Precise Assembly of Synthetic Carriers of siRNA through a Series of Interlocked Thermodynamically Self‐Regulated Processes

Abstract

A synthetic carrier of small interfering RNA (siRNA) with customizable size, pH-responsive degradability and optimized surface population of cell-targeting agents is constructed precisely through a series of interlocked thermodynamically self-regulated processes. This system consists of a unimolecular polyplex core formed from each individual molecule of networked cationic polymer and a multi-functional shell assembled from a rationally designed triblock copolymer. The core-forming polymer of defined size is synthesized via Zeta potential-regulated condensation of branched and linear multi-amine bearing oligomers through pH-responding imidazole conjugated imine linkages. The shell-forming copolymer consists of a multi-carboxyl saccharide end block to guide to the surface assembly, a hydrophobic central block to form an encapsulating layer, and a poly(ethylene glycol) end block equipped with a highly selective active ester for "hooking" various cell-targeting agents. Animal assays confirm the flexibility and convenience of this system to equip with selected functional components for systemic delivery of siRNA.

Rationally designed synthetic vectors for therapeutic nucleic acid delivery against human cytomegalovirus infection

“…We synthesized networked cationic polymers of various molecular diameters in the range of 100−400 nm through the condensation of spermine imidazole-4, 5-aldehyde oligomer, and branched polyethyleneimine-800 in an aqueous medium at pH 9 and in the presence of NaCl at respective concentrations (Figure 1A). 14 On the basis of our previous work in formulating polywraplex, we used a unique method to covalently conjugate an amino group bearing the targeting molecule with the rationally designed F 5 C 6 -O − CO − PEGpolycaprolactone (PCL)-saccharide-(COO − ) n block copolymer (Figure 1B). 12,14 The successful synthesis of the polymer was confirmed by 1 H NMR (Figures 1 and 2).…”

“…14 On the basis of our previous work in formulating polywraplex, we used a unique method to covalently conjugate an amino group bearing the targeting molecule with the rationally designed F 5 C 6 -O − CO − PEGpolycaprolactone (PCL)-saccharide-(COO − ) n block copolymer (Figure 1B). 12,14 The successful synthesis of the polymer was confirmed by 1 H NMR (Figures 1 and 2). For the treatment of HCMV infection, a mutant chemokine molecule, CX 3 CL 1 , was selected and modified by conjugating with lysine to generate an extra amino group at its C terminus (the amino group at N terminus will be protected to avoid losing its bioactivity) (Figure 2).…”

“…8,10,13 We further assembled this polymer to a network structure of designable size through a unique Zeta potential regulated condensation by which small interfering RNA (siRNA) may be packed inside a single molecule of the polymer via electrostatic interactions to form size-defined carrier particles. 14 Finally, we developed a selfassembly process to enable a rationally designed tri-block copolymer to align on the surface of particle to form a functional shell to encapsulate the nucleic acid core and immobilize cell-targeting molecules in optimized surface density. 14 The shell-forming triblock copolymer possesses a multi-carboxyl-saccharide block to guide surface assembly, a hydrophobic central block forms a layer to prevent pre-phagocytic leakage of small RNAs, and a polyethylene glycol (PEG) end-block forms a steric stabilization outer-layer.…”

“…14 Finally, we developed a selfassembly process to enable a rationally designed tri-block copolymer to align on the surface of particle to form a functional shell to encapsulate the nucleic acid core and immobilize cell-targeting molecules in optimized surface density. 14 The shell-forming triblock copolymer possesses a multi-carboxyl-saccharide block to guide surface assembly, a hydrophobic central block forms a layer to prevent pre-phagocytic leakage of small RNAs, and a polyethylene glycol (PEG) end-block forms a steric stabilization outer-layer. The targeting agents may be conjugated to the tri-block copolymer through the selective reaction with the distal end of the PEG block and be mixed with the tri-block copolymer without the modification with desired ratio before the shell assembling.…”

“…In this work, we used the universal carrier system developed in our previous study 12,14 to encapsulate both siRNA and small complementary RNA (scRNA), which target the major immediateearly genes of HCMV. HCMV has two major immediate-early genes, IE1 and IE2.…”