RESUMEN
When N,N'-didansyl cystine binds to the cell membranes of vertebrate rods and cones its fluorescence efficiency increases about 20-fold. The entire outer segments of living cones become brilliantly fluorescent. Stained live rods, as well as most freshly detached outer segments, are only weakly fluorescent, but they become brightly fluorescent within a few seconds if their plasma membranes are osmotically ruptured. The difference in staining of rod and cones suggests that disk membranes of rods are not continuous with the plasma membranes are osmotically ruptured. The difference in staining of rod and cones plasma membrane on outer segments of photoreceptors in electrophysiological and biochemical experiments, and to study the infolding pattern of rod and cone disks.
Asunto(s)
Colorantes Fluorescentes , Células Fotorreceptoras/citología , Animales , Membrana Celular/metabolismo , Cistina/metabolismo , Compuestos de Dansilo/metabolismo , Carpa Dorada , Microscopía Electrónica , Microscopía Fluorescente , Ósmosis , Rana pipiens , Ratas , Coloración y Etiquetado/métodosRESUMEN
Ribonucleic acids from the bacteriophage R17 and from R17 amber mutant AmB2 have been digested with ribonuclease T1. Of the products isolated, only one was different. It codes for the first six amino acid residues of the viral coat protein. The probable base sequence of the wild-type oligonucleotide is CpUpUpCpUpApApCpUpUpUpApCpUpCpApGp.
Asunto(s)
Colifagos , Código Genético , Proteínas Virales/análisis , Secuencia de Aminoácidos , Cromatografía , ARN Viral/análisis , Ribonucleasas/metabolismoRESUMEN
Indolicidin is a tridecapeptide amide isolated from the cytoplasmic granules of bovine neutrophils. It has potent, broad spectrum microbicidal activities in vitro that are thought to be related to the membrane-disruptive properties of the peptide. Based on the putative membrane-targeted mode of action, we postulated that indolicidin would be active against HIV-1, an enveloped virus. Indolicidin was reproducibly virucidal against HIV-1 at a concentration of 333 microg/mL (174 microM) with a 50% inhibitory dose between 67 and 100 microg/mL. At 37 degrees C, killing was rapid with >50% killing of HIV occurring within 5 min, and nearly 100% viral inactivation achieved by 60 min. The anti-HIV activity of indolicidin was temperature-sensitive, a finding consistent with a membrane-mediated antiviral mechanism. Parallel experiments revealed that indolicidin lysed cultured lymphoblastoid cells at concentrations similar to those required for antiviral activity. However, a des-R13-amide indolicidin analog (R12-OH), previously shown to have less antibacterial activity than indolicidin, was significantly less active against HIV and was non-toxic to lymphoid target cells at concentrations up to 333 microg/mL, the highest level tested.
Asunto(s)
Fármacos Anti-VIH/farmacología , Péptidos Catiónicos Antimicrobianos , Infecciones por VIH/tratamiento farmacológico , Neutrófilos/fisiología , Péptidos/farmacología , Animales , Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/toxicidad , Linfocitos T CD4-Positivos/efectos de los fármacos , Bovinos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Humanos , Péptidos/síntesis química , Péptidos/toxicidad , Relación Estructura-Actividad , TemperaturaRESUMEN
Over the past year significant progress has been made in mapping those regions of the HIV-1 envelope glycoproteins involved in virus neutralization and virus enhancement. These functional, antigenic domains of the gp160 are illustrated in Fig. 1. The role of neutralization in vaccine development is still unresolved, although high-titer antibody to the V3 loop of HIV-1 appears to be correlated with the ability to prevent HIV-1 infection by the homologous strain in chimpanzees. Therefore, the mechanism of type-specific neutralization of HIV appears to be clearly defined. The problem of group-specific neutralization of HIV-1 is still a mystery. Nevertheless, the finding that secondary structure is important for the generation of group-specific neutralizing mAbs and that carbohydrate is an important determinant for group-specific neutralization suggests that non-linear determinants are important. Answers to the question of group-specific neutralization should be available in the next few years. The role of HIV-1-enhancing antibodies in pathogenesis is not well understood. Nevertheless, the ability of patient serum to only enhance the patient's own isolate and not neutralize that isolate suggests that enhancing antibodies are important. Furthermore, the findings that enhancing antibody titers increase in SIV infection and peak immediately prior to the death of the animal suggests that such antibodies may play a role in SIV pathogenesis. With the identification and domain mapping of enhancing hu-mAbs, it should be possible to evaluate directly the role of enhancing domains in HIV and SIV pathogenesis by challenging animals in the presence of pure enhancing antibody. Only when these experiments are performed will it be possible to evaluate what role, if any, enhancing antibodies play in HIV pathogenesis. The above questions, when answered, are likely to provide important insights into lentivirus pathogenesis and help researchers to produce a safe and effective anti-HIV vaccine.
Asunto(s)
Anticuerpos Anti-VIH/inmunología , Infecciones por VIH/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Secuencia de Aminoácidos , Animales , Humanos , Datos de Secuencia Molecular , Pruebas de Neutralización , Receptores de Complemento 3d/inmunología , Receptores Fc/inmunologíaRESUMEN
OBJECTIVE: The purpose of this study was to develop and characterize human monoclonal antibodies (HuMAb) that neutralize HIV-1. DESIGN: Based upon previous studies involving the generation of HuMAb that neutralize other enveloped viruses, we thought it feasible to generate HuMAb that might neutralize HIV-1. METHODS: A HuMAb was generated by fusing splenic B-cells from an HIV-positive patient with a mouse myeloma cell line. Flow cytometry was used to determine surface reactivity of the HuMAb on HIV-infected and non-infected cells. Radioimmunoprecipitation was employed to elucidate the antigen recognized by the HuMAb. A cell survival assay was used to determine the ability of the HuMAb to neutralize divergent isolates of HIV-1 in the presence or absence of complement. A gp120-CD4 inhibition enzyme-linked immunosorbent assay (ELISA) was developed in order to initiate studies to determine the mechanism of neutralization by the HuMAb. RESULTS: An anti-HIV HuMAb was generated that neutralized two HIV-1 isolates (IIIB and MN) without complement and which neutralized one divergent isolate (RF) and one clinical isolate in the presence of complement. This HuMAb, designated S1-1, was found, by flow cytometric analysis, to react with the surface of HIV-1-infected but not with uninfected cells. Radioimmunoprecipitation analysis demonstrated that S1-1 binds to native HIV gp120, but not dithiothreitol (DTT)-treated gp120. In addition, HuMAb S1-1 did not bind to denatured HIV antigens in Western blot analysis. HuMAb S1-1 effectively inhibited the binding of gp120 to soluble CD4 in ELISA. CONCLUSIONS: These results suggest that the epitope recognized by S1-1 is conformational and conserved among diverse HIV-1 isolates and may represent an uncharacterized HIV neutralizing domain within or close to the CD4 binding domain on gp120. HuMAb S1-1 might have a role to play in vaccine development or passive immunotherapy.
Asunto(s)
Anticuerpos Monoclonales/inmunología , VIH-1/inmunología , Pruebas de Neutralización , Anticuerpos Monoclonales/metabolismo , Western Blotting , Antígenos CD4/metabolismo , Línea Celular , Citometría de Flujo , Anticuerpos Anti-VIH/inmunología , Proteína gp120 de Envoltorio del VIH/inmunología , Proteína gp120 de Envoltorio del VIH/metabolismo , Humanos , Ensayo de Radioinmunoprecipitación , SolubilidadRESUMEN
OBJECTIVE: To demonstrate that antibodies against amino acids (aa) 603-622 of the SIV gp41 transmembrane glycoprotein enhance infection of SIV in vivo. DESIGN: A synthetic peptide derived from aa 603-622 of SIVmac251 gp41 was synthesized and tested for immunogenicity in rabbits and SIV-infected rhesus macaques. Next, SIV-naive animals were immunized with either a recombinant vaccinia virus expressing the SIV gp160 envelope glycoprotein (VVrgp160) and boosted three times with aa 603-622 (group 1, four animals), wild-type vaccinia virus and boosted with aa 603-622 (group 2, two animals), or VVrgp160 followed by three doses of an irrelevant peptide (group 3, two animals). Animals were challenged with SIVmac251. RESULTS: Peptide aa 603-622 was immunogenic in rabbits. SIV-infected rhesus monkeys immunized with the peptide developed two-three log increases in antibodies to this peptide and antibodies that could enhance SIV infection in vitro. SIV-naive rhesus macaques in group 1 had higher levels of antibody to the peptide by enzyme-linked immunosorbent assay and higher levels of enhancing antibodies at the time of SIV challenge than the animals in groups 2 or 3. Following challenge with SIVmac251 the group 1 animals had detectable p27 antigen longer than animals in group 2 and 3 and died of simian AIDS before the respective animals in the two control groups (P < 0.05 by log-rank test). CONCLUSIONS: aa 603-622 of SIV gp41, like aa 579-613 of HIV gp41, can stimulate production of antibodies that enhance SIV and HIV infection in vitro. Furthermore, immunization with this peptide suppressed beneficial effects of a gp160 vaccine and appeared to enhance SIV infection in vivo.
Asunto(s)
Anticuerpos Antivirales/inmunología , Productos del Gen env/inmunología , Glicoproteínas de Membrana/inmunología , Proteínas de los Retroviridae/inmunología , Vacunas contra el SIDAS/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Virus de la Inmunodeficiencia de los Simios/inmunología , Proteínas del Envoltorio Viral/inmunología , Animales , Relación Dosis-Respuesta Inmunológica , Macaca mulatta , ConejosRESUMEN
Antibody-dependent enhancement (ADE) of human immunodeficiency virus type 1 (HIV-1) infection in vitro has been described recently and was shown to occur by two mechanisms: either participation of the alternative pathway of complement or to involve an Fc receptor-mediated, complement-independent mechanism. Complement-mediated ADE results in an accelerated cytopathic effect in target cells that can abrogate the protective properties of neutralizing antibodies. This study characterizes the surface antigens of MT-2 cells using flow cytometric analysis and shows that these cells express high levels of both CD4 and complement receptor type 2 (CR2) while several CD4+ cell lines that do not demonstrate complement-mediated ADE lack high levels of complement receptors. Further, utilizing MT-2 cell cultures, it is demonstrated that complement-mediated ADE of HIV-1 infection is conferred by the sera from more than 80% of HIV-1 antibody-positive individuals (N = 85). Complement-mediated ADE of HIV-1 infection causes an acceleration of several parameters indicative of HIV-1 infection in vitro including increased HIV-1 antigen synthesis as detected by indirect immunofluorescence, RNA accumulation as measured by a solution hybridization protocol, reverse transcriptase release, and progeny virus production.
Asunto(s)
Efecto Citopatogénico Viral , Anticuerpos Anti-VIH/inmunología , VIH-1/fisiología , ARN Viral/biosíntesis , Receptores de Complemento/fisiología , Línea Celular , Citometría de Flujo , Antígenos VIH/inmunología , VIH-1/inmunología , VIH-1/metabolismo , Humanos , Sondas ARN , ADN Polimerasa Dirigida por ARN/biosíntesis , Receptores de Complemento/análisis , Replicación ViralRESUMEN
The dicaffeoylquinic acids (DCQAs) and dicaffeoyltartaric acids (DCTAs) are potent and selective inhibitors of human immunodeficiency virus type 1 (HIV-1) integrase. They also inhibit HIV-1 replication at nontoxic concentrations. Since integrase is an excellent target for anti-HIV therapy, structure-activity relationships were employed to synthesize compounds with: (1) improved potency against HIV-1 integrase, (2) improved anti-HIV effect in tissue culture, and (3) increased selectivity as indicated by low cellular toxicity. Thirty-four analogues of the DCTAs and DCQAs were synthesized and tested for cell toxicity, anti-HIV activity, and inhibition of HIV-1 integrase. Seventeen of the 34 analogues had potent activity against HIV-1 integrase ranging from 0. 07 to >10 microM. Seventeen analogues that were synthesized or purchased had no inhibitory activity against integrase at concentrations of 25 microM. Of the biologically active analogues, 7 of the 17 inhibited HIV replication at nontoxic concentrations. The most potent compounds were D-chicoric acid, meso-chicoric acid, bis(3,4-dihydroxydihydrocinnamoyl)-L-tartaric acid, digalloyl-L-tartaric acid, bis(3,4-dihydroxybenzoyl)-L-tartaric acid, dicaffeoylglyceric acid, and bis(3, 4-dihydroxyphenylacetyl)-L-tartaric acid. Anti-HIV activity of the active compounds in tissue culture ranged from 35 to 0.66 microM. Structure-activity relationships demonstrated that biscatechol moieties were absolutely required for inhibition of integrase, while at least one free carboxyl group was required for anti-HIV activity. These data demonstrate that analogues of the DCTAs and the DCQAs can be synthesized which have improved activity against HIV integrase.
Asunto(s)
Fármacos Anti-VIH/farmacología , Ácidos Cafeicos , Ácido Clorogénico/análogos & derivados , Ácido Clorogénico/farmacología , Inhibidores de Integrasa VIH/farmacología , Succinatos , Tartratos/farmacología , Replicación Viral/efectos de los fármacos , Fármacos Anti-VIH/química , Secuencia de Bases , Línea Celular , Supervivencia Celular/efectos de los fármacos , Ácido Clorogénico/química , Clonación Molecular , Cartilla de ADN , Farmacorresistencia Microbiana/genética , Inhibidores de Integrasa VIH/química , VIH-1/efectos de los fármacos , VIH-1/genética , VIH-1/fisiología , Humanos , Espectroscopía de Resonancia Magnética , Relación Estructura-Actividad , Tartratos/químicaRESUMEN
Multiple drug effect analyses with mismatched double-stranded RNA (mismatched dsRNA or Ampligen) as a core drug were performed to identify other agents and mechanisms through which mismatched dsRNA may potentiate effective therapeutic intervention in human immunodeficiency virus (HIV) infection. Antiviral activities were defined by a microtiter infection assay utilizing MT-2 cells as targets and HTLV-III-B produced in H9 cells as a virus source. The scope of agents tested included rIFN-alpha A, rIFN-beta Ser 17, and rIFN-gamma as cytokines; azidothymidine and phosphonoformate (Foscarnet) as inhibitors of reverse transcription; ribavirin as a putative inhibitor of proper HIV mRNA capping; amphotericin B as a lipophile; and castanospermine as a glycoprotein processing (glucosidase I) inhibitor. Separately, each drug demonstrated dose-dependent anti-HIV activity and, when used in combination with mismatched dsRNA, demonstrated synergism. Although mismatched dsRNA was synergistic with all three IFNs for anti-HIV activity in microtiter infection assays, it did not potentiate the transient inhibition of virus production observed for IFN in cultures of H9/HTLV-III-B cells. The results of these studies suggest that the pleiotropic activities of dsRNAs differ from those of IFN and may provide synergism in combination therapy with a wide range of antiviral drugs for the treatment of the acquired immunodeficiency syndrome (AIDS).
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/tratamiento farmacológico , Antivirales/farmacología , VIH/efectos de los fármacos , Indolizinas , Interferón beta , Interferones/farmacología , ARN Bicatenario/farmacología , Alcaloides/farmacología , Anfotericina B/farmacología , Antivirales/uso terapéutico , Quimioterapia Combinada , Foscarnet , VIH/crecimiento & desarrollo , Humanos , Técnicas In Vitro , Interferón Tipo I/farmacología , Interferón beta-1a , Interferon beta-1b , Interferón gamma/farmacología , Pruebas de Sensibilidad Microbiana , Ácido Fosfonoacético/análogos & derivados , Ácido Fosfonoacético/farmacología , Proteínas Recombinantes/farmacología , Ribavirina/farmacología , Zidovudina/farmacologíaRESUMEN
The Human Immunodeficiency Virus (HIV), the causative agent of AIDS, is thought to bind to T4+ (CD4+) target cells through the heavily glycosylated gp120 envelope glycoprotein. Plant lectins bind glycoproteins through noncovalent interaction with specific hexose residues; therefore, lectins were evaluated for their ability to inactivate HIV in vitro. The mannose-specific lectins concanavalin-A and succinyl concanavalin-A completely inactivated HIV while lentil lectin, wheat germ agglutinin, and phytohemagglutinin-P substantially inactivated HIV. BS-II, Vicia villosa (hairy vetch), and Ptilota plumosa (red marine algae) failed to alter the infectibility of HIV. Neither simple stearic hindrance, viral aggregation, nor lectin-cell interactions served to explain this phenomenon. Glycoprotein glycosylation was evaluated by differential lectin binding as well as molecular weight changes in gp120 when virus was produced in the presence of swainsonine, a glycosylation inhibitor. Lentil lectin bound gp120 better than concanavalin-A, suggesting the majority of glycosylation sites are fucosylated. The apparent molecular weight of gp120 was reduced by swainsonine, although HIV infectivity and concanavalin-A inactivation were retained. Thus, at least some N-glycosylation sites are complex-type glycoproteins but regions external to the (GlcNAc)2(Man)3 "core" pentasaccharide region are not required for HIV infectivity. It appears that the site or sites involved are nonfucosylated, high mannose and/or biantennary, nonsialylated, N-glycosylated regions of gp120 or gp41. Alternatively, they may be in close approximation to such carbohydrate regions.
Asunto(s)
VIH/patogenicidad , Lectinas/farmacología , Manosa/fisiología , Alcaloides/farmacología , Sitios de Unión , Glicosilación , VIH/efectos de los fármacos , Proteína gp120 de Envoltorio del VIH , Proteína gp41 de Envoltorio del VIH , Proteínas de los Retroviridae/metabolismo , Swainsonina , Proteínas del Envoltorio Viral/metabolismoRESUMEN
Antibodies that are useful in the treatment of HIV infection should result in virus neutralization or lysis of infected cells but should not enhance infection. In this study, the potential clinical use of 20 HIV-1-specific human monoclonal antibodies (HuMAbs) was determined by measuring their enhancing (C-ADE) activities using HIVLAI as the target virus. Two HuMAbs mediated both C-ADE and ADCC, two exclusively neutralized, and five exclusively mediated ADCC. Ten HuMAbs demonstrated no activity in any of the three assays. Three antibodies that neutralized HIVLAI were tested against HIVSF2; all three also neutralized HIVSF2. Four of five HuMAbs mediating ADCC against HIVLAI that were also tested against HIVSF2 had ADCC activity against HIVSF2. These results demonstrate that many HuMAbs have unique functions, allowing the separation of potentially beneficial and harmful activities. Combinations of HuMAbs with ADCC and neutralizing functions may have therapeutic utility.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Anticuerpos Anti-VIH/farmacología , VIH-1/inmunología , Anticuerpos Monoclonales/uso terapéutico , Especificidad de Anticuerpos , Citotoxicidad Celular Dependiente de Anticuerpos , Línea Celular , Proteínas del Sistema Complemento/metabolismo , Anticuerpos Anti-VIH/uso terapéutico , Infecciones por VIH/inmunología , Infecciones por VIH/terapia , Humanos , Pruebas de NeutralizaciónRESUMEN
Previously it was established that L1210 mouse leukemia and a variety of other tumor cell types produced a soluble factor(s), designated tumor-derived recognition factor (TDRF), which synergized with interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) or lipopolysaccharide (LPS) to promote increased tumor necrosis factor-alpha (TNF-alpha) and nitric oxide synthase (NOS) mRNA synthesis by murine macrophages (M phi). Other work revealed that pretreatment of L1210 tumor targets with IFN-gamma rendered them more susceptible to NO-mediated killing by LPS-activated M phi. Now we have combined these observations to determine if pretreatment of L1210 or P815 tumor targets with IFN-gamma and/or IL-2 would augment production of TDRF to enhance M phi activation. Results confirmed that pretreatment of either L1210 or P815 targets with 200 u/ml of IFN-gamma or 1,000 u/ml of IL-2 significantly increased their susceptibility to M phi-mediated cytotoxicity owing to increased NO production. Similar pretreatment of L1210 targets with suboptimal concentrations of IFN-gamma and IL-2 in combination resulted in additive rather than synergistic augmentation of NO-mediated cytotoxicity by cytotoxic M phi. Pretreatment of L1210 targets with IFN-gamma or IL-2 alone or in combination increased the production of TDRF above constitutive levels as demonstrated by increased production of NO and induction of NOS mRNA expression by cytotoxic M phi. Thus IFN-gamma and/or IL-2 promoted increased TDRF production by tumor targets which in turn promoted M phi generation of tumor cytotoxic NO. It appears that M phi activating cytokines have a dual role in acting on certain tumor targets to augment the process of M phi activation through the increased elicitation of immunopotentiating tumor-derived soluble factor(s).
Asunto(s)
Interferón gamma/farmacología , Interleucina-2/farmacología , Leucemia L1210/inmunología , Leucemia Experimental/inmunología , Activación de Macrófagos , Animales , Citocinas/inmunología , Citotoxicidad Inmunológica , Inmunidad Celular , Masculino , Ratones , Ratones Endogámicos C3H , Proteínas de Neoplasias/inmunología , Óxido Nítrico/fisiologíaRESUMEN
Cytolytic activity of human mononuclear peripheral blood leukocytes from healthy donors, cultured in interleukin-2 conditioned medium, was abrogated by in vitro infection with the lymphadenopathy associated virus (LAV) isolate of the human immunodeficiency virus (HIV). Although viral antigens are not expressed in cultured cells until 14 days postinfection, cytolytic activity was lost as early as 3 days after infection. Loss of cytolytic function was not a result of the release of suppressive factors from either infected cells or uninfected CEM cells since supernatants from neither infected cultures nor CEM cell cultures had any inhibitory effects on the function of uninfected cells. Cultured lymphocytes expressing Leu 11b were also shown to express HIV antigens via immunofluorescence after 14 days in culture. These results suggest that natural killer (NK) cells, as defined by expression of Leu 11b, were infected by HIV in vitro and the loss of lytic function was likely a direct consequence of that infection.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/patología , VIH/fisiología , Células Asesinas Naturales/metabolismo , Síndrome de Inmunodeficiencia Adquirida/fisiopatología , Línea Celular , Citotoxicidad Inmunológica , Técnica del Anticuerpo Fluorescente , Humanos , Células Asesinas Naturales/fisiología , Leucocitos Mononucleares/fisiología , Factores de TiempoRESUMEN
Combinations of anti-human immunodeficiency virus (HIV) drugs, including reverse transcriptase inhibitors and protease inhibitors, have proven immensely potent in the therapy of acquired immune deficiency syndrome (AIDS). To determine whether HIV integrase is a suitable target for combination therapy, the ability of an HIV integrase inhibitor, L-chicoric acid, to work in combination with a protease inhibitor and Zidovudine was tested in vitro. The addition of L-chicoric acid to either Zidovudine or protease inhibitor improved upon the observed anti-HIV activity of either compound alone. When all three drugs were combined, the anti-HIV activity was substantially better than either of the three compounds alone or any combination of two inhibitors. Doses of both Zidovudine and protease inhibitor could be reduced by more than 33% for an equivalent anti-HIV effect if L-chicoric acid was added. The improved anti-HIV activity was observed with a tissue culture adapted strain of HIV (HIV(LAI)) and with limited passage clinical isolates of HIV (HIV(R19) and HIV(R45)). These data demonstrate that a first generation HIV integrase inhibitor, L-chicoric acid, is at least additive in combination with existing multi-drug regimens and suggest that HIV integrase will be an excellent target for combination therapy of HIV infection.
Asunto(s)
Fármacos Anti-VIH/farmacología , Ácidos Cafeicos , Inhibidores de Integrasa VIH/farmacología , Inhibidores de la Proteasa del VIH/farmacología , VIH-1/efectos de los fármacos , Succinatos/farmacología , Zidovudina/farmacología , Sinergismo Farmacológico , Inhibidores de la Transcriptasa Inversa/farmacologíaRESUMEN
Combinations of anti-HIV agents including one or two reverse transcriptase inhibitors with a protease inhibitor are potent and effective. However, toxicities, costs and the emergence of drug-resistant organisms have compromised their long-term efficacy in people. A next, likely, target for anti-HIV therapy is HIV-1 integrase. Viral integration, catalyzed by integrase, is absolutely required for HIV replication. L-chicoric acid is a potent and selective inhibitor of HIV-1 integrase that also inhibits HIV-1 replication in cell culture. As a first step in understanding the potential role for integrase inhibitors in clinical medicine, the activities of L-chicoric acid alone and in combination with 2', 3'-dideoxycytidine, zidovudine, and a protease inhibitor, nelfinavir, were tested in vitro against molecular clones of HIV-1 resistant to reverse transcriptase inhibitors. L-chicoric acid was equally effective against a wild-type clone of HIV-1, HIV(NL4-3), or against HIV-1 resistant to either zidovudine or dideoxycytidine. L-chicoric acid was largely synergistic with zidovudine and synergistic with both dideoxycytidine and nelfinavir.
Asunto(s)
Fármacos Anti-VIH/farmacología , Ácidos Cafeicos , Inhibidores de Integrasa VIH/farmacología , VIH-1/efectos de los fármacos , Inhibidores de la Transcriptasa Inversa/farmacología , Succinatos/farmacología , Clonación Molecular , Farmacorresistencia Microbiana , Sinergismo Farmacológico , Quimioterapia Combinada , Inhibidores de la Proteasa del VIH/farmacología , VIH-1/genética , HumanosRESUMEN
Human immunodeficiency virus type 1 (HIV-1) infectivity and cytopathic effect require proper maturation of the viral envelope glycoprotein carbohydrate moieties. We have found that fresh human serum enhances the infectivity of HIV-1 in MT-2 cell infection assays when virus is synthesized in the presence of the mannosidase I inhibitor, 1-deoxymannojirimycin, or the mannosidase II inhibitor, swainsonine, but has no enhancing effect on virus synthesized in the presence of the glucosidase I inhibitors, castanospermine and 1-deoxynojirimycin, or the glucosidase II inhibitor, bromoconduritol. Enhanced infections were characterized by cytopathic effect, antigen synthesis and reverse transcriptase release, all which occurred sooner than in control-infected cultures. This enhancement of infection was also observed in C1q-deficient serum but was not observed in serum that was heat-inactivated or depleted of complement components C3 or factor B, thus suggesting a requirement for the alternate pathway of complement.
Asunto(s)
Alcaloides/farmacología , Proteínas del Sistema Complemento/inmunología , VIH-1/patogenicidad , Manosidasas/antagonistas & inhibidores , 1-Desoxinojirimicina , Células Cultivadas , Complemento C3/inmunología , Factor B del Complemento/inmunología , Efecto Citopatogénico Viral/efectos de los fármacos , Técnica del Anticuerpo Fluorescente , Glucosamina/análogos & derivados , Glucosamina/farmacología , Glucosidasas/antagonistas & inhibidores , Glicosilación , VIH-1/efectos de los fármacos , VIH-1/inmunología , VIH-1/aislamiento & purificación , Humanos , ADN Polimerasa Dirigida por ARN/metabolismo , SwainsoninaRESUMEN
Mismatched double-stranded RNA of the form r(I)n.r(C12-U)n (Ampligen) has been shown to be active against human immunodeficiency virus type 1 (HIV-1) using CEM and C3 cells as targets for infection by the highly similar HIV-1 isolates HTLV-IIIB and LAV (Montefiori, D.C. and Mitchell, W.M., 1987, Proc. Natl. Acad. Sci. U.S.A., 84, 2985-2989). The scope of Ampligen's anti-HIV-1 activity was examined in this study using the genetically divergent HIV-1 isolate HTLV-IIIRF, two additional target T-cell lines, H9 and MT-2, and a monocyte/macrophage cell line, U937. As judged by indirect immunofluorescence, reverse transcriptase activity and vital dye uptake, Ampligen was active against HTLV-IIIRF in H9, MT-2, C3 and U937 cells in addition to being active against HTLV-IIIB in U937 cells. A minimum of 1 h preincubation of cells (MT-2) with Ampligen was required for maximum activity. These results suggest that Ampligen's potential clinical efficacy may not be limited by either the highly variable nature or host cell range of HIV-1.
Asunto(s)
Antivirales/farmacología , VIH/efectos de los fármacos , Poli I-C , Poli U , Polirribonucleótidos/farmacología , ARN Bicatenario/farmacología , Línea Celular , Variación Genética , VIH/genética , Humanos , Macrófagos , Monocitos , Linfocitos T Colaboradores-InductoresRESUMEN
Although highly active anti-retroviral therapy (HAART) is successful in the treatment of HIV infection, problems with toxicity, drug-resistant variants, and therapeutic failures have compromised the long-term utility of existing combination regimens. Mismatched double-stranded RNA (polyI-polyC(12)U) is an immune modulator with inherent anti-HIV activity. Cell toxicities and anti-HIV activities of fourteen anti-HIV agents were determined alone and in combination with polyI-polyC(12)U. Combination analyses for anti-HIV activity were performed at three drug ratios. Using Mixed Dose Effect analyses and the CalcuSyn for Windows software package, combination indeces were determined for all drug combinations. In general, polyI-polyC(12)U was synergistic in combination with abacavir, zidovudine, zalcitabine, didanosine, stavudine, efavirenz, indinavir, ritonavir, nelfinavir, and amprenavir. It was synergistic to antagonistic with lamivudine, delavirdine, nevirapine, and saquinavir. Thus, polyI-polyC(12)U is synergistic with most anti-HIV agents at most drug ratios and across most effective concentrations in vitro, although, certain members of each class were exceptions. PolyI-polyC(12)U alone was equally active against wild-type HIV and HIV resistant to nevirapine, protease inhibitors, or nucleoside analogue reverse transcriptase inhibitors. These results suggest that polyI-polyC(12)U should be re-evaluated as a potential adjunct therapy in patients who have failed current anti-retroviral therapeutic regimens.
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Fármacos Anti-VIH/farmacología , VIH-1/efectos de los fármacos , Poli I-C/farmacología , Poli U/farmacología , ARN Bicatenario/farmacología , Animales , Fármacos Anti-VIH/toxicidad , Disparidad de Par Base , Línea Celular , Relación Dosis-Respuesta a Droga , Farmacorresistencia Microbiana , Sinergismo Farmacológico , Inhibidores de la Proteasa del VIH/farmacología , Inhibidores de la Proteasa del VIH/toxicidad , Humanos , Técnicas In Vitro , Pruebas de Sensibilidad Microbiana , Poli I-C/toxicidad , Poli U/toxicidad , ARN Bicatenario/toxicidad , Inhibidores de la Transcriptasa Inversa/farmacología , Inhibidores de la Transcriptasa Inversa/toxicidadRESUMEN
The antiseptics 4% chlorhexidine gluconate detergent formulation containing 4% isopropyl alcohol (Hibiclens/Hibiscrub) and 0.5% chlorhexidine gluconate in 70% isopropyl alcohol with emollients (Hibistat/Hibisol) efficiently inactivated human immunodeficiency virus (HIV) produced in cell culture within 15 seconds. These antiseptics were completely effective at 1:100 and 1:5 dilutions, respectively. Therefore, use of these products according to the manufacturer's instructions (i.e. undiluted) for routine disinfection of hands following contact with HIV-contaminated materials, as well as immediate disinfection of abrasions or cuts exposed to HIV, should have significant protective effects.
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1-Propanol/farmacología , Antisepsia/métodos , Clorhexidina/farmacología , Detergentes/farmacología , VIH-1/efectos de los fármacos , Tensoactivos/farmacología , Antisepsia/normas , Línea Celular Transformada , VIH-1/patogenicidad , Virus Linfotrópico T Tipo 2 Humano , Humanos , Linfocitos TRESUMEN
Fits to the vanadium K-edge X-ray absorption spectra (XAS) of five whole blood cell samples from the tunicate Phallusia nigra revealed unprecedented forms of intracellular vanadium. Endogenous vanadium was divided between the V(III) ion (74.2+/-5.1% of total V) and the vanadyl ion [V(IV)=O](2+) (25.2+/-5.4% of total V). The V(III) fraction included both [V(H(2)O)(6)](3+) (36.7+/-5.5%) modeled as VCl(3) in 1 M HCl, and three previously unprecedented chelated V(III) forms (37.5+/-4.6%). Two of these could be represented by the model ligand environments V(acetylacetonate)(3) (17.9+/-3.2%) and K(3)V(catecholate)(3) (13.1+/-4.7%), implying DOPA-like complexation. The third chelated form was represented by the 7-coordinate N(2)O(5) complex Na[V(edta)(H(2)O)] (8.0+/-1.8%). This coordination array, suggestive of a novel mononuclear V(III) protein site, contributed only to fits to samples 1, 2, 3 and 5, which were prepared in the presence of DTT. Endogenous V(IV) (25.2+/-5.4%) was principally modeled as VOCl(2) in 1 M HCl. EPR spectra (averages: A(parallel)=(1.842+/-0.006)x10(-2) cm(-1); A( perpendicular)=(0.718+/-0.007)x10(-2) cm(-1); g(parallel)=1.936+/-0.002; g( perpendicular)=1.990+/-0.001) confirmed the predominance of the aquated vanadyl ion. Blood cell sample five uniquely required the XAS spectrum of VOSO(4) in 0.1 M H(2)SO(4) solution (13.0%) and of [OV(V)(pivalate)(3)] (3.1%) to successfully fit the XAS pre-edge energy region. This endogenous V(V) signal is also unprecedented. These results are compared with those of analogous fits to the blood cells of Ascidia ceratodes and may support assignment of P. nigra to a different genus.