Hepatitis C Virus

Abstract: Simeprevir (TMC435, Olysio™), a second-generation hepatitis C virus (HCV) protease inhibitor, has been recently approved for the treatment of genotype 1 chronic hepatitis C in combination with pegylated interferon and ribavirin. This molecule has very different char- acteristics from first-generation protease inhibitors. Results from trials show that simeprevir is highly effective and safe, with few adverse events. We discuss the specific features of this new treatment option for HCV infection, in terms of in vitro data, pharmacological data, and clini- cal trials. We also discuss the impact of Q80K polymorphism at baseline. Studies evaluating interferon-free regimens with simeprevir are ongoing. Future combinations of two or more direct-acting antiviral agents, targeting different viral enzymes and with synergistic antiviral effects, will be approved, allowing treatment of pan-genotypic HCV with optimized sustained virologic responses. Simeprevir will undoubtedly be part of future treatment strategies. Keywords: simeprevir, protease inhibitor, direct-acting antiviral agent, hepatitis C virus

24. Takahashi K, Asabe S, Wieland S, Garaigorta U, Gastaminza P, Isogawa M, Chisari FV. 2010. Plasmacytoid dendritic cells sense hepatitis C virus- infected cells, produce interferon, and inhibit infection. Proc. Natl. Acad. Sci. U. S. A. 107:7431–7436. http://dx.doi.org/10.1073/pnas.1002301107. 25. Negash AA, Ramos HJ, Crochet N, Lau DT-Y, Doehle B, Papic N, Delker DA, Jo J, Bertoletti A, Hagedorn CH, Gale M. 2013. IL-1 ␤ production through the NLRP3 inflammasome by hepatic macrophages links hepatitis C virus infection with liver inflammation and disease. PLoS Pathog. 9:e1003330. http://dx.doi.org/10.1371/journal.ppat.1003330. 26. Shrivastava S, Mukherjee A, Ray R, Ray RB. 2013. Hepatitis C virus

Abstract: The prevalence of hepatitis C virus (HCV) infection in hemodialysis units (HD) is higher than among normal population. Seroconversion was included in many previous studies which constitute a great problem against infection control policies. The aim of this study was to evaluate seroconversion rate and the effect of isolation of hepatitis C positive patients as infection control method. This is controlled prospective study that included 90 patients for 2 years. Isolation policy of hepatitis C positive patients was implemented in the second year of the study. The prevalence of HCV was 48.9% among hemodialysis patients. Seroconversion rate decreased from 15.2% in the first year to 5.1%in the second year after application of isolation. The duration of hemodialysis in months, positive history of blood transfusion and the amount of transfused blood were considered significant factors affecting seroconversion. so we concluded that Isolation of hepatitis C positive patients as an infection control policy is mandatory to control HCV seroconversion in Egypt.

The E2 envelope glycoprotein of hepatitis C virus (HCV) binds to the host entry factor CD81 and is the principal target for neu- tralizing antibodies (NAbs). Most NAbs recognize hypervariable region 1 on E2, which undergoes frequent mutation, thereby allowing the virus to evade neutralization. Consequently, there is great interest in NAbs that target conserved epitopes. One such NAb is AP33, a mouse monoclonal antibody that recognizes a conserved, linear epitope on E2 and potently neutralizes a broad range of HCV genotypes. In this study, the X-ray structure of AP33 Fab in complex with an epitope peptide spanning residues 412 to 423 of HCV E2 was determined to 1.8 Å. In the complex, the peptide adopts a ␤-hairpin conformation and docks into a deep binding pocket on the antibody. The major determinants of antibody recognition are E2 residues L413, N415, G418, and W420. The structure is compared to the recently described HCV1 Fab in complex with the same epitope. Interestingly, the anti- gen-binding sites of HCV1 and AP33 are completely different, whereas the peptide conformation is very similar in the two struc- tures. Mutagenesis of the peptide-binding residues on AP33 confirmed that these residues are also critical for AP33 recognition of whole E2, confirming that the peptide-bound structure truly represents AP33 interaction with the intact glycoprotein. The slightly conformation-sensitive character of the AP33-E2 interaction was explored by cross-competition analysis and alanine- scanning mutagenesis. The structural details of this neutralizing epitope provide a starting point for the design of an immuno- gen capable of eliciting AP33-like antibodies.

The hepatitis C virus NS2/3 protein is a highly hydrophobic protease responsible for the cleavage of the viral polypeptide between non-structural proteins NS2 and NS3. However, many aspects of the NS2/3 protease’s role in the viral life cycle and mechanism of action remain unknown. Based on the recently elucidated crystal structure of NS2, NS2/3 has been proposed to function as a cysteine protease despite its lack of sequence homology to proteases of known function. In addition, although shown to be required for HCV genome replication and persistent infection in a chimpanzee, the role of NS2/3 cleavage in the viral life cycle has not yet been fully investigated. However, several recent studies are beginning to clarify possible roles of the cleaved NS2 protein in modulation of host cell gene expression and apoptosis.

Viral hepatitis is one of the major global public health concerns. According to international reports, the prevalence of hepatitis B virus (HBV) in Iran is moderate (1, 2). Although they are hepatic viruses, these viruses have also been found in extrahepatic tissues such as kidneys, ovaries, and testis in the semen. Viral cirrhosis or hepatocellular carcinoma and even death had been reported as pathological complications among patients with chronic viral hepatitis (3). Infertility is a common issue in the world and according to the World Health Organization (WHO) reports, there are around 80 million infertile couples around the world (4). Overall infertility rate and its specific rate among 21-26 years old Iranian male population is 10-15% and 17.2%, respectively (4, 5). The relationship between HBV and hepatitis C virus (HCV) infection and male infertility has been a considerable interest, as the impact of HBV and HCV on the sperm parameters have been studied by many researchers. Lorusso and co-workers reported that sperm concentration, morphology, and viability have been damaged among HBV seropositive infertile men (6). Other studies in this filed showed that some of sperm parameters were impaired in infertile men with chronic HBV infection, and even there was a negative correlation between HBV viral load and sperm parameters (7, 8). Antisperm antibodies were detected among 12% of men who were assessed for infertility in Egyptain patients with HCV infection (9), and in the most of viral hepatitis patients, and antisperm antibodies had been bound to their spermatozoa (10). Hepatic viruses have a negative impact on the passing of the sperm through the female genitalia and reaching to fertilizing space (11). In some cases, exposing sperms with HBV can induce oxidative stress in the cells and may lead to apoptosis. However, there are few studies about the impact of HBV and HCV infection on sperm parameters of infertile men in the national infertility clinics in Iran.

35. Xess A, Kumar M, Minz S, Sharma H, Shahi S. Prevalence of hepatitis B and hepatitis C virus coinfection in chronic liver disease. Indian Journal of Pathology and Microbiology. 2001;44(3):253-5. 36. Pramoolsinsap C, Sirikulchayanonta V, Busakorn W, Poovorawan Y, Hirsch P, Theamboonlers A, et al. Coinfections with hepatitis g and/or c virus in hepatitis B-related chronic liver disease. J Trop Med Public Health. 1999;30:741-9. 37. Semnani S, Roshandel G, Abdolahi N, Besharat S, Keshtkar AA, Joshaghani H, et al. Hepatitis B/C virus co-infection in Iran: a seroepidemiological study. Turk J Gastroenterol. 2007;18(1):20-1.

Hepatitis C virus (HCV) is a major causative agent of chronic liver disease and thus a major public health problem, infecting at least 3% of the world population (47). HCV in- fection proceeds to the persistent stage in approximately 80% of patients, leading to the development of cirrhosis in 20% to 50% of patients, of whom approximately 5% eventually de- velop hepatocellular carcinoma (12). HCV encompasses a sin- gle-stranded positive-sense RNA genome of approximately 9.6 kb, which encodes a large precursor polyprotein comprising approximately 3,000 amino acids (26). The structural proteins are cleaved from the N-terminal one-fourth of the polyprotein by the host signal peptidase and signal peptide peptidase (23, 32, 33), resulting in the maturation of the capsid protein, two envelope proteins and viroporin p7. The NS2 protease cleaves after the carboxyl terminus, and then NS3 cleaves the appro- priate downstream positions to produce NS4A, NS4B, NS5A, and NS5B (8, 42), all of which form the replication complex along with several host proteins (5, 21). NS5B is the RNA- dependent RNA polymerase, which is a main enzymatic com- ponent of the replication complex of HCV (3), while NS5A is a membrane-anchored zinc-binding phosphoprotein that ap- pears to possess diverse functions, including the suppression of host defense and the regulation of the virus’s replication (1, 4, 6, 41), although its biological function remains unclear.

Using a cell-based replicon screen, we identified a class of compounds with a thiazolidinone core structure as inhibitors of hepatitis C virus (HCV) replication. The concentration of one such compound, BMS-824, that resulted in a 50% inhibition of HCV replicon replication was ⬃ 5 nM, with a therapeutic index of >10,000. The compound showed good specificity for HCV, as it was not active against several other RNA and DNA viruses. Replicon cells resistant to BMS-824 were isolated, and mutations were identified. A combination of amino acid substitutions of leucine to valine at residue 31 (L31V) and glutamine to leucine at residue 54 (Q54L) in NS5A conferred resistance to this chemotype, as did a single substitution of tyrosine to histidine at amino acid 93 (Y93H) in NS5A. To further explore the region(s) of NS5A involved in inhibitor sensitivity, genotype-specific NS5A inhibitors were used to evaluate a series of genotype 1a/1b hybrid replicons. Our results showed that, consistent with resistance mapping, the inhibitor sensitivity domain also mapped to the N terminus of NS5A, but it could be distinguished from the key resistance sites. In addition, we demonstrated that NS5A inhibitors, as well as an active-site inhibitor that specifically binds NS3 protease, could block the hyperphosphorylation of NS5A, which is believed to play an essential role in the viral life cycle. Clinical proof of concept has recently been achieved with derivatives of these NS5A inhibitors, indicating that small molecules targeting a nontra- ditional viral protein like NS5A, without any known enzymatic activity, can also have profound antiviral effects on HCV-infected subjects.

The chimpanzee is the only animal model for investigating the pathogenesis of viral hepatitis types A through E in humans. Studies of the host response, including microarray analyses, have relied on the close relationship between these two primate species: chimpanzee samples are commonly tested with human-based reagents. In this study, the host responses to two dissimilar viruses, hepatitis E virus (HEV) and hepatitis C virus (HCV), were compared in multiple experimentally infected chimpanzees. Affymetrix U133 ⴙ 2.0 human microarray chips were used to assess the entire transcriptome in serial liver biopsies obtained over the course of the infections. Respecting the limitations of microarray probes designed for human target transcripts to effectively assay chimpanzee transcripts, we conducted probe-level analysis of the microarray data in conjunc- tion with a custom mapping of the probe sequences to the most recent human and chimpanzee genome sequences. Time points for statistical comparison were chosen based on independently measured viremia levels. Regardless of the viral infection, the alignment of differentially expressed genes to the human genome sequence resulted in a larger number of genes being identified when compared with alignment to the chim- panzee genome sequence. This probably reflects the lesser refinement of gene annotation for chimpanzees. In general, the two viruses demonstrated very distinct temporal changes in host response genes, although both RNA viruses induced genes that were involved in many of the same biological systems, including interferon- induced genes. The host response to HCV infection was more robust in the magnitude and number of differentially expressed genes compared to HEV infection.

T he Flaviviridae family is composed of four genera: Flavivirus, Pestivirus, Pegivirus, and Hepacivirus. Flaviviridae family vi- ruses are enveloped and contain a single-stranded, positive-sense RNA genome, which encodes a single large precursor polyprotein composed of approximately 2,800 to 3,000 amino acids. The ge- nus Hepacivirus had included only two species, hepatitis C virus (HCV) and GB virus B (GBV-B), until 2010. GBV-B was isolated from serum samples obtained from laboratory tamarins by 11 passages of serum obtained from a human patient with idiopathic hepatitis (1). Although GBV-B experimentally infects tamarins and common marmosets, but not chimpanzees, in vivo (2, 3), the natural host of GBV-B has not yet been clarified. Several hepaci- virus species were recently detected in dogs, horses, bats, and ro- dents and tentatively designated nonprimate hepaciviruses (NPHVs). Bat hepaciviruses have been isolated from some species of bats in Kenya (4), while rodent hepaciviruses have been isolated from several species of rodents in Germany, the Netherlands, South Africa, and Namibia (5, 6). GBV-B is phylogenetically more

Hepatitis C virus (HCV) is a hepatotropic, positive-stranded RNA virus belonging to the family Flaviviridae; it is a major cause for chronic liver disease with an estimated 170 million people infected (19). Although both Huh7 and HepG2 cells are derived from human hepatocytes, HCV RNA constructs can only replicate in Huh7 cells. To explore whether this could be related to the presence of miR-122 in permissive Huh7 cells, we inspected the 9600-nt, positive-strand, viral RNA genome for potential miR-122 binding sites that fulfill the rules for a successful miRNA-target mRNA interaction. We searched for sequences in the viral mRNA that could engage in Watson and Crick base pairing with nucleotides 2 through 8, the B seed sequence [ of miR-122 (6, 20), and we noted two predicted binding sites for miR- 122 (Fig. 1B) in the viral NCRs. One is located

The role of viral factors in the pathogenesis of chronic hepatitis C is unknown. The objective of the present study was to characterize markers of hepatitis C virus (HCV) infection and replication in liver biopsy specimens obtained from 65 genotype 1-infected subjects, including 31 who were coinfected with human immunodeficiency virus (HIV), and to analyze associations between intrahepatic viral markers and hepatitis C disease severity. The percentages of liver cells harboring HCV genomes (%G) and replicative-intermediate RNAs (%RI) were evaluated using strand-specific in situ hybridization, while HCV core and NS3 antigens were assessed by immunocytochemistry. HIV-positive and HIV-negative subjects had similar mean grades and stages of liver disease and had similar indices of HCV infection and replication in liver, even though coinfected subjects had significantly shorter mean disease duration (P ⴝ 0.0003). Multivariate analysis showed that %G was not associated with grade or stage of liver disease (P ⴝ 0.5 and 0.4, respectively), while %RI was strongly associated with liver inflammation (P < 0.001), liver fibrosis (P < 0.001), and serum alanine aminotransferase levels (P ⴝ 0.01). NS3 antigen (but not core) was more frequently detected in HCV RI-positive versus RI-negative specimens (P ⴝ 0.028). These findings demonstrate a link between HCV proliferation and hepatitis C disease severity and suggest similar pathogenic mechanisms in HIV-positive and HIV-negative individuals.

Abstract: Hepatitis C virus (HCV) infection is associated with a number of extrahepatic disorders. The most studied conditions associated with HCV are type II mixed cryoglobulinemia and B cell lymphoma. However, many reports suggest that HCV might also be associated with a number of autoimmune disorders, both organ-specific and not organ-specific. Although concomitant treatment of HCV infection is a confounding factor when ascertaining the actual role of HCV in inducing autoimmune disease, a considerable amount of experimental data indicates that HCV is able to subvert the immune system and consequently induce autoimmunity. In the present review, we report a series of observations which associate chronic HCV infection with the onset of autoimmune disorders.

110. Instructions regarding these clinical criteria indicate that the criteria implement new requirements pursuant to the New York Drug Utilization Review Board meeting on Sept. 18. At that meeting, proposed criteria also included the following restrictions related to continuation of treatment, although these do not appear in the actual criteria or PA form: “in order to continue therapy once initiated, the patient must not exhibit any signs of high-risk behavior (recurring alcoholism, IV drug use, etc.) or failure to complete HCV disease evaluation appointments and procedures should be evident in follow-up reviews.” It is not clear from these criteria what was meant by “readiness and ability to adhere to drug regimen,” or how it might be documented, nor is it clear what was meant by “substance abuse potential,” or how it might or might not affect eligibility for treatment. Similarly, the standards for demonstrating signs of “high-risk behavior” and how that might impact continuation of therapy were also unclear. It is not known to what degree these proposed criteria are actually being used to evaluate continuation, as they are not actually reflected in these new guidelines. See Hepatitis C Virus Clinical Criteria Update, September 18, 2014, 19-23, New York State Department of Health (Sept. 2014), available at http://nychepbc.org/wp-content/uploads/sites/50/2014/09/HCV-DAA-Clinical-Criteria-2014_17_09_ Final1.pdf?200c4a. However, the criteria indicate that “Other recommendations made by the DURB at the September meeting will be implemented at a future date.” New York State Medicaid Update, “Medicaid Pharmacy Prior Authorization Update,” New York State Department of Health (Oct. 2014) available at http://www.health.ny.gov/health_care/medicaid/program/ update/2014/2014-10.htm.

Hepatitis C virus (HCV) infection is a major cause of chronic hepatitis and cirrhosis and may lead to hepatocellular carcinoma. With an estimated 170 million people worldwide chronically infected with HCV, this disease has emerged as a serious global health problem since the cloning of the viral genome in 1989 (6). HCV is an enveloped RNA virus belong- ing to the genus Hepacivirus of the Flaviviridae family. Its genome is a 9.6-kb single-stranded RNA of positive polarity with a 5⬘ untranslated region (UTR) that functions as an in- ternal ribosome entry site, a single long open reading frame encoding a polyprotein of approximately 3,000 amino acids (aa) and a 3⬘ UTR (1). This polypeptide is posttranslationally cleaved by host cell peptidases to yield three structural proteins and by viral proteases, which generate the five nonstructural proteins. The structural proteins, which are located in the amino-terminal region of the polyprotein, include the core protein and the envelope glycoproteins E1 and E2. The non- structural proteins (NS) 2 to 5B are separated from the struc- tural proteins by the short hydrophobic polypeptide p7, the function of which is unknown (1). By analogy to related posi- tive-strand RNA viruses, replication occurs by means of a negative-strand RNA intermediate and is catalyzed by the NS proteins, which form a cytoplasmic membrane-associated rep- licase complex.

Two direct-acting antivirals (DAAs) against hepatitis C virus (HCV): telaprevir and boceprevir, are now available in combination with peginterferon plus ribavirin for the treatment of chronic hepatitis C infection. Although these drugs are potent inhibitors of HCV replication, they occasionally result in severe adverse events. In the present clinical trials, in their stead, several second-generation DAAs are being investigated. Most of them are being viewed with high expectations, but they also require the combination with peginterferon plus ribavirin. In the near future, we might be using all-oral DAAs and interferon-free regimens for the treatment of HCV-infected patients, and these would be potent inhibitors of HCV and have less adverse events.

Results: There was no significant difference for risk factors for hepatitis C virus infection in both the groups except for the increase in number of surgical procedures being carried out in the control group. There was no significant difference in the presence of rheumatoid factor antibody in both the groups and cryoglobulins were not positive in any individual. Five percent patients with lymphoproliferative disorders and 3.4% with non- hematological malignancies were positive for anti HCV. HCV RNA was detected in 29.2% cases and 31.0% in controls.

The blood-borne pathogen, hepatitis C virus (HCV), chronically infects approximately 62 – 79 million persons worldwide [1, 2]. HCV infection is one of the causes of several morbidities including fibrosis, cirrhosis, and liver cancer, placing a strain on healthcare systems [2 – 5]. The recently available and highly efficacious direct-acting anti- virals (DAA) can treat the infection and reduce its disease burden [6]. As such, global targets for elimination of HCV infection as a global health concern by 2030 have been set by the World Health Organization (WHO) [7, 8].

Fig. 1 Pathogenesis of extrahepatic manifestations in chronic hepati- tis C infection (according to Ferri [8]) outlining the complex patho- genetic process of extrahepatic manifestations. Depending to the host (e.g. genetic factors) and unknown environmental factors, hepatitis C virus infection leads through several mechanisms (molecular mimic- ry, bystander activation and HCV lymphotropism) to the formation of autoreactive T cells and polyclonal B-cell proliferation. Autoreactive T cells and autoantibody formation cause cryoglobulinaemic vasculi- tis, organ-specific autoimmune diseases and systemic autoimmune diseases (SAD). In addition, continuous B-cell stimulation generates the development or rather selection of a B-cell clone (e.g. the t(14;18) translocation responsible for prolonged B-cell survival). Further genetic alterations (e.g. myc oncogenes) are responsible for the devel- opment of lymphomas