Multimeric coding nucleic acid and uses thereof

Abstract

The present invention provides, among other things, multimeric coding nucleic acids that exhibit superior stability for in vivo and in vitro use. In some embodiments, a multimeric coding nucleic acid (MCNA) comprises two or more encoding polynucleotides linked via 3′ ends such that the multimeric coding nucleic acid compound comprises two or more 5′ ends.

1 . A multimeric coding nucleic acid (MCNA) comprising two polynucleotides linked via 3′ ends via an oligonucleotide bridge comprising a stable 3-3′ inverted phosphodiester linkage such that the multimeric coding nucleic acid compound comprises two or more 5′ ends, and
 wherein at least one of the two polynucleotides is an encoding polynucleotide. 
 
     
     
         2 .- 4 . (canceled) 
     
     
         5 . The MCNA of  claim 1 , wherein the at least one encoding polynucleotide encodes a protein of interest. 
     
     
         6 . The MCNA of  claim 5 , wherein the two polynucleotides are encoding polynucleotides, and each of the two encoding polynucleotides encodes the same protein. 
     
     
         7 . The MCNA of  claim 5 , wherein the two polynucleotides are encoding polynucleotides, and each of the two encoding polynucleotides encodes a distinct protein. 
     
     
         8 .- 11 . (canceled) 
     
     
         12 . The MCNA of  claim 1 , wherein the at least one encoding polynucleotide comprises a 3′ UTR. 
     
     
         13 . (canceled) 
     
     
         14 . The MCNA of  claim 12 , wherein the 3′ UTR comprises a plurality of multi-A segments with spacers in between. 
     
     
         15 .- 26 . (canceled) 
     
     
         27 . The MCNA of  claim 1 , wherein the nucleosides comprising the oligonucleotide bridge are selected from the group consisting of 2′-OMe-A, 2′-OMe-G, 2′-OMe-C, 2′-OMe-U, 2′-F-A, 2′-F-G, 2′-F-C, 2′-F-U, LNA-A, LNA-G, LNA-C, LNA-U, N6-methyl-adenosine, 2-thiouridine (2sU), 5-methyl-cytidine (5mC), pseudouridine (ψU), and 1-methyl-pseudouridine. 
     
     
         28 .- 30 . (canceled) 
     
     
         31 . The MCNA of  claim 1 , wherein the at least one encoding polynucleotide comprises one or more modified nucleosides. 
     
     
         32 . The MCNA of  claim 31 , wherein the modified nucleosides are selected from the group consisting of 2′-OMe-A, 2′-OMe-G, 2′-OMe-C, 2′-OMe-U, 2′-F-A, 2′-F-G, 2′-F-C, 2′-F-U, LNA-A, LNA-G, LNA-C, LNA-U, N6-methyl-adenosine, 2-thiouridine (2sU), 5-methyl-cytidine (5mC), pseudouridine (ψU), and 1-methyl-pseudouridine. 
     
     
         33 .- 38 . (canceled) 
     
     
         39 . The MCNA of  claim 1 , wherein the at least one or more encoding polynucleotides comprise a polynucleotide that encodes an enzyme, a receptor, a ligand, a light chain or heavy chain of an antibody, a nuclease, or a DNA-binding protein. 
     
     
         40 . (canceled) 
     
     
         41 . A composition comprising the MCNA of  claim 1  encapsulated or complexed with a delivery vehicle. 
     
     
         42 . The composition of  claim 41 , wherein the delivery vehicle is selected from the group consisting of liposomes, lipid nanoparticles, solid-lipid nanoparticles, polymers, viruses, sol-gels, and nanogels. 
     
     
         43 .- 47 . (canceled) 
     
     
         48 . The MCNA of  claim 12 , wherein the 3′ UTR does not include a polyA tail. 
     
     
         49 . The MCNA of  claim 1 , at least one of the two polynucleotides is unmodified. 
     
     
         50 . The MCNA of  claim 1 , wherein the oligonucleotide bridge is unmodified. 
     
     
         51 . The MCNA of  claim 1 , wherein at least one of the two polynucleotides has a 5′ cap. 
     
     
         52 . The MCNA of  claim 1 , wherein the second polynucleotide is an non-coding polynucleotide. 
     
     
         53 . The MCNA of  claim 1 , wherein the MCNA comprises a second encoding polynucleotide.