Biomedical subjects
E Alteri
Publications and source records attributed to E Alteri.
CGP 53437, an orally bioavailable inhibitor of human immunodeficiency virus type 1 protease with potent antiviral activity.
CGP 53437 is a peptidomimetic inhibitor of human immunodeficiency virus type 1 (HIV-1) protease containing a hydroxyethylene isostere. The compound inhibited recombinant HIV-1 protease with a Ki of 0.2 nM. The inhibition constant versus human cathepsin D and human cathepsin E was 4 nM. Human pepsin and gastricsin were inhibited with Kis of 8 and 500 nM, respectively, and human renin was inhibited with a Ki of 190 microM. The replication of HIV-1/LAV, HIV-1/Z-84, and HIV-1/pLAI was inhibited with a 90% effective dose of 0.1 microM in acutely infected MT-2 cells. The 50% cytotoxic dose was 100 microM. Similar antiviral activity was observed when the compound was added up to 10 h after infection. At the effective concentration, processing of Gag precursor protein p55 was greatly reduced, confirming an action on the late stage of the virus life cycle, as expected. The efficacy of the inhibitor was also demonstrated by using primary human peripheral blood lymphocytes infected with the HIV-1/LAV strain, low-passage clinical isolates obtained from HIV-1-seropositive individuals (including a zidovudine-resistant strain), and HIV-2/ROD. In these cells, CGP 53437 delayed the onset of HIV replication in a dose-dependent fashion (substantial effects with concentrations of > or = 0.1 microM) as long as the inhibitor was maintained in the culture. CGP 53437 was orally bioavailable in mice. Concentrations in plasma 10-fold in excess of the in vitro antiviral 90% effective dose could be sustained for several hours after oral application of 120 mg/kg. Therefore, CGP 53437 has the potential to be a therapeutically useful anti-HIV agent for the treatment of AIDS.
The replicative restriction of lymphocytotropic isolates of HIV-1 in macrophages is overcome by TGF-beta.
In vitro exposure of human blood monocyte-derived macrophages to T-cell tropic human immunodeficiency virus (HIV) isolates fails to establish a productive viral infection. Several studies have shown that such preferential HIV-1 replication in T cells or in mononuclear phagocytes (HIV tropism) may be determined by distinct viral characteristics. In the present study it was demonstrated that transforming growth factor-beta (TGF-beta), a factor known to be produced by platelets, macrophages, and other cells present at a wound site, can act as a mediator in overcoming the lymphocytotropic restriction of several well-characterized viral isolates of HIV-1 (i.e., LAV, Z84, pLAI, NY5). Macrophages infected with these isolates show cytopathic changes comparable to those seen upon infection with the monocytotropic isolate ADA. To achieve this effect with TGF-beta, the factor must be present after the infection period. The emerging virus retains its original cellular tropism. Based on these observations the authors propose a role for TGF-beta in the establishment and progression of HIV infection and disease.
Lack of effect of desferrioxamine on in-vitro HIV-1 replication.
Explore the source record for details and available documents.
In vitro effect of transforming growth factor-beta on progression of HIV-1 infection in primary mononuclear phagocytes.
In vitro-differentiated monocytes can be infected with the monocytotropic isolate of HIV-1/ADA. The infection is characterized by formation of giant cells and production of virus that can be found in cell supernatants or cell-associated. In this study, we demonstrate that the above described parameters of infection can be enhanced by a factor present in acidified M phi supernatants, suggesting that it might be transforming growth factor beta-1 (TGF-beta 1). When recombinant or purified TGF-beta were examined, similar activities were detected. This effect apparently is not because of changes in the cellular phenotype that could favor infection. The effect of TGF-beta is exerted on cells once infection is established or on cells with active virus production. The activity can be also demonstrated using U-937 cells.
Use of human monocytes in the evaluation of antiviral drugs: quantitation of HSV-1 cytopathic effects.
An assay for the evaluation of antiviral and immunomodulator potency was developed using pure populations of cultured human monocytes. The assay involved culturing of human monocytes until they were fully susceptible (15-20 days) to lytic infection with HSV-1. When susceptible cells were cultured with recombinant interferon-alpha or a synthetic interferon inducer such as polyinosinic:polycytidylic acid prior to infection, a significant enhancement in resistance to the cytopathic effects of HSV-1 was observed. Likewise, a dose dependent reduction in cell lysis was observed when acyclovir was added immediately after virus infection. Monocyte resistance to HSV-1 was determined by the retention of pinocytic activity as determined by the uptake of neutral red dye. Relative pinocytic activity was quantitated using a simple colorimetric procedure. This antiviral assay can be completed in 48 h; is easy to perform, highly sensitive and reproducible.
The lipophilic muramyl peptide MTP-PE is a potent inhibitor of HIV replication in macrophages.
Cultured monocyte-derived macrophages were productively infected with human immunodeficiency virus in vitro. Treatment of these cells shortly after infection and several times thereafter with the free form of MTP-PE had an inhibitory effect on virus production. When the liposomal formulation of MTP-PE was used, higher levels of protection were achieved. The drug was not only effective when added to cells immediately after infection, but it also reduced virus production by cells with an established infection. When the liposomal formulation of MTP-PE was used only one treatment was required to achieve maximal effects. During these studies it was noted that the placebo liposomes had some effect in reducing the reverse transcriptase levels found in the supernatants of infected cells. This reduction could not be explained by direct cytotoxic effect. Both free and liposomal MTP-PE lipid significantly prevented formation of giant cells during the course of infection as well as reduced the cell associated viral antigen.
A derivative of staurosporine (CGP 41 251) shows selectivity for protein kinase C inhibition and in vitro anti-proliferative as well as in vivo anti-tumor activity.
Analogues of staurosporine were synthesized and their ability to inhibit protein kinases was examined. Staurosporine is a potent but non-selective inhibitor of in vitro protein kinase C(PKC) activity (IC50 6.0 nM). The derivative CGP 41 251 had reduced PKC activity with an IC50 of 50 nM but showed a high degree of selectivity when assayed for inhibition of cyclic AMP-dependent protein kinase (IC50 2.4 microM), S6 kinase (IC50 5.0 microM) and tyrosine-kinase-specific activity of epidermal growth factor receptor (IC50 3.0 microM). Staurosporine and CGP 41 251 exerted growth inhibition in the human bladder carcinoma line T-24, human promyelocytic leukemia line HL-60 and bovine corneal endothelial cells at concentrations which correlated well with in vitro PKC inhibition. In addition, both compounds inhibited the release of H2O2 from human monocytes pre-treated with 12-O-tetradecanoyl-phorbol-13-acetate at non-toxic concentrations. In vivo anti-tumor activity was examined in T-24 human bladder carcinoma xenografts in athymic nude mice. Tumor growth inhibition tests revealed significant anti-tumor activity (2p less than 0.001) at 1/10 of the maximum tolerated doses for both compounds. By contrast, a closely related derivative of staurosporine (CGP 42 700) was inactive at concentrations of over 100 microM in all in vitro enzyme and anti-proliferative assays as well as in animal tumor models. Our data suggest an association between PKC inhibition and anti-proliferative and anti-tumor activity.
An assay for the detection of interleukin-1 synthesis inhibitors: effects of antirheumatic drugs.
The monokines interleukin-1 beta and alpha (IL-1) play a central role in the connective tissue destruction of many chronic inflammatory diseases. A high capacity screening assay for the detection of inhibitors of IL-1 biosynthesis has been established. Normal human monocytes were obtained by leukapheresis and elutriation. IL-1 beta and alpha biosynthesis was stimulated with LPS, and cell-associated and secreted IL-1 beta and IL-1 alpha were measured by specific immunoassays (ELISA). The mean total IL-1 beta (cell-associated and secreted) production in 18 different donors was 11 ng/10(6) cells (range 1.2-28.8). Secreted IL-1 beta represented 31 to 86% of the total IL-1 beta. More IL-1 alpha than IL-1 beta was produced but, unlike IL-1 beta, IL-1 alpha was poorly secreted. The steroids prednisolone and dexamethasone, gold (sodium aurothiomalate) and chloroquine were potent inhibitors of the IL-1 production. Mean IC50 values of 180 nM (range 2.5 nM-1 microns), 10 microM (range 6-20 microM) and of 75 microM were found for prednisolone, gold and chloroquine, respectively. Above 5 microM, the non-steroidal anti-inflammatory compounds indomethacin and BW755C increased IL-1 beta biosynthesis. Nordihydroguaiaretic acid inhibited the level of the secreted form of IL-1 beta, but tended to increase the cell-associated level. D-Penicillamine (up to 6 mM), cyclosporin A (up to 1 microM) and methotrexate (up to 12 microM) inhibited neither cell-associated nor secreted IL-1 beta levels. This high capacity assay, which is insensitive to classical NSAIDs, may serve in the detection and characterization of new classes of anti-inflammatory compounds.
In vitro interactions of serotonin (5-HT) with mononuclear cells from migraine patients: alterations related to the phase of the attack.
Serotonin (5-HT) binds in a saturable and specific manner to high affinity binding sites present on the surface of human mononuclear cells. The serotonin binding to mononuclear cells has been assayed in patients with migraine, during and outside the crisis period. In these patients, unlike the episodic cluster headache patients that show a constant trend, the results are variable. This variability may depend on the fluctuation of 5-HT levels in the blood, during and after the crisis. A possible explanation for these observations is that at the beginning of the crisis the binding of 5-HT is still similar to the normal level. As the crisis goes on, there is at first an increase in 5-HT binding that saturates all the available sites and a subsequent phenomenon of desensitization, with a loss of high affinity sites. Further investigations are needed in order to assess whether or not the mononuclear cell functions are affected by the modifications of the binding of 5-HT, thought to be a regulatory molecule.
Defect of serotonin binding to mononuclear cells from episodic cluster headache patients.
Explore the source record for details and available documents.
N-formylmethionyl-leucyl-[3H]phenylalanine binding, superoxide release, and chemotactic responses of human blood monocytes that repopulate the circulation during leukapheresis.
Human blood monocytes comprise two subpopulations: one migrates to the chemoattractant, N-formylmethionyl-leucyl-phenylalanine (fMet-Leu-Phe), and has saturable binding sites for this peptide; the other does not migrate and exhibits little peptide binding. To determine if expression of binding sites was a function of monocyte maturation, we depleted human subjects of blood monocytes by leukapheresis so that the circulation was repopulated by monocytes released from the bone marrow. Pre- and postleukapheresis monocytes were then compared for fMet-Leu-[3H]Phe binding, superoxide generation, and chemotactic responses. No significant differences in peptide binding curves were found, suggesting that receptor expression was stable over the maturational span represented by these two groups of cells. This supports the hypothesis that there are two distinct lineages of monocytes with respect to expression of receptors for fMet-Leu-Phe. An additional finding of interest was that the number of chemotactically responsive cells immediately postleukapheresis was half the control. This was a transient state; monocyte responses were normal 3 hr after termination of leukapheresis, suggesting that they rapidly become functionally mature.
Differences in superoxide production by nonmigrating and migrating human monocyte subpopulations.
Explore the source record for details and available documents.
TGF-beta: upregulator of HIV replication in macrophages.
TGF-beta at physiological concentrations, when added to monocyte-derived macrophages following HIV1 infection, has an enhancing effect upon the rate of virus production. This effect is observed with the monocytotropic isolate ADA, as well as with HIV1 IIIB, which poorly replicates in macrophages.