Role of immune responses against the envelope and the core antigens of simian immunodeficiency virus SIVmne in protection against homologous cloned and uncloned virus challenge in Macaques

Abstract

We previously showed that envelope (gp160)-based vaccines, used in a live recombinant virus priming and subunit protein boosting regimen, protected macaques against intravenous and intrarectal challenges with the homologous simian immunodeficiency virus SIVmne clone E11S. However, the breadth of protection appears to be limited, since the vaccines were only partially effective against intravenous challenge by the uncloned SIVmne. To examine factors that could affect the breadth and the efficacy of this immunization approach, we studied (i) the effect of priming by recombinant vaccinia virus; (ii) the role of surface antigen gp130; and (iii) the role of core antigens (Gag and Pol) in eliciting protective immunity. Results indicate that (i) priming with recombinant vaccinia virus was more effective than subunit antigen in eliciting protective responses; (ii) while both gp130 and gp160 elicited similar levels of SIV-specific antibodies, gp130 was not as effective as gp160 in protection, indicating a possible role for the transmembrane protein in presenting functionally important epitopes; and (iii) although animals immunized with core antigens failed to generate any neutralizing antibody and were infected upon challenge, their virus load was 50- to 100-fold lower than that of the controls, suggesting the importance of cellular immunity or other core-specific immune responses in controlling acute infection. Complete protection against intravenous infection by the pathogenic uncloned SIVmne was achieved by immunization with both the envelope and the core antigens. These results indicate that immune responses to both antigens may contribute to protection and thus argue for the inclusion of multiple antigens in recombinant vaccine designs.

FIG. 1

ELISA analysis of SIV-specific antibody responses in immunized macaques prior to challenge. Dilutions of macaque sera collected at the indicated times were incubated with disrupted, sucrose gradient-purified SIVmne virions proteins on microtiter plates. Endpoint ELISA (EIA) titers were defined as the reciprocal of the highest dilution that gave an optical absorbance value at least threefold higher than the average values obtained with SIV-negative macaque sera. Panels A through F correspond to experimental groups A through F (Table 1). Positions of the arrows indicate the times of primary (solid arrows) and booster (open arrows) immunizations.

FIG. 2

PBMC provirus load in macaques after intravenous challenge with E11S. Proviral load was determined by PCR analysis with radiolabeled primer incorporation as described in Materials and Methods. Proviral DNA was measured by using an external E11S DNA standard of known quantity. Panels A through F correspond to experimental groups A through F (Table 1). Solid lines indicate infected animals.

FIG. 2

PBMC provirus load in macaques after intravenous challenge with E11S. Proviral load was determined by PCR analysis with radiolabeled primer incorporation as described in Materials and Methods. Proviral DNA was measured by using an external E11S DNA standard of known quantity. Panels A through F correspond to experimental groups A through F (Table 1). Solid lines indicate infected animals.

FIG. 3

SIV-specific antibody responses in immunized macaques on day of challenge with E11S. Sera collected on the day of challenge were analyzed for neutralizing antibody (NAb) activities against the homologous challenge virus E11S or a heterologous virus, SIVmac251. Neutralizing titers are expressed as the reciprocal serum dilutions that results in >90% reduction of syncytium formation by SIVmac251 on HuT78 cells (A) or >50% reduction of cytopathicity of E11S-infected CEMx174 cells (B). Serum reactivity with disrupted SIVmne E11S virion proteins was analyzed by ELISA, and the results are expressed as endpoint titers (C). Designations of experimental groups (A through F) are indicated at the top.

FIG. 4

Plasma viral load in macaques after uncloned SIVmne challenge. Plasma viral load was determined by RT-QC-PCR using an internally controlled template as described in Materials and Methods. Panels B, E, and F correspond to experimental groups B, E, and F. Solid lines indicate infected animals.

FIG. 5

ELISA analysis of SIV-specific antibodies in macaques after uncloned SIVmne challenge. Animals in experimental groups B, E, and F (represented by panels B, E, and F) that were protected against E11S challenge were held for 8 to 9 months to confirm their virus-negative status. They were then boosted again and challenged 4 weeks later with uncloned SIVmne (week 0 on the abscissa). Endpoint titers were defined as the reciprocal of the highest dilution that gave an optical absorbance value at least threefold higher than the average values obtained with SIV-negative macaque sera. Solid lines indicate infected animals.

FIG. 6

SIV-specific antibody responses in immunized macaques on day of challenge with uncloned SIVmne. Sera collected on the day of rechallenge were analyzed for neutralizing antibody (NAb) activities against the homologous challenge virus uncloned SIVmne or a heterologous virus, SIVmac251. Neutralizing titers are expressed as the reciprocal serum dilutions that results in >90% reduction of syncytium formation by SIVmac251 on HuT78 cells (A) or >50% reduction of cytopathicity of uncloned SIVmne-infected CEMx174 cells (B). Serum reactivity with disrupted SIVmne E11S virion proteins was analyzed by ELISA, and the results are expressed as endpoint titers (C). Designations of experimental groups (B, E, and F) are indicated at the top.

FIG. 7

Peripheral blood CD4⁺ T-lymphocyte numbers in immunized and control macaques infected with SIVmne E11S. (A), animals sacrificed because of AIDS; (E), animals euthanized at the end of the experiment; (R), animals reassigned for challenge with uncloned SIVmne. The last datum point for each animal represents the time of death or reassignment. Panels A through F represent data from individual animals groups A through F (Table 1). Cumulative data from 15 naive control animals infected with E11S (Control) were obtained from three separate experiments previously described (76). Data for concurrent controls are indicated by solid symbols; solid lines indicate infected animals.

FIG. 7

Peripheral blood CD4⁺ T-lymphocyte numbers in immunized and control macaques infected with SIVmne E11S. (A), animals sacrificed because of AIDS; (E), animals euthanized at the end of the experiment; (R), animals reassigned for challenge with uncloned SIVmne. The last datum point for each animal represents the time of death or reassignment. Panels A through F represent data from individual animals groups A through F (Table 1). Cumulative data from 15 naive control animals infected with E11S (Control) were obtained from three separate experiments previously described (76). Data for concurrent controls are indicated by solid symbols; solid lines indicate infected animals.

FIG. 8

Peripheral blood CD4⁺ T-lymphocyte numbers in immunized and control macaques infected with uncloned SIVmne. Designations (A) and (E) are as described in the legend to Fig. 7; the animal that died of causes unrelated to AIDS is labeled (U). The last datum point for each animal represents the time of death or termination of the experiment. Panels B, E, and F represent data from individual animals in groups B, E, and F (Table 1). Cumulative data from 10 naive control animals infected with uncloned SIVmne (Control) are obtained from three separate experiments previously described (76). Data for concurrent controls are indicated by solid symbols; solid lines indicate infected animals.