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

Abstract

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)NH 2 . 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.

Efficient Synthesis of 2’‐O‐Methoxyethyl Oligonucleotide‐Cationic Peptide Conjugates

“…These bifunctional molecules are generally considered difficult to synthesize which, at least partly explains their lack of development to date. Indeed, most advanced CPPs are weakly charged [7a] or combined with PMO chemistry. A significant finding of the work is that highly cationic CPPs can be conjugated to 2’‐MOE PS oligonucleotides and isolated in good yields, provided that denaturing conditions are used for conjugation and purification steps.…”

Innovative developments and emerging technologies in RNA therapeutics

“…Recent synthetic improvements have been reported [ 30 , 31 ], but it is no surprise that the most advanced peptides for delivery of PS ASOs are weakly charged (e.g. GLP [ 32 ] and neurotensin peptides). Antibodies (Abs) are another class of ligands with great potential for ASO delivery.…”

“…Despite the now firmly established success of ASO therapies, challenges related to delivery remain, and further research efforts should be undertaken to develop structural modifications that optimize their therapeutic index [ 15 ] and to understand the cellular mechanisms responsible for intracellular trafficking. It is well known that a number of tissues and biological barriers are refractory to ssON uptake, but novel, tailored peptide [ 32 ] or antibody [ 33 ] conjugates may lead to significant improvements. It remains to be seen whether novel modifications such as stereodefined PS bonds or constrained ribose chemistries will succeed in the clinic and lead to approved drugs.…”

Kidney Effects by Alternative Classes of Medicines in Patients and Relationship to Effects in Nonclinical Toxicity Studies

“…Specific organ and tissue targeting has been achieved via GalNAc conjugation and glucagon like peptide 1 receptor (GLP1r) agonist conjugation to selectively target liver and pancreas, respectively. 50,51 In contrast, double-stranded siRNAs, represented by the recently approved treatment patisiran, are 20- to 25-nucleotide-long duplexes that load into the RNA-induced silencing complex (RISC) for mRNA inhibition. 52 They are cleared rapidly in urine if delivered as unconjugated molecules, and therefore must either be formulated in lipids or nanoparticles, or be chemically modified or conjugated to cell surface ligands (e.g., GalNAc) for effective tissue delivery.…”