The Significance of the Direction of the HLA Mismatch in Cord Blood Matching and the Implication of Graft-Specific Anti-HLA Antibodies

1
The Significance of the Direction of the HLA Mismatch in
Cord Blood Matching and the Implication of Graft-
Specific Anti-HLA Antibodies
Marcelo A. Fernández Viña, Ph.D.
Department of Pathology
Medical School
Stanford University

2
Goals of Allogeneic HSCT
• Achieve Engraftment
• Absence of Graft versus Host Disease
• Prevent Relapse
• Effective Immune reconstitution

3
Search for Allogeneic
HSC
Nuclear family
Extended family
CBU and Unrelated Donors

44
Factors Influencing Decision to Transplant
 Quality of HLA match
 Recipient factors
 Diagnosis and stage of disease
 Age
 Major organ function
 CMV serology
 Donor/CBU factors
 Age – Period of collection
 Body size - Cell Dose
 CMV serology – Infectious Disease Markers
 Pregnancy and transfusion history

5
Patient and Donor/CBU are Heterozygous in all HLA loci
Difference (mismatch) in One allele of HLA-A
• The HLA-A mismatched antigen in the Donor/CBU can be recognized as
foreign by the Patient’s Immune System (Host versus Graft; rejection)
• The HLA-A mismatched antigen in the Patient can be recognized as foreign by
the Donor/CBU’s Immune System (Graft versus Host; GvHD and GvL)
33
Molecules encoded in the HLA systemMolecules encoded in the HLA system
Maternal
origin
PATIENT
Encoded in HLA-DRB1
locus
(Class II)
Encoded in HLA-A
locus
(Class I)
Paternal
origin
Encoded in HLA-B
locus
(Class I)

6
Patient is Homozygous in HLA-A and Donor/CBU is
Heterozygous in HLA-A
Mismatch Only in the HvG direction
• The HLA-A mismatched antigen in the Donor/CBU can be recognized as
foreign by the Patient’s Immune System (Host versus Graft; rejection)
• No mismatch in the Graft versus host direction (No Graft versus Host; No
GvHD/ No GvL)
33
Maternal
origin
DONOR/CBU
Encoded in HLA-DRB1
locus
(Class II)
Encoded in HLA-A
locus
(Class I)
Paternal
origin
Encoded in HLA-B
locus
(Class I)

7
Patient is Heterozygous in HLA-A, Donor/CBU is
Homozygous in HLA-A for one allele present in the Patient
Mismatch Only in the HvG direction
• The HLA-A mismatched antigen in the Patient can be
recognized as foreign by the Donor/CBU ’s Immune
System (Graft versus Host; GvHD and GvL)
• No mismatch int Host versus Graft direction
33
Maternal
origin
PATIENT
Encoded in HLA-DRB1
locus
(Class II)
Encoded in HLA-A
locus
(Class I)
Paternal
origin
Encoded in HLA-B
locus
(Class I)

8
“HLA mismatch direction in cord blood transplantation: impact on
outcome and implications for cord blood unit selection”
Stevens CE, Carrier C, Carpenter C, Sung D, Scaradavou A
Blood. 2011 Oct 6;118(14):3969-78.
• 1202 single CBU TX
• 1993-2006 (57% before 2000)
• Engraftment at day 77
• 75 % <16 y/o
• 67% Malignancies
• Early (23%), Intermediate
(46%), Advanced 30%)
• 6/6 – 6%
• 5/6 – 38%
• 364 1 bidirectional MM
• 58 GvH-only MM
• 40 HvG-Only MM
• 76% Neutrophil engraftment
• 51 % Platelet engraftment
• Match grade (p<0.001) and cell
dose (p<0.006) changed over
time
• Engraftment Faster in the GvH-
only (HR = 1.6, P = .003)
• GvH-only had Less TRM,
Mortality, Treatment Failure in
patients with Maligancy
• Comparable to Matched
transplants

Cumulative probability of achieving ANC 500 during the first 77 days after transplantation.
Stevens C E et al. Blood 2011;118:3969-3978
©2011 by American Society of Hematology

The Significance of the Direction of the HLA Mismatch in Cord Blood Matching and the Implication of Graft-Specific Anti-HLA Antibodies

Cumulative probability of TRM during the first 3 years after transplantation.
Stevens C E et al. Blood 2011;118:3969-3978
Patients with malignancies
HLA-matched or GVH-only
mismatched CB grafts
Patients with other diseases
1 bidirectional mismatch

12
• “Evaluation of HLA matching in unrelated
hematopoietic stem cell transplantation for
nonmalignant disorders”
• John Horan, Tao Wang, Michael Haagenson,
Stephen R. Spellman, Jason Dehn, Mary Eapen,
Haydar Frangoul, Vikas Gupta, Gregory A.
Hale, Carolyn K. Hurley, Susana Marino,
Machteld Oudshoorn, Vijay Reddy, Peter Shaw,
Stephanie J. Lee and Ann Woolfrey
• Blood. 2012 Oct 4;120(14):2918-24.

14
BMT in Non-Malignant Diseases
CBU
• Similar to HSCT in non-Malignant
Diseases, absence of Engraftment may have
played a major role in causing morbidity
and mortality

15
Goals of Allogeneic BMT
Achieve Engraftment:
• Patient’s immune system may cause
rejection
• Donor’s immune system may enhance the
engraftment through the destruction of
patient’s immune cells that cause rejection

16
Histocompatibility Factors
Affecting Engraftment
• HLA-mismatches in the Host versus graft
direction (HvG mismatch)
• Patient’s Homozygosity (HvG mismatch)
• Donor-specific anti-HLA Antibodies
• ABO Major Mismatch

17
Homozygosity at a given locus vs
Heterozygosity
Major-Histocompatibility-Complex class I alleles and antigens in hematopoietic-
cell transplantation.
Petersdorf EW, Hansen JA, Martin PJ, Woolfrey A, Malkki M, Gooley T, Storer
B, Mickelson E, Smith A, Anasetti C.
N Engl J Med. 2001 Dec 20;345(25):1794-800
• CML, No ATG
• RESULTS: A single HLA allele mismatch did not increase the risk of graft
failure, whereas a single antigen mismatch significantly increased the risk
• The risk was also increased if the recipient was HLA homozygous at the
mismatched class I locus or if the donor had two or more class I mismatches
• CONCLUSIONS: HLA class I antigen mismatches that are serologically
detectable confer an enhanced risk of graft failure after hematopoietic-cell
transplantation. Transplants from donors with a single class I allele mismatch that
is not serologically detectable may be used without an increased risk of graft
failure

Impact of High Resolution Mismatches (Allele Level) & Broad
Mismatches (Serologically Detectable) in Alleles of HLA Loci

20
The impact of HLA unidirectional mismatches on
the outcome of myeloablative hematopoietic stem
cell transplantation with unrelated donors
• Hurley CK, Woolfrey A, Wang T, Haagenson M,
Umejiego J, Aljurf M, Askar M, Battiwalla M, Dehn J,
Horan J, Oudshoorn M, Pidala J, Saber W, Turner V, Lee
SJ, Spellman S
• Blood. 2013 May 1. [Epub ahead of print] PubMed PMID:
23637130
• 2,687 myeloablative MUD; malignant disease
• 7/8 bidirectional MM transplants, n=1393
• 7/8 host vs. graft (HVG) vector MM , n=112
• 7/8 graft vs. host (GVH) vector MM, n=119
• 8/8 matches, n=1063

21
HvG and GvH mismatches
(Conclusions)
• The 7/8 groups differed only for grades 3-4 acute GVHD
where HVG MM had less GVHD than the 7/8 bidirectional
MM (HR 0.52, p=0.0016) and GVH MM (HR 0.43,
p=0.0009) but not the 8/8 group (HR 0.83, p=0.39)
• There were no differences between the 7/8 groups for
relapse, chronic GVHD, neutrophil engraftment or graft
failure
• Unidirectional GVH vector mismatches have the same risk
as 7/8 bidirectional mismatches
• Recipients with a 7/8 HVG MM have a reduced risk of
acute GVHD without an increased risk of disease relapse
or graft failure compared to a 7/8 bi-directional MM at a
heterozygous locus

23
Causes of Death
• The 7/8 bi-directional and GVH MM
groups had slightly higher deaths attributed
to GVHD, compared to the 8/8 and 7/8
HVG MM groups, 18.2% and 18.0% vs.
14.9% and 14.3%, respectively
• The 7/8 HVG MM group had more deaths
attributed to graft rejection than the 8/8, 7/8
GVH MM and 7/8 bidirectional groups,
9.5% vs. 0.6%, 1.1% and 1.6%, respectively

24
Differences in the number of Mismatched Epitopes when the Patient
is Homozygous or Heterozygous at the Mismatched locus

25
Heterozygosity
• Patients homozygous at a given locus when presenting
humoral sensitization tend to make anti-HLA antibodies
reactive with many (in some instances all but self antigens)
HLA antigens of the same locus
• Novel anti-HLA antibody screening techniques allow to
precisely identify anti-HLA antibodies defining unacceptable
or high risk mismatches. Therefore, donors can be excluded
or given lower priority on the basis of their HLA mismatch
and the patient’s antibody reactivity.
• Current conditioning/immunosupression (ATG) this risk for
primary graft loss mediated by T-cells may be greatly
reduced

26
Heterozygosity
• Current conditioning/immunosupression (ATG) this risk for
primary graft loss mediated by T-cells may be greatly
reduced

DSA in TCD Haploidentical Stem Cell Transplantation
Ciurea SO, de Lima M, Cano P, Korbling M, Giralt S, Shpall EJ,
Wang X, Thall PF, Champlin RE, Fernandez-Vina M
 DSA were detected in 5 of 24 consecutive patients
(21%)
 4/24 patients had primary graft failure (PGF)
 3/4 (75%) patients with DSA failed to engraft as
compared with 1/20 (5%) without DSA (P=0.008)
DSAEngraftment YES NO
YES 1 3
NO 19 1
“High risk of graft failure in patients with anti-HLA antibodies undergoing
haploidentical stem-cell transplantation”
Transplantation. 2009 Oct 27;88(8):1019-24

Recently published work: Anti-HLA
Antibodies and Virtual Cross Match in MUDs
 Retrospective study - DSA found in 9/37 MUDT patients with
graft failure
 1/78 had DSA in the matched control group of patients who
engrafted (TCR grafts, 85% mismatched at DP locus)
Spellman S, Bray R, Rosen-Bronson S, Haagenson M, Klein J,
Flesch S, Vierra-Green C, Anasetti C.
The detection of donor-directed, HLA-specific alloantibodies in
recipients of unrelated hematopoietic cell transplantation is
predictive of graft failure.
Blood. 2010 Apr 1;115(13):2704-8
PGF Control
DSA 9/37 (24%) 1/78 (1%)
Class I +/- Anti-DP 5 1
Anti-DP alone 4 0
DSA and PGF
RR = 24.8 – 28.1

Ciurea SO, Thall PF, Wang X, Wang SA, Hu Y, Cano P, Aung F, Rondon G,
Molldrem JJ, Korbling M, Shpall EJ, de Lima M, Champlin RE, Fernandez-Vina M.
Donor-specific anti-HLA Abs and graft failure in matched unrelated donor
hematopoietic stem cell transplantation.
Blood. 2011 Nov 24;118(22):5957-64
592 MUD TX
75 % of the 8/8 transplants present at least one
mismatch in DP, DQ, DRB3/4/5 in the HvG vector
 20 % of HSC patients are immunized against HLA
HLA Immunization in FEMALE HSCT patients: 30%
HLA Immunization in MALE HSCT patients: 10%
3.3 % present antibodies against HLA-DP (1/2 MUD TX
have one or two DP mismatches and DSA anti HLA-DP)

Other Associations: Gender, Number of Pregnancies
and anti-HLA antibodies
Variable N
Coefficie
nt SE P-value OR 95% CI
Intercept -2.71 0.60 <0.0001 - -
Male vs. Females 0
pregnancies
356 0.73 0.62 0.24 2.1 0.62 - 6.94
Female: Number of
pregnancies = 1
(vs. 0)
37 1.85 0.70 0.008 6.3 1.62 -
24.85
Female: Number of
pregnancies ≥ 2
(vs. 0)
152 2.25 0.62 0.0003 9.5 2.83 -
32.02
Significant association between gender and the presence of
AHA: 30.8% females vs 12.1% males had anti-HLA
antibodies (p<0.0001)
7/8 pts with DSA were females

31
Lower Expression of HLA in CBU-
CD34
Bone
Marrow
CD34
Cord Blood
CD34

Anti-HLA antibodies in CBU
transplantation
 DSA in Single Unit CBU TX
 Double CBU and DSA:
• DSA against both Units
• DSA against one Unit
• No DSA

33
Evaluation of DSA by a Solid Phase Assay
with Single Antigen Beads

34
“The impact of anti-HLA antibodies on unrelated cord blood
transplantations”
Takanashi M, Atsuta Y, Fujiwara K, Kodo H, Kai S, Sato H,
Kohsaki M, Azuma H, Tanaka H, Ogawa A, Nakajima K,
Kato S
Blood. 2010 Oct 14;116(15):2839-46
• 386 Single CBT; myeloablative, Median age 33
• HLA Immunization: 89 patients (23%); 20 cases with
DSA
• Neutrophil recovery @ day +60 :
• 83% for the ab-negative
• 73% for ab-positive/No-DSA
• 32% ab-positive/Yes-DSA
• Anti-HLA antibodies should be tested and considered
pre-transplantation in the selection of cord blood

Cumulative incidence of neutrophil/platelet recovery and survival for 386 CBT cases. ab-
negative indicates patient does not have anti-HLA antibody; ab-positive, patient has anti-HLA
antibody but the CB does not have the corresponding antigen for the antibod...
Takanashi M et al. Blood 2010;116:2839-2846

36
“Donor-specific anti-HLA antibodies predict outcome in double umbilical
cord blood transplantation”
Cutler C, Kim HT, Sun L, Sese D, Glotzbecker B, Armand P, Koreth J,
Ho V, Alyea E, Ballen K, Ritz J, Soiffer RJ, Milford E, Antin JH
Blood. 2011 Dec 15;118(25):6691-7
• 73 Double CBT; myeloablative (27%)or
RIC/ATG (73%); Median age 48
• 18 patients with DSA (11 Single Unit DSA,
7 DSA against both CBU)

Cumulative incidence of graft failure.
Cutler C et al. Blood 2011;118:6691-6697
no DSA
DSA against one unit
DSAs against both units

Cumulative incidence of neutrophil engraftment.
no DSA
DSA against one or both units

Cumulative incidence of early death or relapse.
no DSA
DSA against one unit
DSA against both units

DSA in the Study by Brunstein et al
• Lower cut-off for calling positve anti-HLA antibodies
• DSA against 1 Unit: n = 12; 2 graft failures
• DSA> 3000 MFI: n = 3; 1/3 had graft failure
• DSA against both Units: n= 6; 1 graft failure
• One had DSA> 3000 MFI : n = 4 ; 1/3 had graft failure

Impact of donor specific anti-HLA antibodies on graft
failure and survival after reduced intensity conditioning-
unrelated cord blood transplantation. A Eurocord,
Societe Francophone d'Histocompatibilite et
d'Immunogenetique (SFHI) and Societe Francaise de
Greffe de Moelle et de Therapie Cellulaire (SFGM-TC)
analysis
Boudifa A, Coeffic B, Devys A, De Matteis M, Dubois V,
Hanau D, Hau F, Jollet I, Masson D, Pédron B, Perrier P,
Picard C, Ramouneau-Pigot A, Volt F, Charron D, Gluckman
E, Loiseau P
Haematologica. 2012 Dec 14. [Epub ahead of print] PubMed
PMID: 23242594

43
transplantation
• HvG vector, Serologic Level mismatches (class I)
• Sensitivity, specificity and quantitation of antibody detection are greatly
enhanced by novel solid phase assays
• Variations in end points and cut-off values
• Cross-match testing is not be practical in HSCT
• High risk of graft rejection in patients with D.S. Antibodies
• Higher mortality and morbidity associated to DSA
• Allo-antibodies directed against both class I and II HLA antigens
• Less mismatches in MUD than in CBU
• Anti-DP antibodies in MUD (mostly matched at other HLA loci)
• Anti-HLA antibody testing is warranted to all HSCT with known or presumed
HLA mismatches
• Further analysis for different graft sources and diseases is warranted

“Complement (C1q) fixing solid-phase screening for HLA antibodies
increases the availability of compatible platelet components for refractory
patients”
Fontaine MJ, Kuo J, Chen G, Galel SA, Miller E, Sequeira F, Viele M,
Goodnough LT, Tyan DB.
Transfusion. 2011 Dec;51(12):2611-8
• 13 highly sensitized refractory patients received 177 PLT units
incompatible by the IgG-SAB method
• The mean CPRA value was significantly lower by C1q-SAB (60%)
than by IgG-SAB (94%; p < 0.05)
• Patients showed significantly better corrected count increment with
C1q-compatible than with C1q-incompatible PLTs
• Results show that 75% of PLT units previously considered
incompatible were actually compatible
• For highly refractory patients to PLT transfusion, the C1q-based SAB
binding assay may be a better method for identifying clinically
relevant HLA antibodies and selecting PLT units that will result in
acceptable CCI

0
5000
10000
15000
20000
25000
A*23:01
A*80:01
A*01:01
A*24:02
A*36:01
A*29:02
A*24:03
A*66:02
A*33:01
A*33:03
A*29:01
A*34:01
A*32:01
A*11:02
A*43:01
A*11:01
A*03:01
A*74:01
A*26:01
A*34:02
A*69:01
A*68:02
A*25:01
A*02:06
A*66:01
A*02:01
A*68:01
A*02:03
A*30:01
A*30:02
A*31:01
IgG C1Q
FluorescenceIntensity
Self HLA-A
A*02, A*24, A*25, A*26,A*34, A*68, A*69, A*43
may be acceptable by the C1q Assay
Antibody
Tests by
Assays

46
transplantation
• Only 16-24 % PGF have DSA
• T-cell immunity
• Conditioning, Immunosuppressive regimens
(ATG, post TX Cyclophosphamide)
• Preservation of patient’s Immune System (Primary
Disease and Stage)
• Graft type and variations in HLA expression
• Graft Manipulation

4747
Goals of Allogeneic BMT
• Absence of Graft versus Host Disease:
GvHD is mediated by donor’s T-cells
• Effective Immune reconstitution:
T-cells of donor origin may interact with patient’s tissues
and in some instances patient’s APCs
• Prevent Relapse:
GvL may be mediated by both, T-cells and NK cells of
donor origin; donor’s T cells causing GvHD may also
prevent relapse
• GvH: Major factor in Mortality and Morbidity: Once
engraftment is achieved; the mismatches in the GvH vector
play a major role in transplant outcome

Heterozygosity
• However, in light of current conditioning/immunosupression
(ATG) this risk for primary graft loss mediated by T-cells
may be greatly reduced

49
Differences in the number of Mismatched Epitopes when the Patient
is Homozygous or Heterozygous at the Mismatched locus

Neutrophil Engraftment
(absence @ day+28)
Mismatch n RR L-R HR p
8/8 4779 1.00 - - 0.0009
C*03 PMM 134 0.84 0.33 2.16 0.72
C-allele 61 0.41 0.04 4.69 0.47
C-antigen 700 1.66 1.27 2.18 0.0002
A, B or DRB1 959 1.46 1.13 1.89 0.0034

Histocompatibility Factors
Affecting Engraftment
• HLA-mismatches in the Host versus graft
direction (HvG mismatch)
• Serologic level HLA-class I mismatches
• Patient’s Homozygosity (HvG mismatch)
• Donor-specific anti-HLA Antibodies
• ABO Major Mismatch

Univariate Logistic Regression Model for Graft Failure in MUDT
Variable
Parameter
Estimate
Parameter
Standard Error
Univariate
P-value Odds Ratio
Male (vs. female) -0.53 0.47 0.26 0.59
Black (vs. white) 1.17 0.78 0.14 3.22
Others (vs. white) -12.15 316.51 0.97 0.00
AHA= yes (vs. no) 0.91 0.49 0.06 2.48
DSA = yes (vs. no) 3.06 0.77 0.0001 21.33
HEL HvG = 8 (vs. 7) -0.31 0.64 0.62 0.73
LEL HvG = 6 (vs. <6) -0.53 0.64 0.41 0.59
CD34 cell numbers
infused
-0.51 0.38 0.18 0.60
CMV mismatch = yes (vs.
no)
0.27 0.47 0.56 1.31
Sex mismatch = yes (vs.
no)
0.60 0.47 0.21 1.82
ABO mismatch = yes (vs.
no)
1.35 0.63 0.03 3.86
AHA – anti HLA antibodies; DSA – donor-specific AHA; HEL – high expression loci; LEL – low expression loci;
HvG – host versus graft direction
MVA: DSA (p=0.0001) and ABO mm (p=0.04) remained significantly associated with GF

Cumulative incidence of neutrophil and platelet recovery in subgroups of CD34 cell dose. ab-
negative indicates patient does not have anti-HLA antibody; ab-positive, patient has anti-HLA
antibody but the CB does not have the corresponding antigen for the ant...
Takanashi M et al. Blood 2010;116:2839-2846

Progression-free (A) and overall survival (B)
no DSA
no DSA

transplantation
 DSA in Single Unit CBU TX
 Double CBU and DSA
 DSA against both Units
 DSA against one Unit
 Even though the authors do not agree:
Higher mortality and morbidity associated to
DSA. Variations in end points and cut-off
values

The Significance of the Direction of the HLA Mismatch in Cord Blood Matching and the Implication of Graft-Specific Anti-HLA Antibodies

More Related Content

What's hot (20)

Similar to The Significance of the Direction of the HLA Mismatch in Cord Blood Matching and the Implication of Graft-Specific Anti-HLA Antibodies (20)

More from cordbloodsymposium (20)

Recently uploaded (20)

The Significance of the Direction of the HLA Mismatch in Cord Blood Matching and the Implication of Graft-Specific Anti-HLA Antibodies