Therapeutic RNA

Abstract

This disclosure relates to the field of therapeutic RNAs for treatment of solid tumor cancers.

1 .- 88 . (canceled) 
     
     
         89 . A composition comprising RNA encoding an IL-12sc protein, RNA encoding an IL-15 sushi protein, RNA encoding an IFNα protein, and RNA encoding a GM-CSF protein. 
     
     
         90 . The composition of  claim 89 , wherein the IFNα protein is an IFNα2b protein. 
     
     
         91 . The composition of  claim 89 , wherein (i) the RNA encoding an IL-12sc protein comprises the nucleotide sequence of SEQ ID NO: 17 or 18, or a nucleotide sequence having at least 80% identity to the nucleotide sequence of SEQ ID NO: 17 or 18 and/or (ii) the IL-12sc protein comprises the amino acid sequence of SEQ ID NO: 14, or an amino acid sequence having at least 80% identity to the amino acid sequence of SEQ ID NO: 14. 
     
     
         92 . The composition of  claim 89 , wherein (i) the RNA encoding an IL-15 sushi protein comprises the nucleotide sequence of SEQ ID NO: 26, or a nucleotide sequence having at least 80% identity to the nucleotide sequence of SEQ ID NO: 26 and/or (ii) the IL-15 sushi protein comprises the amino acid sequence of SEQ ID NO: 24, or an amino acid sequence having at least 80% identity to the amino acid sequence of SEQ ID NO: 24. 
     
     
         93 . The composition of  claim 89 , wherein (i) the RNA encoding an IFNα protein comprises the nucleotide sequence of SEQ ID NO: 22 or 23, or a nucleotide sequence having at least 80% identity to the nucleotide sequence of SEQ ID NO: 22 or 23 and/or (ii) the IFNα protein comprises the amino acid sequence of SEQ ID NO: 19, or an amino acid sequence having at least 80% identity to the amino acid sequence of SEQ ID NO: 19. 
     
     
         94 . The composition of  claim 89 , wherein (i) the RNA encoding a GM-CSF protein comprises the nucleotide sequence of SEQ ID NO: 29, or a nucleotide sequence having at least 80% identity to the nucleotide sequence of SEQ ID NO: 29 and/or (ii) the GM-CSF protein comprises the amino acid sequence of SEQ ID NO: 27, or an amino acid sequence having at least 80% identity to the amino acid sequence of SEQ ID NO: 27. 
     
     
         95 . The composition of  claim 89 , wherein at least one RNA comprises a modified nucleoside in place of at least one uridine, wherein the modified nucleoside is pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), 5-methyl-uridine (m5U), or a combination thereof. 
     
     
         96 . The composition of  claim 89 , wherein each RNA comprises a modified nucleoside in place of at least one uridine, wherein the modified nucleoside is pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), 5-methyl-uridine (m5U), or a combination thereof. 
     
     
         97 . The composition of  claim 89 , wherein at least one RNA comprises the 5′ cap m 2   7,3′-O Gppp(m1 2′-O ApG, or 3′-O-Me-m 7 G(5′)ppp(5′)G, or m 2   7,3′-O Gppp(m 1   2′-O )ApG, or 3′-O-Me-m 7 G(5′)ppp(5′)G. 
     
     
         98 . The composition of  claim 89 , wherein at least one RNA comprises a 5′ UTR comprising (i) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2, 4, and 6, or (ii) a nucleotide sequence having at least 80% identity to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2, 4, and 6 and/or a 3′ UTR comprising (i) the nucleotide sequence of SEQ ID NO: 8, or (ii) a nucleotide sequence having at least 80% identity to the nucleotide sequence of SEQ ID NO: 8. 
     
     
         99 . The composition of  claim 89 , wherein at least one RNA comprises a poly-A tail of at least 100 nucleotides. 
     
     
         100 . The composition of  claim 99 , wherein the poly-A tail comprises the poly-A tail shown in SEQ ID NO: 78. 
     
     
         101 . The composition of  claim 89 , wherein one or more RNA comprises:
 a 5′ cap comprising)m 2   7,3′-O Gppp(m 1   2′-O )ApG or 3′-O-Me-m 7 G(5′)ppp(5′)G;   a 5′ UTR comprising (i) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2, 4, and 6, or (ii) a nucleotide sequence having at least 80% identity to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2, 4, and 6;   a 3′ UTR comprising (i) the nucleotide sequence of SEQ ID NO: 8, or (ii) a nucleotide sequence having at least 80% identity to the nucleotide sequence of SEQ ID NO:8; and   a poly-A tail comprising at least 100 nucleotides.   
     
     
         102 . The composition of  claim 101 , wherein the poly-A tail comprises SEQ ID NO: 78. 
     
     
         103 . A pharmaceutical composition comprising the composition of  claim 89  and a pharmaceutically acceptable carrier, diluent and/or excipient. 
     
     
         104 . A method for treating or reducing the likelihood of a solid tumor comprising administering to a subject in need thereof the composition of  claim 89 . 
     
     
         105 . A method for treating or reducing the likelihood of a solid tumor comprising administering to a subject in need thereof a first RNA, wherein the first RNA encodes an IL-12sc protein, an IL-15 sushi protein, an IFNα protein, or a GM-CSF protein, and additional RNA, wherein:
 if the first RNA encodes an IL-12sc protein, then the additional RNA encodes an IL-15 sushi protein, an IFNα protein, and a GM-CSF protein; 
 if the first RNA encodes an IL-15 sushi protein, then the additional RNA encodes an IL-12sc protein, an IFNα protein, and a GM-CSF protein; 
 if the first RNA encodes an IFNα protein, then the additional RNA encodes an IL-15 sushi protein, an IL-12sc protein, and a GM-CSF protein; and 
 if the first RNA encodes a GM-CSF protein, then the additional RNA encodes an IL-15 sushi protein, an IFNα protein, and an IL-12sc protein. 
 
     
     
         106 . The method of  claim 105 , wherein the first RNA is administered to the subject at the same time as the additional RNA. 
     
     
         107 . The method of  claim 104 , wherein the RNA is administered intra-tumorally or peri-tumorally. 
     
     
         108 . The method of  claim 104 , wherein the subject is further treated with an additional therapy comprising (i) surgery to excise, resect, or debulk a tumor, (ii) immunotherapy, (iii) radiotherapy, or (iv) chemotherapy. 
     
     
         109 . The method of  claim 104 , wherein the subject is further treated with a checkpoint modulator. 
     
     
         110 . The method of  claim 109 , wherein the checkpoint modulator is an anti-PD1 antibody, an anti-CTLA-4 antibody, or a combination of an anti-PD1 antibody and an anti-CTLA-4 antibody. 
     
     
         111 . The method of  claim 109 , wherein the RNA is administered intra-tumorally or peri-tumorally via injection, and the checkpoint modulator is administered systemically. 
     
     
         112 . The method of  claim 104 , wherein the solid tumor is a sarcoma, carcinoma, or lymphoma. 
     
     
         113 . The method of  claim 104 , wherein the solid tumor is in the lung, colon, ovary, cervix, uterus, peritoneum, testicles, penis, tongue, pancreas, bone, breast, prostate, soft tissue, connective tissue, kidney, liver, brain, thyroid, or skin. 
     
     
         114 . The method of  claim 104 , wherein the solid tumor is an epithelial tumor, Hodgkin lymphoma (HL), non-Hodgkin lymphoma, prostate tumor, ovarian tumor, renal cell tumor, gastrointestinal tract tumor, hepatic tumor, colorectal tumor, tumor with vasculature, mesothelioma tumor, pancreatic tumor, breast tumor, sarcoma tumor, lung tumor, colon tumor, brain tumor, melanoma tumor, small cell lung tumor, neuroblastoma tumor, testicular tumor, carcinoma tumor, adenocarcinoma tumor, glioma tumor, seminoma tumor, retinoblastoma, or osteosarcoma tumor. 
     
     
         115 . The method of  claim 104 , wherein treating or reducing the likelihood of the solid tumor comprises reducing the size of a tumor, reducing the likelihood of a reoccurrence of cancer in remission, or reducing the likelihood of cancer metastasis in the subject. 
     
     
         116 . A method for treating or reducing the likelihood of a solid tumor, comprising administering to a subject in need thereof RNAs and a further therapy, wherein the RNAs encode an IL-12sc protein, an IL-15 sushi protein, an IFNα protein, and a GM-CSF protein, and the further therapy comprises immunotherapy, chemotherapy, or a checkpoint modulator. 
     
     
         117 . An isolated nucleic acid comprising a sequence encoding an IL-12sc protein, wherein:
 the sequence encoding the IL-12sc protein comprises a) contiguous nucleotides having at least 78% identity to nucleotides 1-984 of SEQ ID NO: 16 or 18, b) contiguous nucleotides having at least 81% identity to nucleotides 1027-1623 of SEQ ID NO: 16 or 18, and c) nucleotides encoding a linker between the nucleotides of a) and b).   
     
     
         118 . An isolated nucleic acid comprising a sequence encoding an IFNα protein, wherein:
 the sequence encoding the IFNα protein has at least 80% identity to the nucleotide sequence of SEQ ID NO: 21 or 23. 
 
     
     
         119 . A method for treating or reducing the likelihood of a solid tumor comprising administering to a subject in need thereof the nucleic acid of  claim 117 , wherein the nucleic acid is RNA, and wherein:
 the subject is further treated with additional RNA encoding an IL-15 sushi protein, an IFNα protein, and a GM-CSF protein.   
     
     
         120 . A method for treating or reducing the likelihood of a solid tumor comprising administering to a subject in need thereof the nucleic of  claim 118 , wherein the nucleic acid is RNA, and wherein:
 the subject is further treated with additional RNA encoding an IL-15 sushi protein, an IL-12sc protein, and a GM-CSF protein.   
     
     
         121 . A method of producing an RNA encoding IL-12sc, comprising contacting an expression construct comprising the DNA nucleic acid of  claim 117  operably linked to a promoter with an RNA polymerase under conditions permissive for transcription. 
     
     
         122 . A method of producing an RNA encoding IFNα, comprising contacting an expression construct comprising the DNA nucleic acid of  claim 118  operably linked to a promoter with an RNA polymerase under conditions permissive for transcription. 
     
     
         123 . An expression construct comprising the nucleic acid of  claim 117  operably linked to a promoter. 
     
     
         124 . An expression construct comprising the nucleic acid of  claim 118  operably linked to a promoter. 
     
     
         125 . A kit comprising the composition of  claim 89 . 
     
     
         126 . A kit comprising RNA encoding an IL-12sc protein, RNA encoding an IL-15 sushi protein, RNA encoding an IFNα protein, and RNA encoding a GM-CSF protein, wherein the RNAs are not in the same container. 
     
     
         127 . A method for treating or reducing the likelihood of a solid tumor comprising administering to a subject in need thereof RNA encoding an IL-12sc protein, RNA encoding an IL-15 sushi protein, RNA encoding an IFNα protein, and RNA encoding a GM-CSF protein, wherein the solid tumor is in a lymph node. 
     
     
         128 . A method for treating or reducing the likelihood of a solid tumor comprising administering to a subject in need thereof RNA encoding an IL-12sc protein, RNA encoding an IL-15 sushi protein, RNA encoding an IFNα protein, and RNA encoding a GM-CSF protein, wherein the RNA is administered via intra- or peri-tumoral injection into a first tumor, and wherein the injected first tumor and a non-injected second tumor are both reduced in size.