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Vaccination is the most prevalent prophylactic means for controlling seasonal influenza infections. However, an effective vaccine usually takes at least 6 months to develop for the circulating strains. Therefore, new therapeutic options are needed for the acute treatment of influenza infections to control this virus and prevent epidemics/pandemics from developing. We have discovered fast-acting, orally bioavailable acylated 4-aminopiperidines with an effective mechanism of action targeting viral hemagglutinin (HA). Our data show that these compounds are potent entry inhibitors of influenza A viruses. We present docking studies that suggest an HA binding site for these inhibitors on H5N1. Compound 16 displayed a significant decrease of viral titer when evaluated in the infectious assays with influenza virus H1N1 (A/Puerto Rico/8/1934) or H5N1 (A/Vietnam/1203/2004) strains and the oseltamivir-resistant strain with the most common H274Y mutation. In addition, compound 16 showed significant synergistic activity with oseltamivir in vitro.

Glioblastoma multiforme (GBM) is the most common and aggressive malignant primary brain tumor in humans. Over the past several decades, despite improvements in neurosurgical techniques, development of powerful chemotherapeutic agents, advances in radiotherapy, and comprehensive genomic profiling and molecular characterization, treatment of GBM has achieved very limited success in increasing overall survival. Thus, identifying and understanding the key molecules and barriers responsible for the malignant phenotypes and treatment resistance of GBM will yield new potential therapeutic targets. We review the most recent development of receptor tyrosine kinase targeted therapy for GBM and discuss the current status of several novel strategies with the emphasis on blood–brain barrier penetration as a major obstacle for small-molecule drugs to achieve their therapeutic goals. Likewise, a major opportunity for the treatment of GBM lies in the use of biomarkers for the discovery and development of new receptor tyrosine kinase targeted therapy.

Many small molecules have been identified as entry inhibitors of filoviruses. However, a lack of understanding of the mechanism of action for these molecules limits further their development as anti-filoviral agents. Here we provide evidence that toremifene and other small molecule entry inhibitors have at least three distinctive mechanisms of action and lay the groundwork for future development of anti-filoviral agents. The three mechanisms identified here include: (1) direct binding to the internal fusion loop region of Ebola virus glycoprotein (GP); (2) the HR2 domain is likely the main binding site for Marburg virus GP inhibitors and a secondary binding site for some EBOV GP inhibitors; (3) lysosome trapping of GP inhibitors increases drug exposure in the lysosome and further improves the viral inhibition. Importantly, small molecules targeting different domains on GP are synergistic in inhibiting EBOV entry suggesting these two mechanisms of action are distinct. Our findings provide important mechanistic insights into filovirus entry and rational drug design for future antiviral development., Author summary Filoviruses are among the deadliest pathogens known to mankind with case-fatality rates ranging from 25–90%. New outbreaks in central Africa and the identification of novel filoviruses in other regions highlight the urgent need to develop novel therapeutics. Although many novel anti-filovirus compounds have been reported as entry inhibitors, to date, none have made to market. This high rate of failure is in part due to a lack of knowledge of the mechanisms of action. In this report, we provide a molecular basis for the multiple mechanisms of action by which small molecule inhibitors of Ebola virus and Marburg virus block virus entry, which provides new mechanistic insight to guide design for next-generation viral entry inhibitors.

The recent Ebola epidemics in West Africa underscore the great need for effective and practical therapies for future Ebola virus outbreaks. We have discovered a new series of remarkably potent small molecule inhibitors of Ebola virus entry. These 4-(aminomethyl)benzamide-based inhibitors are also effective against Marburg virus. Synthetic routes to these compounds allowed for the preparation of a wide variety of structures, including a conformationally restrained subset of indolines (compounds 41–50). Compounds 20, 23, 32, 33, and 35 are superior inhibitors of Ebola (Mayinga) and Marburg (Angola) infectious viruses. Representative compounds (20, 32 and 35) have shown good metabolic stability in plasma and liver microsomes (rat and human), and 32 did not inhibit CYP3A4 nor CYP2C9. These 4-(aminomethyl)benzamides are suitable for further optimization as inhibitors of filovirus entry, with potential to be developed as therapeutic agents for the treatment and control of Ebola virus infections.

Background: Influenza viruses cause severe upper respiratory illness in children and the elderly during seasonal epidemics. Influenza viruses from zoonotic reservoirs can also cause pandemics with significant loss of life in all age groups. Although vaccination is one of the most effective methods to protect against seasonal epidemics, seasonal vaccines vary in efficacy, can be ineffective in the elderly population, and do not provide protection against novel strains. Small molecule therapeutics are a critical part of our antiviral strategies to control influenza virus epidemics and pandemics as well as to ameliorate disease in elderly and immunocompromised individuals. Objective: This review aims to summarize the existing antiviral strategies for combating influenza viruses, the mechanisms of antiviral resistance for available drugs, and novel therapeutics currently in development. Methods: We systematically evaluated and synthesized the published scientific literature for mechanistic detail into therapeutic strategies against influenza viruses. Results: Current IAV strains have developed resistance to neuraminidase inhibitors and nearly complete resistance to M2 ion channel inhibitors, exacerbated by sub-therapeutic dosing used for treatment and chemoprophylaxis. New tactics include novel therapeutics targeting host components and combination therapy, which show potential for fighting influenza virus disease while minimizing viral resistance. Conclusion: Antiviral drugs are crucial for controlling influenza virus disease burden, but their efficacy is limited by human misuse and the capacity of influenza viruses to circumvent antiviral barriers. To relieve the public health hardship of influenza virus, emerging therapies must be selected for their capacity to impede not only influenza virus disease, but also the development of antiviral resistance.

Influenza A viruses (IAVs) cause seasonal flu and occasionally pandemics. The current therapeutics against IAVs target two viral proteins - neuraminidase (NA) and M2 ion-channel protein. However, M2 ion channel inhibitors (amantadine and rimantadine) are no longer recommended by CDC for use due to the emergence of high level of antiviral resistance among the circulating influenza viruses, and resistant strains to NA inhibitors (oseltamivir and zanamivir) have also been reported. Therefore, development of novel anti-influenza therapies is urgently needed. As one of the viral surface glycoproteins, hemagglutinin (HA) mediates critical virus entry steps including virus binding to host cells and virus-host membrane fusion, which makes it a potential target for anti-influenza drug development. In this study, we report the identification of compound CBS1116 with a 4-aminopiperidine scaffold from a chemical library screen as an entry inhibitor specifically targeting two group 1 influenza A viruses, A/Puerto Rico/8/34 (H1N1) and recombinant low pathogenic avian H5N1 virus (A/Vietnam/1203/04, VN04Low). Mechanism of action studies show that CBS1116 interferes with the HA-mediated fusion process. Further structure activity relationship study generated a more potent compound CBS1117 which has a 50% inhibitory concentration of 70 nM and a selectivity index of ~4000 against A/Puerto Rico/8/34 (H1N1) infection in human lung epithelial cell line (A549).

Filoviruses, consisting of Ebola virus, Marburg virus and Cuevavirus, cause severe hemorrhagic fevers in humans with high mortality rates up to 90%. Currently, there is no approved vaccine or therapy available for the prevention and treatment of filovirus infection in humans. The recent 2013-2015 West African Ebola epidemic underscores the urgency to develop antiviral therapeutics against these infectious diseases. Our previous study showed that GPCR antagonists, particularly histamine receptor antagonists (antihistamines) inhibit Ebola and Marburg virus entry. In this study, we screened a library of 1220 small molecules with predicted antihistamine activity, identified multiple compounds with potent inhibitory activity against entry of both Ebola and Marburg viruses in human cancer cell lines, and confirmed their anti-Ebola activity in human primary cells. These small molecules target a late-stage of Ebola virus entry. Further structure-activity relationship studies around one compound (cp19) reveal the importance of the coumarin fused ring structure, especially the hydrophobic substituents at positions 3 and/or 4, for its antiviral activity, and this identified scaffold represents a favorable starting point for the rapid development of anti-filovirus therapeutic agents.

Influenza A virus (IAV) causes seasonal epidemics and occasional but devastating pandemics, which are major public health concerns. The putative antiviral therapeutics are useful for the treatment of influenza, however, the emerging resistant strains necessitate a constant search for new drug candidates. Here we report the discovery of a novel antiviral agent, compound CBS1194, which was identified by a parallel high-throughput screening (HTS) campaign using two retroviral pseudotypes bearing H7 or H5 hemagglutinins (HAs). Subsequent analyses demonstrated that CBS1194 is specific to IAVs of group 2, while it has no effect against those of group 1. In a time-of-addition assay, CBS1194 showed a significant inhibitory effect during the early phase of viral infection. In addition, HA-mediated hemolysis can be inhibited by CBS1194 treatment, indicating that this compound may target the HA stalk region, which is responsible for membrane fusion. Escape mutant analyses and in silico docking further revealed that CBS1194 fits into a pocket near the fusion peptide, causing steric hindrance that blocks the low-pH induced rearrangement of HA. In summary, our study identifies a novel fusion inhibitor of group 2 IAVs, which has the potential as lead compound for further development.

Diels–Alder reactions of five-membered heterocycles containing one heteroatom with an N-arylmaleimide were studied. Cycloaddition of 2,5-dimethylfuran (4) with 2-(4-methylphenyl)maleimide (3) in toluene at 60 °C gave bicyclic adduct 5. Cycloadditions of 3 with 2,5-dimethylthiophene (11) and 1,2,5-trimethylpyrrole (14) were also studied. Interestingly, the bicyclic compound 5 cleanly rearranged, with loss of water, when treated with p-toluenesulfonic acid in toluene at 80 °C to give 4,7-dimethyl-2-p-tolylisoindoline-1,3-dione (6).

Herein we describe the synthesis and antibacterial evaluation of a new, unsymmetrical triaryl bisamidine compound series, [Am]-[indole]-[linker]-[HetAr/Ar]-[Am], in which [Am] is an amidine or amino group, [linker] is a benzene, thiophene or pyridine ring, and [HetAr/Ar] is a benzimidazole, imidazopyridine, benzofuran, benzothiophene, pyrimidine or benzene ring. When the [HetAr/Ar] unit is a 5,6-bicyclic heterocycle, it is oriented such that the 5-membered ring portion is connected to the [linker] unit and the 6-membered ring portion is connected to the [Am] unit. Among the 34 compounds in this series, compounds with benzofuran as the [HetAr/Ar] unit showed the highest potencies. Introduction of a fluorine atom or a methyl group to the triaryl core led to the more potent analogs. Bisamidines are more active toward bacteria while the monoamidines are more active toward mammalian cells (as indicated by low CC50 values). Importantly, we identified compound P12a (MBX 1887) with a relatively narrow spectrum against bacteria and a very high CC50 value. Compound P12a has been scaled up and is currently undergoing further evaluations for therapeutic applications.

We have previously attempted to prepare 5-(4-morpholinyl)-2-aminothiophene ( 3 ) by reducing 5-(4-morpholinyl)-2-nitrothiophene ( 2 ), to prepare a precursor for the preparation of thienyloxazolidinones as potential linezolid-like antibiotics. This strategy failed, but allowed us to define an interesting reductive aromatization reaction. We have now succeeded in producing thienyloxazolidinones using an alternative strategy, initiating from 5-(4-morpholinyl)thiophene-2-carboxaldehyde ( 8 ). This Letter describes the synthesis of thienyloxazolidinone 14 , which is a key intermediate for the preparation of new oxazolidinone antibiotics with a thiophene core.

This manuscript describes the preparation of new small molecule inhibitors of Bacillus anthracis lethal factor. Our starting point was the symmetrical, bis-quinolinyl compound 1 (NSC 12155). Optimization of one half of this molecule led to new LF inhibitors that were desymmetrized to afford more drug-like compounds.

Methylenecyclopropane nucleoside (MCPN) analogs are being investigated for treatment of human cytomegalovirus (HCMV) infection because of favorable preclinical data and limited ganciclovir cross-resistance. Monohydroxymethyl MCPNs bearing ether and thioether functionalities at the purine 6 position have antiviral activity against herpes simplex virus (HSV) and varicella-zoster virus (VZV) in addition to HCMV. The role of the HCMV UL97 kinase in the mechanism of action of these derivatives was examined. When tested against a kinase-inactive UL97 K355M virus, a moderate 5- to 7-fold increase in 50% effective concentration (EC 50 ) was observed, in comparison to a 13- to 25-fold increase for either cyclopropavir or ganciclovir. Serial propagation of HCMV under two of these compounds selected for three novel UL97 mutations encoding amino acid substitutions D456N, C480R,and Y617del. When transferred to baseline laboratory HCMV strains, these mutations individually conferred resistance to all of the tested MCPNs, ganciclovir, and maribavir. However, the engineered strains also demonstrated severe growth defects and abnormal cytopathic effects similar to the kinase-inactive mutant. Expressed and purified UL97 kinase showed in vitro phosphorylation of the newly tested MCPNs. Thus, HCMV UL97 kinase is involved in the antiviral action of these MCPNs, but the in vitro selection of UL97-defective viruses suggests that their activity against more typical ganciclovir-resistant growth-competent UL97 mutants may be relatively preserved.

Influenza viruses are a major public health threat worldwide, and options for antiviral therapy are limited by the emergence of drug-resistant virus strains. The influenza virus glycoprotein hemagglutinin (HA) plays critical roles in the early stage of virus infection, including receptor binding and membrane fusion, making it a potential target for the development of anti-influenza drugs. Using pseudotype virus-based high-throughput screens, we have identified several new small molecules capable of inhibiting influenza virus entry. We prioritized two novel inhibitors, MBX2329 and MBX2546, with aminoalkyl phenol ether and sulfonamide scaffolds, respectively, that specifically inhibit HA-mediated viral entry. The two compounds (i) are potent (50% inhibitory concentration [IC 50 ] of 0.3 to 5.9 μM); (ii) are selective (50% cytotoxicity concentration [CC 50 ] of >100 μM), with selectivity index (SI) values of >20 to 200 for different influenza virus strains; (iii) inhibit a wide spectrum of influenza A viruses, which includes the 2009 pandemic influenza virus A/H1N1/2009, highly pathogenic avian influenza (HPAI) virus A/H5N1, and oseltamivir-resistant A/H1N1 strains; (iv) exhibit large volumes of synergy with oseltamivir (36 and 331 μM 2 % at 95% confidence); and (v) have chemically tractable structures. Mechanism-of-action studies suggest that both MBX2329 and MBX2546 bind to HA in a nonoverlapping manner. Additional results from HA-mediated hemolysis of chicken red blood cells (cRBCs), competition assays with monoclonal antibody (MAb) C179, and mutational analysis suggest that the compounds bind in the stem region of the HA trimer and inhibit HA-mediated fusion. Therefore, MBX2329 and MBX2546 represent new starting points for chemical optimization and have the potential to provide valuable future therapeutic options and research tools to study the HA-mediated entry process.

Benzobisthiazole derivatives were identified as novel helicase inhibitors through high throughput screening against purified Staphylococcus aureus (Sa) and Bacillus anthracis (Ba) replicative helicases. Chemical optimization has produced compound 59 with nanomolar potency against the DNA duplex strand unwinding activities of both B. anthracis and S. aureus helicases. Selectivity index (SI=CC50/IC50) values for 59 were greater than 500. Kinetic studies demonstrated that the benzobisthiazole-based bacterial helicase inhibitors act competitively with the DNA substrate. Therefore, benzobisthiazole helicase inhibitors represent a promising new scaffold for evaluation as antibacterial agents.

Reduction of a 2-nitrothiophene 4 and the corresponding 2-nitrofuran 20 with zinc and acetic acid produced a remarkable dearomatization and fragmentation reaction to give acyclic nitriles 6 and 22, respectively. The intermediate 2-aminothiophene 5 was trapped by acylation with acetic anhydride to give acetamide 7. An additional acrylonitrile intermediate 17 was also trapped by Michael­ addition with benzyl mercaptan to give adduct 18. An alternate synthesis of nitrile 22 produced in the reduction of nitrofuran 20 provided an authentic sample of the product. The conversion of nitroaromatic heterocycles 4 and 20 into aliphatic nitriles 6 and 22 are remarkable and unprecedented reactions.

Pyridine is frequently found as the core structure or a component in a variety of drugs, for example, ethionade,[ 1 ] etoricoxib,[ 2 ] dimetindene,[ 3 ] brompheniramine,[ 4 ] isoniazid,[ 5 ] and me...

6-Cyanobenzo[b]furan-2-boronic acid pinacol ester (10) is a potentially useful two-point scaffold for the construction of specific compounds or compound libraries with benzofuran cores. Using a per-iodination/de-iodination strategy coupled with Sonogashira alkynylation and Cu-catalyzed heteroannulation, we have developed a procedure that allows the preparation of benzo[b]furan-6-carbonitrile (9) and 6-cyanobenzo[b]furan-2-boronic acid pinacol ester (10) in gram quantities. Supplemental materials are available for this article. Go to the publisher's online edition of Synthetic Communications® to view the free supplemental file.

Ebola virus has caused 26 outbreaks in 10 different countries since its identification in 1976, making it one of the deadliest emerging viral pathogens. The most recent outbreak in West Africa from 2014-16 was the deadliest yet and culminated in 11,310 deaths out of 28,616 confirmed cases. Currently, there are no FDA-approved therapeutics or vaccines to treat Ebola virus infections. The slow development of effective vaccines combined with the severity of past outbreaks emphasizes the need to accelerate research into understanding the virus lifecycle and the development of therapeutics for post exposure treatment. Here we present a summary of the major findings on the Ebola virus replication cycle and the therapeutic approaches explored to treat this devastating disease. The major focus of this review is on small molecule inhibitors.

We previously reported the synthesis and biological activity of a series of cationic bis-indoles with potent, broad-spectrum antibacterial properties. Here, we describe mechanism of action studies to test the hypothesis that these compounds bind to DNA and that this target plays an important role in their antibacterial outcome. The results reported here indicate that the bis-indoles bind selectively to DNA at A/T-rich sites, which is correlated with the inhibition of DNA and RNA synthesis in representative Gram-positive ( Staphylococcus aureus ) and Gram-negative ( Escherichia coli ) organisms. Further, exposure of E. coli and S. aureus to representative bis-indoles resulted in induction of the DNA damage-inducible SOS response. In addition, the bis-indoles were found to be potent inhibitors of cell wall biosynthesis; however, they do not induce the cell wall stress stimulon in S. aureus , suggesting that this pathway is inhibited by an indirect mechanism. In light of these findings, the most likely basis for the observed activities of these compounds is their ability to bind to the minor groove of DNA, resulting in the inhibition of DNA and RNA synthesis and other secondary effects.

The plasmodial surface anion channel (PSAC) increases erythrocyte permeability to many solutes in malaria but has uncertain physiological significance. We used a PSAC inhibitor with different efficacies against channels from two Plasmodium falciparum parasite lines and found concordant effects on transport and in vitro parasite growth when external nutrient concentrations were reduced. Linkage analysis using this growth inhibition phenotype in the Dd2 × HB3 genetic cross mapped the clag3 genomic locus, consistent with a role for two clag3 genes in PSAC-mediated transport. Altered inhibitor efficacy, achieved through allelic exchange or expression switching between the clag3 genes, indicated that the inhibitor kills parasites through direct action on PSAC. In a parasite unable to undergo expression switching, the inhibitor selected for ectopic homologous recombination between the clag3 genes to increase the diversity of available channel isoforms. Broad-spectrum inhibitors, which presumably interact with conserved sites on the channel, also exhibited improved efficacy with nutrient restriction. These findings indicate that PSAC functions in nutrient acquisition for intracellular parasites. Although key questions regarding the channel and its biological role remain, antimalarial drug development targeting PSAC should be pursued.

The Cadogan–Sundberg indole synthesis produces two major products from the treatment of 2-nitrostilbenes with triethyl phosphite, which are the 2-arylindoles and the corresponding 2-aryl-N-ethoxyindoles. We were interested in determining the origin of the oxygen atom in the 2-aryl-N-ethoxyindoles. In this study, we verify that this oxygen atom originates from the nitro group and not from the triethyl phosphite.

Botulinum neurotoxins (BoNTs) are the most lethal of biological substances, and are categorized as class A biothreat agents by the Centers for Disease Control and Prevention. There are currently no drugs to treat the deadly flaccid paralysis resulting from BoNT intoxication. Among the seven BoNT serotypes, the development of therapeutics to counter BoNT/A is a priority (due to its long half-life in the neuronal cytosol and its ease of production). In this regard, the BoNT/A enzyme light chain (LC) component, a zinc metalloprotease responsible for the intracellular cleavage of synaptosomal-associated protein of 25 kDa, is a desirable target for developing post-BoNT/A intoxication rescue therapeutics. In an earlier study, we reported the high throughput screening of a library containing 70,000 compounds, and uncovered a novel class of benzimidazole acrylonitrile-based BoNT/A LC inhibitors. Herein, we present both structure-activity relationships and a proposed mechanism of action for this novel inhibitor chemotype.

Botulinum neurotoxins (BoNTs) are the most potent of known toxins and are listed as category A biothreat agents by the U.S. CDC. The BoNT-mediated proteolysis of SNARE proteins inhibits the exocytosis of acetylcholine into neuromuscular junctions, leading to life-threatening flaccid paralysis. Currently, the only therapy for BoNT intoxication (which results in the disease state botulism) includes experimental preventative antibodies and long-term supportive care. Therefore, there is an urgent need to identify and develop inhibitors that will serve as both prophylactic agents and post-exposure ‘rescue’ therapeutics. This review focuses on recent progress to discover and develop small molecule inhibitors as therapeutic countermeasures for BoNT intoxication.

Unexpected difficulty in the conversion of a bromobenzofuran to the corresponding formylbenzofuran led us to develop a new synthesis for 5-formylbenzo[b]furan-2-carbonitrile (1).

Treatment of racemic 2-hydroxy-3-(1H-indol-3yl)propionic acid methyl ester (5) with isopropyl magnesium chloride provided the title compound 1 and its isomer, 3-hydroxy-1-(indol-3-yl)-4-methylpentan-2-one (9). Both enantiomers (>96% ee) of each component were obtained via semi-preparative chiral supercritical fluid chromatography (SFC). In contrast to previous reports, these compounds, as well as their acetate derivatives, were not active or very weakly active against 16 bacterial strains, including Escherichia coli, Bacillus subtilis and Staphylococcus aureus.

Antimicrobial susceptibilities of 233 Gram-positive and 180 Gram-negative strains to two novel bis-indoles were evaluated. Both compounds were potent inhibitors of Gram-positive bacteria, with MIC 90 values of 0.004 to 0.5 μg/ml. One bis-indole, MBX 1162, exhibited potent activity against all Gram-negative strains, with MIC 90 values of 0.12 to 4 μg/ml, even against high-level-resistant pathogens, and compared favorably to all comparator antibiotics. The bis-indole compounds show promise for the treatment of multidrug-resistant clinical pathogens.

The type III secretion system (T3SS) is a clinically important virulence mechanism in Pseudomonas aeruginosa that secretes and translocates up to four protein toxin effectors into human cells, facilitating the establishment and dissemination of infections. To discover inhibitors of this important virulence mechanism, we developed two cellular reporter assays and applied them to a library of 80,000 compounds. The primary screen was based on the dependence of the transcription of T3SS operons on the T3SS-mediated secretion of a negative regulator and consisted of a transcriptional fusion of the Photorhabdus luminescens luxCDABE operon to the P. aeruginosa exoT effector gene. Secondary assays included direct measurements of the T3SS-mediated secretion of a P. aeruginosa ExoS effector-β-lactamase fusion protein as well as the detection of the secretion of native ExoS by the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of culture supernatants. Five inhibitors in three chemical classes were demonstrated to inhibit type III secretion selectively with minimal cytotoxicity and with no effects on bacterial growth or on the type II-mediated secretion of elastase. These inhibitors also block the T3SS-mediated secretion of a YopE effector-β-lactamase fusion protein from an attenuated Yersinia pestis strain. The most promising of the inhibitors is a phenoxyacetamide that also blocks the T3SS-mediated translocation of effectors into mammalian cells in culture. Preliminary studies of structure-activity relationships in this phenoxyacetamide series demonstrated a strict requirement for the R -enantiomer at its stereocenter and indicated tolerance for a variety of substituents on one of its two aromatic rings.

The bis-indoles are a novel class of compounds with potent antibacterial activity against a broad spectrum of Gram-positive and Gram-negative pathogens. The mechanism of action of these compounds has not been clearly defined. To study the mechanism of action of bis-indoles, selections for mutants of Staphylococcus aureus NCTC 8325 with reduced susceptibility to several chemically related bis-indoles were carried out using serial passages in subinhibitory compound concentrations. Resistant mutants were only obtained for one of the four bis-indoles tested (MBX-1090), and these appeared at concentrations up to 16X MIC within 10–12 passages. MBX-1090 resistance mutations produced a truncated open reading frame of mepR (SAOUHSC_00314), a gene encoding a MarR-like repressor. MepR regulates expression of mepA (SAOUHSC_00315), which encodes a member of the Multidrug and Toxic Compound Extrusion (MATE) family of efflux pumps. MBX-1090 resistance was reverted when mepR (wild type) was provided in trans. Microarray experiments and RT-PCR experiments confirmed that over-expression of mepA is required for resistance. Interestingly, MBX-1090 resistant mutants and strains overexpressing mepA from an expression vector did not exhibit cross-resistance to closely related bis-indole compounds. MBX-1090 did not induce expression of mepA, suggesting that this compound does not directly interact with MepR. Conversely, the bis-indoles that were not substrates of MepA strongly induced mepA expression. The results of this study suggest that MepA and MepR exhibit remarkably distinct substrate specificity for closely related bis-indoles.

Treatment of 5-chloro-2-aminobenzophenone (1) with o-phenylenediamine, sodium acetate, and acetic acid gave 2-(acetyl)amino-5-chlorobenzophenone (5) rather than N-[(2-amino-5-chlorophenyl)phenylmethylene]-1,2-benzenediamine (3), as reported by Kulkarni et al, [1]. Authentic 3 was prepared and treated with chloroacetic acid and polyphosphoric acid (PPA) to give 1, recovered 3, 2,8-dichloro-6,12-diphenyldibenzo[b,f][1,5]diazocine (9) and 2-chloro-6-(chloromethyl)-13-phenyl-5H-dibenzo[d,h][1,3,6]triazonine (10). Treatment of 5 with PPA, with or without chloroacetic acid, gave [5-chloro-2-[(6-chloro-4-phenyl-2-quinolinyl)amino]phenyl]phenylmethanone (11) as the sole product in 90% yield. Treatment of other benzophenones, acetophenones, and anilines with sodium acetate and acetic acid provided acetanilides in 78-96% yield, with the exception of 2′-aminoacetophenone (20), which gave a quantitative yield of 2-[((2-acetyl)phenyl)amino]-4-methylquinoline (21). The mechanism of acetanilide formation with sodium acetate and acetic acid is discussed. The structure of 21 was established using high resolution 1H nmr techniques. Attempts to prepare an authentic sample of 21 from 2-chlorolepidine (26) and (20) gave 4-methyl-N-[2-(4-methyl-2-quinolinyl)phenyl]-2-quinolinamine (29) as the major product.

An efficient and reliable three-step synthesis of 7-amino-3-hydroxyindan-1-one (7) is described. Compound 7 is a versatile, three-dimensional, three-point scaffold that is useful for the construction of focused compound libraries.

Herein, we describe the antifungal evaluation of 43 bisamidine compounds, of which 26 are new, having the scaffold [Am]–[HetAr]–[linker]–[HetAr]–[Am], in which [Am] is a cyclic or acyclic amidine group, [linker] is a benzene, pyridine, pyrimidine, pyrazine ring, or an aliphatic chain of two to four carbon, and [HetAr] is a 5,6-bicyclic heterocycle such as indole, benzimidazole, imidazopyridine, benzofuran, or benzothiophene. In the head-to-head series the two [HetAr] units are oriented such that the 5-membered rings are connected through the linker, and in the head-to-tail series, one of the [HetAr] systems is connected through the 6-membered ring; additionally, in some of the head-to-tail compounds, the [linker] is omitted. Many of these compounds exhibited significant antifungal activity against Candida albicans, Candida krusei, Candida glabrata, Candida parapsilosis, and Cryptococcus neoformans (MIC ⩽ 4 μg/ml). The most potent compounds, for example, P10, P19 and P34, are comparable in antifungal activities to amphotericin B (MIC 0.125 μg/ml). They exhibited rapid fungicidal activity (>3 log10 decrease in cfu/ml in 4 h) at concentrations equivalent to 4× the MIC in time kill experiments. The bisamidines strongly inhibited DNA, RNA and cell wall biosynthesis in C. albicans in macromolecular synthesis assays. However, the half-maximal inhibitory concentration for DNA synthesis was approximately 30-fold lower than those for RNA and cell wall biosynthesis. Fluorescence microscopy of intact cells of C. albicans treated with a bisamidine exhibited enhanced fluorescence in the presence of DNA, demonstrating that the bisamidine was localized to the nucleus. The results of this study show that bisamidines are potent antifungal agents with rapid fungicidal activity, which is likely to be the result of their DNA-binding activity. Although it was difficult to obtain a broad-spectrum antifungal compound with low cytotoxicity, some of the compounds (e.g., P9, P14 and P43) exhibited favorable CC50 values against HeLa cells and maintained considerable antifungal activity.

2-Pyridones are well-known ambident nucleophiles which are capable of reacting with electrophiles through either the nitrogen or oxygen atom to form N-alkyl-2-pyridones or 2-alkoxypyridines, respectively. It has been shown that the ratio of these products can be affected by a number of factors including the nature of the electrophile, the base used for deprotonation, and the solvent. We have now discovered a relationship between the ratio of N- and O-alkylation products and the nature of substituents on the pyridone ring when the Mitsunobu reaction is used to alkylate 2-pyridones.

Two new syntheses of the phytoalexin methyl 1-methoxyindole-3-carboxylate (1) were developed that employ the same key intermediate. Methyl 2-(2-nitrophenyl)acrylate (2) was directly converted into phytoalexin 1 using modified Cadogan–Sundberg indole synthesis conditions with trimethyl phosphite. Acrylate 2 also underwent reductive cyclization with stannous chloride to give methyl 1-hydroxyindole-3-carboxylate (3), which was methylated to produce phytoalexin 1.

Many genetic (gene deletion, interruption or mutation), epigenetic (such as antisense or small interfering RNA) and immunological methods are being applied in 'high-throughput target validation' studies of the novel potential targets arising from whole genome sequencing. Such applications often focus on 'loss of function' approaches. However, target validation is most reliable when multiple orthogonal approaches are used. Initiating a target-based discovery project based on correlative evidence is faster than awaiting causative evidence. Indeed, the multiple tools needed to generate firm proof usually include methods and reagents only generated after starting a discovery project with little evidence beyond correlations. Robust and rigorous tests of whether a drug candidate is efficacious in vivo because of its effects on a specific molecular particular target are best made by simultaneously applying multiple orthogonal tools. Examples of the orthogonal tools approach will be discussed.

20-Fluoro-17(20)-pregnenolone derivatives were designed as enol mimics of pregnenolone. All of the targeted, novel fluoroolefins were potent inhibitors of C 17(20) lyase.

A new route to esters of ( E)-2-(styrylsulfonyl)aceticacid has been developed. Treatment of the methyl ester with hydrazine hydrategave the six-membered 4-amino-5-phenylthiomorpholine-3-one 1,1-dioxide( 1), and not the seven-membered 6-phenyl-1,4,5-thiadiazepane-3-one1,1-dioxide ( 2) as previously describedby others. In contrast, treatment of the ethyl ester with the samereagent provided a separable 1:1.2 mixture of 1 and 2. The NMR spectra of both 1 and 2 differed substantially from that describedfor 2 in the previous report.

The in vitro activity of 5 novel Microbiotix bis-indole agents (MBXs) (Microbiotix, Worcester, MA) against 30 multidrug-resistant (MDR) Acinetobacter baumannii (including 18 resistant to carbapenems) was evaluated. Overall, MIC 90 's ranged from 1 to 8 μg/mL, whereas those for imipenem were >64 μg/mL. MBX 1196 was the most potent (MIC 90 , 1 μg/mL). MBXs are compounds that are highly effective against MDR A. baumannii .

The described synthesis starts from the commercially available furan-3-one derivative (I) which undergoes a reductive dehydration and conversion to a triflate derivative.

Cyclin-dependent kinases (CDKs) belong to a class of enzymes that control the ability of a cell to enter into and proceed through the cell division cycle. Using purine as a scaffold, we have synthesized a number of nanomolar inhibitors of CDK-2/cyclin E. In this report, the synthesis of a series of piperidine-substituted purine analogs will be presented, as well as some of their in vitro and in vivo biological effects.

Studies on tachykinin peptides and the corresponding neurokinin receptors (NKr) have increased dramatically recently due to the discovery of selective, orally-active, metabolically stable and sometimes CNS penetrating NKr antagonists. After demonstrating the potential use for NKr antagonists in animal models, some compounds have recently progressed into clinical trials and a few results have been published. NKr antagonists have demonstrated efficacy for the treatment of emesis and depression, while results in other areas have been disappointing. Nonetheless, this area is coming to the exciting time of proof of concept in humans. Demonstration of the involvement of tachykinin peptides in biological functions continues to grow, as do the potential indications for NKr antagonists. More drug candidates are undergoing clinical trials for various conditions and these results could widen the potential use for NKr antagonists.

Inhibition of human neutrophil elastase (HNE), (EC 3.4.21.37), as an approach to the treatment of inflammatory diseases in the chronic obstructive pulmonary disease (COPD) continuum, such as emphysema and chronic bronchitis, is still an area of intense research within the pharmaceutical industry. This review summarises some recent developments in the design and evaluation of HNE inhibitors as therapeutic agents since the subject was last reviewed in this publication in 1997.

The structurally related neuropeptides, substance P (SP), neurokinin A (NKA), and neurokinin B (NKB), which belong to a family of molecules termed tachykinins and are widely distributed in the central and peripheral nervous systems, influence the function of many tissues. SP and NKA have links to the fol)owing chronic diseases: asthma, inflammatory bowel disorders, rheumatoid arthritis, pain and psychiatric disorders. These peptides exert their effects through three G-protein coupled receptor sub types, namely, the NK1, NK2, and NK3 receptors. Non-peptide antagonists of these receptors may provide opportunities for disease treatments. In this review, the very recent advances in nonpeptide neurokinin receptor antagonists will be described with an emphasis on structure-activity relationships which have been developed.

A new method for attaching aldehydes to solid supports has been developed employing a 2,2-bis(hydroxymethyl)propionic acid (DMPA®) functionalized resin. High loading levels are obtained for both aryl and alkyl aldehydes protected as their respective acetals. Treatment of the derivatized resin with 95% TFA then cleanly affords the recovered aldehyde in high yield.

A series of P2-modified, orally active peptidic inhibitors of human neutrophil elastase (HNE) are reported. These pentafluoroethyl ketone-based inhibitors were designed using pentafluoroethyl ketone 1 as a model. Rational structural modifications were made at the P3, P2, and activating group (AG) portions of 1 based on structure-activity relationships (SAR) developed from in vitro (measured Ki) data and information provided by modeling studies that docked inhibitor 1 into the active site of HNE. The modeling-based design was corroborated with X-ray crystallographic analysis of the complex between 1 and porcine pancreatic elastase (PPE) and subsequently the complex between 1 and HNE.

The type III secretion system (T3SS) is a clinically important virulence mechanism in Pseudomonas aeruginosa that secretes and translocates effector toxins into host cells, impeding the host's rapid innate immune response to infection. Inhibitors of T3SS may be useful as prophylactic or adjunctive therapeutic agents to augment the activity of antibiotics in P. aeruginosa infections, such as pneumonia and bacteremia. One such inhibitor, the phenoxyacetamide MBX 1641, exhibits very responsive structure-activity relationships, including striking stereoselectivity, in its inhibition of P. aeruginosa T3SS. These features suggest interaction with a specific, but unknown, protein target. Here, we identify the apparent molecular target by isolating inhibitor-resistant mutants and mapping the mutation sites by deep sequencing. Selection and sequencing of four independent mutants resistant to the phenoxyacetamide inhibitor MBX 2359 identified the T3SS gene pscF , encoding the needle apparatus, as the only locus of mutations common to all four strains. Transfer of the wild-type and mutated alleles of pscF , together with its chaperone and cochaperone genes pscE and pscG , to a Δ pscF P. aeruginosa strain demonstrated that each of the single-codon mutations in pscF is necessary and sufficient to provide secretion and translocation that is resistant to a variety of phenoxyacetamide inhibitor analogs but not to T3SS inhibitors with different chemical scaffolds. These results implicate the PscF needle protein as an apparent new molecular target for T3SS inhibitor discovery and suggest that three other chemically distinct T3SS inhibitors interact with one or more different targets or a different region of PscF.