Assessment Of A High-Throughput Sequencing Assay For Measurable Residual Disease (Mrd) Monitoring In Patients With T-Cell Malignancies

(MRD) Monitoring in Patients with T-Cell Malignancies J. Tung1, C. Ho2, J. Zehnder2, B. Zhang2 1Stanford University, Mountain View, CA; 2Stanford Healthcare, Stanford, CA. Introduction: High-throughput sequencing of the T-cell receptor (TCR) beta and gamma loci is becoming more widely utilized due to its high sensitivity, specificity, and versatility in the diagnosis of T-cell malignancies. Application of these technologies for tracking disease burden can be valuable in detecting recurrence, determining response to therapy, guiding future management of patients, and establishing endpoints for clinical trials. In this study, we assessed the performance of a commercially available highthroughput sequencing assay for determining residual disease burden in patients with various T-cell malignancies receiving care at our institution. Methods: Peripheral blood samples from 56 patients previously diagnosed with various T-cell malignancies were sequenced with the commercially available LymphoTrack T-cell MRD assay (InvivoScribe, Inc), which includes the TRB and TRG assay kits, internal and low positive controls, and the LymphoTrack MRD analysis software. Samples were sequenced and analyzed according to the manufacturer’s recommendations. Test performance characteristics including linearity, analytic sensitivity, specificity, and precision were assessed with both contrived and patient samples. The accuracy of the assay was further assessed by tracking 96 previously identified TRB clones and 130 previously identified TRG clones, and comparing results with concurrent MRD assessment by the clonoSEQ assay (Adaptive Biotechnologies). Results: The LymphoTrack T-cell MRD assay demonstrated excellent test performance characteristics for the DNA inputs tested. Assessment of intrarun and inter-run precision revealed coefficients of variation averaging less than 20% for samples containing 1,000 or more T-cell equivalents, whereas greater variances were seen for samples containing fewer than 100 T-cell equivalents. Both TRB and TRG assays reliably detected tracked clonotypes down to an order of 10 T-cell equivalents. Tracked clonotypes were also found to be highly specific to malignant cells when interrogated in pooled normal controls. Despite some notable differences with the clonoSEQ assay in the ability to detect certain rearrangements, MRD status was highly concordant with the LymphoTrack T-cell MRD assay at the sample level. Conclusions: High-throughput sequencing of the TCR beta and gamma loci is a highly sensitive, specific, precise, and accurate method for determining disease burden for various T-cell malignancies and can be readily implemented in molecular diagnostic laboratories capable of performing high-throughput sequencing.