(116bc) In Vitro Characterization of IL-10-Pdna Microparticles for Therapeutic Administration

Use of Interleukin-10-plasmid-DNA as gene therapy alleviates chronic neuropathic pain. Incorporating this DNA into biodegradable microparticles provides a sustained release of DNA into the tissue and reduces required injection requirements. These microparticles, however, have not been fully characterized as of yet. Additionally, positively charged containing PEI microparticles have better encapsulation efficiency and higher transfection rates, but are documented to be much more cytotoxic than standard microparticles. It is desired to design positively charged microparticles that have greater transfection rates while retaining a lower cytotoxicity. The standard microparticles were found not to be toxic to any of the tested cell lines. However, the microparticles were not able to transfect cells regardless of additive used. The DNA was extracted from the microparticles and the bioactivity of the DNA was tested, inducing a strong production of interlukin-10. Cellular internalization of the microparticles was investigated and was found that RAW cells internalize the microparticles very well, but HEK-293 cells do not. It was also found that cell internalization for RAW reaches a maximum between 4 to 6 hours of incubation and that the presence of lipopolysaccharide does not affect internalization. Positively charged particles were designed using various methods. Particles containing CTAB were successfully produced, but the formation protocol required more optimization before encapsulating viable DNA into these microparticles will be feasible. Poly-lysine coated microparticles were successfully designed and displayed a strong positive charge. These microparticles displayed little difference in cytotoxicity when compared to standard microparticles. However, the poly-lysine microparticles did not transfect any cell line. More optimization and characterization is required for the poly-lysine microparticles to determine why transfection failed. It is also suggested to test how long poly-lysine remains on the microparticle surface as well as testing if the proteins within the serum used is interacting with the microparticles.