Practical considerations for using RNA sequencing in management of B-lymphoblastic leukemia Malaysia-Singapore acute lymphoblastic leukemia 2020 implementation strategy
Document Type
Article
Publication Date
10-1-2021
Abstract
Despite the immense genetic heterogeneity of B-lymphoblastic leukemia or precursor B-cell acute lymphoblastic leukemia (B-ALL)], RNA sequencing (RNA-Seq) could comprehensively interrogate its genetic drivers, assigning a specific molecular subtype in >90% of patients. However, study groups have only started to use RNA-Seq. For broader clinical use, technical, quality control, and appropriate performance validation are needed. We describe the development and validation of an RNA-Seq workflow for subtype classification, TPMT/NUDT15/TP53 variant discovery, and immunoglobulin heavy chain (IGH) disease clone identification for Malaysia-Singapore acute lymphoblastic leukemia (ALL) 2020. We validated this workflow in 377 patients in our preceding Malaysia-Singapore ALL 2003/ Malaysia-Singapore ALL 2010 studies and proposed the quality control measures for RNA quality, library size, sequencing, and data analysis using the International Organization for Standardization 15189 quality and competence standard for medical laboratories. Compared with conventional methods, we achieved >95% accuracy in oncogene fusion identification, digital karyotyping, and TPMT and NUDT15 variant discovery. We found seven pathogenic TP53 mutations, confirmed with Sanger sequencing, which conferred a poorer outcome. Applying this workflow prospectively to the first 21 patients in Malaysia-Singapore ALL 2020, we identified the genetic drivers and IGH disease clones in >90% of patients with concordant TPMT, NUDT15, and TP53 variants using PCR-based methods. The median turnaround time was 12 days, which was clinically actionable. In conclusion, RNA-Seq workflow could be used clinically in management of B-cell ALL patients.
Keywords
Minimal residual disease, Abnormalities, Polymorphisms, Children, Gene, DUX4
Divisions
fac_med
Funders
Singapore National Medical Research Council Clini-cian Scientist Awards[NMRC/CSA/0053/2013],Singapore National Medical Research Council Clini-cian Scientist Awards[MOH-000277],Children's Cancer Foundation, Singapore,Singapore Tote Board,Goh Foundation, Singapore
Publication Title
Journal of Molecular Diagnostics
Volume
23
Issue
10
Publisher
Elsevier
Publisher Location
STE 800, 230 PARK AVE, NEW YORK, NY 10169 USA