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Introduction. Recent genome profiling studies have increased our understanding of the mutation landscapes of myeloid malignancies. Molecular testing of AMLs (NPM1, FLT3-ITD, KIT) and MPNs (JAK2, CALR) constitute current diagnostic standard-of-care. Evidence for the diagnostic, prognostic and/or therapeutic impact of a growing set of genes and variants in myeloid malignancies allows for more accurate patient stratification and enhanced patient management. This has led to consideration of next-generation sequencing (NGS) approaches to simultaneously detect multiple variants in myeloid malignancies for use in the clinical diagnostic setting, to supplant single-gene molecular assays. We designed the Princess Margaret Advanced Genomics in Leukemia (AGILE) trial to prospectively assess the utility of NGS molecular profiling in the management of patients with myeloid malignancies. Methods. Patients for the AGILE trial are consented at the time of diagnosis using an REB approved written consent. Bone marrow or peripheral blood samples are collected at consent, accessioned within CoPath, and DNA extracted for NGS testing. NGS molecular profiling was performed using the TruSight Myeloid Sequencing Panel (TMSP; Illumina) on the MiSeq benchtop genome sequencer (Illumina) by the University Health Network Advanced Molecular Diagnostics Laboratory. The TMSP enables profiling of 54 genes (39 hotspot region; 15 complete coding region coverage) using amplicon-based library preparation and sequencing by synthesis. The TMSP detects the CALR 52 base pair deletion relevant to myelofibrosis, but not FLT3 internal tandem duplications greater than 30 base pair in size. Data were analyzed by NextGENe (v.2.3.1, SoftGenetics) and MiSeq Reporter v2.4.60. A specific script enabling alignment and calling of CALR deletions was added to the analysis to ensure there were no false negative calls. Additional testing and verification of CEBPA variants was performed by Sanger sequencing. Variants were interpreted according to Sukhai et al (Genetics in Medicine, 2015), reviewed by lab directors and reported in the Electronic Patient Record. Impact on patient care was defined as: potential for post-consolidation clinical trials; changes to frequency of monitoring; and, changes to transplant management. Cases were discussed in an interdisciplinary Genomic Tumor Board setting, at which NGS profiling data were reviewed in the context of all other diagnostic information for the patient, to determine impact on patient care. Results. Between February 11 and July 24, 2015, 162 patients were consented for AGILE; 148/162 were profiled by NGS, and to date 124/148 have been reviewed and interpreted. 62/124 (50%) of interpreted cases had a diagnosis of acute myeloid leukemia (AML); 21/124 (20%) with myeloproliferative neoplasms (MPNs); 13/124 (10%) with myelodysplastic syndromes (MDS); 6/124 (5%) with MDS/MPN; and, 15% with other hematologic malignancies. 90% of all cases profiled were informative for at least one variant (range 1-9 variants, average 3.1 variants/case). AML, MDS and MDS/MPN cases exhibited slightly more variants (3.4-4.4 variants/case) than did MPN cases (2.6 variants/case). Overall, 69% of variants were potentially actionable (Sukhai et al, 2015: 23% class 1; 8% class 2; 38% class 3), with a large fraction of cases (90/124, 72.6%) demonstrating at least one class 1 or class 3 variant. Additionally, 73/124 (58.9%) of patients exhibited actionable, class 1, variants not currently being identified by routine molecular diagnostics. In AMLs and MPNs, 88-90% of cases exhibited at least one potentially actionable variant; NGS profiling was more informative in AMLs (62% of cases exhibiting potentially actionable variants not profiled in standard of care testing, compared to 12% of MPN cases). Conclusions. We report the results of a prospective analysis of integrated NGS profiling in the context of diagnosis and management of patients with myeloid malignancies. Using a targeted NGS panel, molecular profiling of patients yielded significant information benefit over current standard approaches in 58.9% of cases analyzed, enabling potential impact on patient management. These data highlight the utility of NGS profiling to complement the initial diagnostic evaluation of myeloid malignancies. Disclosures Gupta: Incyte: Honoraria, Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees.