Recent advances in biotechnology have allowed the production of a variety of peptide and protein drugs on a commercial scale that are used in the treatment of life-threatening diseases. Peptides and proteins have drawn a lot of attention due to their macromolecular nature and structural complexity, that offers a very high receptor specificity and potency. However, direct use of native polypeptides as biopharmaceuticals is limited due to their poor pharmacokinetic properties, such as short half-lives and negligible oral bioavailability. The short half-lives of most peptides and small proteins (< 30 kDa) is due to their extensive renal clearance and high susceptibility to enzymatic degradation. Several half-life extension strategies for modification of peptides and proteins have been pursued, and the first project of this thesis was focused on an alternative half-life extension method that directs the nano-scale self-assembly. The poor oral bioavailability of peptides is also associated with enzymatic degradation, in addition to the limited permeation across cell plasma membranes. A strategy to improve the permeation of peptides across the epithelial barrier is to co-administer the potential peptide drugs with permeation enhancers.
Project I aimed to investigate in vivo the size effects of the metal-ion induced self-assembly of a modified Peptide YY_3-36 (PYY_3-36) analogue in a biodistribution context. For this purpose, the peptide analogue was synthesized and modified with a metal-ion-binding 2,2’-bipyridine ligand that enabled self-assembly through metal complexation. Upon addition of CuII, the bipyridine-modified PYY_3-36 peptide bound stoichiometric quantities of metal ions and contributed to the organization of higher-order assemblies that were studied using light scattering. The size effects of the self-assembled peptide were investigated in vivo by using non-invasive quantitative single-photon emission computed tomography/computed tomography (SPECT/CT) imaging. SPECT/CT imaging and biodistribution experiments showed fast renal clearance and accumulation at the kidney cortex. The radiolabeled bipyridyl-PYY_3–36 conjugates with and without Cu^II demonstrated a slightly slower excretion 1 h post injection compared to the unmodified PYY_3–36, thus presenting that higher order self-assemblies of the peptide might have an effect on the pharmacokinetics.
Project II explored a novel class of synthesized neo-glycolipids as potential intestinal permeation enhancers for oral drug delivery. Sixteen novel neo-glycolipids were designed and synthesized based on the known permeation-enhancing characteristics of different medium chain fatty acids (MCFAs; C₈, C₁₀ and C₁₂) and their varying hydrophobicity. To increase the surfactant properties, the lipids were anchored to various unprotected carbohydrate molecules (glucose, lactose, cellobiose and maltose), which have different polarities and configurations, via an oxime linkage. The potential of the synthesized neo-glycolipids on epithelial cell permeation was investigated in vitro across an intestinal epithelium model, the Caco-2 monolayer, by using two marker molecules (Fluorescein-isothiocyanate dextran of 4000 Da (FD4) and ³H-mannitol). Most of the synthesized neo-glycolipids induced an enhanced permeation of the marker molecules when compared to untreated controls. In certain cases, the investigated neo-glycolipids improved FD4 permeation compared to (pre-)clinically known enhancers and showed lower cytotoxic effects on the Caco-2 monolayers. These findings illustrate the potential of the synthesized neo-glycolipids as permeation enhancers for oral drug delivery.