Virion-like delivery particles for self-replicating rna molecules

Abstract

Nucleic acid immunisation is achieved by delivering a self-replicating RNA encapsulated within a small particle. The RNA encodes an immunogen of interest, and the particle may deliver this RNA by mimicking the delivery function of a natural RNA virus. Thus the invention provides a non-virion particle for in vivo delivery of RNA to a vertebrate cell, wherein the particle comprises a delivery material encapsulating a self-replicating RNA molecule which encodes an immunogen. These particles are useful as components in pharmaceutical compositions for immunising subjects against various diseases.

1 . A formulation comprising self-replicating ribonucleic acid (RNA) molecules and liposomes; the self-replicating RNA molecules comprising an open reading frame that encodes an immunogen and an open reading frame that encodes a RNA-dependent RNA polymerase; the immunogen comprising a respiratory syncytial virus (RSV) immunogen, an Epstein-Barr virus (EBV) immunogen, a cytomegalovirus (CMV) immunogen, a herpes simplex virus (HSV) immunogen, a human immunodeficiency virus (HIV) immunogen, a coronavirus immunogen, an influenza virus immunogen, a Varicella zoster virus (VZV) immunogen, or a flavivirus immunogen; the liposomes comprising a cationic lipid and a first lipid; the first lipid comprising an anionic lipid or a zwitterionic lipid; the liposomes comprising within at least half of the self-replicating RNA molecules. 
     
     
         2 . The formulation of  claim 1 , the liposomes comprising a polyethylene glycol-conjugated (PEG-conjugated) lipid. 
     
     
         3 . The formulation of  claim 2 , the self-replicating RNA molecules comprising modified nucleotides. 
     
     
         4 . The formulation of  claim 1 , at least 80% by number of the liposomes have diameters from 20 nm to 220 nm. 
     
     
         5 . The formulation of  claim 4 , the self-replicating RNA molecules comprising modified nucleotides. 
     
     
         6 . The formulation of  claim 1  being immunogenic against the immunogen in vivo, and the self-replicating RNA molecules comprising a 3′ poly(adenosine monophosphate) (poly(A)) tail. 
     
     
         7 . The formulation of  claim 6 , the self-replicating RNA molecules comprising modified nucleotides. 
     
     
         8 . The formulation of  claim 2 , the PEG-conjugated lipid comprising a PEG that has a molecular weight of 2000 Da. 
     
     
         9 . The formulation of  claim 2 , the self-replicating RNA molecules further comprising a 7′-methylguanosine, a triphosphate bridge, and a 5′ first ribonucleoside, the 7′-methylguanosine linked 5′-to-5′ to the first 5′ ribonucleoside by the triphosphate bridge. 
     
     
         10 . The formulation of  claim 9 , the first 5′ ribonucleoside comprising a 2′-methylated ribose. 
     
     
         11 . The formulation of  claim 4 , the self-replicating RNA molecules further comprising a 7′-methylguanosine, a triphosphate bridge, and a 5′ first ribonucleoside, the 7′-methylguanosine linked 5′-to-5′ to the first 5′ ribonucleoside by the triphosphate bridge. 
     
     
         12 . The formulation of  claim 11 , the first 5′ ribonucleoside comprising a 2′-methylated ribose. 
     
     
         13 . The formulation of  claim 6 , the self-replicating RNA molecules further comprising a 7′-methylguanosine, a triphosphate bridge, and a 5′ first ribonucleoside, the 7′-methylguanosine linked 5′-to-5′ to the first 5′ ribonucleoside by the triphosphate bridge. 
     
     
         14 . The formulation of  claim 13 , the first 5′ ribonucleoside comprising a 2′-methylated ribose. 
     
     
         15 . The formulation of  claim 2 , the immunogen comprising the coronavirus immunogen. 
     
     
         16 . The formulation of  claim 4 , the immunogen comprising the coronavirus immunogen. 
     
     
         17 . The formulation of  claim 6 , the immunogen comprising a coronavirus immunogen; the self-replicating RNA molecules being positive-stranded; the open reading frame that encodes the RNA-dependent RNA polymerase comprising a nucleic acid that encodes Venezuelan equine encephalitis virus non-structural proteins 1-4 (nsP1-4). 
     
     
         18 . The formulation of  claim 15 , the coronavirus immunogen comprising a spike protein. 
     
     
         19 . A formulation comprising self-replicating ribonucleic acid (RNA) molecules and liposomes; the self-replicating RNA molecules comprising an open reading frame that encodes an immunogen and an open reading frame that encodes a RNA-dependent RNA polymerase; the liposomes comprising a cationic lipid and a first lipid; the first lipid comprising an anionic lipid or a zwitterionic lipid; the liposomes comprising within at least half of the self-replicating RNA molecules. 
     
     
         20 . The formulation of  claim 19 , the immunogen comprising a tumor polypeptide.