Humoral and CD4+ T helper (Th) cell responses to the hepatitis C virus non-structural 3 (NS3) protein: NS3 primes Th1-like responses more effectively as a DNA-based immunogen than as a recombinant protein

The non-structural 3 (NS3) protein is one of the most conserved proteins of hepatitis C virus, and T helper 1 (Th1)-like responses to NS3 in humans correlate with clearance of infection. Several studies have proposed that DNA-based immunizations are highly immunogenic and prime Th1-like responses, although few head-to-head comparisons with exogenous protein immunizations have been described. A full-length NS3/NS4A gene was cloned in eukaryotic vectors with expression directed to different subcellular compartments. Inbred mice were immunized twice in regenerating tibialis anterior (TA) muscles with either plasmid DNA or recombinant NS3 (rNS3). After two 100 μg DNA immunizations, specific antibody titres of up to 12960 were detected at week 5, dominated by IgG2a and IgG2b. NS3-specific CD4+ T cell responses in DNA-immunized mice peaked at day 13, as measured by proliferation and IL-2 and IFN-γ production. Mice immunized with 1–10 μg rNS3 without adjuvant developed antibody titres comparable to those of the DNA-immunized mice, but dominated instead by IgG1. CD4+ T cell responses in these mice showed peaks of IL-2 response at day 3 and IL-6 and IFN-γ responses at day 6. With adjuvant, rNS3 was around 10-fold more immunogenic with respect to speed and magnitude of the immune responses. Thus, immunization with rNS3 in adjuvant is superior to DNA immunization with respect to kinetics and quantity in priming specific antibodies and CD4+ T cells. However, as a DNA immunogen, NS3 elicits stronger Th1-like immune responses, whereas rNS3 primes a mixed Th1/Th2-like response regardless of the route, dose or adjuvant.

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The Clearance of Hepatitis C Virus Infection in Chimpanzees May Not Necessarily Correlate with the Appearance of Acquired Immunity

Journal of Virology ◽

10.1128/jvi.77.2.862-870.2003 ◽

2003 ◽

Vol 77 (2) ◽

pp. 862-870 ◽

Cited By ~ 63

Author(s):

Michael Thomson ◽

Michelina Nascimbeni ◽

Michael B. Havert ◽

Marian Major ◽

Sophia Gonzales ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

T Cell Responses ◽

Distinct Pattern ◽

Hcv Infection ◽

Interleukin 4 ◽

Mrna Levels ◽

Cell Responses ◽

Ifn Γ

ABSTRACT Clearance of hepatitis C virus (HCV) infection in humans and chimpanzees is thought to be associated with the induction of strong T-cell responses. We studied four chimpanzees infected with HCV derived from an infectious full-length HCV genotype 1b cDNA. Two of the chimpanzees cleared the infection to undetectable levels for more than 12 months of follow-up; the other two became persistently infected. Detailed analyses of HCV-specific immune responses were performed during the courses of infection in these chimpanzees. Only weak and transient T helper responses were detected during the acute phase in all four chimpanzees. A comparison of the frequency of gamma interferon (IFN-γ)-producing CD4+ and CD8+ T cells in peripheral blood by ELISpot assay did not reveal any correlation between viral clearance and T-cell responses. In addition, analyses of IFN-γ, IFN-α, and interleukin-4 mRNA levels in liver biopsies, presumably indicative of intrahepatic T-cell responses, revealed no distinct pattern in these chimpanzees with respect to infection outcome. The present study suggests that the outcome of HCV infection in chimpanzees is not necessarily attributable to HCV sequence variation and that chimpanzees may recover from HCV infection by mechanisms other than the induction of readily detectable HCV-specific T-cell responses.

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Corrigendum to “Hepatitis C Virus NS3/NS4A DNA Vaccine Induces Multiepitope T Cell Responses in Rhesus Macaques Mimicking Human Immune Responses”

Molecular Therapy ◽

10.1038/mt.2011.238 ◽

2012 ◽

Vol 20 (5) ◽

pp. 1075

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

Immune Responses ◽

Dna Vaccine ◽

Rhesus Macaques ◽

T Cell Responses ◽

Cell Responses ◽

Human Immune

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Hepatitis C Virus NS3/NS4A DNA Vaccine Induces Multiepitope T Cell Responses in Rhesus Macaques Mimicking Human Immune Responses

Molecular Therapy ◽

10.1038/mt.2011.188 ◽

2012 ◽

Vol 20 (3) ◽

pp. 669-678 ◽

Cited By ~ 25

Author(s):

Krystle A Lang Kuhs ◽

Arielle A Ginsberg ◽

Jian Yan ◽

Roger W Wiseman ◽

Amir S Khan ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

Immune Responses ◽

Dna Vaccine ◽

Rhesus Macaques ◽

T Cell Responses ◽

Cell Responses ◽

Human Immune

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Transmitted/Founder Viruses Rapidly Escape from CD8+T Cell Responses in Acute Hepatitis C Virus Infection

Journal of Virology ◽

10.1128/jvi.03717-14 ◽

2015 ◽

Vol 89 (10) ◽

pp. 5478-5490 ◽

Cited By ~ 23

Author(s):

Rowena A. Bull ◽

Preston Leung ◽

Silvana Gaudieri ◽

Pooja Deshpande ◽

Barbara Cameron ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

Immune Responses ◽

Primary Infection ◽

T Cell Responses ◽

Viral Population ◽

Cell Responses ◽

Cellular Immune Responses ◽

Cellular Immune

ABSTRACTThe interaction between hepatitis C virus (HCV) and cellular immune responses during very early infection is critical for disease outcome. To date, the impact of antigen-specific cellular immune responses on the evolution of the viral population establishing infection and on potential escape has not been studied. Understanding these early host-virus dynamics is important for the development of a preventative vaccine. Three subjects who were followed longitudinally from the detection of viremia preseroconversion until disease outcome were analyzed. The evolution of transmitted/founder (T/F) viruses was undertaken using deep sequencing. CD8+T cell responses were measured via enzyme-linked immunosorbent spot (ELISpot) assay using HLA class I-restricted T/F epitopes. T/F viruses were rapidly extinguished in all subjects associated with either viral clearance (n= 1) or replacement with viral variants leading to establishment of chronic infection (n= 2). CD8+T cell responses against 11 T/F epitopes were detectable by 33 to 44 days postinfection, and 5 of these epitopes had not previously been reported. These responses declined rapidly in those who became chronically infected and were maintained in the subject who cleared infection. Higher-magnitude CD8+T cell responses were associated with rapid development of immune escape variants at a rate of up to 0.1 per day. Rapid escape from CD8+T cell responses has been quantified for the first time in the early phase of primary HCV infection. These rapid escape dynamics were associated with higher-magnitude CD8+T cell responses. These findings raise questions regarding optimal selection of immunogens for HCV vaccine development and suggest that detailed analysis of individual epitopes may be required.IMPORTANCEA major limitation in our detailed understanding of the role of immune response in HCV clearance has been the lack of data on very early primary infection when the transmitted viral variants successfully establish the acute infection. This study was made possible through the availability of specimens from a unique cohort of asymptomatic primary infection cases in whom the first available viremic samples were collected approximately 3 weeks postinfection and at regular intervals thereafter. The study included detailed examination of both the evolution of the viral population and the host cellular immune responses against the T/F viruses. The findings here provide the first evidence of host cellular responses targeting T/F variants and imposing a strong selective force toward viral escape. The results of this study provide useful insight on how virus escapes the host response and consequently on future analysis of vaccine-induced immunity.

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Optimal Induction of T-Cell Responses against Hepatitis C Virus E2 by Antigen Engineering in DNA Immunization

Journal of Virology ◽

10.1128/jvi.77.21.11596-11602.2003 ◽

2003 ◽

Vol 77 (21) ◽

pp. 11596-11602 ◽

Cited By ~ 17

Author(s):

Jin-Won Youn ◽

Su-Hyung Park ◽

Jae Ho Cho ◽

Young Chul Sung

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

Immune Responses ◽

Transmembrane Domain ◽

T Cell Responses ◽

Dna Immunization ◽

Cell Responses ◽

Immunodeficiency Virus ◽

Cellular Immune

ABSTRACT Although DNA immunization is a safe and efficient method for inducing cellular immune responses, it generates relatively weak and slow immune responses. Here, we investigated the effect of hepatitis C virus (HCV) antigen modifications on the induction of T-cell responses in DNA immunization. It is likely that the strength of T-cell responses has an inverse relationship with the length of the insert DNA. Interestingly, a mixture of several plasmids carrying each gene induced a higher level of T-cell responses than a single plasmid expressing a long polyprotein. Moreover, the presence of a transmembrane domain in HCV E2 resulted in stronger T-cell responses against E2 protein than its absence. Taken together, our results indicate that the tailored modifications of DNA-encoded antigens are capable of optimizing the induction of T-cell responses which is required for eliminating the cells chronically infected with highly variable viruses such as HCV and human immunodeficiency virus.

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Hepatitis C Virus Structural Proteins Impair Dendritic Cell Maturation and Inhibit In Vivo Induction of Cellular Immune Responses

Journal of Virology ◽

10.1128/jvi.77.20.10862-10871.2003 ◽

2003 ◽

Vol 77 (20) ◽

pp. 10862-10871 ◽

Cited By ~ 98

Author(s):

Pablo Sarobe ◽

Juan José Lasarte ◽

Aintzane Zabaleta ◽

Laura Arribillaga ◽

Ainhoa Arina ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

Immune Responses ◽

T Cell Responses ◽

Cell Responses ◽

Cellular Immune Responses ◽

Cell Immunity ◽

Cellular Immune

ABSTRACT Hepatitis C virus (HCV) chronic infection is characterized by low or undetectable cellular immune responses against HCV antigens. Some studies have suggested that HCV proteins manipulate the immune system by suppressing the specific antiviral T-cell immunity. We have previously reported that the expression of HCV core and E1 proteins (CE1) in dendritic cells (DC) impairs their ability to prime T cells in vitro. We show here that immunization of mice with immature DC transduced with an adenovirus encoding HCV core and E1 antigens (AdCE1) induced lower CD4+- and CD8+-T-cell responses than immunization with DC transduced with an adenovirus encoding NS3 (AdNS3). However, no differences in the strength of the immune response were detected when animals were immunized with mature DC subsequently transduced with AdCE1 or AdNS3. According to these findings, we observed that the expression of CE1 in DC inhibited the maturation caused by tumor necrosis factor alpha or CD40L but not that induced by lipopolysaccharide. Blockade of DC maturation by CE1 was manifested by a lower expression of maturation surface markers and was associated with a reduced ability of AdCE1-transduced DC to activate CD4+- and CD8+-T-cell responses in vivo. Our results suggest that HCV CE1 proteins modulate T-cell responses by decreasing the stimulatory ability of DC in vivo via inhibition of their physiological maturation pathways. These findings are relevant for the design of therapeutic vaccination strategies in HCV-infected patients.

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Induction of Broad CD4+ and CD8+ T-Cell Responses and Cross- Neutralizing Antibodies against Hepatitis C Virus by Vaccination with Th1-Adjuvanted Polypeptides Followed by Defective Alphaviral Particles Expressing Envelope Glycoproteins gpE1 and gpE2 and Nonstructural Proteins 3, 4, and 5

Journal of Virology ◽

10.1128/jvi.02743-07 ◽

2008 ◽

Vol 82 (15) ◽

pp. 7492-7503 ◽

Cited By ~ 44

Author(s):

Yinling Lin ◽

Taewoo Kwon ◽

John Polo ◽

Yi-Fei Zhu ◽

Stephen Coates ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

T Cell ◽

Neutralizing Antibodies ◽

Antiviral Treatment ◽

Acute Infection ◽

T Cell Responses ◽

T Helper ◽

Nonstructural Proteins ◽

Cell Responses

ABSTRACT Broad, multispecific CD4+ and CD8+ T-cell responses to the hepatitis C virus (HCV), as well as virus-cross-neutralizing antibodies, are associated with recovery from acute infection and may also be associated in chronic HCV patients with a favorable response to antiviral treatment. In order to recapitulate all of these responses in an ideal vaccine regimen, we have explored the use of recombinant HCV polypeptides combined with various Th1-type adjuvants and replication-defective alphaviral particles encoding HCV proteins in various prime/boost modalities in BALB/c mice. Defective chimeric alphaviral particles derived from the Sindbis and Venezuelan equine encephalitis viruses encoding either the HCV envelope glycoprotein gpE1/gpE2 heterodimer (E1E2) or nonstructural proteins 3, 4, and 5 (NS345) elicited strong CD8+ T-cell responses but low CD4+ T helper responses to these HCV gene products. In contrast, recombinant E1E2 glycoproteins adjuvanted with MF59 containing a CpG oligonucleotide elicited strong CD4+ T helper responses but no CD8+ T-cell responses. A recombinant NS345 polyprotein also stimulated strong CD4+ T helper responses but no CD8+ T-cell responses when adjuvanted with Iscomatrix containing CpG. Optimal elicitation of broad CD4+ and CD8+ T-cell responses to E1E2 and NS345 was obtained by first priming with Th1-adjuvanted proteins and then boosting with chimeric, defective alphaviruses expressing these HCV genes. In addition, this prime/boost regimen resulted in the induction of anti-E1E2 antibodies capable of cross-neutralizing heterologous HCV isolates in vitro. This vaccine formulation and regimen may therefore be optimal in humans for protection against this highly heterogeneous global pathogen.

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