Tailored Polymers for Delivery of Gene Medicines

点击翻译按钮获取中文摘要

Molecular therapy offers the best promise of a cure for a wide range of genetic disorders. The ability to permanently ‘fix’ defective genes is desirable over current drug therapy, which typically do little to eradicate the underlying cause of the disease. Successful molecular therapy involves a precise knowledge of a gene defect in a disease, followed by delivery of nucleic acids to modulate gene expression. It is possible to enhance gene expression by delivering plasmid DNAs (pDNA) or silence gene expression by delivering short interfering RNA (siRNA). But therapeutic success is absolutely dependent on the use of gene carriers since nucleic acids cannot be internalized by cells. Cationic polymers are safe delivery vehicles for this purpose but they suffer from low transfection efficiencies and high toxicities. This Discovery Project aims to create ‘engineered’ polymers that can self-assemble with nucleic acids to create nanoparticles ideal for cellular delivery. We have been engineering a class of amphiphilic polymers constructed from hydrophobic lipids and cationic polymers. Our work has led us to low molecular weight (0.6 – 2.0 kDa) polyethylenimines (PEI) as the optimal starting material. Hydrophobic moieties were found to create bridges among self-assembled nanoparticles, stabilizing the structure via lipid-lipid associations and enhancing delivery of the cargo into cells by as much as 10-fold. A specific issue not addressed in the previous funding period was controlled delivery and release of the cargo, which is needed for optimal functional outcomes. The proposed project is intended to address this issue. Our overall hypothesis is: amphiphilic polymers can be engineered from cationic polymers and lipids for controlled delivery and release of nucleic acids in cells, ultimately leading to improved functional outcomes mediated by nucleic acids. Structure-function relationships will be pursued to better understand limiting barriers for delivery of nucleic acids and approaches to overcome these barriers. Specific aims of this project are: Aim-1. To design a library of small molecular weight (<2 kDa) amphiphilic polymers with improved lipid substituents for endosomal escape. Our previous work explored a limited range of lipids and established preliminary information on functional features for endosomal escape. This effort will be expanded by exploring other lipid substituents with promising functional features. The relevant physicochemical properties of the polymer library will be characterized as well as cellular delivery of nucleic acids. Aim-2. To conjugate effective lipids to polymers via labile linkages. Using short polymeric backbones, the chosen lipids will be anchored onto polymer backbones via cleavable linkages. The stability of the polymers will be characterized as well as the intracellular trafficking of resultant nanoparticles. Aim-3. To determine functional performance of polymers for nucleic acid delivery. We anticipate the properties of self-assembled complexes formed between polymeric carriers and nucleic acids to dictate the functional outcomes. This aim will probe the functional outcomes in relevant cellular systems in vitro. The outcome of the proposed project will have significant impact on development of molecular therapies for human diseases. A foundation for polymer-guided delivery of genetic agents will be established by exploring the effectiveness of safer alternatives (i.e., polymers) to current delivery vehicles (i.e., viruses). While exploring molecular assembly into nanoparticles, functional materials for effective and safe application of molecular therapy will be developed as a result of successful completion of this project.