Sensitive NPM1 mutation quantitation in acute myeloid leukemia using ultradeep next-generation sequencing in the diagnostic laboratory

Abstract

Context.—Detection of measurable residual disease after therapy is an important predictor of outcome in acute myeloid leukemia. Objective.—To investigate the feasibility of using next-generation sequencing (NGS) in the diagnostic laboratory to perform quantitative NPM1 mutation assessment using ultradeep (approximately 300 0003–500 0003) sequencing (NGS-qNPM1) as a method of assessing residual disease burden in patients with acute myeloid leukemia. Design.—A flexible NGS-based assay for the detection and quantitation of NPM1 mutations was developed by polymerase chain reaction amplification of target DNA sequences, sequencing on an Illumina (San Diego, California) MiSeq, and analyzing data w..