Title:  Materials and methods for intracellular delivery of biologically active molecules

United States Patent:  6,171,863

Assignee:  University of Florida (Gainesville, FL)

Filed:  September 8, 1998

The subject invention finds utility in the area of gene therapy of diseases. More specifically, the invention concerns the making of a novel non-viral vector which can bind to desired DNA to form a combination useful to transfect diseased mitochondria of human or animal cells. The non-viral vector comprises a dequalinium salt subjected to standard liposome production procedures to obtain the vector names DQAsomes.

DETAILED DISCLOSURE OF THE INVENTION

The subject invention provides materials and methods useful in delivering biologically active molecules to mitochondria. In a preferred embodiment, the subject invention provides a method for selectively transforming mitochondrial DNA. This method can be used to correct defects in mitochondrial DNA.

In a specific embodiment, the subject invention pertains to the use of an amphiphilic dicationic compound complexed with DNA to deliver the DNA specifically to the mitochondria. In a preferred embodiment, the amphiphilic dicationic compound is a salt of dequalinium (DQA). The salt may be, for example, dequalinium chloride (available from Sigma Chemical Company, St. Louis, Mo.). Using standard liposome production procedures, combined with the teachings provided herein, dequalinium chloride can be transformed into an effective non-viral gene therapy vector (DQAsomes). This is a novel use for DQA. This is the first disclosure that DQAsomes are effective as a transfection system.

The gene therapy vectors of the subject invention can be used to treat diseases associated with mitochondrial DNA, for example, Alzheimer's disease, Parkinson's disease, Leber's Hereditary Optic Neuropathy, myoclonic epilepsy and ragged-red fiber disease, Leigh's syndrome dystonia, adult-onset chronic progressive external opthalmoplegia, Kearns-Sayre syndrome and Pearson's marrow/pancreas syndrome. The DNA delivery vectors of the subject invention are particularly advantageous because these amphipathic dicationic compounds will specifically deliver DNA to the mitochondria. Thus, in a specific embodiment of the subject invention, DQAsomes can be used as a mitochondria-specific polynucleotide delivery system.

Those skilled in the art, having the benefit of the instant disclosure, will appreciate that other salts of dequalinium can be used. In one specific embodiment, dequalinium acetate (Sigma Chemical Company, St. Louis, Mo.) can be used. Other amphiphilic dicationic compounds which can be used according to the subject invention include all derivatives of dequalinium with varying substituents at the aromatic ring systems including 1,1'-Decamethylene bis-quinolinium-salts, which have no substituents at all. The critical characteristics of the compounds which can be used according to the subject invention include the presence of two positive charge centers separated by a relatively long hydrocarbon chain. The hydrocarbon chain may have, for example, from about 5 to about 20 carbons. In a preferred embodiment, the hydrocarbon chain may have from about 8 to about 12 carbons.

Claim 1 of 3 Claims

What is claimed is:

1. A method for delivering DNA into a cell wherein said method comprises administering to said cell a complex of DNA and a molecule which comprises two positive charge centers separated by a hydrocarbon chain; wherein said hydrocarbon chain has 8 to 20 carbons and wherein said molecule with two positive charge centers is a salt of dequalinium; wherein upon administration of said complex to said cell, said complex enters said cell and delivers said DNA into said cell.

____________________________________________
If you want to learn more about this patent, please go directly to the U.S. Patent and Trademark Office Web site to access the full patent.