Measurable residual disease monitoring for patients with acute myeloid leukemia following hematopoietic cell transplantation using error corrected hybrid capture next generation sequencing.

Vidya Balagopal,Lauren L Ritterhouse,Pankhuri Wanjari,Wenjun Kang,Chao Jie Zhen,Andrew Hantel,Wendy Stock,Sabah Kadri,Jeremy P Segal,George Steinhardt,Alfons Navarro

doi:10.1371/journal.pone.0224097

Abstract

Improved systems for detection of measurable residual disease (MRD) in acute myeloid leukemia (AML) are urgently needed, however attempts to utilize broad-scale next-generation sequencing (NGS) panels to perform multi-gene surveillance in AML post-induction have been stymied by persistent premalignant mutation-bearing clones. We hypothesized that this technology may be more suitable for evaluation of fully engrafted patients following hematopoietic cell transplantation (HCT). To address this question, we developed a hybrid-capture NGS panel utilizing unique molecular identifiers (UMIs) to detect variants at 0.1% VAF or below across 22 genes frequently mutated in myeloid disorders and applied it to a retrospective sample set of blood and bone marrow DNA samples previously evaluated as negative for disease via standard-of-care short tandem repeat (STR)-based engraftment testing and hematopathology analysis in our laboratory. Of 30 patients who demonstrated trackable mutations in the 22 genes at eventual relapse by standard NGS analysis, we were able to definitively detect relapse-associated mutations in 18/30 (60%) at previously disease-negative timepoints collected 20–100 days prior to relapse date. MRD was detected in both bone marrow (15/28, 53.6%) and peripheral blood samples (9/18, 50%), while showing excellent technical specificity in our sample set. We also confirmed the disappearance of all MRD signal with increasing time prior to relapse (>100 days), indicating true clinical specificity, even using genes commonly associated with clonal hematopoiesis of indeterminate potential (CHIP). This study highlights the efficacy of a highly sensitive, NGS panel-based approach to early detection of relapse in AML and supports the clinical validity of extending MRD analysis across many genes in the post-transplant setting.

Highlights

Due to the high mortality rate and frequency of treatment failures, improved methods of disease status monitoring are clearly needed for acute myeloid leukemia (AML) patients during therapy
To assess the performance and utility of next-generation sequencing (NGS)-based measurable residual disease (MRD) detection in patients with AML following hematopoietic cell transplantation (HCT), we designed a retrospective case-control study taking advantage of samples and data collected during routine clinical engraftment analysis of bone marrow (BM) and peripheral blood (PB) using short tandem repeat (STR) polymerase chain reaction (PCR)
To identify patients suitable for inclusion in the study, we reviewed our results from the University of Chicago Medicine (UCM) Molecular Diagnostics Laboratory post-transplant engraftment testing from 2014–2018, with approval from the University of Chicago Institutional Review Board

Summary

Introduction

Due to the high mortality rate and frequency of treatment failures, improved methods of disease status monitoring are clearly needed for acute myeloid leukemia (AML) patients during therapy. MRD flow cytometry for AML can require highly multiplexed analysis and is often complicated by variable sensitivity due to patient-specific marker expression profiles. These analyses can be subject to inter-assay and inter-operator variability[2,3,4,5]. STR-based assays do not measure recurrent disease but instead offer a percentage of recipient DNA as a surrogate measure for recurrence. This impairs specificity, as non-malignant recipient cell lineages may be present in various sample types and conditions without disease relapse[10]

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