Jaquelyn F. Fleckenstein

Naturally Occurring Dominant Resistance Mutations to Hepatitis C Virus Protease and Polymerase Inhibitors in Treatment-Naïve Patients

Resistance mutations to hepatitis C virus (HCV) nonstructural protein 3 (NS3) protease inhibitors in <1% of the viral quasispecies may still allow >1000‐fold viral load reductions upon treatment, consistent with their reported reduced replicative fitness in vitro. Recently, however, an R155K protease mutation was reported as the dominant quasispecies in a treatment‐naïve individual, raising concerns about possible full drug resistance. To investigate the prevalence of dominant resistance mutations against specifically targeted antiviral therapy for HCV (STAT‐C) in the population, we analyzed HCV genome sequences from 507 treatment‐naïve patients infected with HCV genotype 1 from the United States, Germany, and Switzerland. Phylogenetic sequence analysis and viral load data were used to identify the possible spread of replication‐competent, drug‐resistant viral strains in the population and to infer the consequences of these mutations upon viral replication in vivo. Mutations described to confer resistance to the protease inhibitors Telaprevir, BILN2061, ITMN‐191, SCH6 and Boceprevir; the NS5B polymerase inhibitor AG‐021541; and to the NS4A antagonist ACH‐806 were observed mostly as sporadic, unrelated cases, at frequencies between 0.3% and 2.8% in the population, including two patients with possible multidrug resistance. Collectively, however, 8.6% of the patients infected with genotype 1a and 1.4% of those infected with genotype 1b carried at least one dominant resistance mutation. Viral loads were high in the majority of these patients, suggesting that drug‐resistant viral strains might achieve replication levels comparable to nonresistant viruses in vivo. Conclusion: Naturally occurring dominant STAT‐C resistance mutations are common in treatment‐naïve patients infected with HCV genotype 1. Their influence on treatment outcome should further be characterized to evaluate possible benefits of drug resistance testing for individual tailoring of drug combinations when treatment options are limited due to previous nonresponse to peginterferon and ribavirin. (HEPATOLOGY 2008;48:1769–1778.)

The Induction of Type I Interferon Production in Hepatitis C–Infected Patients

Chronic infection with hepatitis C virus (HCV) is a major global health problem. One way HCV may evade the host immune response is by inhibiting the production of type I interferon (IFN). In addition, the standard treatment for chronic HCV infection involves treatment with IFN-alpha (or its pegylated derivative), alone or in combination with ribavirin. Therefore, it is believed that an important reason that most HCV-infected individuals progress from acute to chronic infection is due to a defect in the host response. In this study, we examined the host response to HCV infection in a cohort of patients enrolled in the UTHSC Cooperative HCV Research Center by determining levels of biologically active IFN in the sera of patients. We found that 15 of 35 enrolled HCV-infected patients show serum levels of IFN (ranging from 2 to 40 IU/mL) before initiation of therapy. Uninfected individuals do not have circulating levels of IFN. Basal IFN levels do not correlate with the clinical response to therapy, nor do they reflect the age, sex, or race of patients. These results suggest that the differential response of patients most likely reflects a defect in the later stages of the host innate immune response, such as the cellular response to endogenous or exogenous IFN. In contrast, the early stage of the host immune response in vivo of many HCV-infected patients (approximately 40%) is intact as determined by IFN production.

Thrombocytopenia Associated with Octreotide

A 42-year-old woman with a history of hepatitis C-induced cirrhosis, gastrointestinal bleeding, and alcohol abuse presented to the hospital with hematemesis and melena. Based on our previous experience, octreotide (Sandostatin) therapy was started at 50 mg/hr and continued for 5 days. Platelet count on admission (122 x 10(9)/L) dropped immediately after octreotide therapy was started; upon discontinuation, platelet count began trending up from 72 x 10(9)/L. However, octreotide was not suspected at this point as the cause of thrombocytopenia. In a subsequent admission, octreotide was again administered with a resultant prompt decrease in platelet count. To our knowledge, this is only the second case report of octreotide-induced thrombocytopenia, and the first case of this adverse effect demonstrated by inadvertent rechallenge.

An interferon response gene signature is associated with the therapeutic response of hepatitis C patients

Infection with the hepatitis C virus (HCV) is a major cause of chronic liver diseases and hepatocellular carcinoma worldwide, and thus represents a significant public health problem. The type I interferon (IFN), IFNα, has been successful in treating HCV-infected patients, but current IFN-based treatment regimens for HCV have suboptimal efficacy, and relatively little is known about why IFN therapy eliminates the virus in some patients but not in others. Therefore, it is critical to understand the basic mechanisms that underlie the therapeutic resistance to IFN action in HCV-infected individuals, and there is an urgent need to identify those patients most likely to respond to IFN therapy for HCV. To characterize the response of HCV-infected patients to treatment with IFNα, the expression of an IFN-response gene signature comprised of IFN-stimulated genes and genes that play an important role in the innate immune response was examined in liver biopsies from HCV-infected patients enrolled in a clinical trial. In the present study we found that the expression of a subset of IFN-response genes was dysregulated in liver biopsy samples from nonresponsive hepatitis C patients as compared with virologic responders. Based on these findings, a statistical model was developed to help predict the response of patients to IFN therapy, and compared to results obtained to the IL28 mutation model, which is highly predictive of the response to IFN-based therapy in HCV-infected patients. We found that a model incorporating gene expression data can improve predictions of IFN responsiveness compared to IL28 mutation status alone.

Hepatitis C Hypervariable Region 1: Association of Reduced Selection Pressure in African Americans with Treatment Failure

In a prospective therapeutic trial, features of the hepatitis C quasispecies were investigated as possible markers of therapeutic response. Individuals chronically infected with hepatitis C genotype 1 received antiviral therapy consisting of α-interferon plus ribavirin. The study targeted the most rapidly evolving segment of the viral genome, hypervariable region 1 within the envelope-2 gene. Among individuals failing to clear virus in response to therapy, significant differences were observed between quasispecies of African-American and Caucasian subjects. While distance measures for synonymous substitutions were similar between racial subgroups, measures of distance at the amino acid level (nonsynonymous substitutions) varied significantly. Taken together, the observed patterns of variability corresponded to reduced host selection pressure against hypervariable region 1 in African-American nonresponders. Reduced selection pressure was present at baseline and persisted through treatment and follow-up, suggesting population stratification of host factors that influence selection pressure on hepatitis C virus.

Vitamin D‐binding protein gene polymorphisms may contribute to the racial disparity in genotype 1 chronic hepatitis C treatment outcome

We read with great interest the study by Falleti et al. in which the authors report that the rs7041 G>T and rs4588 C>A single nucleotide polymorphisms (SNPs) of the vitamin D-binding protein gene are predictors of the treatment outcome of patients with chronic hepatitis C. The authors found that patients with any combination of three or more rs7041 G and rs4588 C alleles (wild type [WT1]) achieve a sustained virologic response (SVR) at a greater rate than patients with other genotypes (WT2). Additionally, they found that the relative frequency of the rs7041 T allele was increased significantly among the patients when compared with its frequency among healthy controls, raising the possibility that rs7041 T alleles are associated with increased susceptibility to infection or a lower rate of spontaneous viral clearance. Importantly, there is biologic plausibility for these findings in that each of the protein isoforms encoded by the variants of the vitamin D-binding protein gene studied by Falleti et al. differentially affect certain components of the immune response. In light of these findings, we sought to determine if there are racial differences in the distribution of the alleles of these SNPs among patients with chronic hepatitis C that could potentially contribute to the racial disparity in treatment outcome. We genotyped the rs7041 G>T and rs4588 C>A SNPs in 48 Caucasian and 95 African American genotype 1 chronic hepatitis C patients. Most notable were our findings concerning the rs7041 G>T SNP. The frequency of the rs7041 G>T alleles among the Caucasians was G 5 0.542 and T 5 0.458. This is similar to the frequency among the patients studied by Falleti et al., all of whom were Caucasian, of G 5 0.553 and T 5 0.447. However, the rs7041 G>T allele frequency among the African Americans we studied was G 5 0.147 and T 5 0.853, which is significantly different than that of the Caucasians we studied (P< 0.0001) and the patients studied by Falleti et al. (P< 0.0001). We then grouped our patients into the WT1 and WT2 categories as defined by Falleti et al. (Table 1). Of the Caucasian patients we studied, 26/48 (54.2%) were WT1. This is similar to the finding by Falleti et al. that 100/206 (48.5%) of their patients were WT1. In contrast, however, only 25/95 (26.3%) of the African American patients in our study were WT1, which is significantly less than the frequency of the WT1 genotype in both our group of Caucasian patients (P 5 0.0016) and the patients examined by Falleti et al. (P 5 0.0003). Although our study was not sufficiently powered to confirm that vitamin D-binding protein genotype predicts antiviral treatment outcome independently of race, when considered with the findings of Falleti et al. our findings raise the possibility that racial variations in distribution of vitamin D-binding protein gene SNP alleles contribute to the racial disparity in treatment outcome in patients with chronic hepatitis C. Further studies are warranted.