Biodegradable Polymeric Materials for Gene and Drug Delivery
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In the past two decades, gene therapy has been recognized as a powerful tool in cancer treatment as the genetic links to tumor growth have been identified and studied. For instance, small interfering RNAs (siRNAs) can induce down-regulation and knockdown expression of mutated genes and overexpressed proteins, respectively; as a result, RNA interference (RNAi) using siRNAs as therapeutic gene materials has been developed. Unfortunately, therapeutic genes are poorly internalized and are susceptible to degradation by ubiquitous nucleases, and there is serious biosafety concern on viral vectors. Thus, the development of safe and efficient non-viral gene therapy vectors addressing the aforementioned delivery problems is of high importance. Based on the practical needs of non-viral vectors, well-defined tertiary amine-functionalized cationic polylactides (CPLAs) were synthesized by thiol-ene click functionalization of an allyl-functionalized polylactide (APLA). They were further utilized for the delivery of interleukin-8 (IL-8) siRNA to prostate cancer cells (PC3 cells), via the formation of CPLA-IL-8 siRNA nanocomplexes by electrostatic interaction. The CPLAs possess high hydrolytic degradability and low cytotoxicity. The CPLA-IL-8 siRNA nanocomplexes are readily taken up by PC3 cells, resulting in statistically significant IL-8 gene silencing. It was determined that the degradability, as well as transfection efficiency of CPLAs, positively correlate with the amine mol% of CPLAs. In addition to siRNA delivery, CPLAs were successfully used to deliver plasmid DNA (pDNA) to two physiological distinct cell lines (macrophage and fibroblast) via the formation of CPLA/pDNA nanocomplexes. Biophysical characterization of charge densities at various CPLA/pDNA weight ratios revealed a positive correlation between surface charge and gene delivery. To be more relevant for clinical applications, well-defined poly(ethylene glycol)- block -cationic polylactides (PEG- b -CPLAs) functionalized with tertiary amine-based cationic groups were synthesized by thiol-ene functionalization of a PEG- b -APLA. Subsequently, the application of PEG- b -CPLAs as biodegradable vectors for the pDNA delivery was investigated. Via the formation of PEG- b -CPLA/pDNA nanocomplexes by spontaneous electrostatic interaction, pDNAs encoding luciferase or enhanced green fluorescent protein were successfully delivered to four physiologically distinct cell lines (including macrophage, fibroblast, epithelial, and stem cell). Biophysical characterization of charge densities of nanocomplexes at various polymer:pDNA weight ratios revealed a positive correlation between surface charge and gene delivery. With PEG shielding layers, these nanocomplexes exhibited much higher serum resistance as compared to positive controls using an in vitro serum gene delivery inhibition assay. Recently, the simultaneous delivery of small molecular drug with gene via a single platform has emerged as a novel strategy to combat cancer diseases. Cross-linked biodegradable CPLA nanocapasules (CPLA NCs) were synthesized by miniemulsion interfacial thiol-ene cross-linking of CPLA. CPLA NCs exhibit high hydrolytic degradability without noticeable cytotoxicity. With size ranging from 20 to 50 nm, these NCs can avoid fast clearance in a mouse model. Multidrug resistance (MDR) by cancer cells remains a major challenge in chemotherapy, in which P-glycoprotein (Pgp) plays a key role by enhanced efflux of anticancer drug. Through the encapusulation by NCs, Doxorubicin (Dox) can successfully bypass Pgp-mediated efflux pump, thereby resulting in increased intracellular drug concentration and enhanced therapeutic efficacy. Relative to free Dox, Dox-loaded CPLA NCs exhibited a significantly enhanced anti-proliferation effect in MCF7/ADR cells (a stable MDR model). Additionally, these NCs can in situ encapsulate Dox with inner cavities and bind IL-8 siRNA with cationic shells, thereby enabling the codelivery of drugs with genes. In vitro studies showed that the NCs loaded with Dox, IL-8 siRNA and both agents can be readily taken up by PC3 prostate cancer cells, resulting in significant chemotherapeutic effect and/or IL-8 gene silencing. In addition to CPLA-based nanocapsules, we are interested in synthesizing chitosan-based NCs due to the low synthetic cost and facile preparation procedures. Well-defined chitosan nanocapsules (CSNCs) with tunable sizes were synthesized through interfacial cross-linking of N -maleoyl-functionalized chitosan (MCS) in miniemulsions, and their application in the delivery of Dox was investigated. Thiol-ene interfacial cross-linking was conducted in oil-in-water miniemulsions at room temperature under UV irradiation for 1 h, using MCS as both surfactant and precursor polymer, 1,4-butanediol bis(3-mercapto-propionate) as cross-linker and D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) as co-surfactant. With the increase of co-surfactant concentration in the reaction systems, the sizes of the resulting CSNCs decreased steadily. Dox-loaded CSNCs were readily prepared by in situ encapsulation of Dox during miniemulsion cross-linking. With acid-labile β-thiopropionate cross-linkages, the Dox-loaded CSNCs demonstrated faster Dox release rate under acidic conditions. Relative to free Dox, Dox-loaded CSNCs exhibited enhanced cytotoxicity towards MCF-7 breast cancer cells while the empty CSNCs did not show noticeable cytotoxicity. Effective delivery of Dox into the cancer cells via Dox-loaded CSNCs was also observed. Overall, the results suggest promising potential applications of these novel CSNCs as nanoscopic scaffolds for therapeutic delivery.