Lipid and carbohydrate-based systems to enhance the bioavailability and immunogenicity of therapeutic peptides

Goodwin, Daryn (2013). Lipid and carbohydrate-based systems to enhance the bioavailability and immunogenicity of therapeutic peptides PhD Thesis, School of Chemistry and Molecular Biosciences, The University of Queensland. doi:10.14264/uql.2014.150

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Author Goodwin, Daryn
Thesis Title Lipid and carbohydrate-based systems to enhance the bioavailability and immunogenicity of therapeutic peptides
School, Centre or Institute School of Chemistry and Molecular Biosciences
Institution The University of Queensland
DOI 10.14264/uql.2014.150
Publication date 2013
Thesis type PhD Thesis
Supervisor Istvan Toth
Kirill Alexandrov
Pavla Simerska
Total pages 207
Language eng
Subjects 1115 Pharmacology and Pharmaceutical Sciences
0304 Medicinal and Biomolecular Chemistry
Formatted abstract
Peptide and protein based therapeutics represent the most rapidly developing segment of the pharmaceutical market. Inadequate absorption and rapid breakdown by proteolytic enzymes are amongst some of the problems associated with peptide delivery. This project was designed to counter the obstacles in developing novel synthetic peptide and vaccine systems by harnessing the properties of lipids and carbohydrates.

The benefits of lipidation with lipoamino acids (LAAs) were explored for the delivery of Luteinising Hormone-Releasing Hormone (LHRH). It was demonstrated that the addition of LAAs to the N-terminus of LHRH significantly enhanced the stability of the peptide. The specific cleavage of the LAA-peptide bond was observed using LHRH, and this finding was investigated further. By using a library of lipodipeptides as a model, it was determined that hydrolysis between LAAs and large, polar amino acids suggested there is an element of residue specificity. This action was determined to be likely due to an aspartyl protease. The lipidation of LHRH also appeared to confer an increase in permeability across Caco-2 cell monolayers, as long as solubility was not compromised. LAA conjugation to the side chain central to the LHRH sequence also appeared to enhance receptor binding, inferred by the anti-proliferative activity of the conjugate on androgen-dependant tumour cells. Further conjugation of glucose moieties to the LHRH lipopeptides increased proteolytic stability in most cases, however a decrease in epithelial permeability across Caco-2 monolayers was observed.

The design and synthesis of a number of potential LHRH vaccine candidates to eliminate or substantially lower natural levels of LHRH was sought. LAA conjugation was used to render the LHRH sequence immunogenic. These studies confirmed the self-adjuvanting activity of LAA moieties by eliciting significant anti-LHRH IgG titers in mice when the LHRH epitope was incorporated with T-helper epitopes or as a dimer. This work was then expanded to the development of an LHRH vaccine candidate for use in rams. When co-administered with AdjuVac™, an LHRH and T-helper system produced sustainably high anti-LHRH antibody titers, which resulted in a regression in testicular growth and body weight. This scheme was expanded further via the use of rapid biological peptide production systems for the expression of an LHRH tandem-repeat polypeptide. LHRH polypeptides were expressed successfully using a variety of in vivo and in vitro expression methods. The LHRH tandem-repeat polypeptide was shown to elicit significant anti-LHRH IgG titers when co-administered with CFA in mice.

It is evident that incorporating LAAs and/or carbohydrates can enhance the delivery of peptide therapeutics. The specificity, stability and pliability of these systems hold tremendous potential. 
Keyword Peptide
Drug Delivery
Peptide Synthesis
Protein Expression

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Created: Mon, 16 Jun 2014, 19:14:10 EST by Daryn Goodwin on behalf of Scholarly Communication and Digitisation Service