Case Study: Epitope Discovery in Prostate Cancer

The Use of H-2Db / HCIRNKSVI (PSA 65-73) Custom Pro5® MHC Class I Pentamer in the Characterization of an Immunodominant CTL Epitope of PSA in Mice

Pavlenko, M. et al. (2005) Identification of an immunodominant H-2Db-restricted CTL epitope of human PSA. Prostate 64: 50-59. [PubMedID: 15651071]

Prostate cancer is a serious condition affecting 1 in 6 men. Prostate specific antigen (PSA) expression is increased in prostate cancer and so is exploited not only for diagnosing and monitoring prostate cancer, but also as a potential target for immunotherapy. By working with Custom Pro5® Pentamers from ProImmune, Pavlenko et al. have identified and validated an immunodominant cytotoxic T lymphocyte (CTL) epitope of PSA in C57BL/6 mice. A combined bioinformatics approach using the SYFPEITHI website (, and biochemical MHC-peptide stabilization assays was used to define the candidate epitope (H-2Db / HCIRNKSVI) and a custom Pro5® Pentamer was synthesized.

PSA-specific CTLs were induced by immunizing mice with a plasmid expressing PSA (pVax-PSA). By using both functional assays, intracellular cytokine staining and detection of PSA-specific CD8+ T cells with Pro5® Pentamer staining, the authors were able to demonstrate correlating frequencies of both IFN-g positive and Pentamer positive T cells following in vitro stimulation with the specific peptide.

The authors conclude that “Pentamer technology enables detection of T cell receptor specific T-cell populations and allows, in combination with functional assays, the discrimination between anergy and tolerance induction during effector responses. H-2Db Pentamers assembled with this peptide are an efficient tool for the monitoring of PSA-specific CTL responses after DNA vaccination”.


Figure 1: 12 days after immunization as described, the splenocytes were restimulated for 5 days in vitro with 1nM of psa65-73 (HCIRNKSVI) peptide (A). The PSA-specific T cells were detected by staining with the H-2Db/HCIRNKSVI Pentamer (B). The splenocytes were stimulated for 5hr with 10nM of the psa65-73 or GP33 (negative control) peptides and stained with the antibodies against CD8 and IFN-gamma.