Small molecule activates citrullination through targeting PAD2.

Citrullination is a post-translational modification catalysed by peptidyl arginine deiminase (PAD) enzymes, and dysregulation of protein citrullination is involved in various pathological disorders. During the past decade, a panel of citrullination inhibitors has been developed, while small molecules activating citrullination have rarely been reported so far. In this study, we screened citrullination activator using an antibody against citrullinated histone H3 (cit-H3), and a natural compound demethoxycurcumin (DMC) significantly activated citrullination. The requirement of PAD2 for DMC-activated citrullination was confirmed by a loss of function assay. Notably, DMC directly engaged with PAD2, and showed binding selectivity among PAD family enzymes. Point mutation assay indicated that residue E352 is essential for DMC targeting PAD2. Consistently, DMC induced typical phenotypes of cells with dysregulation of PAD2 activity, including citrullination-associated cell apoptosis and DNA damage. Overall, our study not only presents a strategy for rationally screening citrullination activators, but also provides a chemical approach for activating protein citrullination. This article is part of the Theo Murphy meeting issue 'The virtues and vices of protein citrullination'.

Liquidambaric acid inhibits Wnt/ß-catenin signaling and colon cancer via targeting TNF receptor-associated factor 2.

Wnt/ß-catenin signaling is a well-established driver of colon cancer; however, a targeted therapeutic agent has not reached clinics yet. In the present study, we report that the natural compound liquidambaric acid (LDA) inhibits oncogenic Wnt/ß-catenin signaling in vitro and in vivo through its direct target tumor necrosis factor receptor-associated factor 2 (TRAF2). Mechanistically, TRAF2 positively regulates Wnt signaling by interacting with the N-terminal of ß-catenin via its TRAF-C domain; this interaction is disrupted in presence of LDA. Particularly, a TRAF2/ß-catenin/TCF4/TNIK complex is present in colon cancer cells, where TRAF2 is indispensable for the complex formation, and TRAF2/ß-catenin and ß-catenin/TCF4 interactions are disrupted upon LDA treatment. Our findings not only highlight that TRAF2 is an oncogenic regulator of Wnt/ß-catenin signaling and colon cancer but also provide a lead compound targeting TRAF2 for cancer therapy.

Small molecule targeting topoisomerase 3ß for cancer therapy.

DNA topoisomerases are proved cancer therapeutic targets with clinically successful anticancer drugs for decades. However, the role of RNA topoisomerase (TOP3ß) remained mysterious especially in cancer, and no targeted agent has been reported yet. In a target identification assay of anti-cancer compound using a modified DrugTargetSeqR strategy, mutation of TOP3B was detected in cancer cells acquired resistance to cinobufagin (CBG), a key compound of Huachansu that has been approved for cancer therapy in China. We demonstrated that CBG directly engaged with TOP3ß, and promoted TOP3ß depletion in wildtype but not mutant cancer cells. Notably, knockout of TOP3ß in cancer cells significantly reduced tumor enlargement but not initiation, and inhibited colony formation upon nutrient deprivation. We also demonstrated that CBG induced formation of stress granule, RNA-loop and asymmetric DNA damages in cancer cells, and all these phenotypes were significantly attenuated in TOP3B knockout cells. Of note, examination of a panel of cancer cell lines revealed associations among cell growth inhibition and induction of DNA damage as well as TOP3B depletion upon CBG treatment. Our findings not only highlighted TOP3ß as a promising therapeutic target of cancer, but also identified CBG as a lead chemical inhibitor of TOP3ß for cancer therapy.