Zhang, Yu

Elucidating the structure and function of enteropeptidase (EP) is essential for advancing our understanding of its biological significance, particularly in regulating trypsinogen activation. Using cryo-EM and enzymatic activity, we uncovered the significance of the CUB2 domain in mediating the cleavage of macromolecular substrates. We identified crucial binding loops and key residues for EP’s proteolytic function. The mutation E574A enhanced the proteolytic activity of EP, whereas the mutation N619A diminished its cleavage efficiency, highlighting the importance of surface-charged interactions in modulating EP’s activity. A proteolytic cycle was proposed to deepen our understanding of the trypsinogen activation by EP. This work offers valuable insights into the molecular mechanisms underlying EP’s interaction with its substrate, and opens up avenues for therapeutically modulating EP-mediated proteolysis.

Abstract Pancreatic cancer is the fourth leading cause of cancer related deaths in the United States and continues to be hindered with limited treatment options. One of the current therapeutic regimens for pancreatic cancer often include the nucleoside analog gemcitabine (Gem), while it has been used for many years, the overall survival rate lingers around 6%. New therapies are needed to treat this disease. Fatty acid synthase (FASN) has received increased attention as a potential target for cancer therapy in the last decade. FASN is the key enzyme in the de novo synthesis of palmitate from malonyl-CoA. Palmitate is the precursor other long-chain fatty acids (FA). However, the precise mechanism by which pharmacological interference with endogenous FA biogenesis might facilitate apoptosis in tumor cells remains unresolved. We herein explored the molecular relationship between FASN activity, oxidative stress/redox balance, and the intrinsic apoptotic pathway in FASN overexpressing-dependent pancreatic cancer cells. About 68% of Pancreatic Ductal Carcinomas (PDA) expresses high levels of Fatty acid synthase (FASN), and its expression is correlated with disease free survival. A newly developed FASN inhibitor with excellent pharmaceutical properties was used for our studies TVB-3166 (in vitro) and TVB-3664 (in vivo) and ten different PDX derived cells were used for the studies. Pharmacological blockade of FASN activity significantly increased the NADPH/NADP+ ratio, promoted the production of reactive oxygen species (ROS). Accordingly, the inhibition of endogenous FA synthesis promoted mitochondrial membrane permeabilization, cytochrome c release, and apoptotic cell death. Each step of the FASN inhibition/BH3-only protein axis could be reversed with a ROS scavenger or the FASN end-product palmitate, thus confirming that the delineated pro-apoptotic cascade likely reflects on-target effects of FASN inhibition. Most importantly, the in vivo studies using PaCa-PDX reveal a significant decrease in tumor growth and furthermore a synergistic effect of between TVB-3664 and Gem. In conclusion, our findings uncover a novel FASN-dependent mitochondrial priming that links de novo FA biosynthesis with the intrinsic apoptotic threshold in cancer cells. The discovery that FASN-inhibited cancer cells exist in an apoptosis-prone state highly sensitive to BH3 mimetics warrants clinical exploration in FASN-overexpressing pancreatic cancer patients. Citation Format: Travis Vander Steen, Li Ding, Yu Zhang, George Kemble, Daniel Billadeau, Ruth Lupu. Fatty acid synthase: A new therapeutic target for pancreatic cancer [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 3723.