Distinct hepatitis C virus core and F protein quasispecies in tumoral and nontumoral hepatocytes isolated via microdissection

Sobesky, Rodolphe; Féray, Cyrille; Rimlinger, F.; Dérian, Nicolas; Santos, Alexandre Dos; Roque–Afonso, Anne–Marie; Bréchot, Christian; Thiers, Valérie

doi:10.1002/hep.21898

Cited by 48 publications

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“…Our data also show that the differences in quasispecies distribution are not the result of differences in host selective pressure in malignant hepatocytes, as was suggested previously (20). The molecular mechanisms leading to high viral diversity within the tumor despite the significant drop in viral replication remain to be elucidated.…”

Section: Discussionsupporting

confidence: 84%

“…Consistent with this hypothesis, Mantel test analysis demonstrated significant differences in HCV quasispecies distribution between the tumorous and nontumorous compartments, providing evidence of HCV compartmentalization between these two liver areas. In contrast, analysis of multiple liver areas in the control patients with non-HCC cirrhosis demonstrated no evidence of HCV compartmentalization within the liver or between the liver and serum, further indicating that the presence of HCV compartmentalization is unique to tumor-containing livers (20,41). Interestingly, viral compartmentalization between plasma and extrahepatic sites, including leukocytes (42), PBMCs (43), and the brain (44), also has been reported in HCV-infected patients.…”

Section: Discussionmentioning

confidence: 95%

“…This remains controversial, however, in part because of the difficulty in obtaining multiple paired liver specimens from the tumor and adjacent nontumorous tissue. Although some previous investigations have shown no differences in the presence and levels of HCV RNA between the tumor and nontumorous liver tissues (15)(16)(17), others have reported low to undetectable levels of HCV RNA within the tumor (18)(19)(20). Several reports have indicated that miR-122 is an essential cellular host factor for HCV replication (21,22).…”

mentioning

confidence: 98%

“…however, very limited information is available on the composition of the HCV quasispecies and the possible compartmentalization between the tumor and the surrounding nontumorous areas or serum from the same individuals with HCV-associated HCC (20,(28)(29)(30).…”

mentioning

confidence: 99%

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Diminished viral replication and compartmentalization of hepatitis C virus in hepatocellular carcinoma tissue

Harouaka

Engle

Wollenberg

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Analysis of hepatitis C virus (HCV) replication and quasispecies distribution within the tumor of patients with HCV-associated hepatocellular carcinoma (HCC) can provide insight into the role of HCV in hepatocarcinogenesis and, conversely, the effect of HCC on the HCV lifecycle. In a comprehensive study of serum and multiple liver specimens from patients with HCC who underwent liver transplantation, we found a sharp and significant decrease in HCV RNA in the tumor compared with surrounding nontumorous tissues, but found no differences in multiple areas of control non-HCC cirrhotic livers. Diminished HCV replication was not associated with changes in miR-122 expression. HCV genetic diversity was significantly higher in livers containing HCC compared with control non-HCC cirrhotic livers. Tracking of individual variants demonstrated changes in the viral population between tumorous and nontumorous areas, the extent of which correlated with the decline in HCV RNA, suggesting HCV compartmentalization within the tumor. In contrast, compartmentalization was not observed between nontumorous areas and serum, or in controls between different areas of the cirrhotic liver or between liver and serum. Our findings indicate that HCV replication within the tumor is restricted and compartmentalized, suggesting segregation of specific viral variants in malignant hepatocytes.hepatitis C virus | hepatocellular carcinoma | HCV RNA levels | HCV quasispecies | liver H epatitis C virus (HCV) is a hepatotropic, single-stranded RNA virus that replicates in the cytoplasm of hepatocytes and does not integrate into the host genome (1). HCV circulates in vivo as a dynamic distribution of closely related viral variants that are commonly referred to as "quasispecies" (2). Such diversity confers a remarkable advantage to the virus under host selective constraints (3-5). One of the most important features of HCV is its extraordinary ability to persist in up to 80% of infected individuals (6). Approximately 20-30% of chronic HCV carriers develop cirrhosis and its long-term sequelae, including hepatic decompensation and hepatocellular carcinoma (HCC), leading to orthotopic liver transplantation (OLT) or liver-related death (6). The incidence of HCC in the United States has more than tripled over the past 3 decades (7), an alarming trend due primarily, if not exclusively, to HCV infection (8).Although epidemiologic evidence has linked chronic HCV infection to a significantly elevated risk of developing HCC (9), the mechanisms whereby HCV promotes hepatocarcinogenesis remain to be elucidated (10). Whether HCV elicits liver cancer indirectly, through chronic inflammation, fibrosis, and continuous liver regeneration (11), or directly, through the expression of tumor-promoting viral proteins, in a manner analogous to other oncogenic viruses, such as human papillomaviruses and EpsteinBarr virus (12, 13), remains unknown. The major challenges in defining the role of HCV in the pathogenesis of HCC include the inherent limitations of available experimental...

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Section: Discussionsupporting

confidence: 84%

Section: Discussionmentioning

confidence: 95%

mentioning

confidence: 98%

mentioning

confidence: 99%

See 2 more Smart Citations

Diminished viral replication and compartmentalization of hepatitis C virus in hepatocellular carcinoma tissue

Harouaka

Engle

Wollenberg

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Studies have shown that HCV compartmentalizes within the host, with quasispecies compositions differing between different organs (10,11). The primary target of HCV is the liver, where it forms distinct foci of infection (12), but it also infects extrahepatic sites, including the brain (10,11).…”

mentioning

confidence: 99%

Determining the Cellular Diversity of Hepatitis C Virus Quasispecies by Single-Cell Viral Sequencing

Leitch

McLauchlan

2013

J Virol

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Single-cell genomics is emerging as an important tool in cellular biology. We describe for the first time a system to investigate RNA virus quasispecies diversity at the cellular level utilizing hepatitis C virus (HCV) replicons. A high-fidelity nested reverse transcription (RT)-PCR assay was developed, and validation using control transcripts of known copy number indicated a detection limit of 3 copies of viral RNA/reaction. This system was used to determine the cellular diversity of subgenomic JFH-1 HCV replicons constitutively expressed in Huh7 cells. Each cell contained a unique quasispecies that was much less diverse than the quasispecies of the bulk cell population from which the single cells were derived, suggesting the occurrence of independent evolution at the cellular level. An assessment of the replicative fitness of the predominant single-cell quasispecies variants indicated a modest reduction in fitness compared to the wild type. Real-time RT-PCR methods capable of determining single-cell viral loads were developed and indicated an average of 113 copies of replicon RNA per cell, correlating with calculated RNA copy numbers in the bulk cell population. This study introduces a single-cell RNA viral-sequencing method with numerous potential applications to explore host-virus interactions during infection. HCV quasispecies diversity varied greatly between cells in vitro, suggesting different within-cell evolutionary pathways. Such divergent trajectories in vivo could have implications for the evolution and establishment of antiviral-resistant variants and host immune escape mutants.

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Next‐generation sequencing sheds light on the natural history of hepatitis C infection in patients who fail treatment

Abdelrahman

Hughes

Main³

et al. 2014

Hepatology

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High rates of sexually transmitted infection and reinfection with hepatitis C virus (HCV) have recently been reported in human immunodeficiency virus (HIV)-infected men who have sex with men and reinfection has also been described in monoinfected injecting drug users. The diagnosis of reinfection has traditionally been based on direct Sanger sequencing of samples pre- and posttreatment, but not on more sensitive deep sequencing techniques. We studied viral quasispecies dynamics in patients who failed standard of care therapy in a high-risk HIV-infected cohort of patients with early HCV infection to determine whether treatment failure was associated with reinfection or recrudescence of preexisting infection. Paired sequences (pre- and posttreatment) were analyzed. The HCV E2 hypervariable region-1 was amplified using nested reverse-transcription polymerase chain reaction (RT-PCR) with indexed genotype-specific primers and the same products were sequenced using both Sanger and 454 pyrosequencing approaches. Of 99 HIV-infected patients with acute HCV treated with 24-48 weeks of pegylated interferon alpha and ribavirin, 15 failed to achieve a sustained virological response (six relapsed, six had a null response, and three had a partial response). Using direct sequencing, 10/15 patients (66%) had evidence of a previously undetected strain posttreatment; in many studies, this is interpreted as reinfection. However, pyrosequencing revealed that 15/15 (100%) of patients had evidence of persisting infection; 6/15 (40%) patients had evidence of a previously undetected variant present in the posttreatment sample in addition to a variant that was detected at baseline. This could represent superinfection or a limitation of the sensitivity of pyrosequencing. Conclusion: In this high-risk group, the emergence of new viral strains following treatment failure is most commonly associated with emerging dominance of preexisting minority variants rather than reinfection. Superinfection may occur in this cohort but reinfection is overestimated by Sanger sequencing. (Hepatology 2015;61:88–97)

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Distinct hepatitis C virus core and F protein quasispecies in tumoral and nontumoral hepatocytes isolated via microdissection

Cited by 48 publications

References 49 publications

Diminished viral replication and compartmentalization of hepatitis C virus in hepatocellular carcinoma tissue

Diminished viral replication and compartmentalization of hepatitis C virus in hepatocellular carcinoma tissue

Determining the Cellular Diversity of Hepatitis C Virus Quasispecies by Single-Cell Viral Sequencing

Next‐generation sequencing sheds light on the natural history of hepatitis C infection in patients who fail treatment

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