RNAi Validation of Pancreatic Cancer Antigens Identified by Cell Surface Proteomics

Pancreatic cancer is one of the most aggressive malignancies that is often diagnosed at a late stage following tumor metastasis. Given the low response rate to current therapies, there is a clear and urgent need for new therapeutic and diagnostic approaches. Proteomics is an emerging tool that offers the opportunity to discover novel antigens elevated in cancer. Considering the large number of marketed drugs that target plasma membrane proteins, and given their amenability to both small molecule and antibody-based technologies, an approach that combines a cell membrane labeling strategy with proteomic (LC-MS/MS) methodology would enable novel cell surface antigens that can act as novel biomarkers and/or targets for drug development to be identified. One major consideration when selecting targets for therapeutics is whether or not the antigen plays a functional role in a disease context. RNA interference (RNAi) permits the downregulation of any given gene, and therefore allows the identification of functionality in the target disease. As such, this tool permits the selection of targets prior to investment in a therapeutic antibody program or an extensive small molecule–screening program. In this chapter, we describe the utility of the cell membrane labeling strategy and LC-MS/MS method that was developed at Celera. Subsequent application of RNAi allowed us to assign functionality to cell surface antigens we identified that were upregulated in pancreatic cancer. By using a combination of a proteomics and an RNAi platform, we demonstrate that it is possible to discover novel therapeutic and diagnostic targets for this devastating disease.