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

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.

Molecular Minimal Residual Disease Detection in Acute Myeloid Leukemia

“…By utilizing the UMIs, mutations at a VAF of <0.1% could be reliably detected in 22 frequently mutated genes in AML. With this improved sensitivity, previously undetected residual mutations associated with an eventual relapse were found in 18 out of 30 AML patients [ 57 ].…”

Molecular testing for acute myeloid leukemia

“…In recent years, NGS assay has been used in monitoring minimal (measurable) residual disease (MRD). Using unique molecular identification (UMI) barcode, NGS sequencing error and noise background have been reduced, which makes NGS assay a good tool in monitoring MRD [9][10][11][12][13][14][15][16] . NGS assay has also been used in single cell sequencing, which is a powerful tool for revealing information about clonal evolution 17,18 .…”

Sequencing-Based Measurable Residual Disease Testing in Acute Myeloid Leukemia

“…In another recent publication, Balagopal et al (2019) used a UMI error corrected hybrid capture enrichment panel targeting 22 recurrently mutated genes and tested 30 post-transplant AML patients who were negative for MRD by conventional short tandem repeat (STR)-based testing. STR testing is frequently used to monitor post bone marrow transplant patients in the clinical setting and works by comparing differences in highly polymorphic alleles between recipient and donor bone marrow via PCR and capillary-based fragment sizing.…”