S004 HPV-Infected Cancers Process and Present HLA-A2–p53(264-272) Peptide Complexes: Implications for p53-Based Immunotherapy

Abstract

Objective: We have shown that the human papillomavirus (HPV) E6 oncoprotein in HPV-16 head and neck squamous cell carcinoma (HNSCC) cells induces proteasomal degradation of p53, which leads to greater CD8 T-cell lysis mediated by enhanced expression of HLAA2-p53(264-272) complexes on the tumor cell surface. By using a staining reagent capable of recognizing these complexes, we will be able to quantify and compare expression of HLA-A2p53(264-272) complexes in HPVinfected HNSCC cells and what the effect of p53 destabilization has on this expression. Methods: We used a multimeric, soluble T-cell receptor– like reagent to quantitate HLA-A2–p53(264-272) complexes (264scTCR/multimer) and confirmed its specificity and sensitivity using T2 cells pulsed with various peptides. We then utilized the 264scTCR/multimer to directly measure tumor cell presentation of HLA-A2–p53 (264-272) complexes in HPV-infected cancer cells, which rapidly degraded p53. Various strategies are currently being used to improve staining of tumor cell lines by the 264scTCR, especially in cases where the cells are known to present HLA-A2–p53(264-272) complexes. Results: We observed enhanced staining of CaSki cells compared with other cell lines known to express the HLA-A2– p53(264-272) complex, including several head and neck cell lines. Pretreatment with interferon gamma exhibited the strongest staining, likely due to up-regulation of HLA and antigen-processing proteins. HPV E6–transfected PCI-13 cells exhibited improved staining when stained with 264scTCR/multimer-PE followed by anti-PE antibody. Conclusions: This novel tool, which can be improved upon, provides a method for selecting appropriate candidates best suited for immunotherapy, and also identifies those who may require further pretreatment before beginning an immunotherapeutic regimen.