Lobucavir is phosphorylated in human cytomegalovirus-infected and -uninfected cells and inhibits the viral DNA polymerase

Abstract

Lobucavir (LBV) is a deoxyguanine nucleoside analog with broad-spectrum antiviral activity. LBV was previously shown to inhibit herpes simplex virus (HSV) DNA polymerase after phosphorylation by the HSV thymidine kinase. Here we determined the mechanism of action of LBV against human cytomegalovirus (HCMV). LBV inhibited HCMV DNA synthesis to a degree comparable to that of ganciclovir (GCV), a drug known to target the viral DNA polymerase. The expression of late proteins and RNA, dependent on viral DNA synthesis, was also inhibited by LBV. Immediate-early and early HCMV gene expression was unaffected, suggesting that LBV acts temporally coincident with HCMV DNA synthesis and not through cytotoxicity. In vitro, the triphosphate of LBV was a potent inhibitor of HCMV DNA polymerase with a Ki of 5 nM. LBV was phosphorylated to its triphosphate form intracellularly in both infected and uninfected cells, with phosphorylated metabolite levels two- to threefold higher in infected cells. GCV-resistant HCMV isolates, with deficient GCV phosphorylation due to mutations in the UL97 protein kinase, remained sensitive to LBV. Overall, these results suggest that LBV-triphosphate halts HCMV DNA replication by inhibiting the viral DNA polymerase and that LBV phosphorylation can occur in the absence of viral factors including the UL97 protein kinase. Furthermore, LBV may be effective in the treatment of GCV-resistant HCMV.

3′‐Fluorine‐Substitution in 3‐Fluorocarbocyclic Oxetanocin A Augments the Drug's Inhibition of HHV‐6B Propagation in Chronically Infected Cultures

“…It has been reported that GCV is significantly phosphorylated by HCMV UL97, a viral protein kinase, in the virus-infected cells (19,20,23,35), like the phosphorylation by HSV-l TK in the HSV-l-infected cells. The finding that the ECso of C.OXT-G in HCMV-infected cells as well as in UL97-deficient HCMV-infected cells was equivalent to that observed in HSV-l TK --infected cells (21,36) indicated that C.OXT-G was not efficiently phosphorylated by the viral kinases, including UL97, whereas the basal phosphorylation might consistently occur. Neither was additional phosphorylation of C.OXT-A expected in HCMV-infected cells because of the similar level of its ECso to that of C.OXT-G (21); however, 3'F-C.OXT-A exhibited a l-log-more potent ECso than that of C.OXT-A (29).…”

“…This suggested that C.OXTs might exert their antiviral activities mainly at steps preceding their incorporation into viral DNA strands. It has been reported previously that the triphosphate form of C.OXT-G, C.OXT-G-Tp, acts as a potent competitive inhibitor of HCMV polymerase, of which the Ki was estimated to 5 nM in contrast to the 0.22-14~M for human DNA polymerase-a; C.OXT-G-TP also acts as a potential chain terminator as a consequence of its incorporation into viral DNA strands (II, 15,36,37). The present study suggested that the inhibition of viral polymerase is very likely to be of greater importance than the chain termination function in accounting for the antiviral potency.…”

“…Among the C.OXTs, the guanosine analog carbocyclic oxetanocin G (C.OXT-G) has been extensively studied regarding its antiviral activity. The 50% effective concentration (EC so) of C.OXT-G against HCMV has been shown to be comparable to that of GCV against HSV (21,36), despite its lower level of phosphorylation in the HCMV-infected cells (36). The finding that the triphosphate form of C.OXT-G is a potent competitive inhibitor of HCMV polymerase with a 2-log-more potent Ki than that of triphosphorylated GCV (33,36), suggested that the antiviral potency of C.OXTs is based on their ability to inhibit viral DNA polymerases.…”

New Strategies for Prevention and Therapy of Cytomegalovirus Infection and Disease in Solid-Organ Transplant Recipients

“…Novel antiviral compounds such as benzimidazole (508), adefovir (504), and lobucavir (461) are currently under investigation and might further improve the management of CMV infection. Moreover, antisense oligonucleotides complementary to RNA transcripts of IE genes provide a novel mechanism of inhibition for viral replication (11).…”

MODERN ETHIOTROPIC CHEMOTHERAPY OF HERPESVIRUS INFECTIONS: ADVANCES, NEW TRENDS AND PERSPECTIVES. ALPHAHERPESVIRINAE (part I)

“…табл. 2), активен в отношении ВПГ-1, ВПГ-2, ВВЗ, ЦМВ, вируса гепатита В и ВИЧ [42][43][44]. Концентрации ЛБВ, эффективные в отношении ВПГ-1, ВПГ-2 и ЦМВ in vitro [42], ниже аналогичных величин для АЦВ.…”