Aldar S. Bourinbaiar

Anti-HIV and anti-tumor activities of recombinant MAP30 from bitter melon.

MAP30 is an anti-HIV plant protein that we have identified and purified to homogeneity from bitter melon (Momordica charantia). It is capable of acting against multiple stages of the viral life cycle, on acute infection as well as replication in chronically infected cells. In addition to antiviral action, MAP30 also possesses anti-tumor activity, topological inactivation of viral DNA, inhibition of viral integrase and cell-free ribosome-inactivation activities. We have cloned and expressed the MAP30 gene. The objective of this study is to characterize recombinant MAP30 (re-MAP30) and to determine its anti-HIV, anti-tumor and other activities. We report here that re-MAP30 inhibits HIV-1 and certain human tumors to the same extent as its native counterpart, natural MAP30 (nMAP30). The anti-HIV activity was measured by quantitative focal syncytium formation on CEM-ss cell monolayers, viral core protein p24 expression and viral-associated reverse transcriptase activity in HIV-1-infected H9 cells. The anti-tumor activity was measured by metabolic labeling of protein synthesis in tumor cells. In the dose range of the assay, re-MAP30 exhibits little toxicity to the uninfected viral target cells and other normal human cells. Identical to nMAP30, re-MAP30 is also active in topological inactivation of viral DNA, inhibition of viral DNA integration and cell-free ribosome inactivation. The cloning and expression of the gene encoding biologically active re-MAP30 provides an abundant source of homogeneous material for clinical investigations, as well as structure-function studies of this novel antiviral and anti-tumor agent.

Comparative in vitro study of contraceptive agents with anti-HIV activity: Gramicidin, nonoxynol-9, and gossypol

Gramicidin, a polypeptide antibiotic derived from Bacillus brevis, was compared in vitro with the established contraceptive virucidal agents nonoxynol-9 and gossypol for activity against human immunodeficiency virus (HIV) infection. The effective antiviral 10 ng/ml concentration of gramicidin required for complete HIV inactivation was a thousand-fold lower than the dose observed for nonoxynol-9 or gossypol. Gramicidin, routinely used as a contraceptive agent in the former Soviet Union, should be considered for in vivo trials as a spermicide with potent antiviral activity.

Anti-HIV effect of beta subunit of human chorionic gonadotropin (βhCG) in vitro

Abstract Human chorionic gonadotropin (hCG)—a pregnancy-associated immunomodulating hormone—has been recently shown in vitro to suppress reverse transcriptase activity in chronically HIV-infected lymphocytes and monocytes and to block viral transmission resulting from cell-cell contact between virus-carrying lymphocytes and placental trophoblasts. In further pursuit of the query into the mechanism of action, purified α and β subunits of hCG were tested for the inhibition of p24 gag protein synthesis in virus-producing ACH-2 lymphocytes and U1 monocytes. Unlike the α subunit, β-hCG displayed a distinct U-shaped dose response, characteristic of the effect of dimer hCG. Maximum inhibition of viral expression has been achieved at 10–100 ng/ml, the concentration corresponding to blood levels of β-hCG in pregnant women. The doses that were several logs higher of normal levels seemed to increase viral production in monocytes. The data presented supports our original observations regarding the effect of intact hCG on HIV replication. While the mechanism of action remains to be established, the results suggest that the virus-interfering activity of hCG is determined by hormone-specific β chain but not by the α subunit—shared with the family of glycoprotein hormones from the pituitary—follicle-stimulating hormone, luteinizing hormone and thyrotropin.

Pregnancy hormones, estrogen and progesterone, prevent HIV‐1 synthesis in monocytes but not in lymphocytes

Increase in levels of estrogen and progesterone during pregnancy may affect intra‐uterine HIV‐1 injection through their effect on maternal immunocompetent cells. These hormones were examined for containing HIV‐1 production from ACH‐2 lymphocytes and U1 monocytes. Neither of the hormones has an effect on ACH‐2, but with U1, the physiological concentrations (0.1 μg‐0.1 ng) of progesterone and estrogen demonstrate significant inhibition of HIV‐1 release. Except for the highest dose of 1 μg/ml, the dose—response to progesterone and estrogen was not correlated with the negative influence on proliferation of both types of cells. The results suggest that in vivo low doses of female steroids may display specific antiviral activity in monocytes but not in lymphocytes.

Anti-HIV effect of gramicidin in vitro: potential for spermicide use.

Gramicidin, cation channel forming ionophore with antibacterial properties, was studied in vitro for inhibition of human immunodeficiency virus (HIV) infection of MT-4 lymphocytes. Effective antiviral concentrations required for complete HIV inactivation were three orders of magnitude lower than 10 micrograms/ml cytotoxic dose. Gramicidin, routinely used as a contraceptive agent, should be considered for clinical application as a spermicide with antiviral activity.

Inhibitory effect of human chorionic gonadotropin (hCG) on HIV-1 transmission from lymphocytes to trophoblasts

It has been demonstrated that human chorionic gonadotropin (hCG) inhibits HIV production in vitro, suggesting that this soluble placental glycoprotein can control viral replication and spread in vivo, hCG ‐ the major product of fetal trophoblasts ‐ was tested on an in vitro model consisting of choriocarcinoma‐derived ENAMI trophoblasts exposed to HIV‐infected MOLT‐4 lymphocytes. The results show a U‐shaped antiviral dose‐ effect and suggest that hCG may contribute to protection against intrauterine transmission of HIV‐1.

The non-steroidal anti-inflammatory drug, indomethacin, as an inhibitor of HIV replication

Indomethacin, a common non‐steroidal anti‐inflammatory drug (NSAID), has been used to treat rheumatoid arthritis. Although indomethacin has also been used as an immunopotentiator and symptomatic NSAID in AIDS, its effect on HIV replication is unknown. MT‐4 lymphocytes were inoculated with HIV in the presence of indomethacin and tested for p24 expression by ELISA. The 50% inhibition (IC50) was 10 μM, corresponding to plasma levels after administration of 50 mg oral indomethacin. The antiviral effect appears to be specific since no toxicity has been observed at the IC50 dose, and unrelated NSAIDs have not shown the activity at clinical doses. Indomethacin may, thus, represent a new class of anti‐HIV drug.

Effect of human chorionic gonadotropin (hCG) on reverse transcriptase activity in HIV-1 infected lymphocytes and monocytes

The majority of infants born to HIV-positive mothers are not infected in utero, suggesting that the pregnancy factors produced by fetal trophoblasts may provide protection against HIV-1 infection. Except for steroid female hormones, little is known of other pregnancy factors that may regulate either resistance or susceptibility to HIV-1. Human chorionic gonadotropin (hCG)--the major glycoprotein produced by the placental trophoblast throughout the pregnancy--was tested on reverse transcriptase activity in HIV-infected ACH-2 lymphocytes and U1 monocytes. The results suggest that low non-cytotoxic doses of hCG (0.01-1.0 IU range) may inhibit viral replication in maternal blood cells.

Acrosin inhibitor, 4′-acetamidophenyl 4-guanidinobenzoate, and experimental vaginal contraceptive with anti-HIV activity

Serine proteases are involved in a wide variety of seemingly unrelated physiological functions including capacitation of the spermatozoa and potentiation of human immunodeficiency virus (HIV) infection. The experimental vaginal contraceptives derived from 4-guanidinobenzoic acid act through inhibition of acrosin--a serine protease from the sperm. The serial ten-fold dilutions of 4-acetamidophenyl 4-guanidinobenzoate (AGB) were tested in vitro for the effect against HIV infection by assaying the suppression of de novo p24 synthesis in virus-inoculated MT-4 T lymphocytes. The results reveal that complete inhibition of HIV occurred at 100 micrograms/ml--a dose corresponding to previously reported concentrations responsible for preventing fertilization in rabbits. These findings suggest that serine protease inhibitors and in particular the guanidinobenzoates, reported to be up to 100-fold more potent and less irritating than nonoxynol-9, can be potentially operative against sexual transmission of HIV.

Synergistic effect of gramicidin and EDTA in inhibiting sperm motility and cervical mucus penetration in vitro

Gramicidin, a linear polypeptide with antiviral and antimicrobial properties, was compared in vitro with a commonly used spermicidal detergent-nonoxynol-9 (N9). The inhibition of sperm functions was evaluated by computer-assisted semen analysis (CASA) for sperm motility, in cervical mucus penetration assay, and by colorimetric tetrazolium salt and lactate dehydrogenase release assays routinely employed for testing the toxicity of drugs. The effective 100% inhibitory concentration (IC100) of gramicidin in a 2-min sperm immobilization assay by CASA was equal to 4 micrograms/ml, whereas IC100 of N9 was equal to 200 micrograms/ml. The presence of 0.1% of chelating agent, EDTA, reduced IC100 of gramicidin to 10 ng/ml, while less than a twofold enhancement in N9 activity was observed upon combination with EDTA. Likewise, the gramicidin/EDTA combination was 100,000 times more potent than N9/EDTA in the sperm penetration assay. Quantitative toxicity tests confirmed that gramicidin is a potent spermostatic rather than spermicidal agent. Further development of a gramicidin/EDTA formulation is warranted as a nontoxic topical contraceptive with activity against viral and microbial sexually transmitted diseases (STDs).