Pancreatic ductal adenocarcinoma (PDAC) is resistant to virtually all chemo-and targeted therapeutic approaches. Epigenetic regulators represent a novel class of drug targets. Among them, BET and HDAC proteins are central regulators of chromatin structure and transcription, and preclinical evidence suggests effectiveness of combined BET and HDAC inhibition in PDAC. Here, we describe that TW9, a newly generated adduct of the BET inhibitor (+)-JQ1 and class I HDAC inhibitor CI994, is a potent dual inhibitor simultaneously targeting BET and HDAC proteins. TW9 has a similar affinity to BRD4 bromodomains as (+)-JQ1 and shares a conserved binding mode, but is significantly more active in inhibiting HDAC1 compared to the parental HDAC inhibitor CI994. TW9 was more potent in inhibiting tumor cell proliferation compared to (+)-JQ1, CI994 alone or combined treatment of both inhibitors. Sequential administration of gemcitabine and TW9 showed additional synergistic antitumor effects. Microarray analysis revealed that dysregulation of a FOSL1-directed transcriptional program contributed to the antitumor effects of TW9. Our

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The genome is exposed daily to many deleterious factors. Ubiquitination is a mechanism that regulates several crucial cellular functions, allowing cells to react upon various stimuli in order to preserve their homeostasis. Ubiquitin ligases act as specific regulators and actively participate among others in the DNA damage response (DDR) network. UBE4B is a newly identified member of E3 ubiquitin ligases that appears to be overexpressed in several human neoplasms. The aim of this review is to provide insights into the role of UBE4B ubiquitin ligase in DDR and its association with p53 expression, shedding light particularly on the molecular mechanisms of carcinogenesis.

The tumor suppressor protein p53 is inactivated in the majority of human cancers and remains a prime target for developing new drugs to reactivate its tumor suppressing activity for anticancer therapies. The oncogenic p53 mutant Y220C accounts for approximately 125,000 new cancer cases per annum and is one of the most prevalent p53 mutants overall. It harbors a narrow, mutationally induced pocket at the surface of the DNA-binding domain that destabilizes p53, leading to its rapid denaturation and aggregation. Here, we present the structure-guided development of high-affinity small molecules stabilizing p53-Y220C in vitro, along with the synthetic routes developed in the process, in vitro structure− activity relationship data, and confirmation of their binding mode by protein X-ray crystallography. We disclose two new chemical probes displaying sub-micromolar binding affinity in vitro, marking an important milestone since the discovery of the first small-molecule ligand of Y220C in 2008. New chemical probe JC744 displayed a K d = 320 nM, along with potent in vitro protein stabilization. This study, therefore, represents a significant advance toward highaffinity Y220C ligands for clinical evaluation.

The extremophile Alvinella pompejana, an annelid worm living on the edge of hydrothermal vents in the Pacific Ocean, is an excellent model system for studying factors that govern protein stability. Low intrinsic stability is a crucial factor for the susceptibility of the transcription factor p53 to inactivating mutations in human cancer. Understanding its molecular basis may facilitate the design of novel therapeutic strategies targeting mutant p53. By analyzing expressed sequence tag (EST) data, we discovered a p53 family gene in A. pompejana. Protein crystallography and biophysical studies showed that it has a p53/p63-like DNA-binding domain (DBD) that is more thermostable than all vertebrate p53 DBDs tested so far, but not as stable as that of human p63. We also identified features associated with its increased thermostability. In addition, the A. pompejana homolog shares DNA-binding properties with human p53 family DBDs, despite its evolutionary distance, consistent with a potential role in maintaining genome integrity. Through extensive structural and phylogenetic analyses, we could further trace key evolutionary events that shaped the structure, stability, and function of the p53 family DBD over time, leading to a potent but vulnerable tumor suppressor in humans.