Measurable residual disease detection by high-throughput sequencing improves risk stratification for pediatric B-ALL

Abstract

Early response to induction chemotherapy is an important prognostic factor in B-lymphoblastic leukemia (B-ALL). Here, we compare high-throughput sequencing (HTS) of and genes vs flow cytometry (FC) for measurable residual disease (MRD) detection at the end of induction chemotherapy in pediatric patients with newly diagnosed B-ALL. Six hundred nineteen paired pretreatment and end-of-induction bone marrow samples from Children's Oncology Group studies AALL0331 (clinicaltrials.gov #NCT00103285) (standard risk [SR]; with MRD by FC at any level) and AALL0232 (clinicaltrials.gov #NCT00075725) (high risk; with day 29 MRD <0.1% by FC) were evaluated by HTS and FC for event-free (EFS) and overall survival (OS). HTS and FC showed similar 5-year EFS and OS for MRD-positive and -negative patients using an MRD threshold of 0.01%. However, there was a high discordant rate with HTS identifying 55 (38.7%) more patients MRD positive at this threshold. These discrepant patients have worse outcomes than FC MRD-negative patients. In addition, the increased analytic sensitivity of HTS permitted identification of 19.9% of SR patients without MRD at any detectable level who had excellent 5-year EFS (98.1%) and OS (100%). The higher analytic sensitivity and lower false-negative rate of HTS improves upon FC for MRD detection in pediatric B-ALL by identifying a novel subset of patients at end of induction who are essentially cured using current chemotherapy and identifying MRD at 0.01% in up to one-third of patients who are missed at the same threshold by FC.

Clinical benefit of a high‐throughput sequencing approach for minimal residual disease in acute lymphoblastic leukemia

“…Advantages/disadvantages of HTS screening and MRD monitoring have been discussed previously, particularly in tandem with flow cytometry MRD monitoring . HTS MRD monitoring may lead to assessment at different time points post treatment as well as potentially in blood samples . In the imminent future, it is likely that HTS will be the technique of choice for both screening and follow‐up MRD, but at a minimum, HTS needs to be adopted by MRD laboratories for screening in cases where traditional methods have failed.…”

“…The “gold standard” molecular MRD assessment is based on the allele‐specific amplification of immunoglobulin and T‐cell receptor gene (IG/TCR) rearrangements followed by real‐time quantitative PCR (RT‐qPCR) performed on bone marrow (BM). Recent studies have shown the applicability and potential benefits of a high‐throughput sequencing (HTS) approach to MRD measurement . We aimed to utilize a targeted HTS strategy on diagnostic/relapse BM to identify novel clonal rearrangements in patients proving difficult to analyze using conventional screening.…”

“…Recent studies have shown the applicability and potential benefits of a high-throughput sequencing (HTS) approach to MRD measurement. 6,7 We aimed to utilize a targeted HTS strategy on diagnostic/relapse BM to identify novel clonal rearrangements in patients proving difficult to analyze using conventional screening. We present detailed results on an illustrative cohort of six patients, demonstrating the feasibility of HTS to provide timely markers to use in subsequent RT-qPCR in cases where conventional molecular screening failed.…”

Advances in measurable residual disease monitoring for adult acute lymphoblastic leukemia

“…A correct interpretation requires substantial expertise and familiarity on the part of the interpreting pathologist and it is limited by the potential for false‐positive or false‐negative results especially with post‐induction bone marrow with normal blasts (so called “hematogones”) or too few blasts, respectively. Standard (eg, steroids) and new monoclonal antibody (CD19, 20, 22)‐based treatments can also influence the detectability of residual blasts as it can cause marker shifts . Another major limitation is the lack of standardization of MFC across US, especially in adult ALL although some evidence exists in the pediatric realm, whereas in Europe extensive efforts have made significant progress, again in the pediatric population .…”

“…This technique relies on the high‐level multiplex PCR capability of NGS to allow the design of balanced primers that simultaneously amplify all possible combinations of the IGH/TR rearrangements . NGS has been compared to MFC as well as RT‐qPCR with NGS showing better sensitivities and discordant results favoring NGS. However, just like MFC, it is limited by the requirement of pretreatment diagnostic samples to obtain an index sequence for IGH/TR MRD detection.…”