Rna formulation for immunotherapy

Abstract

The present invention is in the field of immunotherapy, in particular tumor immunotherapy. The present invention provides pharmaceutical formulations for delivering RNA to antigen presenting cells such as dendrite cells (DCs) in the spleen after systemic administration. In particular, the formulations described herein enable to induce an immune response after systemic administration of antigen-coding RNA.

1 - 53 . (canceled) 
     
     
         54 . A method for inducing an immune response in a subject comprising administering to the subject a pharmaceutical composition comprising nanoparticles that comprise:
 (i) at least one cationic lipid, and   (ii) at least one RNA molecule encoding at least one antigen, wherein:
 at physiological pH, an overall charge ratio of positive charges to negative charges of the nanoparticles is between 1:1.2 and 1:2, wherein the positive charges are contributed by the at least one cationic lipid, and the negative charges are contributed by the at least one RNA molecule; and 
 the at least one antigen is a disease-associated antigen, whereby a specific immune response against a disease-associated antigen or cells expressing a disease-associated antigen is elicited. 
   
     
     
         55 . The method of  claim 54 , wherein the disease-associated antigen is an antigen of an infectious disease or a tumor antigen. 
     
     
         56 . The method of  claim 55 , wherein the infectious disease is caused by an infection of a pathogen selected from the group consisting of human immunodeficiency virus, hepatitis A, hepatitis B, hepatitis C, herpes, herpes zoster, rubella, flaviviruses, influenza, Marburg viruses, Ebola viruses,  Legionella, Helicobacter , Vibrio, E.  coli ,  Staphylococci ,  Salmonella ,  Streptococci , Plasmodium, Trypanosoma, Leishmania, Toxoplasma,  Cryptococcus neoformans ,  Histoplasma capsulatum ,  Coccidioides immitis ,  Blastomyces dermatitidis  and  Candida albicans . 
     
     
         57 . The method of  claim 55 , wherein the disease-associated antigen is a tumor antigen. 
     
     
         58 . The method of  claim 57 , wherein the tumor antigen is selected from the group consisting of p53, ART-4, BAGE, beta-catenin/m, Bcr-abL CAMEL, CAP-1, CASP-8, CDC27/m, CDK4/m, CEA, CLAUDIN-6, CLAUDIN-18.2, CLAUDIN-12, c-MYC, CT, Cyp-B, DAM, ELF2M, ETV6-AML1, G250, GAGE, GnT-V, Gap100, HAGE, HER-2/neu, HPV-E7, HPV-E6, HAST-2, hTERT, LAGE, LDLR/FUT, MAGE-A, MAGE-Al, MAGE-A2, MAGE-A3, MAGE-A4, MAGEA5, MAGE-A6, MAGE-A7, MAGE-A8, MAGE-A9, MAGE-A1 0, MAGE-A11, MAGEA12, MAGE-B, MAGE-C, MART-1/Melan-A, MC1R, Myosin/m, MUC1, MUM-1, MUM-2, MUM-3, NA88-A, NF1, NY-ESO-1, NY-BR-1, p190 minor BCR-abL, Pml/RARa, PRAME, proteinase 3, PSA, PSM, RAGE, RU1, RU2, SAGE, SART-1, SART-3, SCGB3A2, SCP1, SCP2, SCP3, SSX, SURVIVIN, TEL/AML1, TPI/m, TRP-1, TRP-2, TRP-2/INT2, TPTE WT, and WT-1. 
     
     
         59 . The method of  claim 54 , wherein the specific immune response is an immune response against cancer. 
     
     
         60 . The method of  claim 59 , wherein the cancer is selected from the group consisting of leukemias, seminomas, melanomas, teratomas, lymphomas, neuroblastomas, gliomas, rectal cancer, endometrial cancer, kidney cancer, adrenal cancer, thyroid cancer, blood cancer, skin cancer, cancer of the brain, cervical cancer, intestinal cancer, liver cancer, colon cancer, stomach cancer, intestine cancer, head and neck cancer, gastrointestinal cancer, lymph node cancer, esophagus cancer, colorectal cancer, pancreas cancer, ear, nose and throat (ENT) cancer, breast cancer, prostate cancer, cancer of the uterus, ovarian cancer, lung cancer, and the metastases thereof. 
     
     
         61 . The method of  claim 60 , wherein the cancer is melanoma. 
     
     
         62 . The method of  claim 60 , wherein the cancer is pancreas cancer. 
     
     
         63 . The method of  claim 54 , wherein the at least one RNA molecule is or comprises mRNA. 
     
     
         64 . The method of  claim 54 , wherein the at least one cationic lipid is or comprises 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA) and/or 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). 
     
     
         65 . The method of  claim 54 , wherein the nanoparticles further comprise at least one helper lipid. 
     
     
         66 . The method of  claim 65 , wherein at physiological pH, an overall charge ratio of positive charges to negative charges of the nanoparticles is about 1.3:2, wherein the positive charges are contributed by the at least one cationic lipid, and the negative charges are contributed by the at least one RNA molecule;
 the at least one cationic lipid is 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA);   the at least one helper lipid is 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE); and   a molar ratio of DOTMA to DOPE is about 2:1.   
     
     
         67 . The method of  claim 65 , wherein at physiological pH the overall charge ratio of positive charges to negative charges of the nanoparticles is 1.3:2, wherein the positive charges are contributed by the at least one cationic lipid, and the negative charges are contributed by the at least one RNA molecule;
 the nanoparticles have an average diameter in the range of from about 300 nm to about 500 nm as measured by dynamic light scattering; and   the at least one cationic lipid is 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA);   the at least one helper lipid is 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE); and   the molar ratio of DOTMA to DOPE is about 2:1.   
     
     
         68 . The method of  claim 65 , wherein at physiological pH the overall charge ratio of positive charges to negative charges of the nanoparticles is between 1:1.2 and 1:2;
 the nanoparticles have an average diameter in the range of from about 250 nm to about 550 nm as measured by dynamic light scattering; and   the molar ratio of the at least one cationic lipid to the at least one helper lipid is from 9:1 to 3:7.   
     
     
         69 . A method for stimulating, priming and/or expanding T-cells in a subject comprising administering to the subject a pharmaceutical composition comprising nanoparticles that comprise:
 (i) at least one cationic lipid, and   (ii) at least one RNA molecule encoding at least one antigen, wherein:
 at physiological pH, an overall charge ratio of positive charges to negative charges of the nanoparticles is between 1:1.2 and 1:2, wherein the positive charges are contributed by the at least one cationic lipid, and the negative charges are contributed by the at least one RNA molecule; and 
 wherein the antigen elicits an immune response against a disease-associated antigen or cells expressing a disease-associated antigen, whereby T-cells in the subject are stimulated, primed, and/or expanded. 
   
     
     
         70 . The method of  claim 69 , wherein the disease-associated antigen is an antigen of an infectious disease or a tumor antigen. 
     
     
         71 . The method of  claim 70 , wherein the infectious disease is caused by an infection of a pathogen selected from the group consisting of human immunodeficiency virus, hepatitis A, hepatitis B, hepatitis C, herpes, herpes zoster, rubella, flaviviruses, influenza, Marburg viruses, Ebola viruses,  Legionella ,  Helicobacter ,  Vibrio , E.  coli ,  Staphylococci ,  Salmonella ,  Streptococci , Plasmodium, Trypanosoma, Leishmania, Toxoplasma, Cryptococcus neoformans, Histoplasma capsulatum,  Coccidioides immitis ,  Blastomyces dermatitidis  and  Candida albicans . 
     
     
         72 . The method of  claim 70 , wherein the disease-associated antigen is a tumor antigen. 
     
     
         73 . The method of  claim 72 , wherein the tumor antigen is selected from the group consisting of p53, ART-4, BAGE, beta-catenin/m, Bcr-abL CAMEL, CAP-1, CASP-8, CDC27/m, CDK4/m, CEA, CLAUDIN-6, CLAUDIN-18.2, CLAUDIN-12, c-MYC, CT, Cyp-B, DAM, ELF2M, ETV6-AML1, G250, GAGE, GnT-V, Gap100, HAGE, HER-2/neu, HPV-E7, HPV-E6, HAST-2, hTERT, LAGE, LDLR/FUT, MAGE-A, MAGE-Al, MAGE-A2, MAGE-A3, MAGE-A4, MAGEA5, MAGE-A6, MAGE-A7, MAGE-A8, MAGE-A9, MAGE-A1 0, MAGE-A11, MAGEA12, MAGE-B, MAGE-C, MART-1/Melan-A, MC1R, Myosin/m, MUC1, MUM-1, MUM-2, MUM-3, NA88-A, NF1, NY-ESO-1, NY-BR-1, p190 minor BCR-abL, Pml/RARa, PRAME, proteinase 3, PSA, PSM, RAGE, RU1, RU2, SAGE, SART-1, SART-3, SCGB3A2, SCP1, SCP2, SCP3, SSX, SURVIVIN, TEL/AML1, TPI/m, TRP-1, TRP-2, TRP-2/INT2, TPTE WT, and WT-1. 
     
     
         74 . The method of  claim 69 , wherein the immune response is an immune response against cancer. 
     
     
         75 . The method of  claim 74 , wherein the cancer is selected from the group consisting of leukemias, seminomas, melanomas, teratomas, lymphomas, neuroblastomas, gliomas, rectal cancer, endometrial cancer, kidney cancer, adrenal cancer, thyroid cancer, blood cancer, skin cancer, cancer of the brain, cervical cancer, intestinal cancer, liver cancer, colon cancer, stomach cancer, intestine cancer, head and neck cancer, gastrointestinal cancer, lymph node cancer, esophagus cancer, colorectal cancer, pancreas cancer, ear, nose and throat (ENT) cancer, breast cancer, prostate cancer, cancer of the uterus, ovarian cancer, lung cancer, and the metastases thereof. 
     
     
         76 . The method of  claim 75 , wherein the cancer is melanoma. 
     
     
         77 . The method of  claim 75 , wherein the cancer is pancreas cancer. 
     
     
         78 . The method of  claim 69 , wherein the at least one RNA molecule is or comprises mRNA. 
     
     
         79 . The method of  claim 69 , wherein the at least one cationic lipid is or comprises 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA) and/or 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). 
     
     
         80 . The method of  claim 69 , wherein the nanoparticles further comprise at least one helper lipid. 
     
     
         81 . The method of  claim 80 , wherein at physiological pH, an overall charge ratio of positive charges to negative charges of the nanoparticles is about 1.3:2, wherein the positive charges are contributed by the at least one cationic lipid, and the negative charges are contributed by the at least one RNA molecule;
 the at least one cationic lipid is 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA);   the at least one helper lipid is 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE); and   a molar ratio of DOTMA to DOPE is about 2:1.   
     
     
         82 . The method of  claim 80 , wherein at physiological pH the overall charge ratio of positive charges to negative charges of the nanoparticles is 1.3:2, wherein the positive charges are contributed by the at least one cationic lipid, and the negative charges are contributed by the at least one RNA molecule;
 the nanoparticles have an average diameter in the range of from about 300 nm to about 500 nm as measured by dynamic light scattering; and   the at least one cationic lipid is DOTMA and the at least one helper lipid is DOPE, and the molar ratio of DOTMA to DOPE is about 2:1.   
     
     
         83 . The method of  claim 80 , wherein at physiological pH the overall charge ratio of positive charges to negative charges of the nanoparticles is between 1:1.2 and 1:2;
 the nanoparticles have an average diameter in the range of from about 250 nm to about 550 nm as measured by dynamic light scattering; and   the molar ratio of the at least one cationic lipid to the at least one helper lipid is from 9:1 to 3:7.