A Chimeric HIV-1 Envelope Glycoprotein Trimer with an Embedded Granulocyte-Macrophage Colony-stimulating Factor (GM-CSF) Domain Induces Enhanced Antibody and T Cell Responses

Abstract

An effective HIV-1 vaccine should ideally induce strong humoral and cellular immune responses that provide sterilizing immunity over a prolonged period. Current HIV-1 vaccines have failed in inducing such immunity. The viral envelope glycoprotein complex (Env) can be targeted by neutralizing antibodies to block infection, but several Env properties limit the ability to induce an antibody response of sufficient quantity and quality. We hypothesized that Env immunogenicity could be improved by embedding an immunostimulatory protein domain within its sequence. A stabilized Env trimer was therefore engineered with the granulocyte-macrophage colony-stimulating factor (GM-CSF) inserted into the V1V2 domain of gp120. Probing with neutralizing antibodies showed that both the Env and GM-CSF components of the chimeric protein were folded correctly. Furthermore, the embedded GM-CSF domain was functional as a cytokine in vitro. Mouse immunization studies demonstrated that chimeric Env GM-CSF enhanced Env-specific antibody and T cell responses compared with wild-type Env. Collectively, these results show that targeting and activation of immune cells using engineered cytokine domains within the protein can improve the immunogenicity of Env subunit vaccines.

Immunosilencing a Highly Immunogenic Protein Trimerization Domain

“…Anti-Env and Anti-HA ELISA-Anti-gp120 Ab titers were measured by D7324-capture ELISA as described previously (34). Anti-HA Ab titers were measured using HA-Fd immobilized on Ni-NTA HisSorb 96-well plates.…”

Targeting HIV-1 Envelope Glycoprotein Trimers to B Cells by Using APRIL Improves Antibody Responses

“…In a second study, fusion of gp120 to Flt-3 ligand or CTLA4 increased murine Env-specific CD8 ϩ T-cell and binding antibody responses, but nAbs were again not assessed (70,82). Trimers of gp140 containing an embedded granulocytemacrophage colony-stimulating factor (GM-CSF) domain induced antibody and T helper responses in mice, which resulted in enhanced neutralization of the highly sensitive SF162 strain (100). Finally, fusing monomeric gp120 and trimeric gp140 to the complement component C3d modestly improved nAb responses in rabbits (12,37,50).…”

Enhanced Immunogenicity of HIV-1 Envelope gp140 Proteins Fused to APRIL

“…The chimeric molecules, denoted Env hAPRIL , Env rAPRIL and Env mAPRIL , respectively, are secreted from mammalian cells, and form trimers efficiently but they are uncleaved (Fig. 1B and C, and data not shown) [13], [18]. These proteins bind CD4 and NAbs against conformational epitopes that constitute protein and glycan (Fig.…”

“…We used rabbits for immunizations because rabbits, in contrast to mice, can produce Abs with long CDRH3 domains that are often found in HIV-1 bNAbs (reviewed in [1]). Furthermore, APRIL does not function very efficiently as an adjuvant in mice [18], [57], [58]. Env APRIL –immunized rabbits had significant increases in anti-gp120 binding Ab titers compared to Env wt at week 6 (after the first boost), week 8, week 10 (after the second boost) and week 12 (terminal bleed).…”