Several antigen-specific vaccines targeting the E7 protein of HPV-16 are capable of preventing and treating the growth of murine model tumors expressing E7. These improved results from murine models led us to apply these vaccines to human subjects at the Johns Hopkins Hospital (JHH). One of the important focuses of human vaccine trials is the characterization of immunologic parameters that may serve as indicators for effective clinical responses. Since cell-mediated immunity has been shown to play an important role in controlling HPV-associated neoplasms, we will focus on characterizing immunological parameters associated with cell mediated. Two components of T cell-mediated immune response will be monitored in vitro including E7-specific CD8+ cytotoxic T lymphocyte (CTL) responses and E7- specific CD4+ T helper (Th) cell responses. The HPV-16 E7-specific CD8+ T cell activities and their precursors will be determined using cytotoxic T lymphocyte (CTL) assays, the enzyme-linked immunospot (ELISPOT) assays and intracellular cytokine staining with flow cytometry analysis. The E7- specific CD4+ Th cells derived from peripheral blood mononuclear cells (PBMCs) will be used for in vitro proliferation assays, ELISPOT assays and intracellular cytokine staining with flow cytometry analyses. In addition, cytokine profiles related to Th1 (IFN-gamma and IL-2) cytokine staining with flow cytometry analyses. In addition, cytokine profiles related to Th1 (IFN-gamma and IL-2) and Th2 (IL-4 and IL-10) will be determined using quantitative ELISA and intracellular cytokine staining. Two additional in vivo parameters related to T cell-mediated immunity will be measured, including delayed-type hypersensitivity (DTH) responses against HPV-16 E7 antigen in patients and characterization of immune cells and their cytokine expression in biopsies of cervical lesions before and after vaccination. The results of these immunological assays will be correlated with the pathologic, virologic and clinical outcomes of vaccination in these patients. These relevant immunological parameters may be used as predictors of vaccine effects. In addition, they may provide insight into the mechanism of these vaccine effects, and facilitate the generation of better HPV vaccines in the future.
