PHARMACEUTICAL COMPOSITION CONTAINING A STABILISED mRNA OPTIMISED FOR TRANSLATION IN ITS CODING REGIONS

Abstract

The present invention relates to a pharmaceutical composition comprising a modified mRNA that is stabilised by sequence modifications and optimised for translation. The pharmaceutical composition according to the invention is particularly well suited for use as an inoculating agent, as well as a therapeutic agent for tissue regeneration. In addition, a process is described for determining sequence modifications that promote stabilisation and translational efficiency of modified mRNA of the invention.

1 - 28 . (canceled) 
     
     
         29 . A method of providing dystrophin expression in a subject having cystic fibrosis, the method comprising administering an effective amount of a pharmaceutical composition comprising an mRNA encoding dystrophin to the subject. 
     
     
         30 . The method of  claim 29 , wherein the subject has insufficient levels of dystrophin. 
     
     
         31 . The method of  claim 29 , wherein the pharmaceutical composition is administered by injection. 
     
     
         32 . The method of  claim 29 , wherein the pharmaceutical composition is administered intravenously, intradermally, subcutaneously, intramuscularly, topically, or orally. 
     
     
         33 . The method of  claim 32 , wherein the pharmaceutical composition is administered intravenously. 
     
     
         34 . The method of  claim 29 , wherein the mRNA encoding dystrophin comprises a sequence wherein at least one codon of a wild-type sequence recognized by a rare cellular tRNA is replaced with a codon recognized by an abundant cellular tRNA, and wherein said rare cellular tRNA and said abundant cellular tRNA recognize the same amino acid. 
     
     
         35 . The method of  claim 29 , wherein the mRNA encoding dystrophin comprises an increased G/C content relative to a wild-type RNA encoding dystrophin. 
     
     
         36 . The method of  claim 29 , wherein the mRNA encoding dystrophin comprises a stabilizing 5′ untranslated region (UTR) or 3′ UTR. 
     
     
         37 . The method of  claim 29 , wherein the mRNA comprises a 5′ cap structure and/or a poly-A tail of at least 50 nucleotides. 
     
     
         38 . The method of  claim 29 , wherein the mRNA encoding dystrophin comprises at least one chemical modification of the mRNA. 
     
     
         39 . The method of  claim 29 , wherein the mRNA encoding dystrophin comprises at least one nucleotide that is substituted with an analog of the naturally occurring nucleotide. 
     
     
         40 . The method of  claim 29 , wherein the mRNA encoding dystrophin comprises at least one nucleotide position replaced with a nucleotide analogue selected from the group consisting of phosphorous amidates, phosphorous thioates, peptide nucleotides, methylphosphonates, 7-deazaguanosine, 5-methylcytosine, and inosine. 
     
     
         41 . The method of  claim 29 , wherein the mRNA further encoding a secretion signal. 
     
     
         42 . The method of  claim 29 , wherein the mRNA is dissolved in an aqueous carrier. 
     
     
         43 . The method of  claim 42 , wherein the aqueous carrier is water for injection (WFI), a buffered solution, or a salt solution. 
     
     
         44 . The method of  claim 43 , wherein the salt solution comprises sodium chloride or potassium chloride solution. 
     
     
         45 . The method of  claim 29 , wherein the pharmaceutical composition comprises a component selected from the group consisting of human serum albumin, a polycationic protein, polysorbate 80, a sugar, and an amino acid. 
     
     
         46 . The method of  claim 45 , wherein the pharmaceutical composition comprises a polycationic protein. 
     
     
         48 . The method of  claim 29 , wherein the mRNA is provided in a liposome complex. 
     
     
         50 . The method of  claim 29 , further comprising administering the pharmaceutical composition to the subject two or more times. 
     
     
         52 . The method of  claim 29 , wherein the composition is administered by injection and wherein the mRNA is provided in a liposome complex.