New HCV Antiviral in the Pipeline

New HCV Antiviral in the Pipeline

excertped from new Hepatology textbook recently published

Table 1. Antivirals in the pipeline.

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HCV life cycle and targets for STAT-C

HCV is a positive-sense single-stranded RNA virus of approximately 9600 nucleotides.

The HCV genome contains a single large open reading frame encoding for

a polyprotein of about 3100 amino acids. From this initially translated polyprotein,

the structural HCV protein core (C) and envelope 1 and 2 (E1, E2); p7; and the

six non-structural HCV proteins NS2, NS3, NS4A, NS4B, NS5A and NS5B, are

processed by both viral and host proteases. The core protein forms the viral nucleocapsid

carrying E1 and E2, which are receptors for viral attachment and host cell

entry. The non-structural proteins are mainly enzymes essential for the HCV life

cycle (Bartenschlager 2004; Pawlotsky 2007). P7 is a small hydrophobic protein

that oligomerises into a circular hexamer, most likely serving as an ion channel

through the viral lipid membrane (Carrere-Kremer 2002; Clarke 2006). The large

translated section of the HCV genome is flanked by the strongly conserved HCV

3´ and 5´ untranslated regions (UTR). The 5´ UTR is comprised of four highly

structured domains forming the internal ribosome entry site (IRES), which plays an

important role in HCV replication (Figure 2).

Figure 2. Genomic organisation of HCV.
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Figure 9. Antiviral activity of NS3/4A protease inhibitors.

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Resistance to NS3/4A inhibitors

Because of the high replication rate of HCV and the poor fidelity of its RNA-dependent

RNA polymerase, numerous variants (quasispecies) are continuously produced

during HCV replication. Among them, variants carrying mutations altering the conformation

of the binding sites of STAT-C compounds can develop. During treatment

with specific antivirals, these drug-resistant variants have a fitness advantage and

can be selected to become the dominant viral quasispecies. Many of these resistant

mutants exhibit an attenuated replication with the result that, after stopping exposure

to specific antivirals, the wild type may displace the resistant variants (Tong

2006; Sarrazin 2007). Nevertheless, HCV quasispecies resistant to NS3/4A protease

inhibitors or non-nucleoside polymerase inhibitors can be detected at low levels in

some patients who were never treated with specific antivirals before (Gaudieri 2009;

Kuntzen 2008; Rodriguez-Frias 2009; Le Pogam 2008). The clinical relevance of

these pre-existing mutants is not completely understood, although there is evidence

that they may reduce the chance of achieving an SVR after treatment with STAT-C

compounds.

Table 5. Resistance mutations to HCV NS3 protease inhibitors.

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* mutations associated with resistance in vitro but not described in patients.

Telaprevir

To date, mutations conferring telaprevir-resistance have been identified at four positions,

V36A/M/L, T54A, R155K/M/S/T and A156S//T (Lin 2005; Lin 2007; Sarrazin

2007; Welsch 2008; Zhou 2008) (Table 5). The A156 mutation was revealed

by in vitro analyses in the replicon while the other mutations were detected in vivo

by a clonal sequencing approach during telaprevir administration in patients with

chronic hepatitis C. A detailed kinetic analysis of telaprevir-resistant variants was

performed in genotype 1 patients during 14 days of telaprevir monotherapy and

combination therapy with PEG-IFN a-2a (Sarrazin 2007). Telaprevir monotherapy

initially led to a rapid HCV RNA decline in all patients due to a strong reduction in

wild type virus. In patients who developed a viral rebound during telaprevir monotherapy,

mainly the single mutation variants R155K/T and A156T were uncovered

by wild type reduction and became dominant after day 8. These single mutant

variants were selected from preexisting quasispecies. During the viral rebound

phase these variants typically were replaced by highly resistant double-mutation

variants (e.g., V36M/A +R155K/T). The combination of telaprevir and PEG-IFN

a-2a was sufficient to inhibit the breakthrough of resistant mutations in a 14-day

study (Forestier 2007). It is important to note that after up to 3 years of telaprevir

treatment low to medium levels of V36 and R155 variants were observed in single

patients (Forestier 2008).

As shown also for other NS3/4A protease inhibitors (e.g., ITMN-191), the genetic

barrier to telaprevir resistance differs significantly between HCV subtypes. In all clinical

studies of telaprevir alone or in combination with PEG-IFN a and ribavirin, viral

resistance and breakthrough occurred much more frequently in patients infected with

HCV genotype 1a compared to genotype 1b. This difference was shown to result from

nucleotide differences at position 155 in HCV subtype 1a (aga, encodes R) versus

1b (cga, also encodes R). The mutation most frequently associated with resistance to

telaprevir is R155K; changing R to K at position 155 requires 1 nucleotide change in

HCV subtype 1a and 2 nucleotide changes in subtype 1b isolates (McCown 2009).

Boceprevir

In the replicon system, mutations have been seen at three positions that confer boceprevir

resistance (Table 5). T54A, A156S and V170A confer low level resistance to

boceprevir whereas A156T, which also confers telaprevir and ciluprevir resistance,

exhibits greater levels of resistance (Tong 2006). In patients with chronic hepatitis

C three additional mutations were detected during boceprevir monotherapy (V36G/

M/A, V55A, R155K) (Susser 2009). In a number of these patients at one year and in

single patients at even 4 years after stopping boceprevir treatment resistant variants

could still be detected in the HCV quasispecies by clonal sequence analysis (Susser

2009). However, another study revealed that the antiviral activity of boceprevir was

not different in people whether they had or had not been previously treated with

PEG-IFN a (Vermehren 2009).


Compounds targeting HCV replication

NS5B polymerase inhibitors

NS5B RNA polymerase inhibitors can be divided into two distinct categories. Nucleoside

analogue inhibitors (NIs) like valopicitabine (NM283), R7128, R1626, PSI-7851

or IDX184 mimic the natural substrates of the polymerase and are incorporated into

the growing RNA chain, thus causing direct chain termination by blocking the active

site of NS5B (Koch 2006; Koch 2007). Because the active centre of NS5B is a highly

conserved region of the HCV genome, NIs are potentially effective against different

genotypes. Single amino acid substitutions in every position of the active centre may

result in loss of function. Thus, there is a relatively high genetic barrier in the development

of resistances to NIs.

In contrast to NIs, the heterogeneous class of non-nucleoside inhibitors (NNIs)

achieves NS5B inhibition by binding to different allosteric enzyme sites, which results

in conformational protein change before the elongation complex is formed (Beaulieu

2007). For allosteric NS5B inhibition high chemical affinity is required. NS5B is

structurally organized in a characteristic “right hand motif”, containing finger, palm

and thumb domains, and offers at least four NNI binding sites, a benzimidazole-

(thumb 1)-, thiophene-(thumb 2)-, benzothiadiazine-(palm 1)- and benzofuran-(palm

2)-binding site (Lesburg 1999; Beaulieu 2007) (Figure 12). Because of their distinct

binding sites, different polymerase inhibitors can theoretically be used in combination

or in sequence to manage the development of resistance. Because NNIs bind distantly

to the active centre of NS5B, their application may rapidly lead to the development of

resistant mutants in vitro and in vivo. Moreover, mutations at the NNI binding sites do

not necessarily lead to impaired function of the enzyme.


Figure 13. Antiviral activity of nucleoside analogue NS5B polymerase inhibitors.

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Non-nucleoside analogs

At least 4 different allosteric binding sites have been identified for the inhibition of the

NS5B polymerase by non-nucleoside inhibitors. An overview of the antiviral activities of

non-nucleoside polymerase inhibitors in monotherapy studies is shown in Figure 14.

NNI site 1 inhibitors (thumb 1 / benzimidazole site)

BILB1941, BI207127 and MK-3281 are NNI site 1 inhibitors investigated in phase

I clinical trials and have shown little to modest antiviral activity (Erhard 2009; Shi

2009; Sarrazin 2009). No viral breakthrough via selection of resistant variants was

seen after 5 days of treatment with BILB1941 or BI207127.

NNI site 2 inhibitors (thumb 2 / thiophene site)

Filibuvir (PF-00868554) is a NNI site 2 inhibitor with modest antiviral activity in a

phase I study. In a subsequent triple therapy trial with filibuvir, pegylated interferon

a-2a and ribavirin for 4 weeks viral breakthrough was observed in 5/26 patients.

VCH-759, VCH-916 and VCH-222 are three other NNI site 2 inhibitors with

antiviral activity in monotherapy studies (Cooper 2009; Sarrazin 2009). For VCH-

759 as well as VCH-916 viral breakthroughs via selection of resistant variants

were observed.

NNI site 3 inhibitors (palm 1 / benzothiadiazine site)

ANA598 is a NNI site 3 inhibitor that displayed antiviral activity during treatment of

genotype 1 infected patients. Viral breakthrough was not observed during this short

monotherapy trial.

NNI site 4 inhibitors (palm 2 / benzofuran site)

Monotherapy with the NNI site 4 inhibitor HCV-796 showed low antiviral activity

in genotype 1 infected patients (Kneteman 2009; Villano 2007). Viral breakthrough

was associated with selection of resistant variants conferring a medium to high level

of phenotypic resistance. For GS-9190 low antiviral activity was observed in a clinical

study and variants conferring resistance were identified in the beta-hairpin of the

polymerase. ABT-333, another palm site inhibitor, demonstrated antiviral activity in

patients with genotype 1 infection and from in vitro replicon as well as clinical studies

specific variants were observed as main resistance mutations.




Figure 14. Antiviral activity of non-nucleoside analogue NS5B polymerase inhibitors.

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NS5A inhibitor

In a single ascending dose study it was shown that inhibition of the NS5A protein with

BMS-790052 leads to a sharp initial decline of HCV RNA concentrations (Nettles

2008). BMS-790052 is the first NS5A inhibitor binding to domain I of the NS5A protein,

which was shown to be important for regulation of HCV replication. No clinical

data on resistance to this class of drugs have been presented yet and results of multiple

dose studies are eagerly anticipated. (from Jules: clinical data, in patients was presented at the EASL meeting in April 2010 and previously at AASLD 2 years ago)

Once-daily NS5A Inhibitor (BMS-790052) Plus Peginterferon-alpha-2a And Ribavirin Produces High Rates Of Extended Rapid Virologic Response In Treatment-naïve HCV-genotype 1 Subjects: Phase 2a Trial - Bristol-Myers Squibb Study AI444014 - (04/20/10)

BMS-790052 is a First-in-class Potent Hepatitis C Virus (HCV) NS5A ...
Nov 1, 2008 ... This new class of drug, the BMS NS5A inhibitor, attracted quite a lot of discussion because of it potent viral load reduction of -3.6 logs, ...
www.natap.org/2008/AASLD/AASLD_06.htm

The Most recent data updates on new HCV antivirals and new interferons were reported at the recent EASL:

EASL 45th Annual Meeting
(European Association for the Study of the Liver)
April 14-18, 2010
Vienna, Austria