Real-World Landscape of KMT2A and NPM1 Variants in Hematologic Malignancies

Background: KMT2A rearrangements and NPM1 mutations are key genomic drivers in myeloid neoplasms, informing diagnosis, prognostication, MRD assessment, and emerging targeted therapies. Large-scale real-world data are needed to clarify testing trends, positivity rates, and co-mutational patterns across hematologic indications. Methods: We analyzed all KMT2A and NPM1 tests performed in a national reference laboratory cohort from 10/1/2020-9/30/2025. Test modality, indication, and positivity were evaluated, as well as variant classification, fusion partners, and aggregated co-mutations for positive cases. Results: Across the study period, 201,381 KMT2A and 202,902 NPM1 tests were performed, with positivity rates of 2.6% and 3.1%, respectively. Annual testing volume increased substantially for both markers, driven by hospital-based ordering and expanding NGS utilization. Testing spanned both sexes and all ages, with most samples originating from patients ≥65 years. Top testing indications were MDS, AML, and MPN/CMML. KMT2A positivity was most frequent in AML (1,964), MPN (1,369), and MDS (353). NPM1 positivity was enriched in MDS (6,170), AML (1,191), and CMML (541). Over time, use of NGS expanded significantly from 2021 onward, becoming the dominant modality for NPM1 testing and complementing ongoing FISH testing for KMT2A, with positivity detected across both platforms. Among KMT2A-positive cases, SNVs (86%) and fusions (9%) were observed, with most pathogenic findings occurring in fusion-positive samples. Common fusion partners included MLLT3, AFF1, ELL, MLLT4, and MLLT10. Recurrent co-mutations involved TP53, KRAS/NRAS, TET2, FLT3, DNMT3A, and ASXL1. Within AML, 273 of 1,964 KMT2A-positive patients harbored a fusion. NPM1-positive variants were predominantly pathogenic (65%), with co-alterations spanning SNVs, insertions/deletions, and rare fusions. The most frequent co-mutations involved TET2, DNMT3A, FLT3, IDH2, NRAS, and SRSF2. Conclusions: Testing for KMT2A and NPM1 nearly doubled over five years, driven by widespread adoption of NGS, continued FISH utilization, and increasing clinical relevance of genomically guided therapies for KMT2A- and NPM1-mutated disease, particularly in AML. These findings define a comprehensive real-world genomic profile of KMT2A-rearranged and NPM1-mutated hematologic malignancies and support ongoing efforts in diagnosis, prognostication, and therapeutic development. Citation Format: Roisin Puentes, Madhuri Paul, Ryan Bender, Amber Chevalier, Nathan Montgomery. Real-world landscape of KMT2A and NPM1 variants and fusions in hematologic malignancies (2020-2025) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1993.