Induction of t-cell responses against PSA by plasmid DNA immunization

Abstract

This thesis is focused on using plasmid DNA encoding human prostate-specific antigen (PSA) for immunotherapy of prostate cancer. PSA is a secreted serine protease, which is produced almost exclusively by prostate epithelial cells. It is also expressed by prostate epithelial cells after malignant transformation and thus can be considered a tumor-associated antigen for prostate cancer. In our studies we investigated the potency of DNA vaccines to induce PSA-specific CD8+ cytotoxic T lymphocytes (CTLs), which can efficiently target and eliminate PSA-producing tumor cells.

In the first study we evaluated the efficacy of PSA-encoding plasmid DNA (pVax-PSA) in inducing T cell responses in patients with hormone-refractory prostate cancer. The plasmid DNA was administered during five cycles in doses of 100, 300 and 900μg, with three patients in each cohort. We demonstrated in this study that the pVax-PSA vaccine given in doses of 900μg is safe and can induce cellular immune responses against PSA.

The major aim of the following studies was to characterize the potency of pVax-PSA plasmid in inducing CD8+ T-cell responses in a mouse model. We demonstrated that the pVax-PSA plasmid is highly immunogenic in mice, with just one DNA immunization being sufficient to induce potent CTL responses against PSA. Additional modifications of human PSA, by linking it to heat shock proteins of different origin (pVax-PSA-HSPs plasmids), did not further increased the efficacy of pVax-PSA plasmid in inducing CTL responses or mediating protection against tumor challenge. We have also characterized the epitope specificity of CTLs induced after immunization of C57Bl/6 mice with the pVax-PSA plasmid.

We demonstrated that the majority of PSA-specific CTLs were directed against a single H-2Db-restricted epitope HCIRNKSVIL65-74 of PSA. However, immunization of mice with pVax-PSA plasmid does not properly reflect the clinical settings, where endogenous expression of PSA may potentially lead to depletion of high-avidity CD8+ T cell precursors with PSA-specific reactivity. Nevertheless, our findings in this study, will allow us to address this relevant question in experiments with transgenic mice expressing human PSA in prostate tissue, once such model is available on C57Bl/6 background.

In the last study, we investigated the role of activation of antigen-presenting cells (APCs) by the plasmid DNA backbone in the induction of CTL responses by DNA vaccines. A single immunization with pVax-PSA plasmid induced stronger CTL responses in wild-type than in TLR9- or MyD88-deficient mice. These results suggest that immunostimulatory activity of CpG-DNA is essential for efficient priming of CTLs after a single DNA immunization.

Collectively, our findings demonstrate that DNA vaccines represent a promising modality for induction of PSA-specific T-cell responses. Detailed characterization of immunogenicity of pVax-PSA plasmid in C57Bl/6 mice establishes an experimental background for further evaluation of various strategies to increase its potency in inducing CTL responses in a PSA-transgenic murine model.