Abstract
Hepatitis C virus (HCV) is a member of Ηepacivirus genus within the Flaviviridae family. It infects the human liver and is a major cause of acute and chronic hepatitis, which in turn leads to severe liver diseases, such as chronic active hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). An estimated 170 million people (3% of world population) is infected by HCV. Comparison of nucleotide sequence data has led to the classification of HCV into six major types (genotypes) that differ from each other by 30% over the entire genome and several closely related subtypes that differ from each other by around 20%. Genotypes 1, 2 and 3 are spread worldwide. In Western Europe, HCV genotype 1 is the most prevalent genotype in individuals aged over 40 years, whereas genotype 3 is mostly common in young people, especially in intravenous drug users. In Greece, HCV genotype 1 is the most prevalent (46.9%), followed by genotype 3 (28.1%), 4 (13.2%), 2 (6.9%) and 5 (0.4%). Moreover, a sharp (13-f ...
Hepatitis C virus (HCV) is a member of Ηepacivirus genus within the Flaviviridae family. It infects the human liver and is a major cause of acute and chronic hepatitis, which in turn leads to severe liver diseases, such as chronic active hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). An estimated 170 million people (3% of world population) is infected by HCV. Comparison of nucleotide sequence data has led to the classification of HCV into six major types (genotypes) that differ from each other by 30% over the entire genome and several closely related subtypes that differ from each other by around 20%. Genotypes 1, 2 and 3 are spread worldwide. In Western Europe, HCV genotype 1 is the most prevalent genotype in individuals aged over 40 years, whereas genotype 3 is mostly common in young people, especially in intravenous drug users. In Greece, HCV genotype 1 is the most prevalent (46.9%), followed by genotype 3 (28.1%), 4 (13.2%), 2 (6.9%) and 5 (0.4%). Moreover, a sharp (13-fold) increase for genotype 3 was observed from 1970 to 1990, suggesting that genotype 3 and intravenous drug use are the major forces for HCV epidemic in the recent cohort of HCV-infected individuals in Greece.
Although there is no vaccine or broadly effective therapy, treatment of HCV infection with standard combination therapy, which includes pegylated interferon-a (Peg-IFN-a-2a) plus ribavirin, has an overall estimated success rate of 50% for genotype 1 and about 80% for genotype 2 and 3. Viral load of less than 600,000 IU/mL at the beginning of the therapy is also a major predictor of a sustained virological response (SVR). Host factors such as age, race, gender, obesity and degree of hepatic fibrosis, also affect the chance of an SVR.
HCV genome consists of a 9.6 kb single-strand, positive-sense RNA, which encodes for a single immature polyprotein of about 3,000 amino acids. This translation product is cleaved by cellular and viral proteases to yield at least four structural (C, E1, E2, p7) and six non structural (NS2, NS3, NS4A, NS4B, NS5A, NS5B) mature proteins. Recent studies by different groups of investigators, have provided strong evidence for the presence of a functional alternative open reading frame (ORF) within the core genomic region in the +1 reading frame (known as ARF or core+1 ORF), that leads to the production of ‘core overlapping’ peptides known as alternative reading frame proteins (ARFPs) or frameshift (F) or core+1 proteins.
Since its discovery, different isoforms of ARF/core+1 protein have been reported based on differences in the translation initiation mechanism that are found to stimulate expression of ARF/core+1 ORF in cultured cells. The biological significance of the ARF/core+1 expression remains elusive. However, several studies imply the presence of this protein in HCV patients via its specific recognition by serum antibodies (anti-ARF/core+1 Abs). Some of these studies suggest a correlation between the presence of anti ARF/core+1 Abs and specific stages of HCV infection, such as the latest stages of liver steatosis and hepatocellular carcinoma (HCC).
In the present study we estimated the prevalence of specific anti-ARF/core+1 Abs in HCV infected patients belonging to genotypes 1a/1b or 3a, treated with Peg-IFN-a-2a plus ribavirin and its association with patients’ age and HCV-RNA levels at baseline. The anti-ARF/core+1 Ab’s titer dynamics during therapy follow up were also investigated.
For these purposes, highly purified HCV core and ARF/core+1 recombinant proteins were expressed in Escherichia coli. Using an enzyme-linked immunosorbent assay (ELISA), we assessed the prevalence of anti-ARF/core+1 antibodies in 90 chronic hepatitis C patients. Samples derived from 92 healthy blood donors were used as negative controls. All HCV-RNA positive serum samples reacted with core 1a antigen while 15 (37.5%) out of 40 and 14 (28%) out of 50 patients infected with HCV-1a/1b and HCV-3a respectively, were found to have anti-ARF/core+1 antibodies into their serum before treatment initiation. These antibodies were persistently present during treatment follow up and linked to elevated levels of HCV RNA at baseline.
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