Abstract 1514: Comparative quantitation of Warburg Effect proteins in FFPE tumor and stroma tissue from 10 breast carcinoma tumors

Abstract

Abstract The observation that cancer cells can undergo aerobic glycolosis (the Warburg Effect) has been the target of a great deal of research in recent years, specifically directed to development of novel agents which can disrupt this pathway and potentially retard cancer growth. Recent studies, by Lisanti and colleagues, have suggested that the critical site of aerobic glycolysis may not be the tumor epithelial cells, but instead the ‘Warburg Effect’ is occurring in the tumor associated fibroblasts and other stromal cells. In order to better define the importance of ‘tumor’ vs. ‘stroma’ components of the Warburg effect, we have developed a quantitative multiplexed assay which can be performed directly in formalin fixed paraffin embedded (FFPE) patient tissue. This multiplex assay is based on the Liquid Tissue®-SRM technology platform, a combination of tissue microdissection, Liquid Tissue® processing which renders dissected tissue to a completely solubilized tryptic digest, and mass spectrometry-based selected reaction monitoring (SRM). For mass spectrometric analysis, we developed 9 different quantitative assays for each of the Warburg Effect components (LDHA, PKM2, FBA(a), PGKI, LDHA, PMI, EnolaseA, and TPI) as well as Caveolin-1 which was identified as a marker of active aerobic glycolysis in tumor fibroblasts. We then quantitated all of these proteins in multiplex on matched tumor and stromal tissue laser microdissected from 10 different FFPE breast carcinoma tumor samples. This analysis demonstrated broadly divergent levels of glycolytic enzyme expression across the set of tumor samples, and in most cases, much higher enzyme expression in tumor cells as opposed to tumor associated stroma cells. However there were several cases where the stromal level of Warburg protein expression was elevated above matched tumor epithelial cell expression. These results demonstrate that by using the Liquid Tissue® technology, we can separately microdissect tumor and stroma in FFPE tumor sections and characterize the specific tumor/stroma components, be they pharmacodynamic analyses, drug target analysis, or seed vs. soil studies. Second, this technology reinforces the capability of SRM based mass spectrometry studies to function in massively multiplexed fashion, where 20 to 40 or more specific analytes can be quantitated by analysis of 0.5ug of protein lysate from a single FFPE tumor section. Finally, we are in the process of clinically validating this Warburg Effect multiplex assay and will then perform the assay on a larger, completely annotated tumor cohort in order to extend our knowledge of how tumor vs. stroma expression of Warburg effect components impacts tumor progression and therapeutic response. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1514. doi:10.1158/1538-7445.AM2011-1514