Abstract: Purpose: The tumor immune microenvironment determines clinical outcome. Whether the original tissue in which a primary tumor develops influences this microenvironment is not well understood. Experimental Design: We applied high-dimensional single-cell mass cytometry (CyTOF) analysis and functional studies to analyze immune cell populations in human papillomavirus (HPV)-induced primary tumors of the cervix (CxCa) and oropharynx (OPSCC). Results: Despite the same etiology of these tumors, the composition and functionality of their lymphocytic infiltrate substantially differed. CxCa displayed a 3-fold lower CD4:CD8 ratio, contained more activated CD8+CD103+CD161+ effector T-cells and less CD4+CD161+ effector memory T-cells than OPSCC. CD161+ effector cells produced the highest cytokine levels among tumor-specific T-cells. Differences in CD4+ T-cell infiltration between CxCa and OPSCC were reflected in the detection rate of intratumoral HPV-specific CD4+ T-cells and in their impact on OPSCC and CxCa survival. The PBMC composition of these patients, however, was similar. Conclusions: The tissue of origin significantly impacts the overall shape of the immune infiltrate in primary tumors.