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. 2000 Dec;74(23):10903-10.
doi: 10.1128/jvi.74.23.10903-10910.2000.

Glycoprotein exchange vectors based on vesicular stomatitis virus allow effective boosting and generation of neutralizing antibodies to a primary isolate of human immunodeficiency virus type 1

N F Rose  1 , A Roberts, L Buonocore, J K Rose
Affiliations

Glycoprotein exchange vectors based on vesicular stomatitis virus allow effective boosting and generation of neutralizing antibodies to a primary isolate of human immunodeficiency virus type 1

N F Rose et al. J Virol. 2000 Dec.

Abstract

Live recombinant vesicular stomatitis viruses (VSVs) expressing foreign antigens are highly effective vaccine vectors. However, these vectors induce high-titer neutralizing antibody directed at the single VSV glycoprotein (G), and this antibody alone can prevent reinfection and boosting with the same vector. To determine if efficient boosting could be achieved by changing the G protein of the vector, we have developed two new recombinant VSV vectors based on the VSV Indiana serotype but with the G protein gene replaced with G genes from two other VSV serotypes, New Jersey and Chandipura. These G protein exchange vectors grew to titers equivalent to wild-type VSV and induced similar neutralizing titers to themselves but no cross-neutralizing antibodies to the other two serotypes. The effectiveness of these recombinant VSV vectors was illustrated in experiments in which sequential boosting of mice with the three vectors, all encoding the same primary human immunodeficiency virus (HIV) envelope protein, gave a fourfold increase in antibody titer to an oligomeric HIV envelope compared with the response in animals receiving the same vector three times. In addition, only the animals boosted with the exchange vectors produced antibodies neutralizing the autologous HIV primary isolate. These VSV envelope exchange vectors have potential as vaccines in immunizations when boosting of immune responses may be essential.

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Figures

FIG. 1
FIG. 1
Diagram of VSV G protein exchange vectors. A schematic representation of the negative-strand recombinant VSV genomic RNAs of the G protein exchange vectors is shown at the top, indicating the gene order 3′ to 5′. The proteins expressed by the recombinants are depicted by the symbols above the RNA genome. A gene encoding the HIV (89.6) envelope with its cytoplasmic tail replaced by the cytoplasmic tail of VSV G was cloned into the three different infectious VSV plasmids downstream from the indicated VSV G protein genes. The different G protein trimers are shown with distinct shading patterns, as indicated. Diagrams of the three recombinant viruses are shown below. The HIV EnvG protein has been shown to be incorporated into the VSV virion, as indicated (18).
FIG. 2
FIG. 2
Proteins encoded by VSV vectors. Plates of BHK cells were infected with each of the three recombinant envelope exchange viruses expressing HIV envelope for 4 h and then labeled for 1 h with 100 μCi of [35S] methionine. Cell extracts were prepared and electrophoresed on SDS–10% PAGE. All five VSV proteins (N, P, M, G, and L) are expressed by each recombinant virus. The different mobilities of each of the three VSV glycoproteins, indicated as G(I), G(Ch), and G(NJ), can be seen, and all three constructs express the Env G protein. Recombinant wt VSV (rwt) was used as a control.
FIG. 3
FIG. 3
Pathogenesis (weight loss) caused by VSV vectors in mice. Four groups of five to seven mice were inoculated i.n. on day 0 with DMEM (weight control) or with 105 PFU of each of the VSV G protein exchange vectors diagrammed in Fig. 1. After the inoculation, the mice were weighed daily, and average weights are presented.
FIG. 4
FIG. 4
Neutralizing titers to VSV G proteins induced by VSV vectors after initial inoculation and boost with the same vector. Sera from mice inoculated i.n. with the three G protein exchange vectors expressing EnvG 89.6 (Fig. 1) were assayed for neutralizing titer to the homologous G protein at 1 month after inoculation. The mice were boosted with the identical vector at 1 month, and sera were assayed again at 2 months. Neutralizing titers induced to the VSV G proteins at 1 month after inoculation (bars numbered 1) were 1:10,240 to VSV G(I) and 1:5,120 to VSV G(Ch) or VSV G(NJ) and were unchanged 1 month after boost (bars numbered 2). There was no detectable cross-neutralization by heterologous sera (titer less than 1:8).
FIG. 5
FIG. 5
Neutralizing titers induced to VSV vectors following sequential boosting with G protein exchange vectors. Groups of five to seven mice were inoculated sequentially i.n. with the G(I) vector, followed by boosting with the G(Ch) vector at 1 month and boosting with the G(NJ) vector at 2 months. Neutralizing antibody titers to all three VSV G proteins were determined in sera taken at 1, 2, and 3 months, as indicated. Bars at the baseline indicate undetectable cross-neutralization (titer of less than 1:8).
FIG. 6
FIG. 6
Antibody titers to oligomeric HIV Env 89.6 determined by ELISA. Oligomeric HIV 89.6 gp140 envelope protein was bound to plates coated with ConA. Serial dilutions of sera from mice inoculated and boosted with the recombinant VSVs were added to the wells, and antibodies were detected using the ELISA method described in Materials and Methods. Antibody responses to HIV Env after three inoculations given 1 month apart with each of the recombinant viruses [×, G(I); ▴, G(Ch); and ▪, G(NJ)] were virtually identical. Sequential boosting with the three different vectors showed antibody titers to HIV Env that were fourfold higher at an absorbance of 0.25 (♦).
FIG. 7
FIG. 7
Neutralizing antibody titers to HIV envelope protein. Mice were inoculated sequentially with all three VSV G protein exchange vectors encoding HIV EnvG 89.6 and then given a boost (i.p.) with a combination of all three vectors according to the time line shown in panel A. Panel B shows the titers of neutralizing antibody to HIV Env 89.6 measured using an assay based on neutralization of VSVΔG-89.6G-GFP, a virus which lacks the VSV G protein but expresses HIV EnvG 89.6 and GFP (2). Diluted mouse sera were mixed and incubated in duplicate 96-well plates with approximately 100 infectious units of VSVΔG-89.6G-GFP. Then this mixture of serum and virus was added to HeLa T4 cells. A reduction of >50% in the number of GFP-positive cells compared to control mouse serum was scored as positive neutralization.
FIG. 8
FIG. 8
ELISA showing minimal effect of boosting with a vaccinia virus recombinant encoding 89.6 Env protein. Mice were inoculated sequentially with all three VSV G protein exchange vectors encoding HIV EnvG 89.6 and then boosted i.p. with a mixture of all three vectors as in Fig. 7. Mice were rested for 1 month and then boosted i.p. with 2 × 106 PFU of vaccina virus recombinant vBD3 encoding the HIV Env 89.6 protein. One month later, sera were collected from boosted and unboosted animals and assayed by ELISA for antibodies to oligomeric HIV Env 89.6 gp140. Results for serum from control mice vaccinated with vBD3 alone or VSV (GI) EnvG alone and assayed after 1 month are included for comparison.
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