title: Evaluation of Lipopeptides as Toll-like Receptor 2 Ligands.
abstract: 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.
  
  
  METHODS
  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.
  
  
  CONCLUSIONS
  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.
authors: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Waleed M
authors: WM
authors: Hussein
authors: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Phil M
authors: PM
authors: Choi
authors: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Cheng
authors: C
authors: Zhang
authors: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Mei
authors: M
authors: Su
authors: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Emma
authors: E
authors: Sierecki
authors: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Wayne
authors: W
authors: Johnston
authors: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Vincent
authors: V
authors: Fagan
authors: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Kirill
authors: K
authors: Alexandrov
authors: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Mariusz
authors: M
authors: Skwarczynski
authors: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Yann
authors: Y
authors: Gambin
authors: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Istvan
authors: I
authors: Toth
authors: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD 4072. Australia.
authors: Pavla
authors: P
authors: Simerska
mesh_terms: D000276:Adjuvants, Immunologic / Q000737:chemistry / Q000378:metabolism; D001710:Biotin / Q000737:chemistry / Q000378:metabolism; D018985:Epitopes, B-Lymphocyte / Q000737:chemistry / Q000378:metabolism; D008024:Ligands; D055666:Lipopeptides / Q000737:chemistry / Q000378:metabolism*; D051195:Toll-Like Receptor 2 / Q000378:metabolism*; D051200:Toll-Like Receptor 8 / Q000378:metabolism
chemical_list: D000276:Adjuvants, Immunologic; D018985:Epitopes, B-Lymphocyte; D008024:Ligands; D055666:Lipopeptides; D051195:Toll-Like Receptor 2; D051200:Toll-Like Receptor 8; D001710:Biotin
keywords: AlphaScreen assay; TLR2 agonists; lipoamino acid; lipopeptide; peptide synthesis; vaccines