Deuterium-Stabilised Ribonucleic Acid (RNA) Molecules Displaying Increased Resistance to Thermal and Enzymatic Hydrolysis, Aqueous Compositions Comprising Stabilised RNA Molecules and Methods for Making Same

Abstract

The invention relates to the field of RNA stabilisation, and more particularly to the use of deuterium oxide (D 2 O) during storage and/or synthesis of RNA molecules. Described herein are deuterium-stabilised ribonucleic acid (RNA) molecules that display an increased resistance to thermal and enzymatic hydrolysis. Also described are aqueous compositions comprising stabilized RNA molecules and methods for making same. The invention is particularly useful for in the manufacture of RNA-based therapeutics, such as mRNA vaccines, to render them less sensitive to temperature fluctuations.

1 . An aqueous composition comprising deuterium-stabilised RNA molecules that have been synthesised in presence of deuterium, wherein said deuterium-stabilised RNA molecules are selected from the group consisting of total RNA, siRNA, and shRNA. 
     
     
         2 . The aqueous composition of  claim 1 , wherein said deuterium is present at concentration of at least 85 atom % D. 
     
     
         3 . The aqueous composition of  claim 1 , wherein said deuterium is present at a concentration of about 85 atom % D to about 99.9 atom % D. 
     
     
         4 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules incorporates deuterium. 
     
     
         5 . The aqueous composition of  claim 4 , wherein said deuterium-stabilised RNA molecules comprise deuterated ribonucleosides. 
     
     
         6 . The aqueous composition of  claim 4 , wherein said deuterium-stabilised RNA molecules comprise substitution of protium atoms by deuterium atoms. 
     
     
         7 . The aqueous composition of  claim 4 , wherein said deuterium-stabilised RNA molecules comprise a deuterium atom in the 2′OH-group on the ribose sugar moiety. 
     
     
         8 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules display at least one of: i) increased structural integrity of their primary and/or secondary structure, compared to non-stabilized RNA molecules; and ii) increased resistance to degradation compared to non-stabilised RNA molecules. 
     
     
         9 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules display increased resistance to at least one of (i) hydrolysis or degradation by endonucleases, and (ii) thermal degradation. 
     
     
         10 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules display increased resistance to thermal hydrolysis after 1 day of exposure to 37° C. 
     
     
         11 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules display increased resistance to thermal hydrolysis after a challenge at 45° C. 
     
     
         12 . The aqueous composition of  claim 1 , wherein said aqueous composition further comprises deuterium in solution. 
     
     
         13 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules consists essentially of small interfering RNA (siRNA) molecules. 
     
     
         14 . The aqueous composition of  claim 1 , wherein said deuterium-stabilised RNA molecules are components of a therapeutic. 
     
     
         15 . An aqueous ribonucleic acid (RNA) composition comprising:
 (i) an aqueous solution comprising deuterium at a concentration sufficient for stabilising RNA molecules; and   (ii) deuterium-stabilised RNA molecules that have been synthesised in presence of deuterium, wherein said deuterium-stabilised RNA molecules are selected from the group consisting of total RNA, siRNA, and shRNA.   
     
     
         16 . The aqueous RNA composition of  claim 15 , wherein said deuterium-stabilised RNA molecules are components of a therapeutic. 
     
     
         17 . An aqueous composition comprising deuterium-stabilised siRNA molecules that have been synthesised in presence of deuterium. 
     
     
         18 . The aqueous composition of  claim 17 , wherein said deuterium-stabilised siRNA molecules are components of a therapeutic. 
     
     
         19 . A therapeutical product comprising the aqueous composition of  claim 1 . 
     
     
         20 . A therapeutical product comprising the aqueous composition of  claim 17 .