Background: BRAF V600E mutations can lead to activation of the MEK/ERK pathway. The combination of dabrafenib plus trametinib was recently FDA-approved for use in these BRAF V600E altered unresectable or metastatic cancers. BRAF non-V600 alterations also increase phosphorylation of MEK and ERK and are potential candidates for MEK/ERK inhibitors. Here we examine the frequency of BRAF mutations and their genomic background across GI cancers. Methods: The Oncomap ExTra genomic profiling assay utilizes tumor-normal, whole-exome, whole-transcriptome DNA and RNA sequencing to identify somatic alterations in tumors. The assay detects single-nucleotide substitutions, indels, copy number alterations, alternative transcripts, and gene fusions. Tumor mutational burden (TMB) and microsatellite instability (MSI) are also assessed. The frequency of BRAF and other gene alterations were identified across all GI cancers. Reported BRAF alterations were limited to those linked to an FDA approved therapy, clinical trial eligibility, and those deemed significant from review of the literature. Results: A total of 1927 patients with GI cancers were assayed between April 2018 to July 2022. The BRAF gene was mutated in 85 (4.4%) patients, which included 53 (2.8%) that were V600E and 32 (1.5%) that were non-V600, representing 37.6% of BRAF mutations. Colorectal and small bowel cancers had the highest frequencies of BRAF mutations at 7.6% and 7.1% respectively, and the frequency was lower in other GI cancers (0-2.3%). In addition to BRAF V600E, other mutations occurred across the kinase domain, including D594 (0.6%), K601E (0.3%), G466 (0.3%) and G469 (0.2%). BRAF fusions were found in 1 CRC and 2 pancreatic cancer cases. BRAF mutational status (altered versus not) was significantly associated with mutations in several other genes. In addition, BRAF status was significantly associated with TMB-high and MSI-high. Conclusions: Analysis of whole-exome and whole transcriptome sequencing data found actionable BRAF mutations in 4.4% of GI cancer patients, 37.6% of whom had BRAF mutations not at V600. Several biomarkers were significantly associated with BRAF alterations. Assays that are limited to DNA sequencing, or to targeted gene panels, may not detect mutations and fusions that could aid in therapy selection and prognosis.

^ap-value for Fisher’s Exact test, ^aq-value is the Benjamini-Hochberg false discovery rate.