In vitro generated cytolytic T lymphocytes reactive against head and neck cancer recognize multiple epitopes presented by HLA-A2, including peptides derived from the p53 and MDM-2 proteins.
In previous studies, we were successful in generating HLA-A2-restricted CD8+ CTLs reactive with head and neck carcinomas (HNCs) in 4/10 cases using traditional mixed lymphocyte tumor cultures (MLTCs) employing a semi-allogeneic HLA-A2+ HNC cell line, PCI-13, as the stimulator of normal HLA-A2+ donor T lymphocytes. However, these T cell lines contained only 1-1.5% HLA-A2-restricted, tumor-reactive CD8+ CTLs, as assessed by both limiting dilution and IFN-gamma ELISPOT assays. In order to increase the success rate in generating such HNC-reactive CTL lines, we modified the procedure to allow for T cell crosspriming by autologous DCs pulsed with PCI-13 lysates. In all three attempts, HLA-A2-restricted effector T cell lines were obtained that contained PCI-13-reactive CD8+ T cells at frequencies as high as 1 in 6. These cultured bulk lines recognized at least five predominant HLA-A2-restricted epitopes based on ELISPOT fingerprinting of HPLC-fractionated, naturally presented PCI-13-derived peptides. Two of these epitopes appear to be derived from the p53 and MDM-2 proteins overexpressed by the PCI-13 cell line. Interestingly, the synthetic wild type sequence p53 (264-272) and MDM-2 (53-61) peptides were able to drive in vitro generation of tumor-specific CTLs from the PBMCs of normal HLA-A2+ donors. However, this MDM-2 peptide was not able to elicit responses from HLA-A2+ patients with HNC in short-term in vitro cultures. Overall, these data suggest that tumor lysate-loaded DCs elicit a broad repertoire of CTL responses, some of which are directed against peptides derived from cell cycle regulatory proteins that may prove to be of clinical significance in the therapy of HNC.
University of Pittsburgh Cancer Institute, Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
Tadao
T
Asai
Walter J
WJ
Storkus
Jan
J
Mueller-Berghaus
William
W
Knapp
Albert B
AB
DeLeo
Kazuaki
K
Chikamatsu
Theresa L
TL
Whiteside
D000595:Amino Acid Sequence; D017951:Antigen Presentation; D000951:Antigens, Neoplasm / Q000737:chemistry / Q000276:immunology*; D002277:Carcinoma / Q000276:immunology; D002460:Cell Line; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000276:immunology; D018984:Epitopes, T-Lymphocyte / Q000737:chemistry / Q000276:immunology; D015789:HLA-A2 Antigen / Q000032:analysis / Q000502:physiology; D006258:Head and Neck Neoplasms / Q000276:immunology*; D006801:Humans; D013058:Mass Spectrometry; D009687:Nuclear Proteins; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000276:immunology*; D051736:Proto-Oncogene Proteins c-mdm2; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000276:immunology*
D000951:Antigens, Neoplasm; D018984:Epitopes, T-Lymphocyte; D015789:HLA-A2 Antigen; C474137:MDM-2 peptide 53-61, human; D009687:Nuclear Proteins; D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; C474136:p53 peptide 264-272, human; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2