title: Evaluation of hepatitis C virus glycoprotein E2 for vaccine design: an endoplasmic reticulum-retained recombinant protein is superior to secreted recombinant protein and DNA-based vaccine candidates.
abstract: Hepatitis C virus (HCV) is the leading causative agent of blood-borne chronic hepatitis and is the target of intensive vaccine research. The virus genome encodes a number of structural and nonstructural antigens which could be used in a subunit vaccine. The HCV envelope glycoprotein E2 has recently been shown to bind CD81 on human cells and therefore is a prime candidate for inclusion in any such vaccine. The experiments presented here assessed the optimal form of HCV E2 antigen from the perspective of antibody generation. The quality of recombinant E2 protein was evaluated by both the capacity to bind its putative receptor CD81 on human cells and the ability to elicit antibodies that inhibited this binding (NOB antibodies). We show that truncated E2 proteins expressed in mammalian cells bind with high efficiency to human cells and elicit NOB antibodies in guinea pigs only when purified from the core-glycosylated intracellular fraction, whereas the complex-glycosylated secreted fraction does not bind and elicits no NOB antibodies. We also show that carbohydrate moieties are not necessary for E2 binding to human cells and that only the monomeric nonaggregated fraction can bind to CD81. Moreover, comparing recombinant intracellular E2 protein to several E2-encoding DNA vaccines in mice, we found that protein immunization is superior to DNA in both the quantity and quality of the antibody response elicited. Together, our data suggest that to elicit antibodies aimed at blocking HCV binding to CD81 on human cells, the antigen of choice is a mammalian cell-expressed, monomeric E2 protein purified from the intracellular fraction.
authors.affiliation: IRIS Research Center, Chiron, 53100 Siena, Italy.
authors.forename: J M
authors.initials: JM
authors.lastname: Heile
authors.forename: Y L
authors.initials: YL
authors.lastname: Fong
authors.forename: D
authors.initials: D
authors.lastname: Rosa
authors.forename: K
authors.initials: K
authors.lastname: Berger
authors.forename: G
authors.initials: G
authors.lastname: Saletti
authors.forename: S
authors.initials: S
authors.lastname: Campagnoli
authors.forename: G
authors.initials: G
authors.lastname: Bensi
authors.forename: S
authors.initials: S
authors.lastname: Capo
authors.forename: S
authors.initials: S
authors.lastname: Coates
authors.forename: K
authors.initials: K
authors.lastname: Crawford
authors.forename: C
authors.initials: C
authors.lastname: Dong
authors.forename: M
authors.initials: M
authors.lastname: Wininger
authors.forename: G
authors.initials: G
authors.lastname: Baker
authors.forename: L
authors.initials: L
authors.lastname: Cousens
authors.forename: D
authors.initials: D
authors.lastname: Chien
authors.forename: P
authors.initials: P
authors.lastname: Ng
authors.forename: P
authors.initials: P
authors.lastname: Archangel
authors.forename: G
authors.initials: G
authors.lastname: Grandi
authors.forename: M
authors.initials: M
authors.lastname: Houghton
authors.forename: S
authors.initials: S
authors.lastname: Abrignani
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D015195:Drug Design; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005069:Evaluation Studies as Topic; D005260:Female; D006031:Glycosylation; D006168:Guinea Pigs; D016174:Hepacivirus / Q000276:immunology*; D006526:Hepatitis C / Q000517:prevention & control*; D018937:Hepatitis C Antibodies / Q000097:blood; D006801:Humans; D007114:Immunization; D008565:Membrane Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D060225:Tetraspanin 28; D019444:Vaccines, DNA / Q000276:immunology*; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D014761:Viral Hepatitis Vaccines / Q000276:immunology*
chemical_list: D015703:Antigens, CD; C554798:CD81 protein, human; C554799:Cd81 protein, mouse; D018937:Hepatitis C Antibodies; D008565:Membrane Proteins; D011994:Recombinant Proteins; D060225:Tetraspanin 28; D019444:Vaccines, DNA; D014759:Viral Envelope Proteins; D014761:Viral Hepatitis Vaccines; C091015:glycoprotein E2, Hepatitis C virus