Abstract 1373: PeptidePCR: Direct identification of cancer vaccine targets from scant clinical samples

Abstract

Abstract Background: Peptides presented by Human Leukocyte antigen (HLA) class I and II molecules form an important component of the adaptive immune response against tumors. Identifying HLA-bound peptides is therefore crucial to understand the specificity of T cell responses in cancer. Due to sample limitations, it is often difficult or impossible to confirm epitope presentation in clinical biopsy material. To develop truly personal medicine, target epitopes need to be directly identified or validated in patient tumors at the peptide antigen level. Methodology: We have developed a microscale HLA immunoprecipitation protocol which when combined with a tandem mass tag (TMT)-based barcoding approach was able to identify HLA-bound peptides. HLA-complexes were affinity purified from either cultured cells, patient-derived xenograft or melanoma biopsies and the peptide cargo was tagged using TMT barcodes and analyzed using high-resolution mass spectrometry. Like primers in PCR, the carrier peptides and multiplexing of samples amplifies the signal enabling sensitive detection. Results: By using this strategy, we identified HLA-bound peptides from as few as ~1000 cultured cells. We demonstrate the clinical utility of this approach by confirming the presentation of around 1000 peptides from a challenging melanoma biopsy (~1-20mg in total), including 11 well-known T-cell response-inducing epitopes and other potential epitopes derived from Melanoma-associated antigens and neoantigens. Conclusion: This strategy will be useful for studying peptidome subsets in other clinical samples where the amount of tissue available via sampling is limiting, including biopsies taken for autoimmune diseases and infections, or to sample rare cell types in tissue/cell samples, such as different antigen-presenting cell subsets. The approach can directly isolate antigens that are already presented by HLA molecules, thereby increasing the precision of personalized approaches to cancer treatment with less than 1-week turnaround times. Citation Format: Sri H. Ramarathinam, Pouya Faridi, Angela Peng, Pacman Szeto, Nicholas C. Wong, Andreas Behren, Mark Shackleton, Anthony W. Purcell. PeptidePCR: Direct identification of cancer vaccine targets from scant clinical samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1373.