Development of a Novel Methodology for Improving CTL Recognition of Prostate Specific Antigen (PSA) for the Immunotherapy of Prostate Cancer
Final rept. 1 Mar 2000-28 Feb 2003
ROCHESTER UNIV NY
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The major goal of the research was to develop a novel methodology for improving T cell epitopes. The underlying hypothesis is that T cells capable of recognizing tumor-associated antigens TAA are present but often difficult to activate. However, once activated such T cells might be effective against tumors due to the less stringent triggering requirements of mature effectors. We developed a novel bacterial expression system for screening the epitopes of PSA, a known TAA. We have employed a saturation mutagenesis technique to a PSA peptide epitope identified. Expression libraries constructed corresponding to each position in the peptide were screened functionally. Plasmids from clones showing enhanced activity were sequenced. Peptides were synthesized and tested in a functional assay and showed enhanced activity with the T cell hybridoma. Using a competitive binding assay for MHC binding, we showed that some mutants enhance T cell activation by binding better to MHC whereas others appear to improve interaction with the T cell receptor. Immunization with a mixture of altered peptide ligand and the wild-type peptide can enhance the frequency of responding cells. The findings show that altered peptide ligands can be discovered using this novel methodology and should be applicable to other tumor antigens.
- Anatomy and Physiology
- Medicine and Medical Research