Disintegration of retroviruses by chelating agents

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Disintegration of retroviruses by chelating agents

V. Wunderlich1 and G. Sydow1(1) Central Institute for Cancer

Research, -Rössle-Institute, Academy of Sciences of the German

Democratic Republic, Berlin, German Democratic Republic

Summary Exposurein vitro of various mammalian retroviruses to the chelating

agents EDTA or EGTA in millimolar concentrations resulted in partial

disintegration of viral membranes as measured by accessibility or even

release of reverse transcriptase, an internal viral protein, without any

other treatment usually required. Among the viruses responding to chelators

were mammalian type C viruses, primate type D viruses and bovine leukemia

virus. The effect was dose-dependent. The avian type C virus AMV, however,

was found to be not susceptible to the agents. Rauscher mouse leukemia virus

treatedin vitro with EDTA or EGTA showed reduced infectivity in mice. The

results are considered as evidence for some association of divalent cations

with membranes of mammalian retroviruses. The disintegrating activity of

EGTA suggests that Ca2+ is an integral constituent of viruses but Mg2+ may

also be involved. These cations seem to be responsible for maintaining

integrity of retroviral membranes which, after chelation of ions, are either

disrupted or become permeable for the exogenous template of reverse

transcriptase. In addition, the distintegrating activity of trifluoperazine

may indicate that a calmodulin-like protein occurs in retroviral membranes.

Received: 1 March 1982 Accepted: 2 April 1982

Metal chelation underlies anti-HIV integrase activity of beta-diketo acids

[no abstract] http://www.accessmylibrary.com/coms2/summary_0286-803606_ITM

Use of DMSA as antiretroviral treatment:

http://www.freepatentsonline.com/5464869.html

J Med Chem. 1998 Aug 13;41(17):3202-9.

Salicylhydrazine-containing inhibitors of HIV-1 integrase: implication for a

selective chelation in the integrase active site.

Neamati N, Hong H, Owen JM, Sunder S, Winslow HE, Christensen JL, Zhao

H, Burke TR Jr, Milne GW, Pommier Y. Laboratories of Molecular

Pharmacology and Medicinal Chemistry, Division of Basic Sciences, National

Cancer Institute, Bethesda, land 20892-4255, USA.

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. A common feature for activity

among these inhibitors is the hydroxyl group of the salicyl moiety. Although

the active inhibitors must contain this hydroxyl group, other structural

modifications can also influence potency. Removal of this hydroxyl group or

replacement with an amino, bromo, fluoro, carboxylic acid, or ethyl ether

totally abolished potency against integrase. Several asymmetric structures

exhibited similar potency to the symmetric lead inhibitor 1. The

superimposition of the lowest-energy conformations upon one another revealed

three sites whose properties appear important for ligand binding. Site A is

composed of the 2-hydroxyphenyl, the alpha-keto, and the hydrazine moieties

in a planar conformation. We propose that this site could interact with

HIV-1 integrase by chelation of the metal in the integrase active site as

inhibition of HIV-1 integrase catalytic activity and DNA binding were

strictly Mn2+-dependent. The hydrophobic sites B and C are probably

responsible for complementarity of molecular shape between ligand and

receptor. Our data indicate that only those compounds which possessed sites

A, B, and C in a linear orientation were potent inhibitors of HIV-1

integrase. Although all the active inhibitors possessed considerable

cytotoxicity and no apparent antiviral activity in CEM cells, the study

presents useful information regarding ligand interaction with HIV-1

integrase protein. PMID: 9703465 [PubMed - indexed for MEDLINE]

Mol Pharmacol. 1997 Dec;52(6):1041-55.

Potent inhibitors of human immunodeficiency virus type 1 integrase:

identification of a novel four-point pharmacophore and tetracyclines as

novel inhibitors.

Neamati N, Hong H, Sunder S, Milne GW, Pommier Y. Laboratory of

Molecular Pharmacology, National Cancer Institute, Bethesda, land 20892,

USA.

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). Each retrieved structure was fit to the

pharmacophore, and the conformation that afforded the best fit was

identified. Twenty of the 39 compounds tested exhibited IC50 values of < 20

microM. Among the most potent inhibitors, tetracyclines emerged as a new

class of inhibitors. Although the parent tetracycline exhibited marginal

potency against purified IN, all substituted tetracyclines tested showed

5-100-fold increased potency. Disintegration assays with truncated IN

mutants indicated that tetracyclines inhibit the IN catalytic core domain.

To investigate whether chelation of divalent metals is implicated in

differential potency of tetracyclines, enzyme assays were performed in the

presence of both Mn2+ or Mg2+; no significance difference in potency was

observed. Rolitetracycline inhibited IN/DNA complex formation in the

presence of EDTA, which suggests that inhibition was metal independent.

Rolitetracycline reversed DNA binding of IN after the complex was allowed to

form before the addition of drug. Selectivity of tetracyclines was also

examined in an assay specific for topoisomerase I, and none of the

tetracyclines tested induced topoisomerase I-mediated cleavable complex or

inhibited camptothecin-induced cleavable complex. Remarkable potency against

the IN in the absence of divalent metals and the core enzyme coupled with

water solubility makes tetracyclines potential candidates for X-ray crystal

structure determination with IN. PMID: 9415714

Biochem Biophys Res Commun. 1995 Dec 26;217(3):802-10.

Characterization of the DNA-binding activity of HIV-1 integrase using a

filter binding assay.

Haugan IR, Nilsen BM, Worland S, Olsen L, Helland DE. Laboratory of

Biotechnology, University of Bergen, Norway.

Based on the selective binding of proteins and DNA to distinct filter

materials a double-layered dot blot radio assay was developed to evaluate

the binding of DNA to HIV-1 integrase. In this assay the DNA-binding was

found to be independent of Mn2+ concentration, inhibited by concentrations

of Mg2+ above 5 mM, abolished by zinc chelation and inhibited by monoclonal

antibodies reacting with either the N-terminal or C-terminal regions of

integrase. Atomic absorption spectroscopy revealed the molar ratio between

integrase and zinc to be close to 1. It is concluded that both the

N-terminal and the C-terminal regions of integrase are involved in

DNA-binding and that the reported double-layered dot blot radio assay is

well suited for further characterization of the integrase. PMID: 8554601

Advances in two-metal chelation inhibitors of HIV integrase

Authors: s, A; Svolto, Angilique C Source: Expert Opinion on

Therapeutic Patents, Volume 18, Number 11, November 2008 , pp. 1225-1237(13)

Abstract:

Background: Small-molecule inhibitors of the retroviral enzyme HIV integrase

are among the most exciting discoveries of the past 10 years in the area of

infectious disease research. The field of two-metal chelating inhibitors of

this nucleic-acid-modifying enzyme has exploded since the early diketo-acid

discoveries in the late 1990s and the advent of the more drug-like

heterocyclic templates in the early 2000s. The field of research has now

expanded across multiple companies, resulting in several clinical

candidates, and ultimately delivered the first licensed inhibitor -

raltegravir - in late 2007. Objective: This review is an attempt to assess

the most relevant patent literature concerning two-metal chelators from the

pioneering examples in 1999 that first displayed antiviral activity through

mid-2008. Methods: The existing integrase inhibitor patent literature was

surveyed for chemical series that consisted of a two-metal binding

pharmacaphore, which were further broken down into structural subclasses for

presentation. Results/conclusions: The integrase inhibitor field has matured

rapidly over the past decade into a major area of focus for new HIV

therapeutics. This work has resulted in one licensed drug and clearly

defined needs for future directions centered around viral resistance, for

which solutions that are contained within the reviewed patents will

undoubtedly emerge.