Class I tetramer technology takes advantage of the exquisite specificity of the T cell-peptide- MHC interaction to label antigen specific T cells. This project addresses the use of HLA-A2 tetramer probes for the measurement of antigen specific T cells as an immunologic marker for predicting effective immunization. Over the last several years, multiple investigators have identified CD8+ peptide epitopes in both infectious disease and tumor antigen systems. Identification of antigen specific epitopes makes it feasible to design specific MHC (HLA) tetramers. Epitopes that are restricted by Class I HLA-A2 have been identified for various melanomas (MART, gpl00, tyrosinase, MAGEs), breast cancer (HER2) and colon cancer (CEA) antigens, among others. It has also been shown that tetramer probes are capable of quantitating circulating CD8+ T cells specific for some of these epitopes (Mart-l , tyrosinase, CEA). Anti-viral CTL responses (CMV, Flu, EBV) have been quantitated using tetramers, indicating that this method is suitable for measuring anti-viral index immune responses in the context of anti-tumor antigen immunization. Based on these findings, we propose to use tetramers to establish baseline responses to several tumor antigens, as well as to foreign viral proteins from CMV and influenza. Following these determinations, post-immunization monitoring will determine whether tetramers may be used to detect immune augmentation of anti-tumor antigen and anti-viral index responses, and whether these measurements can predict immunization to cancer antigens. To determine whether MHC tetramers can be widely adapted to clinical use, several issues must be delineated concerning their production and application. First, as a biologic reagent, tetramers must prove reproducibility from lot to lot. Secondly, methods must be developed to determine assay reproducibility and limits of detection. The type of sample analyzed, whole blood vs. purified cells and the handling and shipping of the sample can effect the analysis. Finally, conditions established to detect high affinity responses, such as those seen against infectious disease antigens may be different from conditions needed to detect CD8+ T cells specific for tumor antigens, i.e. low affinity responses. The purpose of this proposal is to elucidate the clinical use of MHC-tetramer technology in preparation for wide-scale application to the readout of vaccine trial.
The specific aims of this proposal are to: (1) determine whether HLA-A2 tetramers can identify the T cell response to foreign protein antigens from the PBMC of volunteer donors and cancer patients, (2) determine whether HLA-A2 tetramers can identify the T cell response to tumor antigens from the PBMC of volunteer donors and cancer patients, and, (3) determine whether HLA-A2 tetramers can predict immunization to cancer antigens and whether levels achieved are similar to infectious disease index antigens in human clinical trials of HER2, CEA, MAGE3, and gp100 vaccines.
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