Abstract 4545: Quantitative profiling of Cytochrome P450 2S1 in colorectal cancer by PRM assay

Introduction: Cytochromes P450s (CYPs) constitute a superfamily of xenobiotic metabolising enzymes responsible for metabolism of many pharmaceuticals in the liver. Elevated mRNA levels of specific isoforms, such as CYP2S1, are associated with poor prognosis in colorectal cancer (CRC), and represent novel therapeutic targets for biotransformation of prodrugs to potent cytotoxics at the cancer site. In order to understand the expression of CYP2S1 protein, we have developed a parallel reaction monitoring mass spectrometry (PRM MS) assay to screen CRC samples. Method: Peptides (n=3) uniquely associated with CYP2S1 were synthesized and used as standards to optimise analytical performance (fragmentation conditions, LC retention time, LOQ, LOD, dynamic range) in an Orbitrap Fusion-based PRM MS assay. Levels of CYP2S1 were then determined in protein extracts of CRC, relative to the standards. Results: The PRM MS assay yielded quantitative data over 3 orders of magnitude. CYP2S1 was detected in C106, CaCO2, HCC2998, HT55 and DLD1 cell lines (0.05-1.08pg) and HT55 and DLD1 xenografts (0.29-0.41pg). CYP2S1 was also detected in patient-specific CRC tissues, with elevated levels in Stage III tumours. Conclusions: The PRM MS assay provides a specific, sensitive, high throughput method for identifying CYP2S1 compared to established enzyme and immunoassays. CYP2S1 levels vary considerably across CRC sources highlighting the importance of screening to identify the correct models for new drug development and the right patients for subsequent treatment. Citation Format: Sadr ul Shaheed, Laurence H. Patterson, Klaus Pors, Chris W. Sutton. Quantitative profiling of Cytochrome P450 2S1 in colorectal cancer by PRM assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4545.