Abstract 7171: Comparison of clinical next generation sequencing and whole genome sequencing for acute myeloid leukemia characterization and classification
Authors: Annette A. Prah, Derek Van Delden, Joan Alexander, Asya Stepansky, Erin Boyle, Nicholas Chiorazzi, Jonathan Kolitz, Stephanie Boisclair, Steven L. Allen, David Chitty, Andrea B. Moffitt
Journal: Cancer Research (2025)

Abstract Acute myeloid leukemia (AML) is a hematopoietic neoplasm characterized by uncontrolled proliferation of myeloid blast cells. AML is the most common acute leukemia, with over 20, 000 new U.S. cases diagnosed annually. The 5-year survival rate varies greatly by age, from 63% in younger patients to 11% in older patients. AML is a genetically heterogeneous disease, with over 25 genes implicated in classification. Clinical next-generation sequencing (NGS) is crucial for AML management, aiding in relapse risk assessment, therapy guidance, and understanding of the genetic landscape. In AML, this typically involves sequencing a panel of 50-500 genes. In practice, only diagnostic samples are tested, without germline controls. In research settings, whole genome sequencing (WGS) is performed on paired diagnosis and remission samples, with remission serving as a germline control. This study assessed the accuracy and utility of AML clinical NGS reports by comparing with matched WGS data. We explored the benefits of germline controls and how discrepancies impact risk classification. 65 AML patients treated in the Northwell Health System and enrolled in an IRB-approved biospecimen study were analyzed. Clinical NGS testing was performed using FoundationOne Heme (n=51) or GenPath OnkoSight (n=14). We analyzed mutation data from clinical NGS reports (median 12 variants per report; range 1-26), identifying DNMT3A, FLT3, TP53, and NPM1 as the most frequently mutated genes, consistent with prior studies. Matched WGS data from 15 patients classified clinically reported variants as somatic (27%; n=57), germline (65%; n=135), or not observed (8%; n=17), using variant allele frequencies (VAFs). There was a strong but imperfect correlation between somatic/germline labels and known/unknown status, provided in the clinical NGS reports: 89% of germline variants were of unknown significance (n=120), while 83% of somatic variants were of known significance (n=43). Clinical NGS reports lacked data such as population variant frequency, pathogenicity scores, and VAFs. Germline and unknown significance variants had higher population frequencies and lower pathogenicity scores than somatic and known significance variants (p&amp;lt;0.0001). European LeukemiaNet (ELN) risk groups assigned using clinical NGS data (Adverse: 51%, n=33; Favorable: 29%, n=19) were affected by the inclusion of germline or unobserved variants; excluding those reclassified two patients from Adverse to Intermediate risk. This study finds general agreement between AML clinical NGS and WGS in the overlapping genes. The clinical NGS reports studied here lack annotation of germline and subclonal variant status. We highlight the limitations of clinical NGS without germline controls and its implications for AML classification, clinical decision making, and broader scientific understanding. Citation Format: Annette A. Prah, Derek Van Delden, Joan Alexander, Asya Stepansky, Erin Boyle, Nicholas Chiorazzi, Jonathan Kolitz, Stephanie Boisclair, Steven L. Allen, David Chitty, Andrea B. Moffitt. Comparison of clinical next generation sequencing and whole genome sequencing for acute myeloid leukemia characterization and classification [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7171.