Anti-HLA Antibodies and Outcomes after

Cord Blood Transplantation Annalisa Ruggeri, MD

Eurocord, Hôpital Saint Louis, Paris

Background

• Delayed hematopoietic recovery and graft failure (GF) are critical complications of cord blood transplantation (UCBT) and are associated with TNC or CD34 cell dose and HLA disparities

• Other factors such as anti HLA-antibodies in the recipient may be associated with transplant outcomes

• Anti-HLA antibodies in the recipient may be unspecific or donor specific (DSA), directed to an antigens found in the graft

Background

• In solid organ transplant, DSA is associated with organ rejection and poor outcomes

• In unrelated donor recipients, Spellman et al. reported pre-transplant anti-HLA antibodies associated to GF and mortality. Interestingly, in this case control study, higher frequency of DSA anti HLADP was reported

• In UCBT, DSA has also been associated with transplant outcomes, including GF. However, authors report different findings regarding the impact of DSA in UCBT

Takanashi, Blood 2010

•386 patients with malignant diseases

•single UCBT and MAC regimen

•89 (23%) pts had anti HLA antibody positive screening

•20 pts had DSA against the CB (15 vs. HLA class I, 5

vs. class II)

•Assays with 1000 mean fluorescent intensity (MFI)

above baseline were considered positive

Results

Neutrophil and plt recovery Survival: OS and EFS

Takanashi, Blood 2010

In multivariate analysis, presence of DSA was associated with lower neutrophil and platelet engraftment, treatment failure and survival

Conclusion • Transplants with CBU with an antigen

corresponding to DSA had higher risk of graft failure and overall mortality

• The authors suggest that the presence and specifity of DSA should be determined and use if CBU with the corresponding antigens should be avoided

Takanashi, Blood 2010

• 73 adult patients transplanted from 2004-2008

• dUCBT after MAC (FluCyTBI) or RIC (FluMelATG)

• Anti-HLA DSA were detected in 18/73 pts, 4 against both CBU Cutler, Blood 2011

Methods • Patients plasma samples were analyzed for the presence of anti-

HLA antibodies (vs. HLA-A, B, C, DR, DQ)

• Assays with 1000 mean fluorescent intensity (MFI) above baseline were considered positive

• Anti-HLA antibody were considered for analysis if directed against class I or II HLA specificities found in CBU

Cutler, Blood 2011

Results- Graft Failure

Cumulative Incidence of GF was 12.3%

DSA vs both CBU (n=7), 57%

DSA vs one CBU (n=11), 18%

No DSA (n=55), 5%

The presence of DSA against one CBU (n=11) or DSA against both CBU (n=7) was independently associated with lower incidence of engraftment (p=0.001 and p=0.015)

Cutler, Blood 2011

Results- Early Mortality

DSA vs both CBU (n=7), 71%

DSA vs one CBU (n=11), 36%

No DSA (n=55), 23%

• Relapse with or without death was also associated with DSA status

In multivariable analysis, the presence of DSA against both CBU (n=7) was independently associated with relapse or early death (p=0.03)

Cutler, Blood 2011

Results- Survival

3y EFS was 30% 3y OS was 43%

The presence of a single DSA was not statistically significant, however comparing pts with DSA vs both CBU with those without any DSA,

both EFS and OS were significantly shorter (3y EFS: 0% vs 33%, p=0.004; 3y OS 0% vs 45%, p=0.04)

DSA vs both CBU (n=7), 0%

No DSA (n=55), 33%

DSA vs both CBU (n=7), 0%

No DSA (n=55), 45%

Cutler, Blood 2011

Conclusion

• The presence of pre-formed DSA against CBU increases the incidence of graft failure, delays neutrophil and platelet engraftment

• Findings demonstrate the necessity of screening of DSA before CBU selection. The presence of DSAs should be an important factor in the selection algorithm

Cutler, Blood 2011

Brunstein, BBMT 2011

•297 patients received a dUCBT from 2004-2009

•126 patients had stored plasma for testing for anti-HLA ab

•Anti-HLA ab were positive in 50 of the 126 pts (41%)

•Of the 50 patients with one or more anti-HLA ab:

-12 (24%) had a DSA that targeted 1 CBU

-6 (12%) had a DSA that targeted both CBU

Assays with 500 MFI above baseline were considered

positive

Results

• The CI of neutrophil engraftment for patients with DSA against 1 CBU was 78%, (median 24.5 days) and was 86% in patients with no antibody, (median 19 days), p=0.25

• Of the 12 patients with a DSA vs 1 or 2 CBU, the targeted unit was detectable in 9 patients

• Of the 6 patients with DSA vs both CBU, 5 engrafted, with the predominance of only 1 CBU in 4 cases, whereas in the other 2 cases both CBU coexisted at day +21

Brunstein, BBMT 2011

Results

Brunstein, BBMT 2011

Conclusion

• Contrarily two the other two studies, Brunstein showed no association between the presence of DSA and transplant outcomes after double UCBT, however the threshold for DSA positivity was different

• Homogeneous techniques are needed

Brunstein, BBMT 2011

Haematologica 2013

Selection Criteria • UCBT from 2000 to 2010

• Single and double UCBT, performed in France

• Reduced intensity conditioning regimen • Availability of pre-transplant samples to evaluate DSA

n=294 patients

Patients and disease characteristics, n=294

Diagnosis

Acute Leukemias

39%

MDS/MPD 12% CML

4%

CLL/ Lymphoma

s 19%

Plasma Cell

Disorder 8%

Solid Tumor

1% BMFS 14%

Immune Deficiency

3%

Median Follow-up, months (range) 36 (3- 98)

Children, n 60, 20%

Female gender, n 136, 46% Non malignant

disease, n 50, 17%

Previous Auto-HSCT, n 112, 38%

Anti HLA antibodies

• Pre-transplant serum was tested for HLA-Ab using LuminexTM platform

• Assays with 1000 mean fluorescent intensity (MFI) above baseline were considered positive

• 54/294 patients (18%) had positive anti-HLA antibodies before UCBT and of these 14 patients had donor specific anti-HLA (DSA)

Type of Graft and Conditioning Regimen, n=294

*HLA A, B antigenic level - DRB1 allelic level

No DSA (n=280) DSA (n=14)

Malignant disease 83% 80%

Type of transplant: Double UCBT 64% 50%

HLA match*: >2 mismatches

70%

78%

Median TNC (107/Kg) 3,6 (1,5-17) 3,7 (1,5-5,20)

Median CD34 (105/Kg) 2,4 (1-10)

2,4 (1-4,30)

Conditioning regimen: CyFluTBI<6 77% 78%

GvHD prophylaxis: CsA+MMF

74% 74%

ATG before day 0 34% 36%

Diagnosis DSA DSA specificity for UCB unit MFI HLA disparities TNC/Kg

(107)

1 BMF, 12y Class II DRB1*1301, DQB1*0301, 0603, DPB1*0301

2490, 4110, 4189, 1723, 3422 4/6 (DRB1) 3,30

2 MDS, 52y Class I- II A*03, B*51, DP*04:01 11076, 6672, 5000 4/6 (A, DRB1) 3,00

3 CML, 51y Class I B*44:03 1226 5/6 (B) 5,10

4 BMF, 9y Class I- II C*0102, C*0501, DPB1*0201 1620, 1115, 2449 5/6 (A) 3,20

5 ALL, 55y Class I A*26 2032 4/6 (A, B) 4,30

6 AML, 47y Class I- II B*51, DPB1*0402 12695, 19969 4/6 (A, B) 4,00

7 MDS, 40y Class I B*07 6500 4/6 (B, DRB1) 4,50

8* HD, 34y Class I A*02 B*44

3000 8500 4/6 (A, B) 2,52

9* BMF, 22y Class I A*24, B*35 B*37

9000 3800 4/6 (A, B) 5,20

10* HD, 18y Class I B*44 _

2100 _ 4/6 (A, B) 3,76

11* AML, 19y Class II _ DP*0104

_ 3587 4/6 (A, B) 3,62

12* AML, 42y Class II DRB1*1501 _

3650 _ 4/6 (B, DRB1) 3,20

13* AML, 36y Class I B*57 _

2500 _ 4/6 (A, B) 3,40

14* AML, 61y Class I _ A*01

_ 7000 4/6 (A, B) 4,70

DSA positive patients- characteristics, n=14

*dUCBT recipients

0 10 20 30 40 50 60

Days

0.0

0.2

0.4

0.6

0.8

1.0

Cum

Inc

Neu

troph

il En

graf

tmen

t

No DSA, n=280DSA, n=14

Results- Neutrophil Engraftment by DSA

No DSA: 78±3%

p=0.003

DSA: 44±8%

•The intensity of DSA measured by MFI correlates with the occurrence of graft failure All the 6 patients with DSA who engrafted had MFI lower than median (3900). The median MFI among the DSA patients who experienced graft failure was 7750 (range 2032-19969), and it was 2474 (range 1226-3650) in the DSA patients who engrafted (P=0.004)

Results- Neutrophil Engraftment and Survival for DSA positive patients

MFI Chimerism Engraftment (days)

Other treatment Outcome Follow-up

(months)

1 2490, 4110, 4189,

1723, 3422 NT No 2nd UCBT Dead- Rejection 3

2 11076, 6672,

5000 NT No Auto HSCT Dead- Rejection 1

3 1226 Full Donor Day 17 Alive 13

4 1620, 1115,

2449 Full Donor Day 18 Dead- Infection 3

5 2032 Autologous No Dead- Relapse 8 6 12695, 19969 Autologous No Alive 59 7 6500 Autologous No Dead- Relapse 3

8* 3000 8500 NT No 2nd UCBT Alive 56

9* 9000 3800 NT No Dead- Relapse 2

10* 2100 Full Donor Day 15 Alive 23

11* 3587 Full Donor Day 8 Dead- Relapse 3

12* 3650 Full Donor Day 24 Dead- GvHD 5

13* 2500 Mixed Chimera Day 17 Dead- Infection 7

14* 7000 Autologous No Dead- Rejection 1

DSA Negative patients n=280

Treatment failure

• Of the non DSA population, 65 patients failed to engraft – 14 received a 2nd HSCT

– 24 had autologous reconstitution (12 were alive with a median FU of 26 months, 7 died of relapse and 5 of TRM)

– 27 did not received any subsequent treatment • 26 died in a median time of 53 days (6 of relapse and

20 of TRM)

0 2 4 6 8 10 12Months

0.0

0.2

0.4

0.6

0.8

1.0

Cum

ulat

ive

Inci

denc

e of

TR

M

No DSA, n=280DSA, n=14

Transplant-Related Mortality by DSA

p=0.06

No DSA: 32±3%

DSA: 46±9%

p=0.07

Overall Survival by DSA

No DSA: 42±3%, n=280

DSA: 29±12%, n=14

Conclusion •Our findings confirm that the presence of DSA in recipients measured by Luminex assay (>1000 MFI), is associated with significantly higher graft failure after single or double UCBT with RIC

•In the different published studies the frequency of DSA ranges from 0.5% in the Japanese group to 20% in the American reports

•This difference in frequency may due to Tx center policy (recipients systematic screening) and use of leukocyte reduced cellular blood component

•Considering feasibility, in France, the average cost for screening for DSA identification is 250 euros

Summary- DSA and UCBT outcomes

Authors n Type of UCBT

and Conditioning Regimen

AntiHLA Ab, n DSA, n MFI Outcomes

Takanashi, Blood 2007 386 sUCBT, MAC 89 20 1000 Higher GF and

Overall mortality

Cutler, Blood 2011 73 dUCBT, MAC& RIC (73%) -- 18 1000 Higher GF

Brunstein, BBMT 2011 126 dUCBT, MAC&

RIC 50 18 500 No association with Tx outcomes

Ruggeri, Haematol. 2012 294 Single & dUCBT,

RIC 54 14 1000 Higher GF and Overall mortality

Summary •Pre-formed DSA directed against CBU have a deleterious impact on transplant outcomes

•Recipient screening and identification of DSA should be performed using standardized methodology as part of donor selection

•Strategies such as plasma or B-cell depletion to minimize the detrimental effect of DSA on transplant outcomes might be considered. However, the benefit of long lasting immune depletion should be carefully considered

•Avoid the selection of DSA positive-CB unit is recommended whenever possible

•Other factors such as cell dose and HLA matching must always be taken in consideration in the donor selection algorithm

Criteria of CB unit choice- EUROCORD >> Patients screening for antibodies against HLA antigens of the cord blood unit >> Look at the number of cells in MAC, RIC: >2.5x107 NC/kg and or >1x105 CD34+/kg >> Look at HLA matches:

0-1 mm better than 2 avoid 3-4 mm Prefer class I mismatches than class II Include HLA C typing, avoiding C mismatches Allele typing of HLA -A and –B (++ in case of 4/6 CBU)

>> Then adapt to graft indication: Malignant diseases: cell dose is the best prognostic factor because HLA

differences reduce relapse (GVL) Non malignant diseases: increase cell dose (>4.0x107 NC/kg ) and find the best

HLA match If the criterion for the minimum number of cells for a single UCBT is not achieved,

a double UCBT should be considered >> Other considerations, if several CBU are available consider:

Cord Blood Bank accreditation status and location ABO compatibility NIMA and KIR status

Acknowledgments

EBMT, CIBMTR Transplant Centers

(Data Managers, Nurses and Physicians) Cord Blood Banks and Netcord

Eliane Gluckman MD FRCP Project Leader

Vanderson Rocha MD, PhD

Scientific Director Annalisa Ruggeri, MD

Agnès Devergie, MD

Federica Giannotti , MD

Myriam Pruvost, PA

Fernanda Volt, MT Chantal Kenzey Data Manager

EUROCORD TEAM 2012-2013

Erick Xavier, MD