Allogeneic hematopoietic stem cell transplantation (allo-HCT) remains one of the few potentially curative treatment options for patients with leukemia and other hematologic malignancies.1,2 By replacing the patient's diseased blood-forming system with healthy donor stem cells, transplantation restores blood formation and establishes a donor-derived immune system capable of recognizing and eliminating residual leukemia cells through the graft-versus-leukemia (GVL) effect. 3,4
Advances in conditioning regimens and GVHD prophylaxis, particularly the adoption of post-transplant cyclophosphamide (PTCy), have broadened the applicability of allo-HCT and enabled the safer use of alternative donors, including haploidentical and mismatched unrelated donors (MMUDs). 5,6
Yet relapse remains the leading cause of treatment failure, affecting approximately 25–30% of transplant recipients. 7,8 When relapses occur, treatment commonly involves donor lymphocyte infusion (DLI) or a second allogeneic transplant. 9
Not all relapses are biologically the same. One important mechanism is HLA loss, in which leukemia cells lose the mismatched HLA haplotype, thereby evading recognition by the donor immune system. 10,11 By losing the mismatched HLA haplotype, leukemic cells can evade donor T-cell recognition and escape the GVL effect. Therefore, the EBMT Acute Leukemia Working Party recommends assessing HLA loss before DLI. 12,13 Despite its clinical importance, routine HLA loss testing remains limited by the lack of standardized, readily available assays. 14
To better understand this implementation gap, researchers conducted an international survey of transplant physicians and, in parallel, evaluated an NGS-based HLA loss assay in patients who relapsed after haploidentical or MMUD transplantation. The findings provide valuable insights into current HLA loss testing practices and highlight how integrated NGS-based approaches may help bring guideline recommendations into routine clinical practice.
The international survey revealed considerable variation in post-transplant relapse management. While DLI was the preferred strategy for molecular relapse, most clinicians favored a second allogeneic transplant for hematologic relapse. Most respondents (85.7%) would choose a different donor rather than reuse the original donor. Despite the recognized role of HLA loss in guiding these decisions, routine testing remained uncommon, with only 21% testing before a second transplant and 17.7% before DLI. Testing rates were consistently low across center types and geographic regions. 15
Despite recommendations to assess HLA loss at the time of relapse, routine implementation remains limited. According to the survey, the most frequently reported barriers to HLA loss testing were cost (88.1%), limited assay availability (87.8%), and long turnaround times (44.5%). These findings highlight the practical challenges laboratories face when implementing HLA loss testing and underscore the need for testing solutions that are accessible, standardized, and easily integrated into routine workflows. 15
To complement the physician survey, the researchers evaluated an NGS-based indel chimerism approach using the One Lambda Devyser Chimerism and Devyser HLA Loss assays in a retrospective cohort of 36 samples from individuals who relapsed after haploidentical or MMUD transplantation. The study investigated the ability of the approach to distinguish classical relapse from HLA loss-mediated relapse, while also exploring its potential to address some of the practical barriers to HLA loss testing identified in the survey.
The approach combines two complementary assays. One Lambda Devyser Chimerism establishes genome-wide chimerism using indel markers across 16 chromosomes, while Devyser HLA Loss focuses on the HLA region of chromosome 6 using indel markers spanning HLA-A to HLA-DP with additional markers on the chromosome 6 p- and q-arms. By comparing the two, laboratories can distinguish classical relapse, in which chimerism remains consistent across the genome, from HLA loss-mediated relapse, in which reduced chimerism is observed specifically within the HLA region.
In the study, the assay successfully identified HLA loss, with representative findings confirmed by independent HLA typing. Beyond its analytical performance, the authors highlight several practical advantages of this NGS-based approach. Other laboratory methods used for HLA loss detection each have distinct technical limitations. Some require leukemic cell purification, while others are limited by marker availability or rely on separate HLA typing workflows. 16,17 In contrast, the NGS-based indel chimerism approach evaluated in this study integrates HLA loss assessment directly into routine chimerism monitoring, without requiring a separate HLA typing run while supporting rapid turnaround. 18 These features have the potential to help overcome practical barriers, such as assay availability and workflow complexity, identified in the physician survey.
The findings highlight a clear opportunity to improve post-transplant relapse management. Although the clinical importance of HLA loss is increasingly recognized, routine testing remains limited by practical barriers. As the use of haploidentical and mismatched donor transplantations continues to expand, making HLA loss testing practical for routine clinical use will become increasingly important.
By making HLA loss testing more accessible, laboratories can provide clinicians with timely information that may support donor selection and inform decisions around donor lymphocyte infusion in post-transplant relapse management.
Learn more about Devyser HLA Loss and discover how it can help simplify HLA loss testing in your laboratory.
Disclaimer: Devyser HLA Loss is for research use only (RUO). Not for use in diagnostic procedures.