Integration of Spatial Transcriptomics and Mendelian Randomization Identifies Candidate Molecular Regulators of Endometrial Cancer Progression

Abstract

Highlights What are the main findings? MM0 was identified as a putative stemness-associated malignant epithelial transcriptional subpopulation, and MM0-enriched regions showed higher inferred JAK–STAT and hypoxia scores in the cross-sectional spatial analysis. Integrative Mendelian randomization and colocalization analyses prioritized three candidate genes: DNAJA4, HSPA6, and LMNA. Subsequent experiments showed that knockdown of either DNAJA4 or HSPA6 suppressed estrogen-stimulated cell proliferation and migration, with reduced CDK1 and Cyclin B expression. What are the implications of the main findings? These findings illustrate a multi-omics approach linking malignant epithelial heterogeneity and spatial tumor biology to genetically supported candidate genes associated with endometrial cancer risk. DNAJA4 and HSPA6 warrant further validation as candidate functional genes in endometrial cancer and may inform future studies of therapeutic strategies. Abstract Endometrial cancer (EC) is a common gynecologic malignancy arising from the epithelial cells of the endometrium. The marked cellular heterogeneity of EC and features of its tumor immune microenvironment (TIME) contribute to disease complexity and have been associated with poor prognosis. This study integrates single-cell RNA sequencing, spatial transcriptomics, and Mendelian randomization (MR) to identify candidate genes associated with EC. Single-cell analysis identified MM0 as a putative stemness-associated transcriptional subpopulation with the highest CytoTRACE-inferred score; irGSEA indicated enrichment of proliferation- and stress-response pathways. Spatial data were analyzed with RCTD, MISTy, and stLearn to estimate spatial associations and pathway activities. MR and colocalization analyses integrating eQTL data and EC GWAS summary statistics prioritized DNAJA4, HSPA6, and LMNA as candidate genes with potential causal associations with EC risk. The expression patterns of these candidate genes were further examined in patient-derived samples. In Ishikawa cells cultured under high-estrogen conditions, siRNA-mediated knockdown of DNAJA4 and HSPA6 significantly suppressed proliferation and migration, accompanied by reduced CDK1 and Cyclin B expression. Collectively, these findings provide insight into EC heterogeneity and support further mechanistic investigation of candidate genes associated with malignant epithelial proliferation.