Inhibition on HIV-1 integrase activity and nitric oxide production of compounds from Ficus glomerata

An ethanol Ficus glomerata wood extract and its purified components were investigated for their HIV-1 integrase (IN) and nitric oxide (NO) inhibitory activities. From bioassay-guided isolation, five compounds: beta-sitosterol-D-glucoside (1), aloe-emodin (2), genistein (3), 1,3,6-trihydroxy-8-methyl-anthraquinone (4) and 3-(1-C-beta-D-glucopyranosyl)-2,6-dihydroxy-5-methoxybenzoic acid (5) were isolated. Among the tested samples, at concentrations of 100 microM; compound 2 showed 31.9% inhibition of HIV-1 IN, followed by 4 (19.5%), whereas other compounds were inactive. With regard to the inhibitory effect on NO production, 3 possessed the highest activity with an IC50 value of 27.5 microM, followed by 4 (IC50 = 34.7 microM) and 2 (IC50 = 41.8 microM), respectively. This is the first time that compounds 2-5 have been isolated from Ficus glomerata.     

Two new triterpenes from the Rhizome of Dryopteris crassirhizoma, and inhibitory activities of its constituents on human immunodeficiency virus-1 protease

Two new hopane type triterpenes, named dryopteric acids A (1) and B (2), were isolated from the Rhizome of Dryopteris crassirhizoma (Aspiadaceae) together with sixteen known compounds (3-18). Of isolated compounds, ursolic acid (15), and dryopteric acid A (1) and B (2) showed potent inhibitory activities against HIV-1 protease with IC50 values of 8.9-44.5 microM. In addition, acetylated compounds 1 and 2 appreciably increased inhibitory activities with their IC50 values of 1.7 and 10.8 microM, respectively.     

Polyoxometalate HIV-1 protease inhibitors. A new mode of protease inhibition.

Nb-containing polyoxometalates (POMs) of the Wells-Dawson class inhibit HIV-1 protease (HIV-1P) by a new mode based on kinetics, binding, and molecular modeling studies. Reaction of alpha(1)-K(9)Li[P(2)W(17)O(61)] or alpha(2)-K(10)[P(2)W(17)O(61)] with aqueous H(2)O(2) solutions of K(7)H[Nb(6)O(19)] followed by treatment with HCl and KCl and then crystallization affords the complexes alpha(1)-K(7)[P(2)W(17)(NbO(2))O(61)] (alpha(1)()1) and alpha(2)-K(7)[P(2)W(17)(NbO(2))O(61)] (alpha(2)()1) in 63 and 86% isolated yields, respectively. Thermolysis of the crude peroxoniobium compounds (72-96 h in refluxing H(2)O) prior to treatment with KCl converts the peroxoniobium compounds to the corresponding polyoxometalates (POMs), alpha(1)-K(7)[P(2)W(17)NbO(62)] (alpha(1)()2) and alpha(2)-K(7)[P(2)W(17)NbO(62)] (alpha(2)()2), in moderate yields (66 and 52%, respectively). The identity and high purity of all four compounds were confirmed by (31)P NMR and (183)W NMR. The acid-induced dimerization of the oxo complexes differentiates sterically between the cap (alpha(2)) site and the belt (alpha(1)) site in the Wells-Dawson structure (alpha(2)()2 dimerizes in high yield; alpha(1)()2 does not). All four POMs exhibit high activity in cell culture against HIV-1 (EC(50) values of 0.17-0.83 microM), are minimally toxic (IC(50) values of 50 to >100 microM), and selectively inhibit purified HIV-1 protease (HIV-1P) (IC(50) values for alpha(1)()1, alpha(2)()1, alpha(1)()2, and alpha(2)()2 of 2.0, 1.2, 1.5, and 1.8 microM, respectively). Thus, theoretical, binding, and kinetics studies of the POM/HIV-1P interaction(s) were conducted. Parameters for [P(2)W(17)NbO(62)](7)(-) were determined for the Kollman all-atom (KAA) force field in Sybyl 6.2. Charges for the POM were obtained from natural population analysis (NPA) at the HF/LANL2DZ level of theory. AutoDock 2.2 was used to explore possible binding locations for the POM with HIV-1P. These computational studies strongly suggest that the POMs function not by binding to the active site of HIV-1P, the mode of inhibition of all other HIV-1P protease inhibitors, but by binding to a cationic pocket on the "hinge" region of the flaps covering the active site (2 POMs and cationic pockets per active homodimer of HIV-1P). The kinetics and binding studies, conducted after the molecular modeling, are both in remarkable agreement with the modeling results: 2 POMs bind per HIV-1P homodimer with high affinities (K(i) = 1.1 +/- 0.5 and 4.1 +/- 1.8 nM in 0.1 and 1.0 M NaCl, respectively) and inhibition is noncompetitive (k(cat) but not K(m) is affected by the POM concentration).     

CCR5 antagonists as anti-HIV-1 agents. 1. Synthesis and biological evaluation of 5-oxopyrrolidine-3-carboxamide derivatives

A novel lead compound, N-(3-[4-(4-fluorobenzoyl)piperidin-1-yl]propyl)-1-methyl-5-oxo-N-phenylpyrrolidine-3-carboxamide (1), was identified as a CCR5 antagonist by high-throughput screening using [(125)I]RANTES and CCR5-expressing CHO cells. The IC(50) value of 1 was 1.9 microM. In an effort to improve the binding affinity of 1, a series of 5-oxopyrrolidine-3-carboxamides was synthesized. Introduction of 3,4-dichloro substituents to the central phenyl ring (10i, IC(50)=0.057 microM; 11b, IC(50)=0.050 microM) or replacing the 1-methyl group of the 5-oxopyrrolidine moiety with a 1-benzyl group (12e, IC(50)=0.038 microM) was found to be effective for improving CCR5 affinity. Compound 10i, 11b, and 12e also inhibited CCR5-using HIV-1 envelope-mediated membrane fusion with IC(50) values of 0.44, 0.19, and 0.49 microM, respectively.     

Inhibitory effects of constituents from Cynomorium songaricum and related triterpene derivatives on HIV-1 protease

From CH2Cl2 and MeOH extracts of the stems of Cynomorium songaricum RUPR. (Cynomoriaceae), ursolic acid and its hydrogen malonate were isolated as inhibitors of human immunodeficiency virus type 1 (HIV-1) protease, with 50% inhibitory concentrations (IC50) of 8 and 6 microM, respectively. Amongst various synthesized dicarboxylic acid hemiesters of related triterpenes, inhibitory activity tended to increase in the order of oxalyl, malonyl, succinyl and glutaryl hemiesters, for triterpenes such as ursolic acid, oleanolic acid and betulinic acid. The most potent inhibition was observed for the glutaryl hemiesters, with an IC50 of 4 microM. From the water extract of the stems of C. songaricum, flavan-3-ol polymers, consisting of epicatechin as their extender flavan units, were also found to be potent inhibitory principles against HIV-1 protease.     

A new sesquiterpenoid hydroquinone from the marine sponge Dysidea arenaria

Detailed chemical investigation of the South China sponge Dysidea arenaria resulted in the isolation of a new sesquiterpenoid hydroquinone, 19-hydroxypolyfibrospongol B (1), along with five known compounds: polyfibrospongol B (2), isosemnonorthoquinone (3), ilimaquinone (4), smenospongine (5) and smenotronic acid (6). The structures were determined by extensive spectroscopic analysis. The in vitro anti- HIV activity on HIV-1 RT was evaluated. Compounds 3 -6 displayed moderate inhibitory activity, with IC(50)values of 239.7, 16.4, 176.1, and 130.4 microM, respectively, while 1 and 2 were found to be inactive against the same biological target.     

Fluorescent HIV-1 Dimerization Initiation Site: design, properties, and use for ligand discovery

The HIV-1 Dimerization Initiation Site (DIS) is an intriguing, yet underutilized, viral RNA target for potential antiretroviral therapy. To study the recognition features of this target and to provide a quantitative, rapid, and real-time tool for the discovery of new binders, a fluorescence-based assay has been constructed. It relies on strategic incorporation of 2-aminopurine, an isosteric fluorescent adenosine analogue, into short hairpin RNA constructs. These oligomers self-associate to form a kissing loop that thermally rearranges into a more stable extended duplex, thereby mimicking the association and structural features of the native RNA sequence. We demonstrate the ability of two fluorescent DIS constructs, DIS272(2AP) and DIS273(2AP), to report the binding of known DIS binders via changes in their emission intensity. Binding of aminoglycosides such as paromomycin to DIS272(2AP) results in significant fluorescence enhancement, while ligand binding to DIS273(2AP) results in fluorescence quenching. These observations are rationalized by comparison to the sequence-analogous bacterial A-site, where the relative emission of the fluorescent probe is dependent on the placement of the flexible purine residues inside or outside the helical domain. Analysis of binding isotherms generated using DIS272(2AP) yields submicromolar EC50 values for paromomycin (0.5 +/- 0.2 microM) and neomycin B (0.6 +/- 0.2 microM). Other neomycin-family aminoglycosides are less potent binders with neamine, the core pharmacophore, displaying the lowest affinity of 21 +/- 1 microM. Screening of additional aminoglycosides and their derivatives led to the discovery of new, previously unreported, aminoglycoside binders of the HIV DIS RNA, among them butirosin A (5.5 +/- 0.6 microM) and apramycin (7.6 +/- 1.0 microM). A conformationally constrained neomycin B analogue displays a rather high affinity to the DIS (1.9 +/- 0.2 microM). Among a series of nucleobase aminoglycoside conjugates, only the uracil derivatives display a measurable affinity using this assay with EC50 values in the 2 microM range. In addition, similarity between the solution behavior of HIV-1 DIS and the bacterial decoding A-site has been observed, particularly with respect to the intra- and extra-helical residence of the conformationally flexible A residues within the bulge. Taken together, the observations reported here shed light on the solution behavior of this important RNA target and are likely to facilitate the design of new DIS selective ligands as potential antiretroviral agents.     

Rational design and synthesis of a novel class of active site-targeted HIV protease inhibitors containing a hydroxymethylcarbonyl isostere. Use of phenylnorstatine or allophenylnorstatine as a transition-state mimic.

A novel class of HIV-1 protease inhibitors containing a hydroxymethylcarbonyl (HMC) isostere were designed from the substrate transition state and synthesized. Phenylnorstatine [Pns; (2R,3S)-3-amino-2-hydroxy-4-phenylbutyric acid] and the 2S diastereomer, (2S,3S)-3-amino-2-hydroxy-4-phenylbutyric acid, named allophenylnorstatine (Apns) were effective transition-state mimics, and incorporation of Pns-Pro or Apns-Pro at the P1-P1' site gave potent and specific HIV-1 protease inhibitors. In the inhibitory assays, the chemically synthesized [Ala67,95] HIV-1 protease was used.     

Three new lignans, longipedunins A-C, from Kadsura longipedunculata and their inhibitory activity against HIV-1 protease

Three new lignans, longipedunins A (1), B (2) and C (3), together with three known compounds, benzoyl-binankadsurin A (4), acetyl-binankadsurin A (5) and schisanlactone A (6), were isolated from Kadsura longipedunculata. Their structures and stereochemistry were determined by spectral and single-crystal X-ray analyses. Compounds 1 and 6 showed appreciable inhibitory activity against HIV-1 protease with IC50 values of 50 and 20 microM, respectively.     

The C-disaccharide alpha-C(1-->3)-mannopyranoside of N-acetylgalactosamine is an inhibitor of glycohydrolases and of human alpha-1,3-fucosyltransferase VI. Its epimer alpha-(1-->3)-mannopyranoside of N-acetyltalosamine is not

The radical C-glycosidation of (-)-(1S,4R,5R, 6R)-6-endo-chloro-3-methylidene-5-exo-(phenylseleno)-7-ox abi cyclo[2. 2.1]heptan-2-one ((-)-4) with 2,3,4, 6-tetra-O-acetyl-alpha-D-mannopyranosyl bromide gave (+)-(1S,3R,4R, 5R,6R)-6-endo-chloro-5-exo-(phenylseleno)-3-endo-(1',3',4', 5'-tetra-O-acetyl-2', 6'-anhydro-7'-deoxy-D-glycero-D-manno-heptitol-7'-C-yl)-7-oxabi cyc lo[ 2.2.1]hept-2-one ((+)-5) that was converted into (+)-(1R,2S,5R, 6R)-5-acetamido-3-chloro-2-hydroxy-6-(1',3',4',5'-tetra-O-acetyl)-2', 6'-anhydro-7'-deoxy-D-glycero-D-manno-heptitol-7'-C-yl)cyclohex -3-en- 1-yl acetate ((+)-10) and into (+)-(1R,2S,5R, 6S)-5-bromo-3-chloro-2-hydroxy-6-(1',3',4',5'-tetra-O-acetyl-2', 6'-anhydro-7'-deoxy-D-glycero-D-manno-heptitol-7'-C-yl)cyclohex -3-en- 1-yl acetate ((+)-19). Ozonolysis of (+)-10 and further transformations provided 2-acetamido-2,3-dideoxy-3-C-(2', 6'-anhydro-7'-deoxy-D-glycero-D-manno-heptitol-7'-C-yl)-D-galac tos e (alpha-C(1-->3)-D-mannopyranoside of N-acetylgalactosamine (alpha-D-Manp-(1-->3)CH(2)-D-GalNAc): 1). Displacement of the bromide (+)-19 with NaN(3) in DMF provided the corresponding azide ((-)-20) following a S(N)2 mechanism. Ozonolysis of (-)-20 and further transformations led to 2-acetamido-2,3-dideoxy-3-C-(2', 6'-anhydro-7'-deoxy-D-glycero-D-manno-heptitol-7'-C-yl)-D-talose (alpha-C(1-->3)-D-mannopyranoside of N-acetyl D-talosamine (alpha-D-Manp-(1-->3)CH(2)-D-TalNAc): 2). The neutral C-disaccharide 1 inhibits several glycosidases (e.g., beta-galactosidase from jack bean with K(i) = 7.5 microM, alpha-L-fucosidase from human placenta with K(i) = 28 microM, beta-glucosidase from Caldocellum saccharolyticum with K(i) = 18 microM) and human alpha-1, 3-fucosyltransferase VI (Fuc-TVI) with K(i) = 120 microM whereas it 2-epimer 2 does not. Double reciprocal analysis showed that the inhibition of Fuc-TVI by 1 displays a mixed pattern with respect to both the donor sugar GDP-fucose and the acceptor LacNAc with K(i) of 123 and 128 microM, respectively.     

Synthesis of new 1H-1,2,4-triazolylcoumarins and their antitumor and anti-HIV activities

A variety of 7-[(1,5-dialkyl-1H-1,2,4-triazolyl)methoxy(and methyl)]coumarins were synthesized from cycloaddition of 2-(2H-benzopyran-7-yloxy)acetonitrile and 2-(5-methoxy-4-methyl-2H-benzopyran-7-yloxy)acetonitrile, respectively, with various reactive cumulene intermediates via spontaneous rearrangements. The anticancer (breast, lung, CNS cancers) and antiviral (HIV-1, HIV-2) properties of some compounds were investigated in vitro. 5-Methoxy-4-methyl-7-[(6,7,8,9-tetrahydro-5H-[1,2,4]triazolo[1,5-a]azepin-2-yl)methyl]coumarin showed some inhibition of HIV-1. Published in Khimiya Geterotsiklicheskikh Soedinenii, No. 5, pp. 669–678, May, 2006.     

Phenolics with antiviral activity from Millettia erythrocalyx and Artocarpus lakoocha

From the leaves of Millettia erythrocalyx, a new flavone named 3',5'-dimethoxy-[2",3": 7,8]-furanoflavone and three known compounds were isolated. Assays for anti-herpes simplex virus activity (HSV-1 and HSV-2) were performed on 24 phenolic compounds obtained from M. erythrocalyx and Artocarpus lakoocha. It was found that the flavones ovalifolin, pongol methyl ether and millettocalyxin A, and the stilbene oxyresveratrol possessed moderate activity against both types of HSV. In addition, oxyresveratrol was evaluated for potential anti-HIV activity against a wild-type human immunodeficiency virus type 1 (HIV-1/LAI) isolate and was found to be a modest inhibitor of HIV (EC50 28.2 microM), showing no toxicity in PBM, CEM and Vero cells at 100 microM. The heartwood of A. lakoocha, which contains a large amount of oxyresveratrol, could be considered as a source of starting material for the development of new natural product-based anti-HSV and anti-HIV agents.     

3-芳氧基-1-丙胺类HIV-1 Tat/PCAF BRD抑制剂的合成及生物活性

为了研究HIV-1 Tat/PCAF BRD抑制剂的构效关系, 合成了6个3-芳氧基-1-丙胺类化合物. 以取代的2-硝基苯酚为起始原料, 在常规加热和微波辐射加热下与1,3-二溴丙烷反应合成3-(2-硝基芳氧基)-1-溴丙烷(3), 结果显示, 微波辐射加热比常规加热下的反应速度明显加快, 收率有所提高. 3和邻苯二甲酰亚胺钾进行N-烷基化反应合成了2-[3-(芳氧基)-丙基]二氢异吲哚-1,3-二酮, 再经肼解得到了目标化合物, 所有化合物的结构均经FTIR, ¹H NMR, ¹³C NMR及HRMS确证. ELISA检测法测定了它们体外抑制HIV-1 Tat/PCAF BRD的活性, 并对影响活性的因素进行了讨论.