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Functional genetics-directed identification of novel pharmacological inhibitors of FAS- and TNF-dependent apoptosis that protect mice from acute liver failure.

ABSTRACT: shRNA-mediated gene-silencing technology paired with cell-based functional readouts reveals potential targets directly, providing an opportunity to identify drugs against the target without knowing the precise role of the target in the pathophysiological processes of interest. By screening a lentiviral shRNA library targeting for major components of human signaling pathways and known drug targets, we identified and validated both canonical as well as 52 novel mediators of FAS and TNF ligand-induced apoptosis. Presence of potential therapeutic targets among these mediators was confirmed by demonstration of in vivo activity of siRNAs against four identified target candidates that protected mice from acute liver failure (ALF), a life-threatening disease with known involvement of death receptor (DR)-mediated apoptosis. Network-based modeling was used to predict small-molecule inhibitors for several candidate apoptosis mediators, including somatostatin receptor 5 (SSTR5) and a regulatory subunit of PP2A phosphatase, PPP2R5A. Remarkably, pharmacological inhibition of either SSTR5 or PPP2R5A reduced apoptosis induced by either FASL or TNF in cultured cells and dramatically improved survival in several mouse models of ALF. These results demonstrate the utility of loss-of-function genetic screens and network-based drug-repositioning methods for expedited identification of targeted drug candidates and revealed pharmacological agents potentially suitable for treatment of DR-mediated pathologies.

SUBMITTER: Komarov AP

PROVIDER: S-EPMC4823946 | biostudies-literature |

REPOSITORIES: biostudies-literature

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| S-EPMC7921426 | biostudies-literature

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Project description:Uncontrolled systemic activation of the immune system is an early initiating event that leads to development of acute fulminant liver failure (FLF) in mice after treatment with agonistic Fas mAb. In this study, we demonstrate that treatment of mice with N-acetylcysteine (NAC), an ROS scavenger and glutathione (GSH) precursor, almost completely abolished Fas mAb-induced FLF through suppression of Vα14iNKT cell activation, IFN-γ signaling, apoptosis and nitrotyrosine formation in liver. In addition, enrichment of the liver with GSH due to Vα14iNKT cells deficiency, induced an anti-inflammatory response in the liver of Jα18(-/-) mice that inhibited apoptosis, nitrotyrosine formation, IFN-γ signaling and effector functions. In summary, we propose a novel and previously unrecognized pro-inflammatory and pro-apoptotic role for endogenous ROS in stimulating Th1 signaling in Vα14iNKT cells to promote the development of FLF. Therefore, our study provides critical new insights into how NAC, a ROS scavenger, regulates Th1 signaling in intrahepatic Vα14iNKT cells to impact inflammatory and pathological responses.

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| S-EPMC40509 | biostudies-other

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