Background: High levels of microsatellite instability (MSI-H) is an approved biomarker for the selection of immunotherapy across solid tumor types. Given the high prevalence of MSI high in CRC, we used CGP to uncover additional therapy targets associated with MSI status. Methods: Hybrid capture-based CGP was performed on 8,004 clinically advanced CRC. Tumor mutational burden (TMB) was determined on 1.1 Mbp of sequenced DNA and microsatellite instability status (MSI-H or MS-Stable, MSS) was determined by principal components analysis of optimal homopolymer loci. Results: Of the 8,004 mCRC, 402 (5%) MSI-H and 7,602 (95%) MSS. Patient age and gender distribution did not differ between the 2 groups. Significant GA differences were found: MSS mCRC featured more KRAS, TP53 and APC GA, whereas MSI-H mCRC had more BRAF, PIK3CA, BRCA2 and ALK GA (Table). As expected, GA in the 4 genes associated with heritable CRC (HNPCC) were significantly enriched in MSI-H tumors. In addition, RNF43 correlated with MSI-H status. Median TMB was markedly higher in the MSI-H samples, 96% of which featured ≥20 mutations/Mb, compared to only 1% of MSS mCRC. Mutation frequencies and false discovery rate corrected p values from chi-squared analysis are presented below. Conclusions: MSI-H mCRC feature GA in RNF43, BRAF, PIK3CA, ALK and BRCA2 and near universal high TMB, impacting potential responses to both targeted and immunotherapies. MSS mCRC is characterized by more KRAS and TP53 GA frequencies and low TMB. GA in receptor kinases such as in ERBB2 and ALK are more often found in MSI-H mCRC, but are present in both types and represent additional potential targets for treatment strategies.