Long-Acting DNA Dendrimers and Methods Thereof
This invention provides a unique composition which includes a DNA dendrimer combined with siRNA molecule. Further, methods of preparing a composition which includes a DNA dendrimer combined with a siRNA molecule, methods of protecting a DNA dendrimer siRNA complex against degradation in body fluids, methods of protecting a DNA dendrimer against degradation in bodily fluids, and methods of delivering a DNA dendrimer into bodily fluids are provided.
1 . A composition comprising a DNA dendrimer attached to a siRNA molecule.
2 . The composition of claim 1 , further comprising a protein, a peptide, an aptamer, fluorescein, a fluorescein derivative, a fluorescent dye, digoxigenin, cholesterol, a primary amine, a hydrocarbon spacer of length 3 to 120 carbons, FITC, a PEG molecule, biotin, a biotin derivative, fluorescein or fluorescein derivative, or any combination thereof attached to said DNA dendrimer.
3 . The composition of claim 2 , comprising a) a DNA dendrimer attached to a siRNA molecule and an antibody or an antibody fragment, b) a DNA dendrimer attached to a siRNA molecule and biotin, or c) a DNA dendrimer attached to a siRNA molecule and fluorescein or a fluorescein derivative.
4 .- 7 . (canceled)
8 . A method of preparing a composition comprising a DNA dendrimer and a siRNA molecule, comprising the step of attaching said siRNA to said DNA dendrimer (a) via a disulfide bridging bond; (b) via the use of NHS ester dependent condensation reaction; (c) via the use of bifunctional cross linking reaction; (d) via direct or indirect hybridization of the siRNA to DNA dendrimer sequence; or (e) via the use of polycationic compounds to bridge siRNA molecule to said DNA dendrimer via charge- charge interactions.
9 . The method of claim 8 , further comprising the step of attaching to said DNA dendrimer a protein, fluorescein or a fluorescein derivative, digoxigenin, cholesterol, a primary amine, a hydrocarbon spacer of length of 3 to 120 carbons, a molecule containing a polyethylene glycol (PEG) moiety, biotin or a biotin derivative, or any combination thereof.
10 . The method of claim 9 , which comprises further attaching an antibody or a fragment thereof.
11 .- 12 . (canceled)
13 . A method of protecting a siRNA molecule against degradation in a bodily fluid, comprising the step of attaching (a) said siRNA molecule and (b) a protein, digoxigenin, cholesterol, a primary amine, a hydrocarbon spacer of length of 3 to 120 carbons, a PEG molecule, biotin, a biotin derivative, fluorescein or a fluorescein derivative, or any combination thereof to a DNA dendrimer, thereby protecting the siRNA molecule against degradation.
14 .- 15 . (canceled)
16 . The method of claim 13 , wherein said protein is an antibody or a fragment thereof.
17 .- 18 . (canceled)
19 . The method of claim 13 , wherein the DNA dendrimer is protected against nuclease degradation in the bodily fluid.
20 .- 23 . (canceled)
24 . The method of claim 19 , further comprising the step of attaching to said DNA dendrimer a molecule comprising a DNA molecule, a RNA molecule, a protein, a chemotherapeutic agent, an antiviral agent, an anti-inflammatory agent, a bacteriostatic agent, a psychoactive agent, a statin, a neuropathic agents, a hormone, an ACE inhibitor, an anti-clotting factor, an analgestic, an anti angiogenic agent, a pro angiogenic agent, a growth factor, a growth factor inhibitor, or any combination thereof.
25 .- 32 . (canceled)
33 . The composition of claim 3 , wherein the antibody is a monoclonal antibody, a single-chain Fv fragment or a conjugated antibody.
34 . The composition of claim 2 , wherein the fluorescent dye has a molecular weight less than 10,000 daltons and is attached to the DNA dendrimer by a spacer.
35 . The composition of claim 34 , wherein the fluorescent dye is Cy3, Cy5, Oyster 550, Oyster 650, an Alexa Fluor dye or BODIPY 630/650.
36 . The composition of claim 2 , wherein the fluorescent dye has a molecular weight greater than 50,000 daltons.
37 . The composition of claim 36 , wherein the fluorescent dye is R-phycoerythrin conjugated to streptavidin, B-phycoerythrin conjugated to streptavidin or allophycocyanin conjugated to streptavidin.
38 . The composition of claim 2 which is a pharmaceutical composition.
39 . The composition of claim 38 which is formulated for oral, rectal, transmucosal, transnasal, intestinal, parenteral or local delivery.
40 . The composition of claim 39 which is formulated for intramuscular, subcutaneous, intramedullary, intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, or intraocular injection.
41 . The composition of claim 2 which is a composition for in vitro transfection.
42 . The composition of claim 2 , further comprising an active agent from the group consisting of a DNA molecule, a RNA molecule, a protein, a chemotherapeutic agent, an antiviral agent, an anti-inflammatory agent, a bacteriostatic agent, a psychoactive agent, a statin, a neuropathic agents, a hormone, an ACE inhibitor, an anti-clotting factor, an analgesic, an anti angiogenic agent, a pro angiogenic agent, a growth factor, a growth factor inhibitor, or any combination thereof attached to the DNA dendrimer.