title: Next generation carbon nanoparticles for efficient gene therapy.
abstract: In a pursuit to develop a commercially exploitable and traceable gene delivery vehicle, here, we develop next generation carbon nanoparticle-DNA complex (CNPLex). CNPLexes were used to transfect green fluorescent protein (GFP) reporter gene containing plasmid DNA (pDNA) pEGFP-N1 targeting breast cancer cells MCF-7 and MDA-MB231. Prepared CNPs were optimized for particle size, surface potential, polymer surface decoration, absorbance efficiency, fluorescence efficiency, IR spectroscopic signatures, and DNA loading and release efficiencies. Rigorous biophysical methods were employed to determine the variations in physicochemical properties of CNPs after surface decoration with polymers followed by complexation with pDNA. Optimized CNPLexes were used to deliver pEGFP-N1 plasmid and efficiency of GFP was followed by fluorescence microscopy and quantified by flow assisted cell sorting. Lipofectamine2000 was used as positive control according to manufacturer's protocol and found to be comparative in transfection efficiency with one of our novel formulations. Further evaluation of cell toxicity and cell viability was performed by LDH activity and MTT assay, respectively. It was found that cell toxicity furnished by polymer decorated carbon nanoparticles was significantly low compared to the parent polymer (polyethylenimine, PEI). Similarly cell viability was found to be much higher with CNPLexes compared to PEI alone. This established the developed particles as better transfecting agents for reporter gene plasmid pEGFP-N1 compared to PEI and showed similar efficacy to one of the best known commercial transfection agents Liofectamine2000 in breast cancer cells MCF-7 and MDA-MB231.
authors.affiliation: Department of Bioengineering, University of Illinois Urbana-Champaign , Urbana, Illinois 61801, United States.
authors.forename: Santosh K
authors.initials: SK
authors.lastname: Misra
authors.forename: Ayako
authors.initials: A
authors.lastname: Ohoka
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Kolmodin
authors.forename: Dipanjan
authors.initials: D
authors.lastname: Pan
mesh_terms: D002244:Carbon / Q000737:chemistry*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics / Q000502:physiology; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D061986:MCF-7 Cells; D053758:Nanoparticles / Q000737:chemistry*; D010957:Plasmids / Q000235:genetics; D014162:Transfection
chemical_list: D049452:Green Fluorescent Proteins; D002244:Carbon
keywords: breast cancer; carbon particles; gene delivery; gene therapy; nanometer sized agents