title: Human cytomegalovirus resistance to deoxyribosylindole nucleosides maps to a transversion mutation in the terminase subunit-encoding gene UL89.
abstract: Human cytomegalovirus (HCMV) infection can cause severe illnesses, including encephalopathy and mental retardation, in immunocompromised and immunologically immature patients. Current pharmacotherapies for treating systemic HCMV infections include ganciclovir, cidofovir, and foscarnet. However, long-term administration of these agents can result in serious adverse effects (myelosuppression and/or nephrotoxicity) and the development of viral strains with reduced susceptibility to drugs. The deoxyribosylindole (indole) nucleosides demonstrate a 20-fold greater activity in vitro (the drug concentration at which 50% of the number of plaques was reduced with the presence of drug compared to the number in the absence of drug [EC50] = 0.34 μM) than ganciclovir (EC50 = 7.4 μM) without any observed increase in cytotoxicity. Based on structural similarity to the benzimidazole nucleosides, we hypothesize that the indole nucleosides target the HCMV terminase, an enzyme responsible for packaging viral DNA into capsids and cleaving the DNA into genome-length units. To test this hypothesis, an indole nucleoside-resistant HCMV strain was isolated, the open reading frames of the genes that encode the viral terminase were sequenced, and a G766C mutation in exon 1 of UL89 was identified; this mutation resulted in an E256Q change in the amino acid sequence of the corresponding protein. An HCMV wild-type strain, engineered with this mutation to confirm resistance, demonstrated an 18-fold decrease in susceptibility to the indole nucleosides (EC50 = 3.1 ± 0.7 μM) compared to that of wild-type virus (EC50 = 0.17 ± 0.04 μM). Interestingly, this mutation did not confer resistance to the benzimidazole nucleosides (EC50 for wild-type HCMV = 0.25 ± 0.04 μM, EC50 for HCMV pUL89 E256Q = 0.23 ± 0.04 μM). We conclude, therefore, that the G766C mutation that results in the E256Q substitution is unique for indole nucleoside resistance and distinct from previously discovered substitutions that confer both indole and benzimidazole nucleoside resistance (D344E and A355T).
authors.affiliation: Department of Pharmaceutical, Biomedical and Administrative Sciences, College of Pharmacy and Health Sciences, Drake University, Des Moines, Iowa, USA brian.gentry@drake.edu.
authors.forename: Brian G
authors.initials: BG
authors.lastname: Gentry
authors.affiliation: Department of Pharmaceutical, Biomedical and Administrative Sciences, College of Pharmacy and Health Sciences, Drake University, Des Moines, Iowa, USA.
authors.forename: Quang
authors.initials: Q
authors.lastname: Phan
authors.affiliation: Department of Microbiology and Immunology, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Ellie D
authors.initials: ED
authors.lastname: Hall
authors.affiliation: Department of Biologic and Materials Sciences, School of Dentistry, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Julie M
authors.initials: JM
authors.lastname: Breitenbach
authors.affiliation: Department of Biologic and Materials Sciences, School of Dentistry, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Katherine Z
authors.initials: KZ
authors.lastname: Borysko
authors.affiliation: Department of Microbiology and Immunology, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Jeremy P
authors.initials: JP
authors.lastname: Kamil
authors.affiliation: Department of Medicinal Chemistry, School of Pharmacy, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Leroy B
authors.initials: LB
authors.lastname: Townsend
authors.affiliation: Department of Biologic and Materials Sciences, School of Dentistry, University of Michigan, Ann Arbor, Michigan, USA Department of Medicinal Chemistry, School of Pharmacy, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: John C
authors.initials: JC
authors.lastname: Drach
mesh_terms: D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000494:pharmacology; D001483:Base Sequence; D001562:Benzimidazoles / Q000494:pharmacology*; D003587:Cytomegalovirus / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D003853:Deoxyribonucleosides / Q000494:pharmacology*; D024882:Drug Resistance, Viral / Q000235:genetics*; D007211:Indoles / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009154:Mutation; D012263:Ribonucleosides / Q000494:pharmacology*; D014764:Viral Proteins / Q000235:genetics*
chemical_list: C109864:2-bromo-5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole; C000600717:3-acetyl-2-bromo-5,6-dichloro-1-(2-deoxyribofuranosyl)indole; D000998:Antiviral Agents; D001562:Benzimidazoles; D003853:Deoxyribonucleosides; D007211:Indoles; D012263:Ribonucleosides; C436448:UL89 protein, Cytomegalovirus; D014764:Viral Proteins