Background: Peptide-based vaccines are considered to be the next generation of modern immunizations,
as they are safe, easy to produce and well-defined. However, due to their weak immunogenic
effect, it is important to first develop an appropriate adjuvant for peptide-based vaccines.
Objective: The aim of this work was to synthesize a series of four adjuvanting moieties as alkyne
derivatives, incorporating dipalmitoyl serine (DPS), 1,3-diglyceride (DG), two hexadecane lipoamino
acids (diLAA), and 2,3-dipalmitoyl-S-glycerylcysteine (Pam2Cys). Next aim was to synthesize and attach
the azide derivative of biotinylated J14 peptide (model B-cell epitope) to the alkynes through copper-
catalyzed alkyne-azide 1,3-dipolar cycloaddition (CuAAC) reaction. Final aim was to test the ability
of the final biotin labeled conjugates to directly interact with in vitro expressed TLR2 and 8 using
AlphaScreen proximity assay.
Method: All of the peptides were synthesized by manual stepwise solid phase peptide synthesis (SPPS)
on rink amide MBHA resin using HATU/DIPEA Fmoc-chemistry. The target compounds were synthesized
in a solution phase using CuAAC reaction.
Results: Pam2Cys analogue bound to TLR2 as expected. Analogues of DPS and C16-LAA showed
also affinity to TLR2, while it did not bind to the control protein (TLR8), demonstrating ability of the
DPS and C16-LAA to be recognized by TLR2.
Conclusion: Four alkyne derivatives of lipids were successfully synthesized and coupled to a biotinylated
J14 peptide to give a series of self-adjuvanting ligands. These ligands showed different affinity to
TLR2 upon testing by AlphaScreen assay. The DPS derivative showed the most promising affinity in
comparison to the standard TLR2 agonist, Pam2Cys.