Screening for anti-HIV drugs that can combine virucidal and virustatic activities synergistically

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Abstract

Chlorcyclizine HCl and ciprofloxacin HCl were shown to have anti-HIV activity. They possess virustatic and virucidal activities against HIV, a murine retrovirus (RV) and several other RNA and DNA viruses. These drugs were screened from a large number of compounds on the basis of in vitro mutagenicity and antimetabolite detection tests. Subsequent studies were based on different exo vivo cell cultures. These two compounds were then tested on an animal model, following standard test protocols, using another retrovirus, maintained as Ehrlich’s ascites cell tumour virus (EACTV). The animal mortality and protection tests corroborated the findings obtained in vitro, suggesting that these drugs acted synergistically against HIV, exhibiting both virucidal and virustatic properties.

Introduction

The retroviruses of man and animals constitute a unique organisation of a dual RNA–DNA controlled replicative molecular machinery, whose counterparts are the transposon elements (TnE), occurring in bacteria as well as in plants and other biological systems.

The current anti-HIV drugs in usage have been reviewed [1], [2], [3]. Several categories of drugs exist which act either by causing errors of nucleic acid synthesis, or by acting as inhibitors of key proteins or enzymes responsible for the synthesis of crucial components of the virus, or by causing interference with the process of viral infection of the cells, mediated through interferons, and blocking of the CD4 receptors/sequences. In addition, there are several viral inhibitory chemical compounds of herbal origin.

Despite the above mechanisms and other approaches leading to a design of newer anti-HIV drugs, no drug which can completely stop the multiplication of HIV or other retroviruses exists as yet. This is probably because of the versatility of the recombination processes of the retroviruses which reside in several copies of the same TNE-RV in a single genome [1], [4]. All of the currently known drugs, being virustatic, seem to have limitations as anti-HIV agents, because of their inability to stop the viral replication irreversibly.

The purpose of the present study was to identify newer drugs that may possess anti-viral properties. Though animal models and tissue culture systems are available for evaluation of the anti-viral potentials of these compounds, these methods are often dilatory, costly and not entirely predictable. It is the basic similarity of the biochemical and enzymatic processes between bacterial and mammalian cells that justifies the use of bacteria in some preliminary screening processes, such as the Ames’ test; or Hanka’s anti-metabolite test [5], [6].

Section snippets

Test compounds and reagents

Two agents, namely chlorcyclizine and ciprofloxacin HCl appeared the most active when a number of anti-viral agents were screened [7].

Chlorcyclizine (CCZ), ciprofloxacin (CIP), azidothymidine (AZT) and other chemicals and reagents were dissolved in sterile distilled water, stored at −20°C in aliquots of working units, thawed and added to tissue culture and other media, to a final pH of 7.4–7.6. Addition of these chemicals to tissue/cell culture media directly was avoided as chlorcyclizine forms

Toxicity in cell cultures

It was found that the non-toxic concentrations of chlorcyclizine was <12.5 mg/l as was that of nalidixic acid (NAL), used as control, whilst for ciprofloxacin the concentration was 25.0 mg/l. Resistance of the cells to chlorcyclizine was in the order: Vero cells, KB cells and Hep 2 cells.

The toxic effects of both drugs (concentrations varying between 3.1 to 50 mg/l) on C8166 cells on the basis of cell viability by dye exclusion tests were noted. The viability of cells on exposure to

Discussion

The inadequacy of highly effective anti-retroviral and anti-HIV drugs is well recognised; this is particularly so with respect to virucidal drugs. Research had been centred on derivatives of major classes of virustatic drugs. Several newer compounds with anti-cancer, anti-viral and anti-retroviral potentials may be obtained if research is conducted on drugs designed as antibiotics. The screening of such drugs facilitated by the initial use of bacterial prescreens, could short-list these agents

Acknowledgements

We thank the Heads of the Department of Virology and Virus Research Centre, School of Tropical Medicine, Calcutta; Indian Council of Medical Research, New Delhi and Government of West Bengal; Chittaranjan National Cancer Research Institute, Government of India, and the Department of Academic Virology, Royal London Hospital School of Medicine and Dentistry, UK, who kindly supplied us the Reference strains or cultures of bacteria, viruses and the animals. We are particularly grateful to the

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