Who is the best donor for a related HLA haplotype-mismatched transplant?

Human leukocyte antigen (HLA) haplotype-mismatched transplants from related donors are increasing.^1-3  Several approaches were used including ex-vivo T-cell depletion ^4,5  granulocyte colony-stimulating factor,⁶  posttransplant cyclophosphamide ^7,8  and pre- and posttransplant antithymocyte globulin (ATG).⁹  Kasamon et al⁸  reported degree of HLA disparity on the HLA-mismatched haplotype is not correlated with survival after nonmyeloablative transplants with posttransplant cyclophosphamide. Similarly, we found that number of HLA mismatches was not correlated with survival after T-cell–replete transplants. These data suggest variables other than HLA matching on the mismatched HLA haplotype should be studied to determine their impact on transplant outcomes.^10,11  Donor age and gender correlate with outcomes after transplants from HLA-identical sibling donors and HLA-matched unrelated donor.^12,13  Matching for noninherited maternal antigens (NIMAs) is reported to correlate with incidence of graft-versus-host disease (GVHD) after HLA-identical transplants.^14-16  These data suggest the possible impact of donor-related variables correlated with transplant outcomes in the setting of related HLA haplotype-mismatched transplants.

We reported our approach to related HLA haplotype-mismatched T-cell–replete transplants in >1000 subjects during the last 10 years, an approach also used by others.^10,11  During this interval, pretransplant conditioning regimen, graft type, and GVHD prophylaxis have not changed. Consequently, our dataset provides an opportunity to explore donor-related variables that might correlate with transplant outcomes.

A total of 1256 consecutive subjects with hematological neoplasms receiving related HLA haplotype-mismatched transplants May 2002 through February 2013 at Peking University Institute of Hematology were considered. We excluded 32 subjects with cousin donors and 14 subjects who received porcine ATG. The remaining 1210 subjects were enrolled; 713 were previously reported.¹⁰  The Institutional Review Board of Peking University approved this study, and all subjects and donors gave written informed consent. This study was conducted in accordance with the Declaration of Helsinki.

A human leukocyte antigen (HLA)-matched sibling donor was the first choice for allotransplant. If an HLA-matched sibling donor was unavailable, subjects without a suitable, closely HLA-matched, unrelated donor (>8 of 10 matching HLA-A, B, C, DR, and DQ loci and >5 of 6 matching HLA-A, B, and DR loci or whose disease state left insufficient time for an unrelated donor search) were eligible for HLA haplotype-mismatched transplant. Family members were assessed for degree of HLA mismatch. HLA-A and HLA-B typing were performed by intermediate resolution DNA typing, whereas HLA-DRB1 typing was performed by high-resolution DNA techniques. Each subject received a graft from a family member sharing one HLA haplotype with the recipient but differed to a variable degree for the HLA-A, B, and DR antigens of the unshared HLA haplotype.

Pretransplant conditioning was with cytarabine (4 g/m E+2 per day, days –10 to –9), busulfan (4 mg/kg per day, orally days –8 to –6 before January 2008 and 3.2 mg/kg per day, intravenously days –8 to –6 thereafter), cyclophosphamide (1.8 g/m E+2 per day, days –5 to –4), semustine (250 mg/mE+2, day –3), and rabbit ATG (thymoglobulin; Imtix Sangstat, Lyon, France, days –5 to –2). Before June 2008, subjects received 2.5 mg/kg per day, and thereafter subjects with advanced leukemia (defined below) received 1.5 mg/kg per day. Between December 2010 and May 2012, subjects with standard-risk leukemia (defined below) were randomized to receive 2.5 mg/kg per day or 1.5 mg/kg per day (study registered at www.chictr.org #ChiCTR-TRC-11001761). Overall, 158 subjects received an ATG total dose of 6 mg/kg and 1052 received 10 mg/kg. GVHD prophylaxis was with cyclosporine, mycophenolate mofetil, and short course methotrexate as described.¹⁰  Grafts were granulocyte-colony stimulating factor mobilized bone marrow and blood cells as described.¹⁰ 

Subjects with acute leukemia were categorized as standard-risk if they were in first or second complete remission.¹⁴  Subjects with lymphoma or multiple myeloma were categorized as standard risk if they were in first or second complete remission, partial remission, or had stable disease, as were subjects with chronic myeloid leukemia in first chronic phase and those with myelodysplastic syndrome with <20% bone marrow blasts.¹⁴  Other subjects were classified as high risk.

Transplant outcomes considered were measured in terms of GVHD, nonrelapse mortality (NRM), relapse, progression-free survival (PFS), and survival with the major endpoints of NRM and survival. Cumulative incidences were estimated for engraftment, GVHD, NRM, and relapse to accommodate competing risks. Relapse was a competing risk for NRM, and death from any cause was a competing risk for engraftment, GVHD, and relapse. Probabilities of survival and PFS were estimated by the Kaplan-Meier method. Potential prognostic factors were evaluated in univariate analyses by the log-rank test, with a P value <.05 considered significant. Cox proportional hazards regression was used to identify risk factors associated with outcomes. Multivariate analyses were done for the entire cohort and predefined subgroups. The assumption of proportional hazards for each factor in the Cox model was tested. The test indicated that the proportionality assumptions hold. Variables considered in the multivariate models were donor sex (male vs female, except in the comparison between parental donors), donor age (continuous variable), HLA disparity (3/6 vs 4/6 vs 5/6 or 6/6), donor-recipient sex (match vs mismatch), donor-recipient blood type pair (match vs major mismatch vs minor mismatch), and donor-recipient relationship (father vs mother vs sibling vs children for the entire study population and separate comparison performed in subgroup analysis). Cytomegalovirus (CMV) serological state was not considered as a covariate, because only 1% of subjects were low risk (recipient [R]−, donor [D]−) for CMV reactivity. Killer immunoglobulin-like receptor matching was not analyzed because of too few data. The test indicated that the effect of donor-related factor on relapse is independent of disease type. The final multivariate models were built using a forward stepwise model selection approach with 5% significance level. Final models for each outcome were reported. The end point of the last follow-up for all of survivors was December 1, 2013. The median follow-up of survivors was 1299 days (range; 283-4241 days).

Characteristics of the donors, recipients, and grafts are listed in Table 1. Parental donors account for 60% of the study population. The majority of patient/recipient pairs were cytomegalovirus serology^+/+. The median time to granulocytes >0.5 × 10E+9/L was 13 days (range, 8-49 days) and was 16 days (range, 5-100 days) to platelets >25 × 10E+5/L. A total of 1199 (99%) of subjects had sustained bone marrow recovery. The 3-year cumulative incidences of NRM and relapse were 17% (95% confidence interval [CI] = 15% to 19%) and 17% (95% CI = 15% to 19%). The 100-day cumulative incidences of acute GVHD ≥grade 2 and grade 3 were 40% (95% CI = 37% to 42%) and 12% (95% CI = 10% to 14%), respectively. The 3-year cumulative incidences of total and extensive chronic GVHD were 50% (95% CI = 47% to 53%) and 21% (95% CI = 19% to 24%). The 3-year probabilities of PFS and survival were 67% (95% CI, = 63 −69%) and 70% (95% CI = 67% to 73%).

Overall degree and specific HLA matching or mismatching were not significantly correlated with cumulative incidences of NRM, acute or chronic GVHD, or survival (Table 2).

Younger donors were associated with a lower cumulative incidence of acute GVHD than older donors (Table 2). This association persisted when we considered donor age < or ≥30 years (Table 2). In multivariate analyses, in both models treating donor age as a continuous variable and a binary variable, respectively, donors <30 years old and male donors were independently significantly associated with less NRM and better survival than older or female donors (Table 3; Figure 1). Mother donors were also associated with a higher cumulative incidence of acute GVHD than other pairings in multivariate analyses (Table 3). To determine whether the higher cumulative incidences of acute GVHD associated with older and female donors resulted solely from use of mother donors, we conducted a separate analysis in which mother donors were excluded. In these 909 transplants, we found no significant correlation between donor gender and cumulative incidence of acute GVHD (P = .84). In contrast, donor age <30 years remained significantly associated with less acute GVHD compared with donors ≥30 years with an adjusted hazard ratio (HR) = 0.60 (P < .001).

Family relationship was also significantly correlated with transplant outcomes. Because many families have only 1 child, the most common comparisons we could analyze were grafts from father vs mother, child vs sibling, and of sibling vs parent.

Mother donors were associated with more NRM and acute GVHD and worse survival than father donors in multivariate analyses (Table 3; Figure 2). This adverse impact of mother donors on acute GVHD persisted regardless of whether the recipient was a son (HR = 1.34; 95% CI = 1.07-1.34; P = .03) or a daughter (HR = 1.59; 95% CI = 1.20-2.10; P = .001). In contrast, maternal donors were associated with more NRM and worse survival when the recipient was a son¹⁷  (HR = 2.04; 95% CI = 1.37-3.03; P = .001 and HR = 1.49; 95% CI = 1.09-2.03; P = .01, respectively), but not when the recipient was a daughter. Donor age was not associated with any outcome in this subset analysis. These data favor the use of father donors over mother donors, especially when the recipient is a son.

We also considered outcomes after child vs sibling donors in multivariate analysis, as this situation is obviously confounded by donor age and gender. Offspring donors were associated with less acute GVHD ≥ grade-2 compared with sibling donors (Table 3). In contrast, donor age rather than child vs sibling was more important in the context of NRM and survival (Table 3). Donor gender was not significantly associated with any outcome in this subset analysis.

For many young adults without children, donor choice is between a sibling or a parent. Because we showed better outcomes with fathers vs mother, the critical choice is between a sibling or father donor. In our preliminary analyses, there was no significant difference in outcomes between sibling and father donors (Table 2), and these cohorts were combined in further analyses where father donors comprised >50% of the combined cohort. We next considered whether donor age and/or gender or gender matching were associated with transplant outcomes. Younger donors were associated with less NRM and better survival, whereas female donors (siblings only) were associated with more NRM in multivariate analysis (Table 3). To determine the relative import of donor age and sex on transplant outcomes, we created 4 subsets. Sibling donors <30 years were associated with less acute GVHD ≥grade 2 compared with sibling donors ≥30 years (HR = 0.68; 95% CI = 0.46-0.99; P = .05) or father donors < or ≥30 years (HR = 0.68; 95% CI = 0.48-0.98; P = .04). In contrast, outcomes of transplants from brothers ≥30 years were not significantly different than father donors (data not shown). Sister donors ≥30 years, especially when the recipient was a brother, were associated with more NRM (HR = 1.87; 95% CI = 1.10-3.20; P = .02) and worse survival (HR = 1.59; 95% CI = 1.05-2.40; P = .03) compared with father donors.

The impact of NIMA disparities was studied in 264 subjects who had informative donor, recipient, and parent HLA-typing data. Subjects were divided into 3 subsets: (1) mother to offspring grafts with GvH reaction against inherited father HLA antigens (IPA); (2) offspring to mother grafts with GvH reaction against the NIMAs of offspring; and (3) sibling to sibling grafts, where the siblings are NIMA mismatched but share inherited father HLA antigens. Four models were created to study the NIMA effect as described.¹⁵  Estimates of event frequencies were calculated at 1.5 years because of briefer follow-up (except acute GVHD at 100 days).

In the sibling-only model (N = 53), NIMA-mismatched sibling donors (N = 27) had less acute GVHD ≥grade 2 (19% vs 46%; P = .02) and a trend toward lower cumulative incidence of relapse (7% vs 23%; P = .11) compared with NIPA-mismatched sibling donors (N = 26) (Table 3). Next, in the NIMA/IPA model (N = 89), the subset was divided into a NIMA-targeted subset (N = 34), which included NIMA-mismatched sibling grafts (N = 27) and child to mother grafts (N = 7) and an IPA-targeted subset referred to maternal donors (N = 55). Here, NIMA-mismatched donors and younger donors had less acute GVHD ≥grade 2 (Table 3). Then, in the sibling and parent combined model (N = 238), mother donors (N = 55) and father donors (N = 130) were associated with more acute GVHD ≥grade 2 compared with NIMA-mismatched sibling donors (N = 27; P = .04 and P = .02), but there was no significant difference when compared with NIPA-mismatched sibling donors (P = .93 and P = .58) (Table 3; Figure 3). Finally, the sibling-offspring combined model (N = 79) was divided into subsets including a NIMA-targeted subset (N = 34), which included NIMA-mismatched siblings grafts (N = 27), and offspring to mother grafts (N = 7) and a NIPA-targeted subset (N = 45), which included NIPA-mismatched siblings grafts (N = 26) and offspring to father grafts (N = 19). The NIMA-targeted subset had slightly less acute GVHD ≥grade 2 (23% vs 42%; P = .06). There were no differences in other outcomes, including NRM, chronic GVHD, and survival among any of the above-mentioned subsets regarding NIMA.

Our data confirm degree of HLA disparity on the HLA-mismatched haplotype is not significantly correlated with transplant outcomes among recipients of T-cell–replete–related HLA haplotype-mismatched transplants.^10,11  Whether this is so in all transplant settings requires further study but seems so.^8,19 

In contrast to the lack of impact of HLA disparity on transplant outcomes, we found that older donors were associated with more acute GVHD and that donors >30 years were associated with more NRM and worse survival. Similar findings operate in other transplant settings.^12,13,18  The adverse impact of older donors operates in all subset analyses except the subset of parental donors (Table 3). This effect likely contributes to the lower NRM and better survival of younger sibling donors than parent donors. Why older donors are associated with worse transplant outcomes is uncertain but may reflect age-associated declines in hematopoiesis and immunity.^20,21  However, it is important to recall we report statistically significant associations, which may but need not imply causality.

Donor gender was also significantly associated with transplant outcomes. Transplants from male donors had less NRM, less acute GVHD ≥grade 2, and better survival. This benefit was lost when mother donors were excluded. Similar associations of gender and transplant outcomes are reported in some but not all other transplant settings, especially transplants from parous mother.^8,12,13,16  Some data suggest this effect may reflect immunity to minor histocompatibility antigen encoded genes on the Y chromosome.^22,23  Unfortunately, we have insufficient data on donor parity to explore this further.

Family relationship was also an independent predictor of transplant outcomes. Similar but inconsistent relationships are reported by others.^24-26  Mother donors were associated with more acute GVHD ≥grade 2 compared with father donors similar to some¹⁴  but not other studies.^15,16  The reason for this disparity is uncertain but may reflect differences in several variables, including the pretransplant conditioning, graft composition, posttransplant immune suppression, and others. In the Perugia report,¹⁶  patients who received transplants from mothers had a more favorable risk profile regarding disease state at transplantation, although mother remained a better donor after adjustment in multivariable analysis. Controversial effects of natural killer cell alloreactivity between the European and American study and our study were reviewed in detail by our group.^27,28 

In the sibling-only model and the sibling-parent combined model, our data, consistent with other studies, suggest a tolerance-inducing NIMA effect.²⁹  However, we found no significant impact of NIMA mismatch on NRM, chronic GVHD, relapse, or survival. In a more detailed subset analysis, we found older NIMA-mismatched sibling donors were associated with less acute GVHD ≥grade 2 compared with younger NIPA-mismatched sibling donors. However, there were few subjects in each subset, and the difference was not statistically significant. We also found NIMA-mismatched sibling donors had less acute GVHD ≥grade 2 than father donors even after adjusting for age and gender. In contrast, this NIPA effect did not counterbalance the impacts of donor age and gender discussed above (data not shown). These conclusions should be interpreted cautiously, because of the few subjects in each subset. In the sibling-offspring combined model, the NIMA-targeted subset had a less acute GVHD ≥grade 2 compared with the NIPA-targeted subset. Again, the number of subjects in each cohort was small and analyses were confounded by the shared HLA haplotypes.¹⁵  Furthermore, the immune system of the child is more naive than that of the mother. Other factors may also operate. We were unable to critically analyze child to mother transplants or determine whether donor age or gender are more important than the NIMA effect because of a small sample size.

The strength of our study is the large sample size and relative consistency of transplant variables. Nevertheless, we were unable to analyze some potentially important variables such as CMV serological state and killer immunoglobulin-like receptor disparity.

In conclusion, selection of the best donor in the setting of related HLA haplotype-mismatched T-cell–replete transplants needs to consider donor age, gender, family relationship, NIMA effect, and other variables. Our data suggest choosing young, male, NIMA-mismatched donors is reasonable. Transplants from older mothers and NIPA-mismatched donors should probably be avoided (Table 4). Comparisons of different approaches in the setting of related HLA haplotype-mismatched transplant should probably be adjusted for the variables we identified, which are associated with transplant outcomes.

Professors Robert Peter Gale (Imperial College London), Mei-Jie Zhang (Medical College of Wisconsin), and Nelson Chao (Duke University) kindly reviewed the typescript.

This work was partly supported by grants from Collaborative Innovation Center of Hematology China, the Key Program of National Natural Science Foundation of China (81230013), and Beijing Municipal Science and Technology Commission (nos. Z121107002812033 and Z121107002612035).

Contribution: X.-J.H. designed the study; Y.W. and Y.-J.C. collected data; Y.W., Y.-J.C. and X.-J.H. analyzed the data and wrote the typescript; and all authors contributed to interpretation of the data, preparing the typescript, and approved the final version.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Xiao-Jun Huang, Peking University People’s Hospital, Peking University Institute of Hematology, No 11 Xizhimen South St, Beijing, 100044, China; e-mail: [email protected].