Short interfering rna templated lipoprotein particles (sirna-tlp)

Abstract

Nanostructures for the systemic delivery of nucleic acids, such as RNA, are provided herein. The nanostructures include templated lipoprotein nanoparticles (TLPs) composed of a core decorated with proteins, a lipid bilayer and hydrophobic molecules that self-assemble with nucleic acids, such as RNA. The nanostructures are useful for research, therapeutic and diagnostic applications.

What is claimed is: 
     
         1 . An anionic nanostructure aggregate, comprising:
 an aggregate of cationic lipid-nucleic acid complexes and templated lipoprotein particles (TLP), wherein the TLP comprises a core, and a lipid shell surrounding the core; and the cationic lipid-nucleic acid complex, comprised of nucleic acids, wherein each nucleic acid is complexed with a cationic lipid, and wherein the aggregate of cationic lipid-nucleic acid complexes and TLPs has a negative ζ-potential and forms the anionic nanostructure aggregate.   
     
     
         2 . The nanostructure of  claim 1 , wherein the core is a metal. 
     
     
         3 . The nanostructure of  claim 2 , wherein the core is gold. 
     
     
         4 . The nanostructure of  claim 1 , wherein the lipid shell comprises phospholipids, and wherein the phospholipids are 1,2-dioleoyl-sn-glycero-3-phophocholine (DOPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-[3-(2-pyridyldithio)propionate] (PDP-PE). 
     
     
         5 . The nanostructure of  claim 1 , further comprising an apolipoprotein functionalized to the core. 
     
     
         6 . The nanostructure of  claim 5 , wherein the apolipoprotein is apolipoprotein A-1 (apo A-1). 
     
     
         7 . The nanostructure of  claim 1 , further comprising cholesterol. 
     
     
         8 . The nanostructure of  claim 1 , wherein the nucleic acids are single-stranded. 
     
     
         9 . The nanostructure of  claim 1 , wherein the nucleic acids are RNAs. 
     
     
         10 . The nanostructure of  claim 9 , wherein the RNAs are antisense and sense RNAs of an siRNA duplex. 
     
     
         11 . The nanostructure of  claim 1 , wherein the sense RNA and antisense RNA are present in nearly equimolar amounts. 
     
     
         12 . The nanostructure of  claim 1 , wherein the sense RNA and antisense RNA are present in about a 1:2 ratio. 
     
     
         13 . The nanostructure of  claim 1 , wherein the sense RNA and antisense RNA are present in about a 1:1 ratio. 
     
     
         14 . The nanostructure of  claim 1 , wherein the sense RNA and antisense RNA are present in about a 2:1 ratio. 
     
     
         15 . The nanostructure of  claim 1 , wherein the nucleic acid in the nanostructure is more stable than free nucleic acid. 
     
     
         16 . The nanostructure of  claim 1 , wherein the nanostructure comprises alternating layers of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) and nucleic acid. 
     
     
         17 . The nanostructure of  claim 1 , wherein the nanostructure is solid. 
     
     
         18 . The nanostructure of  claim 1 , wherein the nucleic acid is not chemically modified. 
     
     
         19 . The nanostructure of  claim 1 , wherein the nucleic acid is mixed with TLP in a molar ratio of 5:1, 15:1 or 25:1. 
     
     
         20 . The nanostructure of  claim 19 , wherein the nucleic acid is mixed with TLP in a molar ratio of 25:1. 
     
     
         21 . The nanostructure of  claim 1 , wherein the cationic lipid is DOTAP. 
     
     
         22 . The nanostructure of  claim 21 , wherein the DOTAP is mixed with nucleic acid in a molar ratio of 10:1, 20:1, 30:1 or 40:1. 
     
     
         23 . The nanostructure of  claim 1 , wherein the TLP is a synthetic HDL. 
     
     
         24 . A method for delivering siRNA to a cell in a subject, the method comprising:
 contacting the subject with an anionic nanostructure aggregate, comprising an aggregate of cationic lipid-nucleic acid complexes and templated lipoprotein particles (TLP), wherein the cationic lipid-nucleic acid complex is comprised of antisense and sense RNA of an siRNA duplex, each complexed with a cationic lipid, and wherein the aggregate of cationic lipid-nucleic acid complexes and TLPs has a negative ζ-potential and forms the anionic nanostructure aggregate, to deliver siRNA to the cell.   
     
     
         25 . The method of  claim 24 , wherein the antisense RNA and/or the sense RNA of the siRNA duplex is single-stranded. 
     
     
         26 . The method of  claim 24 , wherein the antisense RNA and the sense RNA of the siRNA duplex are single-stranded. 
     
     
         27 . The method of  claim 24 , wherein the cell is a cancer cell. 
     
     
         28 . The method of  claim 27 , wherein the cancer cell is a prostate cancer cell, a breast cancer cell, a renal cancer cell or an ovarian cancer cell. 
     
     
         29 . The method of  claim 24 , wherein the cell is a LNCaP cell, an enzalutamide resistant LNCaP cell, an MDA-MB-231 cell, a 786-O cell, or a OvCar3 cell. 
     
     
         30 . The method of  claim 24 , wherein the cell is contacted with the nanostructure at a concentration of 1nM, 5 nM, 10 nM, or 20 nM. 
     
     
         31 . The method of  claim 24 , wherein the cell is in contact with the nanostructure for 24, 48, 72, and 96 hours. 
     
     
         32 . The method of  claim 24 , wherein the cell expresses the androgen receptor (AR) or the enhancer of zeste homolog 2 (EZH2) proteins. 
     
     
         33 . The method of  claim 24 , wherein the subject is a mammal. 
     
     
         34 . The method of  claim 33 , wherein the subject is a human. 
     
     
         35 . A method for treating a cancer, the method comprising:
 systemically administering to a subject having a cancer an anionic nanostructure aggregate, comprising an aggregate of cationic lipid-nucleic acid complexes and templated lipoprotein particles (TLP), wherein the cationic lipid-nucleic acid complex is comprised of antisense and sense RNA of an siRNA duplex, each complexed with a cationic lipid, and wherein the aggregate of cationic lipid-nucleic acid complexes and TLPs has a negative ζ-potential and forms the anionic nanostructure aggregate, wherein the siRNA is an anti-cancer siRNA.   
     
     
         36 . The method of  claim 35 , wherein the antisense RNA and/or the sense RNA of the siRNA duplex is single-stranded. 
     
     
         37 . The method of  claim 35 , wherein the antisense RNA and the sense RNA of the siRNA duplex are single-stranded. 
     
     
         38 . The method of  claim 35 , wherein the nanostructure is administered in vivo. 
     
     
         39 . The method of  claim 35 , wherein the subject is a mammal. 
     
     
         40 . The method of  claim 39 , wherein the subject is a human. 
     
     
         41 . The method of  claim 35 , wherein the nanostructure is administered at a dose of about 0.7 mg siRNA/kg. 
     
     
         42 . The method of  claim 35 , wherein the nanostructure is not toxic to a surrounding non-cancerous cell or non-cancerous tissue. 
     
     
         43 . A method of regulating gene expression in a cell with an anionic nanostructure aggregate, comprising an aggregate of cationic lipid-nucleic acid complexes and templated lipoprotein particles (TLP), wherein the cationic lipid-nucleic acid complex is comprised of antisense and sense RNA of an siRNA duplex, each complexed with a cationic lipid, and wherein the aggregate of cationic lipid-nucleic acid complexes and TLPs has a negative ζ-potential and forms the anionic nanostructure aggregate. 
     
     
         44 . The method of  claim 43 , wherein the antisense RNA and/or the sense RNA of the siRNA duplex is single-stranded. 
     
     
         45 . The method of  claim 43 , wherein the antisense RNA and the sense RNA of the siRNA duplex are single-stranded. 
     
     
         46 . The method of  claim 43 , wherein the nanostructure decreases the expression of a gene. 
     
     
         47 . The method of  claim 46 , wherein the nanostructure decreases the expression of the gene that encodes for the AR protein. 
     
     
         48 . The method of  claim 46 , wherein the nanostructure decreases the expression of the gene that encodes for the EZH2 protein. 
     
     
         49 . A method for synthesizing a templated lipoprotein particle (TLP) comprising contacting gold nanoparticles with an apolipoprotein to produce an apolipoprotein coated gold particle, contacting the apolipoprotein coated gold particle with two phospholipids and cholesterol to produce an anionic TLP. 
     
     
         50 . The method of  claim 49 , further comprising mixing the anionic TLP with a DOTAP RNA mixture.