RESUMO
Negatively charged albumins (NCAs, with the prototypes Suc-HSA and Aco-HSA), modified proteins with a potent anti-HIV-1 activity in the nanomolar concentration range, were studied for several aspects of their antiviral mechanism. In addition we investigated the antiviral activity of combinations and covalent conjugates of these NCAs and the reverse transcriptase inhibitor AZT. Addition of NCAs to HIV-1-infected target cells in time-of-addition experiments could be delayed for 30 min after infection before significant loss of activity occurred. Syncytium formation of HIV-infected and uninfected T-cells was inhibited by the NCAs at concentrations of 1-4 microg/ml. The gp120-mediated virus/cell binding, however, was only inhibited by the NCAs at a 10-fold higher concentrations. The combined data are compatible with a preferential influence on virus/cell fusion. A subsynergistic activity against HIV-1 was observed with the non-covalent mixture of Aco-HSA and AZT. When AZT was covalently coupled to the NCAs, and added one hour after infection of the target cells, the anti-HIV-1 activity of NCA-AZTMP was diminished by only 2-6-fold, while this was diminished at least 120-fold for the NCAs. Furthermore the addition of NCA-AZTMP could be delayed up to at least 3 h after infection without loss of antiviral activity. It is concluded that AZT that was coupled to the NCAs significantly contributes to the overall antiviral activity of the conjugates leading to complementary effects. These results highlight the potential of using NCA-AZTMP as dual-targeting preparations against HIV-1 in which both the carrier and the coupled drug contribute to the therapeutic efficacy acting at a different level in the replication cycle.
Assuntos
Ácido Aconítico/análogos & derivados , Fármacos Anti-HIV/farmacologia , Albumina Sérica/farmacologia , Nucleotídeos de Timina/farmacologia , Zidovudina/análogos & derivados , Ácido Aconítico/farmacologia , Anticoagulantes , Técnicas de Cultura de Células , Didesoxinucleotídeos , Combinação de Medicamentos , Estabilidade de Medicamentos , Sinergismo Farmacológico , Células Gigantes/efeitos dos fármacos , Humanos , Albumina Sérica Humana , Eletricidade Estática , Linfócitos T/efeitos dos fármacos , Fatores de Tempo , Zidovudina/farmacologiaRESUMO
A series of new polyanions was synthesized via gamma-polymerization, in aqueous micellar solution, of omega-unsaturated anionic surfactants whose polar head was derived from amino acids or dipeptides. The obtained polyanions were evaluated for their activity against human immunodeficiency virus (HIV-1, HIV-2) and various other RNA and DNA viruses. All the test compounds proved active against HIV-1 and HIV-2, their 50% inhibitory concentration (IC50) being in the range of 0.04-7.5 micrograms/mL, while they were not toxic to the host cells (CEM-4 or MT-4) at concentrations up to 100 micrograms/mL or higher. The HIV-inhibitory effect increased with the hydrophilic character of the amino acid moiety. The compounds were found to interact with both the viral envelope glycoprotein gp120 and the cellular CD4 receptor, thus blocking virus-cell binding and virus-induced syncytium formation. These polyanions also proved active against human cytomegalovirus at about the same IC50 as for HIV. In addition, they were also active, albeit at somewhat higher IC50 values (0.8-20 micrograms/mL), against other enveloped viruses such as respiratory syncytial virus and arenaviruses (Junin and Tacaribe). At yet higher IC50 values ( > or = 20 micrograms/mL), some of the compounds showed activity against influenza A virus. No activity was observed with any of the compounds against vesicular stomatitis virus, Sindbis virus, Semliki forest virus, influenza B, parainfluenza type 3, and the nonenveloped viruses Coxsackie type B4, polio type 1, and reovirus type 1.
Assuntos
Antivirais/síntese química , HIV/efeitos dos fármacos , Tensoativos/síntese química , Animais , Antivirais/farmacologia , Linhagem Celular , Cães , Humanos , Relação Estrutura-Atividade , Tensoativos/farmacologia , Replicação Viral/efeitos dos fármacosRESUMO
The bicyclams represent a new class of highly potent and selective HIV inhibitors. Time-of-addition experiments have previously shown that these compounds interfere with an early event in the viral replicative cycle. Additional experiments have now been carried out in order to investigate in more detail the mechanism of action of these promising compounds. As described in this paper, PCR experiments revealed that no viral DNA was formed following viral infection, thus confining the target(s) of action of the bicyclams to an early stage of HIV infection. An assay, using pseudotype virions containing the envelope of HIV-1 and the genome of a plaque-forming virus (Cocal Virus), pointed to viral entry as the main target of the bicyclams. HIV-1 strains resistant to two prototype bicyclams, JM2763 and SID791 (JM3100), were raised. Results obtained with SID791 with respect to syncytium formation induced by SID791-sensitive and -resistant HIV-1 strains and the cross-resistance observed for dextran sulfate, suggest inhibition of binding and/or fusion as a plausible target of SID791. Additional experiments enabled us to exclude SID791 and JM2763 as binding inhibitors and to conclude that bicyclams block the entry of cell-bound virus. Furthermore, a monoclonal antibody recognising the V3 loop of wild-type gp120 did not bind to this region in the two bicyclam-resistant strains. Our results point to gp120 as a possible target for the HIV-inhibitory effects of the bicyclams.
Assuntos
Antivirais/farmacologia , HIV-1/efeitos dos fármacos , Compostos Heterocíclicos/farmacologia , Anticorpos Monoclonais/imunologia , Sequência de Bases , Benzodiazepinas/farmacologia , Benzilaminas , Ciclamos , Primers do DNA , DNA Viral/biossíntese , DNA Viral/efeitos dos fármacos , Resistência Microbiana a Medicamentos , Células Gigantes , Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , HIV-1/genética , HIV-1/patogenicidade , Humanos , Imidazóis/farmacologia , Dados de Sequência Molecular , Fragmentos de Peptídeos/imunologia , Células Tumorais CultivadasRESUMO
Human herpesvirus 6 (HHV-6) was examined in vitro for its sensitivity to a broad range of nucleoside analogues, including acyclovir (ACV), ganciclovir (GCV), penciclovir (PCV), buciclovir (BCV), brivudin (BVDU), the N7-isomer of 6-deoxyganciclovir (S2242), foscarnet (phosphonoformic acid, PFA), and several acyclic nucleoside phosphonate (ANP) analogues such as (S)-HPMPA, (S)-HPMPC, PMEA and PMEDAP. Antiviral efficacy was monitored microscopically by the inhibitory effect of the compounds on HHV-6-induced cytopathic effect in human T-lymphoblastoid HSB-2 cells. In addition, a newly developed immunofluorescence/flow cytometric assay (FACS) was used to determine HHV-6-specific antigen expression. A close correlation was observed between the antiviral data obtained by the microscopic assay and the flow cytometric assay. Marked antiviral efficacy was noted for S2242, PFA and the ANP analogues (S)-HPMPA, (S)-HPMPC, (S)-cHPMPC, (S)-3-deaza-HPMPA, (S)-3-deaza-cHPMPA, (S)-HPMPG and (R)-HPMPG. Also, PMEA and PMEDAP proved highly active against HHV-6 infection, whereas (S)-FPMPA and (R)-PMPDAP were inactive. ACV was only slightly protective against HHV-6, and no activity was found for GCV, PCV, BCV and BVDU. Overall, the efficacy of the nucleoside analogues against HHV-6 appeared to correlate with their efficacy against human cytomegalovirus (HCMV).
Assuntos
Antivirais/farmacologia , Herpesvirus Humano 6/efeitos dos fármacos , Nucleosídeos/farmacologia , Organofosfonatos , Adenina/análogos & derivados , Adenina/farmacologia , Linhagem Celular , Cidofovir , Efeito Citopatogênico Viral , Citosina/análogos & derivados , Citosina/farmacologia , Citometria de Fluxo , Foscarnet/farmacologia , Ganciclovir/análogos & derivados , Ganciclovir/farmacologia , Humanos , Compostos Organofosforados/farmacologiaRESUMO
T30177, an oligonucleotide composed of only deoxyguanosine and thymidine, is 17 nucleotides in length and contains single phosphorothioate internucleoside linkages at its 5' and 3' ends for stability. This oligonucleotide does not share significant primary sequence homology with or possess any complementary (antisense) sequence motifs to the human immunodeficiency virus type 1 (HIV-1) genome. T30177 inhibited replication of multiple laboratory strains of HIV-1 in human T-cell lines, peripheral blood lymphocytes, and macrophages. T30177 was also found to be capable of inhibiting multiple clinical isolates of HIV-1 and preventing the cytopathic effect of HIV-1 in primary CD4+ T lymphocytes. In assays with human peripheral blood lymphocytes there was no observable toxicity associated with T30177 at the highest concentration tested (100 microM), while the median inhibitory concentration was determined to be in the range of 0.1 to 1.0 microM for the clinical isolates tested, resulting in a high therapeutic index for this drug. In temporal studies, the kinetics of addition of T30177 to infected cell cultures indicated that, like the known viral adsorption blocking agents dextran sulfate and Chicago sky blue, T30177 needed to be added to cells during or very soon after viral infection. However, analysis of nucleic acids extracted at 12 h postinfection from cells treated with T30177 at the time of virus infection established the presence of unintegrated viral cDNA, including circular proviral DNA, in the treated cells. In vitro analysis of viral enzymes revealed that T30177 was a potent inhibitor of HIV-1 integrase, reducing enzymatic activity by 50% at concentrations in the range of 0.050 to 0.09 microM. T30177 was also able to inhibit viral reverse transcriptase activity; however, the 50% inhibitory value obtained was in the range of 1 to 10 microM, depending on the template used in the enzymatic assay. No observable inhibition of viral protease was detected at the highest concentration of T30177 used (10 microM). In experiments in which T30177 was removed from infected cell cultures at 4 days post-HIV-1 infection, total suppression of virus production was observed for more than 27 days. PCR analysis of DNA extracted from cells treated in this fashion was unable to detect the presence of viral DNA 11 days after removal of the drug from the infected cell cultures. The ability of T30177 to inhibit both laboratory and clinical isolates of HIV-1 and the experimental data which suggest that T30177 represents a novel class of integrase inhibitors indicate that this compound is a viable candidate for evaluation as a therapeutic agent against HIV-1 in humans.
Assuntos
Antivirais/farmacologia , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , Oligonucleotídeos/farmacologia , Sequência de Bases , Fusão Celular/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , DNA Nucleotidiltransferases/antagonistas & inibidores , DNA Viral/biossíntese , Citometria de Fluxo , HIV-1/enzimologia , HIV-1/fisiologia , Humanos , Integrases , Linfócitos/efeitos dos fármacos , Linfócitos/virologia , Macrófagos/virologia , Dados de Sequência Molecular , Inibidores da Transcriptase Reversa/farmacologia , Subpopulações de Linfócitos T/virologia , Replicação Viral/efeitos dos fármacosRESUMO
The antiviral activity of water-soluble dextrans derivatized with varying percentages of carboxymethyl, benzylamide, and sulfonate groups was evaluated. Several of the polymers exhibited potent antiviral activity against a variety of enveloped viruses, but not against non-enveloped viruses, and only when present during virus adsorption. The mechanism of activity against retroviruses [i.e. human immunodeficiency virus (HIV)] and herpes viruses (i.e. human cytomegalovirus) could be ascribed to inhibition of virus binding to the cells. An absolute requirement for anti-HSV activity appeared to be a sufficiently high percentage of benzylamide and benzylamide sulfonate groups. This did not, however, apply for human cytomegalovirus, respiratory syncytial virus, and HIV. The sensitivity of the latter viruses appeared to be influenced by factors other than the global chemical composition, which leads us to assume that physical factors such as the distribution and sequence of the substituents on the sugar backbone play an important role in the antiviral activity of the derivatized dextrans.
Assuntos
Antivirais/síntese química , Dextranos/síntese química , Linhagem Celular , Citomegalovirus/efeitos dos fármacos , Dextranos/química , Dextranos/toxicidade , HIV/efeitos dos fármacos , Vírus Sinciciais Respiratórios/efeitos dos fármacos , Simplexvirus/efeitos dos fármacos , Relação Estrutura-Atividade , Vírus da Estomatite Vesicular Indiana/efeitos dos fármacos , Replicação Viral/efeitos dos fármacosRESUMO
Sixteen freshly isolated varicella-zoster virus (VZV) strains were evaluated in vitro, in parallel with two reference strains expressing a functional thymidine kinase (TK+) (Oka and YS) and two thymidine kinase-deficient mutants (TK-) (07-1 and YS-R), for their susceptibility to a broad range of antiviral compounds. The following compounds were included: acyclovir (ACV), brivudine (BVDU), sorivudine (BVaraU), other BVDU congeners such as BTDU, CTDU, CVDC and CVDU, ganciclovir (GCV), FIAC, araT, araA, araC, foscarnet (PFA), phosphonoacetic acid (PAA), the acyclic nucleoside phosphonates HPMPC, cHPMPC, HPMPA, cHPMPA, HPMPc3A, PMEA and PMEDAP, the N7-isomeric acyclic nucleoside analogue N7AP, penciclovir (PCV), compounds 882C87 and H2G and two oxetanocin derivatives OXT-A and OXT-G. Fourteen of the 16 clinical isolates displayed the following order of decreasing selectivity against VZV: BVaraU > BVDU > CVDU approximately CVDC > H2G > N7AP approximately CTDU approximately BTDU approximately OXT-G approximately 882C87 > ACV > FIAC approximately araT > HPMPC approximately cHPMPC approximately HPMPA approximately HPMPc3A approximately cHPMPA > PCV approximately GCV approximately OXT-A > PMEDAP approximately PMEA > PFA approximately PAA approximately araA > araC. Two VZV strains (isolated from the cerebrospinal fluid of an AIDS patient) that were shown to have a truncated TK were clearly resistant to all the compounds that need the viral TK for their phosphorylation, while sensitivity to the acyclic nucleoside phosphonates, PFA, PAA, OXT-A and araA, remained unchanged. A slight (5- and 10-fold) increase was noted in the 50% inhibitory concentration of N7AP and OXT-G, respectively, for the TK- VZV strains as compared to the TK+ VZV strains. Ganciclovir and FIAC also showed a marked decrease in their activity against these two strains, but this was not as pronounced as for the other viral TK-dependent drugs. From our results, it appears that although acyclic nucleoside phosphonates may not have as favourable a therapeutic index as drugs requiring the viral TK, they should be considered for the treatment of TK- VZV life-threatening infections that are resistant to the viral TK-dependent drugs.
Assuntos
Antivirais/farmacologia , Herpesvirus Humano 3/efeitos dos fármacos , Varicela/tratamento farmacológico , Citometria de Fluxo , Herpes Zoster/tratamento farmacológico , Herpesvirus Humano 3/isolamento & purificação , Humanos , Testes de Sensibilidade Microbiana , Valores de Referência , Sensibilidade e EspecificidadeRESUMO
Several new analogues of the novel anti-HIV agent cosalane have been synthesized and evaluated as inhibitors of HIV-1 integrase and protease, HIV-1 replication, HIV-1 and HIV-2 cytopathicity, HIV-1- and HIV-2-mediated syncytium formation, and cytopathicity of a variety of human pathogenic viruses. The congeners displayed enhanced potencies relative to cosalane itself as inhibitors of HIV-1 integrase and protease. The two most potent analogues against HIV-1 integrase displayed IC50 values of 2.2 microM, while the three most potent compounds against HIV-1 protease had IC50 values in the 0.35-0.39 microM range. In addition to its activity against HIV-1 and HIV-2 cytopathicity, cosalane inhibited the cytopathic effects of herpes simplex virus-1, herpes simplex virus-2, and human cytomegalovirus at concentrations that were well below the cytotoxic concentrations. Potentially useful antiviral activities were also revealed for some of the new cosalane congeners against influenza virus, Junin virus, and Tacaribe virus.
Assuntos
Antivirais/farmacologia , Ácido Aurintricarboxílico/análogos & derivados , DNA Nucleotidiltransferases/antagonistas & inibidores , Inibidores da Protease de HIV/farmacologia , HIV-1/efeitos dos fármacos , Sequência de Aminoácidos , Ácido Aurintricarboxílico/química , Ácido Aurintricarboxílico/farmacologia , Linhagem Celular , HIV-1/enzimologia , HIV-1/fisiologia , Herpesviridae/efeitos dos fármacos , Humanos , Integrases , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Replicação Viral/efeitos dos fármacosRESUMO
The antiviral activity of polysaccharide fractions obtained from water extracts of the red seaweed Nothogenia fastigiata was investigated. Fraction F6, corresponding to a sulphated xylomannan, was found to inhibit efficiently the replication of herpes simplex virus type 1 (HSV-1). Furthermore, F6 selectively inhibited the replication of several other enveloped viruses including herpes simplex virus type 2, human cytomegalovirus (HCMV), respiratory syncytial virus, influenza A and B virus, Junin and Tacaribe virus and simian immunodeficiency virus. F6 was only weakly active against human immunodeficiency virus type 1 and 2. The mode of action of F6 against HSV-1 and HCMV could be ascribed to an inhibitory effect on virus adsorption.
Assuntos
Antivirais/isolamento & purificação , Herpesvirus Humano 1/efeitos dos fármacos , Polissacarídeos/isolamento & purificação , Alga Marinha/química , Fracionamento Químico , Testes de Sensibilidade Microbiana , Replicação Viral/efeitos dos fármacosRESUMO
Varicella zoster virus (VZV) is recognized as one of the major viral pathogens reactivated in patients with the acquired immune deficiency syndrome (AIDS). We report the case of meningoradiculoneuritis in an AIDS patient,associated with the isolation in the cerebrospinal fluid (CSF) of a thymidine kinase (TK)-deficient, acyclovir (ACV)-resistant strain of VZV. Although the virus was sensitive in vitro to phosphonoformate (PFA), the patient did not improve during PFA therapy and finally died. Several VZV strains isolated from this patient (including two isolates from the patient's CSF) were analyzed for their TK activity and subsequently the viral TK gene was sequenced showing a major deletion leading to a truncated protein. Their susceptibility to several antiviral agents including ACV, PFA, (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU), 9-beta-D-arabinofuranosyladenine (vidarabine), (S)-1-(3-hydroxy-2-phosphonylmethoxypropyl) cytosine (HPMPC), and (S)-9-(3-hydroxy-2-phosphonyl-methoxypropyl)adenine (HPMPA) was evaluated. All the virus strains isolated from this patient remained sensitive to HPMPA and HPMPC, pointing to the potential usefulness of these acyclic nucleoside phosphonates for the treatment of ACV-resistant VZV infections in immunocompromised patients.
Assuntos
Síndrome da Imunodeficiência Adquirida/complicações , Aciclovir/uso terapêutico , Herpes Zoster/complicações , Meningite/etiologia , Neurite (Inflamação)/etiologia , Radiculopatia/etiologia , Adulto , Sequência de Aminoácidos , Sequência de Bases , Resistência a Medicamentos , Feminino , Herpes Zoster/tratamento farmacológico , Humanos , Dados de Sequência Molecular , Timidina Quinase/metabolismoAssuntos
Síndrome da Imunodeficiência Adquirida/complicações , Aciclovir/uso terapêutico , Infecções por Herpesviridae/tratamento farmacológico , Herpesvirus Humano 3/efeitos dos fármacos , Polirradiculoneuropatia/complicações , Resistência Microbiana a Medicamentos , Humanos , Meninges/patologia , Testes de Sensibilidade MicrobianaRESUMO
The drug Ro5-3335 [7-chloro-5-(2-pyrryl)-3H-1,4-benzodiazepin-2(H)-one] inhibits human immunodeficiency virus type 1 (HIV-1) gene expression at the transcriptional level through interference with Tat-mediated transactivation (M.-C. Hsu, A. D. Schutt, M. Holly, L. W. Slice, M. I. Sherman, D. D. Richman, M. J. Potash, and D. J. Volsky, Science 254:1799-1802, 1991). We confirmed this specific inhibitory effect in a quantitative bioassay based on transactivation of a chimeric gene comprising the HIV-1 long terminal repeat promoter fused to the lacZ gene of Escherichia coli and transfected in a HeLa cell line expressing Tat. Ro5-3335 was found to inhibit HIV-1 long terminal repeat-driven lacZ gene expression at a 50% inhibitory concentration of 0.5 microM. The in vitro anti-HIV-1 activity of Ro5-3335 was highly dependent on the nature of the host cells. The highest selectivity index, 50, was found in phytohemagglutinin-stimulated peripheral blood lymphocytes. The selectivity index was between 1 and 10 in the CD4+ T-cell lines CEM, MOLT-4 (clone 8), and HUT-78. In MT-4 and MT-2 cells, Ro5-3335 had no inhibitory effect on HIV-1 replication. The absence of anti-HIV-1 activity of Ro5-3335 in MT-4 cells was confirmed by using different parameters of virus replication and different multiplicities of infection. In persistently HIV-1-infected HUT-78/IIIB/LAI cells, Ro5-3335 failed to demonstrate any activity at subtoxic concentrations. The cytotoxicity of Ro5-3335 was significantly lower in peripheral blood lymphocytes than in the CD4+ T-cell lines.