James Ramos

Discovery of Cationic Polymers for Non-Viral Gene Delivery Using Combinatorial Approaches

Gene therapy is an attractive treatment option for diseases of genetic origin, including several cancers and cardiovascular diseases. While viruses are effective vectors for delivering exogenous genes to cells, concerns related to insertional mutagenesis, immunogenicity, lack of tropism, decay and high production costs necessitate the discovery of non-viral methods. Significant efforts have been focused on cationic polymers as non-viral alternatives for gene delivery. Recent studies have employed combinatorial syntheses and parallel screening methods for enhancing the efficacy of gene delivery, biocompatibility of the delivery vehicle, and overcoming cellular level barriers as they relate to polymer-mediated transgene uptake, transport, transcription, and expression. This review summarizes and discusses recent advances in combinatorial syntheses and parallel screening of cationic polymer libraries for the discovery of efficient and safe gene delivery systems.

Generation of a focused poly(amino ether) library: polymer-mediated transgene delivery and gold-nanorod based theranostic systems.

A focused library of twenty-one cationic poly(amino ethers) was synthesized following ring-opening polymerization of two diglycidyl ethers by different oligoamines. The polymers were screened in parallel for plasmid DNA (pDNA) delivery, and transgene expression efficacies of individual polymers were compared to those of 25 kDa polyethylenimine (PEI), a current standard for polymer-mediated transgene delivery. Seven lead polymers that demonstrated higher transgene expression than PEI in pancreatic and prostate cancer cells lines were identified from the screen. All seven lead polymers showed highest transgene expression at a polymer:pDNA weight ratio of 5:1 in the MIA PaCa-2 pancreatic cancer cell line. Among the conditions studied, transgene expression efficacy correlated with minimal polymer cytotoxicity but not polyplex sizes. In addition, this study indicated that methylene spacing between amine centers in the monomers, amine content, and molecular weight of the polymers are all significant factors and should be considered when designing polymers for transgene delivery. A lead effective polymer was employed for coating gold nanorods, leading to theranostic nanoassemblies that possess combined transgene delivery and optical imaging capabilities, leading to potential theranostic systems.

Transgene delivery using poly(amino ether)-gold nanorod assemblies

Gold nanorods (GNRs) have emerged as promising nanomaterials for biosensing, imaging, photothermal treatment, and therapeutic delivery for several diseases, including cancer. We have generated poly(amino ether)‐functionalized gold nanorods (PAE‐GNRs) using a layer‐by‐layer deposition approach; polymers from a poly(amino ether) library recently synthesized in our laboratory were employed to generate the PAE‐GNR assemblies. PAE‐GNR assemblies demonstrate long‐term colloidal stability as well as the capacity to bind plasmid DNA by means of electrostatic interactions. Sub‐toxic concentrations of PAE‐GNRs were employed to deliver plasmid DNA to prostate cancer cells in vitro. PAE‐GNRs generated using 1,4C‐1,4Bis, a cationic polymer from our laboratory demonstrated significantly higher transgene expression and exhibited lower cytotoxicities when compared to similar assemblies generated using 25 kDa poly(ethylene imine) (PEI25k‐GNRs), a current standard for polymer‐mediated gene delivery. The roles of polyelectrolyte chemistry and zeta‐potential in determining transgene expression efficacies of PAE‐GNR assemblies were investigated. Our results indicate that stable and effective PAE‐GNR assemblies are a promising engineered platform for transgene delivery. PAE‐GNRs also have the potential to be used simultaneously for photothermal ablation, photothermally enhanced drug and gene delivery, and biological imaging, thus making them a powerful theranostic platform. Biotechnol. Bioeng. 2012; 109:1336–1346.

GOLD NANOPARTICLES IN CANCER IMAGING AND THERAPEUTICS

The use of nanomedicine in the war on cancer diseases has progressed significantly in the recent past. Liposomal- and albumin-based chemotherapeutic agents as well as tumor contrast agents (e.g. Gd-DTPA, ferumoxides) have received FDA approval for human clinical use, while many other agents are in different phases of pre-clinical investigation and clinical trials. Plasmonic gold nanoparticles hold great promise as potential theranostic devices for detection and ablation of cancer diseases. This review discusses recent progress in the imaging, photothermal therapy, and nucleic acid/drug delivery using gold nanoparticles (spheres, shells, rods, cages) in vitro and in vivo. Issues relating to toxicity, biocompatibility, biodistribution, cellular uptake, and targeting efficiency are also discussed.

Parallel synthesis of poly(amino ether)-templated plasmonic nanoparticles for transgene delivery.

Plasmonic nanoparticles have been increasingly investigated for numerous applications in medicine, sensing, and catalysis. In particular, gold nanoparticles have been investigated for separations, sensing, drug/nucleic acid delivery, and bioimaging. In addition, silver nanoparticles demonstrate antibacterial activity, resulting in potential application in treatments against microbial infections, burns, diabetic skin ulcers, and medical devices. Here, we describe the facile, parallel synthesis of both gold and silver nanoparticles using a small set of poly(amino ethers), or PAEs, derived from linear polyamines, under ambient conditions and in absence of additional reagents. The kinetics of nanoparticle formation were dependent on PAE concentration and chemical composition. In addition, yields were significantly greater in case of PAEs when compared to 25 kDa poly(ethylene imine), which was used as a standard catonic polymer. Ultraviolet radiation enhanced the kinetics and the yield of both gold and silver nanoparticles, likely by means of a coreduction effect. PAE-templated gold nanoparticles demonstrated the ability to deliver plasmid DNA, resulting in transgene expression, in 22Rv1 human prostate cancer and MB49 murine bladder cancer cell lines. Taken together, our results indicate that chemically diverse poly(amino ethers) can be employed for rapidly templating the formation of metal nanoparticles under ambient conditions. The simplicity of synthesis and chemical diversity make PAE-templated nanoparticles useful tools for several applications in biotechnology, including nucleic acid delivery.

Delivery of plasmid DNA to mammalian cells using polymer-gold nanorod assemblies

Functionalized and surface-modified gold nanorods (GNRs) have emerged as promising vehicles for the delivery of several therapeutic agents. Ease of functionalization, increased stability, biocompatibility, and size-dependent plasmonic properties make gold nanorods attractive in sensing, imaging, and delivery to different cellular types. Here, we demonstrate the use of polyelectrolyte-coated GNRs (PE-GNRs) for delivering plasmid DNA to mammalian cells for transgene expression.

Comparative Investigation of Polymeric and Nanoparticle Vehicles for Transgene Delivery

Effective design of nanoparticle systems can have a significant impact on therapeutic delivery. Physicochemical factors including size, shape and surface chemistry of nanoparticles can play a significant role in determining the efficacy of drug and gene delivery to cells. Polymeric as well as inorganic nanoparticle systems have been investigated as vehicles for nonviral gene delivery of transgenes. However, a head-to-head comparison of these different systems is largely lacking. In this study, we compare three related delivery systems, polymer, polymer-templated gold nanospheres and polymer-coated gold nanorods, for their respective in vitro transgene expression efficacies. Significant differences were seen in the hydrodynamic diameter and zeta potential for each of these different vehicles. Nevertheless, transgene (luciferase) expression efficacies and cytotoxicities were found to be similar for these three vehicles under different conditions. Our results indicate that polymeric systems can be used for s...