Importance of conditioning regimen intensity, MRD positivity, and KIR ligand mismatch in UCB transplantation

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Minimal residual disease (MRD) is an important factor for the outcomes of allogeneic hematopoietic cell transplantation (alloHCT).1, 2 Milano et al.3 reported that umbilical cord blood transplantation (UCBT) resulted in lower relapse rates in acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS) patients with MRD compared with unrelated donor transplantation (URDT). We have shown that intensity of conditioning regimen (that is, myeloablative conditioning (MAC) vs reduced-intensity conditionings (RIC) is critical to overcome the negative effect of MRD before UCBT.4

Donor natural killer (NK) cells are an important part of the GvL effect after alloHCT and its specificity is determined by the killer-Ig receptors (KIRs) on donor NK cells for recipient MHC class I.5 Alloreactive donor-derived (NK) cells improved survival after KIR mismatched URDT in patients with AML.6 Interestingly, this effect has been controversial after UCBT.7, 8, 9, 10 In this study, we analyzed our AML patients in morphologic CR who gave written informed consent for relevant local Institutional Research Board approved UCBT studies between 1 January 2004 and 31 December 2015 to evaluate the effects of MRD, conditioning intensity, and KIRL mismatching on UCBT outcomes.

Allele-level molecular typing was performed for class I HLA-A, -B and -C by sequence-based typing (SBT) using Visible Genetics (Suwanee, GA, USA) reagents and sequencer for HLA typing. Class II HLA DRB1 molecular typing was performed by sequence-specific primers (SSPs) using One Lambda (Canoga Park, CA, USA) reagents. The presence of KIR ligand (KIRL) mismatching in the graft-versus-host (GvH) direction was assigned as initially described.6 For the analysis of acute GvH disease (GvHD), transplantation-related mortality (TRM), relapse and survival only the KIRL assignment of the dominant engrafting unit was considered. Selection criteria for UCB graft regarding HLA match status, total nucleated cell counts, and our MAC and RIC regimens were previously described.4, 11 The MAC and RIC groups were then classified by the presence (MRD+) or absence (MRD−) based upon flow cytometric evidence of AML. For this study, we categorized cases as MRD+ if they included an atypical population of myeloblasts, comprising at least 0.5% of leukocytes, that had an immunophenotype that was distinct from normal myeloblasts, using the markers included in the panel.

In Tables 1A, 1B and 1C comparison of categorical factors was analyzed by the χ2 or Fisher’s exact test. Continuous variables were analyzed by the general Wilcoxon test. Cumulative incidence was used to estimate relapse, treating non-relapse mortality (NRM) as a competing risk. Cumulative incidence was similarly used to estimate NRM, treating relapse as a competing risk. Kaplan–Meier curves were used to estimate the probability of overall survival (OS) and leukemia-free survival (LFS) through 5 years post transplant. The log-rank test was used to complete the comparisons. Cox regression was used to look at the independent effect of residual disease on survival and LFS. Fine and Gray regression was used to look at the independent effect of residual disease on relapse and NRM. Factors considered in regression analyses were detectable residual disease (MRD+ versus MRD-), age, disease status at transplant (CR1, diagnosis to transplantation<6 months versus CR1 diagnosis to transplantation6 months versus CR2+, diagnosis<1 year versus CR2+, diagnosis1 year), recipient CMV serostatus (negative versus positive), Karnofsky performance score (<90 versus 90–100), comorbidity score (low risk versus intermediate risk versus high risk), cytogenetic risk (good/intermediate versus poor), number of donors (single UCB versus dUCB), use of anti-thymocyte globulin (ATG) (no versus yes), central nervous system leukemia pre-transplant (yes versus no), white blood cell count at diagnosis (<20 × 109/L versus 20) and KIRL match (match versus mismatch). KIRL match was evaluated by the predominant UCB unit only among dUCBT. The predominant UCB unit was defined as the unit showing 70% chimerism by day 100 post HCT or 70% by day 21 for patients who were deceased prior to day 100. Factors were tested for inclusion into the regression models by a forward stepwise selection process in which entry required an individual P-value of <0.25 and inclusion required a P-value <0.10 to remain in the model. Residual disease status was forced into all models. All reported P-values were two-sided. All analyses were performed using SAS 9.3 (SAS Institute, Inc., Cary, NC, USA) and R version 3.3.1.

Table 1A Relapse in MRD+ AML in UCB HCT
Table 1B Inferior LFS was observed in MRD+ AML in RIC UCB HCT
Table 1C Inferior OS was observed in MRD+ AML in RIC UCB HCT

Of 140 patients, 55 and 85 received MAC and RIC, respectively. Eleven of the 55 (20%) MAC patients and 8 of the 85 (9%) RIC patients were MRD+. In both the MAC and RIC groups, MRD+ patients had similar characteristics with MRD− patients.

In univariate and multivariate analyses, MRD positivity was negatively affected relapse, LFS and OS in RIC UCBT. (Tables 1A, 1B and 1C; Figure 1a–c). NRM was not significantly influenced by MRD+ in either the MAC or RIC group (Figure 1d).

Figure 1
figure1

(a) Cumulative incidence of relapse by conditioning regimen intensity and MRD status. (b) Leukemia-free survival by conditioning regimen intensity and MRD status. (c) Overall survival by conditioning regimen intensity and MRD status. (d) Non-relapse mortality by conditioning regimen intensity and MRD status.

KIRL mismatch was observed in 9 (24%), 1(10%), 12(19%) and 0 (0%) in the MAC MRD−, MAC MRD+, RIC MRD−, and RIC MRD+ groups, respectively. The effect of KIRL match status was analyzed only in the MRD− group given only one patient was KIRL mismatched in the MRD+ group. KIRL mismatch had no effect on relapse by 5 years (26% (95% CI: 6–45%) vs 34% (95% CI: 23–45%), P=0.50) in KIRL mismatch vs matched, respectively. There was no competing risk of NRM, P=0.26. Relapse rates at 5-year after MAC and RIC were 11% (95% CI, 0–29%) and 38% (95% CI, 9–68%) in KIRL mismatched transplants vs 25% (95% CI, 9–41%) and 39% (95% CI, 25–54%) in KIRL-matched transplants.

It is well-known that immune reconstitution, especially functional T cell recovery is delayed in UCBT.12, 13 However, numerous studies have already shown that single or dUCBT has a strong GVL effect, even in high risk AML,14 comparable with other graft types. Ponce et al showed that the 3-year relapse risk was decreased after dUCB (9%) compared with HLA full-matched URDT (23%) and with HLA 7/8 matched URDT (20%), P=0.037).15 Brunstein et al.16 reported that the risk of relapse was significantly higher in matched related donor transplantation (RDT; RR: 3.64, 95% CI, 1.1.89–7.01, P<0.01), in matched URDT (RR: 3.36, 95% CI, 1.73–6.53, P=0.02) and in mismatched URDT (RR: 2.61, 95% CI, 1.18–5.77, P=0.02) compared with dUCBT. Our group also showed a lower risk of relapse at 3-years after MAC UCBT (19%, 95% CI 9–28%) compared with MAC matched RDT (35%, 95% CI 22–48%), P=0.05.13 Decrease in relapse after UCBT might in part result from rapid recovery of NK cells.13

UCBT outcomes might be superior to matched or mismatched URDT in MRD+ patients.3 In the current study, we evaluated the effect of MRD on the outcomes of UCBT. Study limitations include a small number of MRD+ patients (an indication of our stringent RIC HCT requirements) and limited availability of molecular markers. Although UCBT patients’ 1-year LFS and OS were >50% after both RIC and MAC, survival was significantly worse in MRD+ patients receiving RIC, primarily due to increased relapse risk. Interestingly, MRD+ patients receiving MAC UCBT had similar LFS and OS compared with MRD− AML patients. GVL after UCBT, as observed in other donor types, may not be adequate enough to overcome the negative effect of MRD+ in patients receiving RICT.

In our cohort of MRD− patients, KIRL mismatching had no effect in decreasing relapse in these patients. Of the 4 prior studies evaluating KIR mismatch in UCBT;7, 8, 9, 10 only one study showed a positive effect:9 the 2-year cumulative incidence of relapse was 37% for the KIRL-compatible group (n=149) compared with 20% for the KIRL -incompatible group (n=69), P=0.03 in 94 patients with AML and 124 patients with acute lymphoblastic leukemia (ALL).9 Different from our study, all patients received a single unit UCBT and most patients received ATG that seemed to be a risk factor for relapse in that cohort. Similar to our current study, conditioning intensity was the most important risk factor for relapse. Our group’s prior study (257 patients, between 1998 and 2006), showed that KIRL mismatching in GvH direction was associated with a higher acute GVHD, and thus lower survival after only RIC UCBT.10 In contrast, relapse rates in this study were not affected by KIRL mismatching, albeit only evaluated in the MRD− group. Garfall et al also showed no effect of KIRL mismatching (n=35) on relapse in 80 patients with various hematologic malignancies after dUCBT.7

In summary, our study confirms and highlights many important findings: (1) OS is substantial for those receiving MAC UCBT and similar for those MRD- AML patients receiving RIC; (2) MRD status is a major determinant in long-term outcome with the greatest influence in the RIC setting; (3) KIRL mismatch had no impact on relapse in MRD- patients. The impact of KIRL mismatch needs to be investigated further by larger scale studies in regard to MRD status in patients with AML undergoing UCBT.

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Acknowledgements

Author contributions

CU: conceived the study idea, collected the data, analyzed the data, searched the literature, wrote the article and edited the article. JM, MAL, DW: conceived the study idea, analyzed the data, wrote the article and edited the article. TD: collected the data, analyzed the data, wrote the article and edited the article. JM, MAL, DW: conceived the study idea, analyzed data, wrote the article and edited the article. TD: collected the data, analyzed the data, wrote the article and edited the article. CB, AR, SY, NB, SC, EW: analyzed the data, wrote the article and edited the article.

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Correspondence to C Ustun.

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Ustun, C., Brunstein, C., DeFor, T. et al. Importance of conditioning regimen intensity, MRD positivity, and KIR ligand mismatch in UCB transplantation. Bone Marrow Transplant 53, 97–100 (2018) doi:10.1038/bmt.2017.212

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