Single-cell analysis unravels divergent gene signatures shaping seminoma stemness and metastasis.

Seminoma is the most common solid malignant tumor of the testis in young males, significantly impacting fertility. Approximately 20% of seminomas metastasize, markedly increasing recurrence risk and compromising quality of life. To investigate the poorly understood mechanisms driving seminoma metastasis, we performed a comprehensive analysis by integrating single-cell RNA sequencing, TCGA data mining, and cell biology assays. We identified significant intratumoral heterogeneity. A subset of tumor cells expressing DPPA4 and PSMA7 showed high stemness, enhanced self-renewal, and association with metastasis. Knockdown of these genes reduced sphere formation, tumor migration and proliferation in Tcam-2 and NCCIT cells. Conversely, tumor cells overexpressing PAGE5 and SAT1 exhibited reduced stemness, migratory capacity and proliferation. In the immune microenvironment, we identified IFNG+ T cells, which recruit and activate other antitumor immune cells. These cells secrete IFN-γ, which promotes tumor differentiation, reduces stemness, and mitigates tumor aggressiveness. Based on these findings, we further developed a molecular panel to aid in the identification of seminomas with a higher risk of metastasis. In conclusion, our study identifies unique molecular signatures that facilitate risk stratification based on metastatic potential, providing valuable insights for improving precision medicine.