Review
Herpes simplex virus resistance to antiviral drugs

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Abstract

Herpes simplex virus (HSV) infections are efficiently treated with antiviral drugs such as acyclovir (ACV). However, resistance has been reported, mainly among immunocompromised patients (prevalence around 5%) and particularly allogeneic bone marrow transplant patients (prevalence reaching 30%). Resistance to ACV is associated with mutations on one of the two viral enzymes involved in the ACV mechanism of action: thymidine kinase (TK) and DNA polymerase. In 95% of the cases, ACV resistance is associated with a mutation in the TK gene as this enzyme is not essential for viral replication, unlike viral DNA polymerase, which is rarely involved in resistance. Strains resistant to ACV are almost always cross-resistant to other TK-dependent drugs such as penciclovir and famciclovir. Resistant infections can be managed by foscarnet or cidofovir but both are more toxic than ACV. These drugs also inhibit viral DNA polymerase but they are active on most ACV-resistant HSV as they do not depend on TK; nevertheless virus resistant to ACV because of a mutation in the DNA polymerase may be cross-resistant to these molecules. Published data on genetic characterization of resistant clinical isolates point out hot spots in viral TK and DNA polymerase genes. TK mutations associated with resistance are either insertion or deletion (codons 92 and 146 of TK gene) or substitution (codon 176–177, 336 of TK gene). DNA polymerase mutations are mainly located in conserved sites of the enzyme. A high level of gene polymorphism has also been reported for these genes, especially for TK. These results are useful for the development of rapid genotypic assays for the detection of mutations associated with resistance.

Introduction

Herpes simplex virus (HSV) infections are common and may be responsible for serious complications among immunocompromised patients. The treatment of choice is acyclovir (ACV) which is a nucleoside analogue of guanosine that has to be phosphorylated three times. The first phosphorylation is completed by the viral encoded thymidine kinase (TK) protein which allows ACV to become active only in virus infected cells. The second and third phosphorylations are achieved by cellular thymidylate kinases. ACV triphosphate acts by competitive inhibition of viral DNA polymerase and it is a DNA chain terminator (Elion, 1993).

This widespread use of ACV has lead to the emergence of HSV strains resistant to ACV. Resistant strains were soon reported and the first cases of clinical resistant strains were published in 1982 (Sibrack et al., 1982, Crumpacker et al., 1982, Burns et al., 1982). Large surveys were conducted and revealed the low incidence of ACV-resistant strains among immunocompetent patients, from 0 to 0.6% (Englund et al., 1990, Nugier et al., 1992, Christophers et al., 1998). Conversely ACV-resistant strains were most often recovered from immunocompromised patients with a whole frequency ranging from 3 to 6%, that reaches 14% among bone marrow transplant recipients (Englund et al., 1990, Nugier et al., 1992, Christophers et al., 1998).

This review deals with clinical and epidemiological features associated with the emergence of ACV-resistant HSV, mechanisms of resistance, management of ACV-resistant HSV infections and presents an analysis of recently published data on mutations associated with resistance.

Section snippets

Epidemiology

Among immunocompetent patients, resistance to ACV is rare. Several reports have described a prevalence below 1% in this population (Nugier et al., 1992, Christophers et al., 1998). In our hospital laboratory, all HSV isolated were tested for their sensitivity to ACV; ACV-resistant strains have never been detected among immunocompetent patients during more than 10 years of surveillance (more than 500 isolates tested) (unpublished results).

According to the literature, most ACV-resistant HSV

Mechanism of resistance

The majority of antiherpetic drugs in clinical use are nucleoside analogues. ACV is a guanosine analogue, as is penciclovir, whereas cidofovir is a phosphonate molecule derived from cytidine. Foscarnet presents a very different structure, analogous to a pyrophosphate.

ACV and penciclovir mechanism of action involves two viral enzymes: TK for the first phosphorylation of the activation step and DNA polymerase, which is the target of the triphosphate form. For cidofovir, only two phosphorylations

Management of acyclovir-resistant HSV infections

To manage ACV-resistant infections, several antiviral drugs may be used. Most of ACV-resistant HSV isolates are also resistant to penciclovir although rarely, some isolates resistant to ACV but susceptible to penciclovir have been reported; the mechanism of resistance to ACV of these strains was either an altered TK (Boyd et al., 1993) or a mutation in viral DNA polymerase (Chiou et al., 1995). Foscarnet and cidofovir act directly on viral DNA polymerase without previous activation by viral TK

In vitro detection of resistance

In vitro evaluation of HSV susceptibility to antiviral drugs is based on the determination of viral replication inhibition in the presence of increasing concentrations of antiviral drug. Three techniques are available to reveal viral replication: plaque reduction assay (PRA) which is the reference technique, DNA hybridization test (Swierkosz et al., 1987) and dye uptake method (Langlois et al., 1986). The two last techniques are less time consuming as the reading of cytopathic effect is

Thymidine kinase mutations associated with resistance

Our interest has first been focused on HSV TK, which is involved in 95% of ACV resistance cases. HSV TK is a 376 amino acid protein that is encoded by a gene of 1128 bp (UL 23; McKnight, 1980). It exhibits an ATP binding site (amino acid 51–63) and a nucleoside binding site (amino acid 168–176) as proposed by Darby et al. (1986). Six regions conserved among herpesviridae TK have also been located at amino acids 50–66, 79–91, 162–178, 212–226 and 281–292 (Balasubramaniam et al., 1990).

Fig. 2

DNA polymerase mutations associated with resistance

HSV DNA polymerase is a 1235 amino acid protein encoded by a 3705 bp gene (UL 30). It includes eight conserved regions in comparison with cellular or viral DNA polymerases; these regions are labeled I–VII according to their degree of conservation (region I being the most conserved), plus one region called A (Wong et al., 1988).

Mutations associated with resistance described in the literature are presented in the upper part of Fig. 3. These data are related to genetic characterization of about 15

Conclusion

The increasing use of ACV, especially in prophylaxis treatments among transplanted patients, has raised the fear of an increasing incidence of ACV-resistant infections. Recent survey studies have shown that this is not the case and that ACV resistance is mainly a concern for severely immunocompromised patients, such as those transplanted with bone marrow from allogeneic origin. When managing ACV-resistant infections, other antiviral drugs with different mechanisms of action may be used, such as

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