Jeffrey R. Kovacs

Characterization of particles fabricated with poly(D,L-lactic-co-glycolic acid) and an ornithine–histidine peptide as carriers of oligodeoxynucleotide for delivery into primary dendritic cells

We report the formulation of particles to deliver oligodeoxynucleotides (ODN) into primary murine dendritic cells (DCs), the most potent antigen-presenting cells (APCs) known, using poly(D,L-lactic-co-glycolic acid) (PLGA) and a small cationic peptide. PLGA polymer and the ODN were fabricated into nano-sized spherical particles with the aid of O10H6 (O = ornithine, H = histidine). We have previously determined that O10H6 condenses DNA and is less toxic to DCs than a similar lysine-based peptide. The colloidal particles are stabilized by negative surface potentials. The peptide and the ODN can be detected in the fabricated particles with reflectance infrared spectroscopy. Significant ODN uptake can be observed in DCs exposed to the particles. Confocal imaging studies reveal that ODN can be internalized and escape from lysosomes in DCs. Taken together, these data suggest that combining PLGA and O10H6 is a feasible method to generate ODN-containing nano-sized particles for applications in DCs.

Characterization of Nickel-Decorated PLGA Particles Anchored with a His-tagged Polycation

The pharmacological impact of oligodeoxynucleotides (ODN) as transcription factors decoys (anti-sense) depends on the efficiency of cellular uptake. In this study, we sought to generate nickel-decorated particles to facilitate the entry of ODN into dendritic cells (DCs), the primary instigators of immune responses. Nickel ions were incorporated into the matrix of poly(D,L-lactide-co-glycolide) (PLGA) particles using the metal chelating lipid DOGS-NTA-Ni. Submicrometer-sized PLGA particles containing nickel ions (PLGA-Ni) were formed using a double-emulsion solvent evaporation method. Infrared spectroscopy provided chemical proofs of nickel incorporation into the PLGA matrix. Binding of the polycation O10H6 raised the surface potential of PLGA-Ni from −17 mV to +13 mV. This change was partially reserved by the presence of free imidazole, suggesting the binding was mediated by nickel-histidine coordination. When compared to PLGA particles without nickel, ODN bound to O10H6-coated PLGA-Ni particles exhibited enhanced capacity to accumulate in DCs cultured in vitro. DCs exhibited cellular stress after exposure to PLGA-Ni complexed with O10H6 and DNA, but this effect can be prevented by serum and was reversed overnight. These data suggest PLGA-Ni should be further explored as a nucleic acid carrier in the context of anti-sense gene down-regulation.

The Use of Non-Viral Nucleic Acids Carriers for the Modulation of Leukocytes

Induction of drug-free permanent organ accommodation is the ultimate goal of transplant therapy. Current pharmacological agents, however, are non-specific in their actions and generally do not confer immunological tolerance. Inhibitors of T cell receptor signaling (tacrolimus and cyclosporine A) represent the mainstay in transplant management. These agents exert their therapeutic effects by dampening the activities of all T cells. Chronic exposure to these agents increases the risk of developing opportunistic infections and malignancies. Given that more than 20,000 Americans undergo organ transplantations each year, there is an urgent need to develop specific therapies to mitigate graft rejection and create conditions conducive for long-term transplant accommodation.Copyright