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Background & Aims: Pancreatic stellate cells (PSCs) regulate the development of chronic pancreatitis (CP) and are activated by the cytokine transforming growth factor β (TGFB). Integrins of the αv family promote TGFB signaling in mice, probably by interacting with the Arg-Gly-Asp (RGD) sequence of the TGFB latency-associated peptide, which frees TGFB to bind its cellular receptors. However, little is known about the role of integrins in the development of CP. We investigated the effects of small-molecule integrin inhibitors in a mouse model of CP. Methods: We induced CP in C57BL/6 female mice by repeated cerulein administration. An active RGD peptidomimetic compound (Center for World Health and Medicine [CWHM]-12) was delivered by continuous infusion, starting 3 days before or 5 days after cerulein administration began. Pancreata were collected and parenchymal atrophy, fibrosis, and activation of PSCs were assessed by histologic, gene, and protein expression analyses. We measured CWHM-12 effects on activation of TGFB in co-culture assays in which rat PSC cells (large T immortalized cells [LTC-14]) activate expression of a TGFB-sensitive promoter in reporter cells. Results: Pancreatic tissues of mice expressed messenger RNAs encoding subunits of RGD-binding integrins. Cerulein administration increased expression of these integrins, altered pancreatic cell morphology, and induced fibrosis. The integrin inhibitor CWHM-12 decreased acinar cell atrophy and loss, and substantially reduced fibrosis, activation of PSCs, and expression of genes regulated by TGFB. CWHM-12 also reduced established fibrosis in mice and blocked activation of TGFB in cultured cells. Conclusions: Based on studies of a mouse model of CP and cultured PSCs, integrins that bind RGD sequences activate PSCs and promote the development of pancreatic fibrogenesis in mice. Small-molecule antagonists of this interaction might be developed for treatment of pancreatic fibrotic diseases. Keywords: Signal Transduction, Pancreas, Inflammation, Peptidomimetic

Racecadotril (acetorphan) is a neutral endopeptidase (NEP) inhibitor with known antidiarrheal activity in animals and humans; however, in humans, it suffers from shortcomings that might be improved with newer drugs in this class that have progressed to the clinic for nonenteric disease indications. To identify potentially superior NEP inhibitors with immediate clinical utility for diarrhea treatment, we compared their efficacy and pharmacologic properties in a rat intestinal hypersecretion model. Racecadotril and seven other clinical-stage inhibitors of NEP were obtained or synthesized. Enzyme potency and specificity were compared using purified peptidases. Compounds were orally administered to rats before administration of castor oil to induce diarrhea. Stool weight was recorded over 4 hours. To assess other pharmacologic properties, select compounds were orally administered to normal or castor oil–treated rats, blood and tissue samples collected at multiple time points, and active compound concentrations determined by mass spectroscopy. NEP enzyme activity was measured in tissue homogenates. Three previously untested clinical NEP inhibitors delayed diarrhea onset and reduced total stool output, with little or no effect on intestinal motility assessed by the charcoal meal test. Each was shown to be a potent, highly specific inhibitor of NEP. Each exhibited greater suppression of NEP activity in intestinal and nonintestinal tissues than did racecadotril and sustained this inhibition longer. These results suggest that newer clinical-stage NEP inhibitors originally developed for other indications may be directly repositioned for treatment of acute secretory diarrhea and offer advantages over racecadotril, such as less frequent dosing and potentially improved efficacy.

Background Facioscapulohumeral dystrophy (FSHD) is a progressive muscle disease caused by mutations that lead to epigenetic derepression and inappropriate transcription of the double homeobox 4 (DUX4) gene in skeletal muscle. Drugs that enhance the repression of DUX4 and prevent its expression in skeletal muscle cells therefore represent candidate therapies for FSHD. Methods We screened an aggregated chemical library enriched for compounds with epigenetic activities and the Pharmakon 1600 library composed of compounds that have reached clinical testing to identify molecules that decrease DUX4 expression as monitored by the levels of DUX4 target genes in FSHD patient-derived skeletal muscle cell cultures. Results Our screens identified several classes of molecules that include inhibitors of the bromodomain and extra-terminal (BET) family of proteins and agonists of the beta-2 adrenergic receptor. Further studies showed that compounds from these two classes suppress the expression of DUX4 messenger RNA (mRNA) by blocking the activity of bromodomain-containing protein 4 (BRD4) or by increasing cyclic adenosine monophosphate (cAMP) levels, respectively. Conclusions These data uncover pathways involved in the regulation of DUX4 expression in somatic cells, provide potential candidate classes of compounds for FSHD therapeutic development, and create an important opportunity for mechanistic studies that may uncover additional therapeutic targets. Electronic supplementary material The online version of this article (doi:10.1186/s13395-017-0134-x) contains supplementary material, which is available to authorized users.

Apolipoprotein L1 (ApoL1) is a human serum protein conferring resistance to African trypanosomes, and certain ApoL1 variants increase susceptibility to some progressive kidney diseases. ApoL1 has been hypothesized to function like a pore-forming colicin and has been reported to have permeability effects on both intracellular and plasma membranes. Here, to gain insight into how ApoL1 may function in vivo, we used vesicle-based ion permeability, direct membrane association, and intrinsic fluorescence to study the activities of purified recombinant ApoL1. We found that ApoL1 confers chloride-selective permeability to preformed phospholipid vesicles and that this selectivity is strongly pH-sensitive, with maximal activity at pH 5 and little activity above pH 7. When ApoL1 and lipid were allowed to interact at low pH and were then brought to neutral pH, chloride permeability was suppressed, and potassium permeability was activated. Both chloride and potassium permeability linearly correlated with the mass of ApoL1 in the reaction mixture, and both exhibited lipid selectivity, requiring the presence of negatively charged lipids for activity. Potassium, but not chloride, permease activity required the presence of calcium ions in both the association and activation steps. Direct assessment of ApoL1-lipid associations confirmed that ApoL1 stably associates with phospholipid vesicles, requiring low pH and the presence of negatively charged phospholipids for maximal binding. Intrinsic fluorescence of ApoL1 supported the presence of a significant structural transition when ApoL1 is mixed with lipids at low pH. This pH-switchable ion-selective permeability may explain the different effects of ApoL1 reported in intracellular and plasma membrane environments.