Abstract B04: Identification of T-cell receptors targeting mutant KRAS in pancreatic cancer
Authors: Bear Adham S. , Rech Andrew J. , Richman Lee P. , O'Hara Mark H. , Linette Gerald P. , Carreno Beatriz M. , Vonderheide Robert H. 
Published: March 2020

Pathologic somatic gene mutations within the KRAS proto-oncogene occur in greater than 90% of pancreatic ductal adenocarcinomas (PDA). These mutations are often conserved involving the codon 12 position and most frequently result in G12D, G12V, or G12R transitions. Unfortunately, no mutant KRAS (mKRAS)-targeted therapies exist to date. We hypothesize conserved hot-spot KRAS mutations may serve as cancer neoantigens for targeted immune-based therapeutic approaches. Using a validated in silico epitope prediction pipeline, we identified mKRAS epitopes with predicted strong binding affinity to the highly prevalent MHC class I alleles HLA-A*03:01, HLA-A*11:01, and HLA-B*07:02. We then confirmed the processing, presentation, and binding of putative mKRAS epitopes using HPLC-tandem mass spectrometry and competitive peptide-binding assays. The immunogenicity of bona fide mKRAS epitopes was confirmed by the in vitro generation of mKRAS-specific CD8+ T cell responses using healthy donor blood samples that were restricted to HLA-A*03:01, -A*11:01, or -B*07:02 as determined by cytokine release assay and peptide/MHC multimer staining. Following T-cell receptor (TCR) sequencing, transgenic expression of mKRAS-specific TCRs on primary CD8+ T cells conferred cytotoxic potential against KRAS-mutated tumor cell lines expressing target HLA/mKRAS combinations. Importantly, mKRAS-specific CD8+ T cells did not cross-react with wild-type or alternatively mutated KRAS epitopes as measured by cytokine release or cytotoxicity assays. Based on these data, we initiated an adjuvant vaccine clinical study at the University of Pennsylvania Abramson Cancer Center targeting mKRAS in patients with resected PDA. The objective of this clinical study is to identify further mKRAS-specific TCR sequences for future adoptive T-cell therapy applications. To date, a total of 7 subjects have been enrolled, and 3 subjects have been vaccinated against targeted mKRAS short peptides utilizing a dendritic cell-based platform. Results from the first vaccinated subject have demonstrated the generation of a KRAS G12V-specific CD8+ T-cell response restricted to HLA-A*11:01 following vaccination with &gt;10% mKRAS-specific T cells isolated following primary in vitro expansion of CD8+ T cells using antigen-pulsed dendritic cells as measured by peptide/MHC multimer staining. TCR sequencing has been completed and functional studies to validate antigen specificity and tumor cytotoxicity are in process. The results of these studies will serve to develop a widely applicable T-cell therapy for PDA patients, and will have implications for the large number of patients with other KRAS-mutated cancers.

Citation Format: Adham S. Bear, Andrew J. Rech, Lee P. Richman, Mark H. O'Hara, Gerald P. Linette, Beatriz M. Carreno, Robert H. Vonderheide. Identification of T-cell receptors targeting mutant KRAS in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr B04.