SUMMARY (30 lines max) The vast majority of cancer mortality is due to metastases. A major unmet need in oncology is how to predict and prevent metastatic progression. To address this need, it is imperative we advance our understanding of how tumor cells acquire metastatic capability. Epithelial-to-mesenchymal transition (EMT) is a normal physiologic process in wound healing and development by which polarized epithelial cells undergo biochemical, metabolic, and epigenetic changes and convert to a mesenchymal phenotype, characterized by enhanced migratory and invasive capacity. Cancers co-opt this reprogramming to acquire metastatic ability. Understanding how tumor cells undergo EMT is critical to unraveling how cancer becomes metastatic. Historically, studies have primarily focused on genetic mutations or gene expression changes that trigger EMT and metastasis. But recent work has suggested that non-genetic factors such as metabolites can promote cancer progression, suggesting unexplored areas of cancer biology. To gain a deeper understanding of EMT and capture the sequential changes necessary for cancer to become metastatic, we performed proteomics, metabolomics and transcriptomics at multiple time points as cells underwent EMT. Our multifaceted approach revealed rich new insights. We found that propionate metabolism was dysregulated during EMT. Indeed, we observed increased propionyl-CoA and methylmalonic acid (MMA), a dicarboxylic acid by-product of propionate metabolism, during early EMT. To test the functional effect of elevated MMA, we treated lung and breast cancer cells with MMA. Strikingly, we found that MMA triggered EMT and enhanced migratory and invasive capacity. Prior to our discovery, little was known about propionate metabolism and MMA other than propionate metabolism is dysregulated in rare inborn errors of metabolism collectively referred to as “methylmalonic acidemias.” Some recent studies have also shown that MMA increases in serum with age and is linked to age-related, all-cause mortality. To our knowledge, our findings are the first to demonstrate that propionate metabolism plays a role in EMT and cancer cell progression. This led us to study how propionate metabolism is altered during EMT. Our long-term goals are to define how this pathway is regulated by multiple inducers of metastasis and at multiple points in the pathway, and to define how MMA produced by cancer cells can also influence the tumor microenvironment. Our proposed studies will address a critical need for a greater understanding of the molecular basis of metastasis. Our expectations are that successful completion of the proposed work will impact cancer biology by shifting the paradigm and highlighting critical non-genetic factors that drive cancer progression. Consequently, our work will identify new biomarkers and therapeutic targets that can be exploited to prevent and inhibit metastases.

Project Narrative / Relevance: (3 sentences) Cancer is the 2nd leading cause of death in the US but importantly, the vast majority of cancer-associated mortality is not due to primary tumor growth but to cancer metastasis. Analysis of the changing metabolome during epithelial-to-mesenchymal transition (EMT), a developmental program hijacked by cancer cells to acquire metastatic and drug resistance properties, has led to the novel discovery that methylmalonic acid (MMA), a by- product of propionate metabolism, is rapidly produced and secreted by tumor cells, driving increased invasiveness and chemoresistance in both autocrine and paracrine manners. We propose to: (i) define how MMA production is regulated in cancer, (ii) determine how secreted MMA activates cancer-associated fibroblasts (CAFs) and (iii) characterize how CAFs in turn secrete factors that further promote tumor metastasis with the ultimate goal of defining a new cancer regulatory process, new biomarkers and new potential targets for drug discovery that are needed for personalized therapeutic interventions in metastatic cancer. PHS 398/2590 (Rev. 06/09) Page Continuation Format Page