HPV-associated oropharyngeal squamous cell cancer (OPSCC) is a distinct clinical entity with a much better prognosis after (chemo)radiotherapy than HPV-negative OPSCC, a feature that has been linked to the local presence of a more dense immune infiltrate. To evaluate the relation between clinical outcome, immune cell infiltration and the possibility of the immune system to react against potent virally-derived antigens, we performed an in-depth analysis of freshly isolated tumor-infiltrating lymphocytes from a cohort of 97 HPV16-positive and –negative OPSCC patients.
Fresh tumor tissue was processed to obtain tumor infiltrated lymphocytes (TILs). Furthermore, tumor tissue was archived and subsequently used for HPV DNA typing, test the expression of p16INK4a and quantification of T-cell infiltration. TILs were phenotypically analyzed and their capacity to expand and produce cytokines following stimulation with HPV16 E6 and E7 antigens or phytohemagglutinin (PHA) was tested using proliferation assays and flow cytometry. Findings were validated in an additional cohort of 75 HPV+ OPSCCs from the publicly available cancer genomic atlas (TCGA) database. The direct growth inhibiting effects of the TIL-produced cytokines was tested on five different OPSCC cell lines, either with or without cisplatin, the commonly used chemotherapeutic agent.
Patients with a HPV+ OPSCC have a better overall survival (OS) than HPV-negative OPSCC patients. Within these HPV+ OPSCC patients the majority displayed a HPV-specific T-cell response, predominated by CD4+ T cells. HPV-immunity was strongly correlated to a more favorable clinical outcome. The HPV-reactive T cells produced IFNg, TNFa and IL-17, thus, exhibiting a T-helper type 1 (Th1)/Th17 profile. A high expression of CD4 and IFNg also correlated with better overall survival in the TCGA dataset. Notably, the OPSCC of patients displaying an intratumoral HPV16-specific T cell response were significantly higher infiltrated by T cells expressing the with IFNg-production associated transcription factor Tbet than the other patients. Culture of OPSCC tumor cell lines in the presence of IFNg and TNFa lowered their proliferation, while the combination with cisplatin resulted in increased apoptosis of the tumor cells.
In conclusion, the presence of Th1-oriented HPV16-specific T cells in pre-treated HPV+ OPSCC better controls tumor growth (lower T and N stage), makes the tumor more susceptible for cisplatin and results in better OS of the patients. Therefore, these HPV16-specific TILs can be used as predictor of clinical outcome upon therapy.