Background – The BRAF V600E mutation is associated with the hypermethylator phenotype CIMP, which can also lead to the MSI-H phenotype. BRAF V600E mutation and MSI-H/dMMR status seem to be biologically intertwined; however, the impact of coexisting BRAFV 600E mutations on the TME and immunometabolomic features of MSI-H/dMMR CRC tumors is not well characterized.

Methods – A retrospective review of deidentified records of patients with MSI-H/dMMR CRC tumors was conducted using next-generation sequencing data (Tempus |xT assay: DNA-seq of 595-648 genes at 500x coverage, and full transcriptome RNA-seq). Several immune markers of tumor immunogenicity in BRAF wild-type (BRAFwt ) vs. V600E-mutated (BRAFV 600E ) tumors were assessed, including tumor mutational burden (TMB), neoantigen tumor burden (NTB, ScanNeo), PD-L1 expression, immune infiltration, and canonical immuno-metabolomic pathways (82 geneset signatures).

Results – A total of 459 MSI-H/dMMR CRC tumors were analyzed, of which 123 (27%) tumors harbored BRAF V600E mutations, and 336 (73%) were BRAF wt. MSI-H/dMMR BRAF V600E tumors were more frequently identified in females (69% vs. 55%; P= 0.01), non-Hispanic or non-Latino (100% vs. 73%; P =0.001), and older patients (median age: 76 yrs vs. 62 yrs; P BRAF WT, BRAF V600E tumors exhibited significantly higher TMB (Median: 49 mut/MB vs. 36 mut/MB; P 10 mut/MB; 100% vs. 95%; P = 0.008); however, no significant differences were observed with tumor NTB, immune score, or T cell infiltration (CD4 or CD8). NK cell infiltration was higher in the BRAF V600E cohort (BRAF WT tumors, BRAF V600E tumors harbored a greater frequency of mutations in MSH6 (42% vs. 20%), B2M (33% vs. 16%), BRCA2 (31% vs. 12%), ATM (23% vs. 12%), and TP53 (30% vs. 19%) but a lower frequency of MSH2 (3.3% vs. 11%), all P BRAF V600E tumors: glycerophospholipid, galactose, cyclin-dependent cell signaling; Nucleotide, and TH1 inflammation. Five pathways were downregulated (Wnt, Notch, TH17 inflammation, amino sugar, and cancer stem cell signaling).

Conclusions – MSI-H/dMMR BRAF V600E CRCs undergo broad metabolic reprogramming (e.g., glycerophospholipidgalactose, nucleotide). A rise in lipid metabolism, particularly with NK inflammation, suggests that BRAF V600E-mutated tumors may be associated with a non-classical immune component. BRAF V600E and BRAF wt CRCs exhibited similar NTB and T cell infiltration, suggesting that both are likely to benefit from immune checkpoint inhibitors.