Intratumoral heterogeneity of somatic alterations between spatially separated differentiated and undifferentiated types in gastric cancer

Gastric cancer exhibits substantial intratumoral heterogeneity; however, genetic differences between spatially separated differentiated and undifferentiated components within the same tumor, particularly in deficient mismatch repair (dMMR) cases, remain poorly defined. We compared somatic mutation and copy number variation (CNV) profiles between these two histologic types within the same gastric tumor, including dMMR cases. Formalin-fixed, paraffin-embedded specimens from five gastric cancers were macrodissected to separate differentiated and undifferentiated components and analyzed using a 225-gene targeted panel. All cases shared one or more somatic mutations between the two components, consistent with a common clonal origin. However, most mutations were type-specific, with greater divergence in dMMR tumors. dMMR cases harbored more mutations than proficient mismatch repair cases, and many were detected at low variant allele frequency; type-specific low-variant allele frequency mutations were enriched in the undifferentiated component. CNV patterns also differed between the two components, and most CNVs were amplifications. NOTCH1 CNV was shared across both components in all cases, whereas CNVs overall were more frequent in the differentiated component. These findings indicate that differentiated and undifferentiated components within the same gastric tumor share truncal mutations but accumulate distinct additional alterations, underscoring spatial heterogeneity and supporting multiregion sampling for molecular profiling.