Mannose-Based mRNA Targeted Delivery System and Use Thereof

Abstract

The present invention provides a mannose-based mRNA targeted delivery system and use thereof. The mRNA can encode one or more target polypeptides and contains at least one mannose modification. The technical solution of the present invention modifies the mRNA molecule with a mannose, so that the mRNA can be directly and efficiently coupled with the mannose, and the targeted delivery of the mRNA is realized without the need for a carrier.

What is claimed is: 
     
         1 . A method of preparing an mRNA-mannose conjugate, comprising
 a. synthesizing a polyadenylic acid, wherein a last adenine of the polyadenylic acid is labeled with a secondary amine;   b. linking the secondary amine with at least one mannose through an isourea bond to obtain a polyadenylic acid-mannose conjugate; and   c. linking the polyadenylic acid-mannose conjugate to a 3′ terminus of an mRNA precursor encoding a polypeptide,   
       thereby obtaining the mRNA-mannose conjugate. 
     
     
         2 . The method of  claim 1 , wherein the at least one mannose is 4-isothiocyanatophenyl-α-D-mannoside. 
     
     
         3 . The method of  claim 1 , wherein the ratio of the polyadenylic acid over the at least one mannose is about 1:5 (in weight). 
     
     
         4 . The method of  claim 3 , wherein the polyadenylic acid is about 20 μg, and wherein the at least one mannose is about 100 μg. 
     
     
         5 . The method of  claim 1 , wherein the step b is reacted under a pH of about 9.0, under about 25° C., and is performed overnight. 
     
     
         6 . The method of  claim 1 , further comprising a step b′ after step b and prior to step c: purifying the polyadenylic acid-mannose conjugate with a desalting column. 
     
     
         7 . A kit for preparing an mRNA-mannose conjugate, comprising:
 a. an mRNA precursor comprising a coding sequence encoding a polypeptide;   b. a polyadenylic acid, wherein a last adenine of the polyadenylic acid is labeled with a secondary amine; and   c. a mannose.   
     
     
         8 . A method of expressing a polypeptide in a subject in need thereof, comprising administering to the subject an effective amount of an mRNA-mannose conjugate, where the mRNA-mannose conjugate comprises an mRNA and at least one mannose, wherein the mRNA encodes the polypeptide and comprises a polyA tail at its 3′ terminus, and wherein the at least one mannose is attached to a last adenine of the polyA tail via a secondary amine. 
     
     
         9 . The method of  claim 8 , wherein the mRNA-mannose conjugate is administered parenterally (e.g., subcutaneously, intravenously, intramuscularly, or intraperitoneally), orally (e.g., ingestion, buccal, or sublingual), or intra-tumorally. 
     
     
         10 . The method of  claim 8 , wherein the subject is diagnosed of or is suspected of having a tumor or a pathogen, and optionally wherein the subject is diagnosed of or is suspected of having melanoma. 
     
     
         11 . A method of treating a subject in need thereof, comprising administering to the subject an effective amount of an mRNA-mannose conjugate, where the mRNA-mannose conjugate comprises an mRNA and at least one mannose, wherein the mRNA encodes the polypeptide and comprises a polyA tail at its 3′ terminus, and wherein the at least one mannose is attached to a last adenine of the polyA tail via a secondary amine. 
     
     
         12 . The method of  claim 11 , wherein the mRNA-mannose conjugate is administered parenterally (e.g., subcutaneously, intravenously, intramuscularly, or intraperitoneally), orally (e.g., ingestion, buccal, or sublingual), or intra-tumorally. 
     
     
         13 . The method of  claim 11 , wherein the subject is diagnosed of or is suspected of having a tumor or a pathogen, and optionally wherein the subject is diagnosed of or is suspected of having melanoma. 
     
     
         14 . A method of triggering or activating an immune-response in a subject in need thereof, comprising administering to a subject an effective amount of an mRNA-mannose conjugate, where the mRNA-mannose conjugate comprises an mRNA and at least one mannose, wherein the mRNA encodes the polypeptide and comprises a polyA tail at its 3′ terminus, and wherein the at least one mannose is attached to a last adenine of the polyA tail via a secondary amine. 
     
     
         15 . The method of  claim 14 , wherein the mRNA-mannose conjugate is administered parenterally (e.g., subcutaneously, intravenously, intramuscularly, or intraperitoneally), orally (e.g., ingestion, buccal, or sublingual), or intra-tumorally. 
     
     
         16 . The method of  claim 14 , wherein the subject is diagnosed of or is suspected of having a tumor or a pathogen, and optionally wherein the subject is diagnosed of or is suspected of having melanoma.