Abstract Background: Although BRAF V600-mutated (BRAFm) colorectal carcinoma (CRC) does not generally respond to BRAF inhibitors, results of preclinical studies suggest strong synergistic activity between a BRAF inhibitor and an EGFR inhibitor or a PI3K inhibitor. The aims of this study were to establish safety and preliminary activity of combination therapy with encorafenib (ENC), a highly selective BRAF V600 inhibitor, the EGFR monoclonal antibody cetuximab (CTX), ± the α-specific PI3K inhibitor alpelisib (ALP) in patients (pts) with BRAFm CRC; and assess the potential clinical significance of biomarkers and microsatellite instability (MSI) status by genomic analysis of tumor samples. Methods: In this phase I study, the maximum tolerated dose/recommended phase II dose (RP2D) was determined and safety, tolerability, and anti-tumor activity of ENC + CTX ± ALP were evaluated, in pts with advanced BRAFm/KRAS wild-type CRC. Archival or fresh screening tumor samples were analyzed for somatic mutations and copy number aberrations to evaluate potential correlation with response. Results: Fifty-four pts were enrolled in the dose-escalation phase; 26 in the dual arm (ENC + CTX) and 28 in the triple arm (ENC + ALP + CTX). RP2D for each treatment arm was established as 200 mg ENC QD + CTX weekly (dual arm) + 300 mg ALP once daily (triple arm). The most common grade 3/4 treatment-related adverse events were fatigue and hypophosphatemia in the dual arm (8% each), and nausea and hypophosphatemia in the triple arm (4% each). Hyperglycemia was observed in one patient (3.8%) in the dual arm and 8 patients (29%) in the triple arm. Overall response rates (≥ partial response [PR]) for the dual and triple arms were 23% (1 CR, 4 PR, and 1 unconfirmed (u) PR) and 32% (4 PR and 5 uPR), respectively. Median progression-free survival was 3.7 and 4.3 months, respectively. Biomarker analysis was performed on 21 tumor samples. Several genetic alterations occurred with similar frequency between patient samples and the FoundationOne database (PIK3CA, TP53, APC). Specifically, the PIK3CA mutation and PTEN alterations were found in 24% and 38% of patient samples, respectively; however, these alterations did not appear to be correlated with response. Non-MSI-high status was associated with a lower PFS for the dual arm (2.8 vs 3.7 months), but not the triple arm (4.4 vs 4.3 months). Updated biomarker analyses, including MSI status, will be presented. Conclusions: These results indicate that both ENC + CTX and ENC + CTX + ALP are well tolerated with promising antitumor activity in pts with advanced BRAFm CRC who failed at least one prior therapy. Additional biomarker analyses will help to clarify the clinical significance of genetic alterations, particularly with MSI status and the PI3K pathway. Enrollment in the phase II portion of the study is ongoing. Citation Format: Jan H. M. Schellens, Robin van Geel, Johanna C. Bendell, Anna Spreafico, Martin Schuler, Takayuki Yoshino, Jean-Pierre Delord, Yasuhide Yamada, Martijn P. Lolkema, Jason E. Faris, Ferry A. L. M. Eskens, Sunil Sharma, Rona Yaeger, Heinz-Josef Lenz, Zev A. Wainberg, Emin Avsar, Arkendu Chatterjee, Savina Jaeger, Tim Demuth, Josep Tabernero. Final biomarker analysis of the phase I study of the selective BRAF V600 inhibitor encorafenib (LGX818) combined with cetuximab with or without the α-specific PI3K inhibitor alpelisib (BYL719) in patients with advanced BRAF-mutant colorectal cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr CT136. doi:10.1158/1538-7445.AM2015-CT136