cyclopropavir and Cytomegalovirus-Infections

Ganciclovir (GCV), the therapy of choice for human cytomegalovirus (CMV) infections and foscarnet, a drug used to treat GCV-resistant CMV infections was approved more than twenty years ago. Although cidofovir and a prodrug of GCV have since been added to the armamentarium, a highly effective drug without significant toxicities has yet to be approved. Such a therapeutic agent is required for treatment of immunocompromised hosts and infants, which bear the greatest burden of disease. The modest antiviral activity of existing drugs is insufficient to completely suppress viral replication, which results in the selection of drug-resistant variants that remain pathogenic, continue to replicate, and contribute to disease. Sustained efforts, largely in the biotech industry and academia, have identified highly active lead compounds that have progressed into clinical studies with varying levels of success. A few of these compounds inhibit new molecular targets, remain effective against isolates that have developed resistance to existing therapies, and promise to augment existing therapies. Some of the more promising drugs will be discussed with an emphasis on those progressing to clinical studies. Their antiviral activity both in vitro and in vivo, spectrum of antiviral activity, and mechanism of action will be reviewed to provide an update on the progress of potential new therapies for CMV infections.

Topics: Antiviral Agents; Cidofovir; Cyclopropanes; Cytomegalovirus; Cytomegalovirus Infections; Cytosine; Foscarnet; Ganciclovir; Guanine; Humans; Organophosphonates; Valganciclovir; Virus Replication

Human cytomegalovirus (HCMV) can cause severe disease in patients with compromised or immature immune systems. Currently approved pharmacotherapies for the treatment of systemic HCMV infections [ganciclovir (GCV), cidofovir (CDV), foscarnet] are limited by a high incidence of adverse effects and/or the development of drug resistance. Given that many of these drugs have the same viral target (HCMV-encoded DNA polymerase), cross-resistance is relatively common. The primary means to combat drug resistance is combination pharmacotherapy using therapeutics with different molecular mechanisms of action with the expectation that those combinations result in an additive or synergistic enhancement of effect; combinations that result in antagonism can, in many cases, be detrimental to the outcome of the patient. We therefore tested select combinations of approved (GCV, CDV, letermovir (LMV)) and experimental (brincidofovir (BCV), cyclopropavir (CPV), maribavir (MBV), BDCRB) drugs with the hypothesis that combinations of drugs with different and distinct molecular mechanisms of action will produce an additive and/or synergistic enhancement of antiviral effect against HCMV in vitro. Using MacSynergy II (a statistical package that measures enhancement or lessening of effect relative to zero/additive), select drug combination studies demonstrated combination indices ranging from 160 to 372 with 95% confidence intervals greater than zero indicating that these combinations elicit a synergistic enhancement of effect against HCMV in vitro. These data suggest that administration of a viral DNA polymerase inhibitor, MBV, and/or a viral terminase inhibitor in combination has the potential to address the resistance/cross-resistance problems associated with currently available therapeutics.

Topics: Antiviral Agents; Benzimidazoles; Cell Line; Cidofovir; Cyclopropanes; Cytomegalovirus; Cytomegalovirus Infections; Cytosine; DNA-Directed DNA Polymerase; Drug Antagonism; Drug Combinations; Drug Resistance, Viral; Drug Synergism; Drug Therapy, Combination; Endodeoxyribonucleases; Fibroblasts; Foscarnet; Ganciclovir; Guanine; Humans; Nucleic Acid Synthesis Inhibitors; Organophosphonates; Ribonucleosides; Viral Proteins; Virus Replication

Human cytomegalovirus UL97 kinase mutations that commonly confer ganciclovir resistance cluster in different parts of the gene than those conferring resistance to maribavir, an experimental UL97 kinase inhibitor. The drug resistance, growth, and autophosphorylation phenotypes of several unusual UL97 mutations in the kinase catalytic domain were characterized. Mutations V466G and P521L, described in clinical specimens from ganciclovir-treated subjects, conferred a UL97 kinase knockout phenotype with no autophosphorylation, a severe growth defect, and high-level ganciclovir, cyclopropavir, and maribavir resistance, similar to mutations at the catalytic lysine residue K355. Mutations F342S and V356G, observed after propagation under cyclopropavir in vitro, showed much less growth attenuation and moderate- to high-level resistance to all three drugs while maintaining UL97 autophosphorylation competence and normal cytopathic effect in cell culture, a novel phenotype. F342S is located in the ATP-binding P-loop and is homologous to a c-Abl kinase mutation conferring resistance to imatinib. UL97 mutants with relatively preserved growth fitness and multidrug resistance are of greater concern in antiviral therapy than the severely growth-impaired UL97 knockout mutants. Current diagnostic genotyping assays are unlikely to detect F342S and V356G, and the frequency of their appearance in clinical specimens remains undefined.

Topics: Antiviral Agents; Benzimidazoles; Catalytic Domain; Cell Line; Cyclopropanes; Cytomegalovirus; Cytomegalovirus Infections; Drug Resistance, Multiple; Drug Resistance, Viral; Ganciclovir; Genotype; Guanine; Humans; Mutation; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Ribonucleosides

Human cytomegalovirus (CMV) UL54 DNA polymerase (pol) mutants with known patterns of resistance to current antivirals ganciclovir (GCV), foscarnet (FOS), and cidofovir (CDV) were tested for cyclopropavir (CPV) susceptibility by a standardized reporter-based yield reduction assay. Exonuclease and A987G (region V) mutations at codons commonly associated with dual GCV-CDV resistance in clinical isolates paradoxically conferred increased CPV susceptibility. Various polymerase catalytic region mutations conferring FOS resistance with variable low-grade GCV and CDV cross-resistance also conferred CPV resistance, with 50% effective concentration (EC(50)) increases of 3- to 13-fold. CPV EC(50) values against several pol mutants were increased about 2-fold by adding UL97 mutation C592G. Propagation of a CMV exonuclease mutant under CPV selected for pol mutations less often than UL97 mutations. In 21 experiments, one instance each of mutations E756D and M844V, which were shown individually to confer 3- to 4-fold increases in CPV EC(50), was detected. Unlike GCV and CDV, exonuclease mutations are not a preferred mechanism of CPV resistance, but mutations in and near pol region III may confer CPV resistance by affecting its recognition as an incoming base for DNA polymerization.

Topics: Alkaline Phosphatase; Antiviral Agents; Cidofovir; Cyclopropanes; Cytomegalovirus; Cytomegalovirus Infections; Cytosine; DNA-Directed DNA Polymerase; Drug Resistance, Viral; Exonucleases; Foscarnet; Ganciclovir; Genes, Reporter; Genotype; Guanine; Humans; Inhibitory Concentration 50; Mutation; Organophosphonates; Phenotype; Viral Proteins; Virus Replication

Synthesis of fluorinated cyclopropavir analogues 13a, 13b, 14a, and 14b is described starting from alkene 15. Addition of carbene derived from dibromofluoromethane gave bromofluoro cyclopropane 16. Reduction (compound 17) followed by desilylation gave intermediate 18, which was transformed to 2-nitrophenylselenenyl derivative 19. Oxidation to selenoxide 20 was followed by beta-elimination to afford methylenecyclopropane 21. Addition of bromine provided compound 22 for alkylation-elimination of adenine and 2-amino-6-chloropurine. The resultant E,Z isomeric mixtures of methylenecyclopropanes 23a + 24a and 23c + 24c were resolved and the individual isomers were deprotected to give adenine analogues 13a and 14a as well as compounds 13c and 14c. Hydrolytic dechlorination of 13c and 14c furnished guanine analogues 13b and 14b. The only significant antiviral effects were observed with analogue 13a against HCMV and 14a against VZV in cytopathic inhibition assays.

Topics: Adenine; Antiviral Agents; Bromine; Chemistry, Pharmaceutical; Cyclopropanes; Cytomegalovirus; Cytomegalovirus Infections; Drug Design; Guanine; Inhibitory Concentration 50; Magnetic Resonance Spectroscopy; Models, Chemical; Nucleic Acids; Nucleosides; Ultraviolet Rays

We reported previously that purine 2-(hydroxymethyl)methylenecyclopropane analogs have good activity against cytomegalovirus infection. A second-generation analog, (Z)-9-[[2,2-bis-(hydroxymethyl)cyclopropylidene]methyl]guanine (ZSM-I-62, cyclopropavir [CPV]), has particularly good activity against murine and human cytomegaloviruses (MCMV and HCMV) in vitro. To determine the oral activity of this compound in vivo, BALB/c or severe combined immunodeficient (SCID) mice infected with MCMV and two models using SCID mice implanted with human fetal tissue and subsequently infected with HCMV were used. In MCMV-infected normal mice, CPV at 10 mg/kg of body weight was highly effective in preventing mortality when administered at 24, 48, or 72 h post-viral inoculation and reduced titers of virus in tissues of SCID mice by 2 to 5 log10. In one HCMV model, human fetal retinal tissue was implanted into the anterior chamber of the mouse eye and inoculated with the Toledo strain of HCMV, and in the second, human fetal thymus and liver tissues were implanted under the kidney capsule of mice and then inoculated with HCMV. In general, replication of HCMV in both types of implant tissue increased from 7 through 21 to 28 days and then gradually decreased to undetectable levels by 8 weeks postinfection. Oral treatment with 45 or 15 mg of CPV/kg initiated 24 h after infection was highly effective in reducing replication to undetectable levels in both models and was generally more effective than ganciclovir. These data indicate that the methylenecyclopropane analog, CPV, was highly efficacious in these four animal models and should be evaluated for use in HCMV infections in humans.

Topics: Animals; Antiviral Agents; Cyclopropanes; Cytomegalovirus Infections; Cytopathogenic Effect, Viral; Endpoint Determination; Guanine; Humans; Immunocompromised Host; Injections, Intraperitoneal; Liver; Mice; Mice, Inbred BALB C; Mice, SCID; Retina; Thymus Gland; Viral Plaque Assay; Virus Replication