Stabilization of the gp120 V3 loop through hydrophobic interactions reduces the immunodominant V3-directed non-neutralizing response to HIV-1 envelope trimers

Abstract

To provide protective immunity against circulating primary HIV-1 strains, a vaccine most likely has to induce broadly neutralizing antibodies to the HIV-1 envelope glycoprotein (Env) spike. Recombinant Env trimers such as the prototype BG505 SOSIP.664 that closely mimic the native Env spike can induce autologous neutralizing antibodies (NAbs) against relatively resistant (tier 2) primary viruses. Ideally, Env immunogens should present broadly neutralizing antibody epitopes but limit the presentation of immunodominant non-NAb epitopes that might induce off-target and potentially interfering responses. The V3 loop in gp120 is such a non-NAb epitope that can effectively elicit non-NAbs when animals are immunized with SOSIP.664 trimers. V3 immunogenicity can be diminished, but not abolished, by reducing the conformational flexibility of trimers via targeted sequence changes, including an A316W substitution in V3, that create the SOSIP.v4.1 and SOSIP.v5.2 variants. Here, we further modified these trimer designs by introducing leucine residues at V3 positions 306 and 308 to create hydrophobic interactions with the tryptophan residue at position 316 and with other topologically proximal sites in the V1V2 domain. Together, these modifications further stabilized the resulting SOSIP.v5.2 S306L/R308L trimers in the prefusion state in which V3 is sequestered. When we tested these trimers as immunogens in rabbits, the induction of V3 non-NAbs was significantly reduced compared with the SOSIP.v5.2 trimers and even more so compared with the SOSIP.664 prototype, without affecting the autologous NAb response. Hence, these additional trimer sequence modifications may be beneficial for immunization strategies that seek to minimize off-target non-NAb responses.

B-cell restriction – an alternative piece to the puzzle

“…44,45 In essence, the use of trimeric spike proteins, as in the case of SOSIP trimers for HIV-1, ensures that the inner surfaces of gp120 "chaff decoys" are avoided. [46][47][48]…”

Stabilizing HIV-1 envelope glycoprotein trimers to induce neutralizing antibodies

“…First, thermophilic proteins often have an increased number of hydrophobic residues at domain and oligomer interfaces, facilitating tighter packing of protein domains [46,47]. A similar strategy was applied to HIV-1 Env trimers to stabilize the trimer and prevent the exposure of non-NAbs [48][49][50][51][52][53]. For example, de Taeye et al avoided the spontaneous exposure of the V3 loop by increasing the hydrophobic interactions within the V3 domain and between the V3 and V1V2 domains, by Amino acid substitutions that help stabilize soluble native-like trimers.…”

“…Second, immunization with BG505 SOSIP.664 trimers induced a strong response against non-NAb V3 epitopes [10,50,53,70], leading to the hypothesis that this immunodominant V3-response interfered with the generation of bNAb responses. When rabbits were immunized with an improved version of the trimer, the BG505 SOSIP.…”

“…v4 trimer, which contained the A316W mutation that sequestered the V3 epitope, these SOSIP trimers induced weaker anti-V3 responses and V3-directed Tier 1A virus NAb responses, without affecting the autologous NAb response [10,16]. In a next iteration of trimer design, two additional hydrophobic residues were incorporated in the V3 loop of the BG505 SOSIP.v4 trimer (R306L and R308L) to completely abolish the responses against the V3 loop [53]. Although these modifications reduced V3 immunogenicity, they did not improve the autologous NAb responses, nor did they result in a broadening of the NAb response.…”

Structure-based immunogen design — leading the way to the new age of precision vaccines

“…The Env V3 loop is a major target of non-neutralizing Abs, and is exposed in the Env open conformation. To reduce the V3 conformational dynamics, structure-based design was employed to strengthen hydrophobic packing, preventing V3 loop to adopt the open conformation [29,30]. Similarly, Kulp et al employed computational design to replace a network of buried hydrophilic residues by hydrophobic amino acids, thereby limiting V3 exposure [16 ].…”