Recurrent genetic HLA loss in AML relapsed after matched unrelated allogeneic hematopoietic cell transplantation

Abstract

Immune evasion is a hallmark of cancer and a central mechanism underlying acquired resistance to immune therapy. In allogeneic hematopoietic cell transplantation (alloHCT), late relapses can arise after prolonged alloreactive T-cell control, but the molecular mechanisms of immune escape remain unclear. To identify mechanisms of immune evasion, we performed a genetic analysis of serial samples from 25 patients with myeloid malignancies who relapsed ≥1 year after alloHCT. Using targeted sequencing and microarray analysis to determine HLA allele-specific copy number, we identified copy-neutral loss of heterozygosity events and focal deletions spanning class 1 HLA genes in 2 of 12 recipients of matched unrelated-donor HCT and in 1 of 4 recipients of mismatched unrelated-donor HCT. Relapsed clones, although highly related to their antecedent pretransplantation malignancies, frequently acquired additional mutations in transcription factors and mitogenic signaling genes. Previously, the study of relapse after haploidentical HCT established the paradigm of immune evasion via loss of mismatched HLA. Here, in the context of matched unrelated-donor HCT, HLA loss provides genetic evidence that allogeneic immune recognition may be mediated by minor histocompatibility antigens and suggests opportunities for novel immunologic approaches for relapse prevention.

Graphical abstract

Figure 1.

Late relapse study design and driver mutation analysis. (A) Time to relapse (in years) after alloHCT for AML, myelodysplastic syndrome (MDS), or myeloproliferative disease between 2001 and 2015 at our institution. (B) Schematic of trio targeted sequencing and microarray analysis for 25 patients. (C) Scatter plot showing variant allele frequency (VAF) at time of paired pretransplantation and relapse samples. (D) Alterations in myeloid driver mutations at relapse. Mutations were categorized as lost if they were present at ≥0.02 VAF pretransplantation and absent or <0.02 VAF at relapse. Similarly, mutations were categorized as gained if they were absent or <0.02 VAF pretransplantation and present at ≥0.02 VAF at relapse. CGH, comparative genome hybridization.

Figure 2.

HLA loss at posttransplantation relapse. (A-C) B allele frequency of mismatched SNPs for each matched class 1 HLA gene in pretransplantation (gray) or relapse (red) specimens (left) and copy-number analysis of chromosome 6p (right). (A) The relapse specimen with 20% myeloblast percentage harbors a subtle allelic imbalance of HLA-A (left). Analysis of sorted myeloblasts confirms loss of HLA-A in the relapsed leukemic cells (right). (B) The relapse specimen harbors allelic imbalance of all class 1 HLA genes as a result of copy-neutral loss of heterozygosity across chromosome 6p (right; supplemental Figure 2). (C) The relapse specimen harbors allelic imbalance of HLA-B, which was the result of a 1-Mb deletion spanning HLA-B and HLA-C. (D) Clinical and sample characteristics for cases with relapse-specific HLA loss. cGVHD, chronic graft-versus-host disease; MAC, myeloablative conditioning; RIC, reduced-intensity conditioning; URD, unrelated donor.