Design, Synthesis and Test of Novel Lipidated Cysteine Analogs as Efficient TLR2/6 Agonists

Abstract

Synthetic vaccines use synthetic molecules as antigens for vaccination. Since real pathogens are no longer needed in synthetic vaccines, they are considered safer than the traditional inactivated or attenuated vaccines. The development of synthetic vaccines can be achieved on the genome sequence of the pathogen, which is much more efficient than the traditional way of sampling pathogens from the outbreak and grow cultures. Despite these advantages, the development of synthetic vaccines is hindered by the poor immunogenicity of synthetic antigens and the lack of effective adjuvants that enhance immune responses [2]. S-[2,3-bis(palmitoyloxy)propyl]cysteine (Pam2Cys) was found to be a promising candidate as a synthetic vaccine adjuvant [1]. The molecule is found as the lipid component of macrophageactivating lipopeptide 2 (MALP-2) on the membrane of mycoplasma [3], and is able to activate the heterodimer TLR2/6 [1]. To make it a peptide vaccine adjuvant, Pam2Cys was conjugated to both a Th epitope and a target epitope through the N-terminal amino group and the e-amino group of a lysine residue [1] (Figure 1). The construct was found to be able to induce both cytotoxic T cell immunity and antibody responses [1]. Nevertheless, the development of Pam2Cys based synthetic vaccine is hindered by the high cost of Pam2Cys synthesis. The synthesis involves orthogonal protection-deprotection techniques [4], which is labor-intensive. Here we report a novel synthetic pathway that can be used to effectively synthesize N-acetyl Pam2Cys analogues which share a similar structure as Pam2Cys (Figure 2). Our results suggest that N-acetyl Pam2Cys analogues are able to activate TLR2/6. Our in silico docking experiment shows that N-acetyl Pam2Cys analogues can interact well with the TLR2/6. N-acetyl Pam2Cys analogues are shown to be embedded deeper into the ligand binding pocket comparing to Pam2Cys. Our results suggest that N-acetyl Pam2Cys analogues, particularly N-acetyl Str2Cys, can serve as synthetic vaccine adjuvants with lower costs.