By measuring the relative inhibitory concentrations required to overcome particular mutations in the helicase and primase proteins, evidence was obtained that BAY 57-1293 interacts with both components of the helicase-primase complex to achieve maximum potency, whereas for BILS 22BS, this may not be the case. Furthermore, our observations suggest that BAY 57-1293 interacts simultaneously with UL5 and UL52. Overall, the results suggest that these two potent HPIs interact differently with the helicase-primase complex.
The authors wish to draw attention to the following errors in their article:
On page 15 an incorrect sequence database was referenced. The sentence ‘A set of eight pairs of overlapping HSV-1-specific primers (designed on the basis of HSV-1 strain 17 sequence; Genbank accession number NC001806) was used (McGeoch et al., 1988).’ should read ‘A set of eight pairs of overlapping HSV-1-specific primers (designed on the basis of HSV-1 strain 17 sequence; RefSeq accession number NC001806) was used (McGeoch et al., 1988).’
On Table 2 an incorrect symbol was used: ‘$reported by Liuzzi et al., 2004’ should be replaced by ‘♠reported by Liuzzi et al., 2004’. A corrected table can be found below.
On Table 3 the 5th column heading is incorrect: ‘BAY 57–1293 concentration, mg/ml’ should read ‘BAY 57–1293 concentration µg/ml’. A corrected table can be found below.
Table 2.Observed frequency of BAY 57-1293 and other HPI-resistant variants in various laboratory isolates of HSV-1 and mutations in the helicase (UL5) protein
*Frequency expressed as observed number of drug-resistant plaque-forming units (PFU) in the total number of PFU screened;
†reported by Kleymann et al., 2002;
‡SC16 crude stock reproducibly contained >10× more drug-resistant variants than that observed for plaque-purified SC16 cl-2;
§novel substitutions, not previously reported as BAY 57-1293-resistant mutations;
¶PDK crude stock reproducibly contained >70 × more drug-resistant variants than that observed for plaque-purified PDK cl-1;
♠reported by Liuzzi et al., 2004;
$reported by Spector et al., 1998. Bold type indicates the BAY 57-1293-resistant variants occurring at high frequency and the corresponding laboratory isolates in which they were detected. ED50, 50% effective dose; HPI, helicase-primase inhibitor; HSV, herpes simplex virus; n/s, not specified.
Table 3.A summary of BAY 57-1293-resistant variants showing the methods used for their selection
Fifty percent effective dose (ED50)data was determined by plaque reduction assay in Vero cells and was significantly different from respective wild-type in an un-paired 2-tailed t-test (P<0.001).
*From Figure 6A;
†from Figure 6B;
‡from Figure 6C;
§from Figure 6D.
Herpes simplex virus (HSV) helicase-primase is the target for a new group of potent antivirals that show great promise in vivo. A claimed advantage of this class of compounds is the low rate of drug resistance, which is reported to occur at a lesser rate than acyclovir (ACV)-resistance in cell culture. We confirmed that BAY 57-1293 is highly active against HSV-1 and superior to ACV when tested in Vero cells. Notably, drug resistance was detected in laboratory working stocks in two different strains of HSV at 10(-4) to 10(-5) and there was evidence that the resistant variants were present in the virus population before the selection was applied. Plaque-purified clones obtained from the parental viruses showed a lower level of resistance selection in the presence of drug (10-6) and this value is similar to published reports. In the case of HSV-1 SC16, no difference was observed between a working stock and a plaque-pure clone in the rate of resistance to the nucleoside analogue ACV. The working stocks were found to contain variants with resistance to BAY 57-1293 ranging from approximately 15-fold to 4,000-fold suggesting that these viruses have the potential to subvert effective therapy. Sequence analysis of HSV-1 helicase protein showed that most of the amino acid substitutions in the variants described in this study tallied with published results, with some interesting exceptions in the case of HSV-1 strain PDK. Resistant variants did not readily revert to a sensitive phenotype in the absence of the inhibitor and representative BAY 57-1293-resistant variants were cross-resistant to an alternative helicase-primase inhibitor, BILS 22 BS. Variants resistant to BAY 57-1293 retained sensitivity to the nucleoside analogue, ACV.
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