Revisiting the relevance of sidedness in colonic tumor molecular profiling.

3531 Background: Colorectal cancer (CRC) is a heterogeneous disease with distinct molecular and clinical differences between right- and left-sided tumors. This study analyzes these variations to understand their impact on tumor behavior and treatment strategies. Methods: A total of 445 colonic tumor samples (132 right-sided, 313 left-sided) were profiled to assess mutations, amplifications, and fusions in key cancer-related genes along with targeted transcriptome analysis of 20,802 genes in a subset using semiconductor based next-generation sequencing (NGS) platform at Datar Cancer Genetics. Immunotherapy biomarkers (TMB, MSI, and PD-L1 22C3 TPS) were analyzed in a subset. Results: Right-sided and left-sided colon cancers exhibit substantial molecular heterogeneity, driven by distinct genetic and epigenetic alterations (Table 1). Right-sided tumors were more frequently associated with MSI and had statistically significant higher incidence of BRAF mutations. KRAS mutations were frequently observed in both right-sided and left-sided tumors at equal rates. ERBB2 amplifications were exclusive to left side tumors, whereas oncogenic ERBB2 mutations were equally distributed. Located around ERBB2, PGAP3 gene co‐amplification too was exclusive to left sided tumors. TFE3 alterations were absent from left sided tumors and common on right side. TP53 mutations, though more common in left-sided tumors, the difference was not statistically significant. Gene expression profiling of a subset, including 103 left-sided and 41 right-sided colon tumors, revealed activation of the Wnt / β-catenin signalling pathway, RAS/MAPK pathway, TGF-β signalling pathway, and immune-related pathways, though these differences were not statistically significant, suggesting that while specific drivers may differ—such as the predominance of APC mutations in left-sided tumors (56.8% vs 37.9%) leading to WNT activation and the higher incidence of RSPO2/3 fusions (7.1% vs 1.7%) in right-sided tumors -eventually some pathways are commonly implicated in colorectal cancer biology. Conclusions: Existing therapies like ICIs, HER2 inhibitors, and emerging molecules such as RSPO2/RSPO3 inhibitors could have differing impact based on tumor sidedness. Integrating these distinctions into drug development and clinical trials holds potential to optimize treatment outcomes. Molecular profiles of right- and left-sided colon tumors. Gene Right (%) Left (%) p-Value(Chi-square test) TP53 64.5% 73.2% 0.075644 APC 37.9% 56.8% 0.001354 KRAS 50.0% 43.6% 0.220194 BRAF 18.8% 3.0% 0.00001 TFE3 9.5% 0% 0.109087 ERBB2 mutation 2.3% 2.0% 0.80941 ERBB2 amplification 0% 5.9% 0.023006 PGAP3 amplification 0% 7.1% 0.673427 RSPO2/3 fusion 7.1% 1.7% 0.827207 Immunotherapy Biomarkers TMB 10-14 24.7% 29.3% 0.458066 TMB ³15 15.6% 7.6% 0.059925 MSI-High 8.1% 3.4% 0.066213