Brucella abortus conjugated with a peptide derived from the V3 loop of human immunodeficiency virus (HIV) type 1 induces HIV-specific cytotoxic T-cell responses in normal and in CD4+ cell-depleted BALB/c mice

Abstract

We have previously shown that immunization of mice with human immunodeficiency virus (HIV)-derived proteins or peptides conjugated to inactivated Brucella abortus induces the secretion of virus-neutralizing antibodies, predominantly of the immunoglobulin G2a (IgG2a) isotype. In addition, B. abortus activates human CD4 ؉ and CD8 ؉ cells to secrete gamma interferon. Since these are both characteristics of a Th1-type immune response, which is associated with the development of cell-mediated immunity, it was important to determine if B. abortus conjugates would also act as a carrier to induce a cytotoxic T-lymphocyte (CTL) response. To test this hypothesis, we conjugated an 18-amino-acid peptide from the V3 loop of the MN strain of HIV-1 gp120 that contains both Band cytotoxic T-cell epitopes to B. abortus (B. abortus-MN 18-mer). A 10-amino-acid fragment of this peptide has been shown to be the minimal CTL determinant presented by murine H-2D d. It was found that two in vivo immunizations with 10 8 organisms of B. abortus-MN 18-mer followed by in vitro stimulation with peptide induced a virus-specific CTL response. Conjugation to B. abortus was required for in vivo priming, since there was no induction of memory CTLs when B. abortus was only mixed with peptide. Targets pulsed with peptide as well as those infected with a vaccinia virus encoding HIV gp160 were killed, demonstrating recognition of naturally processed envelope. Also, major histocompatibility complex-incompatible L cells which were infected with vaccinia viruses that encoded H-2D d , but not H-2K d , and pulsed with peptide were lysed. This demonstrated the appropriate major histocompatibility complex class I restriction. Treatment of the mice with anti-L3T4 prior to immunization caused a severe depletion of CD4 ؉ lymphocytes, yet it did not decrease the CTL priming. Thus, inactivated B. abortus can induce non-CD4 ؉ cells to produce the cytokines required for CTL induction. We conclude that B. abortus stimulates a cellular as well as a humoral immune response, even in the relative absence of CD4 ؉ helper cells. It may be a particularly useful vaccine carrier in HIV-1-infected individuals or others with impaired CD4 ؉ T-cell function.

Down-Regulation of Th2 Responses byBrucella abortus, a Strong Th1 Stimulus, Correlates with Alterations in the B7.2-CD28 Pathway

“…Previously, we and others have shown that the heat-inactivated gram-negative bacterium Brucella abortus in mice (in vitro and in vivo) and in humans (in vitro) promotes T-independent antibody responses (3,13,14,29,39) and production of the Th1 cytokines interleukin-12 (IL-12) and gamma interferon (IFN-␥) and could serve as a potential carrier for vaccine development in situations requiring a strong Th1-like response for protection against infection (39,43,51). As a consequence of Th1 differentiation, B. abortus induces immunoglobulin G2a (IgG2a) antibodies and cytotoxic T-cell responses in mice (13,26).…”

Systemic and mucosal immunity in rhesus macaques immunized with HIV‐1 peptide and gp120 conjugated to Brucella abortus

“…V3‐specific antibody titers were measured by enzyme‐linked immunosorbent assay (ELISA) as described previously [13]. Sera from individual animals were serially diluted then incubated for 1 hour at 37°C.…”

“…This was used, as previously described [13], to measure titers of sera capable of neutralizing HIV‐1 by inhibiting cells infected with HIV‐1 from fusing to CD4 + T cells. Reference human serum, derived from a non‐progressor HIV‐1 infected individual, was obtained from the NIH AIDS Reagent Repository.…”

“…The V3 peptide we used acts as a CTL epitope for CD8 + T cells in H2b mice [13], but its ability to induce CTL in macaques is unknown. Recently it has been shown that IFN‐ γ responses, especially when elicited in CD8 + T cells, correlate with CTL responses [15].…”

“…Our previous studies using heat‐inactivated Brucella abortus (HKBA) as a carrier for HIV‐1 peptides or proteins have provided evidence in mice that anti‐HIV‐1 antibody and cytotoxic T lymphocyte (CTL) memory responses can be induced even when CD4 + T cells are lacking [6, 7, 13, 18]. In addition, we have shown that cellular and humoral responses can be elicited at mucosal sites in mice [8].…”