Structurally diverse mitochondrial branched chain aminotransferase (BCATm) leads with varying binding modes identified by fragment screening

Inhibitors of mitochondrial branched chain aminotransferase (BCATm), identified using fragment screening, are described. This was carried out using a combination of STD-NMR, thermal melt (Tm), and biochemical assays to identify compounds that bound to BCATm, which were subsequently progressed to X-ray crystallography, where a number of exemplars showed significant diversity in their binding modes. The hits identified were supplemented by searching and screening of additional analogues, which enabled the gathering of further X-ray data where the original hits had not produced liganded structures. The fragment hits were optimized using structure-based design, with some transfer of information between series, which enabled the identification of ligand efficient lead molecules with micromolar levels of inhibition, cellular activity, and good solubility

Development of a Series of Kynurenine 3-Monooxygenase Inhibitors Leading to a Clinical Candidate for the Treatment of Acute Pancreatitis

Recently, we reported a novel role for KMO in the pathogenesis of acute pancreatitis (AP). A number of inhibitors of kynurenine 3-monooxygenase (KMO) have previously been described as potential treatments for neurodegenerative conditions and particularly for Huntington’s disease. However, the inhibitors reported to date have insufficient aqueous solubility relative to their cellular potency to be compatible with the intravenous (iv) dosing route required in AP. We have identified and optimized a novel series of high affinity KMO inhibitors with favorable physicochemical properties. The leading example is exquisitely selective, has low clearance in two species, prevents lung and kidney damage in a rat model of acute pancreatitis, and is progressing into preclinical development

Mouse Sphingosine Kinase 1a Is Negatively Regulated through Conventional PKC-Dependent Phosphorylation at S373 Residue

Sphingosine kinase is a lipid kinase that converts sphingosine into sphingosine-1-phosphate, an important signaling molecule with intracellular and extracellular functions. Although diverse extracellular stimuli influence cellular sphingosine kinase activity, the molecular mechanisms underlying its regulation remain to be clarified. In this study, we investigated the phosphorylation-dependent regulation of mouse sphingosine kinase (mSK) isoforms 1 and 2. mSK1a was robustly phosphorylated in response to extracellular stimuli such as phorbol ester, whereas mSK2 exhibited a high basal level of phosphorylation in quiescent cells regardless of agonist stimulation. Interestingly, phorbol ester-induced phosphorylation of mSK1a correlated with suppression of its activity. Chemical inhibition of conventional PKCs (cPKCs) abolished mSK1a phosphorylation, while overexpression of PKC alpha, a cPKC isoform, potentiated the phosphorylation, in response to phorbol ester. Furthermore, an in vitro kinase assay showed that PKC alpha directly phosphorylated mSK1a. In addition, phosphopeptide mapping analysis determined that the S373 residue of mSK1a was the only site phosphorylated by cPKC. Interestingly, alanine substitution of S373 made mSK1a refractory to the inhibitory effect of phorbol esters, whereas glutamate substitution of the same residue resulted in a significant reduction in mSK1a activity, suggesting the significant role of this phosphorylation event. Taken together, we propose that mSK1a is negatively regulated through cPKC-dependent phosphorylation at S373 residueopen

Kynurenine–3–monooxygenase inhibition prevents multiple organ failure in rodent models of acute pancreatitis

Acute pancreatitis (AP) is a common and devastating inflammatory condition of the pancreas that is considered to be a paradigm of sterile inflammation leading to systemic multiple organ dysfunction syndrome (MODS) and death1,2 Acute mortality from AP-MODS exceeds 20%3 and for those who survive the initial episode, their lifespan is typically shorter than the general population4. There are no specific therapies available that protect individuals against AP-MODS. Here, we show that kynurenine-3-monooxygenase (KMO), a key enzyme of tryptophan metabolism5, is central to the pathogenesis of AP-MODS. We created a mouse strain deficient for Kmo with a robust biochemical phenotype that protected against extrapancreatic tissue injury to lung, kidney and liver in experimental AP-MODS. A medicinal chemistry strategy based on modifications of the kynurenine substrate led to the discovery of GSK180 as a potent and specific inhibitor of KMO. The binding mode of the inhibitor in the active site was confirmed by X-ray co-crystallography at 3.2 Å resolution. Treatment with GSK180 resulted in rapid changes in levels of kynurenine pathway metabolites in vivo and afforded therapeutic protection against AP-MODS in a rat model of AP. Our findings establish KMO inhibition as a novel therapeutic strategy in the treatment of AP-MODS and open up a new area for drug discovery in critical illness

Lower crustal vs. mantle wedge fingerprint in the Ecuadorian arc magmas: Contribution of Pb isotopes from the Cotopaxi volcano

International audiencePb isotopes are very sensitive to continental crustal contamination. With these isotopes, we are able to discriminate the contribution of the lower crust from that of the upper crust. With the objective of constraining the genesis of arc magmas by focusing here on the role of the continental crust, we propose a geochemical study on eruptive products of the Cotopaxi volcano. This choice is based on the fact that Cotopaxi is constructed on a thick continental crust and that its magmatic series span a large geochemical diversity, from basaltic andesites to rhyolites (Garrison et al., 2006, Garrison et al., 2011). We provide here 23 new high-precision Pb isotope data obtained on tephras, covering the range from andesites to rhyolites, over a period ranging from pre-Holocene to historical times. Isotopes variations are comprised between 18.980 and 18.923 for 206 Pb/ 204 Pb, 15.629 and 15.640 for 207 Pb/ 204 Pb and 38.717 and 38.765 for 208 Pb/ 204 Pb. Comparing these results with published data, we observe that our data have restricte

The genetic relationship between andesites and dacites at Tungurahua volcano, Ecuador

Co-auteurInternational audienceVolcanic eruptions of intermediary and silica-rich magmas (andesites, dacites and rhyolites) in convergent arcsettings generate voluminous and explosive eruptions that can strongly affect human activity and have signifi-cant environmental impacts. It is therefore crucial to understand how these magmas are generated in order toanticipate their potential impact. At convergent margins, primitive magmas (primitive basalts and/or andesites)are derived from the mantle wedge and they are progressively modified by physical and chemical processes op-erating between the melting zone and the surface to produce silica-rich magmas.In order to elucidate the relationship between andesites and dacites, we focus on Tungurahua volcano, located inthe Ecuadorian Andes. We collected a set of samples comprising such lithologies that were erupted during thelast 3000 year BP. This relatively short period of time allows us to assume that the geodynamic parameters re-main constant. Petrology and major-trace element compositions of these lavas have already been examined,and so we performed a complementary Pb-Sr isotope study in order to determine the nature and origin of thecomponents involved in andesite and dacite genesis. Sr isotopes range from 0.70417 to 0.70431, and Pb isotopecompositions range from 18.889 to 19.154 for206Pb/204Pb, from 15.658 to 15.696 for207Pb/204Pb, and from38.752 to 38.918 for208Pb/204Pb. Dacites display a remarkably homogeneous Pb isotopic composition, withhigher206Pb/204Pb values for a given207-208Pb/204Pb compared to andesites. Andesites show notable207Pb/206Pb variations for a given SiO2content, whereas dacites have lower and homogenous207Pb/206Pb values.Andesite and dacite altogether plot in a roughly triangular distribution, with dacitic magmas systematically plot-ting at the high SiO2and87Sr/86Sr and low207Pb/206Pbfields. Based on our new dataset, we show that at least 3different components are required to explain the Tungurahua compositional and isotope variation: one corre-sponds to the mantle, the second has a deep origin (slab component or lower crust), and a mixture betweenthese two components explains andesite heterogeneity. The third component is derived from the underlyingupper continental crust. While andesites are derived from deep components, dacites are derived from the andes-itic magmas that underwent an assimilation-fractional crystallization (AFC) process with incorporation of thelocal metamorphic basement. Finally, we used the geochemical and isotopic data to produce a model of the mag-matic plumbing system beneath Tungurahua, consistent with geophysical and experimental petrology con-straints. We conclude that melt migration and storage in the upper crust appears to be a key parameter forcontrolling volcanic behavior though time

The genetic relationship between andesites and dacites at Tungurahua volcano, Ecuador

Volcanic eruptions of intermediary and silica-rich magmas (andesites, dacites and rhyolites) in convergent arc settings generate voluminous and explosive eruptions that can strongly affect human activity and have significant environmental impacts. It is therefore crucial to understand how these magmas are generated in order to anticipate their potential impact. At convergent margins, primitive magmas (primitive basalts and/or andesites) are derived from the mantle wedge and they are progressively modified by physical and chemical processes operating between the melting zone and the surface to produce silica-rich magmas. In order to elucidate the relationship between andesites and dacites, we focus on Tungurahua volcano, located in the Ecuadorian Andes. We collected a set of samples comprising such lithologies that were erupted during the last 3000 year BP. This relatively short period of time allows us to assume that the geodynamic parameters remain constant. Petrology and major-trace element compositions of these lavas have already been examined, and so we performed a complementary Pb-Sr isotope study in order to determine the nature and origin of the components involved in andesite and dacite genesis. Sr isotopes range from 0.70417 to 0.70431, and Pb isotope compositions range from 18.889 to 19.154 for Pb-206/Pb-204, from 15.658 to 15.696 for Pb-207/Pb-204, and from 38.752 to 38.918 for Pb-208/Pb-204. Dacites display a remarkably homogeneous Pb isotopic composition, with higher Pb-206/Pb-204 values for a given Pb207-208/Pb-204 compared to andesites. Andesites show notable Pb-207/Pb-206 variations for a given SiO2 content, whereas dacites have lower and homogenous Pb-207/Pb-206 values. Andesite and dacite altogether plot in a roughly triangular distribution, with dacitic magmas systematically plotting at the high SiO2 and Sr-87/Sr-86 and low Pb-207/Pb-206 fields. Based on our new dataset, we show that at least 3 different components are required to explain the Tungurahua compositional and isotope variation: one corresponds to the mantle, the second has a deep origin (slab component or lower crust), and a mixture between these two components explains andesite heterogeneity. The third component is derived from the underlying upper continental crust. While andesites are derived from deep components, dacites are derived from the andesitic magmas that underwent an assimilation-fractional crystallization (AFC) process with incorporation of the local metamorphic basement. Finally, we used the geochemical and isotopic data to produce a model of the magmatic plumbing system beneath Tungurahua, consistent with geophysical and experimental petrology constraints. We conclude that melt migration and storage in the upper crust appears to be a key parameter for controlling volcanic behavior though time