Alanine scan-guided synthesis and biological evaluation of analogues of culicinin D,a potent anticancer peptaibol

Culicinin D (1), a 10 amino acid peptaibol originally isolated from Culicinomyces clavisporus, exhibits potent activity against a range of cancer cell lines. Building on our previous work exploring the structure–activity relationship (SAR) of the unusual (2S,4S,6R)-AHMOD residue, a series of analogues of culicinin D were prepared to further investigate the SAR of these peptaibols. Alanine scanning of a potent and readily accessible analogue 23 revealed the effect of each residue on antiproliferative activity, and a small panel of analogues were prepared to explore the SAR of the non-natural amino acid residue (2S,4R)-AMD. Results from the alanine scan were used to design an expanded library of culicinin D analogues, leading to the discovery of cyclohexylalanine analogue 52, which exhibited better antiproliferative activity than the natural product 1.     

Discovery of indane propanamides as potent and selective TRPV1 antagonists

A series of indane-type acetamide and propanamide analogues were investigated as TRPV1 antagonists. The analysis of structure–activity relationship indicated that indane A-region analogues exhibited better antagonism than did the corresponding 2,3-dihydrobenzofuran and 1,3-benzodioxole surrogates. Among them, antagonist 36 exhibited potent and selective antagonism toward capsaicin for hTRPV1 and mTRPV1. Further, in vivo studies indicated that antagonist 36 showed excellent analgesic activity in both phases of the formalin mouse pain model and inhibited the pain behavior completely at a dose of 1 mg/kg in the 2nd phase.     

New antituberculotics originated from salicylanilides with promising in vitro activity against atypical mycobacterial strains

A new series of 30 N-protected amino acid esters were prepared as a part of ongoing search for new anti-tuberculosis active salicylanilides. The esters possess high in vitro activity against Mycobacterium tuberculosis, Mycobacterium avium, and two strains of Mycobacterium kansasii, where one is an isolate from the patient, with MIC in the range 1–32 μmol/L for all tested strains. The prepared esters can be considered as prodrugs with better bio-availability and as more efficient transport forms through the mycobacterial cell membranes due to the higher lipophilicity. The experimental and calculated lipophilicity, stability, antituberculotic activity, cytotoxicity as well as the quantitative structure–activity relationships (QSARs) explored by the Intelligent Problem Solver (IPS) in Trajan Neural Network Simulator 6.0 are presented.     

Conformation studies on Burkholderia cenocepacia lipase via resolution of racemic 1-phenylethanol in non-aqueous medium and its process optimization

In this study, the effect of various organic solvents on enzyme activity and substrate enantiomeric excess (ee_s) of the lipase from Burkholderia cenocepacia (BCL) was investigated in the enantioselective transesterification of 1-phenylethanol. Secondary structure analysis by Fourier transform-infrared spectroscopy (FT-IR) showed that the variations in secondary structure element content (α-helix, β-sheet, β-turn and random coil) were probably responsible for the changes in enzyme activity and ee_s. Furthermore, the change in fluorescence intensity indicated, to some extent, the alteration in tertiary structure, which may also explain why organic solvents affect enzyme activity and ee_s. Moreover, response surface methodology (RSM) was employed to optimize the reaction parameters. The optimized reaction conditions were: substrate molar ratio 4.7:1; reaction time 18.6 h, and reaction temperature 53.4 °C. Under the optimal reaction conditions, the ee_s and ee_p were respectively 99.22% and 98.74%, and the corresponding enzyme activity was 1392.2 U/min/g protein. Compared with other lipases, BCL exhibited better catalytic efficiency and has significant potential in industrial applications.     

Bioactive metabolites from the Andean flora. Antituberculosis activity of natural and semisynthetic azorellane and mulinane diterpenoids

Natural products are recognized as an important source of new and better pharmaceuticals for the treatment of diseases such as tuberculosis. The azorellane and mulinane diterpenoids represent an interesting group of bioactive metabolites produced by Andean plants belonging to the Azorella, Mulinum, Laretia and Bolax genus. Testing of natural and semisynthetic azorellanes and mulinanes against two Mycobacterium tuberculosis strains showed that while most changes in the structure of the natural metabolites result in the loss of antituberculosis activity, methylation of the C-20 carboxyl group improves the biological activity of the corresponding derivatives.     

α-Haloacetophenone and analogues as potential antibacterial agents and nematicides

A series of α-haloacetophenones and analogues were synthesized. The bioassays show that some target compounds have good antibacterial activity against Xanthomonas oryzae pv. oryzae (Xoo), Xanthomonas axonopodis pv. citri (Xac) and Meloidogyne incognita (M. incognita). Especially, the compound 24 has good in vitro and in vivo antibacterial activities against Xoo, the EC₅₀ value, curative and protection activities are 0.09 mg/L, 48.9%, and 52.3%, respectively, which are better than the thiodiazole copper and bismerthiazol. Meanwhile, the compound 24 has good in vitro antibacterial activity against Xac, and has an EC₅₀ value of 1.6 mg/L. Moreover, the compound 19 exhibits good nematicidal activity M. incognita, with the LC₅₀ value of 1.0 mg/L, which is better than the positive control avermectin. In addition, the compound 24 can inhibit the formation of extracellular polysaccharide and biofilm of Xoo, and change the permeability of cell membrane. α-haloacetophenone and analogues have the advantages of simple structure, high efficiency, broad spectrum of biological activity, and can be used as antibacterial agents and nematicides or lead compounds in the future.     

Synthesis of new pyrazoles and pyrozolo [3,4-b] pyridines as anti-inflammatory agents by inhibition of COX-2 enzyme

New pyrazoles and pyrazolo[3,4-b] pyridines were synthesized and their structure was confirmed by elemental analyses as well as IR, ¹H NMR, ¹³C NMR, and mass spectral data. All the newly synthesized derivatives were evaluated in vitro for inhibitory activity against COX-1 and COX-2 enzymes and their IC₅₀ values were calculated, most of the derivatives showed good inhibitory activity with derivatives IVb, IVh and IVJ showing inhibitory activity better than celecoxib. Moreover, the eight most potent derivatives IVa, IVb, IVc, IVd, IVe, IVh, IVJ, and IVL were selected for in vivo assay to measure their effect on paw edema in rates and their ulcerogenic effect. Compounds IVa, IVb and IVc were found to be the most active and selective as COX-2 inhibitors and most effective in protection from edema, they were also found to have lowest ulcerogenic effect among all derivatives.     

Novel 4-thiazolidinone derivatives as potential antifungal and antibacterial drugs

As part of ongoing studies in developing new antimicrobials, a class of structurally novel 4-thiazolidinone derivatives incorporating three known bioactive nuclei such as thiazole, thiazolidinone and adamantane was synthesized by the multi-step reaction protocol, already reported in the literature. NMR and Molecular Modeling techniques were employed for structure elucidation and Z/E potential isomerism configuration of the analogues. Evaluation of antibacterial and antifungal activity showed that almost all compounds exhibited better results than reference drugs thus they could be promising candidates for novel drugs.     

The 1.9 Å Structure of Human α-N-Acetylgalactosaminidase: The Molecular Basis of Schindler and Kanzaki Diseases

α-N-acetylgalactosaminidase (α-NAGAL; E.C. 3.2.1.49) is a lysosomal exoglycosidase that cleaves terminal α-N-acetylgalactosamine residues from glycopeptides and glycolipids. In humans, a deficiency of α-NAGAL activity results in the lysosomal storage disorders Schindler disease and Kanzaki disease. To better understand the molecular defects in the diseases, we determined the crystal structure of human α-NAGAL after expressing wild-type and glycosylation-deficient glycoproteins in recombinant insect cell expression systems. We measured the enzymatic parameters of our purified wild-type and mutant enzymes, establishing their enzymatic equivalence. To investigate the binding specificity and catalytic mechanism of the human α-NAGAL enzyme, we determined three crystallographic complexes with different catalytic products bound in the active site of the enzyme. To better understand how individual defects in the α-NAGAL glycoprotein lead to Schindler disease, we analyzed the effect of disease-causing mutations on the three-dimensional structure.     

Design and synthesis of C10 modified and ring-truncated canthin-6-one analogues as effective membrane-active antibacterial agents

A series of canthin-6-one analogues were designed and synthesized in order to study their antibacterial activity and structure–activity relationships. Compound 22 showed a broad spectrum of antibacterial activity and exhibited better bactericidal effect (8-fold superiority against Staphylococcus aureus and 2-fold superiority against Ralstonia solanacearum) than fosfomycin sodium and propineb with a minimum inhibitory concentration value of 2?μg/mL. Moreover, it showed low cytotoxicity, stimulation on germination rates and good “drug-like” properties. Membrane-active mechanism was further studied by fluorescent microscopy, scanning electron microscopy, cytoplasmic β-galactosidase leakage assay and evaluation of the molecular docking. The results showed that 22 may exert its bactericidal effect by damaging bacterial cell membranes and influencing the membrane formation, both of which could lead to cell death. The in vivo antibacterial assay with a protective efficacy of 68% demonstrated the potential of C ring-truncated canthin-6-one 22 as a new bactericide.     

Synthesis and cytotoxicity of novel imidazo[4,5-d]azepine compounds derived from marine natural product ceratamine A

A series of novel imidazo[4,5-d]azepine compounds derived from marine natural product ceratamine A were designed and synthesized in 7 steps. Most compounds exhibited comparable cytotoxicity against five human cancer cell lines (HCT-116, HepG2, BGC-823, A549 and A2780) to natural product ceratamine A. Compound 1k, bearing methoxy group at C-14, C-15 and C-16, showed the best in vitro cytotoxicity, which was better than ceratamine A. The structure and activity relationships study showed that the benzyloxymethyl group on N-3 played an important role on the cytotoxicity.     

Discovery and optimization of 5,7-dihydro-6H-pyrrolo[2,3-d]pyrimidin-6-one derivatives as mTORC1/mTORC2 dual inhibitors

New potent mTORC1/mTORC2 dual inhibitors, 5,7-dihydro-6H-pyrrolo[2,3-d]pyrimidin-6-one derivatives, were obtained by optimizing functional groups on our previously reported PI3Kα inhibitor. All the target compounds were synthesized and structural optimization on the structure of the lead compound based on cytotoxic activity. The results showed that some of the target compounds exhibited moderate to high cytotoxic activity against cell line U87MG and PC-3. The activities against mTOR kinase were investigated and the compound 12q showed excellent activity with an IC₅₀ value of 54 nM in the same level of the positive control BEZ235 with IC₅₀ value of 55 nM under the same test conditions. The western blot and cell cycle results demonstrate that compound 12q is a candidate as an mTORC1/mTORC2 dual-target inhibitor. The theoretical calculations were also performed to better understanding the binding modes of the compound 12q in the mTOR active site.     

Plant extracts with anti-inflammatory properties—A new approach for characterization of their bioactive compounds and establishment of structure–antioxidant activity relationships

Geranium robertianum L. (Geraniacea) and Uncaria tomentosa (Willd.) DC. (Rubiaceae) plant extracts, frequently used in traditional medicine for treatment of inflammatory and cancer diseases, were studied to identify potential bioactive compounds that may justify their therapeutic use and their underlying mechanisms of action. Since some of the pharmacological properties of these plant extracts may be linked to their antioxidant potential, the antioxidant activity, in relation to free radical scavenging, was measured by the ABTS/HRP and DPPH assays, presenting U. tomentosa the higher activity. The antioxidant activity was also evaluated by scavenging of HOCl, the major strong oxidant produced by neutrophils and a potent pro-inflammatory agent. U. tomentosa was found to be a better protector against HOCl, which may justify its effectiveness against inflammatory diseases. SPE/LC-DAD was used for separation/purification purposes and ESI-MS/MS for identification/characterization of the major non-volatile components, mainly flavonoids and phenolic acids. The ESI-MS/MS methodology proposed can be used as a model procedure for identification/characterization of unknowns without the prerequisite for standard compounds analysis. The ESI-MS/MS data obtained were consistent with the antioxidant activity results and structure–activity relationships for the compounds identified were discussed.     

Novel benzofuran constituent from the husk of Carya cathayensis Sarg.

In an attempt to find the effective botanical fungicide, investigations were conducted on the husk of Carya cathayensis Sarg. (CCS). The results suggested that the crude extracts from the husk of CCS have obvious germicidal effects on the pathogenic fungi, while screening for new antifungal agents from the husk of C. cathayensis Sarg., a novel benzofuran-type compound, cathayenone A, was obtained. Its structure was elucidated by 1-D and 2-D NMR and MS data analyses. The inhibition rates of 0.1 mg/mL of cathayenone A against the spore germination of Exserohilum turcicum and Pyricularia oryzae were 95.8% and 86.7%, respectively, which showed better antifungal activity than tebuconazole fungicide. Therefore, cathayenone A has the potential to be a fungicide.     

Synthesis, characterization, and antifungal activity of novel quaternary chitosan derivatives

Three novel quaternary chitosan derivatives were successfully synthesized by reaction of chloracetyl chitosan (CACS) with pyridine (PACS), 4-(5-chloro-2-hydroxybenzylideneamino)-pyridine (CHPACS), and 4-(5-bromo-2-hydroxybenzylideneamino)-pyridine (BHPACS). The chemical structure of the prepared chitosan derivatives was confirmed by Fourier transform infrared (FT-IR) and ¹³C nuclear magnetic resonance (¹³C NMR) and their antifungal activity against Cladosporium cucumerinum, Monilinia fructicola, Colletotrichum lagenarium, and Fusarium oxysporum was assessed. Comparing with the antifungal activity of chitosan, CACS, and PACS, CHPACS and BHPACS exhibited obviously better inhibitory effects, which should be related to the synergistic reaction of chitosan itself with the grafted 2-[4-(5-chloro-2-hydroxybenzylideneamino)-pyridyl]acetyl and 2-[4-(5-bromo-2-hydroxybenzylideneamino)-pyridyl]acetyl.     

Anti-AIDS agents 84. Synthesis and anti-human immunodeficiency virus (HIV) activity of 2′-monomethyl-4-methyl- and 1′-thia-4-methyl-(3′R,4′R)-3′,4′-di-O-(S)-camphanoyl-(+)-cis-khellactone (DCK) analogs

In a continuing investigation into the pharmacophores and structure–activity relationship (SAR) of (3′R,4′R)-3′,4′-di-O-(S)-camphanoyl-(+)-cis-khellactone (DCK) as a potent anti-HIV agent, 2′-monomethyl substituted 1′-oxa, 1′-thia, 1′-sulfoxide, and 1′-sulfone analogs were synthesized and evaluated for inhibition of HIV-1 replication in H9 lymphocytes. Among them, 2′S-monomethyl-4-methyl DCK (5a)³ and 2′S-monomethyl-1′-thia-4-methyl DCK (7a) exhibited potent anti-HIV activity with EC₅₀ values of 40.2 and 39.1 nM and remarkable therapeutic indexes of 705 and 1000, respectively, which were better than those of the lead compound DCK in the same assay. In contrast, the corresponding isomeric 2′R-monomethyl-4-methyl DCK (6) and 2′R-monomethyl-1′-thia-4-methyl DCK (8) showed much weaker inhibitory activity against HIV-1 replication. Therefore, the bioassay results suggest that the spatial orientation of the 2′-methyl group in DCK analogs can have important effects on anti-HIV activity of this compound class.     

Production and characterization of genetically modified human IL-11 variants

Interleukin-11 (IL-11) has been expected as a drug on severe thrombocytopenia caused by myelo-suppressive chemotherapy. Whereas, development of IL-11 inhibitor is also expected for a treatment against IL-11 related cancer progression. Here, we will demonstrate the creation of various kinds of genetically modified hIL-11s. Modified vectors were constructed by introducing N- or O-glycosylation site on the region of hIL-11 that does not belong to the core α-helical motif based on the predicted secondary structure. N-terminal (N: between 22 to 23 aa), the first loop (M1:70 to 71 aa), the second loop (M2:114–115 aa), the third loop (M3:160–161 aa) and C-terminal (C: 200- aa) were selected for modification. A large scale production system was established and the characteristics of modified hIL-11s were evaluated. The structure was analyzed by amino acid sequence and composition analysis and CD-spectra. Glycan was assessed by monosaccharide composition analysis. Growth promoting activity and biological stability were analyzed by proliferation of T1165 cells. N-terminal modified proteins were well glycosylated and produced. Growth activity of 3NN with NASNASNAS sequence on N-terminal was about tenfold higher than wild type (WT). Structural and biological stabilities of 3NN were also better than WT and residence time in mouse blood was longer than WT. M1 variants lacked growth activity though they are well glycosylated and secondary structure is very stable. Both of 3NN and OM1 with AAATPAPG on M1 associated with hIL-11R strongly. These results indicate N-terminal and M1 variants will be expected for practical use as potent agonists or antagonists of hIL-11.     

Synthesis and structure–activity relationship analysis of caffeic acid amides as selective matrix metalloproteinase inhibitors

Four series of acid amides were synthesized, and through measurement using a fluorogenic substrate assay with human recombinant MMP-1, MMP-2 and MMP-9, compound 3f showed considerable inhibitory activities against MMP-2, MMP-9 and the best selectivity over MMP-1. Preliminary structure–activity relationship analysis indicated that caffeic acid amides with electron-donating groups at para-position of amino phenyl group showed better inhibitory activities and selectivity than those with electron-withdrawing groups, and the presence of adjacent dihydroxy in the caffeoyl group was very important for the MMP-2 and MMP-9 inhibitory activities.     

Crystal structure of HsEg5 in complex with clinical candidate CK0238273 provides insight into inhibitory mechanism, potency, and specificity

HsEg5 is an important mitotic kinesin responsible for bipolar spindle formation at early mitosis. A rich body of evidence shows that inhibition of HsEg5 can result in mitotic arrest followed by cellular apoptosis. Recently identified HsEg5 inhibitor, CK0238273, exhibits potent antitumor activity and is currently in clinical trial. Here we report the cocrystal structure of the motor domain of HsEg5 in complex with CK0238273 at a 2.15 Å resolution. Compared to the previously published HsEg5-Monastrol complex structure, CK0238273 shares the same induced-fit pocket with similar allosteric inhibitory mechanism. However, CK0238273 shows better fitting to the binding pocket with 65% increase of hydrophobic interaction area than that of Monastrol. Some unique hydrophilic interactions were also observed mostly between the phenyl ring and 8-chloro on quinazolinone of CK0238273 with ARG221 and GLY217. We believe that the combination of these interactions defines the superior potency and specificity of CK0238273.