcomes when proliferative disease (PTLD), it is also now used in SOT as a
receiving a minimal immunosuppressive regimen incorporating treatment of antibody-mediated rejection and to desensitize
alemtuzumab induction; these patients may thus benefit from patients who are receiving ABO-incompatible transplants or
augmented immunosuppression. 38 retransplants. However, depletion of antibody-producing cells
+
Review of rejection episodes that have occurred with alem- may be incomplete because rituximab cannot target CD20 plasma
tuzumab induction, particularly in the absence of CNIs, has cells. It has been hypothesized that the action of rituximab in
highlighted some interesting findings. Several authors have allograft rejection is caused by not only depletion of plasma cells
reported that a number of these rejections demonstrate positive but also the effective depletion of APCs, which thus limits indirect
staining for the complement component C4d, indicative of acute pathway T-cell activation and a sustained immune response.
ABMR. In contrast, in acute rejection, examination of biopsy Clinical administration is in conjunction with maintenance
specimens taken from patients who had received both alemtu- immunosuppressive agents, plasmapheresis, intravenous immu-
zumab induction and tacrolimus revealed none that was humoral noglobulin (IVIG), and even splenectomy in some protocols. 43
27
in origin. These data suggest that tacrolimus may prevent this Recent work with nonhuman primate (NHP) models has
early ABMR. Immune profiles of renal transplant recipients shown that preemptive B-cell depletion with rituximab prevents
receiving alemtuzumab induction, 60 days of a CNI, and sub- both acute cellular rejection and antibody-mediated rejection
sequent sirolimus monotherapy displayed a general increase in of cardiac allografts; alloantibody was prevented from developing
39
the naïve B-cell population. Bloom et al. subsequently showed in blood, and complement-mediated graft injury was avoided.
that B cell–activating factor (BAFF), a B-cell survival cytokine Not only was the B-cell response inhibited, but acute cellular
influencing the threshold of B-cell activation, substantially rejection was also reduced by the combination of cyclosporine
increased in renal transplantation following treatment with and rituximab, suggesting that B cells may play a role in acute
40
alemtuzumab. This observation may help explain and prevalent cellular rejection as well. There may be a role for targeting B
the development of alloantibody in patients treated with depleting cells preemptively in SOT, and additional strategies to down-
antibody therapy at the time of SOT. regulate the B-cell response are under evaluation. Promising
targets include BAFF and its receptor, as well as costimulatory
Signal 1: Blockade of Antigen Recognition molecules, such as CD28, CD154, and CD40. 44
Activation of the rejection response to an allograft hinges on
the recognition of antigen by the host immune system. Targeting Signal 2: Blockade of Costimulation
signal 1 through the use of mAbs has been used in both trans- In the absence of appropriate costimulation, partially activated
plantation and autoimmunity. T cells either become hyporesponsive to subsequent antigen-
specific TCR signals (donor-specific anergy) or die by apoptosis.
Anti-CD3 Monoclonal Antibody It has been hypothesized that the inhibition of full T-cell activation
Muromonab-CD3 (OKT3), a mouse mAb binding to the CD3 by costimulatory blockade rather than total T-cell depletion
component of the TCR signal–transduction complex, was used might more selectively target effector T cells and spare beneficial
successfully as an induction agent for high-risk patients or Tregs while avoiding the many adverse effects of nonspecific
for the treatment of corticosteroid-resistant acute rejection immunodepletion.

CHaPter 81 Concepts and Challenges in Organ Transplantation 1107


days and so may be difficult to appropriately titrate in transplant
CD28:B7 (CD80/CD86) Blockade recipients suffering from infections.
CTLA-4 (CD152) is an inducible, T-cell surface antigen that
when bound to CD80/86 receptor:ligand (B7 molecules) on APCs, Signal 3: Blockade of Proliferation/Differentiation
delivers inhibitory signals to the activated T cell. Belatacept Activated T cells produce IL-2, which, in turn, binds to IL-2Rs,
(LEA29Y) is a fusion protein that combines a mutated version which are expressed only on the surface of activated cells and
of the extracellular binding domain of CTLA-4 with the Fc portion are not present on resting T cells. IL-2R is composed of three
of IgG1, with specificity for CD80/86 expressed on APCs. Ligation high-affinity transmembrane protein subunits: α (CD25), β
of CD80/86 by CD28 (a surface antigen constitutively expressed (CD122), and γ (CD132) subunits, which are covalently linked.
on T cells) usually lowers the activation threshold of T cells. The α subunit is specific to IL-2R only, and it is the binding of
Belatacept has a higher affinity and slower dissociation rate from α and β subunits that is crucial to the IL-2 signal transduction
human B7 molecules (i.e., CD80/86) compared with CD28, and T-cell activation that subsequently leads to proliferation
resulting in inhibition of the costimulation required for effective and clonal expansion of T and B cells specific to alloantigen or
T-cell activation. 43 self antigen. These cells are also stimulated to release more IL-2,
The complexities of the human immune system present sig- further magnifying the immune response.
nificant challenges to the translation of such agents into clinical
+
practice: In vivo work indicates memory and cytotoxic CD8 T cells Anti-IL-2R Monoclonal Antibody
have different costimulation requirements for complete activation (Basiliximab and Daclizumab)
+
compared with CD4 naïve T cells and may therefore be more Anti-IL-2R (anti-CD25) mAbs specifically target activated T cells
resistant to some tolerance induction strategies. Such blockade but do not cause significant lymphocyte depletion and are not
may also affect the function of Tregs as the CD28:B7 pathway associated with major adverse effects compared with lymphocyte-
has been shown to be important for their expansion and survival. depleting agents. However, other T-cell subtypes, including Tregs,
However, reassuringly, data from clinical trials of belatacept in also express CD25, and therefore the use of these agents may
renal transplantation, albeit with small patient numbers, suggest impact some of the natural mechanisms of immunoregulation.
that costimulation blockade does not interfere with Treg homeo- Basiliximab (chimeric form) and daclizumab (humanized form)
stasis. 45,46 In non-human primate models, CTLA-4 Ig prolongs have been used in renal transplantation in low-risk recipients,
pancreatic islet survival and, in combination with CD154:CD40 as defined as first allografts with living-related donors. 50,51
pathway blockade, induces indefinite acceptance of renal and heart Basiliximab is currently the agent of choice and binds to IL-2R
allografts. with similar affinity as IL-2, thereby effectively competing with
BENEFIT (Belatacept Evaluation of Nephroprotection and IL-2 and subsequently inhibiting IL-2–driven T-cell proliferation.
Efficacy as First-line Immunosuppression Trial), a 3-year, phase Basiliximab has a high volume of distribution, almost completely
III clinical trial, randomized patients to three groups: cyclosporine, saturating IL-2R on the peripheral lymphocytes within 24 hours
less-intensive belatacept, and more-intensive belatacept. Patients of a single dose of 2.5–25 mg in renal transplant recipients. The
received induction with basiliximab and were maintained on half-life in adults is approximately 13.4 days, and IL-2R saturation
mycophenolate mofetil (MMF) and corticosteroids. Patient and and suppression can last for 4–6 weeks.
graft survival rates were similar across the three groups at both Two meta-analyses evaluating the efficacy and safety of
1 and 2 years after transplantation. At the end of 1 year, although basiliximab in renal transplant recipients have been published. 51,52
the incidence of acute rejection was greater for more-intensive Both studies showed that basiliximab was more effective than
and less-intensive belatacept compared with cyclosporine, no placebo in reducing acute cellular rejection 6 months after
apparent impact on graft survival was demonstrated. At the end transplantation. However, both meta-analyses illustrated no
of 2 years, however, glomerular filtration rates (GFRs) continued significant differences in patient or graft survival rates between
to be significantly higher (15–17 mL/min) in the belatacept- basiliximab and placebo groups at 1 year after transplantation.
treated patients. Encouragingly, belatacept-treated patients also However, in studies involving high-risk transplant recipients (i.e.,
had sustained benefits in their cardiovascular and metabolic risk high HLA-mismatch) the risk of rejection may be higher with
profiles compared with those treated with cyclosporine, supporting IL-2R mAb in comparison with rATG. 53
the potential role of belatacept as a nonnephrotoxic alternative The highly selective and short-term immunosuppressive effects
to CNIs. 47,48 Patients enrolled in this trial have now been followed of basiliximab, which are confined to the highly immunogenic
up for 7 years, and the long-term data show that patient and period immediately after transplantation, make this class of drug
graft survival and the mean eGFR are significantly higher with a useful substitute to steroids in early steroid-withdrawal or
belatacept (both the more-intensive regimen and the less-intensive steroid-free regimens, as illustrated in studies with liver trans-
regimen) than with cyclosporine. 49 plantation. Several prospective clinical trials using basiliximab
Because of concerns about an increased post-transplant induction to facilitate early steroid withdrawal or complete steroid
lymphoproliferative disease (PTLD) risk in Epstein-Barr virus avoidance after kidney transplantation were tried and proven
(EBV)–seronegative patients, current belatacept trial protocols safe. However, similar to alemtuzumab, the use of complete CNI
have now been modified to enroll EBV-seropositive patients only. avoidance protocols should be practiced with extreme caution;
Unfortunately, this exclusion will complicate the introduction in several studies that withheld CNIs for any more than a very
of this drug in young patients, who might derive the maximum short period, despite adequate immunosuppression with IL-2R
long-term benefit from nonnephrotoxic regimens. A significant mAbs, MMF, and steroids, the acute cellular rejection rates after
54
limitation to note is that administration of belatacept requires renal transplantation were much higher than with CNI use.
intravenous infusion; this is not an ideal characteristic for a Thus as with ATG and alemtuzumab, patients receiving basilix-
maintenance immunosuppressant and raises issues of patient imab are still exposed to chronic administration of maintenance
compliance. Furthermore, the agent has a long half-life of 8–10 immunosuppression and its associated comorbidities.

1108 Part NiNe Transplantation


proliferation and the generation of an effective immune response,
Glucocorticoids including the cytokines IL-2, IL-4, TNF-α, and IFN-γ, and
Corticosteroids have complex immunosuppressive as well as costimulatory molecules, such as CD154. However, CNIs are
antiinflammatory effects (Chapter 86). They act principally by well known for their nephrotoxic properties and are thought to
binding to cytoplasmic glucocorticoid receptors, although at be major contributors to chronic allograft dysfunction in renal
higher doses they can exhibit receptor-independent effects as transplantation. Thus in modern immunosuppressive regimens,
well. The steroid–receptor complex translocates to the nucleus, an attempt is made to minimize or even replace this drug class
where it is able to alter the expression of multiple cytokines with less harmful yet equally effective agents.
through DNA-binding and by targeting transcription factors,
such as AP-1 and NF-κB. Corticosteroids reduce the expression Mechanistic Target of Rapamycin Inhibitors
of many molecules important in the immune response, including Sirolimus (rapamycin) and everolimus bind to the same immu-
IL-1, -2, -3, and -6; TNF-α; IFN-γ; and a number of chemokines. nophilin as tacrolimus (FKBP12), although the complexes they
By inhibiting cyclooxygenase, corticosteroids are also able to form are unable to interact with calcineurin. Instead they bind
reduce the production of inflammatory mediators, such as to the regulatory kinase mTOR, which has a critical role in
leukotrienes and prostaglandins. cytokine receptor signal transduction. The usual actions of mTOR
Corticosteroids have been the mainstay of maintenance are to activate the ribosomal enzyme p70 S6 kinase and block
immunosuppression regimens for several decades. However, an inhibitory protein 4E-BP1, both of which are required for
because of the agents’ wide-ranging effects, not only involving translation of proteins necessary for progression from the G 1
the immune response but also a multitude of organ systems, (growth) phase to the S (DNA synthesis) phase of the cell cycle.
and its direct nephrotoxicity and diabetogenic properties, in Therefore inhibition of this pathway in T cells blocks the action
modern immunosuppressive regimens, an attempt is made to of cytokines, such as IL-2, -4, and -15, preventing cell cycle
avoid or withdraw corticosteroids at the earliest possible time progression and clonal expansion.
after transplantation (Table 81.1). In addition to inhibiting clonal expansion of effector T cells,
there is growing evidence supporting the role of rapamycin in
Antiproliferative Agents promoting the generation and survival of Tregs, which have
Azathioprine and MMF interfere with DNA synthesis and prevent been shown in several animal models to be capable of preventing
56
cell cycle progression. In the context of allograft rejection, this allograft rejection. For instance, rapamycin promotes the de
44
impairs the clonal expansion of alloreactive T cells. novo conversion of alloantigen-specific Tregs. This is in contrast
Azathioprine is metabolized in the liver to the purine analogue to CNIs, which inhibit such conversion. This property makes
6-mercaptopurine and incorporated into DNA. By inhibiting rapamycin an attractive agent to potentially promote tolerance,
purine nucleotide synthesis (and therefore DNA and RNA as discussed further in the following section.
synthesis), it reduces gene transcription and prevents cell cycle
progression. The effects of azathioprine are not lymphocyte TOLERANCE
specific, and patients must be monitored closely for bone marrow
suppression. Billingham et al. first introduced the term transplantation tolerance
MMF is metabolized in the liver to mycophenolic acid, which in 1953, with the report that inoculation of fetal mice with
is a noncompetitive, reversible inhibitor of inosine monophos- lymphoid cells from an allogeneic adult donor mouse of a different
phate dehydrogenase (IMPDH). Cells are able to generate purines strain led to later acceptance of skin grafts from the original
57
either de novo by converting inosine monophosphate to guanosine skin graft donors, and for their seminal work Medawar received
monophosphate (catalyzed by IMPDH) or from guanine via the the Nobel Prize for Medicine in 1960.
salvage pathway. The salvage pathway is less active in lymphocytes The condition in which an SOT recipient exhibits a well-
and therefore are relatively dependent on the de novo pathway functioning graft and lacks histological signs of rejection after
of purine synthesis compared with other cell types. As a result, receiving no immunosuppression for at least 1 year is referred to
58
the effects of MMF are more lymphocyte specific than azathio- as clinical operational tolerance (COT). Importantly, patients
prine, and it is less myelosuppressive. Both agents are commonly demonstrating COT must also be capable of responding to other
used in modern maintenance immunosuppression regimens. non–transplantation-related immune challenges, including infec-
tions. Over recent years, experimental models have shown that it
Calcineurin Inhibitors is possible to exploit the processes by which immune homeostasis
The introduction of cyclosporine in the early 1980s presented and tolerance to self antigens are maintained to induce tolerance
a great step forward in transplant-related immunosuppression to alloantigen (Chapter 12). The optimal outcome for patients
because this was the first drug able to selectively block T-cell after transplantation would be harnessing of these mechanisms
activation. Subsequently a second CNI, the macrolide antibiotic to induce specific immunological unresponsiveness or tolerance
tacrolimus, replaced cyclosporine in most immunosuppressive to the graft, thus avoiding the adverse side effects associated with
regimens. Almost 99% of renal transplant recipients currently current immunosuppressive regimens (see Table 81.1).
55
receive CNIs. Both cyclosporine and tacrolimus bind cytoplasmic Unfortunately, more than 5 decades of clinical experience in
immunophilins (cyclophilin in the case of cyclosporine and SOT have demonstrated that COT in humans is extremely difficult
FK506-binding protein 12 [FKBP12] in the case of tacrolimus) to induce intentionally and may only be possible in a subset of
to form complexes that can inhibit the calcium-dependent transplant recipients. Many successful experimental techniques
phosphatase calcineurin, a rate-limiting enzyme in the TCR can produce durable hyporesponsiveness to mismatched allografts
signal–transduction pathway. By preventing translocation of the in rodent models, but scarcely any have been successfully trans-
transcription factor NFAT to the nucleus, calcineurin inhibition lated into large animal models or clinical trials. Progress in the
impairs upregulation of many molecules important for T-cell field of transplantation tolerance has also been hampered by the

CHaPter 81 Concepts and Challenges in Organ Transplantation 1109



tHeraPeUtiC PriNCiPLeS KeY CONCePtS
Mechanisms of Tolerance Barriers to Transplantation Tolerance

Deletion • T-cell memory
Deletion of alloreactive or autoreactive T cells can be achieved centrally • Presensitization—direct exposure to alloantigen (i.e., pregnancy or
in the thymus or in the periphery. Infusion of donor bone marrow into blood transfusion).
a recipient who has been conditioned with nonmyeloablative irradiation • Heterologous immunity—cross-reactivity in the T-cell repertoire
or immunotherapy enables antigen-presenting cells (APCs) to access among antiviral, antibacterial, environmental, and transplantation
the thymus and trigger the deletion of maturing thymocytes. In the antigens.
periphery, deletion can be triggered by alloantigen recognition under • Homeostatic proliferation—induced by immunodepletory antibodies
suboptimal conditions, including costimulation blockade, as well as (i.e., alemtuzumab).
immunodepletion by activation of apoptotic cell death and cytolysis. • Memory T cells generated by the above mechanisms—can result
in rapidly formed effector immune responses upon rechallenge.
anergy These T cells are less sensitive to T-cell depleting antibodies and
This is the functional inactivation of the T-cell response to restimulation costimulatory blockade and thus may be more resistant to some
tolerance induction strategies.
by alloantigen or self antigen and has been described both in vivo and • B-cell response
in vitro. Some forms of T-cell anergy are also reported to result in the
development of regulatory activity. Costimulation blockade, as well as • Recipients treated with immunodepletory antibodies display a general
increase in the naïve B-cell population.
inhibition of downstream proliferative pathways, can trigger anergic states
in T cells. • There is a prevalent development of alloantibody in recipients treated
with depleting antibody therapy.
• Much focus of tolerogenic strategies is on the T-cell response.
immunoregulation However, recent data suggest that the humoral immune system
This active process results in the regulation of one cell population may play a more significant role than previously thought, possibly
by the activity of another cell population. Various populations of contributing to more long-term outcomes. Further work in this area
leukocytes have been described as having the ability to control immune continues.
responsiveness to alloantigen stimulation in both innate and adaptive • Lack of tolerance signature
immune responses. This mechanism, although well described in experi- • An episode of acute rejection can severely affect graft survival in
mental models, has yet to be introduced therapeutically in a sustained, most transplanted organs.
clinical setting. • There is absence of validated biomarkers of tolerance or predictors
of rejection.
Clonal exhaustion • It is clinically difficult to justify often high-risk tolerizing strategies
This can occur as a result of chronic antigen stimulation or antigen recogni- in patients who would otherwise do moderately well with standard
tion under suboptimal conditions. The consequence is either deletion or immunosuppression.
functional inactivation of the cells responding to the recognized antigen.
An example of such exhaustion can be seen in liver transplantation,
where the large number of donor-derived APCs migrating from the liver
to draining lymphoid tissues after transplantation can trigger this type
of response.
In the next section, we discuss the current clinical strategies
ignorance for tolerance induction as well as much-needed future approaches
This is an uncommon mechanism in the induction phase of unresponsive- to produce more short-acting, antigen-specific agents that can
ness to alloantigen as it is difficult to introduce donor cells or tissue optimize outcomes in the clinic.
without alerting the immune system to their presence, in transplantation.
This mechanism, however, does describe the natural state of some Molecule-Based Tolerogenic Protocols
self-reactive CD4 T-cell populations found in healthy individuals with no The induction of COT through the administration of presumed
autoimmune pathology.
tolerogenic drugs and subsequent immunosuppression minimiza-
tion is one such strategy currently under investigation. Promising
results of the first use of alemtuzumab in human subjects by
Calne et al. 31,62 with low-dose cyclosporine alone as maintenance
therapy introduced the concept of “prope” tolerance (i.e., graft
lack of definitive laboratory parameters able to give a clear acceptance with reduced immunosuppression). Several groups
indication of whether a particular recipient is tolerant of his or have subsequently studied alemtuzumab induction and main-
her graft. Furthermore, COT appears to be somewhat organ tenance therapy without nephrotoxic CNI agents. Unfortunately,
dependent. Recipients of a liver graft are more capable of develop- results of pilot studies have been poor, with high acute rejection
ing COT because of the immune-privileged status of the liver rates (as much as 36% of recipients in one study). As discussed
(see Fig. 81.7). Clinical experience in liver transplantation now above, an attempt to induce donor allograft tolerance using
demonstrates clear evidence that a permanent and stable alemtuzumab alone as an induction therapy with no maintenance
immunosuppressive-free state can be safely attempted and immunosuppression at all resulted in all of the patients developing
31
sometimes achieved in patients who have received a liver trans- acute rejection within the first month after transplantation.
59
plant for nonimmunological liver diseases. However, COT has Encouragingly, physicians from the University of Wisconsin have
never been reported after intestinal, islet, or whole-organ pancreas demonstrated excellent long-term graft survival in 90% of patients
transplantation, whereas two exceptional cases of COT have been maintained with only low-dose rapamycin monotherapy; these
described after lung and heart transplantations. 60,61 Since the recipients had undergone alemtuzumab induction and an initial
first renal transplant over 55 years ago, only sporadic cases of 60-day-only treatment with a CNI after transplantation to prevent
COT have been documented after renal transplantation in the acute rejection. It should be noted, however, that 50% of the
absence of genetic identity between donor and recipient. 59 patients developed alloantibody to either HLA class I or II

1110 Part NiNe Transplantation



Tolerogenesis in the Liver Clinical Observations
The liver is an important site for primary T-cell • Positive cross-match or blood type
activation, but this takes place in an incompatibility has little affect on graft survival
environment biased toward tolerance:
• Liver endothelium expresses adhesion • HLA matching is not a prerequisite to liver
molecules that facilitate the sequestration of transplantation
circulating activated T cells, particularly CD8
T cells. This gives the liver a role in systemic • Reduced incidence of hyperacute rejection
immunoregulation. ompared to other organs in SOT
• These activated T cells may undergo Liver sinusoid
apoptosis owing to FasL and TRAIL TGF-β1 HCS • Frequent spontaneous recovery after severe
expressed on Kupffer cells (KC) rejection
and may also be phagocytosed. PD-L1
• Constitutive exposure of liver cells to traces of • Acute rejection does not impact severely on
endotoxin and other microbial products via T cell long-term graft and patient survival
blood from the systemic circulation and Trapping T cell IL-10
intestine results in down-modulation of Apoptosis • Liver allograft protects other extra-hepatic
costimulatory molecules and the synthesis of Phagocytosis KC FasL allograft from rejection if derived from
IL-10 by Kupffer cells (KC) and liver sinusoidal TRAIL KC the same donor
endothelial cells (LSECS). LSEC
• Non haematopoietic liver cells, including • Lower overall incidence of chronic rejection
LSECs and hepatic stellate cells (HSC) act as that is reversible in up to 30% cases
APCs, presenting antigen to T cells in the
presence of immunosuppressive cytokines • Clinical operational tolerance (COT) has been
(IL-10 and TGF-β1) and inhibitory cell surface achieved most reliably and frequently in liver
igands (PD-L1). Anergy, rather than activation transplantation than with any other solid organ
is promoted. Thus immune responses to liver
antigens can often result in tolerance.
FiG 81.7 Tolerogenesis in the Liver. The liver is an important site for primary T-cell activation,
but this takes place in an environment biased toward tolerance. Liver endothelium expresses
adhesion molecules that facilitate the sequestration of circulating activated T cells, particularly
CD8 T cells. This gives the liver a role in systemic immunoregulation. These activated T cells
may undergo apoptosis as a result of FasL and TRAIL expressed on Kupffer cells (KCs) and may
also be phagocytosed. Constitutive exposure of liver cells to traces of endotoxin and other
microbial products via blood from the systemic circulation and intestine results in downmodulation
of costimulatory molecules and the synthesis of interleukin-10 (IL-10) by KCs and liver sinusoidal
endothelial cells (LSECS). Nonhematopoietic liver cells, including LSECs and hepatic stellate cells,
act as APCs, presenting antigen to T cells in the presence of immunosuppressive cytokines
(IL-10 and transforming growth factor-β 1 [TGF-β 1 ]) and inhibitory cell surface ligands (PD-L1).
Anergy, rather than activation, is promoted. Thus immune responses to liver antigens can often
result in tolerance.



+
molecules and 40% demonstrated C4d graft biopsies indicative It is important to note that other protocols based on a similar
of subacute humoral responses to the allograft. 39 strategy have since been attempted, including post–liver trans-
In 2003, investigators at the University of Pittsburgh published plantation protocols (the liver being an organ more capable of
the results of a single center trial in which they administered an developing COT compared with any other). However, none of
immunosuppressive regimen deemed to be tolerogenic to 82 these protocols has achieved COT, nor have these protocols shown
63
adult kidney, liver, pancreas, and intestinal transplant recipients. any impact on overall outcomes convincingly. In most situations,
Their working hypothesis was that the need for continuous it appears that leukocyte depletion is not accompanied by a
high-dose immunosuppression could be avoided in most cases permanent and complete deletion of alloaggressive donor-reactive
with the use of a strong lymphocyte-depleting regimen before cells, and the establishment of a regulatory network is required
engraftment, followed by the administration of low-dose tacro- to maintain tolerance.
limus monotherapy. The goal of the induction treatment was
the nonspecific removal of clones of immune cells responsible Full Chimerism
for rejection before contact with foreign donor antigens occurred The more robust experimental strategies for the induction of
while minimizing exposure to maintenance immunosuppression. tolerance to foreign antigen utilize the mechanisms of central
At the time of publication of the study, 64% renal, 70% liver, deletion to eliminate T-cell clones with specificity for the foreign
42% pancreas, and 54% intestinal transplant recipients were on antigens in question, thereby preventing them from entering the
spaced doses, but no patient could be weaned completely off periphery. This can be reliably achieved by the establishment of
immunosuppression. Nevertheless, the striking reduction in daily hematopoietic chimerism through bone marrow transplantation
dosing was a step forward because it led to a significant reduction (BMT). Stable engraftment of donor hematopoietic stem cells
in overall immunosuppression-related morbidity. (HSCs) results in repopulation of the recipient thymus with

CHaPter 81 Concepts and Challenges in Organ Transplantation 1111


donor-type thymic DCs, with the result that developing T cells with mild, nonmyeloablative total body irradiation or costimula-
with antidonor specificity are deleted by negative selection. tory blockade with anti-CD154 and/or CTLA-4 Ig. When these
Full chimerism can be obtained rapidly through the ablation induction protocols are followed by BMT, the result is mixed
of the recipient’s marrow and immune system with high-dose chimerism (the continued survival of both donor and recipient
radiation and/or chemotherapy; in addition it can be induced hematopoietic progenitor cells). Animals that have undergone
more slowly by nonablative conditioning regimens, followed by these therapies have demonstrated durable tolerance to donor-type
the infusion of donor’s marrow to colonize the recipient com- allografts and have a much lower incidence of GvHD compared
pletely. This phenomenon paves the way for the onset of tolerance with full chimeras.
in the case of a subsequent SOT from the same donor. 64 In particular, the Massachusetts General Hospital (MGH)
As proof of concept, numerous patients have undergone suc- transplantation group showed in mouse and NHP models that
cessful BMT for hematological indications and have subsequently nonmyeloablative regimens could result in transient mixed
been successfully transplanted with a kidney from the same donor, chimerism with accompanying long-term transplantation toler-
without the requirement for increased immunosuppression. ance. Indeed, the same group has reported the long-term follow-up
It is important to highlight that in all these cases, the use of of human recipients, who underwent combined HLA-matched
BMT was justified on the basis of the need for treatment of bone marrow and kidney transplantation for end-stage renal
hematological malignancies. disease with or without multiple myeloma. 66,67
An alternative approach to achieving full chimerism and hence
tolerance to donor alloantigens while avoiding GvHD or engraft- Regulatory T Cells
ment syndrome, in HLA-mismatched unrelated stem cell/renal There is now abundant evidence for the existence of populations
65
transplant human recipients has been reported. Results were of regulatory lymphocytes with the ability to suppress immune
achieved with the introduction of a mobilized cellular product responses by other leukocytes (Chapter 18). Tregs can be divided
enriched for tolerogenic graft facilitating cells (FCs) as well as into two populations: thymic-derived naturally occurring
+
+
HSCs and T cells (the treatment with HSCs in combination with CD25 CD4 cells (tTregs) and induced Tregs or iTregs that are
+
−
FCs is termed FCRx) rather than just bone marrow graft alone. either differentiated from CD25 CD4 nonregulatory cells or
+
+
+
These bone marrow–derived FCs, which are CD8 but do not expanded from CD25 CD4 cells in response to antigen. Expres-
express a TCR, potently enhance engraftment of allogeneic HSCs sion of the transcription factor FOXP3 is essential for the
in conditioned recipients. FCs are composed predominantly of development and function of Tregs; however, tTregs and iTregs
a plasmocytoid precursor DC subpopulation, induce the genera- differ in origin, antigen experience, methylation patterns of
tion of antigen-specific Tregs in vitro and in vivo, and have been FOXP3, and suppressive mechanisms. Both tTregs and iTregs
found to effectively prevent GvHD in mice. In the first phase of have been demonstrated to play important roles in transplant
this single-center clinical study, eight HLA-mismatched kidney tolerance.
transplant recipients were treated with FCRx and nonmyeloabla- tTregs develop within the thymus under the direction of FOXP3
tive conditioning during the peritransplantation period. One and have a critical role in limiting immune responses to self
year later, five recipients exhibited durable macrochimerism with antigens, as demonstrated by experimental models where mice
+
+
no incidence of GvHD or engraftment syndrome, and were depleted of CD25 CD4 cells subsequently develop inflammatory
immunosuppression-free—that is, these patients were clinically bowel disease (IBD) and widespread autoimmune phenomena.
operationally tolerant. In vitro studies indicated that chimeric Mutation of FOXP3 in humans is responsible for immune
donor lymphocytes were tolerized to the recipient, and a sig- dysfunction/polyendocrinopathy/enteropathy/X-linked (IPEX)
nificant increase in the CD4 Treg/T effector cell population ratio syndrome (Chapter 35). There is evidence for the involvement
was observed in these patients when compared with those who of Tregs in the downregulation of immune responses to tumors
had lost chimerism. The ability to establish high levels of donor and chronic infections, as well as allogeneic transplants. 70
multilineage chimerism in haploidentical and highly mismatched Many strategies exist for the in vivo or ex vivo generation
unrelated donor–recipient pairs without the development of and/or expansion of Tregs. The most common in vivo approaches
GvHD through the use of novel cellular therapies could have are based on the fact that stimulation with anti-CD3 and
exciting therapeutic implications for disorders for which HSC anti-CD28 in the presence of high concentrations of IL-2
transplantation can provide a “functional cure,” including stimulates proliferation and that exposure to antigen increases
inherited metabolic disorders, hemoglobinopathies, and auto- Treg frequency and/or potency by either expanding tTregs or
immune disease, as well as in solid organ transplantation. inducing the generation of iTregs from cells that do not originally
Longer-term follow-up data on these initial patients as well as possess regulatory activity. There is a large body of data dem-
data from the enrollment of additional patients into this protocol onstrating that ex vivo expanded Tregs can protect allografts
continue to be encouraging. A phase III multicenter clinical trial from rejection. 56,71
is planned. Transplant recipients are treated with diverse immunosup-
pressive drug combinations, which may have a different impact
Mixed Chimerism on Tregs. As mentioned in the previous section, it has been
A promising approach is the induction of mixed hematopoietic demonstrated that CNIs, in particular cyclosporine, are detri-
chimerism, which can be achieved in experimental models with mental to Tregs, whereas the mTOR inhibitor rapamycin was
far less toxic induction therapy. Nonmyeloablative conditioning shown to be beneficial to Tregs both in terms of in vivo generation
regimens have been applied in the clinical setting in patients and function in mouse models and in vitro cultures of human
who develop renal failure as a consequence of multiple myeloma. Tregs. Adoptive transfer of a low number of alloantigen-specific
Further refinements of such protocols are being explored in Tregs under a cover of low-dose rapamycin was found to be
experimental models. Examples of these include either a combina- capable of inducing long-term survival of heart transplants in
tion of depleting anti-CD4 and anti-CD8 antibodies together an unmanipulated host, an outcome otherwise difficult to obtain. 72

1112 Part NiNe Transplantation


There are significant differences between human and mouse
Tregs, in particular the differences in FOXP3 expression: In Biomarkers of Rejection or Tolerance
humans, FOXP3 is also expressed transiently by activated non- Concerns have been raised regarding the evaluation of tolerance
regulatory T cells that also upregulate CD25 expression. Thus induction protocols, such as those using cellular therapies in
+
+
+
not all CD25 FOXP3 CD4 cells will be genuine Tregs, and recipients of kidney, heart, or pancreas, as an episode of acute
+
hi
therefore isolation strategies based on CD25 CD4 are likely to rejection could severely affect graft function and survival. In
be imperfect. Stable expression of FOXP3 in Tregs depends on these settings, most physicians are reluctant to withdraw immu-
DNA demethylation at the Treg-specific demethylated region nosuppressive drugs in the absence of validated biomarkers of
(TSDR), a conserved, CpG-rich region within the FOXP3 locus. transplantation tolerance and rejection prediction. Biomarkers
The TSDR is selectively demethylated in tTregs (which are more are thus needed to better evaluate the immune status of transplant
stable than pTregs), and therefore the methylation status of the recipients, determine the “tolerance signature” of recipients,
TSDR region can act as a marker for the quality and stability of predict and diagnose graft rejection noninvasively, and individual-
a Treg population. Cell-surface markers that can enrich for Tregs ize immunosuppressive therapy.
+
+
lo
have also been described. In humans, CD127 CD25 CD4 T Technical advances in multiparameter flow cytometry, antigen-
cells are characterized by a higher level of FOXP3 expression specific lymphocyte assays, and genome-wide analyses have led
and a more pronounced suppressive capacity. 70,71 Importantly, to the development of powerful and more standardized immu-
+
+
+
lo
hi
ex vivo expanded CD25 CD4 and CD127 CD25 CD4 Tregs nomonitoring techniques to characterize alloimmune responses.
are effective at inhibiting vasculopathy in a humanized mouse Histological analysis remains the gold standard method by which
+
+
lo
model, whereas CD127 CD25 CD4 T cells are five times more to detect graft rejection, but graft biopsy is an invasive procedure
efficient than those not expressing a low level of CD127. 56 with inherent risks and so cannot be routinely performed for
A number of strategies for the isolation or enrichment of surveillance purposes. It is also associated with sampling error
human Tregs have been described, but there is no consensus as and interpretation biases.
to which strategy produces the optimal population for use in Molecular or cell-based biomarkers are being developed as
cell therapy applications. The most commonly used expansion alternative surveillance methods. 20,75 At present, it is notable that
protocol at present is based on stimulation by anti-CD3/anti-CD28 for the many biomarkers identified, there are discrepancies among
beads in the presence of high doses of recombinant IL-2, supple- data from different laboratories investigating the same parameter.
mented in some protocols with rapamycin. Despite the generation This is likely caused by differences in recipient and donor popula-
of sufficient numbers of Tregs for cellular therapy, this mode of tions, clinical management, and the laboratory techniques used
expansion is antigen nonspecific without any enrichment steps to perform the assays. Therefore it is of great importance that
for the cells of interest. The concept of expanding or generating the assays used to detect biomarkers are optimized, validated,
donor alloantigen-reactive Tregs is much more appealing and standardized and that candidate biomarkers are investigated
for clinical application in the setting of transplantation and a in large patient cohorts from different transplantation centers
number of studies have shown that donor alloantigen-reactive using the same shared and validated assays. 76
Tregs are more potent on a cell-per-cell basis at controlling
allograft rejection. Further work to ascertain the long-term ON tHe HOriZON
stability and plasticity of Tregs in vivo, as well as the potential
effects of Tregs in antitumor and antiviral immunities, as theoreti- Initiatives in Clinical Utilization of Tolerance
cally the infusion of large numbers of potent suppressive cells • Clinical introduction of antigen-specific regulatory T-cell therapy
may compromise the immune response toward infectious • Identification of tolerance signatures and predictors of rejection
pathogens and tumor cells. 73 • Use of immunomodulatory stem cell populations (i.e., mesenchymal
Several other immunomodulatory cell groups are currently stromal cells)
under investigation as potential tolerogenic therapies, such as
mesenchymal stromal cells (MSCs) and regulatory macrophages. Please check your eBook at https://expertconsult.inkling.com/
MSCs were originally isolated from the bone marrow but can for self-assessment questions. See inside cover for registration
now be isolated from almost any human tissue, and they possess details.
fascinating tissue repair and immunoregulatory properties.
Preclinical models indicate that MSCs can promote engraftment
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CHaPter 81 Concepts and Challenges in Organ Transplantation 1114.e1


MUL ti PL e -CHO i C e QU e S ti ONS

1. The transfer of an organ between genetically identical individu- C. About the same as the frequency directed against a nominal
als is termed: antigen
A. An autograft D. Zero given that T cells are self-restricted
B. An allograft E. Approaching 10% of T cells
C. A xenograft 3. What mechanism is responsible for the presence of memory
D. An isograft T cells reactive against a graft in a patient receiving his or
E. A haplograft
her first transplant?
2. The proportion of T cells that are capable of directly recogniz- A. Previous blood transfusion
ing allogeneic major histocompatibility complex–peptide B. Previous pregnancy
complexes is: C. Heterologous immunity
A. Low before transplantation D. Direct allorecognition
B. Only the memory T-cell subpopulation E. All of the above

82






Immune Reconstitution Therapy for

Immunodeficiency



Luigi D. Notarangelo, Sung-Yun Pai










Following the discovery of the major human leukocyte antigens or T cell–depleted) and the weight of the recipient. Blood
(HLAs) in 1958, hematopoietic stem cell transplantation (HSCT) group matching for ABO antigens is not required for HSCT, as
came into practice to provide treatment for a variety of congenital mature red blood cells (RBCs) or anti-ABO antibodies can be
and acquired disorders. Cure of an infant with severe combined removed by RBC depletion and plasma depletion, respectively.
immunodeficiency (SCID) in 1968 was the first successful experi- In the case of HLA-identical transplantation, marrow stem cells
ence in HSCT, marking the beginning of a new era in medicine. are then injected intravenously without further manipulation
Shortly thereafter, partial success was achieved also with HSCT into a central line in the recipient. In the case of mismatched
in a child with Wiskott-Aldrich syndrome (WAS). transplantation, bone marrow cells are usually T cell depleted in
For many years, the successful use of HSCT in severe vitro (see below); stem cells are then enumerated and injected
primary immune deficiency (PID) was largely restricted intravenously. More recently, injection of unmanipulated
to transplantation from HLA-matched related donors (MRDs), bone marrow cells from MMRD, followed by in vivo admin-
as HSCT from mismatched related family donors (MMRDs), istration of cyclophosphamide to prevent GvHD, has been
usually represented by haploidentical parents, was followed also used. 3
by severe complications, graft-versus-host disease (GvHD) in HSCs can also be retrieved from peripheral blood, following
particular. In the late 1970s, it was demonstrated in animal in vivo administration of granulocyte–colony-stimulating factor
models that removal of mature T lymphocytes from the graft (G-CSF) to the donor, usually at the dosage of 10 µg/kg/day for
obtained from mismatched marrow allowed successful reconstitu- 5 days, which allows mobilization of stem cells. In this case, stem
tion upon injection into lethally irradiated recipient animals. cells can also be purified by positive selection (see below), enumer-
This important achievement opened the way to a broader use ated, and injected.
of HSCT in severe forms of PID. More recently, transplantation Finally, UCB is another rich source of HSCs. At birth, cord
of hematopoietic stem cells (HSCs) from volunteer matched blood is collected in a heparinized medium and stored in liquid
unrelated donors (MUDs) and of umbilical cord blood (UCB) nitrogen, with small aliquots preserved for HLA typing. Whenever
has been increasingly used in individuals with PID. Overall, since sufficient compatibility is identified between a patient and stored
1968, over 2000 transplantations have been performed in patients cord blood, the latter is thawed and injected into the recipient
1,2
with PID, most of them in children with SCID. The increasing without further manipulation. With this procedure, the number
number of transplantations over the years reflects an increased of HSCs that can be transplanted is dictated by their concentration
awareness of PID, the broader availability of diagnostic in the cord blood sample and by the volume of the sample itself.
tools (including newborn screening for SCID), improved out- More recently, in vitro expansion of cord blood stem cells, and
comes as a result of advances in supportive and critical care transplantations with multiple cord blood units, have been
before and after HSCT, increasingly improved strategies to prevent attempted to overcome this limitation.
GvHD, and the greater availability of MUD and UCB for
transplantation. Donor Selection and Manipulation of the Graft
HSCT From a Related HLA-Identical Donor
HEMATOPOIETIC STEM CELL TRANSPLANTATION: The use of unfractionated stem cells from an HLA-identical
GENERAL CONSIDERATIONS sibling offers the best chance of rapid engraftment and immune
Sources of Hematopoietic Stem Cells reconstitution. In such cases, the HLA-identity between recipient
and donor minimizes the risk of GvHD. Furthermore, the mature
for Transplantation T cells contained in the graft provide a first line of immune
Several sources of HSCs are available for transplantation (Table reconstitution after transplantation, as they may expand and
82.1). HSCs can be retrieved from bone marrow, peripheral blood, lead to a rapid increase in the number of circulating T lympho-
or UCB (Chapter 2). Bone marrow stem cells are most commonly cytes as early as 2 weeks after HSCT. Engraftment of these mature
obtained by multiple aspirations along the iliac crests, usually T cells transplanted with the bone marrow of an MRD is par-
while the donor is under general anesthesia. The volume of bone ticularly important in infants with SCID, as it is early evidence
marrow that is obtained may vary between 500 mL and 1 L or of immune reconstitution in a severely immunocompromised
even more, depending on the type of transplant (HLA-identical host.

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