Polyclonal bovine sera but not virus-neutralizing monoclonal antibodies block bovine leukemia virus (BLV) gp51 binding to recombinant BLV receptor BLVRcp1.

Bovine leukemia virus (BLV), a transactivating lymphotropic retrovirus, is the etiologic agent of enzootic lymphosarcoma or leukemia in cattle. Sera from BLV-infected animals possess high BLV-neutralizing antibody titres. The availability of the recombinant BLV receptor candidate, BLVRcp1, allowed us to determine a mechanism of virus neutralization by polyclonal sera and monoclonal antibodies (MAbs). Bovine sera from animals naturally infected with BLV blocked gp51 binding to recombinant BLVRcp1. In contrast, virus-neutralizing MAbs specific for gp51 F, G, and H epitopes did not prevent gp51-receptor attachment. Furthermore, gp51 neutralization epitopes F, G, and H were accessible to antibodies following gp51 attachment to BLVRcp1. This finding implies that virus neutralization by MAbs to defined BLV gp51 epitopes can occur subsequent to virus engagement of the receptor while polyclonal sera can specifically block virus attachment to the receptor. In conclusion, these data suggest that cell infection by BLV is a multistep process requiring receptor binding (inhibited by polyclonal sera) followed by a second, postbinding event(s) at the cell membrane (inhibited by anti-gp51 MAbs).     

T-cell responses to highly conserved CD4 and CD8 epitopes on the outer membrane protein of bovine leukemia virus: relevance to vaccine development.

Bovine leukemia virus (BLV) is a retrovirus that infects cattle and sheep and may provide a model for studying human leukemia. Cell-mediated immune mechanisms may play a major role in protection against BLV infection. We describe here for the first time the identification of proliferative (CD4) and cytotoxic T-lymphocyte (CD8) epitopes of the gp51 envelope (env) protein of BLV. This protein and a recombinant form expressed by a vaccinia virus construct have been shown to be potential vaccine candidates. A complete series of overlapping peptides, 20 amino acids in length, was prepared to identify epitopes from gp51. These peptides were tested for the ability to elicit peripheral blood lymphocyte proliferation and cytotoxic T-lymphocyte responses in infected and uninfected cattle and sheep. Peptides 51-70 and 61-80 produced a proliferative response in lymphocytes from only uninfected animals (both sheep and cattle), and this was shown by T-cell subset deletion to be a CD4-mediated response. Seven BLV-infected sheep did not show a response to either peptide. Cytotoxic T-lymphocyte activity, however, was associated only with peptides 121-140 and 131-150. In this case, the response was demonstrated to be CD8 dependent and was found only in BLV-infected animals (sheep). Knowledge of the location of these T-cell recognition domains will complement data available on B-cell epitopes in gp51 and may be useful in the design of a subunit vaccine.     

The influence of ovine MHC class II DRB1 alleles on immune response in bovine leukemia virus infection

We have reported previously that the alleles of the ovine leukocyte antigen (OLA)-DRB1 gene that encode the Arg-Lys (RK) motif and the Ser-Arg (SR) motif at positions beta70/71 of the OLA-DRbeta1 domain are associated with resistance and susceptibility, respectively, to development of bovine leukemia virus (BLV)-induced ovine lymphoma. Here, to investigate the different immune response in sheep that carried alleles associated with resistance and susceptible for 30 weeks after infection with BLV, we selected sheep that had the RK/RK or SR/SR genotype among the 52 sheep analyzed by polymerase chain reaction-restriction fragment length polymorphism and DNA sequencing of PCR product for the OLA-DRB1 exon 2 and infected them with BLV. Although the number of BLV-infected cells and virus titer had been maintaining low levels throughout the experimental period, the sheep with the RK/RK genotype could induce expansion of CD5- B-cells and rapid production of neutralizing antibody in the early phase of infection. The level of incorporation of [3H]thymidine by peripheral blood mononuclear cells from the sheep with RK/RK genotype gave a strong response to BLV virion antigen and synthetic antigenic peptides that corresponded to T-helper epitope of the BLV envelope glycoprotein gp51. In contrast, the sheep with SR/SR genotype showed a strong response to BLV virion antigen and synthetic antigenic peptides that corresponded to T-cytotoxic and B-cell epitopes. In such cases, the animals with the RK/RK strongly expressed IFN-gamma, the animals with SR/SR genotype strongly expressed IL-2. To determine the proliferating cells, we tried a blocking assay with monoclonal antibodies such as anti-CD4, -CD8 and -DR molecule. We found that these proliferating cells were MHC-restricted CD4+ T-cells.     

Protection of cattle against bovine leukemia virus (BLV) infection could be attained by DNA vaccination

The bovine leukemia virus (BLV) envelope gene encoding extracellular glycoprotein gp51 and transmembrane glycoprotein gp30 was cloned into a vehicle expression vector under the human cytomegalovirus (CMV) intermediate early promoter. The intramuscular injection of this plasmid vector generated a cellular immune response. Seven out of ten cows vaccinated with the DNA construct resisted a drastic challenge (500 BLV-infected lymphocytes as an infectious dose).     

Complete Bovine Leukemia Virus (BLV) Provirus Is Conserved in BLV-Infected Cattle throughout the Course of B-Cell Lymphosarcoma Development

Bovine leukemia virus (BLV) and human T-cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) belong to the same subfamily of oncoviruses. Defective HTLV-1 proviral genomes have been found in more than half of all patients with adult T-cell leukemia examined. We have characterized the genomic structure of integrated BLV proviruses in peripheral blood lymphocytes and tumor tissue taken from animals with lymphomas at various stages. Genomic Southern hybridization with SacI, which generates two major fragments of BLV proviral DNA, yielded only bands that corresponded to a full-size provirus in all of 23 cattle at the lymphoma stage and in 7 BLV-infected but healthy cattle. Long PCR with primers located in long terminal repeats clearly demonstrated that almost the complete provirus was retained in all of 27 cattle with lymphomas and in 19 infected but healthy cattle. However, in addition to a PCR product that corresponded to a full-size provirus, a fragment shorter than that of the complete virus was produced in only one of the 27 animals with lymphomas. Moreover, when we performed conventional PCR with a variety of primers that spanned the entire BLV genome to detect even small defects, PCR products were produced that specifically covered the entire BLV genome in all of the 40 BLV-infected cattle tested. Therefore, it appears that at least one copy of the full-length BLV proviral genome was maintained in each animal throughout the course of the disease and, in addition, that either large or small deletions of proviral genomes may be very rare events in BLV-infected cattle.     

Association of BLV infection profiles with alleles of the BoLA-DRB3.2 gene

Bovine leukaemia virus (BLV) causes lymphosarcoma and persistent lymphocytosis (PL). Some MHC class II gene polymorphisms have been associated with resistance and susceptibility to the development of lymphosarcoma and PL, as well as with a reduced number of circulating BLV-infected lymphocytes. Previously, 230 BLV-infected Holstein cattle were classified into two infection profiles characterized by low and high proviral loads (LPL and HPL respectively). Here, the influence of the polymorphism at the BoLA-DRB3.2* gene of these animals was examined. After genotyping, the association between the BoLA-DRB3.2* alleles and the BLV infection profile was determined as the odds ratio (OR). Two subtypes of allele *11 were identified (ISAG *0901 and *0902 ). Allele ISAG *0902 showed a stronger association with the LPL profile (OR = 8.24; P  <   0.0001) than allele *11 itself (OR = 5.82; P  <   0.0001). Allele ISAG *1701 ( *12 ) also showed significant association with the LPL profile (OR = 3.46; P  <   0.0055). Only one allele, ISAG *1501 or 03 ( *16 ), showed significant association with HPL (OR = 0.36; P  <   0.0005). The DRB3.2* alleles were assigned to three categories: resistant ( R ), susceptible ( S ) and neutral ( N ). Based on their DRB3 genotypes, cattle were classified as homozygous or heterozygous. The RR and RN genotypes were associated with the LPL profile, while the SS and NS genotypes were associated with the HPL profile. The RS genotype could not be associated with any particular profile. Our results show that allele ISAG *0902 appears to be the best BLV resistance marker in Holstein cattle.     

Progression to persistent lymphocytosis and tumor development in bovine leukemia virus (BLV)-infected cattle correlates with impaired proliferation of CD4+ T cells in response to gag- and env-encoded BLV proteins.

The mechanism of leukemogenesis and persistent lymphocytosis (PL; benign expansion of B lymphocytes) in cattle infected with bovine leukemia virus (BLV; a retrovirus closely related to human T-cell leukemia virus type 1) is unknown; however, the immune system likely plays an important role in controlling the outcome of infection. In this study, we compared T-cell competence in serologically positive alymphocytotic (AL) animals with T-cell functions in animals with progressive stages of infection, PL and tumor bearing (TB). Dramatic differences were observed in lymphocyte proliferation to recombinant proteins encoded by BLV gag (p12, p15, and p24) and env (gp30, and gp51) genes in different disease stages. Lymphocytes from AL cattle recognized an average of three of five recombinant proteins per animal. Expansion of antigen pulsed lymphocytes in interleukin-2 increased protein recognition to almost five per animal. In contrast, lymphocytes from PL and TB animals failed to recognize any BLV recombinant proteins. Short-term T-cell cultures from the PL group expanded in interleukin-2, as well as the PL and TB cells cultured in indomethacin (3 to 6 microg/ml), increased the average of recognized proteins per animal to one. Cells proliferating to BLV antigens were CD4+ T lymphocytes, as shown by cell depletion studies. The positive effect of indomethacin suggests involvement of prostaglandin E2 as a negative regulatory factor in the later stages of disease. Thus, for the first time, advancing stages of BLV infection were correlated with decreased T-cell competence, providing deeper insight into pathogenesis of retroviral infections.     

Ex vivo survival of peripheral blood mononuclear cells in sheep induced by bovine leukemia virus (BLV) mainly occurs in CD5- B cells that express BLV

Bovine leukemia virus (BLV) is the etiologic agent of enzootic bovine leukosis (EBL). In a previous report, we found that in a sheep model, only CD5(-) B cells proliferated clonally, while CD5(+) B cells rapidly decreased when the disease progressed to the lymphoma stage. We demonstrate here that, although both CD5(+) and CD5(-) B cells, but not CD4(+) T, CD8(+) T and gammadeltaTCR(+)T cells, are protected from spontaneous ex vivo apoptosis in sheep infected with wild-type and a mutant BLV that encodes a mutant Tax D247G protein with elevated trans-activation activity, only CD5(-) B cells become the main target for ex vivo survival when the disease proceeds to the persistent lymphocytotic stage, which showed an increased expansion of the CD5(-) B cells. In addition, we identified, by four-color flow cytometric analysis, that in CD5(-) B cells, the apoptotic rates of cells that expressed wild-type and mutant BLV were greatly decreased compared with those of BLV-negative cells. There was only a slight reduction in the apoptotic rates in BLV-positive cells from CD5(+) B cells. In addition, supernatants from peripheral blood mononuclear cell (PBMC) cultures from wild-type- and mutant BLV-infected sheep mainly protected CD5(-) B cells from spontaneous apoptosis. Our results suggest that, although BLV can protect both CD5(+) and CD5(-) B cells from ex vivo apoptosis, the mechanisms accounting for the ex vivo survival between these two B-cell subsets differ. Therefore, it appears that the phenotypic changes in cells that express CD5 at the lymphoma stage could result from a difference in susceptibility to apoptosis in CD5(+) and CD5(-) B cells in BLV-infected sheep.     

Augmentation of bovine leukemia virus (BLV)-specific lymphocyte proliferation responses in ruminants by inoculation with BLV env-recombinant vaccinia virus: their role in the suppression of BLV replication.

Lymphocyte proliferation responses were investigated in sheep and cattle, in which the replication of bovine leukemia virus (BLV) had been known to be suppressed by inoculation with recombinant vaccinia virus (rVV) expressing BLV envelope glycoprotein (gp60). Enhanced lymphocyte proliferation responses were observed in animals inoculated with rVV, regardless of whether they were naive or BLV carriers. These responses were roughly inversely correlated to the growth of BLV in the peripheral blood leukocytes. In contrast, there was no apparent correlation between humoral immune response and BLV growth. Based on these results, it was suggested that rVV rendered its suppressive effect of BLV replication primarily via augmentation of cell-mediated immunity.     

Mapping of B-neutralizing and T-helper cell epitopes on the bovine leukemia virus external glycoprotein gp51.

A battery of 19 synthetic peptides was used to characterize efficient neutralizing and helper T-cell epitopes on the bovine leukemia virus (BLV) external envelope glycoprotein gp51. Four of the antipeptide antisera raised in rabbits inhibited the formation of BLV-induced syncytia; these antisera are directed against peptides 64-73, 98-117, and 177-192. Only antisera directed against the 177-192 region also neutralized vesicular stomatitis virus-BLV pseudotypes. This study clearly demonstrates that neutralizing properties can be observed with antibodies raised to regions undescribed so far and included in both the amino-terminal and central parts of the antigen. In addition, some helper T-cell determinants were defined from gp51-immunized mice and from BLV-infected cattle. Although none of the peptides tested behaved as a universal helper T-cell epitope, peptide 98-117 stimulated T-cell proliferation from BALB/c mice and from three infected cows, while peptide 169-188 strongly stimulated T-cell proliferation from one infected cow. Further experiments performed with three peptides overlapping the 169-188 region (177-192, 179-192, 181-192) demonstrated the particular relevance of residue(s) P-177 and/or D-178 in the helper T-cell epitope. These data should assist in the design of an efficient subunit vaccine against BLV infection that contains peptides possessing both B-neutralizing and helper T-cell determinants.     

Pathogenicity of molecularly cloned bovine leukemia virus.

To delineate the mechanisms of bovine leukemia virus (BLV) pathogenesis, four full-length BLV clones, 1, 8, 9, and 13, derived from the transformed cell line FLK-BLV and a clone construct, pBLV913, were introduced into bovine spleen cells by microinjection. Microinjected cells exhibited cytopathic effects and produced BLV p24 and gp51 antigens and infectious virus. The construct, pBLV913, was selected for infection of two sheep by inoculation of microinjected cells. After 15 months, peripheral blood mononuclear cells from these sheep served as inocula for the transfer of infection to four additional sheep. All six infected sheep seroconverted to BLV and had detectable BLV DNA in peripheral blood mononuclear cells after amplification by polymerase chain reaction. Four of the six sheep developed altered B/T-lymphocyte ratios between 33 and 53 months postinfection. One sheep died of unrelated causes, and one remained hematologically normal. Two of the affected sheep developed B lymphocytosis comparable to that observed in animals inoculated with peripheral blood mononuclear cells from BLV-infected cattle. This expanded B-lymphocyte population was characterized by elevated expression of B-cell surface markers, spontaneous blastogenesis, virus expression in vitro, and increased, polyclonally integrated provirus. One of these two sheep developed lymphocytic leukemia-lymphoma at 57 months postinfection. Leukemic cells had the same phenotype and harbored a single, monoclonally integrated provirus but produced no virus after in vitro cultivation. The range in clinical response to in vivo infection with cloned BLV suggests an important role for host immune response in the progression of virus replication and pathogenesis.     

IL-12 delivery from recombinant vaccinia virus attenuates the vector and enhances the cellular immune response against HIV-1 Env in a dose-dependent manner.

To develop vaccination strategies against HIV-1 infection aimed to specifically enhance the cell-mediated immunity (CMI), we have engineered vaccinia virus (VV) recombinants expressing HIV-1 Env (rVVenv) and murine IL-12 (rVVlucIL-12) genes or coexpressing both genes (rVVenvIL-12). In mice inoculated with rVVlucIL-12 there is a rapid clearance of the virus, and this correlates with the induction of high levels of IL-12 and IFN-gamma in serum and spleen early after infection. Enzyme-linked immunospot analysis of mice inoculated with rVVlucIL-12, revealed a nearly 2-fold increase in the number of specific anti-VV CD8+ T cells compared with that in mice given control rVV, and the serum Ab response was biased in favor of a Th1 response. An enhancement of about 2-fold in the number of anti-gp160 IFN-gamma-secreting CD8+ T cells was observed in mice inoculated with rVVenvIL-12, when a dose of 1 x 107 PFU/mouse was used, but this enhancement was not observed when mice were given 5 x 107 PFU. This variation with virus dosage was confirmed in mice immunized simultaneously with different multiplicities of rVV expressing singly the env or IL-12 genes. The highest specific CMI was obtained in mice coadministered a low dose (2 x 104 PFU) of rVVlucIL-12 and 1 x 107 PFU of rVVenv. Our findings provide evidence for specific enhancement of the CMI to HIV-1 Env by the differential expression of IL-12 and env genes delivered from VV recombinants. This approach can be of wide vaccination interest as a means to improve immune responses to other Ags.     

Persistent infection of macaques with simian-human immunodeficiency viruses.

Chimeric simian-human immunodeficiency viruses (SHIV) containing the human immunodeficiency virus type 1 (HIV-1) tat, rev, env, and, in some cases, vpu genes were inoculated into eight cynomolgus monkeys. Viruses could be consistently recovered from the CD8-depleted peripheral blood lymphocytes of all eight animals for at least 2 months. After this time, virus isolation varied among the animals, with viruses continuing to be isolated from some animals beyond 600 days after inoculation. The level of viral RNA in plasma during acute infection and the frequency of virus isolation after the initial 2-month period were higher for the Vpu-positive viruses. All of the animals remained clinically healthy, and the absolute numbers of CD4-positive lymphocytes were stable. Antibodies capable of neutralizing HIV-1 were generated at high titers in animals exhibiting the greatest consistency of virus isolation. Strain-specific HIV-1-neutralizing antibodies were initially elicited, and then more broadly neutralizing antibodies were elicited. env sequences from two viruses isolated more than a year after infection were analyzed. In the Vpu-negative SHIV, for which virus loads were lower, a small amount of env variation, which did not correspond to that found in natural HIV-1 variants, was observed. By contrast, in the Vpu-positive virus, which was consistently isolated from the host animal, extensive variation of the envelope glycoproteins in the defined variable gp120 regions was observed. Escape from neutralization by CD4 binding site monoclonal antibodies was observed for the viruses with the latter envelope glycoproteins, and the mechanism of escape appears to involve decreased binding of the antibody to the monomeric gp120 glycoproteins. The consistency with which SHIV infection of cynomolgus monkeys is initiated and the similarities in the neutralizing antibody response to SHIV and HIV-1 support the utility of this model system for the study of HIV-1 prophylaxis.     

Signature for long-term vaccine-mediated control of a Simian and human immunodeficiency virus 89.6P challenge: stable low-breadth and low-frequency T-cell response capable of coproducing gamma interferon and interleukin-2

In 2001, we reported 20 weeks of control of challenge with the virulent 89.6P chimera of simian and human immunodeficiency viruses (SHIV-89.6P) by a Gag-Pol-Env vaccine consisting of DNA priming and modified vaccinia virus Ankara boosting. Here we report that 22 out of 23 of these animals successfully controlled their viremia until their time of euthanasia at 200 weeks postchallenge. At euthanasia, all animals had low to undetectable viral loads and normal CD4 counts. During the long period of viral control, gamma interferon (IFN-gamma)-producing antiviral T cells were present at unexpectedly low breadths and frequencies. Most animals recognized two CD8 and one CD4 epitope and had frequencies of IFN-gamma-responding T cells from 0.01 to 0.3% of total CD8 or CD4 T cells. T-cell responses were remarkably stable over time and, unlike responses in most immunodeficiency virus infections, maintained good functional characteristics, as evidenced by coproduction of IFN-gamma and interleukin-2. Overall, high titers of binding and neutralizing antibody persisted throughout the postchallenge period. Encouragingly, long-term control was effective in macaques of diverse histocompatibility types.     

Correlation between the immune response elicited in rabbits by env-recombinant avipox vaccines and the inhibition of HIV-1-specific functions

The fine immunoreactivity of the rabbit humoral response elicited by four env-recombinant avipoxviruses and their ability to stimulate a memory T-cell response and a protective immunity have been studied. The antibody specificity was compared with the serum neutralizing activity and virus-specific T-cell proliferative response. Resistance to challenge by cell-associated HIV-1 was monitored by PCR. Canarypox (CP) and fowlpox (FP) constructs, containing the complete env gene (IS(+)) from the HIV-1(SF2) strain, induced a higher profile of epitope recognition than their counterparts expressing the env gene deleted of the putative immunosuppressive region (IS(-)). Serum neutralizing activity was in agreement with fusion inhibition and lymphoproliferative response in rabbits immunized with CPIS(+), and only partially with FPIS(+). Rabbits failed to be infected, but anti- p55 gag-specific antibodies could be demonstrated by Western blot. This study confirms the ability of these non-replicative live recombinant viruses to elicit a complete immune response, capable of inhibiting specific HIV-1 functions.     

Recombinant modified vaccinia virus ankara expressing the surface gp120 of simian immunodeficiency virus (SIV) primes for a rapid neutralizing antibody response to SIV infection in macaques

Neutralizing antibodies were assessed before and after intravenous challenge with pathogenic SIVsmE660 in rhesus macaques that had been immunized with recombinant modified vaccinia virus Ankara expressing one or more simian immunodeficiency virus gene products (MVA-SIV). Animals received either MVA-gag-pol, MVA-env, MVA-gag-pol-env, or nonrecombinant MVA. Although no animals were completely protected from infection with SIV, animals immunized with recombinant MVA-SIV vaccines had lower virus loads and prolonged survival relative to control animals that received nonrecombinant MVA (I. Ourmanov et al., J. Virol. 74:2740-2751, 2000). Titers of neutralizing antibodies measured with the vaccine strain SIVsmH-4 were low in the MVA-env and MVA-gag-pol-env groups of animals and were undetectable in the MVA-gag-pol and nonrecombinant MVA groups of animals on the day of challenge (4 weeks after final immunization). Titers of SIVsmH-4-neutralizing antibodies remained unchanged 1 week later but increased approximately 100-fold 2 weeks postchallenge in the MVA-env and MVA-gag-pol-env groups while the titers remained low or undetectable in the MVA-gag-pol and nonrecombinant MVA groups. This anamnestic neutralizing antibody response was also detected with T-cell-line-adapted stocks of SIVmac251 and SIV/DeltaB670 but not with SIVmac239, as this latter virus resisted neutralization. Most animals in each group had high titers of SIVsmH-4-neutralizing antibodies 8 weeks postchallenge. Titers of neutralizing antibodies were low or undetectable until about 12 weeks of infection in all groups of animals and showed little or no evidence of an anamnestic response when measured with SIVsmE660. The results indicate that recombinant MVA is a promising vector to use to prime for an anamnestic neutralizing antibody response following infection with primate lentiviruses that cause AIDS. However, the Env component of the present vaccine needs improvement in order to target a broad spectrum of viral variants, including those that resemble primary isolates.     

gammadelta(+) T-Lp6phocyte cytotoxicity against envelope-expressing target cells is unique to the alymphocytic state of bovine leukemia virus infection in the natural host

Bovine leukemia virus (BLV) is a complex B-lymphotrophic retrovirus of cattle and the causative agent of enzootic bovine leukosis. Serum antibody in infected animals does not correlate with protection from disease, yet only some animals develop severe disease. While a cytotoxic T-lymphocyte response may be responsible for directing BLV pathogenesis, this possibility has been left largely unexplored, in part since the lack of readily established cytotoxic target cells in cattle has hampered such studies. Using long-term naturally infected alymphocytic (AL) cattle, we have established the existence of cytotoxic T-lymphocyte response against BLV envelope proteins (Env; gp51/gp30). In vitro-expanded peripheral blood mononuclear (PBM) cell effector populations consisted mainly of gammadelta(+) (>40%), CD4(+) (>35%), and CD8(+) (>10%) T lymphocytes. Specific lysis of autologous fibroblasts infected with recombinant vaccinia virus (rVV) delivering the BLV env gene ranged from 30 to 65%. Depletion studies indicated that gammadelta(+) and not CD8(+) T cells were responsible for the cytotoxicity against autologous rVVenv-expressing fibroblasts. Additionally, cultured effector cells lysed rVVenv-expressing autologous fibroblasts and rVVenv-expressing xenogeneic targets similarly, suggesting a lack of genetic restricted killing. Restimulation of effector populations increased the proportion of gammadelta(+) T cells and concomitantly Env-specific cytolysis. Interestingly, culture of cells from BLV-negative or persistently lymphocytic cattle failed to elicit such cytotoxic responses or increase in gammadelta(+) T-cell numbers. These results imply that cytotoxic gammadelta(+) T lymphocytes from only AL cattle recognize BLV Env without a requirement for classical major histocompatibility complex interactions. It is known that gammadelta(+) T lymphocytes are diverse and numerous in cattle, and here we show that they may serve a surveillance role during natural BLV infection.     

Control of viremia and prevention of simian-human immunodeficiency virus-induced disease in rhesus macaques immunized with recombinant vaccinia viruses plus inactivated simian immunodeficiency virus and human immunodeficiency virus type 1 particles

An effective vaccine against the human immunodeficiency virus type 1 (HIV-1) will very likely have to elicit both cellular and humoral immune responses to control HIV-1 strains of diverse geographic and genetic origins. We have utilized a pathogenic chimeric simian-human immunodeficiency virus (SHIV) rhesus macaque animal model system to evaluate the protective efficacy of a vaccine regimen that uses recombinant vaccinia viruses expressing simian immunodeficiency virus (SIV) and HIV-1 structural proteins in combination with intact inactivated SIV and HIV-1 particles. Following virus challenge, control animals experienced a rapid and complete loss of CD4(+) T cells, sustained high viral loads, and developed clinical disease by 17 to 21 weeks. Although all of the vaccinated monkeys became infected, they displayed reduced postpeak viremia, had no significant loss of CD4(+) T cells, and have remained healthy for more than 15 months postinfection. CD8(+) T-cell and neutralizing antibody responses in vaccinated animals following challenge were demonstrable. Despite the control of disease, virus was readily isolated from the circulating peripheral blood mononuclear cells of all vaccinees at 22 weeks postchallenge, indicating that immunologic control was incomplete. Virus recovered from the animal with the lowest postchallenge viremia generated high virus loads and an irreversible loss of CD4(+) T-cell loss following its inoculation into a na?ve animal. These results indicate that despite the protection from SHIV-induced disease, the vaccinated animals still harbored replication-competent and pathogenic virus.