Jean Bedard

Evaluation of VCH-759 monotherapy in hepatitis C infection

BACKGROUND/AIMS VCH-759 is a non-nucleoside inhibitor of HCV RNA-dependent polymerase with sub-micromolar IC(50) values versus genotype 1a/1b replicons. METHODS The antiviral activity, pharmacokinetics and tolerability of VCH-759 administered as monotherapy for 10 days with a 14 day follow-up period were evaluated in 31 treatment-nai ve genotype 1 participants. Three cohorts received: 400mg thrice (t.i.d.), 800 mg twice (b.i.d.), 800 mg t.i.d or placebo. RESULTS VCH-759 was well tolerated with the most frequent adverse event being gastrointestinal upset in both the active and placebo groups attributable, in part, to the dosing vehicle. VCH-759 was rapidly absorbed and trough plasma levels were at or above the IC(90) (non protein-adjusted) for all dosing regimens. The mean maximal decrease in HCV RNA log(10) (IU/mL) was 1.97, 2.30 and 2.46 for 400mg t.i.d., 800 mg b.i.d. and 800 mg t.i.d. doses. Viral polymerase genotypic sequencing revealed emergence of HCV variants in a majority of participants that coincided with on-treatment viral rebound. CONCLUSIONS VCH-759 was well tolerated and achieved a> or =2 log(10) decline in HCV RNA with 800 mg b.i.d. and t.i.d doses. In a subset of participants, viral rebound was observed and associated with resistant variants. This data supports further evaluation of VCH-759 in combination with interferon-ribavirin treatment.

Antiviral properties of a series of 1,6-naphthyridine and 7, 8-dihydroisoquinoline derivatives exhibiting potent activity against human cytomegalovirus.

ABSTRACT A series of 1,6-naphthyridine (L. Chan, H. Jin, T. Stefanac, J. F. Lavallee, G. Falardeau, W. Wang, J. Bedard, S. May, and L. Yuen, J. Med. Chem. 42:3023–3025, 1999) and isoquinoline (L. Chan, H. Jin, T. Stefanac, W. Wang, J. F. Lavallee, J. Bedard, and S. May, Bioorg. Med. Chem. Lett. 9:2583–2586, 1999) analogues exhibiting a high level of anti-human cytomegalovirus (HCMV) activity were investigated in a series of studies aimed at better understanding the mechanism of action of some representatives of this class of compounds. In vitro antiviral profiling revealed that these compounds were active against a narrow spectrum of viruses, essentially the human herpesviruses and type 2 rhinovirus. In HCMV assays, a 39- to 223-fold lower 50% inhibitory concentration was obtained for compound A1 than for ganciclovir against strains AD 169 and Towne. In addition, ganciclovir, foscarnet, cidofovir, and BDCRB (2-bromo-5,6-dichloro-1-β-d-ribofuranosylbenzimidazole)-resistant HCMV strains remained susceptible to 1,6-naphthyridines and 7,8-dihydroisoquinolines tested in this study, supporting the view that a novel mechanism of action could be involved. Drug combination studies showed a small but significant synergistic antiviral effect between compound B2 and ganciclovir. Cytotoxicity profiling of representative compounds under various cell growth conditions indicated a generally similar cytotoxic effect, relative to ganciclovir, in log-phase growing cells. However, in stationary cells, a relatively higher level of toxicity was observed than that for control compound. Effect of time of drug addition showed that the anti-HCMV activity of compound A1, ganciclovir, and cidofovir was lost at approximately the same time (72 h postinfection), indicating that the compound was affecting events at the early and late stage of virus replication. This interpretation is also supported by reduction of de novo synthesis of pp65 tegument protein and lack of any effect of the compound on viral adsorption. A reduction of the HCMV enhancer-promoter-directed luciferase expression was also observed in a stably transfected cell line when compound A1 was present at relatively high concentrations.

A high throughput colorimetric cell proliferation assay for the identification of human cytomegalovirus inhibitors

A colorimetric assay based on the cleavage of the tetrazolium salt WST-1 has been developed for human cytomegalovirus (HCMV) antiviral susceptibility testing and adapted to a microtiter plate format. Optimal conditions were determined and the standard routine assay was calibrated with a viral input of 0.05-0.10 plaque forming unit (p.f.u.)/cell with a density of 2000 cells/well in a 96-well microtiter plate for an incubation period of 7 days. Ganciclovir (9-(2-hydroxy-1(hydroxymethyl) ethyoxymethyl) guanine; DHPG), and cidofovir ((S)-1-(3-hydroxy-2-phosphonylmethoxypropyl) cytosine; HPMPC) were used as positive control test compounds to validate the assay. The effective EC50 concentration values obtained with the two antiviral compounds in the present assay were in good agreement with plaque reduction assay results performed in parallel experiments. This method presents the advantage of being easy and rapid to perform, reliable, reproducible, and convenient for use in a high throughput screening capacity.

Isoquinoline-6-carboxamides as potent and selective anti-human cytomegalovirus (HCMV) inhibitors.

Structure-activity relationship studies on our newly identified anti-HCMV compounds, the 1,6-naphthyridines led to the identification of isoquinoline-6-carboxamides as potent and selective anti-HCMV agents.

Identification of novel nucleotide phosphonate analogs with potent anti-HCMV activity.

We have recently described the discovery of new leads in the area of anti-HCMV research. Further structure-activity relationship studies have allowed us to identify potent and selective anti-HCMV nucleotide analogs. The synthesis as well as structure-activity relationship studies are described.

844 GENOTYPIC ANALYSIS OF HCV NS5B VARIANTS SELECTED FROM PATIENTS TREATED WITH VCH-759

Background: VCH-759 is a novel orally bioavailable non-nucleoside inhibitor of hepatitis C virus RNA-dependent RNA polymerase. In a phase Ib clinical study involving genotype 1a and 1b-infected subjects, VCH-759 has achieved a 2 log10 or greater decline in HCVRNA levels. During the 10-day dosing period, an initial rapid reduction of viral load was observed in all patients dosed with VCH-759. Some patients have experienced a sustained anti-viral response during treatment but viral breakthrough was noticed for others. The aim of this study was to genotypically and phenotypically characterize the selection of resistant variants and to investigate a potential correlation with the viral kinetics observed. Methods: Selected samples collected at day 1 (pre-dosing), day 11 (day after last dose), and days 17 and 24 (short term follow-up) were used for analysis. The entire NS5B gene was amplified by RT-PCR and directly sequenced. In order to determine the prevalence of each mutation within the population, the region corresponding to the compound binding pocket (amino acids 340–539) was also amplified, cloned, and numerous clones sequenced. The phenotypic and the in vitro replication capacity analysis of variants were performed using the HCV replicon system. Results: Analysis of pre-dosing sera has revealed a wild-type genotype for the positions 419, 423, and 482 of the NS5B when compared to the consensus sequence of genotypes 1a and 1b. A mixture of wild-type and M423T/V/I substitution was the common feature found by population sequencing in samples at day 11, 17, and 24. The M423T and M423V mutants were associated with a respective 18 to 21-fold increase in replicon EC50 values when compared to the wild-type. A less predominant mutation namely, L419M was detected by clonal analysis and was found to confer a 23-fold reduction in VCH-759 susceptibility. Overall, the selected mutants were found to have a reduced replication capacity when compared to the wild-type. Conclusions: HCV variants with mutations conferring resistance to VCH759 were selected during the course of a 10-day treatment. This study illustrates that VCH-759 should be used in a combination therapy to maintain viral suppression and prevent emergence of resistance.

Design and synthesis of new potent human cytomegalovirus (HCMV) inhibitors based on internally hydrogen-bonded 1,6-naphthyridines.

1,6-Naphthyridine-2-carboxylic acid benzylamides are potent anti-HCMV compounds. Replacement of the amide moiety by other groups containing internal hydrogen bonds was undertaken to extend the SAR. Our results indicated that the urca derivatives showed very good activity.

Design and evaluation of dihydroisoquinolines as potent and orally bioavailable human cytomegalovirus inhibitors.

Following the identification of first pass metabolism issues with our recently described anti-HCMV compounds, the naphthyridines and isoquinolines, we have designed a class of novel metabolically stable and orally bioavailable anti-HCMV agents, the dihydroisoquinolines.