Studies of the aqueous RAFT polymerization of methacrylamido-monomers and the application of their polymers in the protection and delivery of novel siRNA-based gene silencing agents

The extension of the reversible addition-fragmentation chain transfer (RAFT) process to the controlled polymerization of alpha-substituted, methacrylamido-monomers in aqueous media has been investigated and employed in the preparation of homo- and block-copolymer systems for applications in the effective protection and stabilization of a novel RNA-based gene silencing agent.; Building on the previously reported controlled polymerization of a cationic, methacrylamido, monomer, namely N-[3-(dimethylamino)propyl]methacrylamide (DMAPMA) (65), the first study involves the controlled polymerization of the neutral, hydrophilic monomer, methacrylamide (MAM) (74), and its subsequent chain-extension polymerizations with MAM and acrylamide (AM) (70). Homopolymers of poly(MAM) (PMAM) were obtained using 4-cyanopentanoic acid dithiobenzoate (77) and sodium 4,4'azobis(4-cyanopentanoic acid) (V-501) as the RAFT chain transfer agent (CTA) and primary initiating species, respectively, under buffered aqueous conditions at 70°C.; The second study concerns the synthesis and characterization of homo- and diblock copolymers with a neutral, hydroxy-functional methacrylamido monomer, namely, N-(2-hydroxypropyl)methacrylamide (HPMA), and the previously reported cationic methacrylamido monomer, DMAPMA. HPMA was chosen as the first monomer because its homopolymer is permanently hydrophilic and is well known for its biocompatible properties in drug delivery applications. To test the blocking order dependence of HPMA and DMAPMA a DMAPMA macroCTA was prepared under identical conditions and employed in chain-extension experiments to yield well defined block copolymers with minimal formation of homopolymer impurity.; The third study concerns the formation of block ionomer complexes with a series of the above mentioned HPMA/DMAPMA block copolymers and a 43-nucleotide single-stranded segment of ribonucleic acid (RNA) known as short interfering RNA (siRNA). This siRNA represents the new state-of-the-art in the areas of gene silencing for anticancer and gene therapies. The specific sequence employed is able to silence the gene in human cells that ultimately codes for the synthesis of Human RNA Polymerase II, an enzyme whose production is required for the cell to survive.