Spatial heterogeneity of hotspot mutations and clonal relatedness in ovarian endometrioid carcinoma.

Ovarian endometrioid carcinoma (OEC) is a major histotype of endometriosis-associated ovarian carcinoma, yet the spatial distribution of cancer-associated mutations and the clonal relatedness of synchronous endometrial and ovarian carcinomas (SEOC) remain incompletely understood. In this study, we performed spatially controlled molecular profiling of OEC using laser capture microdissection, droplet digital PCR, targeted amplicon sequencing, and exploratory whole-exome sequencing. We analyzed multiple tumor regions and selected stromal compartments from OEC and SEOC cases to assess the intratumor distribution of hotspot mutations in CTNNB1, PIK3CA, and KRAS and to evaluate molecular relatedness between ovarian and endometrial lesions in SEOC. Cancer-associated mutation signals showed spatially heterogeneous distributions in a subset of histologically uniform OEC. In SEOC, shared alterations supported clonal relatedness in selected cases. Low-level hotspot mutation signals were also detected in microdissected stromal compartments immediately adjacent to tumor glands, although their cellular origin remains to be determined. These findings suggest that single-region molecular testing may underestimate spatial molecular heterogeneity in OEC and provide a framework for understanding the clonal relatedness of endometriosis-associated gynecologic carcinomas.