Liver specific delivery of messenger rna

Abstract

Disclosed herein are compositions and methods of modulating the expression of gene or the production of a protein by transfecting target cells with nucleic acids. The compositions disclosed herein demonstrate a high transfection efficacy and are capable of ameliorating diseases associated with protein or enzyme deficiencies.

1 .- 126 . (canceled) 
     
     
         127 . A method of delivery of messenger RNA (mRNA) for in vivo production of protein, comprising
 administering systemically to a subject in need of delivery a composition comprising an mRNA encoding a protein, encapsulated within a liposome such that the administering of the composition results in the prolonged stable expression of the protein encoded by the mRNA in the liver;   wherein the protein encoded by the mRNA is an enzyme; and   wherein the liposome comprises one or more cationic lipids, one or more non-cationic lipids, one or more cholesterol-based lipids and one or more PEG-modified lipids and has a size less than about 100 nm.   
     
     
         128 . The method of  claim 127 , wherein the one or more non-cationic lipids comprise DOPE. 
     
     
         129 . The method of  claim 127 , wherein the one or more cationic lipids comprise DLinDMA, CLinDMA, or DLin-K-XTC2-DMA. 
     
     
         130 . The method of  claim 127 , wherein the one or more PEG-modified lipids comprise DMG-PEG-2000 or C8-PEG-2000. 
     
     
         131 . The method of  claim 127 , wherein the mRNA is of at least 30 kDa. 
     
     
         132 . The method of  claim 127 , wherein the enzyme is a liver-specific enzyme. 
     
     
         133 . The method of  claim 132 , wherein the enzyme is selected from ornithine transcarbamylase (OTC), carbamyl phosphate synthetase (CPS), argininosuccinate synthetase 1 (ASS1) argininosuccinate lyase (ASL), and arginase (ARG). 
     
     
         134 . The method of  claim 127 , wherein the mRNA is modified to enhance stability. 
     
     
         135 . The method of  claim 127 , wherein the mRNA is modified to include a modified nucleotide, an alteration to the 5′ or 3′ untranslated region, a cap structure or a poly A tail. 
     
     
         136 . The method of  claim 135 , wherein the modified nucleotide is pseudouridine. 
     
     
         137 . The method of  claim 127 , wherein the mRNA is unmodified. 
     
     
         138 . The method of  claim 127 , wherein the protein is retained within the cytosol of the liver cells after expression. 
     
     
         139 . The method of  claim 127 , wherein the protein is secreted extracellularly after expression. 
     
     
         140 . The method of  claim 127 , wherein the protein is systemically distributed. 
     
     
         141 . The method of  claim 127 , wherein the composition is administered intravenously. 
     
     
         142 . The method of  claim 127 , wherein the expression of the protein encoded by the mRNA in the liver is detectable at least 4 hours post administration. 
     
     
         143 . The method of  claim 141 , wherein the expression of the protein encoded by the mRNA in the liver is detectable at least 4 hours post administration. 
     
     
         144 . The method of  claim 127 , wherein the one or more cationic lipids carry a net positive charge at physiological pH. 
     
     
         145 . The method of  claim 144 , wherein the one or more cationic lipids constitute about 20-70% mol % of the total lipids present in the liposome. 
     
     
         146 . The method of  claim 145 , wherein the one or more PEG-modified lipids comprise a polyethylene glycol chain of up to 5 kDa in length covalently attached to a lipid with alkyl chain(s) of C 6 -C 20  length. 
     
     
         147 . The method of  claim 146 , wherein the one or more PEG-modified lipids constitute up to about 20% by molar of the total lipids present in the liposome. 
     
     
         148 . The method of  claim 127 , wherein the subject is suffering from or susceptible to a metabolic disorder resulting from deficiency of the enzyme encoded by the mRNA. 
     
     
         149 . The method of  claim 148 , wherein the composition is administered bi-weekly. 
     
     
         150 . The method of  claim 148 , wherein the composition is administered monthly.