Effect of HIV integrase inhibitors on the RAG1/2 recombinase.

Assembly of functional Ig and T cell receptor genes by V(D)J recombination depends on site-specific cleavage of chromosomal DNA by the RAG1/2 recombinase. As RAG1/2 action has mechanistic similarities to DNA transposases and integrases such as HIV-1 integrase, we sought to determine how integrase inhibitors of the diketo acid type would affect the various activities of RAG1/2. Both of the inhibitors we tested interfered with DNA cleavage and disintegration activities of RAG1/2, apparently by disrupting interaction with the DNA motifs bound specifically by the recombinase.

Arylisothiocyanate-containing esters of caffeic acid designed as affinity ligands for HIV-1 integrase.

Integrase is an enzyme found in human immunodeficiency virus, which is required for the viral life cycle, yet has no human cellular homologue. For this reason, HIV integrase (IN) has become an important target for the development of new AIDS therapeutics. Irreversible affinity ligands have proven to be valuable tools for studying a number of enzyme and protein systems, yet to date there have been no reports of such affinity ligands for the study of IN.

Discovery of a nuclease-resistant, non-natural dinucleotide that inhibits HIV-1 integrase.

Integration of HIV viral DNA into human chromosomal DNA catalyzed by HIV integrase is essential for the replication of HIV. Discovery of novel inhibitors of HIV integrase is of considerable significance in approaches to the development of therapeutic agents against AIDS. We have synthesized a new dinucleotide 1 with an internucleotide phosphate bond that is unusually resistant to exonucleases.

Structure-based HIV-1 integrase inhibitor design: a future perspective.

The pol gene of HIV-1 encodes for three essential enzymes, protease (PR), reverse transcriptase (RT) and integrase (IN). More than 16 drugs, targeting two of these enzymes, PR and RT have been approved by the FDA. At present, there are no clinically useful agents that inhibit the third enzyme, IN.

Identification of HIV-1 integrase inhibitors via three-dimensional database searching using ASV and HIV-1 integrases as targets.

Integration of viral DNA into the host cell genome is a critical step in the life cycle of HIV. This essential reaction is catalyzed by integrase (IN) through two steps, 3'-processing and DNA strand transfer. Integrase is an attractive target for drug design because there is no known cellular analogue and integration is essential for successful replication of HIV.

Retroviral integrase inhibitors year 2000: update and perspectives.

HIV-1 integrase is an essential enzyme for retroviral replication and a rational target for the design of anti-AIDS drugs. A number of inhibitors have been reported in the past 8 years. This review focuses on the recent developments in the past 2 years.

Viral entry as the primary target for the anti-HIV activity of chicoric acid and its tetra-acetyl esters.

The antiviral activity of L-chicoric acid against HIV-1 has been attributed previously to the inhibition of HIV-1 integration. This conclusion was based on the inhibition of integrase activity in enzymatic assays and the isolation of a resistant HIV strain with a mutation (G140S) in the integrase gene. Here we show that the primary antiviral target of L-CA and its analogs in cell culture is viral entry.

Recognition and inhibition of HIV integrase by a novel dinucleotide.

The viral enzyme, HIV integrase, is involved in the integration of viral DNA into host cell DNA. In the quest for a small nucleotide system with nuclease stability of the internucleotide phosphate bond and critical structural features for recognition and inhibition of HIV-1 integrase, we have discovered a conceptually novel dinucleotide, pIsodApdC, which is a potent inhibitor of this key viral enzyme.

Retroviral integrase : a novel target in antiviral drug development and basic in vitro assays with the purified enzyme.

Two critical events are the signature of the life cycle of retroviruses (1). The first is reverse transcription, whereby the single-stranded RNA genome of the retrovirus is copied into double-stranded DNA. The second of these events is integration, whereby this viral DNA is inserted into a chromosome of the host cell, establishing what is known as the proviral state.

Solution structure of anti-HIV-1 and anti-tumor protein MAP30: structural insights into its multiple functions.

We present the solution structure of MAP30, a plant protein with anti-HIV and anti-tumor activities. Structural analysis and subsequent biochemical assays lead to several novel discoveries. First, MAP30 acts like a DNA glycosylase/apurinic (ap) lyase, an additional activity distinct from its known RNA N-glycosidase activity toward the 28S rRNA.

Thiazolothiazepine inhibitors of HIV-1 integrase.

A series of thiazolothiazepines were prepared and tested against purified human immunodeficiency virus type-1 integrase (HIV-1 IN) and viral replication. Structure-activity studies reveal that the compounds possessing the pentatomic moiety SC(O)CNC(O) with two carbonyl groups are in general more potent against purified IN than those containing only one carbonyl group. Substitution with electron-donating or -withdrawing groups did not enhance nor abolish potency against purified IN.

Inhibitors of human immunodeficiency virus integrase.

Integration of the viral DNA into a host cell chromosome is an essential step for HIV replication and maintenance of persistent infection. Two viral factors are essential for integration: the viral DNA termini (the att sites) and IN. Accruing knowledge of the IN structure, catalytic mechanisms, and interactions with other proteins can be used to design strategies to block integration.

Chicoric acid analogues as HIV-1 integrase inhibitors.

The present study was undertaken to examine structural features of L-chicoric acid (3) which are important for potency against purified HIV-1 integrase and for reported cytoprotective effects in cell-based systems. Through a progressive series of analogues, it was shown that enantiomeric D-chicoric acid (4) retains inhibitory potency against purified integrase equal to its L-counterpart and further that removal of either one or both carboxylic functionalities results in essentially no loss of inhibitory potency. Additionally, while two caffeoyl moieties are required, attachment of caffeoyl groups to the central linking structure can be achieved via amide or mixed amide/ester linkages.

Inhibition of HIV integrase by novel nucleotides bearing tricyclic bases.

5'-Monophosphates of several novel dideoxynucleosides bearing tricyclic nucleobases were synthesized. Both linear and angular ring-extended analogs of isomeric dideoxyadenosine 5'-monophosphate were discovered to have moderate to good inhibition of the viral-encoded enzyme, HIV integrase. The results suggest that the nucleotide binding site of HIV integrase can accommodate major modifications in the nucleobase, which is in stark contrast to the nucleotide binding site on HIV reverse transcriptase.

Identification of HIV-1 integrase inhibitors based on a four-point pharmacophore.

The rapid emergence of human immunodeficiency virus (HIV) strains resistant to available drugs implies that effective treatment modalities will require the use of a combination of drugs targeting different sites of the HIV life cycle. Because the virus cannot replicate without integration into a host chromosome, HIV-1 integrase (IN) is an attractive therapeutic target. Thus, an effective IN inhibitor should provide additional benefit in combination chemotherapy.

Salicylhydrazine-containing inhibitors of HIV-1 integrase: implication for a selective chelation in the integrase active site.

In previous studies we identified N,N'-bis(salicylhydrazine) (1) as a lead compound against purified recombinant HIV-1 integrase. We have now expanded upon these earlier observations and tested 45 novel hydrazides. Among the compounds tested, 11 derivatives exhibited 50% inhibitory concentrations (IC50) of less than 3 microM.

Highly potent synthetic polyamides, bisdistamycins, and lexitropsins as inhibitors of human immunodeficiency virus type 1 integrase.

Alignment of the available human immunodeficiency virus type 1 (HIV-1) viral DNA termini [U5 and U3 long terminal repeats (LTRs)] shows a high degree of conservation and the presence of a stretch of five or six consecutive adenine and thymine (AT) sequences approximately 10 nucleotides away from each LTR end. A series of AT-selective minor-groove binders, including distamycin and bisdistamycins, bisnetropsins, novel lexitropsins, and the classic monomeric DNA binders Hoechst 33258, 4'-diamino-2-phenylindole, pentamidine, berenil, spermine, and spermidine, were tested for their inhibitory activities against HIV-1 integrase (IN). Although netropsin, distamycin, and all other monomeric DNA binders showed weak activities in the range of 50-200 microM, some of the polyamides, bisdistamycins, and lexitropsins were remarkably active at nanomolar concentrations.

Structure of the catalytic domain of avian sarcoma virus integrase with a bound HIV-1 integrase-targeted inhibitor.

The x-ray structures of an inhibitor complex of the catalytic core domain of avian sarcoma virus integrase (ASV IN) were solved at 1.9- to 2.0-A resolution at two pH values, with and without Mn2+ cations.

Identification of a nucleotide binding site in HIV-1 integrase.

HIV-1 integrase is essential for viral replication and can be inhibited by antiviral nucleotides. Photoaffinity labeling with the 3'-azido-3'-deoxythymidine (AZT) analog 3',5-diazido-2', 3'-dideoxyuridine 5'-monophosphate (5N3-AZTMP) and proteolytic mapping identified the amino acid 153-167 region of integrase as the site of photocrosslinking. Docking of 5N3-AZTMP revealed the possibility for strong hydrogen bonds between the inhibitor and lysines 156, 159, and 160 of the enzyme.

Cellular pharmacology of mitoxantrone in p-glycoprotein-positive and -negative human myeloid leukemic cell lines.

Previous reports suggest that resistance to mitoxantrone in different tumor cell lines is unrelated to the overexpression of p-glycoprotein. In order to determine the role of p-glycoprotein in the cellular pharmacology of mitoxantrone flow cytometry and confocal microscopy were used to study two human myeloid leukemia cell lines selected for resistance to mitoxantrone (HL-60MX2) and doxorubicin (HL-60DOX). To optimize the detection of intracellular mitoxantrone, we determined the maximum excitation (607 nm) and emission (684 nm) wavelength by fluorescence spectroscopy.

Potent inhibitors of human immunodeficiency virus type 1 integrase: identification of a novel four-point pharmacophore and tetracyclines as novel inhibitors.

A four-point pharmacophore was constructed from energy-minimized structures of chicoric acid and dicaffeoylquinic acid. The search of 206,876 structures in the National Cancer Institute 3D database yielded 179 compounds that contain this pharmacophore. Thirty-nine of these compounds were tested in an in vitro assay specific for human immunodeficiency virus type 1 integrase (IN).

Curcumin analogs with altered potencies against HIV-1 integrase as probes for biochemical mechanisms of drug action.

We have previously reported the inhibitory activity of curcumin against human immunodeficiency virus type one (HIV-1) integrase. In the present study, we have synthesized and tested analogs of curcumin to explore the structure-activity relationships and mechanism of action of this family of compounds in more detail. We found that two curcumin analogs, dicaffeoylmethane (6) and rosmarinic acid (9), inhibited both activities of integrase with IC50 values below 10 microM.