Gene Expression Profiling of the Peritumoral Immune Cell Infiltrate of Penile Squamous Cell Carcinomas

Penile carcinomas are rare tumors in Europe and need further investigations due to their inferior prognosis in late tumor stages. The presence of disparate immune cell infiltrates was observed in these tumors, which were subsequently demonstrated to give rise to divergent tumor prognoses. The objective was to further characterize this immune cell infiltrate with the use of immunohistochemistry and RNA expression. A total of twelve well-characterized cases of penile squamous cell carcinomas with known infection status by human papillomavirus (HPV) and p16 status were assessed. The cases were classified according to their morphological characteristics, including those exhibiting a pronounced peritumoral immune cell infiltrate and those with less peritumoral immune cell infiltration. The generation of RNA expression data was conducted using the nCounter^® PanCancer Immune Profiling Panel. Computational models were employed to calculate the proportions of immune cells. To corroborate the findings, an immunohistochemical analysis was conducted using antibodies against CD20, CD3, CD4, CD8, MUM1, CD68, and CD117. Our cases were clustered according to the immune cell infiltrate detected via histology in a group with less immune cell infiltrate density and in a group with increased immune cell infiltrate density. Generally, all immune cells showed an increased amount in the group with pronounced immune cell infiltrate density. The clusters were found to relate to cell functions, the complement system, cytotoxicity, pathogen defense, regulation, and T-cell functions. In cases exhibiting a pronounced immune cell infiltrate, the top three genes that exhibited the greatest upregulation were GZMA, MICB, and GNLY. No relationship to HPV infection status was demonstrated. Immunohistochemistry validated the data gained via RNA expression and showed a correlation with EPIC and Cibersort. The clustering of cases based on immune cell infiltrate density revealed significant distinctions between groups with lower and higher immune cell infiltrate density. The group with increased immune cel infiltrate density showed a greater abundance of immune cells, aligning with key functions like cytotoxicity, pathogen defense, and T-cell regulation. Among these cases, the genes GZMA, MICB, and GNLY were significantly upregulated, suggesting their involvement in an increased immune response. The role of HPV infection status in our cases with regard to the peritumoral immune cell infiltrate remains inconclusive.