scholarly journals Antiviral activity of the 3′;-amino derivative of (E)-5-(2-bromovinyl)-2′;-deoxyuridine

1983 ◽  
Vol 211 (2) ◽  
pp. 439-445 ◽  
Author(s):  
E De Clercq ◽  
J Descamps ◽  
J Balzarini ◽  
T Fukui ◽  
H S Allaudeen
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Thymidine Kinase ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Amino Derivative ◽  
Dna Polymerase Alpha ◽  
Simplex Virus ◽  
Hsv 1

3′-NH2-BV-dUrd, the 3′-amino derivative of (E)-5-(2-bromovinyl)-2′-deoxyuridine, was found to be a potent and selective inhibitor of herpes simplex virus type 1 (HSV-1) and varicella-zoster virus (VZV) replication. 3′-NH2-BV-dUrd was about 4-12 times less potent but equally selective in its anti-herpes activity as BV-dUrd. Akin to BV-dUrd, 3′-NH2-BV-dUrd was much less inhibitory to herpes simplex virus type 2 than type 1. It was totally inactive against a thymidine kinase-deficient mutant of HSV-1. The 5′-triphosphate of 3′-NH2-BV-dUrd (3′-NH2-BV-dUTP) was evaluated for its inhibitory effects on purified herpes viral and cellular DNA polymerases. Among the DNA polymerases tested, HSV-1 DNA polymerase and DNA polymerase alpha were the most sensitive to inhibition by 3′-NH2-BV-dUTP (Ki values 0.13 and 0.10 microM, respectively). The Km/Ki ratio for DNA polymerase alpha was 47, as compared with 4.6 for HSV-1 DNA polymerase. Thus, the selectivity of 3′-NH2-BV-dUrd as an anti-herpes agent cannot be ascribed to a discriminative effect of its 5′-triphosphate at the DNA polymerase level. This selectivity most probably resides at the thymidine kinase level. 3′;-NH2-BV-dUrd would be phosphorylated preferentially by the HSV-1-induced thymidine kinase (Ki 1.9 microM, as compared with greater than 200 microM for the cellular thymidine kinase), and this preferential phosphorylation would confine the further action of the compound to the virus-infected cell.

scholarly journals Differential Mutation Patterns in Thymidine Kinase and DNA Polymerase Genes of Herpes Simplex Virus Type 1 Clones Passaged in the Presence of Acyclovir or Penciclovir

2003 ◽  
Vol 47 (5) ◽  
pp. 1707-1713 ◽  
Author(s):  
Tatsuo Suzutani ◽  
Ken Ishioka ◽  
Erik De Clercq ◽  
Kei Ishibashi ◽  
Hisatoshi Kaneko ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Thymidine Kinase ◽  
Dna Polymerase ◽  
Herpes Simplex Virus Type ◽  
Nucleotide Substitution ◽  
Single Nucleotide ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT A total of 21 clones of acyclovir (ACV)-resistant (ACVr) herpes simplex virus type 1 (HSV-1) and 23 clones of penciclovir (PCV)-resistant (PCVr) HSV-1, emerging during serial passages in the presence of ACV or PCV, were isolated under conditions excluding contamination of resistant mutants in the starting virus culture, and their mutations in the thymidine kinase (TK) and DNA polymerase (DNA Pol) genes were analyzed comparatively. Mutations in the TK genes from ACVr mutants consisted of 50% single nucleotide substitutions and 50% frameshift mutations, while the corresponding figures for the PCVr mutants were 4 and 96%, respectively (P < 0.001). Eight of the 21 ACVr clones, but none of the 23 PCVr clones, had mutations in DNA Pol. Only nucleotide substitution(s) could be detected in the DNA Pol gene, as the gene is essential for virus replication. Therefore, the results for the DNA Pol mutants are concordant with those for the TK mutants in that a single nucleotide substitution was commonly observed in the ACVr, but not in the PCVr, mutants. These results clearly point to differential mutation patterns between ACVr and PCVr HSV-1 clones.

scholarly journals 3′ to 5′ Exonuclease Activity of Herpes Simplex Virus Type 1 DNA Polymerase Modulates Its Strand Displacement Activity

2003 ◽  
Vol 77 (18) ◽  
pp. 10147-10153 ◽  
Author(s):  
Yali Zhu ◽  
Kelly S. Trego ◽  
Liping Song ◽  
Deborah S. Parris
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dna Polymerase ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Strand Displacement ◽  
Displacement Activity ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Using a minicircle DNA primer-template, the wild-type catalytic subunit of herpes simplex virus type 1 (HSV-1) DNA polymerase (pol) was shown to lack significant strand displacement activity with or without its processivity factor, UL42. However, an exonuclease-deficient (exo−) pol (D368A) was capable of slow strand displacement. Although UL42 increased the rate (2/s) and processivity of strand displacement by exo− pol, the rate was slower than that for gap-filling synthesis. High inherent excision rates on matched primer-templates and rapid idling-turnover (successive rounds of excision and polymerization) of exo-proficient polymerases correlated with poor strand displacement activity. The results suggest that the exo activity of HSV-1 pol modulates its ability to engage in strand displacement, a function that may be important to the viability and genome stability of the virus.

scholarly journals Contrasting Effects of W781V and W780V Mutations in Helix N of Herpes Simplex Virus 1 and Human Cytomegalovirus DNA Polymerases on Antiviral Drug Susceptibility

2015 ◽  
Vol 89 (8) ◽  
pp. 4636-4644 ◽  
Author(s):  
Jocelyne Piret ◽  
Nathalie Goyette ◽  
Brian E. Eckenroth ◽  
Emilien Drouot ◽  
Matthias Götte ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Antiviral Drug ◽  
Drug Susceptibility ◽  
Herpes Simplex Virus 1 ◽  
Simplex Virus ◽  
Antiviral Drug Susceptibility ◽  
Hsv 1

ABSTRACTDNA polymerases of theHerpesviridaeand bacteriophage RB69 belong to the α-like DNA polymerase family. In spite of similarities in structure and function, the RB69 enzyme is relatively resistant to foscarnet, requiring the mutation V478W in helix N to promote the closed conformation of the enzyme to make it susceptible to the antiviral. Here, we generated recombinant herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV) mutants harboring the revertant in UL30 (W781V) and UL54 (W780V) DNA polymerases, respectively, to further investigate the impact of this tryptophan on antiviral drug susceptibility and viral replicative capacity. The mutation W781V in HSV-1 induced resistance to foscarnet, acyclovir, and ganciclovir (3-, 14-, and 3-fold increases in the 50% effective concentrations [EC50s], respectively). The recombinant HCMV mutant harboring the W780V mutation was slightly resistant to foscarnet (a 1.9-fold increase in the EC50) and susceptible to ganciclovir. Recombinant HSV-1 and HCMV mutants had altered viral replication kinetics. The apparent inhibition constant values of foscarnet against mutant UL30 and UL54 DNA polymerases were 45- and 4.9-fold higher, respectively, than those against their wild-type counterparts. Structural evaluation of the tryptophan position in the UL54 DNA polymerase suggests that the bulkier phenylalanine (fingers domain) and isoleucine (N-terminal domain) could induce a tendency toward the closed conformation greater than that for UL30 and explains the modest effect of the W780V mutation on foscarnet susceptibility. Our results further suggest a role of the tryptophan in helix N in conferring HCMV and especially HSV-1 susceptibility to foscarnet and the possible contribution of other residues localized at the interface between the fingers and N-terminal domains.IMPORTANCEDNA polymerases of theHerpesviridaeand bacteriophage RB69 belong to the α-like DNA polymerase family. However, the RB69 DNA polymerase is relatively resistant to the broad-spectrum antiviral agent foscarnet. The mutation V478W in helix N of the fingers domain caused the enzyme to adopt a closed conformation and to become susceptible to the antiviral. We generated recombinant herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV) mutants harboring the revertant in UL30 (W781V) and UL54 (W780V) DNA polymerases, respectively, to further investigate the impact of this tryptophan on antiviral drug susceptibility. The W781V mutation in HSV-1 induced resistance to foscarnet, whereas the W780V mutation in HCMV slightly decreased drug susceptibility. This study suggests that the different profiles of susceptibility to foscarnet of the HSV-1 and HCMV mutants could be related to subtle conformational changes resulting from the interaction between residues specific to each enzyme that are located at the interface between the fingers and the N-terminal domains.

scholarly journals Sequence Analysis of Herpes Simplex Virus 1 Thymidine Kinase and DNA Polymerase Genes from over 300 Clinical Isolates from 1973 to 2014 Finds Novel Mutations That May Be Relevant for Development of Antiviral Resistance

2015 ◽  
Vol 59 (8) ◽  
pp. 4938-4945 ◽  
Author(s):  
Susanne Schmidt ◽  
Kathrin Bohn-Wippert ◽  
Peter Schlattmann ◽  
Roland Zell ◽  
Andreas Sauerbrei
Keyword(s):  
Herpes Simplex Virus ◽  
Amino Acid ◽  
Herpes Simplex ◽  
Thymidine Kinase ◽  
Dna Polymerase ◽  
Herpes Simplex Virus 1 ◽  
Immunosuppressed Patients ◽  
Resistant Strains ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACTA total of 302 clinical herpes simplex virus 1 (HSV-1) strains, collected over 4 decades from 1973 to 2014, were characterized retrospectively for drug resistance. All HSV-1 isolates were analyzed genotypically for nonsynonymous mutations in the thymidine kinase (TK) and DNA polymerase (Pol) genes. The resistance phenotype against acyclovir (ACV) and/or foscarnet (FOS) was examined in the case of novel, unclear, or resistance-related mutations. Twenty-six novel natural polymorphisms could be detected in the TK gene and 69 in the DNA Pol gene. Furthermore, three novel resistance-associated mutations (two in the TK gene and one in the DNA Pol gene) were analyzed, and eight known but hitherto unclear amino acid substitutions (two encoded in TK and six in the DNA Pol gene) could be clarified. Between 1973 and 2014, the distribution of amino acid changes related to the natural gene polymorphisms of TK and DNA Pol remained largely stable. Resistance to ACV was confirmed phenotypically for 16 isolates, and resistance to ACV plus FOS was confirmed for 1 isolate. Acyclovir-resistant strains were observed from the year 1995 onwards, predominantly in immunosuppressed patients, especially those with stem cell transplantation, and the number of ACV-resistant strains increased during the last 2 decades. The data confirm the strong genetic variability among HIV-1 isolates, which is more pronounced in the DNA Pol gene than in the TK gene, and will facilitate considerably the rapid genotypic diagnosis of HSV-1 resistance.

A new N-methyl thymine derivative comprising a dihydroxyisobutenyl unit as ligand for thymidine kinase of herpes simplex virus type 1 (HSV-1 TK)

2011 ◽  
Vol 21 (20) ◽  
pp. 6161-6165 ◽  
Author(s):  
Svjetlana Krištafor ◽  
Ivana Novaković ◽  
Tatjana Gazivoda Kraljević ◽  
Sandra Kraljević Pavelić ◽  
Pero Lučin ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Thymidine Kinase ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Simplex Virus ◽  
Hsv 1

Interaction of 5-Methoxymethyl-2′-Deoxyuridine Triphosphate with DNA Polymerases: Effects of the 5-Substituent and Comparison with the Deoxycytidine Derivative

1992 ◽  
Vol 3 (4) ◽  
pp. 243-247 ◽  
Author(s):  
P. J. Aduma ◽  
S. V. Gupta ◽  
A. L. Stuart
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Competitive Inhibitor ◽  
E Coli ◽  
The Difference ◽  
Simplex Virus ◽  
Selective Activity ◽  
Hsv 1

5-Methoxymethyl-2′-deoxyuridine (MMdUrd) is a selective anti-herpes agent that is dependent upon initial phosphorylation by Herpes simplex virus-induced deoxythymidine kinase. In order to determine its mechanism of action, MMdUrd was converted to the 5′-triphosphate (MMdUTP) and the nature of interaction of MMdUTP and dTTP with DNA polymerase of E. coli, HSV-1, and human α was investigated. The order of utilization of deoxyuridine analogues by bacterial and HSV-1 DNA polymerases for DNA synthesis was: dTTP > MMdUTP. In contrast, 5-methoxymethyl-2′-deoxycytidine-5′-triphosphate (MMdCTP) was a better substrate for HSV DNA polymerase compared to dCTP. MMdUTP is a competitive inhibitor of HSV-1 DNA polymerase with respect to dTTP incorporation (Ki = 2.9 × 10−6M). The IC50 values of MMdUTP for both HSV and human αDNA polymerases were 4.5 × 10 −6M. These data suggest that the selective activity of MMdUrd is due to its preferential phosphorylation by viral thymidine kinase and not at the DNA polymerase level. These results may also account for the difference in anti-HSV activity between MMdUrd and its deoxycytidine analogue.

scholarly journals The Epstein-Barr Virus Thymidine Kinase Does Not Phosphorylate Ganciclovir or Acyclovir and Demonstrates a Narrow Substrate Specificity Compared to the Herpes Simplex Virus Type 1 Thymidine Kinase

1998 ◽  
Vol 42 (11) ◽  
pp. 2923-2931 ◽  
Author(s):  
Erik A. Gustafson ◽  
Antoinette C. Chillemi ◽  
David R. Sage ◽  
Joyce D. Fingeroth
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Substrate Specificity ◽  
Thymidine Kinase ◽  
Epstein Barr Virus ◽  
Barr Virus ◽  
Epstein Barr ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT The Epstein-Barr virus (EBV) thymidine kinase (TK) was expressed in mammalian 143B TK− cells to investigate its substrate specificity. The herpes simplex virus type 1 (HSV-1) TK was similarly expressed for comparison. Both viral TKs conferred a TK+ phenotype on 143B TK− cells. The nucleoside analog ganciclovir (GCV) did not affect the growth of 143B EBV TK or 143B TK− cells but effectively killed 143B HSV-1 TK cells. Furthermore, lysates of 143B EBV TK cells could not phosphorylate GCV, which was confirmed by high-performance liquid chromatography. EBV TK, HSV-1 TK, and EBV TK N−, a truncated EBV TK missing 243 N-terminal amino acids, were purified as fusion proteins expressed in bacteria, and all had TK activity. In addition, EBV TK was observed to have a thymidylate kinase activity but could not phosphorylate GCV, acyclovir, or 2′-deoxycytidine. In competition assays, only nucleoside analogs of thymidine significantly inhibited thymidine phosphorylation by EBV TK, with the following rank order: 5-bromodeoxyuridine > zidovudine > stavudine > sorivudine. These results demonstrate that EBV TK substrate specificity is narrower than those of alphaherpesvirus TKs and that thymidine analogs may be the most suitable nucleoside antivirals to target the enzyme. Clinical implications for gammaherpesviruses are discussed.

scholarly journals Evidence of Muller’s ratchet in herpes simplex virus type 1

2013 ◽  
Vol 94 (2) ◽  
pp. 366-375 ◽  
Author(s):  
Nacarí Jaramillo ◽  
Esteban Domingo ◽  
María Carmen Muñoz-Egea ◽  
Enrique Tabarés ◽  
Ignacio Gadea
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Dna Polymerase ◽  
Herpes Simplex Virus Type ◽  
Rna Viruses ◽  
Population Bottlenecks ◽  
Dna Viruses ◽  
Simplex Virus ◽  
Hsv 1

Population bottlenecks can have major effects in the evolution of RNA viruses, but their possible influence in the evolution of DNA viruses is largely unknown. Genetic and biological variation of herpes simplex virus type 1 (HSV-1) has been studied by subjecting 23 biological clones of the virus to 10 plaque-to-plaque transfers. In contrast to large population passages, plaque transfers led to a decrease in replicative capacity of HSV-1. Two out of a total of 23 clones did not survive to the last transfer in 143 TK– cells. DNA from three genomic regions (DNA polymerase, glycoprotein gD and thymidine kinase) from the initial and passaged clones was sequenced. Nucleotide substitutions were detected in the TK and gD genes, but not in the DNA polymerase gene. Assuming a uniform distribution of mutations along the genome, the average rate of fixation of mutations was about five mutations per viral genome and plaque transfer. This value is comparable to the range of values calculated for RNA viruses. Four plaque-transferred populations lost neurovirulence for mice, as compared with the corresponding initial clones. LD50 values obtained with the populations subjected to serial bottlenecks were 4- to 67-fold higher than for their parental clones. These results equate HSV-1 with RNA viruses regarding fitness decrease as a result of plaque–to-plaque transfers, and show that population bottlenecks can modify the pathogenic potential of HSV-1. Implications for the evolution of complex DNA viruses are discussed.

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