Hepatitis C virus molecular evolution: Transmission, disease progression and antiviral therapy

Hepatitis C virus (HCV) infection represents an important public health problem worldwide. Reduction of HCV morbidity and mortality is a current challenge owned to several viral and host factors. Virus molecular evolution plays an important role in HCV transmission, disease progression and therapy outcome. The high degree of genetic heterogeneity characteristic of HCV is a key element for the rapid adaptation of the intrahost viral population to different selection pressures (e.g., host immune responses and antiviral therapy). HCV molecular evolution is shaped by different mechanisms including a high mutation rate, genetic bottlenecks, genetic drift, recombination, temporal variations and compartmentalization. These evolutionary processes constantly rearrange the composition of the HCV intrahost population in a staging manner. Remarkable advances in the understanding of the molecular mechanism controlling HCV replication have facilitated the development of a plethora of direct-acting antiviral agents against HCV. As a result, superior sustained viral responses have been attained. The rapidly evolving field of anti-HCV therapy is expected to broad its landscape even further with newer, more potent antivirals, bringing us one step closer to the interferon-free era.Fil: Preciado, María Victoria. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños ; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Valva, Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños ; ArgentinaFil: Escobar Gutierrez, Alejandro. Instituto de Diagnóstico y Referencia Epidemiológicos; MéxicoFil: Rahal, Paula. Universidade Estadual Paulista Julio de Mesquita Filho; BrasilFil: Ruiz Tovar, Karina. Instituto de Diagnóstico y Referencia Epidemiológicos; MéxicoFil: Yamasaki, Lilian. Universidade Estadual Paulista Julio de Mesquita Filho; BrasilFil: Vazquez Chacon, Carlos. Instituto de Diagnóstico y Referencia Epidemiológicos; MéxicoFil: Martinez Guarneros, Armando. Instituto de Diagnóstico y Referencia Epidemiológicos; MéxicoFil: Carpio Pedroza, Juan Carlos. Instituto de Diagnóstico y Referencia Epidemiológicos; MéxicoFil: Fonseca Coronado, Salvador. Universidad Nacional Autónoma de México; MéxicoFil: Cruz Rivera, Mayra. Universidad Nacional Autónoma de México; Méxic

Replicative Homeostasis III: implications for antiviral therapy and mechanisms of response and non-response

While improved drug regimens have greatly enhanced outcomes for patients with chronic viral infection, antiviral therapy is still not ideal due to drug toxicities, treatment costs, primary drug failure and emergent resistance. New antiviral agents, alternative treatment strategies and a better understanding of viral pathobiology, host responses and drug action are desperately needed. Interferon (IFN) and ribavirin, are effective drugs used to treat hepatitis C (HCV), but the mechanism(s) of their action are uncertain. Error catastrophe (EC), or precipitous loss of replicative fitness caused by genomic mutation, is postulated to mediate ribavirin action, but is a deeply flawed hypothesis lacking empirical confirmation. Paradoxically ribavirin, a proven RNA mutagen, has no impact on HCV viraemia long term, suggesting real viruses, replicating in-vitro, as opposed to mathematical models, replicating in-silico, are likely to resist EC by highly selective replication of fit (~consensus sequence) genomes mediated, in part, by replicative homeostasis (RH), an epicyclic mechanism that dynamically links RNApol fidelity and processivity and other viral protein functions. Replicative homeostasis provides a rational explanation for the various responses seen during treatment of HCV, including genotype-specific and viral load-dependent differential response rates, as well as otherwise unexplained phenomena like the transient inhibition and rebound of HCV viraemia seen during ribavirin monotherapy. Replicative homeostasis also suggests a primarily non-immunological mechanism that mediates increased immune responsiveness during treatment with ribavirin (and other nucleos(t)ide analogues), explicating the enhanced second-phase clearance of HCV ribavirin promotes and, thus, the apparent immunomodulatory action of ribavirin. More importantly, RH suggests specific new antiviral therapeutic strategies

A Multi-Variant, Viral Dynamic Model of Genotype 1 HCV to Assess the in vivo Evolution of Protease-Inhibitor Resistant Variants

Variants resistant to compounds specifically targeting HCV are observed in clinical trials. A multi-variant viral dynamic model was developed to quantify the evolution and in vivo fitness of variants in subjects dosed with monotherapy of an HCV protease inhibitor, telaprevir. Variant fitness was estimated using a model in which variants were selected by competition for shared limited replication space. Fitness was represented in the absence of telaprevir by different variant production rate constants and in the presence of telaprevir by additional antiviral blockage by telaprevir. Model parameters, including rate constants for viral production, clearance, and effective telaprevir concentration, were estimated from 1) plasma HCV RNA levels of subjects before, during, and after dosing, 2) post-dosing prevalence of plasma variants from subjects, and 3) sensitivity of variants to telaprevir in the HCV replicon. The model provided a good fit to plasma HCV RNA levels observed both during and after telaprevir dosing, as well as to variant prevalence observed after telaprevir dosing. After an initial sharp decline in HCV RNA levels during dosing with telaprevir, HCV RNA levels increased in some subjects. The model predicted this increase to be caused by pre-existing variants with sufficient fitness to expand once available replication space increased due to rapid clearance of wild-type (WT) virus. The average replicative fitness estimates in the absence of telaprevir ranged from 1% to 68% of WT fitness. Compared to the relative fitness method, the in vivo estimates from the viral dynamic model corresponded more closely to in vitro replicon data, as well as to qualitative behaviors observed in both on-dosing and long-term post-dosing clinical data. The modeling fitness estimates were robust in sensitivity analyses in which the restoration dynamics of replication space and assumptions of HCV mutation rates were varied

Multi-agent model of hepatitis C virus infection

Objectives: The objective of this study is to design a method for modeling hepatitis C virus (HCV) infection using multi-agent simulation and to verify it in practice. Methods and materials: In this paper, first, the modeling of HCV infection using a multi-agent system is compared with the most commonly used model type, which is based on differential equations. Then, the implementation and results of the model using a multi-agent simulation is presented. To find the values of the parameters used in the model, a method using inverted simulation flow and genetic algorithm is proposed. All of the data regarding HCV infection are taken from the paper describing the model based on the differential equation to which the proposed method is compared. Results: Important advantages of the proposed method are noted and demonstrated; these include flexibility, clarity, re-usability and the possibility to model more complex dependencies. Then, the simulation framework that uses the proposed approach is successfully implemented in C++ and is verified by comparing it to the approach based on differential equations. The verification proves that an objective function that performs the best is the function that minimizes the maximal differences in the data. Finally, an analysis of one of the already known models is performed, and it is proved that it incorrectly models a decay in the hepatocytes number by 40%. Conclusions: The proposed method has many advantages in comparison to the currently used model types and can be used successfully for analyzing HCV infection. With almost no modifications, it can also be used for other types of viral infections

Superinfection and cure of infected cells as mechanisms for hepatitis C virus adaptation and persistence

Copyright © 2018 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND).RNA viruses exist as a genetically diverse quasispecies with extraordinary ability to adapt to abrupt changes in the host environment. However, the molecular mechanisms that contribute to their rapid adaptation and persistence in vivo are not well studied. Here, we probe hepatitis C virus (HCV) persistence by analyzing clinical samples taken from subjects who were treated with a second-generation HCV protease inhibitor. Frequent longitudinal viral load determinations and large-scale single-genome sequence analyses revealed rapid antiviral resistance development, and surprisingly, dynamic turnover of dominant drug-resistant mutant populations long after treatment cessation. We fitted mathematical models to both the viral load and the viral sequencing data, and the results provided strong support for the critical roles that superinfection and cure of infected cells play in facilitating the rapid turnover and persistence of viral populations. More broadly, our results highlight the importance of considering viral dynamics and competition at the intracellular level in understanding rapid viral adaptation. Thus, we propose a theoretical framework integrating viral and molecular mechanisms to explain rapid viral evolution, resistance, and persistence despite antiviral treatment and host immune responses.info:eu-repo/semantics/publishedVersio

A study of novel biomarkers of hepatitis C related liver injury

Chronic hepatitis C virus (HCV) is the most frequent indication for liver transplantation. Unfortunately HCV recurrence is universal following transplantation and many patients will experience aggressive disease recurrence. This thesis comprises 3 related studies with the main focus being an assessment of factors which may impact on the course of HCV recurrence post-transplant. The first study was a retrospective analysis of 118 consecutive HCV-positive liver transplant patients with a median duration of follow-up of 32.4 months. Peak viral RNA ≥ 107 in the first year post-transplant was shown to be an independent predictor of diminished patient survival. The second study was a cross-sectional pilot study using a recently developed CD antibody microarray in patients with various causes for liver disease. This demonstrated disease-specific consensus patterns of expression of CD antigens for patients with chronic liver disease and in particular, the ability to separate major stages of liver disease. In the third study serial CD antigen expression profiles were performed on patients undergoing liver transplantation for HCV infection. Differential antibody expression was most significant in the pre-transplant phase suggesting a significant impact of pre-transplant factors on disease severity post-liver transplantation. The findings in this thesis have demonstrated utility of the CD antibody microarray in the study of human liver disease and have shed light on the importance of post-transplant viral loads in determining the severity of post-transplant HCV recurrence

Modelling the Hepatitis C with Different Types of Virus Genome

Hepatitis C virus (HCV) is one of the leading known causes of liver disease in the world. The HCV is a single-stranded RNA virus. The genomes of HCV display significant sequence heterogeneity and have been classified into types and subtypes. Types from 1 to 11 have so far been recognized, each type having a variable number of subtypes. It has been confirmed that 90% approximately of the isolates HCV infections in Egypt belong to a single subtype (4a) [10]. In this paper, we construct a mathematical model to study the spread of HCV-subtype 4a amongst the Egyptian population. The relation between HCV-subtype 4a and the other subtypes has also been studied. The values of reproduction numbers R01, R02 have been derived [5]. Also, threshold conditions for the value of the transmission rates k1 and k02, in terms of R01, R02 and the mutation factor μ have been determined to insure that the disease will die out. If the conditions fail to happen the disease takes off and becomes endemic

Interfering with interferon: developing a reporter system to study the interaction between hepatitus C viral proteins and the interferon signalling pathway

PhDThe aim of the project was to investigate the mechanism by which HCV evades therapeutic IFN treatment. This involved the development of novel testing systems and their application to patient samples. Initial experiments focused on flavivirus replicons and novel observations on effects of one of these replicons (dengue virus) on interferon signalling were made. The dengue replicon system was demonstrated to inhibit IFNa signalling by reducing the expression of STAT2, an essential component of the type I IFN signalling pathway. This phenomenom was then further examined in dengue virus infected human cells and again it was observed that the expression of STAT2 was reduced. The mechanism of STAT2 degradation was further explored and STAT2 expression was found to be restored using a proteasomal inhibitor. A second flavivirus replicon system involving BVDV was also developed as a reporter system, again with novel observations. The BVDV replicon system was shown to be sensitive to the antiviral effects of I FNa and was not shown to inhibit the IFNa signalling pathway. The BVDV replicon was tested as a reporter system using a well-known viral inhibitor of I FNa. The viral inhibitor, inhibited the antiviral action of IFNa on the BVDV reporter. Having developed and validated this system, the effects of a small number of patient derived samples were assessed and it was demonstrated that NS5a derived from a patient who failed to respond to IFNa treatment inhibited the effects of IFNa on the BVDV reporter. To increase the senstitivity of the assay the reporter cassette was then changed to a destabilised GFP for use in a FACS based assay

The Evolutionary Dynamics of a Rapidly Mutating Virus within and between Hosts: The Case of Hepatitis C Virus

Many pathogens associated with chronic infections evolve so rapidly that strains found late in an infection have little in common with the initial strain. This raises questions at different levels of analysis because rapid within-host evolution affects the course of an infection, but it can also affect the possibility for natural selection to act at the between-host level. We present a nested approach that incorporates within-host evolutionary dynamics of a rapidly mutating virus (hepatitis C virus) targeted by a cellular cross-reactive immune response, into an epidemiological perspective. The viral trait we follow is the replication rate of the strain initiating the infection. We find that, even for rapidly evolving viruses, the replication rate of the initial strain has a strong effect on the fitness of an infection. Moreover, infections caused by slowly replicating viruses have the highest infection fitness (i.e., lead to more secondary infections), but strains with higher replication rates tend to dominate within a host in the long-term. We also study the effect of cross-reactive immunity and viral mutation rate on infection life history traits. For instance, because of the stochastic nature of our approach, we can identify factors affecting the outcome of the infection (acute or chronic infections). Finally, we show that anti-viral treatments modify the value of the optimal initial replication rate and that the timing of the treatment administration can have public health consequences due to within-host evolution. Our results support the idea that natural selection can act on the replication rate of rapidly evolving viruses at the between-host level. It also provides a mechanistic description of within-host constraints, such as cross-reactive immunity, and shows how these constraints affect the infection fitness. This model raises questions that can be tested experimentally and underlines the necessity to consider the evolution of quantitative traits to understand the outcome and the fitness of an infection

Hepatitis C virus compartmentalisation: unravelling the genetic complexity

Hepatitis C virus (HCV) is a global health problem with over 150 million individuals infected worldwide. Many of these patients will develop end-stage liver diseases, such as cirrhosis and hepatocellular carcinoma (HCC), and will require liver transplantation. HCV exists as a heterogeneous population in infected individuals, however, the processes which maintain this genetic complexity are unknown. Recent observations suggest that HCV transmits between hepatocytes via a cell-to-cell route of infection, supporting a “demic” model of evolution where HCV diversity arises from independent evolution in small isolated hepatic populations. To investigate the distribution of HCV within the liver we sampled eight segments of the liver explant from 22 HCV infected subjects undergoing liver transplant and measured viral RNA burden and sequence diversity. Comparable HCV RNA levels were observed across all 8 samples from a single liver, however, between patients we observed a 100-fold range in the hepatic viral load that was independent of hepatic expression of anti-viral and pro-viral interferon stimulated genes (ISGs). Sequence analysis of the viral envelope E1E2 region, obtained from PCR generated single molecules or ultra-deep sequencing approaches, showed minimal evidence of genetic compartmentalisation between hepatic sites or between the liver and plasma. Modelling the HCV population structure in infected patients will have a major impact on our understanding of how HCV escapes host immune responses and anti-viral therapies