BSHI programme v 3

BRITISH SOCIETY FOR
HISTOCOMPATIBILITY
AND IMMUNOGENETICS

CONFERENCE PROGRAMME

BSHI 2016 Organising Committee

Susan Fuggle Chair Local Organising Committee Oxford University Hospitals NHS Trust
Martin Barnardo
Mian Chen Local Organising Committee Oxford University Hospitals NHS Trust
Maggie Sutton
Jeanette Ayers Local Organising Committee Oxford University Hospitals NHS Trust
David Briggs
Luke Foster Local Organising Committee Oxford University Hospitals NHS Trust
Katy Derbyshire
Local Organising Committee Oxford University Hospitals NHS Trust

Chair Research Executive NHSBT Birmingham

Secretary Research Executive NHSBT Birmingham

Meeting Secretary Barts Health NHS Trust

The conference organisers gratefully acknowledge the support of the following:

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Contents Page 2
Page 4
Local organisers and Sponsors Page 5
Welcome from Local Organising Committee Page 8
About Oxford Page 9
Thanks Page 12
Conference Information Page 13
Social Programme Page 17
Conference Programme Page 29
Speaker Biographies and Abstracts Page 33
Oral Presentation Abstracts Page 37
One Lambda and VH Bio Young Scientist Award Abstracts Page 41
Julia Bodmer Award Abstract Page 62
Poster Abstracts Page 66
Abstract Author Index Page 67
BSHI AGM Agenda Page 68
Exhibition Floorplan Page 74
Exhibitor and Sponsors Biographies and Workshop Information Page 77
Delegate List
Notes

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Welcome to the 27th Annual Conference of the British Society
for Histocompatibility and Immunogenetics

Welcome to the 27th Annual British Society for Histocompatibility and Immunogenetics Conference at
Keble College, Oxford. The University of Oxford was founded in 1096 and granted its charter in 1248,
making it the second oldest university in the world (Bologna was founded eight years earlier!). Keble is
one of the 38 colleges comprising the University of Oxford and built between 1868 and 1876 to William
Butterfield’s striking neo-Gothic polychromatic brick design. These buildings have been supplemented by
award winning contemporary developments, including the modern conference facility.

Keble College was one of the venues used for the 7th Histocompatibility Workshop, held in Oxford in
September of 1977. The investigations in this workshop included the serological study of the then
newly-described IA (Immune-associated) determinants, present on B cells, but not on T cells and the
relationship between these specificities and the Dw determinants defined by cellular assays. Based on
workshop results, the HLA-DR locus was officially defined and the first seven HLA-DR antigens identified.
Evidence of further Ia (class II) loci began to emerge from the results of this workshop.

For the BSHI Conference 2016 members of the Research Executive, together with the local organisers, have
developed a programme to cover current areas of interest in H&I. We are delighted to welcome all of our
invited speakers including Professor Paul Moss, Dr John Girdlestone, Professor Paul Johnson, Rachel Johnson,
Professor Tim Elliott and Dr Robert Vaughan. Professor John Forsythe will give the Presidential Lecture;
Professor Kathryn Wood (University of Oxford) will deliver the Festenstein Lecture and Professor Bob
Montgomery (NYU Langone Transplant Institute) the Terasaki Lecture.

Thank you to all who submitted abstracts and we look forward to the oral and poster sessions. As always,
prizes will be awarded for the best abstract, poster and young investigator presentations.

The society gratefully acknowledges the contribution made by all of our sponsors, without which the
conference could not be held. We strongly encourage all delegates to visit the exhibition stands and
attend the lunchtime presentations.

The conference banquet will be held in Keble’s historic Dining Hall with pre-dinner drinks in Liddon Quad.
Calorific content of the banquet can be offset by joining the Fun Run (3.5 km approx) which departs at
07:00 on Wednesday 14th from Keble lodge. This will take in many of Oxford’s historic sites including
Christchurch College, Deadman’s Walk, Radcliffe Camera and Bodleian Library.

We hope that you have an enjoyable and productive conference.

BSHI 2016 Organising Committee

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About Oxford

Oxford, known as the “city of dreaming spires” is alive with history and heritage; a place filled with
incredible architecture and home to a vibrant arts and culture scene. You can tour one of the University of
Oxford’s famous colleges; visit the Ashmolean Museum and see the Egyptian mummies; take to the water
on a punt or boat and enjoy the views along the banks of the River Thames; or alternatively stroll through
the city’s streets – the setting for numerous films and TV series.

Oxford has featured heavily in key periods in British history. Visitors can discover the role of the city in
the English Civil War and climb the Saxon St George Tower at the Oxford Castle.

Oxford’s excellent museums cater for all interests ranging from art and archaeology at the Ashmolean
Museum; the wonder of the Natural History Museum and its collection of dinosaurs through to the
history of the city at the Museum of Oxford.

Oxford’s contribution to literature, film and TV is for all to see. Visit the places frequented by literary
greats such as Tolkien and CS Lewis; tour the filming locations for popular films such as Harry Potter or
retrace the steps of TV’s Inspector Morse, Lewis and Endeavour.

We have listed some specific sites of interest below.

Architecture

Radcliffe Camera is an iconic Oxford building. 'Camera' in Latin means room. It is actually one large
circular room with a spectacular dome and now part of the Bodleian library.

Bodleian Library is one of the oldest libraries in Europe and it is the second largest library in England after
the English library.

Covered market is the place to go to for high end local produce and other useful products. You can get
everything from fresh seafood, wool clothing, gourmet sandwiches, chocolates, coffee, pies, meat and
flowers.

Hertford Bridge (Bridge of Sighs) The arched and covered Hertford Bridge over New College Lane connects
two Hertford College buildings and is more popularly known as the "Bridge of Sighs" since it seems to be
similar to the "Bridge of Sighs" in Venice.

Sheldonian Theatre on Broad Street is an architectural jewel that was designed by Sir Christopher Wren.
It is largely used for University ceremonies including matriculation and graduation. The painted roof is
also a spectacle. The theatre is open to the public at certain times and hosts performances and shows.

Holywell Music Room is the City of Oxford's chamber music hall. It is thought to be the oldest purpose
built music room in Europe and also the oldest concert hall in England. The Music Room is on Holywell
Street.

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Museums

Ashmolean Museum located on Beaumont Street is the University's museum of art and archaeology and is
the world's oldest university museum. Originally housing the collection of Elias Ashmole, its current repertoire
includes priceless paintings, archaeological findings, silver and fine art.
Museum of Natural History is located on Parks Road, this impressive neo-Gothic building displays the
University's collection of natural specimens – zoological, entomological and geological. Highlights include
the Oxford Dodo and dinosaur remains. It is well worth a visit if you have the time.
Pitt Rivers Museum is a treasure trove of ethnographic objects from around the world. The collection is both
overwhelming and astonishing. Popular exhibits include real shrunken human heads and totem poles. Pitt
Rivers museum is located behind, and accessed through, the Museum of Natural History.
Museum of History of Science has a fine collection of mathematical, physical, astronomical and scientific
instruments from various periods. This includes a large collection of astrolabes and microscopes. The museum
is located on Broad Street.
Museum of Oxford This museum gives you an idea of the long history of Oxford, which goes way back before
the beginnings of the University. It is located adjacent to the Town Hall on St Aldate's and is the only museum
in this list which is not run by Oxford University. It is largely managed by volunteers.

Open Spaces

Christ Church Meadow is a vast expanse of green that lies between Christ Church and the River Isis (the
name for the River Thames as it passes through Oxford). This is opposite the visitor entrance to Christ
Church and can also be reached via Rose Lane adjacent to the Botanic Garden.
University Park is another large expanse of green that has numerous walkways, greens and fields, normally
used for cricket, rugby and lawn tennis. It also offers convenient barbecue spots and picnic spots by the
River Cherwell.
The University of Oxford Botanic Garden is the oldest botanic garden in Britain. Plants and flowers are
beautifully arranged in this garden, which is also a research institution of the University. The Botanic Garden
is located at the east end of High Street opposite Magdalen College.
Punting on the river requires a bit of skill and practice to travel in a straight line. If you manage to go punting,
there are memorable views of the Oxford skyline in the background from Christ Church meadow or
Magdalen Bridge.

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Historic Pubs

The Eagle and Child is best known as the meeting place of the Inklings - an informal literary group that
included C.S. Lewis (Chronicles of Narnia) and J.R.R Tolkien (The Lord of the Rings). A plaque in a corner
which was part of the 'Rabbit Room' commemorates where they met regularly. Apparently this pub also
served as the lodgings of the Chancellor of the Exchequer during the Civil War.
The Lamb and Flag is located on St Giles. Apparently Thomas Hardy wrote much of Jude the Obscure here
and the pub was also frequented by C.S. Lewis. The Lamb & Flag used to be a coaching inn on the London
to the North route where stagecoach drivers, horses and travellers used to rest.
The King's Arms pub is located at the corner of Holywell Street and Parks Road, the site of which was
originally an Augustian priory. The name King's Arms was apparently in reference to King James I who was
closely associated with Wadham College next door.
The Turf Tavern a popular 13th century pub, quite hard to find and accessed either a narrow alley off Holywell
Street or a narrow passageway off New College Lane (just past the Bridge of Sighs). Apparently this is
Inspector Morse's favourite watering hole.

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Thanks

The Organising Committee would like to thank the following individuals who have contributed to BSHI
2016:
Special thanks to all the Speakers and Chairs
Reviewers of abstracts:

Martin Barnardo
Mian Chen
Luke Foster
Zareen Deplano
Vivien Hanson
David Lowe

Judges for the One Lambda and VH Bio Young Scientist Award, Best Abstract Presentation and Best Poster:
We’ll keep your identity secret, but you know who you are!
Trade representative liaison:
Rob Cordingley (Alpha Biotech)
We would also like to thank all our corporate sponsors, in particular:

● VH Bio for sponsoring the Terasaki Lecture and providing the BSHI mugs
● Immucor for sponsoring the wine at the Gala Dinner
● VH Bio for sponsoring the One Lambda and VH Bio Young Scientist Award
● R.O.S.E. for sponsoring the delegate bags and notepads
● Takara for sponsoring the lanyards
● Alpha Biotech for sponsoring the pens

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Conference Information

Registration and Information Desk

The Registration and Information Desk, located in the Sloane Robinson building, will be open at the following
times:

Tuesday 13th September 09:00 – 18:00

Wednesday 14th September 08:30 – 16:30

Contact numbers during the Conference

Keble College Conference Office: 01865 282322
Conference Registration Desk: 077650 67306

Conference and Exhibition Venues

The conference sessions will all take place in the auditorium in the Sloane Robinson Building. The poster
displays and some exhibition stands will be in the Douglas Price Room in the Sloane Robinson Building. The
majority of the exhibition stands will be on the ground floor of the ARCO Building.

All refreshments and lunches will be served in the exhibition area in the ARCO Building. If you are attending
one of the lunchtime sessions you can take your lunch in with you. Breakfast for all delegates staying
overnight at Keble will be served in the Dining Hall.

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Checking into your accommodation

Check in is via the Porter’s Lodge which you will enter through on the way to conference. We guarantee
access to the rooms from 2pm but if the rooms are ready then you will be able to get in earlier.
Check out will be by 10am on the day of departure.

Cloakroom and left luggage

There is a cloakroom and left luggage facility available throughout the conference in the Porters' Lodge
at Keble College where bags can be left on arrival.

Exhibitor Presentations

Five companies will be delivering presentations over the lunchtime in the seminar rooms in the Sloane
Robinson Building. As shown on the programme there will be some time to have lunch before the sessions
start, but do please take your lunch along with you if you haven't quite finished!

Conference Dinner

All full delegates are invited to join us for the Gala Dinner in the Dining Hall on Tuesday evening. There is a
set menu but if you have any dietary requirements, please do not hesitate to let the conference desk know.
We invite you to dress to impress! The drinks reception will begin on Liddon Quad at 7:00pm with dinner
being served at 7:30pm. Awards will be presented at the Dinner followed by music and dancing in the College
Bar.

Internet Access

Internet access in bedrooms is hard wired, you will need an Ethernet cable. Ethernet cables are available
for hire in the Porters Lodge, a £5 refundable deposit is charged.
To access hard wired internet, open your web browser to be redirected to the Keble registration website,
click the “Begin Registration” button, select the name of conference you are attending and follow the prompts
on screen.
The password is: BSHI16
Note: If you are not redirected to the Keble registration website then just try to browse to any non-https
webpage (such as www.it.ox.ac.uk) to force the redirection.

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Wireless internet is available in Liddon blocks 4 to 9, Pusey blocks 2 & 4, Hayward & Debreyne blocks
1 – 8, Sloane Robinson and Arco

For Wi-Fi access please connect to: Keble

Then use password: RedBrickOxford
Next open your web browser to be redirected to the Keble registration website, click the “Begin Registration”
button, select the name of the conference you are attending and follow the prompts on screen.

The conference password is: BSHI16
Note: If you are not redirected to the Keble registration website then just try to browse to any non-https
webpage (such as www.it.ox.ac.uk) to force the redirection.

Medical and fire procedures / first aid

In the event of a fire please evacuate the building as quickly as possible, following the standard green
and white emergency exit signs. If you require medical assistance please contact the Porters' Lodge or
registration desk.

Taxis

The Porters' Lodge can arrange taxis on request.

Mobile phones

In consideration of other conference participants and speakers, delegates are requested to switch off all
mobile phones during conference presentations.

Photography at the conference

Photography during the conference sessions is not permitted. Presentations are the property of the speakers.
Copies of presentations, subject to consent from the speaker, will be available on the BSHI website after the
conference. A photographer will be taking photographs during the conference, including at break times,
during the exhibitor reception and at the conference dinner. If you do not wish to have your photograph
taken, please speak to a member of staff on the conference registration desk.

CPD Certificates

Your feedback about this year's conference is important to the organisers to help shape the format and
content of next year's conference. Your CPD certificate will be issued on completion of an electronic feedback
questionnaire. A link will be emailed to you shortly following the conference.

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Social Programme

Trade Reception

Tuesday 13 September (16:45 - 19:00)
Ground Floor ARCO Building
Please take time to chat with our exhibitors over a glass of wine during this time. All of the exhibitors will
be on their stands and wine will be served in the exhibition.

Poster Viewing

Tuesday 13 September (16:45 - 19:00)

Douglas Price Room, Sloane Robinson Building
Alongside the Trade Reception, we will be having a poster viewing session. The posters will be on display for
the duration of the conference, but come along during these times to discuss the posters with their authors.

Drink Reception and Conference Gala Dinner

Tuesday 13 September (19:00 for 19:30) (prompt)

Liddon Quad and Dining Hall
Pre-dinner drinks will be served from 19:00 in the Liddon Quad. Dinner will be served in The Dining Hall
which is a beautiful wood panelled room with tall stained glass windows. We are grateful to Immucor for
sponsoring the pre-dinner wine.
You will be asked to take your seats for dinner at 19:30 prompt. Awards will be presented at the dinner
followed by music and dancing in the college bar (cash bar).

Dress code: We invite you to dress to impress!

Fun Run

Wednesday 14 September (07:00)

Join the BSHI 2016 Fun Run on the morning of 14th September, 2016. The run is approximately 3.5 km in
distance. It will begin at 7am departing from Keble lodge and finishing at the University Park opposite to the
conference venue, making it easy to roll out of bed and get back to the conference. The Fun Run coordinator
will guide you through the centre of Oxford, taking in many of Oxford’s historic sites including Christchurch
College, Deadman’s Walk, Radcliffe Camera and Bodleian Library. Check out the website for full details
including a map of the route and pictures of the sights.

Please bring your running gear and join us.

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Conference Programme

Tuesday 13th September

09:00-10:30 Arrival and Registration

[Sloane Robinson Building]

10:30-10:45 Welcome Address

Dr Kay Poulton, BSHI Chair, Central Manchester University Hospitals NHS Foundation
Trust
Professor Susan Fuggle, Chair of Local Organising Committee, Oxford Transplant Centre

[Auditorium]

10:45-12:15 Plenary Session 1 - Transplantation

Chairs: Prof Steven Marsh and Luke Foster

10:45 Professor Paul Moss, University of Birmingham
“The Immunology of Haemopoietic Stem Cell Transplantation - the Major
Determinant of Clinical Outcome”

11:15 Dr John Girdlestone, NHSBT Stem Cells and Immunotherapies Department, Oxford
“Immunotherapy with Mesenchymal Stromal Cells”

11.45 Professor Paul Johnson, University of Oxford
“Pancreatic Islet Transplantation - Current Status and Future Opportunities”

12:15-13:45 Lunch including Trade Exhibition

[Arco Room]

12:45-13:15 AHCS Equivalence Process – 12:45-13:05 MC Diagnostics Workshop
Amanda Robson, BSHI TDE Chair [Seminar Room 2]

[Music Room]

13:15-13:35 BSHI Trainee Forum – Anouska 13:15-13:35 VH Bio Workshop
Casanova, BSHI Trainee Rep [Seminar Room 1]

[Music Room]

13:45-15:00 Best Abstract Presentations

[Auditorium]

Chairs: Dr Brendan Clark and Mian Chen

13:45 “The Influence of HLA Antigen Matching on Graft Survival in a UK Deceased Donor
Kidney Transplant Cohort From 2009-2014”
David Turner, NHSBT Kidney Allocation HLA Working Group

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14:00 “Evidence to Support Transplant Nephrectomy Fails to Limit Sensitisation”
Anthony Poles, NHSBT Bristol

14:15 “Critical Review of Unacceptable Donor Antigens (UDAs) to Facilitate
Transplantation”
Anthony Poles, NHSBT Bristol

14:30 “Residue 156 in the Heavy Chain of HLA-B*35 Variants Determines HCMV Immune
Specificity Through Distinct Viral Peptide Selection”
Christina Bade-Doeding, Hannover Medical School, Germany

14:45 “Incidence of Graft Versus Host Disease after Multivisceral Transplantation and The Impact
of Longitudinal Monitoring of Donor Peripheral Blood Chimerism on Therapeutic
Interventions: A Single UK Centre Experience”
Sarah Peacock, Cambridge University Hospitals NHS Foundation Trust

15:00-15:30 Plenary Session 2 - National Transplantation Data

Chair: Dr Deborah Sage
Rachel Johnson, Clinical Directorate, NHS Blood and Transplant
“The Latest Trends in Organ Donation and Kidney Transplantation in the UK”

15:30-16:00 Coffee and Trade Exhibition

[Arco Room]

16:00-16:45 Festenstein Lecture

Chair: Dr Andrea Harmer

Professor Kathryn Wood, University of Oxford
“Immune Regulation in Transplantation”

16:45-19:00 Poster Viewing and Judging Event

19:00 [Douglas Price Room, Sloan Robinson Building]
19:30 Trade Exhibition with wine
[Arco Room]
Pre-dinner Drinks - Sponsored by Immucor
[Liddon Quad, Keble College]
Conference Dinner
[Dining Hall, Keble College]
Followed by music and dancing in the College Bar

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Wednesday 14th September

09:00-10:30 Plenary Session 3 - Immunogenetics
[Auditorium]

Chairs: Prof David Briggs and Dr Vivien Hanson
09:00 Professor Tim Elliot, University of Southampton

“A Combined Computational Modelling, Cellular Biochemistry and Structural Biology
Approach to Investigate Peptide Selector Function of MHC I Molecules”

09:30 Dr Robert Vaughan, Viapath, Guy’s Hospital
“Are Higher Resolution HLA Typing and Antibody Analyses Worthwhile for Solid
Organ Transplantation?”

10:00 Professor John Forsythe Associate Medical Director, NHSBT
Presidential Address

10:30-11:00 Coffee and Trade Exhibition
[Arco Room]

11:00-11:50 One Lambda Young Scientist of the Year Award Presentations

Chairs: Dr Kay Poulton and Dr David Turner
11:00 “Post-engraftment Monitoring of JMML”

Simon Mathers, Central Manchester Hospitals NHS Foundation

11:10 “A Novel Pipeline for The Automation of Single Molecule Real Time (SMRT®) for Allelic
Level Resolution of HLA-Class I and Class II Genes”
David Lapore, Anthony Nolan Research Institute

11:20 “A Retrospective Analysis of HLA Sensitisation in Patients Requiring Renal Allograft
Nephrectomy”
Ailish Nemmo, NHS Lothian

11:30 “Single Molecule Real-Time (SMRT®) Sequencing of Full-Length HLA-DRB1, - DDQ1
and -DPB1”
Thomas Turner, Anthony Nolan Research Institute and UCL Cancer Institute

11:40 “Generation of Full-length KIR2DL1/2/3 Gene Sequences Using Single Molecule Real-
Time (SMRT®) DNA Sequencing”
Will Bulltitude, Anthony Nolan Research Institute and UCL Cancer Institute

11.50-13:00 BSHI Annual General Meeting
[Auditorium]

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13:00-14:30 Lunch including Trade Exhibition and Exhibitor Presentations
[Arco Room]

13:30-13:50 Illumina Workshop
[Seminar Room 1]

13:50-14:30 Omixon Workshop 13:50 - 14:30 Gendx Workshop
[Seminar Room 2] [Music Room]

14:30-14:40 Presentation of the One Lambda Young Scientist of the Year Award
[Auditorium]
Dr Kay Poulton, BSHI Chair, Central Manchester University Hospitals NHS Foundation
Trust

Chairs: Natalia Diaz Burlinson and Debbie Pritchard

14:40 Julia Bodmer Award presentation

“Kidney Transplantation - the Catalan Way” Pamela
Hughes - St James’s University Hospital, Leeds

14:50 “Improving Access for Long Waiting Patients in a Future UK Kidney Allocation Scheme”
Lisa Bradbury, NHSBT Kidney Allocation HLA Working Group

15:00 “Improvement in the Efficiency of National Kidney Allocation - A Case for
Inclusion of Further HLA Loci”
Susan Fuggle, NHSBT Kidney Allocation HLA Working Group

15:10 “A Single Centre Analysis of Factors Associated with Early AMR in HLAi Kidney
Transplantation”
David Turner, SNBTS

15:20 “Transplanting Across a HLA-DPB1 Donor-Directed Antibody in Urgent HPCT” Alison
Logan, Transplantation Laboratory, Manchester Royal Infirmary

15:30-16:15 Terasaki Lecture

16:15-16:30 Sponsored by VH Bio Ltd

Chair: Prof Susan Fuggle

Prof Bob Montgomery, Professor of Surgery and Director, NYU Langone
Transplant Institute, USA
“Assessment and Management of Post-Transplant Donor Specific Antibodies”

Closing Session

Dr Kay Poulton, BSHI Chair, Central Manchester University Hospitals NHS Foundation
Trust

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Speakers Biographies and Abstracts

Plenary Session 1: Transplantation

Professor Paul Moss

University of Birmingham, Birmingham, UK

Professor Paul Moss is Professor of Haematology and Director of Research in the College
of Medicine at the University of Birmingham.He is also Chair of the Infections and
Immunity Board and member of Strategy Board at the UK Medical Research Council.
Professor Moss’s research programme is within the immunology of human disease,
where he undertakes a range of translational studies within transplant and cancer
immunology.

Professor Moss played a leading role in the development of HLA-peptide tetramers and
used these to monitor antigen-specific immune responses in the post-transplant setting.
Moreover he went on to use tetramers as a novel form of cell therapy for patients with viral infection in the
post-transplant setting. His team investigate a range of issues in relation to haemopoietic stem cell
transplantation including immune reconstitution, tolerance and cell therapy.
Tuesday 13th September - 10:45 [Auditorium]

The Immunology of Haemopoietic Stem Cell Transplantation - the Major
Determinant of Clinical Outcome

Haemopoietic stem cell transplantation (HSCT)is used widely in the management of many disorders of the
blood system. Substantial progress has been achieved over the last 40 years with increasing donor availability
and improved management of both graft versus host disease and infection. How ever, despite these advances
there are many areas in which substantial progress is required.

HSCT is essentially an immunological procedure which requires the replacement of the immune and
haemopoietic systems of one person by another. Somewhat surprisingly, studies of immune function in the
post-transplant period have been somewhat neglected but it is now clear that this is the major determinant
of clinical outcome. Indeed the major causes of transplant failure are disease relapse, infection and GVHD,
and all of these have a strong immune basis.

The strength and specificity of the alloreactive T cell immune response remains the key factor in determining
transplant outcome and the mechanisms by which this is optimized within transplant protocols is a key
question in protocol design. The role of the innate immune system, including NK and NKT cells, is also gaining
interest and HSCT is now the setting for some of the most interesting cell therapy studies in human medicine.

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In this talk I will discuss the regeneration of the immune system after HSCT and explain how novel
technologies are helping to uncover how the alloreactive immune response develops and is mediated. The
relative importance of homeostatic proliferation and antigenic specificity will be contrasted and the potential
role of alloreactive NK cells will be explained. The mechanisms by which immunological tolerance is
established after transplantation remain uncertain and the talk will explain how different models of
immunological chimerism after transplant can be used to investigate this. Finally, the development and role
of cellular therapy will be covered.
HSCT is one of the most fascinating areas of medicine and one in which the immune system holds the key
to clinical outcome. As advances are made in our understanding of how this is mediated we can be confident
that the clinical outcome for patients will improve substantially in future years.

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Dr John Girdlestone

NHSBT Stem Cells and Immunotherapies Department, John Radcliffe Hospital, Oxford, UK

Dr John Girdlestone obtained a BSc in Biochemistry and a PhD in Developmental Biology, and then joined
Cesar Milstein at the MRC Laboratory of Molecular Biology in Cambridge for post- doctoral work in molecular
immunology. He stayed on at the LMB as a staff scientist investigating the regulation of HLA Class I genes
by Interferons, then continued this work and related projects as an MRC Senior Non-clinical Fellow at the
Centre for Clinical Research in Immunology and Signalling at the University of Birmingham Medical School.
Moving on to biotechnology start-ups, he was Senior Scientist at Gendaq and then Chief Scientist at Plasticell,
developing methods for high-throughput differentiation of stem cells. Looking to contribute more directly
to clinical research and translation, John joined the H&I Research group at NHS Blood and Transplant
Colindale in 2005 as senior research fellow and programme manager, and in 2015 become Head of Lab for
the NHSBT Stem Cells and Immunotherapies Department at the John Radcliffe Hospital, Oxford.

Tuesday 13th September – 11:15 [Auditorium]

Immunotherapy with Mesenchymal Stromal Cells

Mesenchymal stromal cells (MSC) are connective tissue progenitors defined by their ability to differentiate
into bone, fat and cartilage. However, they can also produce stromal elements of bone marrow and lymphoid
tissues that provide trophic support for immune cells. Originally envisioned as a cellular therapy for
regenerating damaged cartilage and bone, it is their intriguing immunoregulatory properties that are being
tested in the majority of clinical trials involving MSC. The main concern over using off-the-shelf allogeneic
cell products is rejection by the host but MSC, while not being totally invisible to the immune system, are
poorly immunogenic and can actually suppress T cell proliferation as well as the activities of other immune
cell types. One of the main therapeutic uses of MSC has been in the treatment of stem cell transplant
recipients who have developed steroid-resistant GvHD, but they are also being tested for efficacy against
autoimmune diseases such as MS and Crohn’s, and in solid organ transplantation.

Although thousands of patients have been treated with MSC to date with no major adverse events, relatively
few have been enrolled in gold standard randomised, controlled trials. Therefore, despite much interest
and anecdotal reports of positive results, robust evidence for efficacy remains to be demonstrated. Due to
the difficulties and expense of running clinical trials there are basic questions that have not been addressed,
such as the dosing regimens that might provide optimal results. Although MSC are relatively easy cells to
expand, and the doses used to date are only about 1x10^6/kg, the costs associated with generating sufficient
numbers of clinical-grade cells are not trivial. In studying the immunoregulatory mechanisms exerted by
MSC, with the aim of increasing their potency and thereby reducing the cell numbers required, we found
that brief pre-treatments with Rapamycin (Sirolimus), Tacrolimus, or Cyclosporin A made them up to 5-fold
more effective at inhibiting the proliferation of T cells. Rather than modifying the physiology of the cells,
we concluded that the drugs were being adsorbed by the cells such that they were acting as delivery vectors.

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The phenomenon is not restricted to MSC and all other cell types tested, including antigen presenting cells,
were also made immunosuppressive by brief exposures to the drugs. Importantly, we found that relatively
low numbers of Rapamycin-treated MSC were superior to higher doses of untreated cells or drug alone in
suppressing GvHD in a pre-clinical model. A strong synergism was evident with MSC + Rapamycin in this
system, suggesting that they are mediating suppression through different mechanisms, and that homing to
sites of inflammation and release of drug may account for the effective inhibition of disease with relatively
low cell numbers.
By enhancing the immunosuppressive properties of MSC with established drugs we believe there is the
potential to achieve clinical end-points that are not reached with current dosing regimens, and to reduce
the costs of production while minimising undesired MSC or drug effects.

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Professor Paul Johnson

University of Oxford, Oxford, UK

Professor Paul Johnson is Professor of Paediatric Surgery at the University of Oxford,
Director of the Oxford Pancreatic Islet Transplant Programme, and Director of the
DRWF Human Islet Isolation Facility. He is also Academic Training Programme
Director for the Oxford University Clinical Academic Graduate School and a Fellow
of St Edmund Hall.

He leads an active research group encompassing basic, translational, and clinical
research, with the ultimate goal of applying beta-cell replacement to children. His
basic research specifically focuses on: 1) understanding the molecular structure and
interactions of the pancreatic matrix (with special emphasis on optimising islet
isolation and islet function), and 2) developing strategies for islet immunoisolation
(islet macroencapsulation and islet pre-conditioning). His Research Group have
major funding from a number of funding bodies including the European Commission,
DRWF, NIHR and are the UK JDRF Centre for Islet Provision.

Paul is a Past-President of the International Pancreas and Islet Transplant Association (IPITA). He is Chair of
the NHSBT UK Islet Transplant Steering Group. He is a member of the Executive Committee of the British
Association of Paediatric Surgeons (BAPS) and Past-Chair of the BAPS Research Committee. He sits / has sat
on a number of Research Advisory Boards, including those of DRWF UK, DRWF Sweden, the Healing
Foundation, the Academic and Research Board of Royal College of Surgeons of England, and the Scientific and
Strategic Advisory Board of the National Institute for Cellular Biotechnology in Ireland. In 2010, he was
awarded an Honorary Fellowship of the American Academy of Paediatrics.

Tuesday 13th September - 11:45 [Auditorium]

Pancreatic Islet Transplantation - Current Status and Future Opportunities

Pancreatic islet transplantation is a minimally invasive procedure that has the potential to reverse type 1
diabetes mellitus. Over the past decade, the results of this treatment have improved significantly, with up
to 80% of selected patients becoming insulin-independent for at least a year post transplant, and nearly all
recipients with functioning grafts achieving restoration of hypoglycaemic unawareness. However, the ultimate
aim for this cellular treatment is to be able to treat children soon after diagnosis, something that the more
invasive whole pancreas alternative is unlikely to realise. To achieve this, there are a number of challenges
to overcome, including the ultimate challenge of being able to transplant islets without the need for
immunosuppression. This talk will provide an overview of the rationale, methods, and current results of islet
transplantation, and discuss the 'state of the art' advances that are helping to make paediatric islet
transplantation a reality.

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Plenary Session 2: National Transplantation Data

Rachel Johnson

Clinical Directorate, NHS Blood and Transplant, Bristol, UK

After gaining a Master’s degree in Medical Statistics at the University of Southampton, I joined NHS Blood
and Transplant (or UK Transplant as it was then) as a statistician in 1992. I have since spent over 20 years
working with clinicians, scientists and others to analyse the UK Transplant Registry database to provide an
evidence base for clinical practice and organ allocation.

I have specialised in the area of renal transplantation and played a key role in development and
implementation of national kidney allocation schemes, including those for paired and non-directed altruistic
living kidney donation.

In 2009 I was appointed to Head of Organ Donation and Transplantation Studies within the Statistics team,
since when I have been involved with statistical support of all areas of organ donation, retrieval and
transplantation in the UK, helping to shape national services and representing the registry at national and
international workshops and conferences. I am a member of the ODT Senior Management Team, a number
of NHSBT’s Organ Advisory Groups and various collaborative working parties. I am also on the Council of the
British Transplantation Society.

Tuesday 13th September – 15:00 [Auditorium]

The Latest Trends in Organ Donation and Kidney Transplantation in the UK

Last year (2015/16) there were more deceased donors in a year in the UK than ever before – 1364. This
represents a 69% increase over the 809 donors in 2007/8 when the Organ Donation Task Force was launched.
Alongside this, there has been a 48% increase in the number of deceased donor transplants from 2384 to
3527. In the same time period the number of patients on the organ transplant waiting lists has fallen by 15%
from 7658 to 6476.

There were 2,945 adult kidney only transplants performed in the UK in 2015/16 and of these, 1,134 (39%)
were from DBD donors, 851 (29%) were from DCD donors and 960 (32%) were from living donors. 129
paediatric patients also received a kidney transplant. There are significant differences in median waiting time
to kidney transplant across the UK, ranging from 14 months for patients listed in Cambridge to 43 months
for patients listed in Birmingham. For the UK as a whole the average is 30 months. A number of factors
contribute to geographical differences eg ethnicity of local population, sensitisation of patients on the list
and the rate at which centres decline offers of kidneys for transplant. Even for standard criteria donors there
is wide variation in offer decline rates.

Once kidneys are accepted and retrieved, time to implantation is improving year on year: median cold
ischaemia time for DBD kidneys was 16 hours in 2006/7, falling to 14 hours in 2015/16 (ranging from 11
hours to 18 hours across transplant centres). For DCD donor transplants the median has fallen from 17
hours to 13 hours over the same time period, with a similar range in centre differences. It is likely that
virtual crossmatching has made an important contribution to that improvement.

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In the last two years the number of living kidney donor transplants has fallen slightly, with a fall seen in both
ABO and HLA incompatible transplants. There have been about 100 altruistic living kidney donor transplants
each year for the last three years, and some of these have triggered altruistic donor chains by donating into
the paired donation pool. In 2015, there were 94 altruistic donors, but they enabled 127 transplants in total.
A further 90 transplants in 2015 were generated from within the paired donation pool (30 through 2-way
exchanges and 60 transplants through 3-way exchanges). The increase in activity in these living donor
‘sharing schemes’may partly explain why there have been fewer incompatible transplants in the last two
years.
Transplantation numbers have increased partly through use of organs from both deceased and living donors
that may not have been deemed suitable in past years. The reassuring thing for patients, however, is that
post transplant outcomes are as excellent as ever, with no significant outlying centres. Five year patient
survival after deceased donor kidney transplant is 88%, with 95% of living donor kidney transplant patients
surviving at least five years.

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Festenstein Lecture

Professor Kathyrn Wood

Nuffield Department of Surgical Sciences, University of Oxford, UK

Professor Kathryn Wood is Professor of Immunology in the Nuffield Department of
Surgical Sciences, University of Oxford where she runs the Transplantation Research
Immunology Group (TRIG – www.nds.ox.ac.uk/trig). Her research focuses on immune
regulation at a cellular and molecular level and interactions between the immune
system and stem cell derived tissues.

Professor Wood is a Fellow of The Academy of Medical Sciences and her research
achievements have been recognised internationally, including receiving a
TheTransplantation Society (Basic Science Award 2012), a Gold Medal awarded by The
Catalan Society of Transplantation (2011), The Maharshi Sushruta Award (2012) and the TTS Women in
Transplantation, Achievement Award (2014). Kathryn was President of The Transplantation Society (2004-
2006); was the founding Chair of the Women in Transplantation initiative (WIT – www.tts.org/women) and
Editor of Transplantation (1992 - 2014).

Tuesday 13th September – 16:00 [Auditorium]

Immune Regulation in Transplantation

Kathryn J Wood1, Andrew Bushell1, Paul Harden1, Fadi Issa1, Joanna Hester1

1 Transplantation Research Immunology Group, Nuffield Department of Surgical Sciences, University of
Oxford, UK

Immune regulation is fundamental to any immune response to ensure that it is appropriate for the perceived
threat to the host. Strategies for the induction of specific unresponsiveness to donor major and minor
histocompatibility antigens currently under investigation in the clinic take advantage of two of the major
mechanisms for the induction of tolerance to self antigens – deletion and immunoregulation/suppression.
We have demonstrated that human regulatory T cells (Treg) expanded ex vivo can protect human allografts
(skin, islets and vessels) from rejection. Treg migrate to the allograft and function in situ. Donor alloantigen
reactive Treg are more effective on a per cell basis than polyclonal Treg. Together with other leukocyte
populations, including regulatory T cells, B cells and macrophages as well as myeloid derived suppressor
cells and dendritic cells, Treg contribute to the regulation of immune responses in vivo after organ
transplantation.

The identification and characterisation of Treg that can control immune responsiveness to alloantigens
has opened up exciting opportunities for new therapies in transplantation. Phase1/2a clinical trials are in
progress – www.onestudy.org.

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Plenary Session 3: Immunogenetics

Professor Tim Elliot

University of Southampton, Southampton, UK

Professor Tim Elliot left the University of Oxford with a first in Biochemistry in 1983
and completed his PhD in cancer immunotherapy at the University of Southampton
in 1986. He did his postdoctoral training at MIT with Herman Eisen at the Center
for Cancer Research. In 1990 he returned to the University of Oxford to join the
Institute for Molecular Medicine as a Wellcome Trust Research Fellow, joining a key
group of immunologists studying antigen presentation at the molecular level: where
he continues to be a world leader with over 130 research articles on the subject. In
1993 he was appointed to a lectureship and later a Professorship at Balliol College,
University of Oxford, as a Wellcome Trust Senior Fellow in Basic Biomedical Science.

In 2000, he moved to the University of Southampton as Professor of Experimental Oncology and five years
later became Associate Dean for the Faculty of Medicine. In 2015 he stepped down from this role to take up
Directorship of the new Southampton Centre for Cancer Immunology which will open in 2017. He is interim
Pro Vice Chancellor (Research) for the University of Southampton, Deputy Director of the interdisciplinary
Southampton Institute for Life Sciences and a Fellow of the Academy of Medical Sciences. He has incorporated
discoveries in the areas of antigen processing, T cell regulation and immunodominance into the development
of new cancer immunotherapies and is the recipient of a Royal Society/Wolfson Research Merit Award.

Wednesday 14th September – 09:00 [Auditorium]

A Combined Computational Modelling, Cellular Biochemistry and Structural
Biology Approach to Investigate Peptide Selector Function of MHC I Molecules

Alistair Bailey¹³⁴ †, Neil Dalchau² †, Rachel Carter¹³, Stephen Emmott², Andrew Phillips2 †, Jörn M. Werner¹⁴
†, Tim Elliott¹³ †
1 Institute for Life Sciences, Building 85, University of Southampton, UK
2 Computational Science Laboratory, Microsoft Research, 21 Station Road, Cambridge, UK
3 Cancer Sciences Unit, Faculty of Medicine, University of Southampton, Southampton, UK
4 Centre for Biological Sciences, Faculty of Natural & Environmental Sciences, Building 85,
University of Southampton, UK
† These authors contributed equally to this work

The selection of peptides for presentation at the surface of most nucleated cells in vertebrates by major
histocompatibility complex class I molecules (MHC I) is crucial to the immune response. However, the precise
mechanisms of the rapid selection of high affinity peptides from amongst many thousands of mostly low
affinity peptides by MHC I are not well understood. To gain a better understanding of the mechanisms of
peptide selection by MHC I,

Page 25 of 84

we developed computational systems models encoding distinct mechanistic hypotheses for two MHC I
molecules HLA-B*44:02 (B*4402) and HLA-B*44:05 (B*4405). These MHC I differ by a single residue, yet they
lie at opposite ends of the spectrum in their intrinsic ability to select high affinity peptides. We fitted these
models to in vivo biochemical data to infer that a conformational intermediate of MHC I is significant for
peptide selection. We also show that peptide selector function correlates with protein plasticity using
molecular dynamics simulations, using site-directed mutagenesis to test predictions arising from the model
directly. This approach also provided evidence that the chaperone molecule tapasin increases MHC I plasticity
by a mechanism of allosteric coupling, resulting in enhanced peptide selector function.

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Dr Robert Vaughan
Guy’s Hospital, London, UK

I have had the rather grand title of Director of the Clinical Transplantation Laboratory
for more than 20 of years. My lack of ambition is illustrated by the simple fact that
I have worked at Guy's Hospital in this laboratory for 45 years. My original tutor was
the late Professor Richard Batchelor, who answered all my questions with expert
knowledge and patience. Subsequently I worked for Dr Ken Welsh who managed not
to answer any of my questions directly, so I suppose he got me thinking.

Wednesday 14th September – 09:30 [Auditorium]

Are Higher Resolution HLA Typing and Antibody Analyses Worthwhile for Solid
Organ Transplantation?

I will present data on a series of nearly 100 historic renal transplants that were described as being lost with
a diagnosis of rejection. The DNA from the donors and recipients were typed to the second field level for
the HLA-A*, B*, C*, DRB1*, DRB3/4/5*, DQA1*, DQB1*, DPA1* and DPB1* loci using standard Sanger
sequencing. The differences between the medium resolution HLA typing currently used for organ allocation
and the second field results are presented and then described in terms of the donor eplet mismatch for
these loci. Sera from each recipient were then analysed using single antigen bead technology to attempt to
describe any HLA specific antibody resulting from the rejection process in relation to the precise donor HLA
mismatch and predicted eplets.
The complexities of the results of this approach will be described; the limitations in my understanding of
the data will be outlined. I will then use this as a platform to ask some questions about our use of
sophisticated technology in relation to the everyday practice of solid organ transplantation in the UK.

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Terasaki Lecture (Sponsored by VH Bio Ltd)

Robert A. Montgomery, M.D., D.Phil., FACS, Professor of Surgery

Director, NYU Langone Transplant Institute, United States

Robert A. Montgomery, MD, DPhil, FACS, is a Professor of Surgery and the Director
of the newly formed NYU Langone Transplant Institute. Before coming to NYU he
was the inaugural recipient of the Margery K. and Thomas Pozefsky endowed
Professorship in Kidney Transplantation. He received his Doctor of Medicine with
Honor from the University of Rochester School of Medicine. He received his Doctor
of Philosophy from Balliol College, The University of Oxford, England in Molecular
Immunology. Montgomery completed his general surgical training, multi-organ
transplantation fellowship, and postdoctoral fellowship in Human Molecular
Genetics at Johns Hopkins.

Dr. Montgomery was part of the team that developed the laparoscopic procedure for live kidney donation, a
procedure that has become the standard throughout the world. He and the Hopkins team conceived the idea
of the Domino Paired Donation, the Hopkins protocol for desensitization of incompatible kidney transplant
patients, and performed the first chain of transplants started by an altruistic donor. He led the team that
performed the first 2-way domino paired donation, 3-way paired donation, 3-way domino paired donation,
4-way paired donation, 4-way domino paired donation, 5-way domino paired donation, 6-way domino paired
donation, 8-way multi-institutional domino paired donation, and co-led the first 10-way open chain. He is
credited in the 2010 Guinness Book of World Records with the most kidney transplants performed in 1 day.
He is considered a world expert on kidney transplantation for highly sensitized and ABO incompatible patients.

Dr. Montgomery is a co-principal investigator for an NIH/ITN sponsored tolerance clinical trial for
simultaneous donor bone marrow and live donor kidney transplantation. He runs multiple investigator
initiated clinical trials of novel desensitization therapies. His research interests

include mechanisms underlying the immunomodulatory effect of plasmapheresis, stem cell therapy for
highly sensitized patients, and gene and cell based therapies in transplantation.

He has received important awards and distinctions including a Fulbright Scholarship and a Thomas J. Watson
Fellowship and memberships in the Phi Beta Kappa and Alpha Omega Alpha academic honor societies. He
has been awarded multiple scholarships from The American College of Surgeons and The American Society
of Transplant Surgeons. The National Kidney Foundation of Maryland has recognized his contributions to the
field of transplantation with the Champion of Hope Award and National Kidney Registry with the Terasaki
Medical Innovation Award.

Wednesday 14th September – 15:30 [Auditorium]

Assessment and Management of Post-Transplant Donor Specific Antibodies

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Oral Presentations

Best Abstract Presentations

Solid Organ Transplantation

O1: The Influence of HLA Antigen Matching on Graft Survival in a UK Deceased Donor Kidney
Transplant Cohort From 2009-2014

David Turner¹, Chloe Brown¹, Linda Shelper¹, Lisa Bradbury¹, Susan Fuggle¹

1 2015 NHSBT Kidney Allocation HLA Working Group, UK

The 2006 National Kidney Allocation Scheme uses HLA-A, B, DRB1 to assess matching. Mismatches (MM) at
these loci are known to influence graft outcome and HLA antibody production. To inform a review of UK
kidney allocation a retrospective analysis of HLA MM has been performed in a patient cohort from 2009-2014.
All UK adult kidney DBD and DCD donor transplants between 01/04/09 and 31/03/14 were included (n=7843).
HLAi transplants were excluded. The following were analysed in relation to graft survival at 1 and 5 years: i)
individual HLA-A, B, DRB1 MM, ii) combined MM at HLA-B/C and HLA-DRB1/DQB1 and iii) grouped antigen
MM (0, 1-3, 4-6 and 7-10). Cox regression modelling adjusted for known variables in graft survival.

For single loci, 1 or 2 MM at HLA-A, B and DRB1 increased the risk of graft loss at 1 and 5 years (HLA-A HR=1.32
(1yr), 1.30 (5yrs); B=1.72, 1.54; DRB1=1.23, 1.24). HLA-C MM increased risk independently in HLA-B matched
pairs (HR=1.86, 1.55) but HLA-DQB1 MM was only significant when combined with DRB1 MM (1.36, 1.31).
As total number of MM increased the risk of graft loss at 1 and 5 years increased (1 yr: 1-3MM=1.73; 4-6=2.25;
7-10=2.32 and 5 yrs: 1-3=1.65; 4-6=1.89; 7-10=1.98). As a linear variable each additional HLA MM increased
the risk of graft loss at 1 and 5 years (1.11, 1.08).

This analysis shows the importance of consideration of HLA-C and DQB1 in future allocation algorithms.
Further analysis is ongoing to establish the influence of epitope MM.

O2: Evidence to Support Transplant Nephrectomy Fails to Limit Sensitisation

Anthony Poles¹, Brian Carey², Andrew Connor³, Imran Saif³, Peter Rowe³
1 NHSBT Bristol, UK
2 H&I Dept, Derriford Hospital NHS trust, Plymouth, UK
3 SouthWest Transplant Centre, Derriford Hospital, Plymouth, UK

British Transplant Society (BTS) guidelines recommend a reduction of immunosuppression following
nephrectomy. We hypothesised that, even when the period of exposure to graft material is very brief
sensitisation to HLA mismatched antigens occurs when

immunosuppression is withdrawn at an early stage. A retrospective audit was performed to include all
graft failures in the past 6 years that resulted in nephrectomy within 6 months of

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transplant. HLA specific antibody data was analysed prior to and post nephrectomy. Incidences of
sensitisation to mismatched HLA antigens were recorded as sensitised to failed graft Data regarding the
duration of graft survival prior to nephrectomy and the timing of immunosuppression withdrawal were also
collected. If HLA antibody data was absent, further testing was performed with bead based assays (MFI
>3000).

There was a high incidence of sensitisation (80% ; 12/15), and this occurred even when the graft was in situ
for 24hrs. The range of graft life was from <24hrs to a maximum of 172 days with a median of 48hrs. Two
patients received 000 mismatched grafts and remain non- sensitised. Eleven patients non-sensitised and one
sensitised to third party antigens prior to transplant developed HLA donor specific antibodies (DSA) post
nephrectomy. These DSAs were present up to three years post nephrectomy. Immunosuppression was ceased
at the time of nephrectomy in 12 patients and weaned over a few weeks in 3 patients.

Longer periods of immunosuppression may be warranted if re-transplantation is likely but must be
balanced against the risks of medications.

O3: Critical Review of Unacceptable Donor Antigens (UDAs) to Facilitate
Transplantation

Anthony Poles¹, Brian Carey², Andrew Connor³, Imran Saif³, Peter Rowe³

1 NHSBT Bristol, UK
2 H&I Dept, Derriford Hospital NHS trust, Plymouth, UK
3 South West Transplant Centre, Derriford Hospital, Plymouth, UK

All patients that have waited >1000 days on the UK waiting list for a deceased donor kidney in the South
West region are classed as “long waiters” (LWs).

All HLA specificity profiles were reviewed for all LWs. All UDAs < 3000 MFI were de-listed. Historic antibody
specificities not detected in the last 12 months were also considered for de- listing on a case by case basis.
Removal of an UDA that correlated with an allele specific antibody was also accepted where there was a
high likelihood of receiving an organ lacking that allele. Where possible, “third party” crossmatches were
performed to confirm that removal of an UDA would not result in a current positive flow cytometric
crossmatch. The cRF and Chance of Transplant web based tools were used to estimate efficacy of this
approach. Proposed changes in UDAs were ratified within transplant multi-disciplinary team meetings.

Within a 14 month period this approach has enabled 12 patients to receive deceased donor kidney
transplants. The first patient possessed an historic Bw6 antibody. Upon de-listing they received an organ
within a few days post removal of this UDA. The second patient possessed DRB1* 09, (matched to DR4 in
O DT algorithm), but had HLA specific antibody to DRB1*04:02 (MFI=14000). Several UDAs including DR4
were removed, they received a DRB1*04:03 positive organ. Both cases have good graft function with a
creatinine level <150µmol/L post- transplant. At this stage, all 12 grafts are functioning, with no episodes of
biopsy proven rejection.

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HLA and Disease

O4: Residue 156 in the Heavy Chain of HLA-B*35 Variants Determines HCMV Immune
Specificity through Distinct Viral Peptide Selection

Wiebke C. Abels¹, Trishna Manandhar¹, Heike Kunze-Schumacher¹, Rainer Blasczyk¹, Christina Bade-Doeding¹

1 Hannover Medical School, Institute for Transfusion Medicine, Germany

The immune evasion strategy of HCMV prevents the presentation of viral antigens; consequently only
peptide-loading-complex (PLC)-independent HLA variants are able to constitutively present viral derived
peptides. An issue in cellular therapy strategies is the emergence of antiviral T-cell anergy over time. The
crucial question is at what stage of HCMV infection viral-peptides are preferentially presented over
self-peptides. Two of those PLC- independent HLA molecules are the common variants B*35:01 and 35:08
distinguished by a single mismatch at residue 156.To analyze the competitive HLA-B*35:01 and B*35:08
restricted viral- and self-peptide repertoire, we utilized soluble HLA technology. sHLA complexes from HCMV
infected BJ/sHLA cells were affinity purified. Kinetic of immune evasions was monitored by mRNA analysis.
sHLA-B*35:01 or sHLA-B*35:08 restricted peptides were recovered and analysed using nano-LC-ESI MS/MS
technology.

We found a distinct self-peptide pattern in the presence of HCMV for B*35 variants, the features and anchor
motifs remained mainly unaltered in comparison to peptides recruited in the absence of HCMV. However,
a different subset of viral peptides from the early to late phase of infection was recruited by B*35:01 and
B*35:08. A high frequency and dominance of certain viral peptides over self-peptides was striking.

These findings explain the discrepancy between predicted and naturally presented immunogenic
epitopes for establishing anti-viral effector cells. Furthermore, peptide prediction based on available
peptide anchors as given in the databases might fail due to structural alteration of HLA variants and
support the need of comprehensive peptide recruitment data for personalized and effective cellular
therapies.

Solid Organ Transplantation

O5: Incidence of Graft Versus Host Disease after Multivisceral Transplantation and the Impact
of Longitudinal Monitoring of Donor Peripheral Blood Chimerism on Therapeutic Interventions:
a Single UK Centre Experience

Sarah Peacock¹, Lisa Sharkey¹, Andrew Butler¹

1 Cambridge University Hospitals NHS Foundation Trust

G raft-versus-host disease (G VHD) is a potentially fatal complication after multivisceral transplantation
occurring in approximately 10% of adult transplant recipients. It is caused by

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alloreactive donor lymphocytes present in the transplanted tissue that migrate into the recipient circulation.
The differential diagnosis can be difficult and optimal therapeutic options are not clear. We retrospectively
analysed patient records to determine the incidence of G VHD, peripheral blood donor lymphocyte
chimerism, treatment used and patient survival in 67 consecutive intestinal or multivisceral transplant
recipients performed between 1998 and 2016. Seven patients (10.4% ) developed G VHD at between 3 and
48 weeks; of these five had received a multivisceral transplant. Three of seven (42.8% ) G VHD patients and
21 of 60 (35% ) patients with no G VHD had died at last follow-up. Presence of donor lymphocyte chimerism
(tested using combinations of SSP, SSO and STR) was detected in all seven patients with G VHD and none of
60 patients tested without G VHD. Treatment of G VHD varied: methylprednisolone (n=1);
methylprednisolone and Campath (n=1); methylprednisolone, Campath and basiliximab (n=2);
extracorporeal photophoresis (ECP) (n=1); calcinurin inhibitor (CNI) withdrawal (n=1) and CNI withdrawal
with ECP (n=1). Quantitative longitudinal donor lymphocyte chimerism monitoring (using Powerplex 16 STR
(Promega) assay) was performed in three patients and demonstrated that CNI withdrawal corresponded
with a substantial decrease in donor lymphocyte chimerism and improving clinical symptoms. O ur results
suggest that quantitative, longitudinal monitoring of donor lymphocyte chimerism provides a useful tool
for the diagnosis of G VHD and monitoring the effectiveness of treatment.

Page 32 of 84

One Lambda and VHBio Young Scientist Award Presentations

Haemopoietic Stem Cell Transplantation

O10: Post-Engraftment Monitoring of JMML

Simon Mathers¹, Rob Wynn¹, Helena Lee¹, Alison Logan¹

1 Central Manchester Hospitals NHS Foundation Trust, Manchester, UK

A four year old boy presented with juvenile myelomonocytic leukaemia (JMML) and required a stem cell
transplant. A transplant was performed using a 5/6 cord (donor 1), and achieved 92.4% donor engraftment
but this quickly dropped due to relapse. The patient was then transplanted with an 8/10 matched unrelated
donor (donor 2). The patient maintained 100% engraftment with donor 2 over the next 6 months. After a
period of 25 months bone marrow studies were performed which were consistent with relapsed JMML
with evolving AML.

Chimerism analysis on a peripheral blood sample from this time-point confirmed a drop in donor cells from
100% to 9.5%, whilst chimerism on the CD3⁺ popula�on was s�ll 96.1%
donor. Chimerism analysis on the CD15⁺ popula�on (myeloid cell marker) demonstrated 41.8% donor and
did not reflect the peripheral blood result (as myeloid cells are the majority cell group in peripheral blood).
A decision was made to test the CD33⁺ cell popula�on, also a myeloid cell marker. The CD33 population
resulted in 6.2% donor cells, which is similar to the result observed with the PBL sample.

While both CD15 and CD33 are both myeloid cell markers, the CD15 marker is more prevalent with mature
myeloid cells. JMML affects immature monocytes known as myelomonocytes. A more appropriate cell
lineage marker may be sought and we are currently investigating CD14 as an alternative monocytic marker
for this disorder.

Methodology

O11: A Novel Pipeline for the Automation of Single Molecule Real Time (SMRT®) For Allelic
Level Resolution of HLA-Class I and Class II Genes

Davide Lepore¹, Francisco Boix¹, Franco Tavarozzi¹, Shem Wallis Jones¹, Reetinder Grewal¹, Katy Latham¹

1 Anthony Nolan Research Institute, Royal Free Hospital, London, UK

Anthony Nolan have implemented Pacific Biosciences’ SMRT® DNA Sequencing allowing
highly multiplexed full length genomic typing of HLA Class I and near full length for Class II. We developed
and implemented an end-to-end automated pipeline, integrating fully with the Laboratory Information
Management System to increase efficiency. Four Hamilton Microlab StarLine workstations were utilised for
all liquid handling stages with bespoke programming for all Pre and Post PCR stages: 1)PCR amplification
preparation, 2) preparation for amplicon sizing and quantification, 3) equinanogram pooling, 4) size selection
by AMPure® PB

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paramagnetic beads, 5)1µg input DNA libraries preparation, 6)MagBead-Complex loading. 190 samples (plus
pos and neg controls) underwent the automated end to end pipeline for -A, -B, -C, -DRB1, -DQB1 and -DPB1.
A total of 1078 out of a possible 1140 DNA sequences were generated. The 62 missing sequences (5.4%)
were due to either PCR failing or to sequencing data not reaching quality standards. This allows for 384
samples to be sequenced for HLA Class I and II in a six day period, with stages on days 1-5 automated utilising
1 FTE increasing capacity by 100% compared to a manual process (192 samples sequenced in a six days period
by a single person). A further increase in capacity is exploited by multiple pipelines run concurrently resulting
in a maximum of 1,536 samples processed in 8 days. From January- June 2016 Anthony Nolan has processed
20,330 samples for all 6 loci with no phase- or coverage-based ambiguities.

Solid Organ Transplantation

Oral 12: A Retrospective Analysis of HLA Sensitisation in Patients Requiring Renal Allograft
Nephrectomy

Ailish Nimmo¹, Sophie McIntyre¹, Lorna Henderson¹, Richard Battle¹

1 NHS Lothian, Edinburgh, UK

Background The development of HLA antibodies towards a failing renal allograft can impact upon chance
of future transplantation. We assessed the formation of HLA antibodies in patients who underwent
transplant nephrectomy at our centre over a 10 year period.

Methods We conducted a retrospective study evaluating patients with a failed transplant who underwent
graft nephrectomy from 2005-2015. Samples were tested for DSA at 5 time points: pre-nephrectomy, post-
nephrectomy, pre-immunosuppression (IS) weaning, post-IS weaning and post-IS cessation. Calculated
reaction frequency (cRF) was determined for each time point. cRF data was entered into the ODT chances
of transplant (CoT) calculator with all other demographics reflecting an average patient at our centre.

Results 24 patients (14 male, mean age 45 years) had sufficient data for analysis. Mean time from
immunosuppression weaning to nephrectomy was 376 days, and from nephrectomy to immunosuppression
cessation 166 days. One patient had no sample post-immunosuppression cessation. One patient remained
on immunosuppression throughout. 7 patients had immunosuppression stopped within 14 days of
nephrectomy. Table 1 shows cRF and chance of transplant at specified time points.

Discussion This analysis investigated changes to sensitisation and chance of future transplant after
nephrectomy and immunosuppression withdrawal. An increase in cRF following nephrectomy and stepwise
increase in cRF as immunosuppression was withdrawn was observed. Immunosuppression changes occur in
close time proximity to transplant nephrectomy which confounds this assessment however it is clear the
risks and benefits of stopping immunosuppression need to be carefully considered on an individual basis to
maximise chance of future transplant.

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Methodology

O13: Single Molecule Real-Time (SMRT®) Sequencing of Full-Length HLA-DRB1, - DQB1 and
-DPB1
Thomas R Turner¹, Cristina Guijarro², James Robinson¹, Alasdair JM McWhinnie², J Alejandro
Madrigal¹, Neema P Mayor¹, Steven GE Marsh¹
1 Anthony Nolan Research Institute and UCL Cancer Institute, London, UK
2 Anthony Nolan Research Institute, London, UK
Allelic level HLA typing facilitates optimal donor selection for haematopoetic stem cell transplantation,
resulting in improved patient survival and reduced side effects. Unambiguous, high resolution typing of HLA
class II genes (HLA-DRB1, -DQB1 and -DPB1) has been challenging due to a lack of reference sequences and
large intronic regions. Additional complexity in HLA-DRB1 arises from allele length variability and the
potential to co-amplify other HLA-DR genes. The majority of HLA class II genes are defined by, and thus
typed based on exon 2 and occasionally exon 3 sequences, ignoring vast regions of potential coding and
non-coding sequence diversity. We have developed a robust method to sequence full-length HLA class II
genes using Pacific Biosciences’ Single Molecule Real Time (SMRT) DNA sequencing. Single-fragment,
long-amplicon sequences generated in isolation allow phase and ambiguities to be resolved without in silico
assembly. To date, we have submitted 22 full- length genomic sequences of HLA-DRB1 to the IPD-IMGT/HLA
Database, adding to the 27 sequences that had been generated since 1997. These include 12 extensions of
known alleles and 10 novel intronic variants of known alleles. Currently, work is on-going to adapt this
strategy for HLA-DQB1 and -DPB1. As we sequence more full-length HLA class II alleles from samples of
diverse ethnicities, we anticipate the discovery of novel polymorphisms and previously unidentified patterns
of linkage disequilibrium. We will show the ability of SMRT DNA sequencing to provide full-length HLA class
II sequences, revealing novel polymorphisms with potential clinical significance or evolutionary interest.

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O14: Generation of Full-Length KIR2DL1/2/3 Gene Sequences Using Single Molecule
Real-Time (SMRT®) DNA Sequencing
Will P Bultitude¹, Cristina Guijarro², Neema P Mayor¹, J Alejandro Madrigal¹, Steven GE Marsh¹
1 Anthony Nolan Research Institute and University College London, London, UK
2 Anthony Nolan Research Institute, London, UK
The KIR genes are a highly polymorphic family of genes involved in regulating natural killer (NK) cells. There
are several well-established protocols for determining the presence of individual KIR genes and numerous
publications demonstrating associations between specific KIR gene content and a wide range of diseases
and haematopoietic stem cell transplantation (HSCT) outcomes. It has also been established that cell surface
KIR expression, a factor widely accepted to be important in NK cell function, is largely dependent on allelic
polymorphism, although this aspect is still relatively poorly understood. A subset of the KIR molecules,
including KIR2DL1, KIR2DL2 and KIR2DL3, have known ligands and are marginally better studied. Herein,
we describe a SMRT long-amplicon sequencing approach to generate accurate sequences of the entire
KIR2DL1, KIR2DL2 and KIR2DL3 genes. To validate the technique, WT47, an IHIW cell line with previously
determined full haplotype DNA sequence, was sequenced in duplicate for KIR2DL1 and KIR2DL2. Specific,
highly accurate (mean SNR = 5.78) consensus sequences were generated with average read depth of 80
reads. Although our results show remarkably similar sequences to those previously published (30067/30083
homologous nucleotides across KIR2DL1 and KIR2DL2, >99.9% homology), we have detected insertion,
deletion and substitution variations at several intronic positions. Subsequent Sanger sequencing at four of
these discrepant sites (4/16) suggests that the sequences generated by SMRT are correct. We hope that
the ability to accurately sequence KIR alleles will allow us to better understand the function of KIR molecules
and their effect on disease and HSCT outcomes.

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Julia Bodmer Award Presentation

To commemorate the life and work of Julia Bodmer and to recognise the seminal contributions she made to
the field of human histocompatibility and immunogenetics, each year BSHI makes available an award to
promote the interaction and exchange of ideas between laboratories, either in the UK or overseas.
The award is aimed at scientists who are at an early stage in their career working in H&I and have been
members of BSHI for over three years. It is intended to facilitate visits of at least two weeks duration to
another laboratory to carry out a specific piece of research or to transfer new technologies.
The successful applicant is expected to write a report of the exchange visit to be published in the BSHI
Newsletter and to give a short presentation at the BSHI annual conference.
This year’s successful applicant for the award was Pamlea Hughes from the Transplant Immunology
Department at St James's University Hospital, Leeds. An overview of the project is given below and we look
forward to hearing Pamela’s report from her trip to Barcelona at the conference.

Kidney Transplantation - The Catalan Way
Pamela Hughes

Transplant Immunology, St James’s University Hospital, Leeds, UK
Catalonia has a large and active renal transplant programme delivered by 8 centres all of which are served
by the H&I laboratory at Hospital Clinic Barcelona. The Barcelona lab was involved in 954 solid organ
transplants in 2015, of which 647 of them were renal transplants.
During the Barcelona laboratory visit, I gained an insight into the running of day to day laboratory case
management as well as their out of hours work and I hope that this will provide some new ideas for Leeds.
Whilst the goals and aims of both services in obtaining safe transplants for their patients are the same, the
way in which this is approached is distinct in the terms of logistics and practice.
My intention is to highlight the differences in approach between the centres in regards to certain aspects
of service delivery and evaluate these in terms of potential service improvement in both the centres.

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Oral Presentations

Solid Organ Transplantation

O6: Improving Access for Long Waiting Patients in a Future UK Kidney Allocation Scheme

Lisa Bradbury¹, Chloe Brown¹, David Turner¹, Susan Fuggle¹

1 2015 NHSBT Kidney Allocation HLA Working Group, UK

One of the objectives of the 2006 National Kidney Allocation Scheme (NKAS) was to improve access to
transplantation for long waiting patients on the transplant list. Within the first year there was a reduction in
long waiting patients from 15% to 10% although it has remained unchanged ever since. To improve access
to transplant a change to NKAS was implemented in September 2014 giving absolute priority for patients
waiting ≥7 years. Since this change, the percentage of long waiting patients has further reduced to 8%.
However, these patients wait more than double the average waiting time before receiving such priority.

An analysis of 9,131 patients registered for a first kidney transplant between 2006 and 2010 showed those
patients with a calculated Reaction Frequency (cRF) of 40% or higher wait exponentially longer for transplant
and that very difficult to match patients wait > 7 years to transplant. A subset analysis of patients with a cRF
of 100% showed that 58% of those still waiting had been blood group and HLA compatible with at least one
donor, but only 25% of these patients had received an offer due to current prioritisation within the scheme.

Currently level 4 mismatched kidneys [2DR or 2B, 1DR] are not allocated nationally and this restricts access
for highly sensitised patients. Consideration should be given to removal of these criteria. Furthermore the
cRF and time from listing at which patients receive priority, together with the scale of priority needs further
consideration.

O7: Improvement in the Efficiency of National Kidney Allocation - A Case for Inclusion of
Further HLA Loci

Susan Fuggle¹, Linda Shelper¹, Lisa Bradbury¹, Chloe Brown¹, David Turner¹

1 2015 NHSBT Kidney Allocation HLA Working Group, UK

The 2006 National Kidney Allocation Scheme (NKAS) includes features to promote equity of access to
transplant for patients nationally and kidneys are allocated to named patients. The nationally agreed
‘Minimum Resolution for Donor HLA Typing’ was introduced with the 2006 NKAS to promote efficient organ
allocation. Patients sensitised to HLA are often difficult to transplant and highly sensitised patients are
compatible with a restricted donor pool. The allocation scheme prioritises highly sensitised patients for an
antibody compatible kidney. However there is a risk that the crossmatch may be positive, resulting in
reallocation of the organ.

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An audit of the reasons for a positive crossmatch has been performed to inform discussions about kidney
allocation.

Kidneys allocated over a six year period, 2010-2015, were included (n=6300) and 150/6300 (2.4%) kidneys
were reallocated because of a positive crossmatch. The percentage of kidneys reallocated because of a
positive crossmatch has decreased over time from 3.7% in 2010 to 1.4% in 2015. The reasons for positive
crossmatches are routinely explored and the audit showed that 36% of positive crossmatches were caused
by antibodies to specificities not currently included in the ‘Minimum Resolution for Donor HLA Typing’,
HLA-DP (30%), HLA- DQA (5%) and DR alleles (1%).

A positive crossmatch is only one of the reasons for reallocation of kidneys, and the incidence has decreased
over the last six years. However, especially in the era of virtual crossmatching, the efficiency of kidney
allocation could be improved if the UK kidney allocation algorithm included HLA-DP, DQA and certain alleles.

O8: A Single Centre Analysis of Factors Associated With Early AMR in HLAi Kidney
Transplantation

David Turner¹, Richard Battle¹, Jennifer McCaughan¹, Lorna Henderson²

1 Scottish National Blood Transfusion Service, UK
2 Renal Unit, Royal Infirmary of Edinburgh, UK

HLA antibody incompatible (HLAi) kidney transplantation is undertaken in many UK centres. Patients with
donor specific antibody (DSA) are at increased risk of antibody mediated rejection (AMR) and graft failure.
This single centre retrospective study analysed factors associated with early AMR in 25 HLAi transplants
performed between 04/04/12 and 10/02/16.

HLA antibodies were identified by IgG single antigen bead assays (One Lambda). AMR was defined as
histological evidence of microcirculation inflammation or arteritis with or without C4d staining and the
presence of circulating DSA. Cumulative MFI (cMFI), FC-XM, DSA class, current vs historic DSA, desensitisation
and immunosuppression were investigated in relation to AMR.

We included HLAi transplants with current (n=18) or historic Luminex defined (n=7) DSA. 12/25 patients were
diagnosed with AMR. All patients were within 6 weeks of transplant. Nine patients had HLA class I, 10 class
II and 6 class I and II DSA. 11/25 patients were desensitised with plasma exchange and intravenous
immunoglobulin. 14/25 patients received a lymphocyte depleting agent at induction. Mean cMFI was 6834
and 5830 in patients with and without AMR (p=ns). A current FC-XM was positive in 87.5% patients with AMR
compared to 33.3% without AMR (p=0.024). All other factors were non-significant in relation to AMR.

In this small single centre retrospective analysis of CDC negative HLAi transplants, only a positive allo FC-XM
was associated with an increased risk of AMR. This analysis has helped to define the future policy in our
centre for the acceptance of risk in HLAi transplantation.

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Haemopoietic Stem Cell Transplantation

O9: Transplanting Across a HLA-DPB1 Donor-Directed Antibody in Urgent HPCT
Alison Logan, Helena Lee¹, Amanda Robson¹, Ben Adams¹, Eleni Tholouli², Muhammad Saif², Kay Poulton¹
1 Transplantation Laboratory, Manchester Royal Infirmary, Manchester, UK
2 Department of Haematology, Manchester Royal Infirmary, Manchester, UK
A 61 year old CMV positive male with MDS was worked up for RIC HPCT. Two male CMV positive 10/10
MUDs were identified. Both donors were HLA-DPB1*06:01 mismatched with the patient (HvG direction).
At the time of donor work-up it was not laboratory policy to screen HPCT patients for HLA antibodies unless
mismatched with a donor at HLA-A, -B, -C , - DRB1 or -DQB1.
HLA antibody screening for blood product support (sample dated 07/03/2016) indicated the patient was
positive for IgG and IgM antibodies directed towards Class I and II (One Lambda LABScreen® Mixed) with a
calculated reaction frequency of 100%. Subsequently LABScreen® Single Antigen screening identified multiple
HLA antibody specificities; notably an IgG donor- directed HLA-DPB1*06:01 specific antibody (MFI 26437).
Subsequent HLA antibody screening on 15/03/2016 showed the patient’s antibody profile had not changed
significantly.
Clinical urgency and the lack of HLA-DPB1 information on registry donors meant there was insufficient time
to identify alternative donors. The decision was made to transplant the patient and monitor the donor-
directed HLA-DPB1 antibody level in conjunction with chimaerism monitoring. The patient was transplanted
on 16/03/2016. HLA antibody monitoring showed circulating HLA-DPB1 antibody decreased post-transplant:
HLA- DPB1*06:01 MFI results from 17/03/2016, 22/03/2016 and 25/04/2016 were 26119, 21017 and 7850
respectively.
By 30/03/2016 the donor had engrafted successfully (neutrophil count 0.62 x 10⁹/l). Chimaerism monitoring
of PBL, CD3 and CD15 cell lineages showed 100% donor engraftment on 13/04/2016, 27/04/2016 and
19/05/2016. The patient is currently well post-transplant with no indication of rejection.

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Poster Presentations

Antibody Detection

P1: A Biotin-Streptavidin Complex (BSC) Modified Single Antigen Bead Assay (SAB) Does Not
Appear To Eliminate Type II Errors in the Identification of HLA-Specific Antibody
Petra Goldsmith¹, David Lowe¹, Matthew Howse¹, Dan Ridgway¹, Abdul Hammad¹, Derek Middleton¹
1 Royal Liverpool Hospital, Liverpool, UK
Introduction The utility of SAB assays in the detection of HLA-specific antibody in renal transplantation
has facilitated virtual crossmatching and improved diagnosis of rejection. However, these assays yield
false negative results because of the prozone effect.
Various techniques aim to reduce this effect, including serial dilutions, EDTA treatment and a modified BSC
assay. Such assays are expensive and there is no consensus on their use in clinical practice. We present a
pilot study examining the different assay modifications.
Methods Sera from 23 patients were tested neat, at 1:10 dilution, using 0.3% EDTA and with BSC, with One
Lambda SABs on the Luminex platform. Fourteen patients were tested for class I, 4 for class II and 5 for both
classes. Mean fluorescence intensity (MFI) data were collated for 3,277 beads.
Results Linear regression compares MFIs between neat and EDTA, neat and BSC and EDTA and BSC
treatment (Figure 1).

MFIs seen with EDTA treatment have some association with neat serum, but a cluster of false negatives is
seen (graph 1a, top left) with high MFIs on EDTA testing but low values on neat testing. BSC MFIs more
closely associate with neat serum indicating similar inhibition of the BSC modification (bottom right cluster,
graph 1c).
Conclusion In this cohort, we have shown that EDTA treatment can help to eliminate false negative results
in SAB testing, whereas the BSC modification shows only a modest improvement in type 2 errors.

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P2: Relationship between the Complement Dependent Cytotoxicity (CDC), Luminex Single
Antigen Bead (SAB) and Luminex C1q Assays
B Rana¹, C Darke¹, M T Rees¹, F Edwards¹, E Burrows¹

1 Welsh Transplantation and Immunogentics Laboratory, Cardiff, UK

The CDC assay detects complement fixing HLA antibodies and CDC-defined DSAs are considered a
contraindication to renal transplantation. In contrast, the highly sensitive SAB detects both complement
fixing and non-complement fixing antibodies but their clinical significance is controversial. The C1q assay
detects antibodies that bind C1q. We have examined the relationship between these three tests.
111 samples from renal patients were tested by CDC using B-cells (with/without DTT), also Class I and II
SAB (with serum EDTA pre-treatment), and the C1q assay (One Lambda).
Overall, 38 IgG specificities were detected by CDC (covering HLA-A,B,C,DR,DQ), and 1,671 by SAB and 447
(26.8% of SAB specificities) by the C1q assay (both covering HLA- A,B,C,DR,DR51/2/3,DQ,DP).
Interestingly, 44% (183/412) of DQ specificities bound C1q while only 6% (5/88) of HLA-C specificities
were C1q assay positive.
Seven IgM specificities were identified by CDC. None were SAB positive but two were C1q assay positive.
The C1q assay failed to detect 26% (10/38) of the CDC-defined IgG antibodies.
82% (414/503) of the SAB specificities with a MFI>10,000 were C1q binding whereas, only 3% (33/1,168) of
SAB specificities with an MFI<10,000 were C1q binding.
Notably, 93% (26/28) of the IgG specificities detected by CDC and the C1q assay had MFI values >10,000
by SAB.
Thus, an important association was found between high Luminex SAB MFIs and the ability to bind C1q.
Overall, a SAB MFI of >10,000 for a given HLA specificity had a positive predictive value of 93% for that
specificity being positive in the C1q assay.

P3: Relationship between HLA Donor Specific Antibodies (DSAs) Detected by Complement
Dependent Cytotoxicity (CDC), Luminex Single Antigen Bead (SAB) and C1q Assays, and
Antibody Mediated Rejection (AMR) and C4d Deposition, in Renal Transplants
B Rana¹, C Darke¹, M T Rees¹, F Edwards¹, E Burrows¹

1 Welsh Transplantation and Immunogenetics Laboratory, Cardiff, UK

To evaluate the clinical significance of DSAs detected by CDC, SAB and C1q assays we examined the
relationship between DSAs and biopsy findings, i.e. AMR and C4d staining. Samples from 37 renal recipients,
taken close to biopsy, because of likely AMR, were tested

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by CDC using B-cells (with/without DTT), also Class I and II SAB (with serum EDTA pre- treatment),
and the C1q assay (One Lambda).

CDC detected DSAs (Class II) in only one patient with biopsy proven AMR and C4d deposition.
In contrast, SAB and C1q assays detected 55 (14 Class I, 41 Class II) and 16 (2 Class I, 14 Class II) DSA
specificities, respectively (both covering HLA-A, -B, -C, -DR, -DR51/2/3,-DQ,-DP). Overall, HLA-DQ specificities
predominated: 23/55(42%) by SAB; 13/16(81%) by C1q.
C4d deposition was significantly associated with biopsy proven AMR (p value <0.001). Eleven patients had
biopsy proven AMR and C4d deposition. 82% (9/11) had DSA by the SAB assay but only 45% (5/11) had C1q
binding DSA.
Conversely, in the AMR and C4d negative group of 20 patients, 40% (8/20) had DSAs by the SAB assay
and only 1 patient (1/20) had C1q binding DSAs.
The C1q assay showed moderate sensitivity (46% and 54%) but high specificity (85% and 90%) for AMR
and C4d deposition, respectively. In contrast, the SAB assay was more sensitive (79% and 86%) but less
specific (52% and 57%) for both AMR and C4d deposition.
Importantly, no significant association was observed between DSA detected by the CDC, Luminex
SAB and/or C1q assay and biopsy findings.

P4: Variability of Luminex Methods: Findings from a Survey of UK NEQAS for H&I Participants
Felicity May¹, Deborah Singleton¹, Tracey Rees¹, Christopher Darke¹

1 UK NEQ AS for H&I, Cardiff, UK

We requested detailed information on Luminex testing procedures from the 80 participants of our ‘HLA
Antibody Specificity Analysis’ scheme (Scheme 3). 95.0% (n=76) of participants responded.
10 different Luminex kits were used: 75.0% (n=57) used LABScreen kits only, 15.8% (n=12) Lifecodes kits
only and 9.2% (n=7) used both.
Bead volumes ranged from 1.5µl to 5µl for LABScreen and 2.5µl to 40µl for Lifecodes; serum volumes from
8µl to 20µl for LABScreen, 5µl to 20µl for Lifecodes.
72.4% (n=55) of participants used the manufacturer’s negative control, 9.2% (n=7) used ‘in house’
negative, 2.6% (n=2) used no negative control, 15.8% (n=12) did not specify.
81.6% (n=62) used a positive control of which 69.4% (n=43) used patients’ sera and 85.5% (n=53) included
this control in every run.
96.1% (n=73) stored samples before testing, including: freezing 82.2% (n=60), refrigeration 16.4% (n=12)
and ambient 1.4% (n=1). 72.6% (n=53) routinely stored samples for >48 hours before testing.

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79.0% (n=60) used serum treatment, 7.9% (n=6) did not, 13.2% (n=10) did not specify. Methods used were
(some applied several methods): Adsorption beads 76.7% (n=46), EDTA 63.3% (n=38), heat inactivation
8.3% (n=5), dilution 6.7% (n=4), and DTT 1.7% (n=1).
All but two laboratories followed the manufacturer’s incubation times and used the supplied wash buffer.
65.8% (n=50) removed buffer using a vacuum manifold, 30.3% (n=23) used ‘centrifuge, flick and blot’, 1.3%
(n=1) used LabXpress, 2.6% (n=2) did not specify.
For data analysis: 69.7% (n=53) used ‘HLA Fusion’, 17.1% (n=13) MatchIT, 7.9% (n=6) used both, 5.3%
(n=4) used ‘HLA Fusion’ with other software.
Minimum ‘positive control’ bead MFIs varied from 400 to 10000 while criteria for ‘high background’
ranged from 300 to 1500 MFI.

These findings indicate disturbing variations in Luminex practices.

Haemopoietic Stem Cell Transplantation

P5: HLA Typing of an Acute Myeloid Leukaemia Patient with UPD Loss of Heterozygosity
Jessica McCappin¹, Brian McIlhatton¹, Michael McDermott¹, Susan Piggott², Damian Finnegan², Jeanie Martin¹
1 H& I Laboratory, Belfast City Hospital, Belfast, UK
2 Haematology Department, Belfast City Hospital, Belfast, UK

A patient newly diagnosed with acute myeloid leukaemia (AML) was human leukocyte antigen (HLA) typed
with a view to proceeding to a HLA matched unrelated donor (MUD) haematopoietic stem cell transplant
(HSCT). The patients initial molecular type using DNA extracted from an EDTA blood sample was HLA-A*01,-,
B*08, -, C*07, -, DRB1*03, -, and DQB1*02, -. As per EFI guidelines, a verification sample was obtained 6
weeks and the HLA type showed the patient to be heterozygous for HLA-A*01, *02. This was subsequently
confirmed from a saliva sample. Further investigations showed that the initial blood sample for HLA typing
was taken at the time of diagnosis when blast cell count was high and the verification blood sample was
taken during remission. This lead to the hypothesis of a possible loss of heterozygosity (LOH) resulting from
acquired uniparental disomy (aUPD). To determine if LOH or allelic deletion had occurred within the HLA
region comparative genomic hybridisation (CGH) was carried out. Array-CGH performed on DNA obtained
from the initial HLA typing sample showed that the copy number was neutral for all loci with no evidence
of deletion, indicating that HLA loss in the initial sample was likely to be due to UPD. Procedures in this H&I
lab have been changed due to this incident. Blast counts are now required for all haematology patients in
blast crisis and are reviewed in association with all homozygous HLA typing results.

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P6: An Alternative UK Cord Blood Unit (CBU) could be Identified in Many Cases of Umbilical
Cord Blood Transplantation (UCBT) here an International Unit was Selected for UK Patients

Irina Evseeva¹, Laila Ramzi¹, Olga Nikolajeva², Susana G Gomez³, Alejandro Madrigal⁴
1Anthony Nolan, London, UK
2Anthony Nolan Research Institute, UCL Cancer Institute, London, UK
3Barcelona Blood and Tissue Bank, London, UK
4Anthony Nolan Research Institute, UCL Cancer Institute, London, UK

Introduction CBU selection practice for UCBT varies between transplant centres. A retrospective analysis
of CBU selection preferences in the UK was performed to define the rationale of CBU choice and availability
of a national option.

Materials and methods For CBU work-up requests received by Anthony Nolan in 2014-15, patient and
CBU characteristics were analysed. In cases where an international CBU was selected, the availability and
suitability of an alternative UK CBU was assessed based on current UK recommendations.

Results 134 patients were considered for single or double UCBT with 205 units requested in total. Of these
69% were international. Out of the total international selection, 19% did not have an acceptable UK CBU and
in 61% of cases an acceptable UK option was available, but not selected due to inferior parameters. In the
remaining 20% of cases, suitable UK CBUs with optimal qualities were not considered due to the following
reasons:

- not considering permissive mismatches when scoring HLA match - selecting units with
higher TNC above acceptable threshold

- high resolution typing needed to confirm the HLA match

- allelic level match for HLA-A,-B,-C,-DRB1 applied as a primary selection criterion for double-unit
UCBT while there is no clear consensus on its benefit in double-unit transplant

Conclusion For 20% of international CBUs selected for single or double UCBT for UK patients a UK CBU of
optimal quality could have been considered. In 80% of cases an international CBU was selected due to
unavailability or inferior parameters of the UK option.

P7: A 71% Reduction in High Resolution HLA Typing Turnaround-Time by Implementation
of One-Step Sequencing and a New Extended Typing Algorithm

Andrew Lemin¹, Rona Millington¹, Shirley Jobson¹, David Briggs¹, Luke Foster¹

1 NHS Blood and Transplant, Birmingham , UK

High resolution (HR) HLA typing is required to initiate a volunteer unrelated donor (VUD) search for
hematopoietic progenitor cell transplantation (HPCT) and for typing of VUDs being considered as donors.

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In 2015 our department implemented a one-step sequencing approach aiming to reduce the turnaround-
time (TAT) and cost of our current HR typing methodology whilst next generation sequencing (NGS) assays
are further explored. Our method utilises group-specific amplification of all known CI and CII HLA alleles
followed by sequencing of positive amplicons, thus omitting the need for an initial low resolution type.

Concurrently we implemented a new algorithm for extended typing of any donors selected from the VUD
search, which meant that typing was only performed where results would augment registry data for
determination of compatibility.

Compared to the mean number of calendar days (CD) taken to obtain a HR type for new HPCT referrals in
2013, our new approach has resulted in a 71% reduction in TAT (mean 32.2 CD in 2013 Vs 9.2 CD in 2015).
This considerable reduction has allowed the VUD search to proceed more quickly. Taken together with our
VUD typing algorithm, this should result in a positive clinical impact because the time taken to select a final
donor has also decreased.

Our approach resulted in a 26% cost reduction in non-pay expenditure in our HLA typing laboratories in
the last financial year despite an 8% increase in workload. As such, this is a cost effective and clinically
beneficial interim approach until the adoption of NGS.

P8: Development of a Systematic Decision Making Algorithm and Supporting Software for
HSCT Donor Selection - RADSS

Catherine Irwin¹, Peter Irwin², Maria Gilleece¹, Neil Marsden¹, Brendan Clark¹
1 Leeds Teaching Hospital Trust, Leeds, UK
2 Private IT Consultant

Introduction A risk adjusted HSCT donor search strategy has been developed to enable a systematic approach
to clinical decision making. The strategy has been refined and validated through audit and has become an
increasingly accurate tool. RADSS Software (Risk adjusted donor search strategy) has been developed to
enable application of the tool.

Methods Patient HLA type and time taken from request for search to receipt of donor samples into the
laboratory were obtained and used to derive an expected and actual chance of finding a donor. Expected
chance was informed using previously identified criteria. Actual chance was defined as time taken to receive
samples into the laboratory. High, intermediate and low chance categories were created with thresholds
<4, 4-6 and >6 weeks respectively. Non-concordant expected/actual outcomes were investigated in
refinement of the model.

Results The results of audit analysis were used to produce an evidence-based risk-adjusted algorithm.
Meeting or failing to meet the set criteria within this algorithm placed the patient in a high, intermediate
or low chance of finding a donor category. A software programme has been developed to enable the analysis
needed to categorise patients and simplify the use of the strategy.

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Conclusion We have established and validated an algorithm for optimal donor selection in the setting of a
clinical programme and have developed software to support its application. The software is highly visual
and shows criteria are that are impacting on chance, which can help when counselling patients about their
prospects of finding a donor.

P9: Identification of a Novel DPB1*105 Allele, By Single Molecule Real-Time (SMRT®) DNA
Sequencing in an African Caribbean Patient And Related Donor.
Cristabel Trujillo¹, Francisco Boix¹, Franco Tavarozzi¹, Finnuala Fowles¹, Katy Latham¹

1 Anthony Nolan Research Institute, Royal Free Hospital, London, UK

Background Allelic level typing for HLA is performed to achieve the best possible match in Haematopoietic
Stem Cell Transplantation (HSCT). Anthony Nolan recently introduced Pacific Biosciences’ Single Molecule
Real Time (SMRT) DNA sequencing to facilitate long range, phased sequencing of HLA. Here we characterize
a novel HLA-DPB1*105 allele identified in both an African Caribbean patient and sibling donor.
Material and Methods Initial typing for both patient and donor for HLA-DPB1 was performed by SBT
(SeCore®, uTYPE) covering exons 2, 3, and 4. Verification typing was performed using SMRT sequencing
resulting in a 5.5kb fragment encompassing exons 2-4. Comparison of the results highlighted a discrepancy
investigated using sequence data and alignment tools available from IMGT/HLA.
Results SBT resulted in HLA-DPB1*23:01:01, *51:01 and subsequent SMRT sequencing HLA-
DPB1*04:01:01:01, *105(variant) revealing a single nucleotide difference in exon 3 (443A>G). SBT was unable
to characterize the novel position due to heterozygous sequence generation and no exon 3 data available
in IMGT/HLA for HLA-DPB1*51:01 causing an error in the analyses. As this allele was identified independently
in both patient and sibling, the novel position was confirmed and the sequence submitted to IMGT/HLA.
Conclusion The identification of the novel allele by SMRT sequencing reflects the importance of phase in
HLA analysis. The advent of such technology will result in an increase in alleles registered in IMGT/HLA. H&I
laboratories must be able to type to true allelic level typing to ensure the best possible match for patients
and contributing to improved outcomes.

P10: Cord Blood Banking: Getting Started and Keeping Going, Collection and Diversification
Victoria Robertson¹, Jordan Harvie¹, Salmaan Dalvi¹, Amr Abid¹, Husein Salem¹

1 Precious CellsGroup, UK

Umbilical cord blood (UCB) transplantation shows comparable outcomes to those of matched unrelated
stem cell transplantation. However, a decrease in uptake of UCB units is affecting the sustainability of public
cord blood banks (CBB). To that end, Precious Cells Group (PCG)

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has developed a hybrid private/public CBB with the target of banking 10000 public UCB units over four years
and supporting the scientific community by providing UCB-derived components for research. The public CBB
started collections in late 2014.

To achieve this, PCG is implementing next generation sequencing to provide unambiguous HLA-A, -B,
-C,-DRB1/3/4/5 and -DQB1 types for all units. PCG is also working to establish a robust standardised CFU
assay for all pre- and post-cryopreservation assays. Along with standard processing data, this information
should facilitate an informed selection process but also a more stringent banking process. However, as
approximately 2/3 of units collected do not meet processing criteria, PCG is also developing processes to
make best use of these altruistic donations to derive components, such as mesenchymal stem cells, platelet
lysate and regulatory T-cells to support transplant research activities.

Since operations started in 2008, PCG has banked just under 14000 UCB units, 93% autologous and 7%
allogeneic, with 44 units released. PCG is currently applying for FACT accreditation and will apply for AABB
accreditation in 2017, with applications for UKAS and EFI accreditations to follow. Achieving accreditations
and providing UCB-derived products will translate into sustainable operations providing high quality cords
for patients worldwide, whilst also supporting transplantation research.

P11: Validation and Optimisation of a 16-Locus Multiplex PCR for Engraftment Monitoring
of Post-Haematopoietic Stem Cell Transplantation Patients

Chris Hughes¹, Helena Lee¹, Sophia Rowlston¹, Simon Mathers¹

1 Transplantation Laboratory, Manchester Royal Infimary, Manchester, UK

In the field of haematopoietic stem cell transplantation, chimaerism monitoring is crucial to determine the
success of donor cell engraftment post-transplant, and also to predict negative clinical outcomes. The
amplification of short tandem repeat (STR) loci via multiplex PCR has long been a leading methodology of
monitoring. The aim was to improve the current protocol by increasing the number of STR loci amplified in
a multiplex reaction. Following primer design, provisional runs were conducted at equi-molar primer pair
concentrations. Subsequently, experimentation with magnesium chloride (MgCl2) concentration, bovine
serum albumin (BSA) concentration, and annealing temperature was carried out to ascertain the reaction
conditions which elicit the greatest amplification efficiency. Provisional runs indicated that amplification of
all 16 target regions was successful, however preferential amplification of certain loci relative to others was
apparent. This issue was rectified by careful adjustment of primer pair concentrations for individual loci.
The results of altering the final concentration of MgCl2 were conclusive: incremental increases in MgCl2
corresponds to increased amplification efficiency. Similarly, increases in BSA concentration improves product
yield, but excess (>0.50µg/µl) inhibited amplification. Published guidelines recommend the monitoring of
at least 3 ‘informative’ markers. The introduction of an additional 8 loci into the multiplex panel will inevitably
increase the probability of meeting this minimum recommendation, which is currently a challenge in certain
circumstances to achieve from the 8-loci system. Further work is needed to fully validate this methodology,
however, once implemented, the increased choice of STR markers for analysis will have an invaluable effect
on analysis efficiency, cost effectiveness, and turnaround time of results.

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Antibody Detection

P12: Technical Limitations of the C1q Single-Antigen Bead Assay to Detect Complement
Binding HLA-Specific Antibodies
Sarah Peacock¹, Craig Taylor¹, Vasilis Kosmoliaptsis¹, Jess Martin¹, Graham Knighton¹, Dermot Mallon¹, John
Andrew Bradley¹

1 Cambridge University HospitalsNHS Foundation Trust
Background Solid-phase assays to distinguish complement binding from non-complement binding HLA-
specific antibodies have been introduced, but technical limitations may compromise their interpretation.
We have examined the extent to which C1q-binding to HLA- class I single-antigen beads (SAB) is influenced
by denatured HLA on SAB, antibody titre, and complement interference that causes a misleading low
assessment of HLA-specific antibody levels.
Methods Sera from 25 highly sensitized patients were tested using Luminex IgG-SAB and C1q- SAB assays.
Sera were tested undiluted, at 1:20 dilution, and after ethylene diamine tetra acetic acid (EDTA) treatment.
Conformational HLA and denatured HLA (dHLA) levels on SAB were determined using W6/32 and HC-10
monoclonal antibodies. dHLA was expressed as HC- 10 binding to untreated SAB as a percentage of
maximal binding to acid-treated SAB.
Results For undiluted sera, Luminex mean fluorescence intensity (MFI) values for IgG-SAB and C1q-SAB
correlated poorly (r = 0.42). EDTA and serum dilution improved the correlation (r = 0.57 and 0.77,
respectively). Increasing levels of dHLA interfered with the detection of C1q binding; the correlation
between IgG-SAB MFI and C1q-SAB MFI was lowest using undiluted sera and SAB with greater than 30%
dHLA (r = 0.40) and highest using diluted sera and SAB with 30% or less dHLA (r = 0.86).
Conclusions Antibody level, complement interference, and dHLA class I on SAB may all affect the clinical
interpretation of the C1q-SAB assay. The C1q-SAB assay represents a substantial additional cost for routine
clinical use, and we question its justification given the potential uncertainty about its interpretation.

Methodology

P13: The Impact of Automation on DNA Library Preparation for Whole Gene HLA Typing By
Next Generation Sequencing (NGS)
Sue Davey¹, Monica Kyriacou¹, Jasmine Rizvi¹, Arthi Anand¹, Colin Brown¹

1 NHS Blood and Transplant, Colindale, London, UK

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NHSBT has HLA typed >10,000 adult and cord blood donor samples by NGS since its introduction in
2015. Although the majority of our in-house NGS protocol had been automated, the process of
fragmenting, adapter ligation and index PCR of whole gene amplicons to form pooled amplicon
libraries (PAL) was still performed manually. Due to the technical and ergonomic demands of
processing large numbers of samples, we recently validated an automated protocol on the Biomek
FXp liquid handling robot during which quality parameters from twenty PALs were determined and
compared with DNA libraries produced manually.

We found no significant difference in the mean concentration of PAL when processed manually (4.9ng/ul)
or using our automated protocol (4.6 ng/ul). Conversely, fragment sizes produced with the automated
protocol were markedly smaller, with an average of 1094bp compared to the 1806bp generated by manual
preparation. Despite this smaller size, automation did produce a more consistent library, ranging from 782bp
to1382bp, compared to 660-2519bp obtained by the manual method. Knowledge of both concentration and
fragment size are necessary to calculate PAL molarity and obtain optimal cluster density on the MiSeq. The
use of robotics requires significant investment but benefits are quickly realised through accurate and
consistent pipetting alongside reduced risk of staff injury due to repetition. It is also anticipated that the
predictable fragment size of the PAL produced by automation will negate the need for expensive and time
consuming quality assessment, and replace with a simple quantification step prior to loading onto the MiSeq.

P14: Evaluation of the HISTO SPOT PCR-SSO System

Mohammad Ali Rafique¹, Mian Chen¹, Susan Fuggle¹, Martin Barnardo¹

1 Oxford Transplant Centre, Oxford, UK

HLA genotyping forms the backbone of most histocompatibility laboratories. Currently employed
methodologies for low/medium-resolution typing in the UK include gel-based and real-time PCR-SSP and
Luminex PCR-SSO. We evaluated the HISTO SPOT PCR-SSO kit (provided FOC by BAG Technology), which uses
SSO probes immobilised in an array at the bottom of microplate wells. The probes in each well are specific
for a single locus and are detected via colourimetric reactions photographed using the Mr.SPOT processor.
This technique is mostly automated and simple to use with minimal operator handling, has low- turnaround
time (3 hours), and samples can be run in batches (medium throughput). DNA samples (concentration range
15-44 ng/ul in dH2O) from 49 individuals were selected to cover all known split antigens of HLA-A, -B,
-Cw,-DRB1, and -DQB1.

Of 238 loci (476 alleles) assayed in total, 229 (96.2%) produced a concordant result after a single attempt.
Of the remaining 9 loci which failed to give a concordant result, 6 had failed and 3 had given an unlikely
allele in the correct antigenic group (which then became correct on retyping). 3/6 failures yielded concordant
results on repeat typing and 2/6 failed again. Onr etyping, the remaining sample yielded the single
non-concordant result in the study (HISTO SPOT – HLA-C*04:01, PCR-SSP – HLA*04:03).

In summary, one HLA allele out of the 472 assigned alleles (99.8%) was non-concordant with the established
method, however this discrepant allele was in the correct antigenic group; sufficient to fulfil the minimum
requirements for NHSBT-ODT donor HLA typing.

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