Human cytomegalovirus UL27 is not required for viral replication in human tissue implanted in SCID mice

Inhibition of the human cytomegalovirus UL97 kinase by maribavir is thought to be responsible for the antiviral activity of this compound. Some mutations that confer resistance to maribavir map to UL97, however additional mutations that also confer resistance to the drug were mapped to UL27. These open reading frames share a low level of homology, yet the function of pUL27 remains unknown. A recombinant virus with a deletion in the UL27 open reading frame was reported previously to exhibit a slight replication deficit, but a more important function in vivo was hypothesized given its homology to the UL97 kinase. The potential for an important function in vivo was investigated by determining if these knockout viruses could replicate in human tissue implanted in SCID mice. None of the AD169 derived viruses replicated well in the implanted thymus/liver tissue, and is consistent with previous observations, although all of the viruses replicated to some degree in retinal tissue implants. Replication of the parent viruses was observed at 7 days post inoculation, whereas no replication was detected with any of the recombinant viruses with deletions in UL27. By day 14, replication was detected in two of the three knockout viruses and in all of the viruses by day 42. These data are consistent with minimal defects observed in cell culture, but are not consistent with an important role for UL27 in vivo. We conclude that UL27 is not required for viral replication in vivo

Oral Activity of a Methylenecyclopropane Analog, Cyclopropavir, in Animal Models for Cytomegalovirus Infections

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 log(10). 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

In Vitro Activities of Benzimidazole d- and l-Ribonucleosides against Herpesviruses

Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2), varicella-zoster virus (VZV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), human herpesvirus 6 (HHV-6), and human herpesvirus 8 (HHV-8) are responsible for a number of clinical manifestations in both normal and immunocompromised individuals. The parent benzimidazole ribonucleosides evaluated in this series, 2-bromo-5,6-dichloro-1-(β-d-ribofuranosyl)benzimidazole (BDCRB) and maribavir (1263W94), are potent and selective inhibitors of human CMV replication. These nucleosides act by two different mechanisms. BDCRB blocks the processing and maturation of viral DNA, whereas 1263W94 inhibits the viral enzyme pUL97 and interferes with DNA synthesis. In the present study, we have evaluated the in vitro antiviral activity of BDCRB, an analog, GW275175X (175X), and 1263W94 against the replication of HSV-1, HSV-2, VZV, CMV, EBV, HHV-6, and HHV-8. By using various methodologies, significant activity was observed against human CMV and EBV but not against HSV-1, HSV-2, VZV, HHV-6, or HHV-8. Plaque reduction assays performed on a variety of laboratory and clinical isolates of human CMV indicated that all strains, including those resistant to ganciclovir (GCV) and foscarnet, were sensitive to all three benzimidazole ribonucleosides, with mean 50% effective concentration values of about 1 to 5 μM compared to that of GCV at 6 μM. The toxicity of these compounds in tissue culture cells appeared to be similar to that observed with GCV. These results demonstrate that the benzimidazole ribonucleosides are active against human CMV and EBV and suggest that they may be useful for the treatment of infections caused by these herpesviruses