Marianna Foldvari

Carbon nanotubes as functional excipients for nanomedicines: II. Drug delivery and biocompatibility issues

Carbon nanotubes (CNTs) have potential novel application in nanomedicine as biocompatible and supportive substrates, and as pharmaceutical excipients for creating versatile drug delivery systems. In the second part of this two-part review we focus on the application of CNTs as potential drug delivery systems via chemical functionalization of CNTs for exterior binding of therapeutic and biologically relevant molecules, and via encapsulation of these molecules within the inner cavities of CNTs. We review experimental results of CNT-mediated delivery of small molecules, DNA, proteins, and vaccines, and the potential of CNTs as matrices to support and stimulate neural growth. Last, we examine some toxicological and biocompatibility issues related to the use of CNTs as pharmaceutical excipients and discuss attributes that affect toxicity, such as structure (single-walled vs. multi-walled CNTs), length and aspect ratio, surface area, degree of aggregation, extent of oxidation, surface topology, bound functional group(s), and method of manufacturing.

Electroporation improves the efficacy of DNA vaccines in large animals.

It is generally recognized that DNA vaccines are often less effective in large animals than in mice. One possible reason for this reduced effectiveness may be transfection efficiency and the low level of expression elicited by plasmid vectors in large animals. A possible way to improve plasmid gene expression in vivo is electroporation. To determine whether we could enhance immune responses in pigs by electroporation, we used plasmids encoding two different genes (bovine herpesvirus glycoprotein D (gD) and hepatitis B surface antigen (HBsAg)) and two different electrodes, a single-needle electrode and a six-needle electrode. Electroporation significantly enhanced immune responses to both antigens. In addition, we demonstrated that co-administration of plasmids coding for two different antigens (pgD and pHBsAg) did not result in significant interference between the plasmids. We also incorporated a DNA prime/protein boost strategy to examine the effect of DNA priming with electroporation on the immune response after a protein boost.

Cutaneous vaccination : The skin as an immunologically active tissue and the challenge of antigen delivery

Vaccination is one of the major achievements of modern medicine. As a result of vaccination, diseases such as polio and measles have been controlled and small pox has been eradicated. However, despite these successes there are still many microbial diseases that cause tremendous suffering because there is no vaccine or the vaccines available are inadequate. In addition, even if vaccines were available for all infectious diseases there is no guarantee that people would use them routinely. One of the major impediments to ensuring vaccine efficacy and compliance is that of delivery. Presently most vaccines are given by intramuscular administration. Unfortunately this is often traumatic, especially in infants. Thus, if it was possible to replace intramuscular immunization by mucosal (oral/intranasal) or transdermal delivery it may be possible to both enhance mucosal immunity as well as improve overall compliance rates. The transdermal route has been used by the pharmaceutical industry for the delivery of various low molecular weight drugs. Some of the approaches used for smaller compounds may also have potential for delivery of either protein or polynucleotide vaccines. However, there is a greater challenge to delivering large molecular weight molecules through the skin due to size, charge and other physicochemical properties. This review will describe the recent advances that have been made in dermal and topical delivery as related to vaccines.

Non-Viral Nucleic Acid Delivery: Key Challenges and Future Directions

Gene therapy holds the promise of correcting a genetic defect. It can be achieved with the introduction of a normal wild-type transgene into specific cells of the patient where the endogenous gene is underexpressing or by the introduction of a therapeutic agent, such as, antisense oligonucleotides (AON) or small interfering RNA (siRNA) to inhibit transcription and/or translation of an overexpressing endogenous gene or a cancer causing oncogene. Gene therapy has been utilized for vaccination and for the treatment of several diseases, such as, cancer, viral infections and dermatological diseases. However, there are many hurdles to overcome in developing effective gene-based therapeutics, including cellular barriers, enzymatic degradation and rapid clearance after administration. Successful transfer of nucleic acids (e.g. plasmid DNA, AON, siRNA, small hairpin RNA and micro RNA) into cells usually relies on the use of efficient carriers, commonly viral or non-viral vectors. Presently, viral vectors are more efficient than non-viral systems. However, immunogenicity, inflammatory reactions and problems associated with scale-up limit their clinical use. The ideal carriers for gene delivery should be safe and yet ensure that the DNA/RNA survives the extra- and intracellular environment and efficiently transfer to the appropriate cellular compartments. This review discusses some of the strategies that have been employed to overcome the barriers towards successful gene delivery.

Gemini surfactants: a new family of building blocks for non-viral gene delivery systems.

Gemini surfactants provide a significant opportunity in the development of new non-viral delivery systems designed for gene therapy applications. This review summarizes the wide range of gemini surfactant structures that have been employed for DNA transfection in vitro. A general observation is that those structures capable of inducing a wide variety of polymorphic structures (lamellar, hexagonal, or cubic phases) demonstrate higher transfection efficiencies. Those compounds whose structures result in pH-dependent changes in aggregate structure similarly show higher levels of transfection. In vivo transfection using gemini surfactants has been demonstrated in only three cases, and in a recent study the transfection was linked to a specific therapeutic response.

In vivo cutaneous interferon‐γ gene delivery using novel dicationic (gemini) surfactant–plasmid complexes

Localized scleroderma (morphea and linear scleroderma) is a connective tissue disease, accompanied by excessive proliferation and deposition of collagen within the skin, inflammation, vasculopathy and a deranged immune system. Interferon γ (IFNγ), an inhibitor of collagen synthesis and an immunomodulator, could be a potential therapeutic agent if it could be delivered into or expressed locally in affected skin in a non‐invasive manner. In this study, the feasibility of topical delivery of the IFNγ gene and expression of IFNγ were investigated in mice.

Non-invasive administration of drugs through the skin: challenges in delivery system design

Vehicles designed to enhance drug delivery through the skin must incorporate specific elements that improve the ability of the delivery system to overcome the barrier posed by the stratum corneum. This review discusses several chemical penetration enhancers that have been investigated as potential tools to increase drug flux. In addition, lipid-based delivery systems offer an attractive alternative to traditional drug vehicles. The relationship between liposome composition and drug permeation is discussed, in addition to the possible mechanism of action of lipid vesicle-mediated drug delivery.

Structural and transfection properties of amine-substituted gemini surfactant-based nanoparticles

Increases in DNA transfection efficiencies for non‐viral vectors can be achieved through rational design of novel cationic building blocks. Based on previous results examining DNA condensation by polyamines, novel gemini surfactants have been designed that incorporate aza or imino substituents within the spacer group in order to increase interactions with DNA and potentially improve their DNA transfection ability.

Advancing nonviral gene delivery: lipid- and surfactant-based nanoparticle design strategies

Gene therapy is a technique utilized to treat diseases caused by missing, defective or overexpressing genes. Although viral vectors transfect cells efficiently, risks associated with their use limit their clinical applications. Nonviral delivery systems are safer, easier to manufacture, more versatile and cost effective. However, their transfection efficiency lags behind that of viral vectors. Many groups have dedicated considerable effort to improve the efficiency of nonviral gene delivery systems and are investigating complexes composed of DNA and soft materials such as lipids, polymers, peptides, dendrimers and gemini surfactants. The bottom-up approach in the design of these nanoparticles combines components essential for high levels of transfection, biocompatibility and tissue-targeting ability. This article provides an overview of the strategies employed to improve in vitro and in vivo transfection, focusing on the use of cationic lipids and surfactants as building blocks for nonviral gene delivery systems.

Enhanced gene expression in epithelial cells transfected with amino acid-substituted gemini nanoparticles

Gemini surfactants are versatile gene delivery agents because of their ability to bind and compact DNA and their low cellular toxicity. Through modification of the alkyl tail length and the chemical nature of the spacer, new compounds can be generated with the potential to improve the efficiency of gene delivery. Amino acid (glycine and lysine) and dipeptide (glycyl-lysine and lysyl-lysine) substituted spacers of gemini surfactants were synthesized, and their efficiency of gene delivery was assessed in epithelial cells for topical cutaneous and mucosal applications. Three different epithelial cell lines, COS-7, PAM212 and Sf 1Ep cells, were transfected with plasmid DNA encoding for interferon gamma and green fluorescent protein complexed with the amino acid-substituted gemini compounds in the presence of 1,2 dioleyl-sn-glycero-phosphatidyl-ethanolamine as a helper lipid. Gene expression was quantified by ELISA. Size, zeta potential and circular dichroism measurements were used to characterize the plasmid-gemini (PG) and plasmid-gemini surfactant-helper lipid (PGL) complexes. Gene expression was found to increase up to 72h and then declined by the 7th day. In general, the glycine-substituted surfactant showed consistently high gene expression in all three cell lines. Results of physicochemical and spectroscopic studies of the complexes indicate that substitution of the gemini spacer does not interfere with compaction of the DNA. The superior performance of these spacer-substituted gemini surfactants might be attributed to their better biocompatibility compared to the surfactants possessing unsubstituted spacers.