Chemoselective Peptide Modification via Photocatalytic Tryptophan ß-Position Conjugation.

Targeting tryptophan is a promising strategy to achieve high levels of selectivity for peptide or protein modification. A chemoselective peptide modification method via photocatalytic tryptophan ß-position conjugation has been discovered. This transformation has good substrate scope for both peptide and Michael acceptor, and has good chemoselectivity versus other amino acid residues. The endogenous peptides, glucagon and GLP-1 amide, were both successfully conjugated at the tryptophan ß-position. Insulin was studied as a nontryptophan control molecule, resulting in exclusive B-chain C-terminal-selective decarboxylative conjugation. This transformation provides a novel approach toward peptide modification to support the discovery of new therapeutic peptides, protein labeling and bioconjugation.

Novel endosomolytic poly(amido amine) polymer conjugates for systemic delivery of siRNA to hepatocytes in rodents and nonhuman primates.

The application of small interfering (si)RNAs as potential therapeutic agents requires safe and effective methods for their delivery to the cytoplasm of the target cells and tissues. Recent studies have shown significant progress in the development of targeting reagents that facilitate the recognition of, and siRNA delivery to, specific cell types. Among recently reported delivery approaches, polymers with amphipathic properties have been used to enable endosome escape and cytosolic delivery. Here, we describe a linear amphipathic poly(amido amine) polymer conjugate system for the efficient siRNA delivery in vitro and in vivo. This polymer contains a novel amine bearing bis-acrylamide monomer designed for increasing amine density, which resulted in substantial improvement in liver uptake and RNAi activity compared to our previously reported poly(amido amine disulfide) polymer.1 The activity for this liver targeted delivery system was demonstrated in rodents and nonhuman primates.

An in vivo evaluation of amphiphilic, biodegradable peptide copolymers as siRNA delivery agents.

A series of amphiphilic, biodegradable polypeptide copolymers were prepared for the delivery of siRNA (short interfering ribonucleic acid). The molecular weight (or polymer chain length) of the linear polymer was controlled by reaction stoichiometry for the 11.5, 17.2, and 24.6 kDa polypeptides, and the highest molecular weight polypeptide was prepared using a sequential addition method to obtain a polypeptide having a molecular weight of 38.6 kDa. These polymers were used to prepare polymer conjugate systems designed to target and deliver an apolipoprotein B (ApoB) siRNA to hepatocyte cells and to help delineate the effect of polymer molecular weight or polymer chain length on siRNA delivery in vivo. A clear trend in increasing potency was found with increasing molecular weight of the polymers examined (at a constant polymer:siRNA (w/w) ratio), with minimal toxicity found. Furthermore, the biodegradability of these polymer conjugates was examined and demonstrates the potential of these systems as siRNA delivery vectors.

Development of a liver-targeted siRNA delivery platform with a broad therapeutic window utilizing biodegradable polypeptide-based polymer conjugates.

The greatest challenge standing in the way of effective in vivo siRNA delivery is creating a delivery vehicle that mediates a high degree of efficacy with a broad therapeutic window. Key structure-activity relationships of a poly(amide) polymer conjugate siRNA delivery platform were explored to discover the optimized polymer parameters that yield the highest activity of mRNA knockdown in the liver. At the same time, the poly(amide) backbone of the polymers allowed for the metabolism and clearance of the polymer from the body very quickly, which was established using radiolabeled polymers to demonstrate the time course of biodistribution and excretion from the body. The fast degradation and clearance of the polymers provided for very low toxicity at efficacious doses, and the therapeutic window of this poly(amide)-based siRNA delivery platform was shown to be much broader than a comparable polymer platform. The results of this work illustrate that the poly(amide) platform has a promising future in the development of a siRNA-based drug approved for human use.

Endosomolytic bioreducible poly(amido amine disulfide) polymer conjugates for the in vivo systemic delivery of siRNA therapeutics.

Efficient siRNA delivery is dependent not only on the ability of the delivery vehicle to target a specific organ but also on its ability to enable siRNA entry into the cytoplasm of the target cells. Polymers with endosomolytic properties are increasingly being used as siRNA delivery vehicles due to their potential to facilitate endosomal escape and intracellular delivery. Addition of disulfide bonds in the backbone of these polymers was expected to provide degradability through reduction by glutathione in cytosol. This paper describes the synthesis of new endosomolytic bioreducible poly(amido amine disulfide) polymers whose lytic potential can be masked at physiological pH, but can be restored at acidic endosomal pH. These polymer conjugates gave good in vitro knockdown (KD) and did not demonstrate cytotoxicity in a MTS assay. Efficient mRNA KD for apolipoprotein B in mouse liver was observed with these polyconjugates following intravenous dosing.

Tetrahydro anthranilic acid as a surrogate for anthranilic acid: application to the discovery of potent niacin receptor agonists.

The design, synthesis, and biological activity of a series of cycloalkene acid-based niacin receptor agonists are described. This led to the discovery that tetrahydro anthranilic acid is an excellent surrogate for anthranilic acid. Several compounds were identified that were potent against the niacin receptor, had enhanced cytochrome P450 selectivity against subtypes CYP2C8 and CYP2C9, and improved oral exposure in mice.

Discovery of biaryl anthranilides as full agonists for the high affinity niacin receptor.

Biaryl anthranilides are reported as potent and selective full agonists for the high affinity niacin receptor GPR109A. The SAR presented outlines approaches to reduce serum shift and both CYPCYP2C8 and CYP2C9 liabilities, while improving PK and maintaining excellent receptor activity. Compound 2i exhibited good in vivo antilipolytic efficacy while providing a significantly improved therapeutic index over vasodilation (flushing) with respect to niacin in the mouse model.

Agonist lead identification for the high affinity niacin receptor GPR109a.

A strategy for lead identification of new agonists of GPR109a, starting from known compounds shown to activate the receptor, is described. Early compound triage led to the formulation of a binding hypothesis and eventually to our focus on a series of pyrazole acid derivatives. Further elaboration of these compounds provided a series of 5,5-fused pyrazoles to be used as lead compounds for further optimization.