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

Hepatitis C virus (HCV) is the leading causative agent of blood-borne chron ic hepatitis and is the target of intensive vaccine research. The virus gen ome encodes a number of structural and nonstructural antigens which could b e 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 f or inclusion in any such vaccine. The experiments presented here assessed t he optimal form of HCV E2 antigen from the perspective of antibody generati on. The quality of recombinant E2 protein was evaluated by both the capacit y to bind its putative receptor CD81 on human cells and the ability to elic it antibodies that inhibited this binding (NOB antibodies). We show that tr uncated 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-glyc osylated secreted fraction does not bind and elicits no NOB antibodies. We also show that carbohydrate moieties are not necessary for E2 binding to hu man cells and that only the monomeric nonaggregated fraction ran bind to CD 81. Moreover, comparing recombinant intracellular E2 protein to several E2- encoding DNA vaccines in mice, we found that protein immunization is superi or to DNA in both the quantity and quality of the antibody response elicite d. Together, our data suggest that to elicit antibodies aimed at blocking E F(TV binding to CD81 on human cells, the antigen of choice is a mammalian c ell-expressed, monomeric E2 protein purified from the intracellular fractio n.