Inhibit HIV-1 Protease Activity Research Articles

Nanopore detects γ-radiation inhibited HIV-1 protease activity

Inhibition of HIV-1 protease (PR) activity is realized by exposure to 60Co γ-radiation. The radiation effects on enzyme kinetics of HIV-1 PR are subsequently monitored using nanopore sensor. Substantial loss of proteolytic efficiency towards GagPol polypeptide is observed due to the radiation treatment. Results shows ~50% of GagPol polypeptide was not involved in HIV-1 PR proteolysis by exposure to ultra-low intensity of γ-radiation (0.1K Gy), and the values reach to over 90% with high γ-ray treatment. Besides, the inactivation effect is also verified in blood samples which contain the virus protease. Our finding provides the potential benefits of γ-radiation to inactivate viral proteinic function, and might be a complementary to the designation of HIV-1 PR inhibitors.

Inhibition of HIV-1 Protease by Carpobrotus edulis (L.).

Carpobrotus edulis (L.) is a plant commonly found in the Eastern Cape Province of South Africa and is used for the general treatment of infections relating to the human immunodeficiency virus (HIV). HIV-1 protease plays an important role during HIV replication and maturation to its infectious form, and therefore inhibition of the enzyme is one of the main focus areas in drug development. The inhibitory effect of a water extract of C. edulis leaves against HIV-1 protease activity was determined using the SensoLyte® 520 HIV-1 protease assay fluorimetric kit and employing a HiLyte Fluor™488/QXL™520 fluorescence resonance energy transfer (FRET) peptide. Cytotoxicity of the extract towards HeLa Chang cell lines was determined using an in vitro MTT assay, and the phytochemical profile of the extract was determined with FT-IR and LC-MS. HIV-1 protease activity was inhibited 83.06% (IC50 1.6 mg/ml) (p < 0.0001) by the pepstatin A inhibitor control. Treatment with all C. edulis extract concentrations (16, 1.6, 0.16, and 0.016 mg/ml) inhibited HIV-1 protease activity significantly (p < 0.0001) in a typical dose response manner. With regards to cytotoxicity, the negative controls containing untreated HeLa Chang cells exhibited high formazan formation rates in contrast with the positive controls, containing curcumin, which reduced formazan formation significantly (p < 0.001), exhibiting cytotoxicity towards the cells. There was no significant (p > 0.05) difference in the formazan formation rates between the negative controls and 1, 0.5, 0.125, 0.065, 0.031, and 0.015 mg/ml plant extract, confirming no toxicity of C. edulis extracts towards HeLa Chang cells. Major functional phytochemical compounds identified included alcohols, phenols, alkanes, amines, carboxylic acids, and esters. LC-ESI-TOF/MS analysis revealed the putative identities of main compounds present in the aqueous leaves extract, including some that contribute to its anti-HIV-1 protease action.

First report on isolation of methyl gallate with antioxidant, anti-HIV-1 and HIV-1 enzyme inhibitory activities from a mushroom (Pholiota adiposa)

In this study, a compound with antioxidant and anti-HIV activities designated as HEB was first isolated from the edible mushroom Pholiota adiposa by extraction with ethanol and ethyl acetate. HEB was then purified by high performance liquid chromatography (HPLC) and identified to be methyl gallate (C8H8O5, 184.1Da) based on data from its mass spectrum (MS) and nuclear magnetic resonance (NMR) spectrum. HEB displayed strong antioxidant potency in inhibiting, at 1.36mM concentration, erythrocyte hemolysis and scavenging DPPH radicals and superoxide anion (O2−) by 82.4%, 85.6% and 71.4%, respectively. Besides exhibiting a low cytotoxicity, compound HEB demonstrated significant anti-HIV activity in that it inhibited HIV-1 replication in TZM-BL cells infected by pseudovirus with an IC50 value of 11.9μM. Further study disclosed that HEB inhibited the viral entry process and activities of key enzymes essential for the HIV-1 life cycle. HEB inhibited HIV-1 reverse transcriptase and integrase activities with an IC50 value of 80.1μM and 228.5μM, respectively, and at 10mM concentration inhibited HIV-1 protease activity by 17.1% which was higher than that achieved by the positive control pepstatin A. Interestingly, this study first revealed that H2O2 stimulation not only activated cell oxidative stress responses, but also accelerated HIV-1 long terminal repeat (LTR) promotion in TZM-BL cells, which was significantly reduced by HEB from 18.2% to about 2%. It implied a direct relationship between the antioxidant and anti-HIV activities of the natural active constituent HEB. Nuclear transcription factor kappa B (NF-κB) signal pathways plays an important role in oxidative stress responses. Meanwhile, there is κB target sequence in HIV promoter LTR which is significant for virus replication and gene expression. In this study, Western Blot assay showed that HEB could inhibit the activation of NF-κB signal pathway stimulated by H2O2 in mouse spleen cells through suppressing NF-κB (p65) translocation into nucleus and NF-kappa-B inhibitor (IκB) degradation in cytoplasm. In summary, the antioxidant HEB from P. adiposa could inhibit HIV-1 replication through multiple target sites. The data suggest that natural antioxidant compounds might have a potential for treatment of AIDS.

Inhibitory effects of Korean plant resources on human immunodeficiency virus type 1 protease activity

Some Korean plants were found to inhibit HIV-1 protease activity. The extracts of Acanthopanax koreanum (stem bark), Berchemia berchemiaefolia (stem), Berchemia berchemiaefolia (bark), Distylium racemosum (leaves), Distylium racemosum (stem), Lindera erythrocarpa (leaves), Physalis alkekengi var. francheti (root), Platycarya strobilacea (stem), Rodiola rosea (root), Rosa davurica (stem), Syringa dilatata (leaves), Viburnum awabuki (stem) and Viburnum awabuki (leaves) showed significant inhibitory effect against HIV-1 protease. Camelliatannin H from Camellia japonica and uvaol from Cratagus pinatrifida were potent active inhibitors of HIV-1 protease with <TEX>$IC_{50}$</TEX> values of <TEX>$0.9\;{\mu}M$</TEX> and <TEX>$5.5\;{\mu}M$</TEX>, respectively. The cure and prevention of AIDS have been a global challenge since it was discovered in the ealy 1980s. However, the development of anti-HIV agent that can effectively treat or prevent this disease are still demanded.

HIV-1 protease inhibition and anti-HIV effect of natural and synthetic water-soluble lignin-like substances.

Water-soluble lignin extracted from natural sources and dehydrogenated polymers of p-coumaric acid and ferulic acid inhibited HIV-1 protease activity. The dehydrogenated polymers, which are thought to be model compounds for lignin, were synthesized and fractionated into four ranges of molecular mass by ultra-filtration: i.e., over 30 kDa, 30-10 kDa, 10-1 kDa and 1 kDa-500 Da. All of these fractions had HIV-1 protease inhibitory activity. The anti-HIV-1 effect of the smallest mass fractions of the dehydrogenated polymers (1 kDa-500 Da) was also tested, and it was found that these fractions inhibited the replication of HIV-1 in MT-4 cells.

A rapid and sensitive bacterial assay to determine the inhibitory effect of ‘interface’ peptides on HIV-1 protease co-expressed in Escherichia coli

The HIV-1 protease is essential for maturation of virus particles and is, therefore, an attractive target for antiviral drugs. The function of this protease depends on the dimerization of two identical subunits. Commonly used protease inhibitors are directed mainly against the active site of the enzyme which often leads to viral resistance. To determine the inhibitory effect of peptides interfering with the dimerization site of the HIV-1 protease, a recombinant bacterial screening assay was established. Escherichia coli was co-transformed with two different plasmids, expressing the ‘interface’ peptide and an active HIV-1 protease toxic for the bacteria. Co-expression of inhibitory peptides overcomes the incomplete membrane transmission of supplemented inhibitors and leads to a direct interaction of the inhibitory peptide and the HIV-1 protease. The inhibitory effect of co-expressed peptides was measured by an increased growth of co-transformed bacteria, compared with a slowly growing E. coli control culture only expressing the HIV-1 protease. Using this assay several penta- and hexa-peptides were screened for their ability to inhibit HIV-1 protease activity. One of these peptides showed a significant inhibitory effect on co-expressed recombinant HIV-1 protease.

Targeting the Dimerization Interface of HIV-1 Protease: Inhibition with Cross-Linked Interfacial Peptides

Agents have been designed and synthesized which target the dimerization interface of HIV-1 protease. These agents, which contain cross-linked peptides from the N- and C-termini of the protease, both inhibit HIV-1 protease activity and decrease the amount of protease dimer in solution as measured by size exclusion chromatography, protein crosslinking, and protease fluorescence studies. Additionally we have shown that active site-targeted agents inhibit HIV-1 protease activity but have little effect on protease dimerization. These data support the claim that inhibition with the crosslinked agents is based on a decrease in the amount of protease homodimer in solution which in turn is responsible for a decrease in the activity of the protease.

Screening of Traditional Medicines for their Inhibitory Activity Against HIV-1 Protease

Aqueous extracts of 75 different traditional medicines, currently used for the treatment of various human diseases in China, Japan and Indonesia, were screened for possible inhibitory activity against HIV-1 protease using fluorogenic assay. Forty extracts exhibited inhibitory activity at a concentration of 250 μg/mL, and 18 of them inhibited HIV-1 protease activity by more than 70%. Among the extracts examined, Belamcanda chinensis, Magnolia fargesii, Paeonia suffruticosa, Phellodendron amurense and Terminalia chebula were also effective at a concentration of 25 μg/mL in the fluorogenic assay.

Screening of Traditional Medicines for their Inhibitory Activity Against HIV-1 Protease

Aqueous extracts of 75 different traditional medicines, currently used for the treatment of various human diseases in China, Japan and Indonesia, were screened for possible inhibitory activity against HIV-1 protease using fluorogenic assay. Forty extracts exhibited inhibitory activity at a concentration of 250 μg/mL, and 18 of them inhibited HIV-1 protease activity by more than 70%. Among the extracts examined, Belamcanda chinensis, Magnolia fargesii, Paeonia suffruticosa, Phellodendron amurense and Terminalia chebula were also effective at a concentration of 25 μg/mL in the fluorogenic assay.

HIV-1 protease inhibitory activity of L-694,746, a novel metabolite of L-689,502

L-689,502 is a potent inhibitor of HIV-1 protease activity in vitro. Microbial biotransformations of L-689,502 by cultures belonging to the genus Streptomyces sp. were performed. Extracts of culture broths were examined for the production of metabolites of L-689,502 that could inhibit HIV-1 protease activity. One culture, MA 6804 ( Streptomyces lavendulae, ATCC 55095), produced L-694,746 that, while being structurally related to L-689,502, is a novel metabolite and a potent inhibitor of HIV-1 protease.