Measurable residual disease detection by high-throughput sequencing improved risk-stratification for pediatric B-ALL.
The initial response to therapy is a key prognostic factor in pediatric B-ALL. This is currently determined as the level of measurable residual disease (MRD) at the end of Induction chemotherapy. Normally determined by flow cytometry (FC), this leads to limitations in the analytic sensitivity and difficulty in clinical standardization of the technique. As well, changes in antigen expression on the lymphoblasts during therapy may obscure detection by FC. High-throughput sequencing (HTS) of the immunoglobulin and T-cell receptor loci may represent a superior methodology for detection of MRD.
Paired pre-treatment and end of induction samples for patients enrolled in front-line childhood B-ALL treatment trials were identified for MRD evaluation. Both FC and HTS were performed on the samples. For HTS, pretreatment samples were assessed for clonality by deep sequencing of the immunoglobulin (IGH@) gene. A dominant sequence determined for identification at end of induction, as an indicator of MRD. The amount of DNA was extrapolated to the equivalent total nucleated cell count, to give a comparable metric to MRD by FC.
607 evaluable samples were studied, and HTC and FC showed good correlation for both event-free and overall survival at an MRD of 0.01%. Samples where MRD was above the threshold by HTS but not FC were found to have an intermediate prognosis, suggesting that it had a higher analytic sensitivity for low levels of MRD. Patients with no trackable rearrangement of the immunoglobulin heavy chain by HTS, suggesting an earlier stage neoplastic transformation, had a poorer prognosis.
HTS required initial access to heavy leukemic involvement to identify an appropriate target sequence, and cannot be used in patients lacking a detectable immunoglobulin or T-cell receptor rearrangement. Turn-around time is also longer than with FC and the technology is currently more expensive.