Concerns About HCV Genotyping: Hepatitis C Virus (HCV) Genotype 1 Subtype Identification in New HCV Drug Development and Future Clinical Practice

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PLoS ONE 4(12): e8209. doi:10.1371/journal.pone.0008209

Published December 8, 2009

"INNO-LiPA HCV 2.0 currently is the best available commercial assay for HCV genotype 1 subtype identification and should be used in clinical trials and practice.....

Trugene HCV Genotyping Kit and INNO-LiPA HCV 1.0, failed to correctly identify HCV subtype 1a in 22.8% and 29.5% of cases, and HCV subtype 1b in 9.5% and 8.7% of cases, respectively....The second-generation line probe assay is currently the best commercial assay for determination of HCV genotype 1 subtypes 1a and 1b. It can therefore be used locally in clinical trials to identify the HCV subtype and stratify the patients at inclusion, as well as to interpret efficacy and resistance data. When reporting final data, direct sequence analysis of the NS5B region and/or another coding region (for instance the region encoding the antiviral drug target HCV protein) should always be performed as it may identify mistyping or mis-subtyping with commercial assays, especially in the case of rare subtypes."

Stephane Chevaliez1,2, Magali Bouvier-Alias1,2, Rozenn Brillet2, Jean-Michel Pawlotsky1,2*

1 French National Reference Center for Viral Hepatitis B, C and delta, Department of Virology, H™pital Henri Mondor, Universite Paris 12, Creteil, France, 2 INSERM U955, Creteil, France Methods based on the sole analysis of the 5'NCR, namely Trugene HCV Genotyping Kit and INNO-LiPA HCV 1.0, failed to correctly identify HCV subtype 1a in 22.8% and 29.5% of cases, and HCV subtype 1b in 9.5% and 8.7% of cases, respectively (Table 1)...... The results clearly show that, although they are by far the most widely used techniques in new HCV drug development trials, genotyping techniques based on the sole analysis of the 5'NCR should be avoided, as they mistype approximately 25% and 10% of HCV subtype 1a and 1b strains, respectively......INNO-LiPA HCV 2.0 displays the same 5'NCR oligonucleotide probes as INNO-LiPA HCV 1.0, plus core-encoded oligonucleotide probes aimed at better discriminating between HCV subtypes 1a and 1b. With INNO-LiPA HCV 2.0, subtype identification was corrected in 64 of the 70 subtypes 1a that were incorrectly typed with INNO-LiPA HCV 1.0. Five samples could not be PCR-amplified in the core-coding region and the result was not interpretable with INNO-LiPA HCV 2.0 in the remaining case (Table 1). INNO-LiPA HCV 2.0 also corrected subtype identification in 13 of 23 subtypes 1b that were incorrectly typed with INNO-LiPA HCV 1.0. Eight samples could not be PCR-amplified in the core-coding region and the result was not interpretable with INNO-LiPA HCV 2.0 in the remaining two cases (Table 1). Overall, the second-generation line probe assay correctly classified 97.5% of subtype 1a and 96.2% of subtype 1b strains. When only samples that could be PCR-amplified with the assay procedure were taken into account, correct subtype determination was achieved in 99.6% and 99.2% of cases, respectively (Table 1)The real-time PCR-based assay targeting both the 5'NCR and the NS5B region, Abbott RealTime HCV Genotype II assay, correctly identified 93.2% of subtype 1a and 88.9% of subtype 1b strains. Only 2 HCV subtype 1b samples could not be PCR-amplified with this method (Table 1)......Novel assays have been recently developed that aim at better discriminating among the different HCV genotype 1 subtypes and between genotypes 1 and 6. Abbott RealTime HCV Genotype II assay is a real-time PCR method using several sets of genotype- and subtype-specific primers and probes located in both the 5'NCR and the NS5B-coding region. As shown in Table 1, adding a second target region for analysis led to substantially improving HCV genotype 1 subtype identification compared to methods targeting the sole 5'NCR. However, in contrast with a previous report [33], we found that this assay failed to correctly identify HCV genotype 1 subtype in approximately 10% of cases"

Table 1. Ability of the different molecular methods tested in this study to correctly identify HCV subtypes 1a and 1b in a series of 500 patients infected by one or the other of these subtypes.

Abstract

Background

With the development of new specific inhibitors of hepatitis C virus (HCV) enzymes and functions that may yield different antiviral responses and resistance profiles according to the HCV subtype, correct HCV genotype 1 subtype identification is mandatory in clinical trials for stratification and interpretation purposes and will likely become necessary in future clinical practice. The goal of this study was to identify the appropriate molecular tool(s) for accurate HCV genotype 1 subtype determination.

Methodology/Principal Findings

A large cohort of 500 treatment-naïve patients eligible for HCV drug trials and infected with either subtype 1a or 1b was studied. Methods based on the sole analysis of the 5' non-coding region (5'NCR) by sequence analysis or reverse hybridization failed to correctly identify HCV subtype 1a in 22.8%-29.5% of cases, and HCV subtype 1b in 9.5%-8.7% of cases. Natural polymorphisms at positions 107, 204 and/or 243 were responsible for mis-subtyping with these methods. A real-time PCR method using genotype- and subtype-specific primers and probes located in both the 5'NCR and the NS5B-coding region failed to correctly identify HCV genotype 1 subtype in approximately 10% of cases. The second-generation line probe assay, a reverse hybridization assay that uses probes targeting both the 5'NCR and core-coding region, correctly identified HCV subtypes 1a and 1b in more than 99% of cases.

Conclusions/Significance

In the context of new HCV drug development, HCV genotyping methods based on the exclusive analysis of the 5'NCR should be avoided. The second-generation line probe assay is currently the best commercial assay for determination of HCV genotype 1 subtypes 1a and 1b in clinical trials and practice.

Funding: The Trugene HCV 5'NC Genotyping kits and the INNO-LiPA HCV kits were kindly provided by Siemens Medical Solutions Diagnostics. The Abbott RealTime HCV Genotype II kits were kindly provided by Abbott Molecular. This work is part of the activity of the VIRGIL European Network of Excellence on Antiviral Drug Resistance supported by a grant (LSHM-CT-2004-503359) from the Priority 1 "Life Sciences, Genomics and Biotechnology for Health" program in the 6th Framework Program of the European Union. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Introduction

Over 170 million individuals are infected with hepatitis C virus (HCV) worldwide. Phylogenetic analyses have shown that HCV strains can be classified into at least 6 major genotypes (numbered 1 to 6), and a large number of subtypes within each genotype [1]. Genotype 1 is by far the most frequent genotype in chronically infected patients worldwide, with subtypes 1a and 1b representing the vast majority of circulating strains [2], [3], [4].

Current treatment of chronic hepatitis C is based on the combination of pegylated interferon (IFN)-α and ribavirin [5]. This treatment fails to eradicate infection in 50%-60% of patients infected with HCV genotype 1 and approximately 20% of those infected with HCV genotypes 2 and 3 [6], [7], [8]. Thus the need for more efficacious therapies is urgent, especially for patients infected with HCV genotype 1. A number of novel antiviral molecules currently are in preclinical or clinical development [9]. The most advanced ones are specific inhibitors of viral enzymes and functions involved in the HCV life cycle. Molecules that have reached clinical development include inhibitors of the nonstructural (NS) 3/4A serine protease and inhibitors of HCV replication that belong to different categories: nucleoside/nucleotide analogue and non-nucleoside inhibitors of the HCV RNA-dependent RNA polymerase (RdRp), NS5A inhibitors and cyclophilin inhibitors [9]. These agents have shown potent antiviral efficacy when used alone, and encouraging results have been recently published showing that HCV clearance can be achieved in approximately 70% of cases when a potent NS3/4A inhibitor is used in combination with pegylated IFN-α and ribavirin [10], [11], [12].

HCV genotype 1 is generally considered as a homogeneous group. There are however biological differences between the different subtypes of HCV genotype 1, which are related to differences in their nucleotide and amino acid sequences. Importantly, differences between subtype 1a and 1b (by far the most frequently encountered genotype 1 subtypes in clinical practice) include different efficacies of antiviral drugs and different resistance profiles to such drugs. Indeed, several HCV inhibitors appear to have selective activity against different HCV genotype 1 subtypes, both in vitro and in vivo. Differences have been observed in vitro with NS3/4A protease inhibitors, non-nucleoside inhibitors of HCV RdRp and NS5A inhibitors [13], [14], [15], [16], [17]. For instance, BILB 1941, a non-nucleoside inhibitor of HCV RdRp, has been shown to have better antiviral efficacy in patients infected with HCV subtype 1b than in those infected with HCV subtype 1a, a finding reflecting in vitro experiments [13].

A major issue that limits the efficacy of direct acting antiviral therapies for HCV is the selection by these drugs of resistant variants upon administration [18]. Recent studies with NS3/4A protease inhibitors have shown that the genetic barrier and resistance profiles substantially differ between the different genotype 1 subtypes. For instance, the Arg to Lys substitution at position 155 of the NS3 protease (R155K) is usually selected in subtype 1a replicons treated with telaprevir, but not in subtype 1b replicons [19]. The reason is that only one nucleotide substitution is needed relative to the subtype 1a sequence to generate this variant, whereas two substitutions are needed relative to the 1b sequence (codon usage bias). Overall, natural polymorphisms at positions R155 and V36 are frequent in subtype 1a, but rare in subtype 1b where substitutions at position A156 are preferentially selected in vitro [19]. This is reflected in vivo by the different resistance profiles in patients infected by HCV subtypes 1a and 1b. In the former, the V36 and R155 substitutions represent the backbone of resistance, whereas in the latter resistance is less frequent as it is preferentially associated with substitutions at position A156 that are associated with a decreased fitness of the variants [19], [20], [21]. Similarly, important differences in the resistance profiles have been described in vitro with HCV-796, a non-nucleoside inhibitor of HCV RdRp. The C316Y amino acid substitution has been reported to be selected in both subtype 1a and 1b replicon cells. However, in genotype 1a replicons, the C316Y substitution has low replication capacity that must be compensated for by additional "compensatory" substitutions, including L392F or M414T, resulting in an increase in replication levels of at least 10-fold [19]. A higher genetic barrier to resistance to HCV-796 and related compounds is therefore expected in patients infected with HCV subtype 1a than 1b. In vivo, HCV-796 monotherapy was however shown to select subtype 1a variants with a single C316Y substitution, whereas the C316Y substitution was associated with a number of additional substitutions in subtype 1b patients [22].

As a result of these findings, correct identification of HCV subtypes 1a and 1b is crucial in clinical trials assessing new HCV drugs in order to correctly stratify and interpret efficacy and resistance data. It may also become important in future clinical practice, as tailoring treatment schedules with HCV inhibitors to HCV genotype 1 subtype might become necessary. A variety of molecular methods can be used to identify the HCV genotype and subtype both in clinical trials and practice. Commercial assays have been developed, most of them targeting the 5' noncoding region (5'NCR) of the HCV genome, although this region is the most conserved one. These methods have been shown to differentiate well the different HCV genotypes (1 to 6), except genotype 1 from genotype 6, a rare HCV genotype in the Western world [23], [24]. The goal of our study was to assess the ability of molecular methods targeting the 5'NCR to correctly identify the HCV genotype 1 subtype in patients eligible for clinical trials, and to identify the best method for this purpose.

Results

Hepatitis C Virus Genotype and Subtype Determination by Phylogenetic Analysis of a Portion of the NS5B Gene

Direct sequence analysis of a sufficiently long portion of the NS5B gene followed by phylogenetic analysis is the reference method for identification of HCV genotype and subtype [1], [25]. It was used to identify the HCV genotype and subtype in 516 treatment-naïve patients included in a multicenter clinical trial assessing different schedules of pegylated IFN-α2a and ribavirin [26]. All of these patients were thought to be infected with HCV genotype 1 at inclusion based on local assessment. In fact, 6 patients were infected with genotype 6, including 2 with subtype 6e, one with subtype 6o, one with subtype 6p, one with subtype 6q and one with subtype 6r. These 6 samples were not considered for further analysis in the present study. The remaining 510 patients were confirmed to be infected with HCV genotype 1: 237 of them (46.5%) were infected with HCV subtype 1a and 263 (51.6%) with subtype 1b (Figure 1). As shown in Figure 1, HCV subtype 1a strains segregated into two distinct clades, that were termed 1a clade I (n = 83, 35.0%) and 1a clade II (n = 154, 65.0%). Eight patients (1.6%) were infected with another HCV genotype 1 subtype, including 4 patients with subtype 1d, 2 with subtype 1e, one with subtype 1i, and one with subtype 1l. The remaining 2 patients (0.3%) were infected with genotype 1 but the subtype could not be determined. The ability of the different molecular methods to correctly identify HCV subtypes 1a and 1b was then tested on the 237 and 263 samples containing HCV subtypes 1a and 1b, respectively.

INNO-LiPA HCV 2.0 displays the same 5'NCR oligonucleotide probes as INNO-LiPA HCV 1.0, plus core-encoded oligonucleotide probes aimed at better discriminating between HCV subtypes 1a and 1b. With INNO-LiPA HCV 2.0, subtype identification was corrected in 64 of the 70 subtypes 1a that were incorrectly typed with INNO-LiPA HCV 1.0. Five samples could not be PCR-amplified in the core-coding region and the result was not interpretable with INNO-LiPA HCV 2.0 in the remaining case (Table 1). INNO-LiPA HCV 2.0 also corrected subtype identification in 13 of 23 subtypes 1b that were incorrectly typed with INNO-LiPA HCV 1.0. Eight samples could not be PCR-amplified in the core-coding region and the result was not interpretable with INNO-LiPA HCV 2.0 in the remaining two cases (Table 1). Overall, the second-generation line probe assay correctly classified 97.5% of subtype 1a and 96.2% of subtype 1b strains. When only samples that could be PCR-amplified with the assay procedure were taken into account, correct subtype determination was achieved in 99.6% and 99.2% of cases, respectively (Table 1).

The real-time PCR-based assay targeting both the 5'NCR and the NS5B region, Abbott RealTime HCV Genotype II assay, correctly identified 93.2% of subtype 1a and 88.9% of subtype 1b strains. Only 2 HCV subtype 1b samples could not be PCR-amplified with this method (Table 1).

5'NCR Sequence Analysis in Misclassified Subtype 1a Strains

Among the HCV subtype 1a strains, 47 were misclassified as subtype 1b by Trugene HCV Genotyping Kit and/or INNO-LiPA HCV 1.0, including 33 that were misclassified by both assays, 7 that were misclassified by Trugene HCV Genotyping Kit only, and 7 that were misclassified by INNO-LiPA HCV 1.0 only (Table 2). Figure 2 shows an alignment of their 5'NCR sequences relative to the consensus sequences of the correctly classified strains (including subtype 1a clade I, subtype 1a clade II and subtype 1b). As shown in Figure 2, misclassification of subtype 1a strains into subtype 1b in one or both assays was related to the presence of natural polymorphisms at nucleotide positions 204 and 243, both of which are located within the sequence of an INNO-LiPA HCV 1.0 probe. At position 243, A is the most frequent nucleotide in HCV subtype 1a, in both subtype 1a clade I and clade II. Substitution into a G, the most frequent nucleotide at position 243 in subtype 1b, was found in all cases that were misclassified as subtype 1b by Trugene HCV Genotyping Kit and/or INNO-LiPA HCV 1.0 (Figure 2). At position 204, A is the most frequent nucleotide for subtype 1a clade I, whereas C is the most frequent nucleotide for subtype 1a clade II, and C or T are the most frequent nucleotides for subtype 1b. In spite of the presence of a G at position 243, the presence of an A at position 204 allowed correct identification of subtype 1a with Trugene HCV Genotyping Kit but not with INNO-LiPA HCV 1.0 (Figure 2). The usual presence of a C at position 204 in subtype 1a clade II explains why misclassifications were far more frequent with this clade than with subtype 1a clade I.

Among the 12 subtype 1a strains that were classified as genotype 1, indeterminate subtype with Trugene HCV Genotyping Kit, one had a G and 5 had mixed A and G populations at position 243. Two additional patients with an A at position 243 had a C at position 248. In the remaining 4 cases, no explanation was found in the 5'NCR sequence for the failure to identify the HCV subtype (data not shown). Among the 25 subtype 1a strains that were classified as genotype 1, indeterminate subtype with INNO-LiPA HCV 1.0 (including 6 with the same profile in Trugene HCV Genotyping Kit), 4 had a G and 4 had mixed A and G populations at position 243. Three additional patients with an A at position 243 had a C at position 248 (C only in two of them, a mixture of C and T in one). In the 14 remaining cases, no explanation was found in the 5'NCR sequence for the failure to identify the HCV subtype (data not shown).

5'NCR Sequence Analysis in Misclassified Subtype 1b Strains

Among HCV subtype 1b strains, 8 were misclassified as subtype 1a by Trugene HCV Genotyping Kit and/or INNO-LiPA HCV 1.0, including 3 that were misclassified by both assays, 4 that were misclassified by Trugene HCV Genotyping Kit only, and 1 that was misclassified by INNO-LiPA HCV 1.0 only (Table 2). Figure 3 shows an alignment of their 5'NCR sequences relative to the consensus sequences of the correctly classified subtype 1a and subtype 1b strains. As shown in Figure 3, and as for misclassified subtype 1a strains discussed above, misclassification of subtype 1b strains into subtype 1a was related to the presence of natural polymorphisms at positions 204 and 243. At position 243, G is the most frequent nucleotide in HCV subtype 1b. Substitution into an A, the most frequent nucleotide at position 243 in subtype 1a, was found in all cases that were misclassified as subtype 1a by both Trugene HCV Genotyping Kit and INNO-LiPA HCV 1.0 and by INNO-LiPA HCV 1.0 only, but not in those that were misclassified by Trugene HCV Genotyping Kit only (Figure 3). In the latter, it is the presence of an A at position 204 instead of a C or a T that was responsible for misclassification in all but one case (Figure 3).

Among the 11 subtype 1b strains that were classified as genotype 1, indeterminate subtype with Trugene HCV Genotyping Kit, one had an A at position 243. In the remaining cases, no explanation was found in the 5'NCR sequence for the failure to identify the HCV subtype (data not shown). Among the 15 subtype 1b strains that were classified as genotype 1, indeterminate subtype with INNO-LiPA HCV 1.0 (none of which were classified as indeterminate in Trugene HCV Genotyping Kit), one had an A and one harbored mixed A and G populations at position 243. Both of them had a C at position 248 (C only in one of them and a mixture of C and T in the other one). In the remaining 13 cases, no explanation was found in the 5'NCR sequence for the failure to identify the HCV subtype (data not shown).

Incorrect Subtyping with Abbott RealTime HCV Genotype II Assay, that Targets Both the 5'NCR and NS5B Region

Among the HCV subtype 1a strains, 16 were incorrectly classified by Abbott RealTime HCV Genotype II assay (Table 1): 2 were misclassified as subtype 1b, 12 were classified as genotype 1, indeterminate subtype, one was identified as a mixed 1a/1b infection, and one gave an indeterminate result. In one case, PCR amplification failed, and in one case, not enough serum volume was available for testing.

Among the HCV subtype 1b strains, 27 were incorrectly classified by Abbott RealTime HCV Genotype II assay (Table 1): 3 were misclassified as subtype 1a, 18 were classified as genotype 1, indeterminate subtype, 5 were identified as a mixed 1a/1b infection, and one gave an indeterminate result. In 2 cases, PCR amplification failed, and in one case, not enough serum volume was available for testing.

Source

Evaluation of Versant Hepatitis C Virus Genotype Assay (LiPA) 2.0

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Journal of Clinical Microbiology, June 2008, p. 1901-1906, Vol. 46, No. 6 doi:10.1128/JCM.02390-07

"The reliability of genotyping methods highly depends on the amount of information (i.e., the number of informative sites) that is utilized for the discrimination of genetic variants......Our results indicate that Versant HCV genotype assay (LiPA) 2.0 yielded an interpretable genotype result for 96.0% of the samples and that 99.4% of the interpretable results agreed with the reference method, rendering it an accurate and reliable assay suitable for large-scale genotyping. This new assay outperforms the previous version of the line probe assay, since Versant HCV genotype assay (LiPA) 1.0 has an overall accuracy of 74%, taking subtype i nformation into account (8, 23).....In conclusion, Versant HCV genotype assay (LiPA) 2.0 provides a rapid, sensitive, and accurate means of HCV genotyping and can be used as a routine tool to distinguish between the different HCV genotypes and subtypes. Considering the importance of genotype determination in understanding the epidemiology of the virus and in the management of hepatitis C treatment strategies, efficient genotyping tools are indispensable in clinical diagnostic settings."

Jannick Verbeeck,1 Mark J. Stanley,2 Jen Shieh,2 Linda Celis,3 Els Huyck,3 Elke Wollants,1 Judy Morimoto,2 Alice Farrior,2 Erwin Sablon,3 Margaret Jankowski-Hennig,4 Carl Schaper,4 Pamela Johnson,4 Marc Van Ranst,1* and Marianne Van Brussel3

Laboratory of Clinical Virology, Rega Institute for Medical Research, Leuven, Belgium,1 Department of Microbiology, Kaiser Permanente, TPMG Regional Laboratory, Berkeley, California,2 Innogenetics NV, Gent, Belgium,3 Siemens Healthcare Diagnostics, Berkeley, California4

ABSTRACT

Hepatitis C virus (HCV) genotyping is a tool used to optimize antiviral treatment regimens. The newly developed Versant HCV genotype assay (LiPA) 2.0 uses sequence information from both the 5' untranslated region and the core region, allowing distinction between HCV genotype 1 and subtypes c to l of genotype 6 and between subtypes a and b of genotype 1. HCV-positive samples were genotyped manually using the Versant HCV genotype assay (LiPA) 2.0 system according to the manufacturer's instructions. For the comparison study, Versant HCV genotype assay (LiPA) 1.0 was used. In this study, 99.7% of the samples could be amplified, the genotype of 96.0% of samples could be determined, and the agreement with the reference method was 99.4% when a genotype was determined. The reproducibility study showed no significant differences in performance across sites (P = 0.43) or across lots (P = 0.88). In the comparison stud y, 13 samples that were uninterpretable or incorrectly genotyped with Versant HCV genotype assay (LiPA) 1.0 were correctly genotyped by Versant HCV genotype assay (LiPA) 2.0. Versant HCV genotype assay (LiPA) 2.0 is a sensitive, accurate, and reliable assay for HCV genotyping. The inclusion of the core region probes in Versant HCV genotype assay (LiPA) 2.0 results in a genotyping success rate higher than that of the current Versant HCV genotype assay (LiPA) 1.0.

INTRODUCTION

Hepatitis C virus (HCV) is a leading cause of chronic liver disease and has already infected at least 170 million people worldwide. Each year, 3 to 4 million people are newly infected. HCV creates an extensive disease burden, since it accounts for 20 to 30% of cases of acute hepatitis, 70 to 80% of cases of chronic hepatitis, 40% of cases of end-stage cirrhosis, 50 to 76% of cases of hepatocellular carcinoma, and 30 to 40% of liver transplants (15, 33, 34).

HCV belongs to the family of the Flaviviridae and can be divided into different genotypes based on phylogenetic analysis of full-length or partial sequences of HCV strains. The most current consensus proposal distinguishes six genotypes based on phylogenetic cluster analysis of complete genomes. The genotype formerly designated as 10a has been reassigned as genotype 3, subtype k. Genotypes 7, 8, 9, and 11, belonging to clade 6, have been reassigned to genotype 6, subtypes c to l (25, 26, 27). These six HCV genotypes have different geographical distributions (21, 30, 32).

Treatment options for chronic HCV infections are poor. At the moment, the only accepted antiviral therapy with proven effectiveness is a combination therapy of (peg)interferon alpha and ribavirin. The overall success rate of this antiviral treatment ranges from 50% to 90% (11). According to a National Institutes of Health (NIH) 2002 panel, several factors are associated with successful treatment response, including lower baseline HCV RNA levels, lower fibrosis and inflammation scores upon liver biopsy, lower body weight, and lower body surface area, but the most important predicting factor is HCV genotype (24). Patients infected with HCV genotype 1 respond least to therapy, while patients infected with genotypes 2 and 3 show the best responses (14, 17, 22). For HCV genotypes 4, 5, and 6, treatment data are scarce, but it is recommended to treat these individuals using the same regimen as for patients infected with genotype 1 (7, 13, 18, 31). Nearly all patients experience side effects with the antiviral therapy. These side effects can be severe and contribute to discontinuation rates of 10 to 14% and dose reductions for 7 to 42% of patients, depending on the type and length of treatment (16). Therefore, it is important that clinicians have the appropriate information to make individual treatment choices in order to maximize the chance of successful treatment outcome for each individual patient, rendering HCV genotyping assays important and useful tools to optimize treatment type, duration, and dose.

In this paper, we evaluate Versant HCV genotype assay (LiPA) 2.0 (CE marked in Europe; for research use only; not for use in diagnostic procedures in the United States) (manufactured by Innogenetics, distributed by Siemens Healthcare Diagnostics), which uses sequence information from the core region in addition to sequence information from the 5' untranslated region (5'UTR), allowing an improved and more accurate distinction between HCV genotype 1 and subtypes c to l of genotype 6 and between subtypes a and b of genotype 1.

RESULTS

Clinical accuracy study. Table 1 summarizes the results for the 326 specimens that were used for the reference method comparison. Upon initial testing, 93.3% (304/326) of the specimens gave interpretable genotype results, 2.1% (7/326) failed to amplify, and 4.6% (15/326) amplified but gave uninterpretable results. Of the 304 specimens that yielded a genotype result, 99.3% (302/304) gave results that agreed with the reference method. After specimens that yielded no genotype result were retested, 96.0% (313/326) of the specimens gave interpretable genotype results, 3.5% (12/326) amplified but remained uninterpretable, and 0.3% (1/326) failed to amplify. Of the 313 specimens that yielded a genotype result after repeat testing, 99.4% (311/313) gave results that agreed with the reference method. Table 2 shows that the specimens that did not amplify or give interpretable results were distributed across ge notypes. The two specimens that initially gave results that disagreed with those obtained by the reference method were retested, and both gave retest results that agreed with those obtained by the reference method.

In order to determine the core amplification efficiency, 156 genotype 1 and genotype 6 (c to l) samples were analyzed. Two samples showed negative AMPL CTRL 1 lines and were excluded from further analysis. Of the remaining 154 genotype 1 and genotype 6 (c to l) samples, 1 sample had a negative AMPL CTRL 2, r esulting in the amplification of 99.4% (153/154) samples.

The clinical subtype efficiency for HCV genotypes 1a and 1b was determined using 129 samples that were genotype 1a or 1b based on reference sequencing and genotype 1, 1a, or 1b based on LiPA genotyping; this determination was based on initial testing only, excluding repeat testing of initial amplification failures and uninterpretable results. Three out of 129 samples were indeterminate at the subtype level, resulting in a clinical HCV genotype 1 subtype efficiency of 97.7% after initial testing. Upon repeat testing, all samples gave a correct consensus subtype result. All of the 126 samples that were genotype 1a or 1b by LiPA were concordant with sequencing.

In order to check whether Versant HCV genotype assay (LiPA) 2.0 was able to determine the correct genotype for samples with viral loads at the upper limit of detection, 22 samples with viral loads ranging from 4.0 x 106 IU/ml to 8.7 x 106 IU/ml were selected, and the genotype success rate and the percentage of agreement with the reference method for these high-concentration specimens were estimated. For all these samples, Versant HCV genotype assay (LiPA) 2.0 produced the same genotype results as the genotype result determined by NS5b sequencing and phylogenetic analysis, resulting in both a genotype success rate and an agreement with the reference method of 100%.

Reproducibility study. Table 3 summarizes the valid, indeterminate, correct, and incorrect genotype results for each reproducibility panel member. In total, 3.3% (16/486) of reactions gave indeterminate results (defined as specimens with either an amplification failure or an uninterpretable result) and 96.7% (LCL, 95.0%) yielded an interpretable genotype result. Of the 470 specimens with interpretable results, 100% (LCL, 99.4%) gave the correct genotype. The indeterminate results occurred at all sites, with all three reagent lots, and in multiple assay runs. There were no significant performance differences seen for the Versant HCV genotype assay (LiPA) 2.0 system across sites/operators (P = 0.43) or across reagent lots (P = 0.88). The genotype success rates at the individual sites were 98.1% for site 1, 97.6% for site 2, and 94.4% for site 3. The genotype success rates for the individual lots were 96.9% using lot 1, 97.5% using lot 2, and 95.7% using lot 3.

Comparison study. Table 4 gives an overview of the results of the comparison study after original testing and after repeat testing. Of the 100 specimens tested, 13 specimens initially produced uninterpretable results by either Versant HCV genotype assay (LiPA) 1.0 or Versant HCV genotype assay (LiPA) 2.0 or both assays. The HCV RNA concentrations of these 13 samples ranged from 14,615 to 2,500,000 IU/ml. These specimens were retested using both ass ays. After repeat testing, three specimens remained uninterpretable by both versions of the assay, five remained uninterpretable by Versant HCV genotype assay (LiPA) 1.0, and one remained uninterpretable by Versant HCV genotype assay (LiPA) 2.0. For all six specimens that gave a genotype result by only one version of the assay, the observed genotype result agreed with that obtained by sequencing the NS5b region of the HCV genome. After repeat testing, 83 specimens were concordant by both assays at the genotype level. Of these, 16 specimens had concordant genotypes by both assays, but one of the assays failed to give a subtype, resulting in a total of 67 concordant specimens when the subtype level is taken into account. Results from eight specimens were discordant between the two assays, and results from nine specimens were uninterpretable by at least one of the assays. The total number of interpretable specimens by both assays was 91, of which 83 had concordant results at the genotype level only (91.2%; LCL, 84.7%). Table 5 shows the number o f genotype and subtype results produced by both assays for the 100 specimens tested after repeat testing.

The eight samples that showed discordant results were sequenced in the NS5b region of the HCV genome (two samples were HCV genotype 6 subtypes c to l, and six samples were HCV genotype 1a). Results indicated that Versant HCV genotype assay (LiPA) 2.0 gave the correct HCV genotype and subtype, as determined by NS5b sequencing. In contrast, Versant HCV genotype assay (LiPA) 1.0 had misclassified all eight samples as HC V genotype 1b. Of the 96 specimens that were interpretable with Versant HCV genotype assay (LiPA) 2.0, 83 showed concordant results with Versant HCV genotype assay (LiPA) 1.0, while 13 showed improved results over Versant HCV genotype assay (LiPA) 1.0, which leads to 100% concordant or improved results (LCL, 96.9%).

DISCUSSION

Phylogenetic analysis of a coding region, or even more, the complete genome, is considered the gold standard for identifying different HCV genotypes (6). However, since this method is expensive and time-consuming, it is impractical for large-scale genotyping projects (8). For this reason, commercial genotyping kits were developed for routine determination of HCV genotypes. Most commercially available HCV genotyping assays, including Versant HCV genotype assay (LiPA) 1.0, use the 5'UTR, since this region is highly conserved and therefore well suited for the development of detection methods. The reliability of genotyping methods highly depends on the amount of information (i.e., the number of informative sites) that is utilized for the discrimination of genetic variants. The 5'UTR is sufficiently variable for discrimination of HCV genotypes 1 to 5 and most subtypes of HCV genotype 6 (12, 28, 29, 32). However, it does not allow discrimination of HCV genotype 6 subtypes c to l from HCV genotype 1 and has only a limited subtyping accuracy (5, 29). To overcome the limitations of the 5'UTR, a new assay which uses additional sequence information from the core region of the HCV genome, Versant HCV genotype assay (LiPA) 2.0, has recently been developed (20). In this study, we evaluated the new assay and compared it with the previous version of the assay.

Our results indicate that Versant HCV genotype assay (LiPA) 2.0 yielded an interpretable genotype result for 96.0% of the samples and that 99.4% of the interpretable results agreed with the reference method, rendering it an accurate and reliable assay suitable for large-scale genotyping. This new assay outperforms the previous version of the line probe assay, since Versant HCV genotype assay (LiPA) 1.0 has an overall accuracy of 74%, taking subtype information into account (8, 23).

In the comparison study, eight specimens showed discordant results when tested with both assays. The NS5b sequencing results for these samples showed that Versant HCV genotype assay (LiPA) 2.0 gave the correct HCV genotype and subtype and thereby showed an improvement in identifying HCV-positive samples which are subtypes c to l of genotype 6 and in identifying the correct subtype of genotype 1. This improvement can be attributed to the additional information available from the core region of the HCV genome, which can better distinguish between genotype 1 and subtypes c to l of genotype 6 and between subtype a and b of genotype 1. This core information is not available in Versant HCV genotype assay (LiPA) 1.0, and this can lead to misinterpretation. For example, in a study by Chinchai et al., this assay could not discriminate HCV genotype 6a variants from HCV genotype 1b, and two samples found to be genotype 1 by the assay contained genotype 3 core sequences (5). Chen and Weck showed that Versant HCV genotype assay (LiPA) 1.0 cannot accurately distinguish HCV genotypes 1a and 1b, since in most cases, the 5'UTR is not heterogeneous enough for use in determining the HCV subtype (4). Several other studies report on moderate distinction at the subtype level (1, 2, 9, 12, 19). This is not surprising, since the 5'UTR is the most highly conserved region of the HCV genome, and only one or two nucleotide changes distinguish unique subtypes. Assigning correct genotypes and subtypes to HCV specimens is important for several research purposes, including epidemiological, phylogenetic, and natural history studies. Some studies even report that there is a slight difference in treatment outcomes between HCV genotype 1a- and HCV genotype 1b-infected patients, showing that correct subtype assignment is indispensable (3, 10, 30).

In conclusion, Versant HCV genotype assay (LiPA) 2.0 provides a rapid, sensitive, and accurate means of HCV genotyping and can be used as a routine tool to distinguish between the different HCV genotypes and subtypes. Considering the importance of genotype determination in understanding the epidemiology of the virus and in the management of hepatitis C treatment strategies, efficient genotyping tools are indispensable in clinical diagnostic settings.

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Drug offerings expand for Merck Helps programs

By Alaric DeArment

WHITEHOUSE STATION, N.J. (Sept. 22) Merck is expanding its patient assistance program, the drug maker said Wednesday.

Merck announced an expansion of the number of drugs it offers under its Merck Helps programs, including the Merck Patient Assistant program, the Merck Vaccine Patient Assistance program, the ACT program for cancer and hepatitis C drugs, and the SUPPORT program for HIV and AIDS drugs. The program provides medicines and vaccines free of charge to eligible patients, primarily the uninsured, who make up to 400% of the federal poverty level and can’t afford Merck drugs without assistance.

“Merck has historically recognized the critical need for people to have access to the prescription medicines and vaccines they require, even if they lose their insurance,” Merck EVP and chief medical officer Michael Rosenblatt said. “Our patient assistance programs now provide access to even more medicines for chronic conditions like asthma, diabetes and high blood pressure, allowing us to reach more people in need.”

Merck also said that its philanthropic arm, the Merck Company Foundation, made a grant to NeedyMeds, a nonprofit group that helps people who can’t afford medicines or healthcare costs by making information about assistance programs available. NeedyMeds plans to use the grant to translate its website into Spanish.

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Study: HIV Doesn’t Increase Bone Fracture Risk in Women

September 22, 2010

HIV-positive women were no more likely to have a bone fracture than HIV-negative women, according to a study published online September 20 in AIDS. The new data run counter to a growing concern that HIV might be causing age-related problems, including bone problems, to occur at a younger age in HIV-positive people.

As the population of people with HIV grows older, the research community is increasingly turning its focus toward diseases and conditions that typically strike the elderly, including cardiovascular disease, certain cancers and bone problems.

Numerous studies have found that people with HIV, men and women alike, often have poorer bone health than their HIV-negative counterparts. What isn’t certain, however, is whether the reduced levels of bone density found in HIV-positive people are resulting in an increase in bone fractures. While one large study did find an increase in fractures of the hip, spine and wrist, it could not account for many bone fracture risk factors, such as body mass, smoking status and hepatitis C virus (HCV) infection.

To further define the actual risk for fractures in HIV-positive women, Michael Yin, MD, from Columbia University Medical Center in New York City, and his colleagues analyzed data from the Women’s Interagency HIV Study (WIHS)—a large cohort study that has been following more than 3,000 HIV-positive and HIV-negative women since at least 2001. This analysis included 1,728 HIV-positive women and 663 HIV-negative women.

Most of the women in both groups were black or Latina and tended to be a bit overweight. About half were smokers, and roughly 20 percent in both groups were diabetic. The groups differed on several counts, however. HIV-positive women tended to be older, to have begun menopause and to also be infected with HCV.

The rate of new fractures was similar between the groups, with 148 HIV-positive women (8.6 percent) and 47 HIV-negative women (7.1 percent) experiencing a new fracture during an average follow-up of 5.4 years.

Yin’s team found that when they accounted for known risk factors—such as smoking, HCV status and the women’s ratio of height to body weight (body mass index or BMI)—HIV-positive women were no more likely to experience a fracture than HIV-negative women. In fact, the only factors associated with bone fracture risk overall were: older age, white race, HCV status and high kidney protein (serum creatinine) levels.

When Yin and his colleagues restricted their analysis only to HIV-positive women, they found that smoking history, opiate-use history and onset of menopause were all associated with a higher fracture risk. Of interest, low CD4 counts were not associated with an increased fracture risk among HIV-positive women, nor was any class of antiretrovirals (ARVs) including tenofovir (found in Viread, Truvada and Atripla), a commonly used HIV medication that has been tied to decreased bone mineral density.

The authors caution that their study results can’t be applied to older and post-menopausal women, but they concluded, “Our data provide some reassurance that fracture risk is modest in predominantly premenopausal HIV-infected women.”

“However, further research is necessary,” they continued, “to assess fracture risk as these women transition through menopause and to clarify whether fracture risk differs among antiretroviral regimens.”

Source

Doctors File Federal Complaint to Halt Transfer of Chimpanzees for Invasive Experiments

Sept. 22, 2010

MEDIA CONTACT:
Jeanne McVey
202-527-7316
jeannem@pcrm.org

Legal Petition Seeks to Compel HHS Chief Kathleen Sebelius to Retire Elderly New Mexico Chimpanzees and Cancel Their Move to Texas Laboratory

WASHINGTON—Elderly chimpanzees living at a nonresearch facility in Alamogordo, N.M., should not be shipped to Texas for use in invasive experiments, says a federal complaint to be filed Sept. 23 with Kathleen Sebelius, secretary of Health and Human Services.

Doctors and scientists with the Physicians Committee for Responsible Medicine (PCRM) seek to halt the National Institutes of Health’s planned transfer of nearly 200 federally owned chimpanzees to a laboratory in San Antonio, Texas. New Mexico Gov. Bill Richardson and primatologist Jane Goodall have also spoken out against the proposed transfer.

The doctors’ legal petition invokes the Chimpanzee Health Improvement Maintenance and Protection (CHIMP) Act, enacted to ensure that chimpanzees used in experiments for many years are retired to sanctuaries. Many Alamogordo chimpanzees are elderly, have been used repeatedly for invasive procedures, and deserve a peaceful retirement, PCRM’s complaint says. For example, Flo, Guy, and James were born in 1957, 1959, and 1960, respectively. Many suffer from heart disease, making them especially unsuitable for medical experiments.

“There can be no scientific, legal, or ethical justification for returning a 50-year-old chimpanzee to laboratory experiments,” says John J. Pippin, M.D., F.A.C.C., senior medical and research adviser for PCRM. “More than five decades of experiments have shown us that chimpanzees are poor models for researching human diseases. Are we such slow learners that we now return to these outdated methods?” In submitting the petition to Secretary Sebelius, Dr. Pippin is joined by 11 other authorities, including Harvard professor Richard Wrangham, Ph.D., and University of New Mexico professor John Gluck, Ph.D.

The doctors and scientists argue in their complaint, “Chimpanzees have repeatedly proved to be poor models for human disease research, including for HIV—a disease for which repeated failures have led most researchers to stop using chimpanzees—as well as for hepatitis, malaria, and cancer. Therefore, these animals are not necessary for this type of research. Superior alternatives are available and researchers continue to develop new, cutting-edge models for human disease research.” The hepatitis C virus behaves very differently in humans and chimpanzees, and decades of experiments have failed to produce a human vaccine. Leading hepatitis C researchers are using human-cell-based research methods.

The mothers of some Alamogordo chimpanzees now live in a sanctuary in Washington State—Chimpanzee Sanctuary Northwest—and have come to the attention of Sen. Maria Cantwell. Sen. Cantwell recently introduced the Great Ape Protection Act (GAPA), S. 3694, which would advance medical research by phasing out wasteful and misleading chimpanzee experiments and releasing federally owned chimpanzees to sanctuaries. A parallel House bill, H.R. 1326, has gained significant momentum and now has 149 co-sponsors. The United States is the last country in the world that permits and funds large-scale chimpanzee research and testing. Earlier this month, the European Union banned great ape experiments.

Founded in 1985, the Physicians Committee for Responsible Medicine is a nonprofit health organization that promotes preventive medicine, conducts clinical research, and encourages higher standards for ethics and effectiveness in research.

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Taribavirin Offers a Safe, Effective Alternative for Chronic Hepatitis C, Study Finds

ScienceDaily (Sep. 22, 2010) — Researchers at Cedars-Sinai Medical Center and 50 other centers found that weight-based dosing of taribavirin reduces rates of anemia while increasing sustained virologic response (SVR) in patients with chronic hepatitis C (HCV). Full details of this study are available in the October issue of Hepatology, a journal published by Wiley-Blackwell on behalf of the American Association for the Study of Liver Diseases (AASLD).

Chronic HCV is typically treated with ribavirin (RBV). When used in combination with peginterferon alfa (peg-IFN), RBV significantly enhances on-treatment virologic response and reduces relapse. However, RBV, particularly the combination of interferon and RBV, is associated with hemolytic anemia, a significant toxicity resulting from the accumulation of RBV in red blood cells. Taribavirin (TBV), formerly known as viramidine, is a nucleoside analog and oral pro-drug of RBV that is less able to enter red blood cells, and should therefore be associated with significantly less anemia.

This theory was demonstrated in two previous phase 3 trials. While statistically less anemia was observed in patients treated with TBV compared to RBV, the primary efficacy endpoint of these studies, a non-inferior SVR between the TBV and RBV, was not achieved. Detailed subgroup analyses of the data suggest fixed dosing as opposed to weight-based dosing, and the selection of an inadequate dose, are to blame. The present multi-center study explored several higher weight-based doses of TBV to determine a dosage regimen that was able to deliver comparable responses to RBV with fewer incidences of anemia.

A phase 2b randomized, open-label, active-controlled, parallel-group study was conducted in 278 treatment-naïve, genotype 1 patients stratified by body weight and baseline viral load at 51 centers in the United States between March 2007 and October 2008. Patients were randomized 1:1:1:1 to receive TBV (20, 25, or 30 mg/kg/day) or RBV (800 -1400 mg/day) with pegylated interferon alfa-2b for 48 weeks.

The primary efficacy endpoint was early virologic response (EVR) defined as the proportion of patients with at least a 2-log decrease from baseline in serum HCV RNA levels at treatment week 12. Additional efficacy endpoints included SVR, undetectable HCV RNA at treatment weeks 4, 24 and 48, and viral relapse for those who were responders at the end of treatment. A total of 86 (41%) of TBV patients and 25 (36%) of the RBV group completed treatment and follow up. The most commonly cited reasons for premature withdrawal were lack of response (29%) and adverse events (20%).

The present study demonstrated that weight-based dosing of TBV achieved comparable efficacy to RBV as demonstrated by SVR. This was observed in all three TBV weight-based dose treatment groups, which met the study's primary end-point. Patients treated with TBV had less than half the anemia compared to RBV treated patients. These results suggest weight-based dosing of TBV can significantly improve the tolerability of HCV treatment while maintaining efficacy. Specifically, the 25 mg/kg dose offered the optimal balance of efficacy and safety in this patient population.

Notably, fewer patients treated with TBV required dose reductions (13-28%) compared to 32% of patients treated with RBV. Less frequent dose modification in patients treated with TBV may alleviate the need to utilize erythropoiesis-stimulating agents (ESAs). Several studies have demonstrated the use of ESAs can significantly decrease the need to dose reduce RBV and leads to an improvement in the quality of life during HCV treatment, but fails to improve the SVR. The use of ESAs also adds significant cost to HCV treatment and is associated with serious adverse events including thrombosis and red cell aplasia.

Lead investigator Dr. Fred Poordad concludes, "These data suggest TBV may be an effective agent to substitute for RBV in the future and could be incorporated in upcoming trials utilizing emerging small molecules for HCV treatment."

Editorial author Dr. Paul Kwo comments, "If TBV can be shown to preserve or improve efficacy rates in combination with direct-acting antiviral agents (DAAs) and Peg IFN, with lower rates of anemia, the use of TBV in these clinical settings would be a welcome addition to the HCV armamentarium as we begin to expand the HCV populations that we treat. TBV may have a role in populations particularly sensitive to ribavirin-related anemia. However, with the commencement of several trials comprising of multiple combinations of DAAs with and without pegIFN/RBV, and the development of newer protease inhibitors with potentially lower rates of anemia, the role of TBV remains less precisely defined and could potentially have a finite life cycle."

Source

NHS hepatitis C guidance updated

NICE reacts to ‘growing hepatitis C problem’
By Tim Locke
WebMD Health News

Reviewed by Dr Rob Hicks

22nd September 2010 - The NHS regulator, NICE, has updated guidance on the treatment of hepatitis C. This covers the use of peginterferon alfa (2a or 2b) and ribavirin for the treatment of chronic hepatitis C.

This new guidance reflects changes in licensing and recommends their wider use and, where appropriate, shorter treatments for adults with the disease.

Hepatitis C

Estimates from the Health Protection Agency suggest that approximately 142,000 people between the ages of 15-59 years had chronic hepatitis C (HCV) in England and Wales in 2003. More than 90% of all newly diagnosed infections in the UK occur in injecting drug users.

People infected with HCV often don’t have any symptoms, but about 20% will develop acute hepatitis and will experience symptoms such as malaise, weakness and anorexia. About 80% of those infected with the virus go on to develop chronic hepatitis.

The rate of progression from mild to severe disease is slow, taking about 20-50 years from the time of infection. About 30% of infected people develop cirrhosis within 20-30 years, and some of these people are at a high risk of developing hepatocellular carcinoma. Some people with end-stage liver disease or hepatocellular carcinoma may require liver transplants.

Revised guidance

The National Institute for Health and Clinical Excellence (NICE) guidance advises that:

• Combination therapy with peginterferon alfa (2a or 2b) and ribavirin is recommended as a treatment option for adults with chronic hepatitis C who have been treated previously with peginterferon alfa (2a or 2b) and ribavirin in combination; or with peginterferon alfa monotherapy, and whose condition either did not respond to treatment, or responded initially to treatment but then relapsed; or for adults who are also infected with HIV
• Shortened courses of combination therapy with peginterferon alfa (2a or 2b) and ribavirin are recommended for the treatment of adults with chronic hepatitis C who have a rapid virological response to treatment at week 4 that is identified by a highly sensitive test, and who are considered suitable for a shortened course of treatment. 

Many people ‘have no idea they have it’

A recent All Party Parliamentary Hepatology Group report on hepatitis C identified the disease as being one of the main reasons for the large rise in the number of people dying from liver disease. The report estimates that there could be as many as 466,000 people living with hepatitis C in the UK.

Sir Andrew Dillon, NICE Chief Executive, says in a statement: “Many people exposed to the disease have no idea they have it since it can remain symptomless for many years. However, hepatitis C is a potentially debilitating condition, and about 80% of those with the virus go on to develop chronic hepatitis, sometimes as long as 50 years after they were first infected. By widening access to these drugs, this guidance will give clinicians and people living with hepatitis C more treatment options.”

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Santaris Pharma A/S Advances miravirsen, the First microRNA-Targeted Drug to Enter Clinical Trials, Into Phase 2 to Treat Patients Infected With Hepatitis C Virus

- Santaris Pharma A/S initiates Phase 2a clinical trial with miravirsen (SPC3649) to assess safety and tolerability in treatment-naive patients with chronic Hepatitis C
 
HOERSHOLM, Denmark and SAN DIEGO, September 22, 2010 /PRNewswire/ -- Santaris Pharma A/S, a clinical-stage biopharmaceutical company focused on the discovery and development of RNA-targeted therapies, today announced that it has advanced miravirsen (SPC3649), the first microRNA-targeted drug to enter clinical trials, into Phase 2 studies to assess the safety and tolerability of the drug in treatment-naive patients infected with the Hepatitis C virus (HCV).
 
Paving the way to conduct the first clinical trials of a microRNA-targeted drug in the United States, Santaris Pharma A/S also received acceptance of its Investigational New Drug (IND) application from the U.S. Food and Drug Administration (FDA). In addition to the United States, the Phase 2a clinical trials will be conducted in the Netherlands, Germany, Poland, Romania, and Slovakia.

The World Health Organization estimates about 3% of the world's population has been infected with HCV and that some 170 million are chronic carriers at risk of developing liver cirrhosis and/or liver cancer(2). Approximately 3-4 million Americans are chronically infected with an estimated 40,000 new infections per year(1). In Europe, there are about 4 million carriers(2). The current standard of care, pegylated interferon in combination with ribavirin, is effective in only about 50% of those treated(1).

Developed using Santaris Pharma A/S proprietary Locked Nucleic Acid (LNA) Drug Platform, miravirsen is a specific inhibitor of miR-122, a liver specific microRNA that the Hepatitis C virus requires for replication. Miravirsen is designed to recognize and sequester miR-122, making it unavailable to the Hepatitis C virus. As a result, the replication of the virus is effectively inhibited and the level of Hepatitis C virus is reduced.

"Advancing miravirsen, the first microRNA-targeted drug to enter clinical trials, into Phase 2 studies in patients with Hepatitis C demonstrates Santaris Pharma A/S leadership in developing RNA-targeted medicines," said Arthur A. Levin, Ph.D., Vice President, Chief Development Officer and President, US Operations. "Receiving IND acceptance from the FDA to conduct the first clinical trials with a microRNA-targeted drug in the United States brings Santaris Pharma A/S one step closer to potentially providing a growing number of patients chronically infected with HCV with a more effective and better tolerated treatment option."

The LNA Drug Platform is the only technology with both mRNA and microRNA targeted drugs in clinical trials, reinforcing the broad utility of the platform. The unique combination of small size and very high affinity, which is only achievable with LNA-based drugs, allows this new class of drugs to potently and specifically inhibit RNA targets in many different tissues without the need for complex delivery vehicles. LNA-based drugs are a promising new type of therapy that enables scientists to develop drugs to attack previously inaccessible pathways.

"Using our LNA Drug Platform to advance the first microRNA-targeted therapy into human clinical trials was certainly a scientific breakthrough," said Henrik Oerum, Ph.D., Vice President and Chief Scientific Officer of Santaris Pharma A/S. "We are extremely pleased with the results of the Phase I trials and excited to progress miravirsen into Phase 2 clinical trials. Because of its unique mechanism of action and tolerability profile, miravirsen has the potential to be an effective treatment option for patients with HCV."

The randomized, double-blind, placebo-controlled, ascending multiple-dose Phase 2a study will assess the safety and tolerability of miravirsen and is designed to enroll up to 55 treatment-naïve patients with chronic Hepatitis C virus genotype 1 infection. Secondary endpoints include pharmacokinetics of miravirsen and its effect on viral load. Miravirsen will be given as subcutaneous injections weekly or every other week for four weeks.

Data from Phase 1 clinical studies with miravirsen in healthy volunteers show that the drug is well tolerated. A recent study published in Science demonstrated that miravirsen successfully inhibited miR-122 and dramatically reduced Hepatitis C virus in the liver and in the bloodstream in chimpanzees chronically infected with the Hepatitis C virus(3). Miravirsen provided continued efficacy in the animals up to several months after the treatment period with no adverse events and no evidence of viral rebound or resistance.

In addition to miravirsen, Santaris Pharma A/S has a robust product pipeline targeting mRNAs and microRNAs both internally as well as in partnerships and collaborations with miRagen Therapeutics (cardiovascular diseases), Shire plc (rare genetic disorders), Pfizer (undisclosed therapeutic areas), GlaxoSmithKline (viral disease) and Enzon Pharmaceuticals (oncology).

About microRNAs

MicroRNAs have emerged as an important class of small RNAs encoded in the genome. They act to control the expression of sets of genes and entire pathways and are thus thought of as master regulators of gene expression. Recent studies have demonstrated that microRNAs are associated with many disease processes. Because they are single molecular entities that dictate the expression of fundamental regulatory pathways, microRNAs represent potential drug targets for controlling many biologic and disease processes.

About Locked Nucleic Acid (LNA) Drug Platform

The LNA Drug Platform and Drug Discovery Engine developed by Santaris Pharma A/S combines the Company's proprietary LNA chemistry with its highly specialized and targeted drug development capabilities to rapidly deliver potent single-stranded LNA-based drug candidates against RNA targets, both mRNA and microRNA, for a range of diseases including metabolic disorders, infectious and inflammatory diseases, cancer and rare genetic disorders. The LNA Drug Platform overcomes the limitations of earlier antisense and siRNA technologies to deliver potent single-stranded LNA-based drug candidates across a multitude of disease states. The unique combination of small size and very high affinity, which is only achievable with LNA-based drugs, allows this new class of drugs to potently and specifically inhibit RNA targets in many different tissues without the need for complex delivery vehicles. LNA-based drugs are a promising new type of therapy that enables scientists to develop drugs to attack previously inaccessible clinical pathways. The most important features of LNA-based drugs include excellent specificity, providing optimal targeting; increased affinity to targets providing improved potency; and strong pharmacology upon systemic delivery without complicated delivery vehicles.

About Santaris Pharma A/S

Santaris Pharma A/S is a privately held clinical-stage biopharmaceutical company focused on the discovery and development of RNA-targeted therapies. The Locked Nucleic Acid (LNA) Drug Platform and Drug Discovery Engine developed by Santaris Pharma A/S combine the Company's proprietary LNA chemistry with its highly specialized and targeted drug development capabilities to rapidly deliver potent single-stranded LNA-based drug candidates across a multitude of disease states. The Company's research and development activities focus on infectious diseases and metabolic disorders, while partnerships with major pharmaceutical companies include a range of therapeutic areas including cancer, cardiovascular disease, infectious and inflammatory diseases, and rare genetic disorders. The Company has strategic partnerships with miRagen Therapeutics, Shire plc, Pfizer, GlaxoSmithKline, and Enzon Pharmaceuticals. As part of its broad patent estate, the Company holds exclusive worldwide rights to all therapeutic uses of LNA. Santaris Pharma A/S, founded in 2003, is headquartered in Denmark with operations in the United States. Please visit http://www.santaris.com/ for more information.

(1) American Association for the Study of Liver Diseases -http://www.aasld.org/patients/Pages/LiverFastFactsHepC.aspx

(2) World Health Organization - http://www.who.int/csr/disease/hepatitis/Hepc.pdf

(3) Science. 2010 Jan 8; 327(5962):198-201. Epub 2009 Dec 3

Source

Prevalence, risk factors and causes of discordance in fibrosis staging by transient elastography and liver biopsy

Liver Int. 2010 Aug 29. [Epub ahead of print]

Myers RP, Crotty P, Pomier-Layrargues G, Ma M, Urbanski SJ, Elkashab M.

Liver Unit, Department of Medicine, Division of Gastroenterology, University of Calgary, Calgary, AB, Canada.

Abstract

Abstract Background and aims: Liver stiffness measurement (LSM) by transient elastography (TE) is widely used for the noninvasive assessment of fibrosis. Our objectives were to examine the prevalence, risk factors and causes of discordance between fibrosis estimated by TE and liver biopsy. Methods: Two hundred and fifty-one patients with hepatitis B, C and nonalcoholic fatty liver disease underwent LSM by TE and liver biopsy. Predictors of discordance (>/=2 fibrosis stages) between measures, which occurred in 14% of patients (n=35), were identified by comparing patient, TE and biopsy characteristics of discordant and nondiscordant cases. Results: According to predefined criteria, 40% of discordances were attributed to TE error and 23% to biopsy error; 37% were indeterminate. In multivariate analysis, mild fibrosis (F0-2 vs. F3-4), and higher body mass index (BMI), ALT and LSM variability [assessed by the ratio of the interquartile range to median LSM (IQR/M)] were independently associated with discordance. Discordance was three-fold more common in patients with obesity (28 vs. 9%), ALT>/=60 U/L (20 vs. 7%) and IQR/M >/=0.17 (22 vs. 7%; all P<0.005). Based on these variables, a discordance risk score assigning 1 point to each factor was developed. The prevalence of discordance in patients with 0, 1, 2 and 3 factors were 2, 7, 20, and 55% respectively (P<0.0005). Conclusions: Discordance between liver fibrosis estimated by TE and biopsy occurs in one in seven patients. In assessing the validity of TE results, clinicians must recognize risk factors for discordance and in at-risk patients, consider alternative measures including biomarkers and possibly biopsy.

PMID: 20807336 [PubMed - as supplied by publisher]

Source

Intensified peginterferon alfa-2a/ribavirin therapy for patients with HCV genotype 1, weight >/=85 kg and high viral load: randomized trial

Gastroenterology. 2010 Sep 2. [Epub ahead of print]

Reddy KR, Shiffman ML, Rodriguez-Torres M, Cheinquer H, Abdurakhmanov D, Bakulin I, Morozov V, Silva GF, Geyvandova N, Stanciu C, Rabbia M, McKenna M, Thommes JA, Harrison SA; PROGRESS Study Investigators.

Division of Gastroenterology, University of Pennsylvania, Philadelphia, PA, USA.

Abstract

BACKGROUND & AIMS: Patients infected with hepatitis C virus (HCV) genotype 1, body weights >/=85 kg and high baseline viral loads respond poorly to standard doses of peginterferon and ribavirin. We evaluated the effects of intensified therapy with peginterferon alfa-2a plus ribavirin.

METHODS: We performed a double-blind, randomized trial of outpatients from hepatology clinics who were infected with HCV genotype 1. Patients in the study had body weights >/=85 kg and HCV RNA titers >/=400,000 IU/mL. Patients were randomized to 180 mug/week peginterferon alfa-2a for 48 weeks in combination with 1200 mg/day ribavirin (standard of care) (group A, N=191) or 1400/1600 mg/day ribavirin (group B, N=189). Additional groups included those who received 360 mug/week peginterferon alfa-2a for 12 weeks and then 180 mug/week peginterferon alfa-2a for 36 weeks, combined with 1200 mg/day ribavirin (group C, N=382) or 1400/1600 mg/day ribavirin (group D, N=383). All patients were followed for 24 weeks after treatment.

RESULTS: Sustained virologic response rates (HCV RNA <15 IU/mL at the end of the follow-up period) in groups A, B, C, and D were 38%, 43%, 44%, and 41%, respectively. There were no significant differences among the 4 groups or between pooled peginterferon alfa-2a regimens (A+B vs. C+D: odds ratio [OR]=1.08, 95% confidence interval [CI]=0.83-1.39, P=0.584) or pooled ribavirin regimens (A+C vs. B+D: OR=1.00, 95% CI=0.79-1.28, P=0.974).

CONCLUSIONS: In patients infected with HCV genotype 1 who are difficult to treat (high viral load and body weight >/=85 kg), a 12-week induction regimen of peginterferon alfa-2a and/or higher-dose ribavirin is not more effective than the standard regimen.

PMID: 20816836 [PubMed - as supplied by publisher]

Source

Amino acid substitution in hepatitis C virus core region and genetic variation near the interleukin 28B gene predict viral response to telaprevir with peginterferon and ribavirin

Hepatology. 2010 Aug;52(2):421-9.

Akuta N, Suzuki F, Hirakawa M, Kawamura Y, Yatsuji H, Sezaki H, Suzuki Y, Hosaka T, Kobayashi M, Kobayashi M, Saitoh S, Arase Y, Ikeda K, Chayama K, Nakamura Y, Kumada H.

Department of Hepatology, Toranomon Hospital, Tokyo, Japan. akuta-gi@umin.ac.jp

Abstract

Genetic variation near the IL28B gene and substitution of amino acid (aa) 70 and 91 in the core region of hepatitis C virus (HCV) genotype 1b can predict the response to pegylated interferon (PEG-IFN)/ribavirin combination therapy, but its impact on triple therapy of telaprevir/PEG-IFN/ribavirin is not clear. The aims of this study were to investigate the predictive factors of sustained virological response to a 12-week or 24-week regimen of triple therapy in 72 of 81 Japanese adults infected with HCV genotype 1. Overall, sustained virological response and end-of-treatment response were achieved by 61% and 89%, respectively. Especially, the sustained virological response was achieved by 45% and 67% in the 12- and 24-week regimens, respectively. Multivariate analysis identified rs8099917 near the IL28B gene (genotype TT) and substitution at aa 70 (Arg70) as significant determinants of sustained virological response. Prediction of response to therapy based on a combination of these factors had high sensitivity, specificity, and positive and negative predictive values. The efficacy of triple therapy was high in the patients with genotype TT, who accomplished sustained virological response (84%), irrespective of substitution of core aa 70. In the patients having genotype non-TT, those of Arg70 gained high sustained virological response (50%), and sustained virological response (12%) was the worst in patients who possessed both genotype non-TT and Gln70(His70). Conclusion: This study identified genetic variation near the IL28B gene and aa substitution of the core region as predictors of sustained virological response to a triple therapy of telaprevir/PEG-IFN/ribavirin in Japanese patients infected with HCV genotype 1b.

PMID: 20648473 [PubMed - indexed for MEDLINE]

Source

Treatment of chronic hepatitis C in HIV-infected patients with compensated liver cirrhosis

J Viral Hepat. 2010 Aug 31. [Epub ahead of print]

Martín-Carbonero L, Tuma P, Vispo E, Medrano J, Labarga P, González-Lahoz J, Barreiro P, Soriano V.

Infectious Diseases Department, Hospital Carlos III, Madrid, Spain.

Abstract

Summary. The greatest benefit of hepatitis C virus (HCV) therapy is seen in cirrhotics attaining sustained virological response (SVR). However, concerns about toxicity and poorer responses often discourage treatment of cirrhotics. This may be particularly relevant in HIV-HCV-coinfected patients, in whom progression of liver fibrosis is faster and treatment responses lower. This is a retrospective analysis of HIV-HCV-coinfected patients who had received peginterferon-ribavirin therapy at our institution. Individuals naïve for interferon in whom liver fibrosis had been assessed using elastometry within the year before being treated were chosen. Response rates and toxicities were compared in cirrhotics (>14.5 KPa) and noncirrhotics. Patients with previous liver decompensation were excluded. Overall, 41 cirrhotics and 190 noncirrhotics entered the study. Groups were similar in age, gender, HCV genotypes and baseline serum HCV-RNA. SVR occurred at similar rates in cirrhotic and noncirrhotics, either considered by intention-to-treat (39%vs 45%; P = 0.4) or as treated (50%vs 52%, P = 0.8). In multivariate analysis (odds ratio, 95% CI, P), SVR was associated with HCV genotypes 2-3 (5, 2.9-11, <0.01) and lower serum HCV-RNA (2, 1.4-3.03 for every log decrease, <0.01) but not with cirrhosis (1.2, 0.4-3.6, 0.6). Treatment discontinuations because of adverse events tended to be more common in cirrhotics than in noncirrhotics (17%vs 12%; P = 0.2), but only severe thrombocytopenia was more frequent in cirrhotics than in non-cirrhotics (20%vs 3% at week 24; P < 0.01). Response to peginterferon-ribavirin therapy is similar in HIV-HCV coinfected patients with and without liver cirrhosis. Therefore, treatment must be encouraged in all compensated cirrhotic patients, although closer monitoring and management of side effects, mainly thrombocytopenia, may be warranted.

PMID: 20819149 [PubMed - as supplied by publisher

Source

The 61st Annual Meeting of the American Association for the Study of Liver Diseases

Boston, MA - Hynes Convention Center
October 30 - November 2, 2010

PR Newswire
ALEXANDRIA, Va., Sept 20

ALEXANDRIA, Va., Sept 20 /PRNewswire/ -- The Liver Meeting® is the premier meeting in the science and practice of hepatology, including the latest findings on new drugs, novel treatments, and the results from pilot and multicenter studies.

Approximately 10 percent of Americans have some form of liver disease, and the diseases strike disproportionately among certain populations but mostly regardless of lifestyle choices. There are now numerous treatments for both hepatitis B and C, and screening, treatment, and prevention of hepatitis remain important issues. Liver cancer is one of the few cancers growing in incidence, and the obesity epidemic has dire consequences for the nation's liver health and wellness.

The 2047 abstracts addressing these issues that will be presented are available to members of the press at our website (http://www.aasld.org/), including 258 abstracts that will be presented in oral sessions.

Boston, MA – October 30 - November 2, 2010
Poster Presentations: October 30 – November 2
Oral Presentations: October 31 – November 2

An AASLD President's press conference highlighting key abstracts and issues presented at the Liver Meeting® is scheduled for Saturday, October 30 at 4:00 pm.

This year's President's Choice Lecture will be given by the 14th Assistant Secretary for Health for the US Department of Health and Human Services, Dr. Howard Koh. The lecture will focus on the findings from the Institute of Medicine (IOM) study, "Hepatitis and Liver Cancer: A National Strategy for Prevention and Control of Hepatitis B and C." Dr. Koh has provided visionary and extraordinary leadership in assembling a US Department of Health and Human Services-wide team to begin to address the epidemic of viral hepatitis in the US. He will speak about his efforts and the steps the US administration is taking in response to the IOM report. AASLD has worked jointly with the Trust for America's Health (TFAH) to translate the IOM report into language that could be used to affect appropriations to support research and health care delivery for liver disease and also to be used in future legislation to make the screening, early detection, and treatment of viral hepatitis a reality. The AASLD/TFAH report will focus on the policy issues that need to be addressed to advance the implementation of the IOM's work.

Founded in 1950, AASLD is the leading organization of scientists and healthcare professionals committed to preventing and curing liver disease. AASLD has grown into an international society responsible for all aspects of hepatology, and our annual meeting attracts 7,500 physicians, surgeons, researchers, and allied health professionals from around the world.

Please contact AASLD at 703-299-9766 for information about the above presentations, or to receive any additional information about The Liver Meeting® – or visit our website at http://www.aasld.org/.

This release was issued through The Xpress Press News Service, merging e-mail and satellite distribution technologies to reach business analysts and media outlets worldwide. For more information, visit http://www.xpresspress.com/.

Contact:
Gregory Bologna, gbologna@aasld.org
Ann Haran, aharan@aasld.org

SOURCE American Association for the Study of Liver Diseases (AASLD)

Read more: http://www.digitaljournal.com/pr/117474#ixzz10CucnSV4
 
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Heterosexual Sex Not a Major Risk Factor for Hepatitis C Virus Transmission

SUMMARY: Sex between women and men does not appear to be a common route of hepatitis C virus (HCV) transmission, according to study results reported last week at the 50th Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC 2010) in Boston. These findings confirm prior research showing a low rate of heterosexual HCV transmission, in contrast with the higher rate reported for HIV positive gay and bisexual men.

By Liz Highleyman

Over the past decade, researchers have reported several outbreaks of acute hepatitis C among men who have sex with men that appear to be due to sexual transmission. This conflicts with public health guidelines stating that sexual transmission of HCV is uncommon, but these were based on studies of monogamous heterosexual couples.

To shed further light on this issue, Monina Klevens from the U.S. Centers for Disease Control and Prevention (CDC) and colleagues collected data from surveillance of new HCV infections reported during 2005-2009 by health departments in Colorado, Connecticut, Minnesota, Oregon, and 34 counties in New York State.

Included cases met clinical criteria (acute illness with at least 1 sign or symptom of viral hepatitis and either jaundice or elevated alanine aminotransferase [ALT]) or laboratory criteria (confirmed positive HCV antibody test) for acute hepatitis C.

The health departments collected demographic and clinical data for each case, and asked patients or their healthcare providers for information about 21 potential HCV risk behaviors occurring 2 weeks to 6 months before the onset of symptoms.

Results

• A total of 575 cases of acute HCV infection were reported.
• 63 cases (11.0%) had no reported risk factors and were excluded from the present analysis.
• Of the remaining 512 cases, 247 patients (48.2%) reported using drugs.
• 202 people (39.5%) reported exposure through heterosexual sex.
• 20 people (3.9%) reported sex with a same-sex partner.
• Most of the infected individuals who reported heterosexual sex (126 of 202) or homosexual sex (14 of 20) also reported drug use.
• Drug use increased with the number of sexual partners: 

          --79.0% of people with > 5 partners;
          --76.5% with 2-5 partners;
          --54.6% with 1 partner.

• 42 out of 202 people (20.8%) reported sexual contact with a person confirmed or suspected to have HCV infection.
• Just 19 out of 202 heterosexuals (9.4%) reported no other risk behaviors other than sex with an opposite-sex partner.
• Individuals who had heterosexual sex as their only risk behavior were significantly older than those with more risk factors (43 vs 35 years, respectively), but otherwise similar including race/ethnicity. 

Based on these findings, the investigators stated that most people with acute HCV infection who had 1 or more heterosexual partners also had other risk factors, and concluded that "heterosexual transmission may not be an important risk factor for HCV in the U.S."

Investigator affiliations: CDC, Atlanta, GA; Colorado Dept. of Public Health and Environment, Denver, CO; Connecticut Dept. of Public Health, Hartford, CT; Minnesota Dept. of Health, St. Paul, MN; New York State Dept. of Health, Albany, NY; Oregon Public Health Div., Portland, OR.

9/21/10

Reference
M Klevens, D Daniels, K Iqbal, and others. Is Heterosexual Transmission an Important Risk Factor for Hepatitis C in the United States? 50th Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC 2010). Boston, September 12-15, 2010. (Abstract V-1787).

Source

NIH Statement on National Gay Men’s HIV/AIDS Awareness Day

From Anthony S. Fauci, M.D., NIAID Director, on September 27, 2010

The third annual National Gay Men’s HIV/AIDS Awareness Day on Sept. 27, 2010, marks an occasion to reflect on how profoundly HIV/AIDS has affected gay and bisexual men. It also is a fitting time to recognize how much this group has influenced the development and implementation of strategies to prevent and treat the virus and the disease.

The HIV/AIDS epidemic continues to exact a terrible toll on gay and bisexual men. In the United States it is estimated that since AIDS was first recognized in 1981, more than half a million gay and bisexual men have been diagnosed with the disease and more than 300,000 have died. Stigma and fear hamper efforts to make HIV/AIDS prevention and treatment accessible to all gay and bisexual men who would benefit from them, both in the United States and abroad. Yet many gay and bisexual men have been instrumental as AIDS activists in raising awareness about the public health impact of HIV/AIDS, shaping the HIV/AIDS research agenda and advocating that this research is well funded. In addition, gay and bisexual men catalyzed the movement to bring treatment and care to people with HIV/AIDS and to promote HIV prevention. Tens of thousands of gay and bisexual men have participated as volunteers in HIV/AIDS research, including several large studies in progress that are funded by NIAID.

To read the full statement, go to http://www.niaid.nih.gov/news/newsreleases/2010/Pages/GayMenAIDSAwareness.aspx.

You are subscribed to News Releases for National Institute of Allergy and Infectious Diseases. This information has recently been updated, and is now available.

Source: Received via email

Importance of Adherence to HCV Regimens

Melissa Palmer, MD
Clinical Professor of Medicine
Department of Hepatology
Director
NYU Hepatology Associates - Plainview
New York University
Plainview, New York

Maximizing sustained virologic response (SVR) rates (an undetectable HCV RNA level 24 weeks after discontinuation of therapy) is the primary goal of therapy for patients with hepatitis C virus (HCV). This endpoint depends on numerous factors (Table), and adherence to therapy is one of the few variables that can be influenced by the patient and/or the healthcare team by using a multidisciplinary approach. Despite the known importance of treatment adherence in achieving viral eradication, adherence continues to be suboptimal, a fact underscored in a recent study demonstrating that only approximately 60% of patients with HCV in the United States adhered to prescribed therapy.[1] The dawn of a new era of HCV treatment is upon us with the anticipated approval of 2 novel direct-acting antivirals (DAAs) in the latter half of 2011. The addition of a DAA to the current standard of care regimen (peginterferon and ribavirin) enhances SVR rates and diminishes length of treatment in many patients.[2-5] However, issues of adherence will likely become even more significant, as the incidence of adverse events may rise, dosing schedules will become more complex, dietary requirements may be needed, and pill burden will increase. Furthermore, the lack of adherence to DAAs may select for HCV drug resistance mutations, a potentially serious consequence not present with the current standard of care regimen.

Table. Factors Affecting SVR Rates

Adherence Affects Response Rates

Suboptimal drug exposure of ribavirin and/or interferon has been demonstrated to result in reduced SVR rates in numerous studies of both treatment-naive and treatment-experienced patients.[6-10] Genotype 1 patients who were unable to complete ≥ 80% of the prescribed doses of both peginterferon and ribavirin ≥ 80% of the time had a 34% SVR rate vs a 51% SVR rate in those who completed > 80% of both drugs > 80% of the time.[7] Consistent with these findings, a retrospective analysis of 188 HCV-infected Veteran Affairs patients found that adherence to ≥ 85% of prescribed peginterferon plus ribavirin positively correlated with degree of viral suppression and ability to achieve early virologic response (defined as a ≥ 2 log reduction from baseline or undetectable serum HCV RNA at Week 12 of therapy).[10]

Ribavirin Adherence

Several studies have shown that suboptimal ribavirin dosing and/or adherence leads to lower response rates. Indeed, studies have demonstrated that exposure to weight-based ribavirin vs fixed-dose ribavirin is associated with improved early virologic response and SVR rates.[11,12] In addition, a retrospective analysis of 5 clinical trials concluded that patients with genotype 1 HCV exposed to > 13 mg/kg/day of ribavirin had a 2.15-times greater chance of achieving an undetectable HCV RNA by Week 4 (rapid virologic response [RVR]) vs patients exposed to < 13 mg/kg/day of ribavirin (Capsule Summary).[13] Furthermore, genotype 1 patients not achieving RVR were statistically significantly less likely to achieve SVR when cumulative ribavirin dose exposure (due to dose reduction, premature cessation, or skipped doses) was < 60%.[6] In a study by Bronowicki and colleagues,[14] Patients with genotype 1 HCV who discontinued ribavirin after achieving an undetectable HCV RNA by Week 24 were significantly less likely to achieve an SVR vs patients who continued their ribavirin therapy (52.8% vs 68.2%, respectively; P = .004). Finally, dose reduction of ribavirin during the initial 12-20 weeks of therapy has been associated with diminished SVR rates in genotype 1 patients.[7,15,16]

Whereas cumulative ribavirin dose exposure was found not to influence RVR and SVR in genotype 2 patients,[17] the importance of maintaining adequate doses of ribavirin throughout the entire course of treatment was emphasized by Andriulli and colleagues[18] who found that patients with genotype 2/3 HCV had a significantly diminished SVR (54% vs 82%) when ribavirin was discontinued by Week 6 of treatment, even if RVR was achieved (Capsule Summary).

The primary role of ribavirin in HCV treatment is to reduce the likelihood of relapse. Indeed, in an evaluation of 984 genotype 1 HCV–infected patients treated with peginterferon alfa-2b plus ribavirin, the degree of ribavirin exposure was found to be inversely correlated with relapse rates. Only 11% of patients receiving ≥ 12 mg/kg/day of ribavirin relapsed compared with 60% of those receiving < 6 mg/kg/day.[19] In addition, only 4% of patients with an early virologic response in this trial relapsed when exposed to ribavirin doses > 12 mg/kg/day, and ribavirin exposure beyond Week 12 continued to affect relapse rates, demonstrating that ribavirin is inversely dose dependent correlated with relapse in patients with genotype 1 HCV responding to peginterferon plus ribavirin. This study demonstrated the importance of adhering to a dose of ≥ 12 mg/kg/day of ribavirin during the entire treatment period, especially in patients achieving an early virologic response.

Despite these findings, ribavirin adherence continues to be a challenge that appears to be more difficult to overcome than interferon adherence.[20-22] The effect of simplifying ribavirin pill burden was assessed in 2 studies comparing adherence rates in HCV-infected patients treated with fewer, higher-dose ribavirin tablets—a 400-mg or 600-mg ribavirin tablet available in a unit dose blister pack—vs the traditional 200-mg bottled ribavirin tablets. Results from an observational study of 92 patients with HCV from a single center found that patients taking peginterferon with the more compact formulation of ribavirin allowing a reduced pill burden (same total dose) experienced fewer adverse events, improved quality of life, better adherence to prescribed HCV therapy, and a trend toward higher SVR rates vs patients taking peginterferon plus the standard dosing of ribavirin.[23] In the multicenter Accurate Dosing in Hepatitis C: Examining the RibaPak Experience (ADHERE) study, patients who received their ribavirin in the form of the higher-dose tablets provided in a unit dose blister pack were less likely to prematurely stop therapy and more likely to adhere to prescribed HCV treatment at Weeks 12 and 24 compared with patients taking traditional 200-mg ribavirin tablets.[24] Streamlining ribavirin dosing regimens by decreasing pill count may become an even more important factor contributing to enhanced adherence once DAAs become part of HCV therapy, further increasing the regimen pill burden.

Assessment of Adherence

Although there is no gold standard of adherence measurement, it is important for all clinicians to incorporate strategies into their practices to assess adherence to prescribed HCV medications and to differentiate nonadherent patients from those who are truly nonresponders. Adherence analysis by a combination of patient self-report and electronic monitoring of the 401 patients who participated in the Viral Resistance to Antiviral Therapy of Chronic Hepatitis C (VIRAHEP-C) trial revealed that patient self-report overestimates compliance, that adherence wanes over time, and that patients are more likely to miss doses of ribavirin than peginterferon.[20] When pharmacy refill data during the initial 12 weeks of treatment was retrospectively evaluated in 188 predominantly HCV-monoinfected US veterans, adherence to peginterferon was found to be better than adherence to ribavirin—a finding consistent with other adherence studies.[10] A prospective, real-life, observational study of genotype 2 and 3 patients revealed that by 3 months of treatment, 20% of HCV patients self-reported missing 1 peginterferon injection whereas approximately 28% of patients self-reported missing 1200-mg ribavirin within the previous 4-week treatment period and that missed dosages of both agents increased at 6 months of treatment.[21] Of importance, this study also demonstrated that adherence at 6 months was better in those receiving patient therapeutic education during the initial 12 weeks of therapy compared with those not receiving patient education.

Suboptimal Adherence Identification and Management

In addition to reinforcing the importance of treatment adherence to patients, identification of individuals at increased risk for adverse events and aggressive management of adverse events are also necessary components of good clinical care that will enhance patient outcomes.

More than 20% of patients must decrease dose, temporarily discontinue, or prematurely stop ribavirin and/or peginterferon because of adverse events.[16,25,26] However, certain subsets of individuals may have more difficulty tolerating treatment compared with others. Mitra and colleagues[1] found that patients with advanced disease are less likely to be adherent to HCV therapy than those with early disease, a finding likely related to an increased susceptibility to adverse events.[27] A recent study showed that 61% of HCV/HIV-coinfected women and 48% of coinfected men experienced adverse events requiring dose modification of standard of care therapy.[28] Whereas traditionally believed to be poor candidates for HCV treatment because of an increased likelihood of nonadherence and susceptibility to psychiatric adverse events associated with treatment, injection drug users, including active drug users, have been found to be adherent to treatment with comparable SVR rates so long as psychosocial support is supplied.[29-31]

Hepatitis C virus is more common among people with psychiatric conditions compared with the general population,[32] and psychiatric illness, in particular depression, has been found to be a risk factor for HCV medication nonadherence. Development of depression while receiving therapy, which has been noted to occur in between 20% to 35% of patients,[25,26] may lead to discontinuation or decrease in dosage of medication with a resultant negative impact on outcome.[33,34] Pretreatment with antidepressants, specifically paroxetine, has been shown to reduce the severity of depression while receiving treatment in individuals displaying depressive symptoms at baseline.[35] The initiation of citalopram after the onset of interferon-induced depression controlled symptoms and enabled patients to complete the full course of standard-of-care treatment.[36] Therefore, to improve adherence, practitioners must evaluate all HCV-infected patients at baseline and during therapy for psychiatric symptoms and initiate antidepressants or antipsychotics promptly, as needed.

Anemia is an adverse effect of both interferon and ribavirin, with a decrease of hemoglobin of > 3 g/dL occurring in more than one half of patients receiving standard-of-care treatment.[37] Anemia often results in dose reduction or discontinuation, particularly of ribavirin, which may negatively affect outcome, especially in patients with genotype 1 HCV.[25] Reduction of the incidence and severity of treatment-induced anemia with erythropoietin has been shown to encourage adherence and to enable patients to remain on higher doses of ribavirin.[37,38] In a prospective, randomized, controlled trial evaluating the effect of erythropoietin on SVR, it was demonstrated that erythropoietin improved quality of life and improved SVR rates in some—but not all—patients with HCV.[8] Rapid virologic response rates did not improve significantly in HCV/HIV-coinfected patients preemptively given erythropoietin.[39] However, in HCV-infected patients achieving a > 2 log decline in HCV RNA after 4 weeks of therapy, SVR rates were significantly improved when anemia was managed with erythropoietin vs ribavirin dose reductions (81.8% vs 45.0%, respectively).[40] Therefore, treatment with erythropoietin should be considered in specific patients as a tool for enhancing medication adherence and improving outcomes, with an eye toward avoiding overcorrection that may result in thrombotic events.[41]

Conclusion

Although great excitement exists surrounding the future of HCV treatment, it must be emphasized that the increased complexity of treatment regimens, coupled with an associated rise in incidence of adverse events and the risk for development of drug resistance, will require even greater attention to adherence issues. Since adherence to prescribed HCV medication has been shown to be enhanced by a combination of patient education, patient motivation, reduced pill burden, and careful and prompt adverse effect management, clinicians should consider a multidisciplinary team approach to assess and enforce adherence and to optimize HCV patient care.

Melissa Palmer, MD, has disclosed that she has received grants for research support from Bristol-Myers Squibb, Gilead Sciences, and Three Rivers Pharmaceuticals; has served as a consultant for Genentech, Gilead Sciences, Merck, and Three Rivers Pharmaceuticals; and has received fees for non-CME services from Genentech, Gilead Sciences, Merck, and Three Rivers Pharmaceuticals.

References

1. Mitra D, Davis KL, Beam C, Medjedovic J, Rustgi V. Treatment patterns and adherence among patients with chronic hepatitis C virus in a US managed care population. Value Health. 2010;13:479-486.

2. McHutchison JG, Manns MP, Muir AJ, et al. Telaprevir for previously treated chronic HCV infection. N Engl J Med. 2010;362:1292-1303.

3. Suzuki F, Akuta N, Suzuki Y, et al. Rapid loss of hepatitis C virus genotype 1b from serum in patients receiving a triple treatment with telaprevir (MP-424), pegylated interferon and ribavirin for 12 weeks. Hepatol Res. 2009;39:1056-1063.

4. Kwo PY, Lawitz EJ, McCone J, et al. Efficacy of boceprevir, an NS3 protease inhibitor, in combination with peginterferon alfa-2b and ribavirin in treatment-naive patients with genotype 1 hepatitis C infection (SPRINT-1): an open-label, randomised, multicentre phase 2 trial. Lancet. 2010;[Epub ahead of print].

5. Sarrazin C, Rouzier R, Wagner F, et al. SCH 503034, a novel hepatitis C virus protease inhibitor, plus pegylated interferon alpha-2b for genotype 1 nonresponders. Gastroenterology. 2007;132:1270-1278.

6. Reddy KR, Shiffman ML, Morgan TR, et al. Impact of ribavirin dose reductions in hepatitis C virus genotype 1 patients completing peginterferon alfa-2a/ribavirin treatment. Clin Gastroenterol Hepatol. 2007;5:124-129.

7. McHutchison JG, Manns M, Patel K, et al. Adherence to combination therapy enhances sustained response in genotype-1-infected patients with chronic hepatitis C. Gastroenterology. 2002;123:1061-1069.

8. Shiffman ML, Ghany MG, Morgan TR, et al. Impact of reducing peginterferon alfa-2a and ribavirin dose during retreatment in patients with chronic hepatitis C. Gastroenterology. 2007;132:103-112.

9. Raptopoulou M, Tsantoulas D, Vafiadi I, et al. The effect of adherence to therapy on sustained response in daily or three times a week interferon alpha-2b plus ribavirin treatment of naive and nonresponder chronic hepatitis C patients. J Viral Hepat. 2005;12:91-95.
10. Lo Re V 3rd, Amorosa VK, Localio AR, et al. Adherence to hepatitis C virus therapy and early virologic outcomes. Clin Infect Dis. 2009;48:186-193.

11. Bain VG, Lee SS, Peltekian K, et al. Clinical trial: exposure to ribavirin predicts EVR and SVR in patients with HCV genotype 1 infection treated with peginterferon alfa-2a plus ribavirin. Aliment Pharmacol Ther. 2008; 28:43-50.

12. Jacobson IM, Brown RS Jr, Freilich B, et al. Peginterferon alfa-2b and weight-based or flat-dose ribavirin in chronic hepatitis C patients: a randomized trial. Hepatology. 2007;46:971-981.

13. Rodriguez-Torres M, Sulkowski M, Chung RT, Hamzeh FM, Jensen DM. Association of pre-treatment and on-treatment factors with rapid virologic response in HCV genotype 1 infected patients treated with peginterferon alfa-2a/ribavirin. Program and abstracts of the 58th Annual Meeting of the American Association for the Study of Liver Diseases; November 2-6, 2007; Boston, Massachusetts. Abstract 1305.

14. Bronowicki JP, Ouzan D, Asselah T, et al. Effect of ribavirin in genotype 1 patients with HCV responding to pegylated interferon alfa a plus ribavirin. Gastroenterology. 2006;131:1040-1048.

15. Davis GL, Wong JB, McHutchison JG, Manns MP, Harvey J, Albrecht J. Early virologic response to treatment with peginterferon alfa-2b plus ribavirin in patients with chronic hepatitis C. Hepatology. 2003;38:645-652.

16. Shiffman ML. Side effects of medical therapy for chronic hepatitis C. Ann Hepatol. 2004;3:5-10.

17. Inoue Y, Hiramatsu N, Oze T, et al. Factors affecting efficacy in patients with genotype 2 chronic hepatitis C treated by pegylated interferon alpha-2b and ribavirin: reducing drug doses has no impact on rapid and sustained virological responses. J Viral Hepat. 2010;17:336-344.

18. Andriulli A, Cursaro C, Cozzolongo R, et al. Early discontinuation of ribavirin in HCV-2 and HCV-3 patients responding to peg-interferon alfa-2a and ribavirin. Program and abstracts of the 58th Annual Meeting of the American Association for the Study of Liver Diseases; November 2-6, 2007; Boston, Massachusetts. Abstract 234.

19. Hiramatsu N, Oze T, Yakushijin T, et al. Ribavirin dose reduction raises relapse rate dose-dependently in genotype 1 patients with hepatitis C responding to pegylated interferon alpha-2b plus ribavirin. J Viral Hepat. 2009;16:586-594.

20. Smith SR, Wahed AS, Kelley SS, Conjeevaram HS, Robuck PR, Fried MW. Assessing the validity of self-reported medication adherence in hepatitis C treatment. Ann Pharmacother. 2007;41:1116-1123.

21. Cacoub P, Ouzan D, Melin P, et al. Patient education improves adherence to peg-interferon and ribavirin in chronic genotype 2 or 3 hepatitis C virus infection: a prospective, real-life, observational study. World J Gastroenterol. 2008;14:6195-6203.

22. Weiss JJ, Bhatti L, Dieterich DT, et al. Hepatitis C patients' self-reported adherence to treatment with pegylated interferon and ribavirin. Aliment Pharmacol Ther. 2008;28:289-293.

23. Palmer M. Improvement in treatment adherence in patients with chronic hepatitis C. Practical Gastroenterology. 2008;32:31-42.

24. Alam I, Stainbrook T, Cecil B, Kistler KD. Enhanced adherence to HCV therapy with higher dose ribavirin formulation: final analyses from the ADHERE registry. Aliment Pharmacol Ther. 2010;32:535-542.

25. Manns MP, McHutchison JG, Gordon SC, et al. Peginterferon alfa 2b plus ribavirin compared with interferon alfa 2b plus ribavirin for the initial treatment of chronic hepatitis C: a randomized trial. Lancet. 2001;358:958-965.

26. Fried MW, Shiffman ML, Reddy KR, et al. Peginterferon alfa-2a plus ribavirin for chronic hepatitis C virus infection. N Engl J Med. 2002;347:975-982.

27. Crippin JS, McCashland T, Terrault N, Sheiner P, Charlton MR. A pilot study of the tolerability and efficacy of antiviral therapy in hepatitis C virus-infected patients awaiting liver transplantation. Liver Transpl. 2002;8:350-355.

28. Bhattacharya D, Umbleja T, Carrat F, et al. Women experience higher rates of adverse events during hepatitis C virus therapy in HIV infection: a meta-analysis. J Acquir Immune Defic Syndr. 2010;[Epub ahead of print].

29. Robaeys G, Van Vlierberghe H, Matheï C, et al. Similar compliance and effect of treatment in chronic hepatitis C resulting from intravenous drug use in comparison with other infection causes. Eur J Gastroenterol Hepatol. 2006;18:159-166.

30. Bruggmann P, Falcato L, Dober S, et al. Active intravenous drug use during chronic hepatitis C therapy does not reduce sustained virological response rates in adherent patients. J Viral Hepat. 2008;15:747-752.

31. Melin P, Chousterman M, Fontanges T, et al. Effectiveness of chronic hepatitis C treatment in drug users in routine clinical practice: results of a prospective cohort study. Eur J Gastroenterol Hepatol. 2010;22:1050-1057.

32. Brunette MF, Drake RE, Marsh BJ, et al. Responding to blood-borne infections among persons with severe mental illness. Psychiatr Serv. 2003;54:860-865.

33. Raison CL, Broadwell SD, Borisov AS, et al. Depressive symptoms and viral clearance in patients receiving interferon-a and ribavirin for hepatitis C. Brain Behav Immun. 2005:19:23-27.

34. Sylvestre D. Treating hepatitis C in active substance users. Clin Infect Dis. 2005;40(suppl 5):S321-S324.

35. Raison CL, Woolwine BJ, Demetrashvili MF, et al. Paroxetine for prevention of depressive symptoms induced by interferon-alpha and ribavirin for hepatitis C. Aliment Pharmacol Ther. 2007;25:1163-1174.

36. Kraus MR, Schäfer A, Schöttker K, et al. Therapy of interferon-induced depression in chronic hepatitis C with citalopram: a randomised, double-blind, placebo-controlled study. Gut. 2008;57:531-536.

37. Dieterich DT, Wasserman R, Bräu N, et al. Once-weekly epoetin alfa improves anemia and facilitates maintenance of ribavirin dosing in hepatitis C virus-infected patients receiving ribavirin plus interferon alfa. Am J Gastroenterol. 2003;98:2491-2499.

38. Afdhal NH. Role of epoetin alfa in maintaining ribavirin dose. Gastroenterol Clin North Am. 2004;33(1 suppl):S25-S35.

39. Vispo E, Labarga P, Guardiola JM, et al. Preemptive erythropoietin plus high ribavirin doses to increase rapid virological responses in HIV patients treated for chronic hepatitis C. AIDS Res Hum Retroviruses. 2010;26:419-424.

40. Falasca K, Ucciferri C, Mancino P, Gorgoretti V, Pizzigallo E, Vecchiet J. Use of epoetin beta during combination therapy of infection with hepatitis c virus with ribavirin improves a sustained viral response. J Med Virol. 2010;82:49-56.

41. Fandrey J, Dicato M. Examining the involvement of erythropoiesis-stimulating agents in tumor proliferation (erythropoietin receptors, receptor binding, signal transduction), angiogenesis, and venous thromboembolic events. Oncologist. 2009;14(suppl 1):34-42.

Source