RNA Fusions and Their Association with DNA Alterations in Myeloid Neoplasia Patients Identified By a Single Tube Multimodal Comprehensive Genomic Profiling Test

• Published in: Blood 2023-11, Vol.142 (Supplement 1), p.1556-1556
• Main Authors: Krawczyk, Michal, Zelinger, Lina, Wong, Cynthie, Nam, Hyunjun, Thomas, Brad, Montgomery, Nathan D, Lyle, Derek D, Lopez-Diaz, Fernando
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood-2023-190505

Background: Recent updates into NCCN professional guidelines have included adding several genomic biomarkers for myeloid disorders. Detecting SNVs, indels, select CNVs and genomic rearrangements in a single comprehensive genomic profiling test is invaluable for clinical care. Moreover, WHO recognizes 23 genomic rearrangements or fusions which define subclasses of AML, MDS/MPN and related neoplasms, and their detection is essential for patient management. Here we present joint prevalence data of gene fusions and other genomic alterations in myeloid disorders in a cohort of 312 patients analyzed by a CLIA grade single-tube NGS assay capable of concurrent analysis of DNA and RNA alterations. Methods: Total nucleic acid (TNA) from bone marrow or peripheral blood was analyzed by a CLIA grade custom amplicon-based multimodal NGS test reporting DNA mutations in 126 genes and gains/losses in 17 genes by DNA-seq, and RNA fusions from 40 genes by RNA-seq. Libraries were sequenced on a NovaSeq6000 instrument, and fusions were called with in-house developed BI pipeline using the distribution of AI-assisted fusion confidence scores to improve the signal to noise discrimination for fusion calls before validation. Deidentified patient data was used according to an IRB approved protocol. Results: Analytical validation of RNA fusion calling against FISH and Sanger-seq in 74 hematologic disorder samples demonstrated 96.7% sensitivity and 98.2% specificity with 100% reproducibility. This improved fusion detection module was added to our CLIA validated NGS assay, which at tumor purity of >20%, already detects SNVs (VAF >3%), Indels (3%) and copy number gains/losses with a sensitivity of 98.3%, 98.9 % and 96 % for SNV, Indels and copy number gains/losses, respectively and a specificity of >99.9% for SNV/Indels and 98.4% for gene gains/losses. Next, data from a total of 789 patients serially tested with this assay was used to study the distribution of myeloid fusion events in community cases which included 312 adult patients with confirmed/suspected myeloid disorders including AML, CML, MDS/MPN and other disorders while 477 patients had a diagnosis of lymphoid leukemias. Fifty five (55) % of the Myeloid leukemia patients were male and 45% female, with a median age of 67.5 (22-87) and 71 (22-89) years, respectively. 27% (84/312) of them presented a gene fusion, of which in 21% of cases it involved a gene from WHO/NCCN fusion gene recommendations. Prevalence for comm