AbstractSeveral molecular aberrations affect the prognosis of patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) with excess blasts (EB). This study aimed to determine the incidence and clinical impact of molecular genetic aberrations in Thai patients with AML and MDS-EB, detected by the next-generation sequencing (NGS) technique. This prospective, observational study was conducted between 2018 and 2020 on newly diagnosed Thai AML or MDS-EB patients aged above 15 years. NGS was performed using a custom amplicon-based targeted enrichment assay for 42 genes recurrently mutated in myeloid neoplasms. The molecular results were correlated with baseline patient and disease characteristics as well as outcomes. Forty-nine patients were enrolled in this study. The median age was 56 years (interquartile range [IQR], 44–64), with nearly equal proportions of males and females. The median number of mutations was 3 (IQR, 2–4). The most frequent alterations were FLT3 internal tandem duplications (ITD) (28.6%), DNMT3A (24.5%), and WT1 (22.4%) mutations. FLT3-ITD was more frequent in the de novo AML group than in the MDS/secondary AML group, whereas in the MDS/secondary AML group, ASXL1, ETV6, and SRSF2 mutations were more frequent. Patients aged greater than 65 years and patients with mutated TP53 were more likely to have inferior overall survival from multivariate analysis. FLT3-ITD was the most common mutation among newly diagnosed Thai AML patients. TP53 mutation and advanced age were independent adverse factors for survival outcome. The genetic landscapes of AML patients vary between national populations. Thai Clinical Trials Registry identifier: TCTR20190227003.