Genetic Defects and Tumor Microenvironment Associated with Left-Sided and Right-Sided Colorectal Cancer

Ryan Gates, BS (Advisor: Raghu Kalluri, MD, PhD)

          Colorectal cancer (CRC) is one of the most common malignancies in the United States, with approximately 150,000 new cases diagnosed annually. CRC can arise from either the left or right side of the colon, and these anatomical subtypes have historically been associated with distinct developmental pathways, mutational profiles, and tumor microenvironment (TME). Left?sided CRC (LCRC) typically follows the classical adenoma–carcinoma sequence and is characterized by recurrent mutations in APC, TP53, and KRAS, whereas many right?sided CRC (RCRC) tumors arise from the serrated neoplasia pathway and are frequently associated with microsatellite instability (MSI) and BRAF mutations. In this study, we evaluated whether primary tumor location or microsatellite instability status is the dominant determinant of genomic and immunologic heterogeneity in CRC. We analyzed whole?exome sequencing (WES) data to identify differentially mutated genes enriched for predicted oncogenic or likely oncogenic variants across tumor subgroups, and single?cell RNA?sequencing (scRNA?seq) data to characterize T cell composition and functional states within the TME. While limited differences in oncogenic mutation frequencies and immune cell composition were observed between LCRC and RCRC, MSS status emerged as a stronger predictor of genomic alterations and immune phenotypes. MSI CRC primary tumors showed enrichment of oncogenic mutations in WNT signaling-associated genes, such as RNF43, and exhibited increased proportions of exhausted CD8⁺ T cells. In contrast, MSS CRC tumors were enriched for oncogenic mutations in APC, P53 and KRAS, and contained higher proportions of naïve CD4⁺ T cells. Finally, compared to MSS RNF43 WT primary tumors, MSI RNF43 Mutant primary tumors had fewer CD4⁺ naïve T cells. Conversely, compared to MSI RNF43 WT primary tumors, MSI RNF43 Mutant primary tumors had more CD4⁺ naïve T cells. Together, these findings indicate that MSS Status is a more influential predictor of an immunosuppressive TME compared to primary tumor anatomical location and MSI RNF43 Mutant CRC tumors exhibit a unique TME.

Advisory Committee:

• Raghu Kalluri, MD, PhD, Chair
• E. Scott Kopetz, MD, PhD
• Kunal Rai, PhD
• Stephanie Watowich, PhD
• Xiling Shen, PhD