Mitochondrial leader sequence-plasmid DNA conjugates delivered into mammalian cells by DQAsomes co-localize with mitochondria Gerard G.M. D’Souza, Sarathi V. Boddapati, Volkmar Weissig * Department of Pharmaceutical Sciences, School of Pharmacy, Bouve ´ College of Health Sciences, Northeastern University, 360 Huntington Avenue, 211 Mugar Building, Boston, MA 02115, USA Received 6 April 2005; received in revised form 7 July 2005; accepted 12 July 2005 Abstract In the last decade the increase in therapeutic strategies aimed at mitochondrial targets has resulted in the need for novel delivery systems for the selective delivery of drugs and DNA into mitochondria. In this study, we have continued our efforts towards the development of the first mitochondriotropic drug and DNA delivery system (DQAsomes). Prepared from derivatives of the self-assembling mitochondriotropic bola-amphiphile dequalinium chloride, these vesicles bind and transport DNA to mitochondria in living mammalian cells where upon they have been shown to release the DNA on contact with mitochondrial membranes. We present data to demonstrate that oligonucleotides as well as plasmid DNA conjugated to a mitochondrial leader sequence (MLS) co-localize with mitochondria when delivered into mammalian cells by DQAsomes. In contrast to a commercially available DNA delivery vector, our vesicles appear to have a pronounced specificity for mitochondria. Further, the data strongly suggest that linear conjugates might be better suited to delivery into mitochondria and that in the absence of a mitochondria specific vector, the presence of a MLS-peptide conjugated to the DNA is alone not sufficient to direct the accumulation of DNA at mitochondria. q 2005 Elsevier B.V. and Mitochondria Research Society. All rights reserved. Keywords: Mitochondria; Drug delivery; Mitochondrial gene therapy; Mitochondrial drug targeting; Dequalinium; Oligonucleotides; Plasmid DNA; Mitochondrial leader sequence peptides; DNA peptide conjugates 1. Introduction It has become progressively more evident that mitochondrial dysfunction contributes to a variety of human disorders such as neurodegenerative and neuromuscular diseases, obesity, diabetes, ischemia- reperfusion injury and cancer. Increased efforts directed towards the study of mitochondria as targets for pharmacological intervention, have made Mitochondrion 5 (2005) 352–358 www.elsevier.com/locate/mito 1567-7249/$ - see front matter q 2005 Elsevier B.V. and Mitochondria Research Society. All rights reserved. doi:10.1016/j.mito.2005.07.001 Abbreviations DOPE, dioleoyl phosphatidyl ethanolamine; DOTAP, dioleoyl oxytrimethylammonio propane; DMRIE, 1-2 dimyristoyloxypropyl 3- dimethylhydroxyethyl ammonium bro- mide; TCEP, Tris (2 carboxy ethyl) phosphine; mOTC, mouse ornithine transcarbamylase; hMDH, human malate dehydrogenase; COX, cytochrome c oxidase. * Corresponding author. Tel.: C1 617 373 3212; fax: C1 617 373 8886. E-mail address: v.weissig@neu.edu (V. Weissig).