Performance Evaluation of a Custom Myeloid Assay on the Genexus Integrated Sequencer from a Wide Spectrum of Clinical...

A custom Genexus myeloid assay (CMA) underwent a technical evaluation for detection of variants from both DNA and RNA in a single assay format. The custom assay was initially verified with commercial DNA and RNA controls containing known myeloid variants. Seventy-five patient specimens with various DNA and RNA variants were selected for replicate testing. The CMA generated consistent data on two Genexus sequencers, with occasional samples failing quality metrics randomly. All 22 control DNA variants were detected, with 95% reported as key. Sensitivity and positive predictive value for clinical variants were 95.60% and 97.60%, respectively, increasing to 98.63% and 99.65%, respectively, for allele frequencies ≥5%. FLT3 duplications were consistently detected at lower expected frequencies. Low-level, false-positive key variants were mostly recurrent and filterable. Fusion calling sensitivity was 100% for control and clinical RNA samples, with 97.09% of replicates reporting expected fusions. The CMA reliably detects key DNA and RNA variants in myeloid specimens within 24 hours on the Genexus platform. Variants and fusion transcripts were identified with high sensitivity and minimal nucleic acid input, providing a rapid and precise workflow for reporting clinically relevant myeloid disease variants.