Biophysical and Transfection Studies of Gemini/DOPE- pEGFP-C3 Plasmid DNA Lipoplexes

MÓNICA MUÑOZ-ÚBEDA; ANA LILIA BARRÁN-BERDÓN; MARÍA BELÉN SIERRA; CLARA AICART-RAMOS; JOYDEEP BISWAS; SANTOSH K. MISHRA; ELENA JUNQUERA; SANTANU BHATTACHARYA; EMILIO AICART

Porto

Congreso; 4th Iberian Meeting on Colloids and Interfaces; 2011

Biophysical and Transfection Studies of Geminii/DOPE- pEGFP-C3 Plasmid DNA Lipoplexes Monica Muñoz Úbeda,*´1 Ana Lilia Barran-Berd6n,1 Maria Belen Sierra,1Clara Aicart Ramos,2 Joydeep Biswas,3 Santosh K. Mishra,3 Elena Junquera,1 Santanu Bhattacharya,3 and Emilio Aicart1 Grupo Química Coloidal y Supramolecular, Dpto. Química Física I, Fac. Ciencias Químicas, Universidad Complutense de Madrid, 28040-Madrid, Spain 2Dpro. de Bioquímica y Biología Molecular I, Fac. Ciencias Químicas, Universidad Complutense de Madrid, 28040-Madrid, Spain ´ Indian Institute of Sciences, Department of Organic Chemistry, 560 012-Bangalore, India *monicamubeda@quim.ucm.es The interaction between DNA and cationic lipids, acting as non viral vehicles in gene therapy, is of great importance from biological and medical perspectives. Cationic liposomes forming lipoplexes (liposome/DNA complexes) have demonstrated their capability to condense and compact the negatively charged DNA. In this work, the influence of the synthesized gemini lipids, C16CnC16 (n = 2, 3, 5 and 12) on the compaction process of DNA has been analyzed. For that purpose, mixtures of four cationic gemini lipids and a zwitterionic helper lipid, the 1,2-dioleoyl-sn-glycero-3- phosphatidylethanolamine (DOPE), have been characterized in aqueous buffered solution (HEPES, pH = 7.4). Electrophoretic mobility/zeta potential and gel electrophoresis allow to determining the isoneutrality ratio, (L/D)f characteristic of each colloidal aggregate-DNA lipoplex. The interaction among DNA and the colloidal vector has been evaluated from gel red fluorescence spectroscopy intercalation assays. The lipoplex structures have been determined through small angle X-ray scattering (SAXS) experiments. Transfection efficiency, cytotoxicity and the ability to release DNA into the cell cytoplasm have been also analyzed. The whole information reveals that these gemini lipids efficiently compact DNA by means of a driven surface electrostatic interaction, as confirmed by zeta potential measurements. At moderate concentrations, SAXS experiments reveal that DNA intercalates into multilamellar structurated bilayers. In addition, the influence of the percentage of the cationic lipid (L+/(L++L0), and the charge ratio (CR) between the cationic lipid and DNA of the lipoplexes have been also analyzed. Results reveal that a short spacer (n = 2) in the gemini lipid yields a more efficient transfection, less toxicity and higher release of DNA into the cell cytoplasm, at mixed liposome compositions from 0.25 to 0.5. Acknowledgements: We thank the European Synchrotron Radiation Facility (ESRF) Beamline 16, Grenoble (France) for SAXS experiments. Bhattacharya and A. Bajaj, Chem. Commun. 2009, 4632-4656 [2] A. Rodriguez-Pulido, F. Ortega, E. Aicart, 0. Llorca and E. Junquera, J. Phys. Chem. B 2008, 112, 12555-12565 [3] M. Munoz-Obeda, A. Rodriguez-Pulido, A. Nogales, A. Martin-Molina, E. Aicart and E. Junquera, Biomacromolecules 2010, 11, 3332-3340 [4]M. Muiioz-Ilbeda, A. Rodriguez-Pulido, A. Nogales, 0. Llorca, M. Quesada-Perez, A. Martin- Molina, E. Aicart and E. Junquera, Soft Matter 2011, (DOI: 10.1039/cIsm 05352c) [5]A. Bajaj, S. K.Mishra, P. Kondaiah, S. Bhattacharya, Biochinzica et Biophysica Ada 2008, 1778, 1222-1236