Glucagon Like Peptide 1 Receptor Agonists for Targeted Delivery of Antisense Oligonucleotides to Pancreatic Beta Cell.

The extra hepatic delivery of antisense oligonucleotides (ASOs) remains a challenge and hampers the widespread application of this powerful class of therapeutic agents. In that regard, pancreatic beta cells are a particularly attractive but challenging cell type because of their pivotal role in diabetes and the fact that they are refractory to uptake of unconjugated ASOs. To circumvent this, we have expanded our understanding of the structure activity relationship of ASOs conjugated to Glucagon Like Peptide 1 Receptor (GLP1R) agonist peptide ligands. We demonstrate the key role of the linker chemistry and its optimization to design maleimide based conjugates with improved in vivo efficacy. In addition, truncation studies and scoping of a diverse set of GLP1R agonists proved fruitful to identify additional targeting ligands efficacious in vivo including native hGLP1(7-36)NH2. Variation of the carrier peptide also shed some light on the dramatic impact of subtle sequence differences on the corresponding ASO conjugate performance in vivo, an area which clearly warrant further investigations. We have confirmed the remarkable potential of GLP1R agonist conjugation for the delivery of ASOs to pancreatic beta cell by effectively knocking down islet amyloid polypeptide (IAPP) mRNA, a potential proapoptotic target, in mice.

Cloning and characterization of the porcine gastrin/cholecystokinin type 2 receptor.

The gastrointestinal hormone cholecystokinin (CCK) regulates digestive processes and satiety in addition to centrally mediated effects on nociception and anxiety. CCK signals through two seven-trans-membrane receptors named the CCK-1 receptor and the CCK-2 receptor. The expression pattern and biological effects mediated by the CCK-1 and CCK-2 receptors are highly divergent. The pig is a widely used preclinical animal model in medical research, but up until recently, the porcine CCK-2 receptor was described as a pseudogene in the publicly available genomic sequence databases. Thus, it was challenging to interpret data from this animal model in studies of CCK biology and pharmacology. Here we describe an in silico prediction of the porcine CCK-2 receptor and the subsequent cloning, expression, and in vitro pharmacological characterization. We find a high degree of sequence homology with the human orthologue as well as CCK-2 receptors of other major species used in pre-clinical research. We also show that the endogenous ligands CCK-8 and Gastrin-17 bind and activate the porcine CCK-2 receptor with similar affinities and potencies as seen for the human CCK-2 receptor. We conclude that the pig has a functional CCK-2 receptor which is highly comparable to the human orthologue and therefore the pig qualifies as a valid preclinical model for the study of human CCK biology and pharmacology.