Combination of HDAC inhibition and cytokine enhances therapeutic HPV vaccine therapy
HPV-associated malignancies continue to present a major health concern despite the development of prophylactic vaccines. Standard therapies offer limited benefit to patients with advanced stage disease. Despite improved outcomes with PD-1 targeted therapies, treatment resistance and modest response rates highlight a significant unmet need to develop novel therapies for these patients. PDS0101 (designated HPV vaccine) is a liposomal nanoparticle HPV16-specific therapeutic vaccine that has been shown to generate strong HPV-specific responses in pre-clinical and clinical studies. Here we assess the efficacy of this HPV vaccine in combination with the tumor-targeting immunocytokine NHS-IL12 (PDS01ADC), plus either aPD-1 or the class I histone deacetylase (HDAC) inhibitor Entinostat. Mice bearing HPV16+, aPD-1 refractory TC-1 and mEER tumors were treated with HPV vaccine, NHS-IL12, and either aPD-1 or Entinostat to determine anti-tumor efficacy and survival benefits. A comprehensive analysis of the tumor microenvironment was performed using flow cytometry, multiplex immunofluorescence, chemokine and cytokine assessment, and scRNAseq with TCR enrichment. Combination of HPV vaccine and NHS-IL12 with either Entinostat or aPD-1 yields significant anti-tumor activity and prolonged survival in aPD-1 refractory models of HPV16+ cancer, with superior activity employing Entinostat versus aPD-1 combination. Entinostat triple therapy increased overall and HPV16-specific tumor CD8+ T cell infiltration with heightened cytotoxicity. TCR sequencing revealed a CD8+ T cell clone unique to vaccine-treated cohorts, which displayed an enriched cytotoxic transcriptional profile with triple therapy. These effects were parallelled by strong differentiation of tumor-associated macrophages (TAMs) towards proinflammatory, anti-tumor M1-like cell states.  Single-cell transcriptomic analysis indicated all three agents were required for highest modulation of both CD8+ T cells and TAMs conducive to tumor control. A biomarker signature reflecting the pre-clinical findings was found to be associated with improved survival in patients with HPV-associated malignancies. Together, these findings provide a rationale for the combination of HPV vaccine, NHS-IL12, and Entinostat in the clinical setting for patients with HPV16-associated malignancies. Overall design: CD45+ cells were isolated from treated TC-1 tumors harvested from C57Bl/6 mice using magentic separation and analyzed for scRNAseq with TCR enrichment