Abstract
The vaccine elicitation of broadly neutralizing antibodies against HIV-1 is a long-sought goal. We previously reported the amino-terminal eight residues of the HIV-1-fusion peptide (FP8) – when conjugated to the carrier protein, keyhole limpet hemocyanin (KLH) – to be capable of inducing broadly neutralizing responses against HIV-1 in animal models. However, KLH is a multi-subunit particle derived from a natural source, and its manufacture as a clinical product remains a challenge. Here we report the preclinical development of recombinant tetanus toxoid heavy chain fragment (rTTHC) linked to FP8 (FP8-rTTHC) as a suitable FP-conjugate vaccine immunogen. We assessed 16 conjugates, made by coupling the 4 most prevalent FP8 sequences with 4 carrier proteins: the aforementioned KLH and rTTHC; the H. influenzae protein D (HiD); and the cross-reactive material from diphtheria toxin (CRM197). While each of the 16 FP8-carrier conjugates could elicit HIV-1-neutralizing responses, rTTHC conjugates induced higher FP-directed responses overall. A Sulfo-SIAB linker yielded superior results over an SM(PEG)2 linker but combinations of carriers, conjugation ratio of peptide to carrier, or choice of adjuvant (Adjuplex or Alum) did not significantly impact elicited FP-directed neutralizing responses in mice. Overall, SIAB-linked FP8-rTTHC appears to be a promising vaccine candidate for advancing to clinical assessment.
Highlights
The fusion peptide (FP) site of vulnerability on the HIV-1 envelope (Env) glycoprotein has recently been shown to be a promising vaccine target[1,2,3]
A total of 16 FP8-conjugate immunogens were made by coupling FP8 peptides, synthesized with an appended C-terminal Cys, with the heterobifunctional linker m-maleimidobenzoyl-N-hydroxysuccinimide ester (MBS), to exposed lysine residues on carrier proteins (Fig. 1d)
With immunization involving FP linked to keyhole limpet hemocyanin (KLH) eliciting cross-clade neutralizing responses of surprising breadth against HIV-1 in standard vaccine-test animals including mice, guinea pigs and rhesus macaques[1,2,12], it is becoming ever more critical to test FP-conjugated carriers clinically, to determine if similar broad responses against FP can be elicited by vaccination of humans
Summary
The fusion peptide (FP) site of vulnerability on the HIV-1 envelope (Env) glycoprotein has recently been shown to be a promising vaccine target[1,2,3]. When the N-terminal 6–10 residues of FP are coupled to keyhole limpet hemocyanin (KLH), a standard protein carrier widely used in biotechnology, the resultant FP-KLH conjugate immunogens are able to induce broadly neutralizing FP-directed immune responses in mice, guinea pigs, and rhesus macaques[1,2,12]. Vaccine-induced FP-directed antibodies from mice or NHP neutralize up to 31% or 59%, respectively, of a cross-clade panel of 208 HIV-1 strains[2] These results (illustrated in Fig. 1a) indicate FP coupled to a carrier protein to be a promising candidate immunogen. One method of coupling peptides to carrier proteins is through a heterobifunctional crosslinker, with a sulfhydryl-reactive group to react with a cysteine side chain on the peptide and an amine-reactive group to react with surface-exposed lysine side chains on the carrier protein Several such crosslinkers are available with varying spacer lengths, flexibility/rigidity, and hydrophilicity. FP8-rTTHC with Sulfo-SIAB linker appeared to be a promising candidate immunogen for advancing to clinical assessment
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