Transport and metabolism of opioid tetrapeptide agonists with a focus on oral and respiratory delivery

Sammanfattning: The clinical development of peptide drugs for oral delivery has often been hindered because of poor transport across the intestinal membranes and degradation in the gastrointestinal tract and liver. Alternative delivery routes through more permeable membranes preferably with less pronounced proteolytic activity, such as the nasal and pulmonary routes, have therefore been considered. The main objective of this thesis was to investigate the transport and metabolism of three model tetrapeptide opioid agonists, TArPN, TArPP and TAPP, in the gastrointestinal and respiratory tracts. Intestinal transport of the tetrapeptides was investigated with the use of a single-pass perfusion technique in anaesthetised rats and across Caco-2 cell monolayers. Metabolism was evaluated using various in vitro models, such as gastrointestinal fluids from humans and rats, rat tissue homogenates, subcellular fractions and purified enzymes. The transport and metabolism of one of the tetrapeptides (TArPP; Tyr-D-Arg-Phe-Phe-NH2) in the respiratory tract were studied using an isolated and perfused rat lung model. The pharmacokinetics of this tetrapeptide were further investigated after respiratory delivery to anaesthetised rats. Liquid chromatography with UV detection or mass spectrometry was used to quantify the tetrapeptides and to identify/quantify peptide metabolites. The tetrapeptides were stable in gastric juice but rapidly metabolised in jejunal fluid from humans and rats, as well as in various rat tissue homogenates, mainly by deamidation. Transport across Caco-2 cell monolayers was low and comparable to that of the paracellular marker mannitol. These investigations suggest that both low intestinal permeation and enzymatic degradation will limit oral delivery of these tetrapeptides. In the isolated and perfused rat lung, TArPP was well absorbed despite some first-pass metabolism. Regional differences in absorption rate, first-pass metabolism and bioavailability were identified when the tetrapeptide was administered to different regions of the rat respiratory tract in vivo. The bioavailability increased with progression from the upper to the lower regions, from about 50% after nasal delivery to complete after pulmonary delivery. Altogether, the promising results following pulmonary administration indicate that this route could be attractive for systemic delivery of small peptides.