Selective inhibition of BET proteins reduces pancreatic damage and systemic inflammation in bile acid- and fatty acid ethyl ester- but not caerulein-induced acute pancreatitis.

OBJECTIVES: To evaluate the therapeutic potential of I-BET-762, an inhibitor of the bromodomain and extra-terminal (BET) protein family, in experimental acute pancreatitis (AP). METHODS: AP was induced by retrograde infusion of taurolithocholic acid sulphate into the biliopancreatic duct (TLCS-AP) or 2 intraperitoneal (i.p.) injections of ethanol and palmitoleic acid 1 h apart (FAEE-AP) or 12 hourly i.p. injections of caerulein (CER-AP). In all treatment groups, I-BET-762 (30 mg/kg, i.p.) was administered at the time of disease induction and again 12 h later. AP severity was assessed at 24 h by serum biochemistry, multiple cytokines and histopathology. RESULTS: TLCS-AP, FAEE-AP and CER-AP resulted in characteristic elevations in serum amylase and cytokine levels, increased pancreatic trypsin and myeloperoxidase activity, typical pancreatic histopathological changes and lung injury. Treatment with I-BET-762 significantly reduced biochemical, cytokine and histopathological responses in TLCS-AP and FAEE-AP, but not CER-AP. CONCLUSIONS: These results suggest that in different forms of AP there are significant differences in the epigenetic control of gene transcription contributing to the severity of disease responses. There is therapeutic potential in targeting bromodomains for the treatment of gallstone- and alcohol-related pancreatitis.

Caffeine protects against experimental acute pancreatitis by inhibition of inositol 1,4,5-trisphosphate receptor-mediated Ca2+ release.

OBJECTIVE: Caffeine reduces toxic Ca2+ signals in pancreatic acinar cells via inhibition of inositol 1,4,5-trisphosphate receptor (IP3R)-mediated signalling, but effects of other xanthines have not been evaluated, nor effects of xanthines on experimental acute pancreatitis (AP). We have determined effects of caffeine and its xanthine metabolites on pancreatic acinar IP3R-mediated Ca2+ signalling and experimental AP. DESIGN: Isolated pancreatic acinar cells were exposed to secretagogues, uncaged IP3 or toxins that induce AP and effects of xanthines, non-xanthine phosphodiesterase (PDE) inhibitors and cyclic adenosine monophosphate and cyclic guanosine monophosphate (cAMP/cGMP) determined. The intracellular cytosolic calcium concentration ([Ca2+]C), mitochondrial depolarisation and necrosis were assessed by confocal microscopy. Effects of xanthines were evaluated in caerulein-induced AP (CER-AP), taurolithocholic acid 3-sulfate-induced AP (TLCS-AP) or palmitoleic acid plus ethanol-induced AP (fatty acid ethyl ester AP (FAEE-AP)). Serum xanthines were measured by liquid chromatography-mass spectrometry. RESULTS: Caffeine, dimethylxanthines and non-xanthine PDE inhibitors blocked IP3-mediated Ca2+ oscillations, while monomethylxanthines had little effect. Caffeine and dimethylxanthines inhibited uncaged IP3-induced Ca2+ rises, toxin-induced Ca2+ release, mitochondrial depolarisation and necrotic cell death pathway activation; cAMP/cGMP did not inhibit toxin-induced Ca2+ rises. Caffeine significantly ameliorated CER-AP with most effect at 25 mg/kg (seven injections hourly); paraxanthine or theophylline did not. Caffeine at 25 mg/kg significantly ameliorated TLCS-AP and FAEE-AP. Mean total serum levels of dimethylxanthines and trimethylxanthines peaked at >2 mM with 25 mg/kg caffeine but at <100 µM with 25 mg/kg paraxanthine or theophylline. CONCLUSIONS: Caffeine and its dimethylxanthine metabolites reduced pathological IP3R-mediated pancreatic acinar Ca2+ signals but only caffeine ameliorated experimental AP. Caffeine is a suitable starting point for medicinal chemistry.

Potent antimyeloma activity of the novel bromodomain inhibitors I-BET151 and I-BET762.

The bromodomain and extraterminal (BET) protein BRD2-4 inhibitors hold therapeutic promise in preclinical models of hematologic malignancies. However, translation of these data to molecules suitable for clinical development has yet to be accomplished. Herein we expand the mechanistic understanding of BET inhibitors in multiple myeloma by using the chemical probe molecule I-BET151. I-BET151 induces apoptosis and exerts strong antiproliferative effect in vitro and in vivo. This is associated with contrasting effects on oncogenic MYC and HEXIM1, an inhibitor of the transcriptional activator P-TEFb. I-BET151 causes transcriptional repression of MYC and MYC-dependent programs by abrogating recruitment to the chromatin of the P-TEFb component CDK9 in a BRD2-4-dependent manner. In contrast, transcriptional upregulation of HEXIM1 is BRD2-4 independent. Finally, preclinical studies show that I-BET762 has a favorable pharmacologic profile as an oral agent and that it inhibits myeloma cell proliferation, resulting in survival advantage in a systemic myeloma xenograft model. These data provide a strong rationale for extending the clinical testing of the novel antimyeloma agent I-BET762 and reveal insights into biologic pathways required for myeloma cell proliferation.

Discovery of epigenetic regulator I-BET762: lead optimization to afford a clinical candidate inhibitor of the BET bromodomains.

The bromo and extra C-terminal domain (BET) family of bromodomains are involved in binding epigenetic marks on histone proteins, more specifically acetylated lysine residues. This paper describes the discovery and structure-activity relationships (SAR) of potent benzodiazepine inhibitors that disrupt the function of the BET family of bromodomains (BRD2, BRD3, and BRD4). This work has yielded a potent, selective compound I-BET762 that is now under evaluation in a phase I/II clinical trial for nuclear protein in testis (NUT) midline carcinoma and other cancers.

Orexins and the treatment of obesity.

Orexin-A and -B are two peptides derived by proteolytic cleavage from a 130-amino acid precursor, prepro-orexin, which were recently isolated from the rat hypothalamus. Orexin-A is fully conserved across mammalian species, whilst rat and human orexin-B differ by two amino acids. These peptides bind to two Gq-coupled receptors, termed orexin-1 and orexin-2. The receptors are 64% homologous and highly conserved across species. Orexin-A is equipotent at orexin-1 and orexin-2 receptors, whilst orexin-B displays moderate (approximately 10 fold) selectivity for orexin-2 receptors. The distribution and pharmacology of the orexin peptides and their receptors indicate that they play a role in various regulatory systems including energy homeostasis and the regulation of feeding, the evidence for which is reviewed here.

Anorectic, thermogenic and anti-obesity activity of a selective orexin-1 receptor antagonist in ob/ob mice.

A single dose of the orexin-1 (OX1) receptor antagonist 1-(2-methylbenzoxazol-6-yl)-3-[1,5] naphthyridin-4-yl urea hydrochloride (SB-334867-A) reduces orexin-A-induced feeding and natural feeding in Sprague Dawley rats. In this study, the anti-obesity effects of SB-334867-A were determined in genetically obese (ob/ob) mice dosed with SB-334867-A (30 mg/kg, i.p.) once daily for 7 days, and then twice daily for a further 7 days. SB-334867-A reduced cumulative food intake and body weight gain over 14 days. Total fat mass gain, determined by Dual Emission X-ray Absorptiometry, was reduced, while gain in fat-free mass was unchanged. Fasting (5 h) blood glucose was also reduced at the end of the study, with a trend to reduced plasma insulin. Interscapular brown adipose tissue (BAT) weight was reduced, the tissue was noticeably darker in colour and quantitative PCR (TaqMan) analysis of this tissue showed a trend to an increase in uncoupling protein-1 mRNA expression, suggesting that SB-334867-A might stimulate thermogenesis. This was confirmed in a separate study in which a single dose of SB-334867-A (30 mg/kg, i.p.) increased metabolic rate over 4 h in ob/ob mice. OX1 receptor mRNA was detected in BAT, and its expression was increased by 58% by treatment with SB-334867-A. This is the first demonstration that OX1 receptor antagonists have potential as both anti-obesity and anti-diabetic agents.