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D C Montefiori

Publications and source records attributed to D C Montefiori.

126 records · Page 7Linked to original sources
PubMed1989

In vitro evaluation of mismatched double-stranded RNA (ampligen) for combination therapy in the treatment of acquired immunodeficiency syndrome.

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).

Acquired Immunodeficiency Syndrome↗
PubMed1989

Complement-mediated, antibody-dependent enhancement of HIV-1 infection in vitro is characterized by increased protein and RNA syntheses and infectious virus release.

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.

Cell Line↗
PubMed1988

Antibody-dependent enhancement of human immunodeficiency virus type 1 infection.

Two components of human serum enhance human immunodeficiency virus type 1 (HIV-1) infection and mask HIV-1 neutralising antibody activity. The first is heat-stable, unique to HIV-1 seropositive sera, and is removed by protein-A chromatography. The second is heat-labile and ubiquitous; it is found in normal serum and is removed by heating at 60 degrees C for 1 h or by treatment with cobra venom anticomplementary protein. Additionally, complement component C3 deficient serum lacks the labile activity although Clq deficient serum contains the labile factor. The data suggest that the two components are antibody and the alternative pathway of complement fixation. The mechanism of action does not involve an increase in either complement-mediated cytolysis or syncytium formation. The activity has been identified in 11 of 16 patients tested to date.

Antibodies, Viral↗
PubMed1988

Natural killer cell infection and inactivation in vitro by the human immunodeficiency virus.

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.

Acquired Immunodeficiency Syndrome↗
PubMed1988

Role of protein N-glycosylation in pathogenesis of human immunodeficiency virus type 1.

Human immunodeficiency virus type 1 (HIV-1), the retrovirus responsible for acquired immunodeficiency syndrome (AIDS), contains two heavily glycosylated envelope proteins, gp120 and gp41, which mediate attachment of virions to glycosylated cell surface receptor molecules (CD4 antigens) and appear to be responsible for syncytium formation and associated cytopathic effects of this virus. A comprehensive study of the effects of N-linked glycoprotein processing inhibitors on HIV-1 replication, infectivity, cytopathicity, target-cell infectibility, syncytium formation, and gp120 electrophoretic mobility was conducted to assess the importance of protein glycosylation in the pathogenesis of HIV-1 in vitro. The electrophoretic mobility of gp120 was decreased when gp120 was synthesized in the presence of castanospermine or 1-deoxynojirimycin (inhibitors of glucosidase I), increased when gp120 was synthesized in the presence of 1-deoxymannojirimycin (mannosidase I) or swainsonine (mannosidase II), and unaffected when gp120 was synthesized in the presence of bromoconduritol (glucosidase II). Inhibition by tunicamycin (lipid-linked oligosaccharide precursor synthesis), castanospermine, 1-deoxynojirimycin, and 1-deoxymannojirimycin attenuated HIV-1 infectivity and blocked HIV-1-induced syncytium formation and cytopathicity, whereas bromoconduritol and swainsonine failed to have such effects. None of the inhibitors interfered with virus replication in acutely infected cells or affected the ability of target cells to form syncytia with untreated HIV-1-infected cells. These results demonstrate that protein N-glycosylation is critical to the pathogenesis of HIV-1 at the levels of viral infectivity and cytopathicity but not at the level of virus replication or of host-cell infectibility.

Acquired Immunodeficiency Syndrome↗
PubMed1988

Evaluation of antiviral drugs and neutralizing antibodies to human immunodeficiency virus by a rapid and sensitive microtiter infection assay.

A 96-well microtiter infection assay for the human immunodeficiency virus (HIV) is described. The assay utilizes human T-cell lymphotropic virus type I-immortalized MT-2 cells as targets for infection and requires only 4 to 5 days for completion. Cytolysis was quantitated by vital dye uptake of poly-L-lysine-adhered cells as an endpoint for infection. The assay's efficacy was proven by the sensitive and accurate assessment of several known anti-HIV agents including two inhibitors of reverse transcription (3'-azido-3'-deoxythymidine and 2',3'-dideoxycytidine), three biological response modifiers (recombinant interferons alpha and beta and mismatched double-stranded RNA), a direct inactivator of HIV virions (amphotericin B), and neutralizing antibodies from two HIV-positive human subjects. Evaluation of data was facilitated by computer-assisted analysis. This assay provides a means for rapid, sensitive, and inexpensive large-scale in vitro testing of potential anti-HIV therapeutic regimens and quantitation of HIV-neutralizing antibody titers.

Antibodies, Viral↗
PubMed1987

A human immunodeficiency virus type 1 (HIV-1) infection-enhancing factor in seropositive sera.

While testing sera for Human Immunodeficiency Virus neutralizing antibody titers, three sera were identified which had the ability to enhance infectivity of the virus. The sera were from three different individuals residing in Nashville, TN. The enhancing factor was not removed by either filtration through 0.05 micron filters or by incubation for one hour with a stoichiometric amount of protein A sepharose. Two of the sera were able to enhance infection by two divergent isolates (HTLV-IIIB and HTLV-IIIRF) while one was only capable of enhancing infection of target cells by HTLV-IIIB. None of the sera induced syncytium formation in chronic HIV-infected cells. The findings suggest that the substance is neither a virus nor an IgG class 1 or 2.

Antibodies, Viral↗
PubMed1987

Mismatched double-stranded RNA (ampligen) reduces concentration of zidovudine (azidothymidine) required for in-vitro inhibition of human immunodeficiency virus.

'Ampligen', a non-toxic, mismatched polymer of double-stranded RNA with antiviral and immunomodulatory activities reduced the concentration of zidovudine (azidothymidine, AZT; 'Retrovir', Wellcome) required for inhibitory activity against human immunodeficiency virus (HIV) in vitro. At the higher doses of AZT tested, the virustatic activity observed seemed to have a synergistic virustatic relation with ampligen. Thus, combined therapy with ampligen and AZT can be expected to be more beneficial than AZT alone to patients with the acquired immunodeficiency syndrome (AIDS) or AIDS-related complex since AZT regimens that seem to be clinically effective are associated with considerable toxicity.

Antiviral Agents↗
PubMed1987

Persistent coinfection of T lymphocytes with HTLV-II and HIV and the role of syncytium formation in HIV-induced cytopathic effect.

We previously demonstrated a high permissiveness of HTLV-II-transformed T lymphocytes (C3) to human immunodeficiency virus (HIV) infection in vitro, and that this infection results in the lysis of cells (D.C. Montefiori and W.M. Mitchell (1986) Virology 155, 726-731). We now show that a small percentage of HIV-infected C3 cells resist cell lysis, grow continuously in culture, and express antigens of both viruses. High levels of reverse transcriptase activity found in the culture fluid of these coinfected cells were associated with the presence of fully infectious HIV and an absence of detectable infectious HTLV-II. Virus production in C3 cells coinfected with the HIV isolate HTLV-III was approximately threefold greater than in C3 cells coinfected with the HIV isolate LAV, a result which suggests that HIV genomic diversity may give rise to differences in replicative capacities. Lysis resistance was found to be a cellular-determined function in that HIV produced in cultures of C3/HTLV-III cells retained the capacity to elicit a lytic response upon repeated infection. Small syncytia (less than five nuclei) were rarely observed in cultures of C3 and nonlytic C3/HIV cells, whereas large syncytia (more than five nuclei) were in abundance during the lytic phase of coinfection, a result which supports a role for syncytium formation in the mechanism of HIV-induced cytopathic effects. The results of these studies further demonstrate that there exists a lack of viral interference by HTLV-II infection, and that HTLV-II-transformed lymphocytes could act as a chronic reservoir for HIV in vivo.

Antigens, Viral↗
PubMed1987

Antiviral activity of mismatched double-stranded RNA against human immunodeficiency virus in vitro.

The biological response modifier r(I)n.r(C12-U)n, referred to here as mismatched double-stranded (ds) RNA, was examined for antihuman immunodeficiency virus (HIV) activity in vitro because of its known antiviral activity and ability to induce interferon (IFN) in other biological systems [Carter, W. A., Strayer, D. R., Hubbell, H. R. & Brodsky, I. (1985) J. Biol. Response Modif. 4, 495-502]. We found that cultures of the highly HIV-permissive T-cell line C3 were afforded significant protection from HIV infection when incubated in growth media supplemented with mismatched dsRNA at 10-50 micrograms/ml prior to virus challenge. Similar results were obtained at 50 micrograms of mismatched dsRNA per ml in cultures of the T-lymphoblastoid cell line CEM. Infections were monitored by indirect immunofluorescence of cells for viral p24 antigen expression, reverse transcriptase activity in culture fluids for virus production, and vital dye uptake for cytopathic effect. Antiviral activity was increased by the continued presence of mismatched dsRNA in cultures following virus challenge. A one-time exposure to mismatched dsRNA (50 micrograms/ml) provided greater antiviral activity than either a one-time exposure to recombinant IFN-alpha [250 international units (IU)/ml], IFN-beta (250 IU/ml), or IFN-gamma (50 IU/ml) in cultures of CEM cells, or a one-time exposure to a combination of all three IFNs (150 IU each per ml) in cultures of C3 cells. Mismatched dsRNA at 50 micrograms/ml had no effect on cell division, RNA and protein synthesis, or virus replication in all T-cell lines examined. A clear distinction between the activities of mismatched dsRNA and IFN was the ability of IFN to suppress the in vitro replication of HIV that occurred at IFN concentrations (150 IU each of alpha, beta, and gamma per ml) that provided less antiviral activity than mismatched dsRNA (50 micrograms/ml). The results of these in vitro studies suggest a potential therapeutic value for mismatched dsRNA in the treatment of acquired immunodeficiency syndrome (AIDS).

Antiviral Agents↗
PubMed1987

Evidence that mannosyl residues are involved in human immunodeficiency virus type 1 (HIV-1) pathogenesis.

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.

Alkaloids↗
PubMed1986

Infection of the HTLV-II-bearing T-cell line C3 with HTLV-III/LAV is highly permissive and lytic.

The recent discovery that HTLV-I infected T-cells have an enhanced susceptibility to HTLV-III/LAV infection with concurrent cell lysis (S. Harada, Y. Koyanagi, and N. Yamamoto (1985) Science, 229, 563-566) led us to investigate a possible role for HTLV-II infected T cells in this respect. HTLV-II-bearing C3 cells were exposed to HTLV-III/LAV and the subsequent infection monitored by indirect immunofluorescence (IIF) of cells, reverse transcriptase (RT) activity in culture fluids, and hemacytometer cell counts. HTLV-III/LAV antigen expression was detected within 2 days whereas RT activity was detected within 3 days postinfection. The presence of immunofluorescent positive cells and RT activity was accompanied by cell lysis. Double infection of C3 cells was confirmed by IIF using a goat polyclonal antibody directed against the core p24 protein for detecting HTLV-II, and high titer serum from a pre-AIDS patient for detecting HTLV-III/LAV. We conclude that HTLV-I and HTLV-II infected T-cells share an enhanced susceptibility to HTLV-III/LAV infection with the promotion of cell lysis.

Antigens, Viral↗
PubMed1981

Regulation of cell division and of tyrosinase in B16 melanoma cells by imidazole: a possible role for the concept of metabolite gene regulation in mammalian cells.

Results of hemacytometer cell counts and of tyrosinase measurements made by the Pomerantz method demonstrate that imidazole added to the medium of cultured B16 mouse melanoma cells can stimulate tyrosinase specific activity and inhibit cell division. These effects are greater than with adenosine 3',5' cyclic monophosphate (cAMP) or the cAMP-phosphodiesterase inhibitor theophylline. The effects of imidazole on cell division and tyrosinase are enhanced by theophylline and antagonized by cAMP. Cyclic AMP-phosphodiesterase activity in cell-free extracts can be inhibited by theophylline and stimulated by imidazole. However, imidazole does not affect cAMP-phosphodiesterase specific activity in vivo, nor does it affect intracellular cAMP concentrations as determined by competitive protein-binding assays. In contrast, the specific activity of cAMP-phosphodiesterase in vivo is stimulated by cAMP and theophylline, supporting the hypothesis that cAMP and agents which increase intracellular cAMP concentrations induce the synthesis of cAMP-phosphodiesterase. Studies with actinomycin-D and cycloheximide support the hypothesis that cAMP can also mediate posttranslational activation of tyrosinase. Similar experiments suggest that imidazole, or a derivative thereof, can induce the synthesis of tyrosinase at the pretranslational level of control. We hypothesize that this type of regulation (pretranslational) by imidazole may define a role for the concept of "Metabolite Gene Regulation" (MGR), in mammalian cells.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1980

Metabolite gene regulation of the L-arabinose operon in Escherichia coli with indoleacetic acid and other indole derivatives.

The ability of indole derivatives to facilitate RNA polymerase transcription of the L-arabinose operon in Escherichia coli was shown to require the catabolite activator protein (CAP) as well as the araC gene product. Adenosine 3',5'-monophosphate (cAMP) was not obligatory for araBAD transcription when the cells were grown in the presence of 1 mM indole-3-acetic acid or in the presence of indole-3-acetamide, indole-3-propionic acid, indole-3-butyric acid, or 5-hydroxyindole-3-acetic acid. However, these indole derivatives were unable to circumvent the cAMP requirement for the induction of the lactose and the maltose operons. Catabolic repression occurred when glucose was added to cells grown in the presence of L-arabinose and 1 mM indoleacetic acid or 1 mM cAMP. This effect was reversed at higher concentrations of indoleacetic acid or cAMP. The induction and the catabolite repression phenomena were quantitated by measuring the differential rate of synthesis of L-arabinose isomerase (the araA gene product). These results indicated that indole metabolites from various living systems may regulate gene expression and may be involved in "metabolite gene regulation."

Arabinose↗
PubMed1980

Metabolite gene regulation: imidazole and imidazole derivatives which circumvent cyclic adenosine 3',5'-monophosphate in induction of the Escherichia coli L-arabinose operon.

Imidazole, histidine, histamine, histidinol phosphate, urocanic acid, or imidazolepropionic acid were shown to induce the L-arabinose operon in the absence of cyclic adenosine 3',5'-monophosphate. Induction was quantitated by measuring the increased differential rate of synthesis of L-arabinose isomerase in Escherichia coli strains which carried a deletion of the adenyl cyclase gene. The crp gene product (cyclic adenosine 3',5'-monophosphate receptor protein) and the araC gene product (P2) were essential for induction of the L-arabinose operon by imidazole and its derivatives. These compounds were unable to circumvent the cyclic adenosine 3',5'-monophosphate in the induction of the lactose or the maltose operons. The L-arabinose regulon was catabolite repressed upon the addition of glucose to a strain carrying an adenyl cyclase deletion growing in the presence of L-arabinose with imidazole. These results demonstrated that several imidazole derivatives may be involved in metabolite gene regulation (23).

Aldose-Ketose Isomerases↗
PubMedDate not supplied

Mismatched dsRNA (ampligen) induces protection against genomic variants of the human immunodeficiency virus type 1 (HIV-1) in a multiplicity of target cells.

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.

Antiviral Agents↗
PubMedDate not supplied

Immunization against SIVmne in macaques using multigenic DNA vaccines.

All structural and regulatory genes of SIVmne were cloned into mammalian expression vectors to optimize expression in vitro and immunogenicity in mice. Macaca fascicularis were immunized four times with plasmid DNA (n = 4), or two DNA priming inoculations followed by two boosts of recombinant gp160 plus Gag-Pol particles (n = 4). Following intrarectal challenge with SIVmne, all macaques became infected. Three monkeys immunized with DNA alone maintained low plasma virus loads by 1 year post-challenge; the fourth exhibited high virus loads and significant CD4+ cell decline. Two of the DNA plus boost and three control macaques had high virus loads and associated CD4+ cell decline. Both vaccine protocols elicited antibodies and comparable helper T-cell proliferative responses to gp160. Cytokine mRNA levels in activated peripheral blood mononuclear cells (PBMC) taken at time of challenge suggested a dominant T helper (Th) 1 state in three DNA-immunized and one protein-boosted macaque, which correlated with low virus loads and high CD4+ cell counts post-challenge.

AIDS Vaccines↗