Nitro-sulfinate Reductive Coupling to Access (Hetero)aryl Sulfonamides.

A method to assemble (hetero)aryl sulfonamides via the reductive coupling of aryl sulfinates and nitroarenes is reported. Various reducing conditions with sodium bisulfite and with or without tin(II) chloride in DMSO were developed using an ultrasound bath to improve reaction homogeneity and mixing. A range of (hetero)aryl sulfonamides bearing a selection of functional groups were prepared, and the mechanism of the transformation was investigated. These investigations have led us to propose the formation of nitrosoarene intermediates, which were established via an independent molecular coupling strategy.

An inverse agonist of orphan receptor GPR61 acts by a G protein-competitive allosteric mechanism.

GPR61 is an orphan GPCR related to biogenic amine receptors. Its association with phenotypes relating to appetite makes it of interest as a druggable target to treat disorders of metabolism and body weight, such as obesity and cachexia. To date, the lack of structural information or a known biological ligand or tool compound has hindered comprehensive efforts to study GPR61 structure and function. Here, we report a structural characterization of GPR61, in both its active-like complex with heterotrimeric G protein and in its inactive state. Moreover, we report the discovery of a potent and selective small-molecule inverse agonist against GPR61 and structural elucidation of its allosteric binding site and mode of action. These findings offer mechanistic insights into an orphan GPCR while providing both a structural framework and tool compound to support further studies of GPR61 function and modulation.

MnII/III and CeIII/IV Units Supported on an Octahedral Molecular Nanoparticle of CeO2.

The preparation of three new heterometallic clusters [Ce6Mn12O17(O2CPh)26] (1), [Ce10Mn14O24(O2CPh)32] (2), and [Ce23Mn20O48(OH)2(tbb)46(H2O)4](NO3)2 (3; tbb- = 4-tBu-benzoate) is reported. They all possess unprecedented structures with a common feature being the presence of an octahedral CeIV-oxo core: a Ce6 in 1, two edge-fused Ce6 giving a Ce10 bioctahedron in 2, or a larger Ce19 octahedron in 3. Complex 1 is the first Ce6 cluster with a central µ6-O2-. 2 and the cation of 3 are molecular nanoparticles of CeO2 (ceria) because they possess the fluorite structure of bulk ceria and are thus ultrasmall ceria nanoparticles in molecular form. The {Ce19O32} octahedral subunit of the cation of 3 had been predicted from density functional theory studies to be one of the stable fragments of the CeO2 lattice, but has never been previously synthesized in molecular chemistry. Around the Ce/O core of 1-3 is an incomplete monolayer of Mnn ions disposed as four Mn3, two Mn7, and four Mn5 units, respectively. This represents a clear structural similarity with composite (phase-separated) CeO2/MnOx mixtures where at high Ce:Mn ratios the Mn atoms segregate on the surface of CeO2 phases. Variable-temperature dc and ac magnetic susceptibility studies have revealed S = 2, S = 1/2, and S = 3/2 ground states for 1-3, respectively. Fitting of the 5.0-300 K dc data for 1 to a two-J model for an asymmetrical V-shaped Mn3 unit with no interaction between the end MnIII ions gave an excellent fit with the following values: J1 = 5.2(3) cm-1, J2 = -7.4(3) cm-1, and g = 1.96(2).

Fluorophosphonates on-Demand: A General and Simplified Approach toward Fluorophosphonate Synthesis.

Herein, we report a general and simplified synthesis of fluorophosphonates directly from p-nitrophenylphosphonates. This FP on-demand reaction is mediated by a commercially available polymer-supported fluoride reagent that produces a variety (25 examples) of fluorophosphonates in high yields while only requiring reagent filtration for pure fluorophosphonate isolation. This reaction protocol facilitates the rapid profiling of serine hydrolases with diverse and novel sets of activated phosphonates with differential proteome reactivity. Moreover, slight modification of the procedure into a reaction-to-assay format has enabled additional screening efficiency.

Photoaffinity Labeling and Quantitative Chemical Proteomics Identify LXRß as the Functional Target of Enhancers of Astrocytic apoE.

Utilizing a phenotypic screen, we identified chemical matter that increased astrocytic apoE secretion in vitro. We designed a clickable photoaffinity probe based on a pyrrolidine lead compound and carried out probe-based quantitative chemical proteomics in human astrocytoma CCF-STTG1 cells to identify liver x receptor ß (LXRß) as the target. Binding of the small molecule ligand stabilized LXRß, as shown by cellular thermal shift assay (CETSA). In addition, we identified a probe-modified peptide by mass spectrometry and proposed a model where the photoaffinity probe is bound in the ligand-binding pocket of LXRß. Taken together, our findings demonstrated that the lead chemical matter bound directly to LXRß, and our results highlight the power of chemical proteomic approaches to identify the target of a phenotypic screening hit. Additionally, the LXR photoaffinity probe and lead compound described herein may serve as valuable tools to further evaluate the LXR pathway.

Phosphocyclocreatine is the dominant form of cyclocreatine in control and creatine transporter deficiency patient fibroblasts.

Creatine transporter deficiency (CTD) is a metabolic disorder resulting in cognitive, motor, and behavioral deficits. Cyclocreatine (cCr), a creatine analog, has been explored as a therapeutic strategy for the treatment of CTD. We developed a rapid, selective, and accurate HILIC ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method to simultaneously quantify the intracellular concentrations of cCr, creatine (Cr), creatine-d3 (Cr-d3), phosphocyclocreatine (pcCr), and phosphocreatine (pCr). Using HILIC-UPLC-MS/MS, we measured cCr and Cr-d3 uptake and their conversion to the phosphorylated forms in primary human control and CTD fibroblasts. Altogether, the data demonstrate that cCr enters cells and its dominant intracellular form is pcCr in both control and CTD patient cells. Therefore, cCr may replace creatine as a therapeutic strategy for the treatment of CTD.

2,2,2-Trifluoroethoxy Aromatic Heterocycles: Hydrolytically Stable Alternatives to Heteroaryl Chlorides.

Herein we describe the 2,2,2-trifluoroethoxy group as an alternative leaving group for hydrolytically unstable heteroaryl chlorides. This group provides improved shelf stability by years while maintaining reactivity toward nucleophiles in SNAr reactions. A highlighted trifluoroethyl ether was shown to be tolerant to aqueous Suzuki conditions, permitting sequential Suzuki/SNAr processes inaccessible to the heterocyclic chlorides. The strategic use of trifluoroethyl ethers enables storage of otherwise unstable heterocyclic chlorides and limits costly decomposition.

A Benzyne Insertion Approach to Hetisine-Type Diterpenoid Alkaloids: Synthesis of Cossonidine (Davisine).

The hetisine-type natural products exhibit one of the most complex carbon skeletons within the diterpenoid alkaloid family. The use of network analysis has enabled a synthesis strategy to access alkaloids in this class with hydroxylation on the A-ring. Key transformations include a benzyne acyl-alkylation to construct a key fused 6-7-6 tricycle, a chemoselective nitrile reduction, and sequential C-N bond formations using a reductive cyclization and a photochemical hydroamination to construct an embedded azabicycle. Our strategy should enable access to myriad natural and unnatural products within the hetisine-type.

Computer-Aided Discovery and Characterization of Novel Ebola Virus Inhibitors.

The Ebola virus (EBOV) causes severe human infection that lacks effective treatment. A recent screen identified a series of compounds that block EBOV-like particle entry into human cells. Using data from this screen, quantitative structure-activity relationship models were built and employed for virtual screening of a ∼17 million compound library. Experimental testing of 102 hits yielded 14 compounds with IC50 values under 10 µM, including several sub-micromolar inhibitors, and more than 10-fold selectivity against host cytotoxicity. These confirmed hits include FDA-approved drugs and clinical candidates with non-antiviral indications, as well as compounds with novel scaffolds and no previously known bioactivity. Five selected hits inhibited BSL-4 live-EBOV infection in a dose-dependent manner, including vindesine (0.34 µM). Additional studies of these novel anti-EBOV compounds revealed their mechanisms of action, including the inhibition of NPC1 protein, cathepsin B/L, and lysosomal function. Compounds identified in this study are among the most potent and well-characterized anti-EBOV inhibitors reported to date.

Late-Stage Microsomal Oxidation Reduces Drug-Drug Interaction and Identifies Phosphodiesterase 2A Inhibitor PF-06815189.

Late-stage oxidation using liver microsomes was applied to phosphodiesterase 2 inhibitor 1 to reduce its clearance by cytochrome P450 enzymes, introduce renal clearance, and minimize the risk for victim drug-drug interactions. This approach yielded PF-06815189 (2) with improved physicochemical properties and a mixed metabolic profile. This example highlights the importance of C-H diversification methods to drug discovery.

Identification of a Potent, Highly Selective, and Brain Penetrant Phosphodiesterase 2A Inhibitor Clinical Candidate.

Computational modeling was used to direct the synthesis of analogs of previously reported phosphodiesterase 2A (PDE2A) inhibitor 1 with an imidazotriazine core to yield compounds of significantly enhanced potency. The analog PF-05180999 (30) was subsequently identified as a preclinical candidate targeting cognitive impairment associated with schizophrenia. Compound 30 demonstrated potent binding to PDE2A in brain tissue, dose responsive mouse brain cGMP increases, and reversal of N-methyl-d-aspartate (NMDA) antagonist-induced (MK-801, ketamine) effects in electrophysiology and working memory models in rats. Preclinical pharmacokinetics revealed unbound brain/unbound plasma levels approaching unity and good oral bioavailability resulting in an average concentration at steady state (Cav,ss) predicted human dose of 30 mg once daily (q.d.). Modeling of a modified release formulation suggested that 25 mg twice daily (b.i.d.) could maintain plasma levels of 30 at or above targeted efficacious plasma levels for 24 h, which became part of the human clinical plan.

Mechanism of a No-Metal-Added Heterocycloisomerization of Alkynylcyclopropylhydrazones: Synthesis of Cycloheptane-Fused Aminopyrroles Facilitated by Copper Salts at Trace Loadings.

A mechanistic study of a new heterocycloisomerization reaction that forms annulated aminopyrroles is presented. Density functional theory calculations and kinetic studies suggest the reaction is catalyzed by trace copper salts and that a Z- to E-hydrazone isomerization occurs through an enehydrazine intermediate before the rate-determining cyclization of the hydrazone onto the alkyne group. The aminopyrrole products are obtained in 36-93% isolated yield depending on the nature of the alkynyl substituent. A new automated sampling technique was developed to obtain robust mechanistic data.

Novel lead structures with both Plasmodium falciparum gametocytocidal and asexual blood stage activity identified from high throughput compound screening.

BACKGROUND: Blocking malaria transmission is an important step in eradicating malaria. In the field, transmission requires the production of sexual stage Plasmodium parasites, called gametocytes, which are not effectively killed by the commonly used anti-malarials allowing individuals to remain infectious after clearance of asexual parasites. METHODS: To identify new gametocytocidal compounds, a library of 45,056 compounds with diverse structures was screened using a high throughput gametocyte viability assay. The characteristics of active hits were further evaluated against asexual stage parasites in a growth inhibition assay. Their cytotoxicity were tested against mammalian cells in a cytotoxicity assay. The chemical scaffold similarity of active hits were studied using scaffold cluster analysis. RESULTS: A set of 23 compounds were identified and further confirmed for their activity against gametocytes. All the 23 confirmed compounds possess dual-activities against both gametocytes responsible for human to mosquito transmission and asexual parasites that cause the clinical symptoms. Three of these compounds were fourfold more active against gametocytes than asexual parasites. Further cheminformatic analysis revealed three sets of novel scaffolds, including highly selective 4-1H-pyrazol-5-yl piperidine analogs. CONCLUSIONS: This study revealed important new structural scaffolds that can be used as starting points for dual activity anti-malarial drug development.

Synergistic drug combination effectively blocks Ebola virus infection.

Although a group of FDA-approved drugs were previously identified with activity against Ebola virus (EBOV), most of them are not clinically useful because their human blood concentrations are not high enough to inhibit EBOV infection. We screened 795 unique three-drug combinations in an EBOV entry assay. Two sets of three-drug combinations, toremifene-mefloquine-posaconazole and toremifene-clarithromycin-posaconazole, were identified that effectively blocked EBOV entry and were further validated for inhibition of live EBOV infection. The individual drug concentrations in the combinations were reduced to clinically relevant levels. We identified mechanisms of action of these drugs: functional inhibitions of Niemann-Pick C1, acid sphingomyelinase, and lysosomal calcium release. Our findings identify the drug combinations with potential to treat EBOV infection.

The Schistosoma mansoni Cytochrome P450 (CYP3050A1) Is Essential for Worm Survival and Egg Development.

Schistosomiasis affects millions of people in developing countries and is responsible for more than 200,000 deaths annually. Because of toxicity and limited spectrum of activity of alternatives, there is effectively only one drug, praziquantel, available for its treatment. Recent data suggest that drug resistance could soon be a problem. There is therefore the need to identify new drug targets and develop drugs for the treatment of schistosomiasis. Analysis of the Schistosoma mansoni genome sequence for proteins involved in detoxification processes found that it encodes a single cytochrome P450 (CYP450) gene. Here we report that the 1452 bp open reading frame has a characteristic heme-binding region in its catalytic domain with a conserved heme ligating cysteine, a hydrophobic leader sequence present as the membrane interacting region, and overall structural conservation. The highest sequence identity to human CYP450s is 22%. Double stranded RNA (dsRNA) silencing of S. mansoni (Sm)CYP450 in schistosomula results in worm death. Treating larval or adult worms with antifungal azole CYP450 inhibitors results in worm death at low micromolar concentrations. In addition, combinations of SmCYP450-specific dsRNA and miconazole show additive schistosomicidal effects supporting the hypothesis that SmCYP450 is the target of miconazole. Treatment of developing S. mansoni eggs with miconazole results in a dose dependent arrest in embryonic development. Our results indicate that SmCYP450 is essential for worm survival and egg development and validates it as a novel drug target. Preliminary structure-activity relationship suggests that the 1-(2,4-dichlorophenyl)-2-(1H-imidazol-1-yl)ethan-1-ol moiety of miconazole is necessary for activity and that miconazole activity and selectivity could be improved by rational drug design.

Tungsten-catalyzed heterocycloisomerization approach to 4,5-dihydro-benzo[b]furans and -indoles.

A W(CO)(5)·THF-catalyzed cycloisomerization of bicyclo[4.1.0] substrates to afford mono C4-substituted 4,5-dihydro-benzo[b]furans and -indoles is reported. The title compounds are versatile intermediates that lead to a range of fused bicycles including the cores of various furan-, benzofuran-, and indole-containing natural products. In many cases, the functionalization of the dihydro-benzo[b]furans and -indoles is orthogonal to that of the corresponding benzofurans and indoles and, thus, offers complementary approaches for synthesis.

Leaving group potential of a substituted cyclopentadienyl anion toward oxidative addition.

The facility with which a substituted cyclopentadienyl anion may function as a leaving group for palladium-catalyzed allylation reactions is demonstrated. Reaction of several allylcyclopentadienyl substrates is shown. Nucleophilic displacement of carbon with nitrogen is achieved in the deallylation of allylpenta-p-acetylphenylcyclopentadiene with N-methylbenzylamine.

Catalytic asymmetric generation of (Z)-disubstituted allylic alcohols.

A one-pot method for the direct preparation of enantioenriched (Z)-disubstituted allylic alcohols is introduced. Hydroboration of 1-halo-1-alkynes with dicyclohexylborane, reaction with t-BuLi, and transmetalation with dialkylzinc reagents generate (Z)-disubstituted vinylzinc intermediates. In situ reaction of these reagents with aldehydes in the presence of a catalyst derived from (-)-MIB generates (Z)-disubstituted allylic alcohols. It was found that the resulting allylic alcohols were racemic, most likely due to a rapid addition reaction promoted by LiX (X = Br and Cl). To suppress the LiX-promoted reaction, a series of inhibitors were screened. It was found that 20-30 mol % tetraethylethylenediamine inhibited LiCl without inhibiting the chiral zinc-based Lewis acid. In this fashion, (Z)-disubstituted allylic alcohols were obtained with up to 98% ee. The asymmetric (Z)-vinylation could be coupled with tandem diastereoselective epoxidation reactions to provide epoxy alcohols and allylic epoxy alcohols with up to three contiguous stereogenic centers, enabling the rapid construction of complex building blocks with high levels of enantio- and diastereoselectivity.

Direct, stereospecific generation of (Z)-disubstituted allylic alcohols.

A one-pot method to prepare highly functionalized (Z)-disubstituted allylic alcohols is introduced. Hydroboration of a variety of 1-bromo-1-acetylenes with dicyclohexyl borane, reaction with t-BuLi, and transmetalation to zinc generates a (Z)-disubstituted vinylzinc reagent. In situ reaction of this reagent with aldehydes generates (Z)-disubstituted allylic alcohols in high yields (81-97%). Addition to chiral protected alpha- or beta-oxygenated aldehydes proceeds with diastereoselectivities between 6:1 and 18:1. The anti-Felkin product is obtained in most cases.