Cancer treatment compositions and uses thereof

Abstract

The present disclosure provides compositions, methods of treatment, and methods of making and using compositions for delivering nucleic acid to a subject. The compositions described herein include lipid carriers, optionally including inorganic particles, that can be mixed with nucleic acid. The nucleic acids provided herein include those that code for cancer antigens (full length proteins or fragments) and antibodies. Methods of using the compositions as therapeutic vaccines for the treatment of cancer are also provided. Cancer is one of the most difficult human diseases to treat or prevent.

Description

[Technical field]

[0001] cross reference This application claims the benefit of priority to U.S. Provisional Patent Application No. 63 / 247,167, filed September 22, 2021, and U.S. Provisional Patent Application No. 63 / 302,360, filed January 24, 2022, the contents of each of which are incorporated by reference herein in their entirety. [Background technology]

[0002] background Cancer is one of the most difficult human diseases to treat or prevent. Cancer cells evade the immune system by suppressing immune cell signaling, avoiding programmed cell death, and becoming resistant to and adapting to chemotherapeutic agents. Cancer cells evade the immune system by behaving as "self" and producing tolerizing signals that are not recognized by the immune system. In addition, solid cancer cells form tight junctions in affected organs that resist the delivery of chemotherapeutic agents and cell therapy, impairing their effectiveness. Thus, there is a need for improved compositions and methods for the prevention and treatment of a wide range of cancers. Summary of the Invention [Means for solving the problem]

[0003] A quick overview Provided herein is a composition comprising a lipid nanoparticle comprising a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer-associated protein region comprising a cell membrane contacting domain or a functional fragment thereof. Further provided herein is a composition in which the cell membrane contacting domain comprises a transmembrane binding domain, an outer cell membrane contacting domain, or an inner cell membrane contacting domain. Further provided herein is a composition in which the cancer-associated protein is a prostein. Further provided herein is a composition in which the at least one nucleic acid comprises a region encoding a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 90. Further provided herein is a composition in which the at least one nucleic acid comprises a region encoding a sequence of SEQ ID NO: 90. Further provided herein is a composition in which the cancer-associated protein is a protein expressed by a solid cancer cell or a blood cancer cell. Further provided herein is a composition in which the blood cancer cell comprises a melanoma cancer cell, a prostate cancer cell, a colon cancer cell, an ovarian cancer cell, a breast cancer cell, or a pancreatic cancer cell. Further provided herein is a composition in which at least one nucleic acid is in a complex with a lipid nanoparticle to form a nucleic acid-lipid nanoparticle complex. Further provided herein is a composition in which at least one nucleic acid further comprises a sequence encoding an RNA-dependent polymerase. Further provided herein is a composition comprising a second nucleic acid encoding an RNA-dependent polymerase. Further provided herein is a composition in which the RNA-dependent polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition in which the sequence encoding the RNA-dependent polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition in which the lipid nanoparticle comprises a hydrophobic core. Further provided herein is a composition in which the lipids present in the hydrophobic core are in a liquid phase at 25 degrees Celsius. Further provided herein is a composition in which the lipid nanoparticle comprisesFurther provided herein is a composition characterized as having a z-average diameter particle size measurement of about 20 nm to about 80 nm as measured using dynamic light scattering. Further provided herein is a composition wherein the hydrophobic core comprises a liquid oil. Further provided herein is a composition wherein the liquid oil is alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition wherein the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglyceryl myristate. Cationic lipids include 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyltrimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N-trimethylammonium, chloride (DOTAP). TMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10,Tetrakis(8-methylnonyl) 3,3',3'',3'''-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate; 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl) Bis(hexane-6,1-diyl)bis(2-hexyldecanoate);ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide;β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol;BAME-O1 6B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl)o (oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE,1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9''''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanoate; Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo Further provided herein are compositions in which the aryl group is N,N-tris(3-(diphenylphosphine)amino)-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9'''Z,12Z,12'Z,12''Z,12'''Z)-tetrakis(octadeca-9,12-dienoate); PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition in which the lipid nanoparticle comprises an inorganic particle. Further provided herein is a composition in which the inorganic particle is within a hydrophobic core. Further provided herein is a composition in which the inorganic particle comprises a metal. Further provided herein is a composition in which the metal comprises a metal salt, a metal oxide, a metal hydroxide, or a metal phosphate. Further provided herein is a composition in which the metal oxide comprises aluminum oxide, aluminum oxyhydroxide, iron oxide, titanium dioxide, or silicon dioxide. Further provided herein is a composition in which the hydrophobic surfactant is sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, or sorbitan trioleate. Further provided herein is a composition in which the hydrophilic surfactant is a polysorbate.Compositions are further provided herein.

[0004] Provided herein is a composition comprising a lipid nanoparticle, the lipid nanoparticle comprising a surface comprising a cationic lipid; and a hydrophobic core; and a nucleic acid, the nucleic acid comprising a sequence encoding a TRP-1 protein or a functional fragment thereof, the nucleic acid complexing with the cationic lipid to form a nucleic acid-lipid nanoparticle complex. Further provided herein is a composition, the nucleic acid comprising a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO:2. Further provided herein is a composition, the nucleic acid comprising a sequence of SEQ ID NO:2. Further provided herein is a composition, the nucleic acid comprising a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO:76. Further provided herein is a composition, the nucleic acid comprising a sequence of SEQ ID NO:76. Further provided herein is a composition, the nucleic acid encoding an amino acid sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO:78, or a functional fragment thereof. Further provided herein is a composition, wherein the nucleic acid encodes an amino acid sequence comprising SEQ ID NO: 78, or a functional fragment thereof. Further provided herein is a composition, further comprising a nucleic acid encoding an RNA polymerase. Further provided herein is a composition, wherein the nucleic acid further comprises a sequence encoding an RNA polymerase. Further provided herein is a composition, wherein the RNA polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the sequence encoding the RNA polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition, wherein the lipids present in the hydrophobic core are in a liquid phase at 25 degrees Celsius. Further provided herein is a composition, wherein the lipid nanoparticles are characterized as having a z-average diameter particle size measurement of about 20 nm to about 80 nm, as measured using dynamic light scattering. Further provided herein is a composition, wherein the hydrophobic core comprises a liquid oil. Liquid oils include α-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil,Further provided herein is a composition comprising a triglyceride, or vitamin E. Further provided herein is a composition wherein the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerol myristate. Further provided herein is a composition wherein the cationic lipid is 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl 3-trimethylammoniumpropane (DMTAP), dipalmitoyl (C16:0) trimethylammoniumpropane (DPTAP), distearoyltrimethylammoniumpropane (DPTAP ... ammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DODAP). propane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'''-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropane onate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate); ALC-0159,2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide; β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol; BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl)3,3'-((3-methyl-9-oxo- 10-Oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl)dipropionate;BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl)oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-amini cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleoyl oxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9'''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanonanoate; lipid H (SM-102),Heptadecane-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9'''Z,12Z,12'Z,12''Z,12 Further provided herein is a composition comprising (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3; or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition wherein the lipid nanoparticle comprises an inorganic particle. Further provided herein is a composition wherein the inorganic particle is within a hydrophobic core. Further provided herein is a composition wherein the inorganic particle comprises a metal. Further provided herein is a composition wherein the metal comprises a metal salt, a metal oxide, a metal hydroxide, or a metal phosphate. Further provided herein is a composition wherein the metal oxide comprises aluminum oxide, aluminum oxyhydroxide, iron oxide, titanium dioxide, or silicon dioxide. Further provided herein is a composition further comprising a hydrophobic surfactant and a hydrophilic surfactant. Further provided herein is a composition wherein the hydrophobic surfactant comprises sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, or sorbitan trioleate. Further provided herein is a composition wherein the hydrophilic surfactant comprises a polysorbate.

[0005] Provided herein is a composition comprising a lipid nanoparticle, the lipid nanoparticle comprising a surface comprising a cationic lipid; and a hydrophobic core; and a nucleic acid, the nucleic acid comprising a sequence encoding a MAGE-A1 protein or a functional fragment thereof, the nucleic acid complexed with the cationic lipid to form a nucleic acid-lipid nanoparticle complex. Further provided herein is a composition, the nucleic acid comprises a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 1. Further provided herein is a composition, the nucleic acid comprises a sequence of SEQ ID NO: 1. Further provided herein is a composition, the nucleic acid comprises a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 88. Further provided herein is a composition, the nucleic acid comprises a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 75. Further provided herein is a composition, wherein the nucleic acid comprises a sequence of SEQ ID NO: 75. Further provided herein is a composition, wherein the nucleic acid comprises a sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 89. Further provided herein is a composition, wherein the nucleic acid comprises a sequence of SEQ ID NO: 89. Further provided herein is a composition, wherein the nucleic acid encodes an amino acid sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 77, or a functional fragment thereof. Further provided herein is a composition, wherein the nucleic acid encodes an amino acid sequence comprising SEQ ID NO: 77, or a functional fragment thereof. Further provided herein is a composition, wherein the nucleic acid encodes an amino acid sequence at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to SEQ ID NO: 87, or a functional fragment thereof. Further provided herein is a composition, wherein the nucleic acid encodes an amino acid sequence comprising ... RNA polymerase. Further provided herein is a composition, wherein the nucleic acid further comprises a sequence encoding an RNA polymerase. The RNA polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase.Further provided herein is a composition. Further provided herein is a composition, wherein the sequence encoding the RNA polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition, wherein the lipids present in the hydrophobic core are in a liquid phase at 25 degrees Celsius. Further provided herein is a composition, wherein the lipid nanoparticles are characterized as having a z-average diameter particle size measurement of about 20 nm to about 80 nm as measured using dynamic light scattering. Further provided herein is a composition, wherein the hydrophobic core comprises a liquid oil. Further provided herein is a composition, wherein the liquid oil comprises alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition in which the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerol myristate. The cationic lipid is selected from the group consisting of 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyltrimethylammonium propane (DSAP), and the like. propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N-trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA).1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'''-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso 5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate); ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide; β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17 -((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol; BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl)3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl)dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,1 0R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl)oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol,3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamide) ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate;DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine;ePC, ethylphosphatidylcholine;FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9'''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azane triyl)) hexanoate; Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl)(9Z,9'Z,9''Z,9''''Z,12 Further provided herein is a composition comprising Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate; PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition wherein the lipid nanoparticle comprises an inorganic particle. Further provided herein is a composition wherein the inorganic particle is within a hydrophobic core. Further provided herein is a composition wherein the inorganic particle comprises a metal. Further provided herein is a composition wherein the metal comprises a metal salt, a metal oxide, a metal hydroxide, or a metal phosphate. Further provided herein is a composition wherein the metal oxide comprises aluminum oxide, aluminum oxyhydroxide, iron oxide, titanium dioxide, or silicon dioxide.Further provided herein is a composition. Further provided herein is a composition comprising a hydrophobic surfactant and a hydrophilic surfactant. Further provided herein is a composition in which the hydrophobic surfactant comprises sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, or sorbitan trioleate. Further provided herein is a composition in which the hydrophilic surfactant comprises a polysorbate.

[0006] Provided herein is a composition comprising a nucleic acid, the nucleic acid comprising a sequence encoding a protein or functional fragment thereof listed in Table 1; and an RNA polymerase complex region. Further provided herein is a composition, wherein the protein or functional fragment thereof is a cancer-associated protein. Further provided herein is a composition, wherein the protein or functional fragment thereof comprises an amino acid sequence referred to in Table 1. Further provided herein is a composition, wherein the protein or functional fragment thereof comprises an antibody or functional fragment thereof. Further provided herein is a composition, wherein the antibody or functional fragment thereof comprises an antibody listed in Table 2. Further provided herein is a composition, wherein the antibody comprises an immunoglobulin (Ig) molecule or functional fragment thereof. Further provided herein is a composition, wherein the immunoglobulin molecule is an immunoglobulin molecule of an IgG, IgE, IgM, IgD, IgA or IgY isotype, or a functional fragment thereof. Further provided herein is a composition, wherein the immunoglobulin molecule of the functional fragment comprises at least a fragment of an immunoglobulin molecule of IgG1, IgG2, IgG3, IgG4, IgGA1, or IgGA2 subclass. Further provided herein is a composition, wherein the antibody or functional fragment thereof specifically binds to a tumor antigen or a viral antigen. Further provided herein is a composition, wherein the antibody or functional fragment thereof is atezolizumab, avelumab, bevacizumab, cemiplimab, cetuximab, daratumumab, dinutuximab, durvalumab, elotuzumab, ipilimumab, isatuximab, mogamulizumab, necitumumab, nivolumab, obinutuzumab, ofatumumab, olaratumab, panitumumab, pembrolizumab, pertuzumab, ramucirumab, rituximab, or trastuzumab. Further provided herein is a composition, wherein the RNA polymerase complex region is downstream of a subgenomic promoter derived from an alphavirus. Further provided herein is a composition, wherein the RNA polymerase complex region encodes an RNA-dependent RNA polymerase. Further provided herein is a composition, wherein the RNA-dependent RNA polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase.Further provided herein is a composition, wherein the sequence encoding the RNA polymerase complex region comprises SEQ ID NO: 71. Further provided herein is a composition, further comprising lipid nanoparticles for complexing with nucleic acid. Further provided herein is a composition, wherein the composition is lyophilized. Further provided herein is a composition, wherein the composition is in the form of a liquid, semi-liquid, solution, spray, or powder. Further provided herein is a composition, wherein the composition is formulated as a suspension.

[0007] Provided herein is a pharmaceutical composition comprising the composition described herein and a pharma- ceutical acceptable excipient.In some embodiments, the pharma-ceutical acceptable excipient comprises water.In some embodiments, the pharma-ceutical acceptable excipient comprises sugar.In some embodiments, the sugar comprises sucrose.

[0008] Provided herein is a method of generating an immune response in a subject, comprising administering to the subject a composition as described herein or a pharmaceutical composition as described herein, thereby generating an immune response against a cancer-associated protein. Provided herein further is a method in which the composition is administered to the subject over at least two doses. Provided herein further is a method in which the at least two doses are administered at least about 28 days apart. Provided herein further is a method in which up to 5 μg, 10 ug, 25 ug, or more of the nucleic acid is present in the composition administered to the subject. Provided herein further is a method in which the composition is administered via intramuscular, intranasal, oral, subcutaneous, intratumoral, intrathecal, or intravenous injection. Provided herein further is a method in which the subject is a livestock or farm animal. Provided herein further is a method in which the subject is a mammal. Provided herein further is a method in which the subject is a human. Provided herein further is a method in which the subject has, is at risk of, or is suspected of having cancer. Further provided herein is a method in which the subject has a solid tumor or a blood cancer. Further provided herein is a method in which the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a method in which the blood cancer is a lymphoma or leukemia. Further provided herein is a method in which the subject has, is at risk of developing, or is suspected of having skin cancer. Further provided herein is a method in which the skin cancer is basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a method in which the subject has, is at risk of developing, or is suspected of having pancreatic cancer. Further provided herein is a method in which the pancreatic cancer is pancreatic adenocarcinoma, pancreatic exocrine carcinoma, pancreatic neuroendocrine carcinoma, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. Further provided herein is a method in which the subject has, is at risk of developing, or is suspected of having colon cancer, prostate cancer, ovarian cancer, or breast cancer.Further provided herein are methods, wherein the cancer expresses TRP-1 protein, a prostein protein, a MAGE-A1 protein, a MAGE-A3 protein, or a combination thereof.

[0009] Provided herein is a method of treating cancer in a subject, comprising receiving a biomarker report indicating that the subject has cancer; classifying the cancer based on the biomarker report; and administering to the subject a composition or pharmaceutical composition described herein. Provided herein is a method in which the composition is administered to the subject in at least two doses. Provided herein is a method in which the at least two doses are administered at least about 28 days apart. Provided herein is a method in which up to 5 μg, 10 ug, 25 ug, or more of nucleic acid is present in the composition administered to the subject. Provided herein is a method in which the composition is administered via intramuscular, intranasal, oral, subcutaneous, intratumoral, intrathecal, or intravenous injection. Provided herein is a method in which the subject is a domestic or farm animal. Provided herein is a method in which the subject is a mammal. Provided herein is a method in which the subject is a human. Provided herein is a method in which the cancer is a solid cancer or a blood cancer. Further provided herein are methods, wherein the cancer expresses TRP-1 protein, a prostein protein, a MAGE-A1 protein, a MAGE-A3 protein, or a combination thereof.

[0010] Provided herein are compositions comprising a lipid carrier, the lipid carrier comprising a liquid oil; and a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer-associated protein.

[0011] Further provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; and a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding an antibody or functional variant thereof.

[0012] Further provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; and a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer therapeutic antibody or functional variant thereof.

[0013] Further provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; inorganic nanoparticles, the inorganic nanoparticles comprising iron oxide present in an amount of about 0.2 mg / ml of 12 nm iron oxide; and a surfactant, the surfactant comprising a cationic lipid; and at least one nucleic acid, the nucleic acid comprising at least one sequence encoding a cancer associated protein sequence or a functional variant thereof.

[0014] Further provided herein is a composition comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising about 30 mg / mL DOTAP chloride; about 37.5 mg / ml squalene; about 37 mg / ml sorbitan monostearate; about 37 mg / ml polysorbate 80; about 10 mM sodium citrate; and about 0.2 mg Fe / ml 12 nm oleic acid coated iron oxide nanoparticles; and (b) at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer associated protein sequence or a functional variant thereof.

[0015] Further provided herein is a composition comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising: DOTAP chloride present in an amount of about 0.75 mg; squalene present in an amount of about 0.94 mg; sorbitan monostearate present in an amount of about 0.93 mg; polysorbate 80 present in an amount of about 0.93 mg; citric acid monohydrate present in an amount of about 1.05 mg; and oleic acid coated iron oxide nanoparticles present in an amount of about 0.005 mg; and (b) at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer associated protein sequence or a functional variant thereof.

[0016] Further provided herein is a composition comprising: a first nucleic acid comprising a sequence encoding an RNA-dependent RNA polymerase; and a second nucleic acid comprising a sequence encoding a cancer associated protein sequence or a functional variant thereof.

[0017] Further provided herein is a composition comprising: a first nucleic acid comprising a sequence encoding an RNA-dependent RNA polymerase; and a second nucleic acid comprising a sequence encoding a cancer associated protein-binding antibody or antibody fragment.

[0018] Further provided herein is a composition comprising: (a) a lipid carrier, wherein the lipid carrier is a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles; and one or more lipids; and (b) at least one nucleic acid sequence, wherein the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, wherein the antigen is a cancer-associated protein.

[0019] Further provided herein is a vaccine comprising: (a) a lipid carrier, wherein the lipid carrier is a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles, and one or more lipids; and (b) at least one nucleic acid sequence, wherein the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, wherein the antigen is a cancer-associated protein.

[0020] Further provided herein is a method of generating an immune response in a subject, the method comprising administering to the subject a composition comprising a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids, and at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding a sequence capable of expressing an antigen, wherein the antigen is a cancer-associated protein.

[0021] Further provided herein is a composition for immunoprotecting a subject, the composition comprising a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; and at least one nucleic acid sequence, where the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, and the antigen is a cancer-associated protein.

[0022] Further provided herein is a dry composition comprising the composition provided herein; and at least one cryoprotectant.

[0023] Further provided herein is a composition for the prevention of cancer, comprising a sorbitan fatty acid ester, an ethoxylated sorbitan ester, a cationic lipid, an immunostimulant, and at least one RNA encoding an antigen sequence or a functional fragment thereof.

[0024] Further provided herein is a composition for the prevention of cancer, the composition comprising sorbitan monostearate (e.g., SPAN® 60), polysorbate 80 (e.g., TWEEN® 80), DOTAP, an immune stimulant, and at least one RNA encoding an antigen sequence, or a functional fragment thereof.

[0025] Further provided herein is a pharmaceutical composition comprising the composition provided herein; and a pharma- ceutically acceptable excipient.Further provided herein is a pharmaceutical composition comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles; and one or more lipids; and (b) at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding a sequence capable of expressing an antigen, the antigen being a cancer-associated protein.

[0026] Further provided herein are kits that include the compositions provided herein.

[0027] Further provided herein is a method for generating an immune response in a subject, comprising administering to the subject a composition as provided herein, thereby generating an immune response against a cancer-associated protein. Further provided herein is a method for generating an immune response in a subject, comprising administering to the subject (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles, and one or more lipids; and (b) at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding a sequence capable of expressing an antigen, the antigen being a cancer-associated protein.

[0028] Further provided herein is a method of prophylactically immunizing a subject against cancer, the method comprising administering to the subject a composition provided herein, thereby immunizing the subject against a cancer that expresses a cancer-associated protein.

[0029] Further provided herein is a method of reducing the severity of cancer, the method comprising administering a composition comprising a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles, and one or more lipids, and at least one nucleic acid sequence, where the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, and the antigen is a cancer-associated protein.

[0030] Further provided herein is a method of immunoprotecting a subject, the method comprising administering to the subject a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids, at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding a sequence capable of expressing an antigen, wherein the antigen is a cancer-associated protein.

[0031] Further provided herein is a method for preparing a lyophilized composition, the method comprising the steps of obtaining a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, where the nucleic acid has a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; and lyophilizing the formulation to form a lyophilized composition.

[0032] Further provided herein is a method for preparing a spray-dried composition, the method comprising the steps of obtaining a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, where the nucleic acid has a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; and spray drying the formulation to form the spray-dried composition.

[0033] The present invention also relates to a method for reconstituting a lyophilized composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, the nucleic acid having a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; lyophilizing the formulation to form a lyophilized composition; and reconstituting the lyophilized composition in a suitable diluent.

[0034] Further provided herein is a method for reconstituting a spray-dried composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, the nucleic acid having a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; spray drying the formulation to form a spray-dried composition; and reconstituting the spray-dried composition in a suitable diluent.

[0035] Additional features of the present invention will become apparent to those skilled in the art in view of the following disclosure and the accompanying drawings and claims.

[0036] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present disclosure, which can be better understood by reference to the drawings in combination with the detailed description of specific embodiments presented herein. [Brief description of the drawings]

[0037] [Figure 1-1]1A-1H show schematic diagrams of exemplary nanoparticle (NP) carriers. FIG. 1A shows an oil-in-water emulsion. FIG. 1B shows a nanostructured lipid carrier (NLC). FIG. 1C shows a nanoparticle with inorganic nanoparticles in liquid oil. FIG. 1D shows a nanoparticle with a cationic lipid membrane and a liquid oil core. FIG. 1E shows an oil-in-water emulsion with two or more RNA or DNA molecules. FIG. 1F shows a nanostructured lipid carrier (NLC) with two or more RNA or DNA molecules. FIG. 1G shows a nanoparticle with inorganic nanoparticles in liquid oil, two or more RNA or DNA molecules. FIG. 1H shows a nanoparticle with a cationic lipid membrane, a liquid oil core, and two or more RNA or DNA molecules. The figures are not to scale. [Figure 1-2] Same as above. [Figure 1-3] Same as above. [Figure 1-4] Same as above.

[0038] [Figure 2A] Figures 2A-2B show the increase in protein production induced by NP-3, a Miglyol lipid carrier formulation. Figure 2A shows the first assay. Figure 2B shows the second assay. [Figure 2B] Same as above.

[0039] [Figure 3A] Figures 3A-3B show the reduction in immune response induced by NP-3, a Miglyol lipid carrier formulation. Figure 3A shows the first assay. Figure 3B shows the second assay. [Figure 3B] Same as above.

[0040] [Figure 4] Figures 4A-4B show the correlation between enhanced protein production observed with NP-3 and low TNF (e.g., TNF alpha) stimulation as a result of the first and second assays. Figure 4A shows the first assay. Figure 4B shows the second assay.

[0041] [Figure 5-1] 5A-5F show SEAP levels in BALB / c mice injected intramuscularly with various embodiments of lipid carrier formulations described herein. FIG. 5A shows SEAP levels at 4 days post-injection. FIG. 5B shows SEAP levels at 6 days post-injection. FIG. 5C shows SEAP levels at 8 days post-injection. FIG. 5D shows SEAP levels at 4 days post-injection. FIG. 5E shows SEAP levels at 6 days post-injection. FIG. 5F shows SEAP levels at 8 days post-injection. X-axis: condition, Y-axis: relative light units (RLU). [Figure 5-2] Same as above. [Figure 5-3] Same as above.

[0042] [Figure 6] FIG. 6 is a bar graph with Z-average and polydispersity index (PDI) measurements on the Y-axis and group number on the X-axis for conditions 1-14.

[0043] [Figure 7-1]7A-7F show the effect of lipid carrier + TRP-1 replicon vaccine composition on tumor volume in the B16 tumor model of melanoma. FIG. 7A shows the tumor volume of untreated B16 mice over time. X-axis: time, Y-axis: tumor volume (mm3). FIG. 7B shows the tumor volume of B16 mice over time for mice treated with lipid carrier + 0.2 micrograms (μg) of TRP-1 repRNA on days 0 and 14. X-axis: time, Y-axis: tumor volume (mm3). FIG. 7C shows the tumor volume of B16 mice over time for mice treated with lipid carrier + 0.2 μg of TRP-1 repRNA only on day 0. X-axis: time, Y-axis: tumor volume (mm3). FIG. 7D shows the tumor volume of B16 mice over time for mice treated with lipid carrier + 1 μg of TRP-1 repRNA on days 0 and 14. X-axis: time, Y-axis: tumor volume (mm3). Figure 7E shows the tumor volume of B16 mice over time for mice treated with lipid carrier + 1 μg TRP-1 repRNA only on day 0. X-axis: time, Y-axis: tumor volume (mm3). Figure 7F shows the survival rate of animals over time. X-axis: days after inoculation, Y-axis: percentage of animals alive. [Figure 7-2] Same as above. [Figure 7-3] Same as above.

[0044] [Figure 8] Figure 8 shows the mean B16 tumor volume over time in untreated and lipid carrier + repRNA-TRP-1 vaccinated mice. X-axis: days after implantation, Y-axis: tumor volume (mm3).

[0045] [Figure 9] FIG. 9 shows individual tumor growth for control (scrambled RNA) and animals that received prophylactic lipid carrier plus 1 μg of TRP-1 repRNA vaccine prior to melanoma cell injection.

[0046] [Figure 10-1]Figures 10A-10D show that immunization with MAGE-expressing replicons induces antigen-specific T cells. Figure 10A shows CD8 T cells expressing Tbet and IFNγ. Figure 10B shows % IFNγ positive CD8 T cells in MAGE-treated animals. Figure 10C shows CD4 T cells producing IFNγ. Figure 10D shows CD4 T cells producing IL-2. Flow cytometry plots are representative, but plots show data from individual animals as well as the mean and SEM. [Figure 10-2] Same as above.

[0047] [Figure 11] Figures 11A-11B show graphs of mean tumor volume and survival of animals immunized with replicons encoding MAGE-A3 (SEQ ID NO: 87) and TRP1 (SEQ ID NO: 78). Figure 11A shows mean tumor volume (Y-axis) as a function of days after tumor implantation (X-axis). Figure 11B shows the probability of survival in animals treated with no RNA, TRP-1 / MAGE RNA before palpability, and TRP-1 / MAGE after palpability. X-axis: probability of survival; Y-axis: days after tumor implantation. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0048] Various aspects are now described more fully hereinafter. Such aspects may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein, but rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey its scope to those skilled in the art. Detailed Description of the Invention Provided herein are compositions, kits, methods and uses thereof for inducing immune response against cancer cells or tumors. Briefly, (1) nucleic acids encoding cancer-associated proteins, neoantigens, antibodies and RNA polymerases; (2) nanoparticle carrier systems; (3) combination compositions; (4) thermally stable, dry and freeze-dried cancer vaccines; (5) pharmaceutical compositions; (6) dosing; (7) administration; (8) therapeutic applications; and (9) kits are further described herein. definition

[0049] All definitions, as defined and used herein, should be understood to control for any dictionary definitions, definitions in documents incorporated by reference, and / or ordinary meaning of the defined terms.

[0050] All references, patents and patent applications disclosed herein are incorporated by reference with respect to the subject matter cited, and in some cases may include the entirety of the document.All references disclosed herein, including patent and non-patent literature, are hereby incorporated by reference in their entirety, as if each were individually incorporated.However, when patents, patent applications or publications containing express definitions are incorporated by reference, it should be understood that these express definitions apply to the incorporated patents, patent applications or publications in which they are found, and do not necessarily apply to the text of this application, particularly the claims of this application, and in that case, the definitions provided herein shall mean priority.

[0051] The indefinite articles "a" and "an," as used herein in the specification and the claims, unless clearly dictated to the contrary, should be understood to mean "at least one."

[0052] The term "and / or" as used herein in the specification and claims should be understood to mean "either or both" of the elements so conjoined, i.e., elements that are conjointly present in some cases and disjunctively present in other cases. Multiple elements listed with "and / or" should be interpreted in the same manner, i.e., "one or more" of the elements so conjoined. Other elements, whether related or unrelated to the specifically identified elements, may optionally be present other than the elements specifically identified by the "and / or" clause. Thus, as a non-limiting example, a reference to "A and / or B", when used in conjunction with an open-ended term such as "comprising", may refer in one embodiment to only A (including elements other than B, if desired), in another embodiment to only B (including elements other than A, if desired), in yet another embodiment to both A and B (including other elements, if desired), etc.

[0053] When used herein in the specification and claims, "or" should be understood to have the same meaning as "and / or" defined above. For example, when separating items in a list, "or" or "and / or" should be interpreted as being inclusive, i.e., including at least one of a number or list of elements, but also including two or more, and including additional unlisted items, if necessary. Only terms clearly indicated to the contrary, such as "only one of" or "exactly one of," or, when used in the claims, "consisting of," refer to the inclusion of exactly one element of a number or list of elements. In general, the term "or," when used herein, shall be interpreted merely as indicating exclusive alternatives (i.e., "either one, but not both") when preceded by an exclusive term, such as "either," "one of," "only one of," or "exactly one of." "Consisting essentially of," when used in the claims, shall have its ordinary meaning as used in the field of patent law.

[0054] As used herein, "as needed" or "as needed" means that the situation described thereafter may or may not occur, such that the description includes instances where the situation occurs and instances where the situation does not occur.

[0055] Unless specifically stated or clear from the context, as used herein, the term "about" in reference to a number or range of numbers is understood to mean 20% below the recited lower limit and 20% above the recited upper limit for the stated number or range and + / - 20% of that number or range.

[0056] The term "effective amount" or "therapeutically effective amount" refers to an amount that is sufficient to achieve or at least partially achieve a desired effect. nucleic acid

[0057] Compositions comprising nucleic acids are provided herein. In some embodiments, compositions provided herein comprise one or more types of nucleic acid sequences. In some embodiments, compositions provided herein comprise two or more types of nucleic acid sequences. In some embodiments, compositions provided herein comprise at least one DNA molecule. In some embodiments, compositions provided herein comprise at least one RNA molecule. In some embodiments, compositions provided herein comprise at least one DNA molecule and at least one RNA molecule.

[0058] In some embodiments, the nucleic acid provided herein is in a complex with a nanoparticle provided herein. In some embodiments, the nucleic acid is in a complex with a surface of a nanoparticle. In some embodiments, the nucleic acid is in a complex with a hydrophilic surface of a nanoparticle. In some embodiments, the nucleic acid is located within a nanoparticle. In some embodiments, the nucleic acid is located within a hydrophobic core of a nanoparticle. In some embodiments, the nanoparticle is a lipid carrier nanoparticle, and the surface may be referred to herein as a membrane.

[0059] In some embodiments, the nucleic acids provided herein include deoxyribonucleic acid (DNA), ribonucleic acid (RNA), peptide nucleic acid (PNA), or a combination thereof. The nucleic acid may be linear or may include a secondary structure (e.g., a hairpin). In some embodiments, the nucleic acid is a polynucleotide that includes modified nucleotides or bases, and / or their analogs. The polynucleotide may include modified nucleotides, such as methylated nucleotides and their analogs. If present, modifications to the nucleotide structure may be imparted before or after assembly of the compositions provided herein.Modified nucleobases that can be incorporated into modified nucleosides and nucleotides and present in RNA molecules include m5C (5-methylcytidine), m5U (5-methyluridine), m6A (N6-methyladenosine), s2U (2-thiouridine), Um (2'-O-methyluridine), m1A (1-methyladenosine); m2A (2-methyladenosine); Am (2-1-O-methyladenosine); ms2m6A (2-methylthio-N6-methyladenosine); i6A (N6-isopentenyl adenosine); ms2i6A (2-methylthio -N6-isopentenyl adenosine;io6A(N6-(cis-hydroxyisopentenyl) adenosine);ms2io6A(2-methylthio-N6-(cis-hydroxyisopentenyl) adenosine);g6A(N6-glycinylcarbamoyl adenosine);t6A(N6-threonylcarbamoyl adenosine);ms2t6A(2-methylthio-N6-threonylcarbamoyl adenosine);m6t6A(N6-methyl-N6-threonylcarbamoyl adenosine);hn6A(N6-hydroxynorvalylcarbamoyl adenosine hn6A(2-methylthio-N6-hydroxynorvalylcarbamoyl adenosine);Ar(p)(2'-O-ribosyladenosine (phosphate));I(inosine);m1I(1-methylinosine);m'Im(1,2'-O-dimethylinosine);m3C(3-methylcytidine);Cm(2T-O-methylcytidine);s2C(2-thiocytidine);ac4C(N4-acetylcytidine);f5C(5-phenylcytidine);m5Cm(5,2-O-dimethylcytidine);ac4Cm(N4-acetyl2TOmethylcytidine) cysidine); k2C (lysidine); m1G (1-methylguanosine); m2G (N2-methylguanosine); m7G (7-methylguanosine); Gm (2'-O-methylguanosine); m22G (N2,N2-dimethylguanosine); m2Gm (N2,2'-O-dimethylguanosine); m22Gm (N2,N2,2'-O-trimethylguanosine); Gr(p) (2'-O-ribosylguanosine (phosphate)); yW (ubutosine); o2yW (peroxyubutosine); OHyW (hydroxyubutosine); OHyW. *(unmodified hydroxyubutosine); imG (ubutosine); mimG (methylguanosine); Q (queuosine); oQ (epoxyqueuosine); galQ (galactosyl-queuosine); manQ (mannosyl-queuosine); preQo (7-cyano-7-deazaguanosine); preQi (7-aminomethyl-7-deazaguanosine); G *(Archaeosin);D(Dihydrouridine);m5Um(5,2'-O-Dimethyluridine);s4U(4-Thiouridine);m5s2U(5-Methyl-2-Thiouridine);s2Um(2-Thio-2'-O-Methyluridine);acp3U(3-(3-Amino-3-Carboxypropyl)uridine);hoSU(5-Hydroxyuridine);moSU(5-Methoxyuridine);cmo5U(Uridine 5-Oxyacetic Acid);mcmo5U(Uridine 5-Oxyacetic Acid Methyl Ester);chm5U(5-(Carboxyhydroxymethyl)uridine) uridine);mchm5U(5-(carboxyhydroxymethyl)uridine methyl ester);mcm5U(5-methoxycarbonylmethyluridine);mcm5Um(S-methoxycarbonylmethyl-2-O-methyluridine);mcm5s2U(5-methoxycarbonylmethyl-2-thiouridine);nm5s2U(5-aminomethyl-2-thiouridine);mnm5U(5-methylaminomethyluridine);mnm5s2U(5-methylaminomethyl-2-thiouridine);mnm5se2U(5-methylaminomethyl-2-selenouridine) ncm5U(5-carbamoylmethyluridine); ncm5Um(5-carbamoylmethyl-2'-O-methyluridine); cmnm5U(5-carboxymethylaminomethyluridine); cnmm5Um(5-carboxymethylaminomethyl-2-LO-methyluridine); cmnm5s2U(5-carboxymethylaminomethyl-2-thiouridine); m62A(N6,N6-dimethyladenosine); Tm(2'-O-methylinosine); m4C(N4-methylcytidine); m4Cm(N4,2-O-dimethylcytidine); hm5C (5-hydroxymethylcytidine);m3U(3-methyluridine);cm5U(5-carboxymethyluridine);m6Am(N6,TO-dimethyladenosine);rn62Am(N6,N6,O-2-trimethyladenosine);m2'7G(N2,7-dimethylguanosine);m2'2'7G(N2,N2,7-trimethylguanosine);m3Um(3,2T-O-dimethyluridine);m5D(5-methyldihydrouridine);f5Cm(5-formyl-2'-O-methylcytidine);m1Gm(1,2'-O-dimethylguanosine);m'Am (1,2-O-dimethyladenosine) irinomethyluridine; tm5s2U (S-taurinomethyl-2-thiouridine); imG-14 (4-demethylguanosine); imG2 (isoguanosine); ac6A (N6-acetyladenosine), hypoxanthine, inosine, 8-oxo-adenine, its 7-substituted derivatives, dihydrouracil, pseudouracil, 2-thiouracil, 4-thiouracil, 5-aminouracil, 5-(C1-C6)-alkyluracil , 5-methyluracil, 5-(C2-C6)-alkenyluracil, 5-(C2-C6)-alkynyluracil, 5-(hydroxymethyl)uracil, 5-chlorouracil, 5-fluorouracil, 5-bromouracil, 5-hydroxycytosine, 5-(C1-C6)-alkylcytosine, 5-methylcytosine, 5-(C2-C6)-alkenylcytosine, 5-(C2-C6)-alkynylcytosine, 5-chlorocytosine, 5-fluorocytosine, 5-bromocytosine, N; 2-dimethylguanine, 7-deazaguanine, 8-azaguanine, 7-deaza-7-substituted guanine, 7-deaza-7-(C2-C6)alkynylguanine, 7-deaza-8-substituted guanine, 8-hydroxyguanine, 6-thioguanine, 8-oxoguanine, 2-aminopurine, 2-amino-6-chloropurine, 2,4-diaminopurine, 2,6-diaminopurine, 8-azapurine, substituted 7-deazapurine, 7-deaza-7-substituted purine, 7-deaza-8-substituted purine, hydrogen (abasic residue), m5C, m5U, m6A, s2U, W, or 2'-O-methyl-U. Any one or any combination of these modified nucleobases may be included in the self-replicating RNA of the present invention. Many of these modified nucleobases and their corresponding ribonucleosides are available from commercial suppliers. Optionally, the nucleic acid can contain phosphoramidate, phosphorothioate, and / or methylphosphonate linkages. The RNA sequence can be modified with respect to its codon usage, for example, to increase the translation efficiency and half-life of the RNA. A poly-A tail (e.g., of about 30 or more adenosine residues) can be attached to the 3' end of the RNA to increase its half-life. The 5' end of the RNA can be capped with a modified ribonucleotide or derivative having m7G(5')ppp(5')N (cap 0 structure), which can be incorporated during RNA synthesis or enzymatically engineered after RNA transcription (e.g., by using vaccinia virus capping enzyme (VCE), which consists of mRNA triphosphatase, guanylyl-transferase, and guanine-7-methyltransferase, which catalyzes the construction of the N7-monomethylated cap 0 structure). The cap structure can provide stability and translation efficiency to the RNA molecule. The 5' cap of the RNA molecule may be further modified by 2'-O-methyltransferase resulting in the generation of the Cap 1 structure (m7Gppp[m2'-O]N), which may further increase translation efficiency. The Cap 1 structure may also increase in vivo potency.

[0060] In some embodiments, the nucleic acid provided herein is present in an amount of about 5 ng to about 1 mg. In some embodiments, the nucleic acid provided herein is present in an amount of up to about 25, 50, 75, 100, 150, 175 ng. In some embodiments, the nucleic acid provided herein is present in an amount of up to about 1 mg. In some embodiments, the nucleic acid provided herein is present in an amount of about 0.05 μg, 0.1 μg, 0.2 μg, 0.5 μg, 1 μg, 5 μg, 10 μg, 12.5 μg, 15 μg, 25 μg, 40 μg, 50 μg, 100 μg, 200 μg, 300 μg, 400 μg, 500 μg, 600 μg, 700 μg, 800 μg, 900 μg, 1 mg. In some embodiments, the nucleic acids provided herein are present in an amount of 0.05 μg, 0.1 μg, 0.2 μg, 0.5 μg, 1 μg, 5 μg, 10 μg, 12.5 μg, 15 μg, 25 μg, 40 μg, 50 μg, 100 μg, 200 μg, 300 μg, 400 μg, 500 μg, 600 μg, 700 μg, 800 μg, 900 μg, or 1 mg. In some embodiments, the nucleic acid is at least about 200, 250, 500, 750, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10,000, 11,000, 12,000, 13,000, 14,000, 15,000, 16,000, 17,000, 18,000, 19,000, or 20,000 nucleotides in length. In some embodiments, the nucleic acid is at most about 7000, 8000, 9000, 10,000, 11,000, 12,000, 13,000, 14,000, 15,000, 16,000, 17,000, 18,000, 19,000, or 20,000 nucleotides in length. In some embodiments, the nucleic acid is at most about 7500, 10,000, 15,000, or 20,000 nucleotides in length. Nucleic acids encoding cancer-related proteins

[0061] Provided herein is a composition, the composition comprising: a lipid carrier provided herein; and one or more nucleic acids, the one or more nucleic acids comprising a sequence encoding an antigen. In some embodiments, the antigen is a cancer-associated protein (also referred to as a tumor protein antigen or tumor antigen). In some embodiments, the nucleic acid provided herein encodes a cancer-associated protein. In some embodiments, the cancer-associated protein is a surface protein, a cytosolic protein, or a transmembrane protein. In some embodiments, the cancer-associated protein is a protein expressed by a cancer cell. In some embodiments, the cancer-associated protein is a protein expressed by a cancer-causing microorganism (e.g., a viral protein).

[0062] In some embodiments, the nucleic acids provided herein encode proteins expressed by solid cancer cells or hematological cancer cells. In some embodiments, the solid cancer cells are melanoma cells. In some embodiments, the proteins expressed by melanoma cells are not expressed by non-cancer cells. In some embodiments, the proteins expressed by melanoma cells include a mutation in an amino acid sequence compared to the equivalent amino acid sequence in a non-cancer cell. In some embodiments, the nucleic acids provided herein encode MAGE-A1 (SEQ ID NO: 1) or a functional fragment thereof. In some embodiments, the nucleic acids provided herein encode MAGE-A3 (SEQ ID NO: 87) or a functional fragment thereof. In some embodiments, the nucleic acids provided herein encode TRP-1 (SEQ ID NO: 2) or a functional fragment thereof. In some embodiments, the nucleic acids provided herein encode TRP-1 and MAGE-A1. In some embodiments, the nucleic acids provided herein encode TRP-1 and MAGE-A3. In some embodiments, the nucleic acids provided herein encode tyrosinase. In some embodiments, the nucleic acids provided herein comprise a sequence that is at least 80% identical to SEQ ID NO: 1, 2, 75, 76, 88, or 89. In some embodiments, the nucleic acids provided herein comprise a sequence that is at least 80% identical to SEQ ID NO: 1, 2, 75, 76, 88, or 89. In some embodiments, the nucleic acids provided herein encode an amino acid sequence listed in Table 1. In some embodiments, the nucleic acids provided herein encode an amino acid sequence that is at least 80% identical to SEQ ID NO: 77 or SEQ ID NO: 78. In some embodiments, the nucleic acids provided herein encode an amino acid sequence that is at least 80% identical to SEQ ID NO: 87. In some embodiments, the compositions provided herein comprise two or more, three or more, four or more, five or more, six or more, or up to seven or more nucleic acids encoding different sequences listed in Table 1.In some embodiments, the nucleic acids provided herein that encode a protein sequence listed in Table 1 are used as part of the treatment or prevention of melanoma. In some embodiments, the nucleic acids provided herein encode a cancer associated protein listed in Table 1. In some embodiments, the compositions provided herein comprise two or more nucleic acids that encode different sequences listed in Table 1. In some embodiments, the nucleic acids provided herein encode a cancer associated protein sequence that comprises at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity to a sequence listed in Table 1. In some embodiments, the compositions provided herein comprise two or more nucleic acids that encode different sequences listed in Table 1. In some embodiments, the nucleic acids provided herein encode a cancer associated protein or functional fragment thereof that comprises at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence similarity to a sequence listed in Table 1. The percent sequence identity (%) of a given sequence compared to a reference sequence is defined as the percentage of identical residues identified after aligning the two sequences and introducing gaps as necessary to achieve the maximum percent sequence identity.Percent identity can be calculated using alignment methods known in the art, for example, the alignment of sequences can be carried out using publicly available software such as BLAST, Align, ClustalW2, etc. Those skilled in the art can determine the appropriate parameters for alignment, but the default parameters for BLAST are specifically intended. [Table 1-1] [Table 1-2]

[0063] In some embodiments, the cancer-associated protein encoded by the nucleic acid provided herein comprises a cell membrane contact domain or a functional fragment thereof. In some embodiments, the cell membrane contact domain comprises a transmembrane binding domain, an outer cell membrane contact domain, or an inner cell membrane contact domain. In some embodiments, the cell membrane contact domain comprises a transmembrane binding domain, an outer cell membrane contact domain, and an inner cell membrane contact domain. In some embodiments, the cancer-associated protein is a protein expressed by melanoma cancer cells, prostate cancer cells, colon cancer cells, ovarian cancer cells, breast cancer cells, pancreatic cancer cells, or blood cells.

[0064] In some embodiments, the nucleic acid provided herein comprises a sequence that encodes a dimer, trimer, or multimer of the cancer-associated protein provided herein.In some embodiments, the nucleic acid provided herein comprises a sequence that encodes an amino acid sequence that is at least about 500 amino acids long or longer.In some embodiments, the nucleic acid provided herein comprises a sequence that encodes an amino acid sequence that is at least about 200, 300, 400, 500, 750, 1000 amino acids long or longer. In some embodiments, the nucleic acids provided herein comprise a sequence encoding one or more cancer associated proteins, wherein the one or more cancer associated proteins comprise a molecular weight of at least about 50 kilodaltons (kDa) or higher, at least about 100 kilodaltons (kDa) or higher, at least about 150 kilodaltons (kDa) or higher, at least about 200 kilodaltons (kDa) or higher, at least about 250 kilodaltons (kDa) or higher, at least about 300 kilodaltons (kDa) or higher, at least about 350 kilodaltons (kDa) or higher, at least about 400 kilodaltons (kDa) or higher, at least about 450 kilodaltons (kDa) or higher, at least about 500 kilodaltons (kDa) or higher, up to 1000 kDa or higher. In some embodiments, the nucleic acids provided herein comprise a sequence encoding one or more cancer associated proteins, wherein the one or more cancer associated proteins comprise a molecular weight of at least about 59 kDa or higher.

[0065] In some embodiments, the nucleic acid provided herein comprises a sequence encoding a cancer associated protein associated with prostate cancer. In some embodiments, the cancer associated protein of prostate cancer comprises prostein. In some embodiments, the cancer associated protein is prostein. In some embodiments, the cancer associated protein is at least about 50%, 60%, 70%, 80%, 90%, 95%, or 100% of full-length prostein. In some embodiments, the prostein is human prostein. In some embodiments, the cancer associated protein comprises an amino acid sequence that is at least about 80%, 85%, 90%, 95%, or 100% identical to SEQ ID NO:90. Cancer antigen-binding molecules

[0066] Provided herein is a composition comprising a lipid carrier provided herein; and a nucleic acid encoding an antibody or functional fragment thereof. In some embodiments, the nucleic acid provided herein encodes a monoclonal antibody. In some embodiments, the nucleic acid provided herein encodes a murine antibody, a humanized antibody, or a fully human antibody. In some embodiments, the antibody is an immunoglobulin (Ig) molecule. In some embodiments, the immunoglobulin molecule is an immunoglobulin molecule of an IgG, IgE, IgM, IgD, IgA, or IgY isotype. Further provided herein is a composition, wherein the immunoglobulin molecule is an immunoglobulin molecule of an IgG1, IgG2, IgG3, IgG4, IgGA1, or IgGA2 subclass. In some embodiments, the antibody is a recombinant antibody, a chimeric antibody, or a multivalent antibody. In some embodiments, the multivalent antibody is a bispecific antibody, a trispecific antibody, or a multispecific antibody. In some embodiments, the antibody or functional fragment is an antigen-binding fragment (Fab), Fab2, F(ab'), F(ab')2, dAb, Fc, Fv, disulfide-linked Fv, scFv, tandem scFv, free LC, half antibody, single domain antibody (dAb), diabody, or nanobody.

[0067] In some embodiments, the antibody or functional fragment thereof specifically binds to a cancer-associated protein. In some embodiments, the antibody or functional fragment thereof is a cancer therapeutic antibody. In some embodiments, the cancer therapeutic antibody is atezolizumab, avelumab, bevacizumab, cemiplimab, cetuximab, daratumumab, dinutuximab, durvalumab, elotuzumab, ipilimumab, isatuximab, mogamulizumab, necitumumab, nivolumab, obinutuzumab, ofatumumab, olaratumab, panitumumab, pembrolizumab, pertuzumab, ramucirumab, rituximab, or trastuzumab. The amino acid sequences for the cancer therapeutic antibodies are provided in Table 2 below.

[0068] In some embodiments, the nucleic acids provided herein encode a cancer associated protein or antibody amino acid sequence listed in Table 2, or a functional fragment thereof. In some embodiments, the compositions provided herein comprise two or more nucleic acids encoding different sequences listed in Table 2. In some embodiments, the nucleic acids provided herein encode an antibody amino acid sequence that comprises at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence identity to a sequence listed in Table 2. In some embodiments, the compositions provided herein comprise two or more nucleic acids encoding different sequences listed in Table 2. In some embodiments, the nucleic acids provided herein encode an antibody or functional fragment thereof that comprises at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% sequence similarity to a sequence listed in Table 2. [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5]

[0069] As an alternative or in addition to delivering RNA as an antigen, combinations can be used, for example RNA that stimulates innate immune response, or RNA that triggers oncolytic virus or live attenuated virus, or RNA antigens combined with general agonists that stimulate immune response, including TLR (Toll-like receptor), RLR (RIG-I-like receptor), or NLR (nod-like receptor). In certain embodiments, the compositions provided herein include a combination of RNA that codes for an antigen and another RNA that can stimulate innate immune response or trigger oncolytic virus or live attenuated virus. Alternatively, the compositions provided herein that contain RNA that codes for an antigen can be combined with a formulation that contains another RNA or other immune response agonist that can stimulate innate immune response or trigger oncolytic virus or live attenuated virus. self-replicating nucleic acids

[0070] Compositions comprising self-replicating nucleic acids are provided herein. The cancer-associated antigens or cancer therapeutic antibodies provided herein, or fragments thereof, can be encoded as part of a self-replicating nucleic acid construct. In some embodiments, the self-replicating nucleic acid molecule comprises at least one or more genes selected from the group consisting of viral replicase, viral protease, viral helicase and other nonstructural viral proteins, and also comprises 5'- and 3'-terminal cis-active replication sequences, and an antigen sequence encoding a cancer-associated antigen. A subgenomic promoter directing the expression of a heterologous sequence can be included in the self-replicating nucleotide sequence. If desired, the heterologous sequence can be fused in frame to other coding regions in the self-replicating RNA and / or under the control of an internal ribosome entry site (IRES).

[0071] In some embodiments, the self-replicating nucleotide sequence is a self-replicating RNA molecule. The self-replicating RNA molecule is designed so that it cannot induce the production of infectious virus particles. This can be achieved, for example, by removing one or more viral genes that code for structural proteins that are necessary for the production of virus particles in the self-replicating RNA. For example, when the self-replicating RNA molecule is based on an alphavirus, such as Sindbis virus (SIN), Semliki Forest virus and Venezuelan equine encephalitis virus (VEE), one or more genes that code for viral structural proteins, such as capsid and / or envelope glycoproteins, can be removed. If desired, the self-replicating RNA molecule of the present invention can be designed to induce the production of infectious virus particles that are attenuated or pathogenic, or to generate virus particles that are capable of a single round of subsequent infection.

[0072] When a self-replicating RNA molecule is delivered to an animal cell without any protein, it can result in the generation of multiple daughter RNAs by transcription from itself (or from its own antisense copy). After delivery to a cell, the self-replicating RNA can be directly translated, which then provides an RNA-dependent RNA polymerase that generates transcripts from the delivered RNA. In this way, the delivered RNA results in the generation of multiple daughter RNAs. These transcripts can be antisense to the delivered RNA and can themselves be translated to provide in situ expression of the encoded cancer-associated antigen, or can be transcribed to provide additional transcripts with the same sense as the delivered RNA that are translated to provide in situ expression of the encoded cancer-associated antigen.

[0073] The self-replicating RNA molecules provided herein may contain one or more modified nucleotides, and thus have improved stability, resistance to in vivo degradation and clearance, and other advantages. In some embodiments, the self-replicating RNA molecules containing modified nucleotides avoid or reduce the stimulation of endosomes and cytoplasmic immune receptors when the self-replicating RNA is delivered to cells. This allows self-replication, amplification, and expression of proteins to occur. This also reduces safety concerns compared to self-replicating RNAs that do not contain modified nucleotides, since the self-replicating RNAs containing modified nucleotides reduce the activation of the innate immune system and the subsequent undesirable consequences (e.g., inflammation at the injection site, irritation at the injection site, pain, etc.). The RNA molecules generated as a result of self-replication are recognized as foreign nucleic acids by cytoplasmic immune receptors. Therefore, the self-replicating RNA molecules containing modified nucleotides provide efficient amplification of RNA in host cells, and expression of cancer-associated antigen spike protein, as well as adjuvant effects.

[0074] In some embodiments, the self-replicating RNA molecules provided herein contain at least one modified nucleotide. Modified nucleotides that are not part of the 5' cap (e.g., in addition to modifications that are part of the 5'' cap) can be used. Thus, the self-replicating RNA molecule can contain a modified nucleotide at a single position, can contain a particular modified nucleotide (e.g., pseudouridine, N6-methyladenosine, 5-methylcytidine, 5-methyluridine) at two or more positions, or can contain 2, 3, 4, 5, 6, 7, 8, 9, 10 or more modified nucleotides (e.g., at each of one or more positions). Preferably, the self-replicating RNA molecule comprises a modified nucleotide that contains a modification on or in the nitrogenous base, but does not contain a modified sugar or phosphate moiety. In some examples, 0.001% to 99% or 100% of the nucleotides in the self-replicating RNA molecule are modified nucleotides. For example, 0.001% to 25%, 0.01% to 25%, 0.1% to 25%, or 1% to 25% of the nucleotides in the self-replicating RNA molecule are modified nucleotides. In other examples, 0.001% to 99% or 100% of the specific unmodified nucleotides in the self-replicating RNA molecule are replaced with modified nucleotides. For example, about 1% of the nucleotides in the self-replicating RNA molecule that contain uridine can be modified, for example, by replacing uridine with pseudouridine. In other examples, a desired amount (percentage) of 2, 3, or 4 specific nucleotides (nucleotides that contain uridine, cytidine, guanosine, or adenine) in the self-replicating RNA molecule are modified nucleotides. For example, 0.001% to 25%, 0.01% to 25%, 0.1% to 25%, or 1% to 25% of the specific nucleotides in the self-replicating RNA molecule are modified nucleotides. In other examples, between 0.001% and 20%, between 0.001% and 15%, between 0.001% and 10%, between 0.01% and 20%, between 0.01% and 15%, between 0.1% and 25%, between 0.01% and 10%, between 1% and 20%, between 1% and 15%, between 1% and 10%, or about 5%, about 10%, about 15%, about 20% of a particular nucleotide in a self-replicating RNA molecule are modified nucleotides.It is preferred that less than 100% of the nucleotides in the self-replicating RNA molecule are modified nucleotides. It is also preferred that less than 100% of a particular nucleotide in the self-replicating RNA molecule is modified nucleotide. Thus, preferred self-replicating RNA molecules contain at least some unmodified nucleotides.

[0075] Self-replicating RNA molecules containing at least one modified nucleotide can be prepared using any suitable method. Several suitable methods are known in the art for generating RNA molecules containing modified nucleotides. For example, self-replicating RNA molecules containing modified nucleotides can be prepared by transcribing (e.g., in vitro transcription) DNA encoding the self-replicating RNA molecule using a suitable DNA-dependent RNA polymerase, such as T7 phage RNA polymerase, SP6 phage RNA polymerase, T3 phage RNA polymerase, etc., or mutants of these polymerases that allow efficient incorporation of modified nucleotides into RNA molecules. The transcription reaction contains nucleotides and modified nucleotides, as well as other components that support the activity of the selected polymerase, such as suitable buffers and suitable salts. The incorporation of nucleotide analogs into self-replicating RNA can be engineered, for example, to modify the stability of such RNA molecules, increase resistance to RNase, establish replication after introduction into suitable host cells (the "infectivity" of RNA), and / or induce or reduce innate and adaptive immune responses. Suitable synthetic methods can be used alone or in combination with one or more other methods (e.g., recombinant DNA or RNA techniques) to generate self-replicating RNA molecules that contain one or more modified nucleotides.

[0076] Nucleic acid synthesis can also be performed using suitable recombinant methods well known in the art and conventional, including cloning, processing and / or expression of polynucleotides and gene products encoded by such polynucleotides. Random fragmentation and DNA shuffling by PCR reassembly of gene fragments and synthetic polynucleotides are examples of known techniques that can be used to design and manipulate polynucleotide sequences. Site-directed mutagenesis can be used to modify nucleic acids and encoded proteins, for example, to insert new restriction sites, change glycosylation patterns, change codon preferences, generate splice variants, introduce mutations, etc.

[0077] In some embodiments, the nucleic acid provided herein encodes an RNA polymerase. In some embodiments, the nucleic acid provided herein encodes a viral RNA polymerase. In some embodiments, the nucleic acid provided herein encodes (1) a viral RNA polymerase; and (2) a cancer-associated protein or a functional fragment thereof. In some embodiments, the composition provided herein comprises a first nucleic acid encoding a viral RNA polymerase; and a second nucleic acid encoding a cancer-associated protein or a functional fragment thereof. In some embodiments, the nucleic acid provided herein encodes (1) a viral RNA polymerase; and (2) a cancer therapeutic antibody or a functional fragment thereof. In some embodiments, the composition provided herein comprises a first nucleic acid encoding a viral RNA polymerase; and a second nucleic acid encoding a cancer therapeutic antibody or a functional fragment thereof.

[0078] The composition provided herein comprises self-replicating RNA. Self-replicating RNA (also called replicon) comprises any genetic element, such as plasmid, cosmid, bacmid, phage, or virus, that can replicate mainly under its own control. Self-replication provides a system for self-amplification of the nucleic acid provided herein in mammalian cells. In some embodiments, the self-replicating RNA is single-stranded. In some embodiments, the self-replicating RNA is double-stranded.

[0079] In some embodiments, the nucleic acid described herein comprises a sequence encoding an infectious disease antigen described herein and an RNA-dependent RNA polymerase. In some embodiments, the RNA-dependent RNA polymerase is a VEEV RNA polymerase. In some embodiments, the two nucleic acid coding elements are present on separate nucleic acids. In some embodiments, the two nucleic acid coding elements are present on the same nucleic acid. The RNA polymerase provided herein may include, but is not limited to, an alphavirus RNA polymerase, an Eastern Equine Encephalitis Virus (EEEV) RNA polymerase, a Western Equine Encephalitis Virus (WEEV), a Venezuelan Equine Encephalitis Virus (VEEV), or a Chikungunya Virus (CHIKV), a Semliki Forest Virus (SFV), or a Sindbis Virus (SINV). In some embodiments, the RNA polymerase is a VEEV RNA polymerase. In some embodiments, the nucleic acid encoding the RNA polymerase comprises at least 85% identity to the nucleic acid sequence of SEQ ID NO:71. In some embodiments, the nucleic acid encoding the RNA polymerase comprises at least 90% identity to the nucleic acid sequence of SEQ ID NO:71. In some embodiments, the nucleic acid encoding the RNA polymerase comprises at least 95% identity to the nucleic acid sequence of SEQ ID NO: 71. In some embodiments, the nucleic acid encoding the RNA polymerase comprises at least 99% identity to the nucleic acid sequence of SEQ ID NO: 71. In some embodiments, the nucleic acid encoding the RNA polymerase is SEQ ID NO: 71.

[0080] In some embodiments, the amino acid sequence for VEEV RNA polymerase comprises at least 85% identity to RELPVLDSAAFNVECFKKYACNNEYWETFKENPIRLTEENVVNYITKLKGP (SEQ ID NO:72), TQMRELPVLDSAAFNVECFKKYACNNEYWETFKENPIRLTE (SEQ ID NO:73), or SEQ ID NO:74. In some embodiments, the amino acid sequence for VEEV RNA polymerase comprises at least 90% identity to SEQ ID NO:72, SEQ ID NO:73, or SEQ ID NO:74. In some embodiments, the amino acid sequence for VEEV RNA polymerase comprises at least 95% identity to SEQ ID NO:72, SEQ ID NO:73, or SEQ ID NO:74. In some embodiments, the amino acid sequence for VEEV RNA polymerase comprises at least 99% identity to SEQ ID NO:72, SEQ ID NO:73, or SEQ ID NO:74. In some embodiments, the amino acid sequence for VEEV RNA polymerase is SEQ ID NO:72, SEQ ID NO:73, or SEQ ID NO:74.

[0081] Compositions and methods are provided herein that include a replicon RNA (repRNA) that encodes one or more structural proteins from a non-enveloped virus. In some embodiments, the repRNA encodes a protease. In some embodiments, the repRNA encodes a 3CD protease. In some embodiments, the structural proteins and the protease are co-expressed. In further embodiments, the repRNA comprises one or more open reading frames. In some embodiments, the open reading frames are separated by an internal ribosome entry site (IRES). In some embodiments, the open reading frames are separated by a ribosome skipping peptide sequence. In some embodiments, the ribosome skipping peptide sequence is from Tosea asigna virus (T2A). Nanoparticles

[0082] Various compositions comprising nanoparticles are provided herein. In some embodiments, the nanoparticles comprise lipid carriers. Nanoparticles are abbreviated herein as NPs. The nanoparticles provided herein can be organic, inorganic, or a combination of inorganic and organic materials with a diameter of less than about 1 micrometer (μm). In some embodiments, the nanoparticles provided herein are used as a delivery system for the bioactive agents provided herein (e.g., nucleic acids encoding cancer-associated proteins or cancer-therapeutic antibodies). Further provided herein are various compositions comprising lipid carrier complexes or nanoparticle complexes, in which multiple lipid carriers or multiple nanoparticles interact physically, chemically, and / or covalently. The specific type of interaction between lipid carriers or nanoparticles depends on the characteristic shape, size, chemical composition, physical properties, and physiological properties. The nanoparticles provided herein may include, but are not limited to, oil-in-water emulsions, nanostructured lipid carriers (NLCs), cationic nanoemulsions (CNEs), vesicular phospholipid gels (VPGs), polymeric nanoparticles, cationic lipid nanoparticles, liposomes, gold nanoparticles, solid lipid nanoparticles (LNPs or SLNs), mixed-phase core NLCs, ionized lipid carriers, magnetic carriers, polyethylene glycol (PEG)-functionalized carriers, cholesterol-functionalized carriers, polylactic acid (PLA)-functionalized carriers, and polylactic-co-glycolic acid (PLGA)-functionalized lipid carriers.

[0083] A variety of nanoparticles and nanoparticle formulations (i.e., nanoemulsions) are used. Exemplary nanoparticles are illustrated in Figures 1A-1H. An oil-in-water emulsion, illustrated in Figure 1A (not to scale), is a stable immiscible fluid containing oil droplets dispersed in a water or aqueous phase. Figure 1B (not to scale) illustrates a nanostructured lipid carrier (NLC) that may include a blend of solid organic lipids (e.g., trimyristin) and liquid oil (e.g., squalene). In the NLC, the solid lipids are dispersed in the liquid oil. The nanodroplets are dispersed entirely in the aqueous (water) phase. In some embodiments, the nanoparticles include inorganic nanoparticles as solid inorganic nanoparticles (e.g., iron oxide nanoparticles) dispersed in liquid oil, as illustrated in Figure 1C (not to scale). Figure 1D (not to scale) illustrates a nanoparticle that includes a cationic lipid membrane and liquid oil without inorganic particles. The nucleic acids provided herein can be complexed in cis (FIGS. 1A-1D) or in trans (FIGS. 1E-1H) with the nanoparticles in Table 3. For example, a first RNA or DNA molecule can include multiple cancer-associated proteins, and a second RNA or DNA molecule can include an RNA polymerase complex. As another example, a first RNA or DNA molecule can include one or more cancer-associated proteins and an RNA polymerase on the same nucleic acid, and a second RNA or DNA molecule can include additional cancer-associated proteins and / or RNA polymerase.

[0084] Provided herein are nanoemulsions and nanodroplets that comprise a number of lipid carriers or nanoparticles, each lipid carrier or nanoparticle comprising a cationic lipid. In some embodiments, the nanoemulsion comprises a number of cationic lipid carriers. In some embodiments, the compositions provided herein comprise cationic nanoemulsions. In some embodiments, the cationic nanoemulsions described herein comprise lipid (or other surfactant) molecules that surround oil particles dispersed in water, providing the oil particles with a cationic (positively charged) surface to which negatively charged RNA molecules can adhere.

[0085] The nanodroplets may be dispersed as a colloid in an aqueous (water) phase or in suspension. In some embodiments, the nanoparticles provided herein are dispersed in an aqueous solution. Non-limiting examples of aqueous solutions include water (e.g., sterile, distilled, deionized, ultrapure, RNAse-free, etc.), saline (e.g., Krebs, Ascaris, Denz, Tet saline), or 1% (w / v) dimethyl sulfoxide (DMSO) in water.

[0086] In some embodiments, the nanoparticles provided herein comprise a hydrophilic surface. In some embodiments, the hydrophilic surface comprises a cationic lipid. In some embodiments, the hydrophilic surface comprises an ionized lipid. In some embodiments, the nanoparticles comprise a membrane. In some embodiments, the membrane comprises a cationic lipid. In some embodiments, the nanoparticles provided herein comprise a cationic lipid. Exemplary cationic lipids for inclusion into hydrophilic surfaces include, but are not limited to, 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N′,N′-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyltrimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC), 1,2-dioleoyl-3-di methylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxy-dodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3 ,3',3'',3'''-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate; 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate;ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate); ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide; β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17 -tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol; BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10, 11,12,13,14,15,16,17-Tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl)oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine;FTT5, hexa(octan-3-yl) 9,9',9'',9''',9'''',9'''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanoate; Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane- 4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl)(9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate); PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Other examples of suitable classes of lipids include, but are not limited to, phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG); and PEGylated lipids (e.g., DSPE-PEG), including PEGylated versions of any of the above lipids. In some embodiments, the nanoparticles provided herein comprise DOTAP;

[0087] In some embodiments, the nanoparticles provided herein comprise a hydrophobic lipid core. In some embodiments, the hydrophobic lipid core is in a liquid phase at 25 degrees Celsius. Non-limiting examples of hydrophobic lipid core components that can be used include alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, solanesol, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. In some embodiments, the nanoparticles provided herein comprise a triglyceride. Exemplary triglycerides include, but are not limited to, capric triglyceride, caprylic triglyceride, caprylic and capric triglycerides, triglyceride esters, and myristate triglyceride. In some embodiments, the hydrophobic lipid is in a solid phase. In some embodiments, the hydrophobic lipid is in a liquid phase, also referred to as an oil. In some embodiments, the hydrophobic lipid comprises squalene. In some embodiments, the hydrophobic lipid comprises solanesol.

[0088] In some embodiments, the nanoparticles provided herein comprise a liquid organic material and a solid inorganic material. In some embodiments, the nanoparticles provided herein comprise inorganic particles. In some embodiments, the inorganic particles are solid inorganic particles. In some embodiments, the nanoparticles provided herein comprise inorganic particles within a hydrophobic core. In some embodiments, the nanoparticles provided herein comprise a metal. In some embodiments, the nanoparticles provided herein comprise a metal within a hydrophobic core. The metal can be, but is not limited to, a metal salt, such as a transition metal salt, a metal oxide, such as a transition metal oxide, a metal hydroxide, such as a transition metal hydroxide, a metal phosphate, such as a transition metal phosphate, or a metalloid (e.g., silicon and silicon-based compounds or alloys). In some embodiments, the nanoparticles provided herein comprise a metal salt, such as an aluminum oxide (Al2O3), an aluminum oxyhydroxide, an iron oxide (Fe3O4, Fe2O3, FeO, or a combination thereof), titanium dioxide, silicon dioxide (SiO2), aluminum hydroxyphosphate (Al(OH)x (PO4) y ), calcium phosphate (Ca3(PO4)2), calcium hydroxyapatite (Ca 10 (PO4)6(OH)2), iron gluconate, or iron sulfate. The inorganic particles may be formed from one or more of the same or different metals (any metal including transition metals). In some embodiments, the inorganic particles are transition metal oxides. In some embodiments, the transition metal is magnetite (Fe3O4), maghemite (y-Fe2O3), wustite (FeO), or hematite (alpha (α)-Fe2O3). In some embodiments, the metal is aluminum hydroxide or aluminum oxyhydroxide, and a phosphate-terminated lipid or surfactant, such as oleic acid, oleylamine, SDS, TOPO, or DSPA, is used to coat the inorganic solid nanoparticles before mixing with the liquid oil to form the hydrophobic core. In some embodiments, the metal can include paramagnetic, superparamagnetic, ferrimagnetic, or ferromagnetic compounds. In some embodiments, the metal is superparamagnetic iron oxide (Fe3O4).

[0089] In some embodiments, the nanoparticles provided herein comprise cationic lipids, oils, and inorganic particles.In some embodiments, the nanoparticles provided herein comprise DOTAP; squalene and / or glyceryl trimyristate-dynasan; and iron oxide.In some embodiments, the nanoparticles provided herein further comprise a surfactant.

[0090] In some embodiments, the nanoparticles provided herein comprise a cationic lipid, an oil, an inorganic particle, and a surfactant.

[0091] Surfactants are compounds that reduce the surface tension between two liquids or between a liquid and a solid component of the nanoparticles provided herein. Surfactants can be hydrophobic, hydrophilic, or amphiphilic. In some embodiments, the nanoparticles provided herein include a hydrophobic surfactant. Exemplary hydrophobic surfactants that can be used include, but are not limited to, sorbitan monolaurate (SPAN® 20), sorbitan monopalmitate (SPAN® 40), sorbitan monostearate (SPAN® 60), sorbitan tristearate (SPAN® 65), sorbitan monooleate (SPAN® 80), and sorbitan trioleate (SPAN® 85).

[0092] Suitable hydrophobic surfactants include those with a hydrophilic-lipophilic balance (HLB) value of 10 or less, e.g., 5 or less, 1-5, or 4-5. For example, the hydrophobic surfactant can be a sorbitan ester with an HLB value of 1-5, or 4-5. In some embodiments, the nanoparticles provided herein include a ratio of esters that results in a hydrophilic-lipophilic balance of 8-11. HLB is used to classify surfactants as hydrophilic or lipophilic. The HLB scale can be, for example, according to the Griffin method:

number

[0093] In some embodiments, the nanoparticles or lipid carriers provided herein comprise a hydrophilic surfactant, also called an emulsifier. In some embodiments, the nanoparticles or lipid carriers provided herein comprise a polysorbate. Polysorbates are oily liquids derived from ethoxylated sorbitan (a derivative of sorbitol) esterified with fatty acids. Exemplary hydrophilic surfactants that can be used include, but are not limited to, polysorbates, such as TWEEN®, Kolliphor, Scattics, Alkest, or Canarcel; polyoxyethylene sorbitan esters (polysorbates); polysorbate 80 (polyoxyethylene sorbitan monooleate, or TWEEN® 80); polysorbate 60 (polyoxyethylene sorbitan monostearate, or TWEEN® 60); polysorbate 40 (polyoxyethylene sorbitan monopalmitate, or TWEEN® 40); and polysorbate 20 (polyoxyethylene sorbitan monolaurate, or TWEEN® 20). In one embodiment, the hydrophilic surfactant is polysorbate 80.

[0094] In some embodiments, the nanoparticles and lipid carriers provided herein comprise a hydrophobic core surrounded by a lipid membrane (e.g., a cationic lipid such as DOTAP). In some embodiments, the hydrophobic core comprises one or more inorganic particles; a phosphate-terminated lipid; and a surfactant.

[0095] The inorganic solid nanoparticles described herein may be surface modified before mixing with liquid oil. For example, if the surface of the inorganic solid nanoparticles is hydrophilic, the inorganic solid nanoparticles may be coated with hydrophobic molecules (or surfactants) to promote the miscibility of the inorganic solid nanoparticles with the liquid oil in the "oil" phase of the nanoemulsion particle. In some embodiments, the inorganic particles are coated with capping ligands, phosphate-terminated lipids, and / or surfactants. In some embodiments, the hydrophobic core comprises a phosphate-terminated lipid. Exemplary phosphate-terminated lipids that may be used include, but are not limited to, trioctylphosphine oxide (TOPO) or distearylphosphatidic acid (DSPA). In some embodiments, the hydrophobic core comprises a surfactant, such as a phosphorus-terminated surfactant, a carboxylate-terminated surfactant, a sulfate-terminated surfactant, or an amine-terminated surfactant. An exemplary carboxylate-terminated surfactant includes oleic acid. A typical amine-terminated surfactant includes oleylamine. In some embodiments, the surfactant is distearyl phosphatidic acid (DSPA), oleic acid, oleylamine, or sodium dodecyl sulfate (SDS). In some embodiments, the inorganic solid nanoparticles are metal oxides, such as iron oxide, and are coated with a surfactant, such as oleic acid, oleylamine, SDS, DSPA, or TOPO, before mixing with the liquid oil to form a hydrophobic core.

[0096] In some embodiments, the hydrophobic core comprises one or more inorganic particles containing at least one metal hydroxide or oxyhydroxide particle, optionally coated with a phosphate-terminated lipid, phosphorus-terminated surfactant, carboxylate-terminated surfactant, sulfate-terminated surfactant, or amine-terminated surfactant; and a naturally occurring or synthetic squalene-containing liquid oil; a cationic lipid comprising DOTAP; a hydrophobic surfactant comprising a sorbitan ester selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate; and a hydrophilic surfactant comprising a polysorbate.

[0097] In some embodiments, the hydrophobic core comprises one or more inorganic nanoparticles containing aluminum hydroxide or aluminum oxyhydroxide nanoparticles, optionally coated with TOPO, as well as a naturally occurring or synthetic squalene-containing liquid oil; the cationic lipid DOTAP; a hydrophobic surfactant including sorbitan monostearate; and a hydrophilic surfactant including polysorbate 80.

[0098] In some embodiments, the hydrophobic core is comprised of one or more inorganic particles containing at least one metal hydroxide or oxyhydroxide particle, optionally coated with a phosphate-terminated lipid, phosphorus-terminated surfactant, carboxylate-terminated surfactant, sulfate-terminated surfactant, or amine-terminated surfactant; and a naturally occurring or synthetic squalene-containing liquid oil; a cationic lipid comprising DOTAP; a hydrophobic surfactant comprising a sorbitan ester selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate; and a hydrophilic surfactant comprising a polysorbate.

[0099] In some embodiments, the hydrophobic core is comprised of one or more inorganic nanoparticles containing aluminum hydroxide or aluminum oxyhydroxide nanoparticles, optionally coated with TOPO, as well as a naturally occurring or synthetic squalene-containing liquid oil; the cationic lipid DOTAP; a hydrophobic surfactant comprising sorbitan monostearate; and a hydrophilic surfactant comprising polysorbate 80. In some embodiments, the nanoparticles provided herein may comprise about 0.2% to about 40% w / v squalene, about 0.001% to about 10% w / v iron oxide nanoparticles, about 0.2% to about 10% w / v DOTAP, about 0.25% to about 5% w / v sorbitan monostearate, and about 0.5% to about 10% w / v polysorbate 80. In some embodiments, the nanoparticles provided herein may comprise about 2% to about 6% w / v squalene, about 0.01% to about 1% w / v iron oxide nanoparticles, about 0.2% to about 1% w / v DOTAP, about 0.25% to about 1% w / v sorbitan monostearate, and about 0.5% to about 5% w / v polysorbate 80. In some embodiments, the nanoparticles provided herein may comprise about 0.2% to about 40% w / v squalene, about 0.001% to about 10% w / v aluminum hydroxide or aluminum oxyhydroxide nanoparticles, about 0.2% to about 10% w / v DOTAP, about 0.25% to about 5% w / v sorbitan monostearate, and about 0.5% to about 10% w / v polysorbate 80. In some embodiments, the nanoparticles provided herein may comprise about 2% to about 6% w / v squalene, about 0.01% to about 1% w / v aluminum hydroxide or aluminum oxyhydroxide nanoparticles, about 0.2% to about 1% w / v DOTAP, about 0.25% to about 1% w / v sorbitan monostearate, and about 0.5% to about 5% w / v polysorbate 80.

[0100] In some embodiments, the compositions described herein comprise at least one nanoparticle formulation described in Table 3. In some embodiments, the compositions described herein comprise any one of NP-1 to NP-30. In some embodiments, the compositions described herein comprise any one of NP-1 to NP-34. In some embodiments, the nanoparticles provided herein are mixed with the nucleic acids provided herein. In some embodiments, the nanoparticles provided herein are made by homogenization and sonication techniques. [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5] [Table 3-6] [Table 3-7] [Table 3-8] [Table 3-9]

[0101] In some embodiments, the nanoparticles provided herein comprise sorbitan monostearate (e.g., SPAN® 60), polysorbate 80 (e.g., TWEEN® 80), DOTAP, squalene, and are free of solid particles. In some embodiments, the nanoparticles provided herein comprise sorbitan monostearate (e.g., SPAN® 60), polysorbate 80 (e.g., TWEEN® 80), DOTAP, squalene, and iron oxide particles. In some embodiments, the nanoparticles provided herein comprise an immune stimulant. In some embodiments, the immune stimulant is squalene. In some embodiments, the immune stimulant is Miglyol 810 or Miglyol 812. Miglyol 810 is a triglyceride ester of saturated caprylic and capric fatty acids and glycerol. Miglyol 812 is a triglyceride ester of saturated coconut oil / palm kernel oil derived caprylic and capric fatty acids and vegetable derived glycerol. In some embodiments, the immune stimulant may reduce the total amount of protein produced but increase the immune response to the compositions provided herein (e.g., when delivered as a vaccine). In some embodiments, the immune stimulant may increase the total amount of protein produced but decrease the immune response to the compositions provided herein.

[0102] The nanoparticles provided herein can be of various average diameter sizes. In some embodiments, the nanoparticles provided herein have an average diameter (z-average hydrodynamic diameter, measured by dynamic light scattering) ranging from about 20 nanometers (nm) to about 200 nm. In some embodiments, the z-average diameter of the nanoparticles ranges from about 20 nm to about 150 nm, about 20 nm to about 100 nm, about 20 nm to about 80 nm, or about 20 nm to about 60 nm. In some embodiments, the z-average diameter of the nanoparticles ranges from about 40 nm to about 200 nm, about 40 nm to about 150 nm, about 40 nm to about 100 nm, about 40 nm to about 90 nm, about 40 nm to about 80 nm, or about 40 nm to about 60 nm. In one embodiment, the z-average diameter of the nanoparticles is about 40 nm to about 80 nm. In some embodiments, the z-average diameter of the nanoparticles is about 40 nm to about 60 nm. In some embodiments, the nanoparticles are up to 100 nm in diameter. In some embodiments, the nanoparticles are 50-70 nm in diameter. In some embodiments, the nanoparticles are 40-80 nm in diameter. In some embodiments, the inorganic particles (e.g., iron oxide) within the hydrophobic core of the nanoparticles can have an average diameter (number weighted average diameter) ranging from about 3 nm to about 50 nm. For example, the inorganic particles can have an average diameter of about 5 nm, about 10 nm, about 15 nm, about 20 nm, about 25 nm, about 30 nm, about 35 nm, about 40 nm, about 45 nm, or about 50 nm. In some embodiments, the ratio of ester and lipid results in a particle size of 30 nm to 200 nm. In some embodiments, the ratio of ester and lipid results in a particle size of 40 nm to 70 nm.

[0103] The nanoparticles provided herein may be characterized by their polydispersity index (PDI), which is an indication of their quality with respect to size distribution. In some embodiments, the average polydispersity index (PDI) of the nanoparticles provided herein ranges from about 0.1 to about 0.5. In some embodiments, the average PDI of the nanoparticles may range from about 0.2 to about 0.5, about 0.1 to about 0.4, about 0.2 to about 0.4, about 0.2 to about 0.3, or about 0.1 to about 0.3.

[0104] In some embodiments, the nanoparticles provided herein comprise a molar ratio of oil to surfactant ranging from about 0.1:1 to about 20:1, from about 0.5:1 to about 12:1, from about 0.5:1 to about 9:1, from about 0.5:1 to about 5:1, from about 0.5:1 to about 3:1, or from about 0.5:1 to about 1:1. In some embodiments, the nanoparticles provided herein comprise a ratio of hydrophilic surfactant to lipid ranging from about 0.1:1 to about 2:1, from about 0.2:1 to about 1.5:1, from about 0.3:1 to about 1:1, from about 0.5:1 to about 1:1, or from about 0.6:1 to about 1:1. In some embodiments, the nanoparticles provided herein comprise a ratio of hydrophobic surfactant to lipid ranging from about 0.1:1 to about 5:1, from about 0.2:1 to about 3:1, from about 0.3:1 to about 2:1, from about 0.5:1 to about 2:1, or from about 1:1 to about 2:1.

[0105] In some embodiments, the nanoparticles provided herein comprise about 0.2% to about 40% w / v liquid oil, about 0.001% to about 10% w / v inorganic solid nanoparticles, about 0.2% to about 10% w / v lipid, about 0.25% to about 5% w / v hydrophobic surfactant, and about 0.5% to about 10% w / v hydrophilic surfactant. In some embodiments, the lipid comprises a cationic lipid, the oil comprises squalene, and / or the hydrophobic surfactant comprises a sorbitan ester. Combination Compositions

[0106] Provided herein is a composition comprising the nanoparticles described herein and the nucleic acid encoding cancer-associated protein or cancer-associated protein binding protein.In some embodiments, the nucleic acid provided herein is incorporated into, associated with, or complexed with the lipid carrier provided herein to form lipid carrier-nucleic acid complex.The lipid carrier-nucleic acid complex is formed through non-covalent interaction or through reversible covalent interaction.

[0107] Further provided herein is a nanoemulsion comprising a number of nanoparticles provided herein. In some embodiments, the nucleic acid further encodes an RNA-dependent polymerase. In some embodiments, the RNA-dependent polymerase is a viral RNA polymerase. In some embodiments, the nucleic acid encoding the RNA polymerase is on the same nucleic acid strand as the nucleic acid sequence encoding the protein (e.g., in cis). In some embodiments, the nucleic acid encoding the RNA polymerase is on a different nucleic acid strand than the nucleic acid sequence encoding the protein (e.g., in trans). In some embodiments, the nucleic acid encoding the RNA polymerase is a DNA molecule. In some embodiments, the nucleic acid sequence encoding the cancer-associated protein, tumor antigen, neo-antigen, cancer therapeutic antibody, or functional fragment thereof is a DNA or RNA molecule. In some embodiments, the cancer-associated protein and cancer therapeutic antibody provided herein are encoded by DNA. Nanoparticles for inclusion include, but are not limited to, any one of NP-1 through NP-31, or any one of NP-1 through NP-34. Nucleic acids for inclusion include, but are not limited to, those that contain a region including any one or more of SEQ ID NOs: 1, 2, 75, 76, 88, 89, and / or those that encode an amino acid sequence set forth in any one of SEQ ID NOs: 3-70, 77, 78, 87. In some instances, the nucleic acid further comprises a region that encodes an RNA polymerase, e.g., a region that includes the sequence of SEQ ID NO: 71.

[0108] The compositions provided herein can be characterized by the molar ratio of nitrogen:phosphate (N:P). The N:P ratio is determined by the amount of cationic lipid in the nitrogen-containing nanoparticle and the amount of nucleic acid used in the composition containing negatively charged phosphate. The molar ratio of lipid carrier to nucleic acid can be selected to increase the delivery efficiency of nucleic acid, increase the ability of the nucleic acid-carrying nanoemulsion composition to induce an immune response against an antigen, or increase the ability of the nucleic acid-carrying nanoemulsion composition to induce the production of antibody titers against an antigen in a subject. In some embodiments, the compositions provided herein have a molar ratio of lipid carrier to nucleic acid, which may be characterized by the molar ratio of nitrogen to phosphate, which may range from about 0.01:1 to about 1000:1, e.g., from about 0.2:1 to about 500:1, from about 0.5:1 to about 150:1, from about 1:1 to about 150:1, from about 1:1 to about 125:1, from about 1:1 to about 100:1, from about 1:1 to about 50:1, from about 1:1 to about 50:1, from about 5:1 to about 50:1, from about 5:1 to about 25:1, or from about 10:1 to about 20:1. In certain embodiments, the molar ratio of lipid carrier to nucleic acid, characterized by the molar ratio of nitrogen to phosphate (N:P), ranges from about 1:1 to about 150:1, from about 5:1 to about 25:1, or from about 10:1 to about 20:1. In one embodiment, the nanoemulsion composition has an N:P molar ratio of about 15: 1. In some embodiments, the nanoparticles comprise a nucleic acid provided herein covalently attached to a membrane.

[0109] The compositions provided herein may be characterized by the molar ratio of oil to surfactant. In some embodiments, the ratio of oil to surfactant is the molar ratio of squalene:DOTAP, hydrophobic surfactant, and hydrophilic surfactant. In some embodiments, the ratio of oil to surfactant is the molar ratio of squalene:DOTAP, sorbitan monostearate, and polysorbate 80. In some embodiments, the molar ratio of oil to surfactant ranges from about 0.1:1 to about 20:1, from about 0.5:1 to about 12:1, from about 0.5:1 to about 9:1, from about 0.5:1 to about 5:1, from about 0.5:1 to about 3:1, or from about 0.5:1 to about 1:1. In some embodiments, the molar ratio of oil to surfactant is at least about 0.1:1, at least about 0.2:1, at least about 0.3:1, at least about 0.4:1, at least about 0.5:1, at least about 0.6:1, or at least about 0.7:1. In some embodiments, the molar ratio of oil to surfactant is at least about 0.4:1 to at most 1:1.

[0110] The compositions provided herein may be characterized by the ratio of hydrophilic surfactant to lipid (e.g., cationic lipid). In some embodiments, the ratio of hydrophilic surfactant to lipid ranges from about 0.1:1 to about 2:1, from about 0.2:1 to about 1.5:1, from about 0.3:1 to about 1:1, from about 0.5:1 to about 1:1, or from about 0.6:1 to about 1:1. The compositions provided herein may be characterized by the ratio of hydrophobic surfactant to lipid (e.g., cationic lipid). In some embodiments, the ratio of hydrophobic surfactant to lipid ranges from about 0.1:1 to about 5:1, from about 0.2:1 to about 3:1, from about 0.3:1 to about 2:1, from about 0.5:1 to about 2:1, or from about 1:1 to about 2:1.

[0111] Provided herein is a dry composition comprising a sorbitan fatty acid ester, an ethoxylated sorbitan ester, a cationic lipid, an immunostimulant, and an RNA.Further provided herein is a dry composition comprising a sorbitan monostearate (e.g., SPAN® 60), a polysorbate 80 (e.g., TWEEN® 80), a DOTAP, an immunostimulant, and an RNA. Heat-stable dry and freeze-dried cancer vaccines

[0112] Provided herein are dry or lyophilized compositions and vaccines.Further provided herein are pharmaceutical compositions comprising the dry or lyophilized compositions provided herein, which are reconstituted in a suitable diluent and a pharma- ceutically acceptable carrier.In some embodiments, the diluent is aqueous.In some embodiments, the diluent is water.

[0113] Lyophilized compositions are made by a low-temperature dehydration process that involves freezing the composition, followed by reducing pressure and removing ice by sublimation. In certain cases, lyophilization also involves removing bound water molecules by a desorption process. In some embodiments, the compositions and vaccine compositions provided herein are spray-dried. Spray-drying is a process by which a solution is fed through an atomizer to create a spray, which is then exposed to a heated gas stream to promote rapid evaporation. Once a sufficient liquid mass has evaporated, the remaining solid material in the droplets forms particles that are then separated from the gas stream (e.g., using a filter or cyclone). Drying helps the compositions and vaccine compositions provided herein to be stored at higher temperatures (e.g., above 4°C) compared to the subzero temperatures required for storage of existing mRNA vaccines. In some embodiments, the dry and lyophilized compositions provided herein comprise: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising: (i) a hydrophobic core; (ii) one or more inorganic nanoparticles; and (iii) one or more lipids; (b) one or more nucleic acids; and (c) at least one cryoprotectant. In some embodiments, the cryoprotectant is selected from the group consisting of sucrose, maltose, trehalose, mannitol, glucose, and any combination thereof. Additional examples of cryoprotectants include, but are not limited to, dimethyl sulfoxide (DMSO), glycerol, propylene glycol, ethylene glycol, 3-O-methyl-D-glucopyranose (3-OMG), polyethylene glycol (PEG), 1,2-propanediol, acetamide, trehalose, formamide, sugars, proteins, and carbohydrates.

[0114] In some embodiments, the compositions and methods provided herein include at least one cryoprotectant. Exemplary cryoprotectants for inclusion are, but are not limited to, sucrose, maltose, trehalose, mannitol, or glucose, and any combination thereof. In some embodiments, additional or alternative cryoprotectants for inclusion are sorbitol, ribitol, erythritol, threitol, ethylene glycol, or fructose. In some embodiments, additional or alternative cryoprotectants for inclusion are dimethylsulfoxide (DMSO), glycerol, propylene glycol, ethylene glycol, 3-O-methyl-D-glucopyranose (3-OMG), polyethylene glycol (PEG), 1,2-propanediol, acetamide, trehalose, formamide, sugars, proteins, and carbohydrates. In some embodiments, the cryoprotectant is present at about 1% w / v to about 20% w / v, preferably about 10% w / v to about 20% w / v, more preferably about 10% w / v. In certain aspects of the present disclosure, the cryoprotectant is sucrose. In some aspects of the present disclosure, the cryoprotectant is maltose. In some aspects of the present disclosure, the cryoprotectant is trehalose. In some aspects of the present disclosure, the cryoprotectant is mannitol. In some aspects of the present disclosure, the cryoprotectant is glucose. In some embodiments, the cryoprotectant is present in an amount of about 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 325, 350, 375, 400, 450, 500 mg or more. In some embodiments, the cryoprotectant is present in an amount of about 50 to about 500 mg. In some embodiments, the cryoprotectant is present in an amount of about 200 to about 300 mg. In some embodiments, the cryoprotectant is present in an amount of about 250 mg. In some embodiments, the cryoprotectant is present in an amount of at least about 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or more percent by weight of the lyophilized composition.In some embodiments, the cryoprotectant is present in an amount of about 95% by weight of the lyophilized composition. In some embodiments, the cryoprotectant is present in an amount of 80-98%, 85-98%, 90-98%, or 94-96% by weight of the lyophilized composition. In some embodiments, the cryoprotectant is a sugar. In some embodiments, the sugar is sucrose, maltose, trehalose, mannitol, or glucose. In some embodiments, the sugar is sucrose. In some embodiments, sucrose is present in an amount of about 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 325, 350, 375, 400, 450, 500 mg or more. In some embodiments, sucrose is present in an amount of about 50 to about 500 mg. In some embodiments, sucrose is present in an amount of about 200 to about 300 mg. In some embodiments, sucrose is present in an amount of about 250 mg. In some embodiments, sucrose is present in an amount of at least about 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or more percent by weight of the lyophilized composition. In some embodiments, sucrose is present in an amount of about 95% by weight of the lyophilized composition. In some embodiments, sucrose is present in an amount of 80-98%, 85-98%, 90-98%, or 94-96% by weight of the lyophilized composition.

[0115] In some embodiments, the cryoprotectant is sucrose. In some embodiments, the cryoprotectant is at a concentration of at least about 0.1% w / v. In some embodiments, the cryoprotectant is at a concentration of about 1% w / v to about 20% w / v. In some embodiments, the cryoprotectant is at a concentration of about 10% w / v to about 20% w / v. In some embodiments, the cryoprotectant is at a concentration of about 10% w / v.

[0116] In some embodiments, the compositions and vaccine compositions provided herein are thermally stable. A composition is considered to be thermally stable if the composition maintains its properties, such as the ability to resist the effects of heat or cold and protect nucleic acid molecules from degradation at a given temperature. In some embodiments, the compositions and vaccine compositions provided herein are thermally stable at about 25 degrees Celsius (°C) or standard room temperature. In some embodiments, the compositions and vaccine compositions provided herein are thermally stable at about 45°C. In some embodiments, the compositions and vaccine compositions provided herein are thermally stable at about -20°C. In some embodiments, the compositions and vaccine compositions provided herein are thermally stable at about 2°C to about 8°C. In some embodiments, the compositions and vaccine compositions provided herein are thermally stable at temperatures of at least about -80°C, at least about -20°C, at least about 0°C, at least about 2°C, at least about 4°C, at least about 6°C, at least about 8°C, at least about 10°C, at least about 20°C, at least about 25°C, at least about 30°C, at least about 37°C, and up to 45°C. In some embodiments, the compositions and vaccine compositions provided herein are thermally stable for at least about 5 days, at least about 1 week, at least about 2 weeks, at least about 1 month, or up to 3 months. In some embodiments, the compositions and vaccine compositions provided herein are stored at a temperature of at least about 4° C. to a maximum of 37° C. for at least about 5 days, at least about 1 week, at least about 2 weeks, at least about 1 month, or up to 3 months. In some embodiments, the compositions and vaccine compositions provided herein are stored at a temperature of at least about 20° C. to a maximum of 25° C. for at least about 5 days, at least about 1 week, at least about 2 weeks, at least about 1 month, or up to 3 months.

[0117] Also provided herein is a method for preparing a lyophilized composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating one or more nucleic acids into the lipid carrier to form a lipid carrier-nucleic acid complex; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; and lyophilizing the formulation to form a lyophilized composition.

[0118] Further provided herein is a method for preparing a spray-dried composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating one or more nucleic acids into the lipid carrier to form a lipid carrier-nucleic acid complex; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; and spray drying the formulation to form the spray-dried composition.

[0119] Further provided herein is a method for reconstituting a lyophilized composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating one or more nucleic acids into the lipid carrier to form a lipid carrier-nucleic acid complex; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; lyophilizing the formulation to form a lyophilized composition; and reconstituting the lyophilized composition in a suitable diluent.

[0120] Further provided herein is a method for reconstituting a spray-dried composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating one or more nucleic acids into the lipid carrier to form a lipid carrier-nucleic acid complex; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; spray drying the formulation to form the spray-dried composition; and reconstituting the spray-dried composition in a suitable diluent. Pharmaceutical Compositions

[0121] Provided herein is a suspension comprising the composition provided herein. In some embodiments, the suspension provided herein comprises a number of nanoparticles or compositions provided herein. In some embodiments, the composition provided herein is a suspension, optionally a homogenous suspension. In some embodiments, the composition provided herein is in the form of an emulsion.

[0122] Also provided herein is a pharmaceutical composition comprising the composition provided herein.In some embodiments, the composition provided herein is combined with pharmaceutically acceptable salts, excipients, and / or carriers to form a pharmaceutical composition.Pharmaceutical salts, excipients, and carriers can be selected based on the route of administration, the location of target tissue, and the time course of drug delivery.Pharmaceutically acceptable carriers or excipients can include solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc., that are compatible with pharmaceutical administration.

[0123] In some embodiments, the pharmaceutical composition is in the form of a solid, semi-solid, liquid or gas (aerosol). Injectable preparations, for example, sterile injectable aqueous or oily suspensions, can be formulated according to known techniques using suitable dispersing or wetting agents and suspending agents. Sterile injectable preparations can also be sterile injectable solutions, suspensions or emulsions in non-toxic parenterally acceptable diluents or solvents. Among the acceptable vehicles and solvents that can be used are water, Ringer's solution, USP, and isotonic sodium chloride solution. In addition, sterile fixed oils are used as solvents or dispersion media. For this purpose, any brand of fixed oil can be used, including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables. Injectable preparations can be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating a sterilizing agent in the form of a sterile solid composition that can be dissolved or dispersed in sterile water or other sterile injectable medium before use.

[0124] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the encapsulated or non-encapsulated conjugates contain at least one inert pharma- ceutically acceptable excipient or carrier, such as sodium citrate or dibasic calcium phosphate, and / or (a) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, and silicic acid; (b) binders, such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia; (c) humectants, such as glycerol; (d) agar, calcium carbonate, and / or (e) agar-agar, calcium carbonate, and / or (f) agar-agar, calcium carbonate, and / or (g) agar-agar, calcium carbonate, and / or (h) agar-agar, calcium carbonate, and / or (i) agar-agar, calcium carbonate, and / or (j) agar-agar, calcium carbonate, and / or (k) agar-agar, calcium carbonate, and / or (l ... The formulation may be mixed with disintegrating agents such as wheat flour, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, (e) dissolution inhibitors such as paraffin, (f) absorption accelerators such as quaternary ammonium compounds, (g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, (h) absorbents such as kaolin and bentonite clay, and (i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may also contain buffering agents. dosage

[0125] The compositions provided herein may be formulated into dosage unit forms for ease of administration and uniformity of dosage. A dosage unit form is a physically discrete unit of the compositions provided herein appropriate for the subject to be treated. However, it will be understood that the total amount of the compositions provided herein to be used will be determined by the attending physician within the scope of sound medical judgment. For any compositions provided herein, the therapeutically effective dose can be estimated initially in cell culture assays or animal models such as mice, rabbits, dogs, pigs, or non-human primates. Subjects include, but are not limited to, domestic or farmed animals (including, but not limited to, pigs, cows, horses, buffalo, pigs, ducks, geese, chickens, turkeys, fish), as well as humans. Dosing may be for veterinary or human therapeutic use. Animal models are also used to achieve a desired concentration range and route of administration. Such information can then be used to determine useful doses and routes for administration in humans. The therapeutic efficacy and toxicity of the compositions provided herein can be determined using standard pharmaceutical procedures in cell cultures or experimental animals, such as ED 50 (the dose that is therapeutically effective in 50% of the population) and LD 50 (the dose that is lethal to 50% of the population). The dose ratio of toxic to therapeutic effects is the therapeutic index, which is the LD 50 / ED 50 The therapeutic index can be expressed as a ratio of 0 to 1. Pharmaceutical compositions that exhibit large therapeutic indices can be useful in some embodiments. The data obtained from cell culture assays and animal studies can be used to formulate a range of dosages for human use. Administration

[0126] Provided herein are compositions and pharmaceutical compositions for administration to a subject in need thereof. In some embodiments, the pharmaceutical compositions provided herein are in a form that allows the compositions provided herein to be administered to a subject.

[0127] In some embodiments, the administration is local or systemic. In some embodiments, the compositions described herein are formulated for / use in administration via subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, or intrathecal routes. In some embodiments, the administration is every 1, 2, 4, 6, 8, 12, 24, 36, or 48 hours. In some embodiments, the administration is daily, weekly, or monthly. In some embodiments, the administration is repeated at least about every 28 or 56 days.

[0128] In some embodiments, a single dose of the composition provided herein is administered to the subject. In some embodiments, the composition or pharmaceutical composition provided herein is administered to the subject by two doses. In some embodiments, the second dose of the composition or pharmaceutical composition provided herein is administered about 28 days or 56 days after the first dose. In some embodiments, the first dose is administered and the second dose is administered about 14 days later, or about 21 days later, or about 28 days later, or about 35 days later, or about 42 days later, or about 49 days later, or about 56 days later, or about 63 days later, or about 70 days later, or about 77 days later, or about 84 days later. In some embodiments, the second dose is administered about 10-90 days after administration of the first dose, or about 15-85 days after administration of the first dose, or about 20-80 days after administration of the first dose, or about 25-75 days after administration of the first dose, or about 30-70 days after administration of the first dose, or about 35-65 days after administration of the first dose, or about 40-60 days after administration of the first dose.

[0129] In some embodiments, additional, eg, three or more, doses of a composition or pharmaceutical composition provided herein are administered to the subject. In some embodiments, the additional dose is administered about 1 month after administration of the second dose, about 2 months after administration of the second dose, about 3 months after administration of the second dose, about 4 months after administration of the second dose, about 5 months after administration of the second dose, about 6 months after administration of the second dose, about 7 months after administration of the second dose, about 8 months after administration of the second dose, about 9 months after administration of the second dose, about 10 months after administration of the second dose, about 11 months after administration of the second dose, about 12 months after administration of the second dose, about 13 months after administration of the second dose, about 14 months after administration of the second dose, about 15 months after administration of the second dose, about 16 months after administration of the second dose, about 17 months after administration of the second dose, or about 18 months after administration of the second dose. method

[0130] Methods of treating or preventing disease in a subject are provided herein. In some embodiments, the compositions described herein are used for the treatment of cancer. In some embodiments, the cancer is a solid cancer or a blood cancer. In some embodiments, the solid cancer is a carcinoma, melanoma, or sarcoma. In some embodiments, the blood cancer is a lymphoma or leukemia. In some embodiments, the cancer is a metastatic cancer. In some embodiments, the cancer is a skin cancer. In some embodiments, the skin cancer is a basal cell carcinoma, a melanoma, a Merkel cell carcinoma, a squamous cell carcinoma, a cutaneous lymphoma, a Kaposi's sarcoma, or a skin adnexal carcinoma. In some embodiments, the subject has lung cancer. In some embodiments, the lung cancer is a non-small cell lung cancer (NSCLC) or a small cell lung cancer (SCLC). In some embodiments, the NSCLC is an adenocarcinoma, a squamous cell carcinoma, a large cell carcinoma, an adenosquamous carcinoma, or a sarcomatoid carcinoma. In some embodiments, the cancer is a pancreatic cancer. In some embodiments, the pancreatic cancer is a pancreatic adenocarcinoma or a pancreatic exocrine carcinoma. In some embodiments, the pancreatic cancer is pancreatic neuroendocrine cancer, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. In some embodiments, the cancer is prostate cancer.

[0131] Provided herein is a personalized treatment for the treatment of cancer in a subject. Provided herein is a personalized treatment that can match the expression of the cancer-associated proteins provided herein and / or the specific genotype of the subject identified as having cancer. For example, the subject can be tested for specific mutations in oncogenic driver genes (e.g., KRAS G12C or G12D) that are known to cause a specific subtype of cancer (e.g., NSCLC lung cancer). Oncogenic driver mutations are genetic mutations that are responsible for both the initiation and maintenance of cancer. In some embodiments, the subject is identified as having a mutation in oncogenic driver genes or biomarkers.Non-limiting examples of oncogenic driver genes / biomarkers and their associated cancer types include: Breast cancer: BRCA1, BRCA2, TP53, TTN, FLG, OBSCN, ERBB2, GATA3, FGFR1, CCND1, PIK3CA, CACNA1C, ARHGAP35, ARID5B, BIRC6, CDH1, CTCF, DSPP, HDAC9, KDM5B, MAST1, MEF2A, NCOR2, SETD1A, SXL2, RID1A, CTNND1, NUP107, CHD8, FANCI, CHD9, CTCF, KEAP1, PCDH18, LAMA2, HDAC9, ARFGEF1, MLLT4, NRK, FOXO3, CDKN2A, MAP3K1, GPS2, ROCK2, RYR2, PGR, STAT6, PIK3CD, CTCF, CDH1, GATA3, AKT1; gastric cancer: ADCY3, BCL6B, CACNA1C, Multi-Registered Polymorphisms (CGPs), NID1, ROCK2; pancreatic cancer: ARHGAP35, CACNA1C, GRIA3, PDAC, PALB2, KRAS, CDKN2A, TP53, and SMAD4; lung cancer: EGFR, MET, KRAS, ALK, ALK L1196M, ALK C1156Y, EML4-ALK, ERBB3, ERBB4, VEGFR, NBPF12, NTRK, ROCK2, RYR2, SCAF11, SDK2, STAT6; prostate cancer: SLC45A3, DNAH12, DSPP, KRAS, PCDH11X; ovarian cancer: DNAH14, PGR, PIK3CD, TTN; colon cancer: LAMA1, PIK3CD, TTN; bladder: RYR2; skin: BRAF V600, NRAS, NRAS Q61L / R, GNAQ, GNA11, AC1, PPP6C, RAC1, PPP6C, STK19.

[0132] In some embodiments, the methods provided herein include modulating an immune response in a subject. In some embodiments, the immune response in a subject is modulated by a method comprising: (a) administering a composition to a subject having cancer, the composition comprising at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a number of cancer-associated proteins, where prior to the administering step, the number of cancer-associated proteins are increased in presence compared to non-cancer cells of the subject or comprise a sequence modification compared to non-cancer cells of the subject; and a number of nanoparticles, each nanoparticle comprising a cationic surface, and at least one nucleic acid complexed to the cationic surface. The methods provided herein may further comprise obtaining nucleic acid or amino acid sequence information from a sample comprising cancer cells obtained from the subject. The cancer cells and non-cancer cells can be obtained from the subject by any method, including, for example, surgery, biopsy, blood draw, nasal swab, Pap test, colonoscopy, urine test, and the like. In some embodiments, the cancer cells are circulating cancer cells. Nucleic acid sequence information can be obtained from a sample containing non-cancer cells from the same subject to serve as a control. The sequence information can be compared between the cancer cell sample and the non-cancer cell sample to identify somatic mutations present in the cancer cell sequence information, thereby identifying one or more cancer-associated proteins or cancer cell markers in the subject. The sequence information can be obtained from the sample by any method, including, but not limited to, for example, sequencing, PCR, reverse transcriptase PCR (RT-PCR), proteomics, immunosorbent assay, and RNA-seq. The sequence information can be used to classify immunogenic epitopes that have one or more of the following properties: (i) the epitope is present in a transcript; (ii) the epitope is present in a protein coding region; (iii) the epitope introduces a change in amino acid sequence; and (iv) the epitope is predicted to exhibit MHC binding.These properties are useful for identifying cancer-associated proteins and cancer cell markers for use in personalized vaccine compositions for administration to subjects.After identifying cancer-associated protein epitopes in the cancer cells of a subject, the methods provided herein include generating at least one nucleic acid encoding a number of the cancer-associated proteins provided herein; and / or a nucleic acid encoding an antibody that binds to the cancer-associated proteins provided herein.

[0133] Provided herein is a method for personalized treatment of cancer in a subject, comprising: (a) receiving an assay result indicating that the subject has a tumor, the tumor comprising a cancer associated protein; and (b) administering a composition to the subject, the composition comprising at least one nucleic acid encoding the cancer associated protein in (a), thereby treating the cancer in the subject. Provided herein is a method for personalized treatment of cancer in a subject, comprising: (a) receiving an assay result indicating that the subject has a tumor, the tumor comprising a cancer associated protein; and (b) administering a composition to the subject, the composition comprising at least one nucleic acid encoding an antibody that specifically binds to the cancer associated protein in (a), thereby treating the cancer in the subject. In some embodiments, the composition further comprises a nanoparticle as described herein. In some embodiments, the nucleic acid further comprises a replicon sequence as described herein. In some embodiments, the method provides a reduction in severity, tumor size, tumor volume, or incidence of tumor development in the subject.

[0134] In some embodiments, the subject is diagnosed with a tumor necrosis factor (TNF) gene that is associated with a leukemia or bronchitis (BOB) or a bronchitis (BOB) disease. In some embodiments, the subject is diagnosed with a tumor necrosis factor (BNE) gene that is associated with a leukemia or bronchitis (BOB) or a ... xxi) alpha lactalbumin (LALBA); (xxii) cyclin D1 (CCND1); (xxiii) folate receptor 1 (FOLR1); (xxiv) telomerase (TERT); (xxv) RecQ protein-like (DNA helicase Q1-like) (RECQL); (xxvi) leptin receptor (LEPR); (xxvii) ERBB receptor feedback inhibitor 1 (ERRFI1); (xxviii) lysosomal protein transmembrane 4 alpha (LAPTM4A); (xxix) Kirsten rat sarcoma virus (K-Ras); (xxx) S100 proteins; (xxxi) cluster of differentiation (CD) family proteins; (xxxii) alpha fetoprotein (AFP); (xxxiii) epithelial tumor antigen (ETA); (xxxiv) tumor protein p53; (xxxv) ephrin receptor; (xxxvi) transferrin receptor; (xxxvii) neoglycoprotein; (xxxviii) tumor necrosis factor (TNF-α) α-receptor;(xxxvix) human papillomavirus-E6;(xl) human papillomavirus-E7;(xli) cytokeratin;(xlii) beta-catenin;(xliii) carboxypeptidase M;(xliv) EP4 receptor;(xlv) human milk fat glomerular antigen;(xlvi) tumor necrosis factor (TNF)-beta (β) receptor;(xlvii) B7-1 protein;(xlviii) B7-2 protein;(xlix) TNF receptor-associated factor 2;(l) has a tumor comprising a cancer associated protein selected from the group consisting of melanoma associated antigen 1 (MART-1) recognized by T cells; and functional fragments thereof. In some embodiments, the subject has a tumor comprising a cancer associated protein selected from Table 1;

[0135] Further provided herein is a method of modulating immune response in a subject, comprising administering a composition provided herein or a pharmaceutical composition provided herein to a subject with cancer. The compositions provided herein can also be administered prophylactically to immunize a subject against cancer. In some embodiments, the compositions described herein are used to prophylactically immunize a subject against skin cancer or lung cancer. In some embodiments, the subject is at risk of developing a cancer described herein. In some embodiments, administration provides a reduction in the occurrence, size, volume, and / or frequency of tumors in the subject, compared to a subject with a tumor without administration. kit

[0136] Provided herein is a kit comprising the composition provided herein, the pharmaceutical composition provided herein; and optionally a delivery system for administering to a subject.The kit described herein may comprise lyophilized reagents, and optionally a reagent for hydration.The kit described herein may also comprise non-lyophilized reagents.In some embodiments, the kit comprises two or more separate units, each comprising lipid carrier and nucleic acid.

[0137] In some embodiments, the kit comprises a unit comprising a lipid carrier and a nucleic acid. In some embodiments, the kit further comprises a unit comprising a reagent for hydration of the dry composition. In some embodiments, the reagent for hydration comprises water.

[0138] In some embodiments, the kit further comprises one or more surfactants. In some embodiments, the formulation of the compositions described herein is prepared in a single container for administration. In some embodiments, the formulation of the compositions described herein is prepared in two containers for administration, separating the nucleic acid from the nanoparticle carrier. As used herein, "container" includes vessels, vials, ampoules, tubes, cups, boxes, bottles, flasks, jars, dishes, wells of single-well or multi-well instruments, reservoirs, tanks, etc., or other devices in which the compositions disclosed herein can be placed, stored, and / or transported, and accessed to remove contents. Examples of such containers include glass and / or plastic sealed or resealable tubes and ampoules, including those with rubber stoppers or other sealing means compatible with withdrawal of contents using a needle and syringe. In some implementations, the container is RNase-free.

[0139] In some embodiments, the kit comprises: (a) a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, one or more lipids, and one or more surfactants; and (b) at least one nucleic acid sequence, where the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, where the antigen is a cancer-associated protein. Exemplary embodiments

[0140] Provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; and a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer associated protein. Further provided herein is a composition, the cancer associated protein is a protein expressed by a melanoma cell. Further provided herein is a composition, the nucleic acid comprises a sequence region that is at least 85% identical to one of SEQ ID NOs: 1-2, 75, 76, 88, or 89. Further provided herein is a composition, the cancer associated protein sequence or a functional variant thereof has an amino acid sequence set forth in one of SEQ ID NOs: 3-47, 77, 78, 87, 90. Further provided herein is a composition, the nucleic acid is in a complex with the lipid carrier. Further provided herein is a composition, the nucleic acid further encodes an RNA polymerase. Further provided herein is a composition, wherein the RNA polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the nucleic acid encoding the RNA polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition, wherein the liquid oil is alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxylglyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition, wherein the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerin myristate. The cationic lipids were 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA);1,2-Dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyl trimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DO EPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'' '-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-hexyldecanoate);ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide;β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol;BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl) (oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-diamino Leoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9''''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl) Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate);Further provided herein is a composition in which the lipid carrier is PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition in which the lipid carrier comprises a hydrophobic core. Further provided herein is a composition in which the lipid carrier comprises an inorganic particle. Further provided herein is a composition in which the inorganic particle is within the hydrophobic core. Further provided herein is a composition in which the inorganic particle comprises a metal. Further provided herein is a composition in which the metal comprises a metal salt, a metal oxide, a metal hydroxide, or a metal phosphate. Further provided herein is a composition in which the metal oxide comprises aluminum oxide, aluminum oxyhydroxide, iron oxide, titanium dioxide, or silicon dioxide. Further provided herein is a composition in which the hydrophobic surfactant is sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, or sorbitan trioleate. Further provided herein is a composition in which the hydrophilic surfactant is a polysorbate. Further provided herein is a composition in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition in which the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm, with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein are compositions in the form of a suspension. Further provided herein are compositions that are lyophilized.

[0141] Further provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; and a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding an antibody or a functional variant thereof. Further provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; and a surfactant, the surfactant comprising a cationic lipid; a hydrophilic surfactant; and a hydrophobic surfactant; and at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding an antibody or a functional variant thereof for cancer treatment. Further provided herein is a composition, the antibody is a cancer treatment antibody or a functional variant thereof. Further provided herein is a composition, the cancer treatment antibody is an antibody listed in Table 2. Further provided herein is a composition in which the cancer therapeutic antibody is an antibody that is atezolizumab, avelumab, bevacizumab, cemiplimab, cetuximab, daratumumab, dinutuximab, durvalumab, elotuzumab, ipilimumab, isatuximab, mogamulizumab, necitumumab, nivolumab, obinutuzumab, ofatumumab, olaratumab, panitumumab, pembrolizumab, pertuzumab, ramucirumab, rituximab, or trastuzumab. Further provided herein is a composition in which the cancer therapeutic antibody has an amino acid sequence set forth in any one of SEQ ID NOs: 48-70. Further provided herein is a composition in which the nucleic acid is in a complex with a lipid carrier. Further provided herein is a composition in which the nucleic acid further encodes an RNA-dependent polymerase. Further provided herein is a composition, wherein the RNA-dependent polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the nucleic acid encoding the RNA-dependent polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition, wherein the liquid oil is alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil,Further provided herein is a composition wherein the triglyceride is paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition wherein the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglyceryl myristate. Cationic lipids include 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyltrimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC), 1,2-dioleoyl-3-dimethylammonium-propane (DOD AP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'''-( ((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315,((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate); ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide; β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydride 1H-cyclopenta[a]phenanthren-3-ol; BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,1 3,14,15,16,17-Tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl)oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z ,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5,Hexa(octan-3-yl) 9,9',9'',9''',9'''',9'''''-((((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanoate; Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(butane-4,1-diyl))bis(hexanoate) Further provided herein is a composition in which the lipid carrier is bis(azanetriyl))tetrakis(ethane-2,1-diyl)(9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate; PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition in which the lipid carrier comprises a hydrophobic core. Further provided herein is a composition in which the lipid carrier comprises an inorganic particle. Further provided herein is a composition in which the inorganic particle is within the hydrophobic core. Further provided herein is a composition in which the inorganic particle comprises a metal. Further provided herein is a composition in which the metal comprises a metal salt, a metal oxide, a metal hydroxide, or a metal phosphate. Further provided herein is a composition in which the metal oxide comprises aluminum oxide, aluminum oxyhydroxide, iron oxide, titanium dioxide, or silicon dioxide. Further provided herein is a composition in which the hydrophobic surfactant is sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, or sorbitan trioleate. Further provided herein is a composition in which the hydrophilic surfactant is a polysorbate. Further provided herein is a composition in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition in which the lipid carrier isFurther provided herein is a composition comprising a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a composition in the form of a suspension. Further provided herein is a composition that is lyophilized.

[0142] Provided herein is a composition comprising a lipid carrier, the lipid carrier comprising a liquid oil; inorganic nanoparticles, the inorganic nanoparticles comprising iron oxide present in an amount of about 0.2 mg / ml of 12 nm iron oxide; and a surfactant, the surfactant comprising a cationic lipid; and at least one nucleic acid, the nucleic acid comprising a sequence encoding a cancer-associated protein sequence or a functional variant thereof. Provided herein is a composition further comprising about 30 mg / mL DOTAP chloride; about 37.5 mg / mL squalene; about 37 mg / ml sorbitan monostearate; about 37 mg / ml polysorbate 80; and about 10 mM sodium citrate. Provided herein is a composition further comprising a lipid carrier comprising a hydrophobic core. Provided herein is a composition further comprising ... Further provided herein is a composition in which the liquid oil is α-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition in which the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerin myristate. The cationic lipid is 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N′,N′-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA);1,2-Dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyl trimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DO EPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'' '-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-hexyldecanoate);ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide;β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol;BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl) (oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-diamino Leoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9''''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl) Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate);Further provided herein is a composition, wherein the nucleic acid is PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition, wherein the nucleic acid is in a complex with a lipid carrier. Further provided herein is a composition, wherein the nucleic acid comprises a sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89. Further provided herein is a composition, wherein the cancer-associated protein sequence or functional variant thereof has an amino acid sequence set forth in one of SEQ ID NOs: 3-47, 77, 78, 88, 89. Further provided herein is a composition, wherein the nucleic acid further encodes an RNA-dependent polymerase. Further provided herein is a composition in which the RNA-dependent polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition in which the nucleic acid encoding the RNA-dependent polymerase comprises the nucleic acid sequence of SEQ ID NO:71. Further provided herein is a composition in which the hydrophilic surfactant is a polysorbate. Further provided herein is a composition in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by a nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition in which the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm, with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein are compositions in the form of a suspension. Further provided herein are compositions that are lyophilized.

[0143] Provided herein is a composition comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising about 30 mg / mL DOTAP chloride; about 37.5 mg / ml squalene; about 37 mg / ml sorbitan monostearate; about 37 mg / ml polysorbate 80; about 10 mM sodium citrate; and about 0.2 mg Fe / ml 12 nm oleic acid coated iron oxide nanoparticles; and (b) at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer associated protein sequence or a functional variant thereof. Provided herein is a composition further comprising sucrose, optionally wherein the sucrose is present at about 50 mg. Provided herein is a composition wherein the nucleic acid is in a complex with the lipid carrier. Provided herein is a composition wherein the nucleic acid comprises a sequence as set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89. Further provided herein is a composition, wherein the cancer-associated protein sequence or functional variant thereof has an amino acid sequence set forth in one of SEQ ID NOs: 3-47, 77, 78, 87, or 90. Further provided herein is a composition, wherein the nucleic acid further encodes an RNA-dependent polymerase. Further provided herein is a composition, wherein the RNA-dependent polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the nucleic acid encoding the RNA-dependent polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition, wherein the lipid carrier comprises a hydrophobic core. Further provided herein is a composition, wherein oleic acid coated iron oxide nanoparticles are within the hydrophobic core. Further provided herein is a composition, wherein the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition, wherein the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition, wherein the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg).Further provided herein is a composition, wherein at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a composition, wherein at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a composition in the form of a suspension. Further provided herein is a composition that is lyophilized.

[0144] Provided herein is a composition comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising: DOTAP chloride present in an amount of about 0.75 mg; squalene present in an amount of about 0.94 mg; sorbitan monostearate present in an amount of about 0.93 mg; polysorbate 80 present in an amount of about 0.93 mg; citric acid monohydrate present in an amount of about 1.05 mg; and oleic acid coated iron oxide nanoparticles present in an amount of about 0.005 mg; and (b) at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a cancer associated protein sequence or a functional variant thereof. Further provided herein is a composition, the nucleic acid being in a complex with the lipid carrier. Further provided herein is a composition, the nucleic acid comprising a sequence as set forth in any one of SEQ ID NOs: 1-2, 75, 76, 88, or 89. Further provided herein is a composition, wherein the cancer-associated protein sequence or functional variant thereof has an amino acid sequence set forth in one of SEQ ID NOs: 3-47, 77, 78, 87, or 90. Further provided herein is a composition, wherein the nucleic acid further encodes an RNA-dependent polymerase. Further provided herein is a composition, wherein the RNA-dependent polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the nucleic acid encoding the RNA-dependent polymerase comprises the nucleic acid sequence of SEQ ID NO: 71. Further provided herein is a composition, wherein the lipid carrier comprises a hydrophobic core. Further provided herein is a composition, wherein oleic acid coated iron oxide nanoparticles are within the hydrophobic core. Further provided herein is a composition, wherein the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition, wherein the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition, wherein the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg).Further provided herein is a composition, wherein at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a composition, wherein at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a composition in the form of a suspension. Further provided herein is a composition that is lyophilized.

[0145] Provided herein is a composition comprising a first nucleic acid comprising a sequence encoding an RNA-dependent RNA polymerase; and a second nucleic acid comprising a sequence encoding a cancer-associated protein sequence or a functional variant thereof, wherein the cancer-associated protein sequence is at least 85% identical to one of SEQ ID NOs: 1-2, 75, 76, 88, or 89. Further provided herein is a composition wherein the cancer-associated protein sequence comprises a sequence listed in Table 1. Further provided herein is a composition wherein the cancer-associated protein sequence comprises a TRP-1 tumor-associated antigen sequence. Further provided herein is a composition wherein the first nucleic acid and the second nucleic acid are present on a common nucleic acid. Further provided herein is a composition wherein the first nucleic acid and the second nucleic acid are present on separate nucleic acids. Further provided herein is a composition wherein the RNA-dependent RNA polymerase comprises VEEV RNA polymerase. Further provided herein is a composition further comprising a nanoparticle carrier system. Further provided herein is a composition wherein the nanoparticle carrier system comprises a cationic lipid and a hydrophobic core. Further provided herein is a composition wherein the hydrophobic core comprises an inorganic nanoparticle. Further provided herein is a composition in which the first nucleic acid and / or the second nucleic acid comprises RNA. Further provided herein is a composition in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition in which the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a composition in the form of a suspension. Further provided herein are compositions that are lyophilized.

[0146] Provided herein is a composition comprising a first nucleic acid comprising a sequence encoding an RNA-dependent RNA polymerase; and a second nucleic acid comprising a sequence encoding a cancer associated protein-binding antibody or antibody fragment. Further provided herein is a composition wherein the cancer associated protein sequence is at least 85% identical to a sequence listed in Table 2. Further provided herein is a composition wherein the first nucleic acid and the second nucleic acid are present on a common nucleic acid. Further provided herein is a composition wherein the first nucleic acid and the second nucleic acid are present on separate nucleic acids. Further provided herein is a composition wherein the RNA-dependent RNA polymerase comprises VEEV RNA polymerase. Further provided herein is a composition further comprising a nanoparticle carrier system. Further provided herein is a composition wherein the nanoparticle carrier system comprises a cationic lipid and a hydrophobic core. Further provided herein is a composition wherein the hydrophobic core comprises an inorganic nanoparticle. Further provided herein is a composition wherein the first nucleic acid and / or the second nucleic acid comprises RNA. Further provided herein are compositions in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein are compositions in which the lipid carrier comprises a z-average hydrodynamic diameter ranging from about 40 nm to about 150 nm, with an average polydispersity index ranging from about 0.1 to 0.4. Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein are compositions in the form of a suspension. Further provided herein are compositions that are lyophilized.

[0147] Provided herein is a composition comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles; and one or more lipids; and (b) at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding an antigen, the antigen being a cancer-associated protein. Provided herein is a composition further comprising an additional nucleic acid comprising a sequence encoding a nucleic acid polymerase or a sequence capable of expressing a nucleic acid polymerase. Provided herein is a composition further comprising an additional nucleic acid comprising a sequence encoding an RNA polymerase or a sequence capable of expressing an RNA polymerase. Provided herein is a composition further comprising a cancer-associated protein associated with melanoma. Provided herein is a composition further comprising a cancer-associated protein sequence comprising a sequence listed in Table 1. Provided herein is a composition further comprising a cancer-associated protein being MAGE-A1. Provided herein is a composition further comprising a cancer-associated protein being MAGE-A3. Further provided herein is a composition in which the cancer-associated protein is TYRP-1 or TRP-1. Further provided herein is a composition in which the hydrophobic core comprises an oil. Further provided herein is a composition in which the oil comprises at least one of alpha-tocopherol, lauroyl polyoxyl glyceride, monoacyl glycerol, propolis, squalene, mineral oil, grape seed oil, olive oil, paraffin oil, peanut oil, soybean oil, sunflower oil, soy lecithin, triglyceride, and vitamin E, and medium chain triglyceride. Further provided herein is a composition in which the one or more inorganic nanoparticles are selected from the group consisting of metal salts, metal oxides, metal hydroxides, metal phosphates, and any combination thereof. Further provided herein is a composition in which the one or more lipids are selected from the group consisting of cationic lipids, anionic lipids, neutral lipids, and any combination thereof. Further provided herein is a composition in which the one or more lipids are cationic lipids.Cationic lipids include 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP); dipalmitoyl (C16:0) trimethylammonium propane (DPTAP); distearoyltrimethylammonium propane (DSTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N trimethylammonium chloride (DOTMA); N,N-dioleoyl-N Further provided herein is a composition in which the lipid carrier is selected from the group consisting of 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA); 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol)(C12-200), and any combination thereof. Further provided herein is a composition in which the lipid carrier optionally comprises one or more surfactants. Further provided herein is a composition in which the one or more surfactants are selected from the group consisting of hydrophobic surfactants, hydrophilic surfactants, and any combination thereof. Further provided herein is a composition in which the hydrophobic surfactant comprises a sorbitan ester selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate; and the hydrophilic surfactant comprises a polysorbate. Further provided herein is a composition in which the lipid carrier has a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm, with an average polydispersity index in the range of about 0.1 to about 0.4. Further provided herein is a composition in which one or more nucleic acids are incorporated into or complexed with the lipid carrier to form a lipid carrier-nucleic acid complex.Further provided herein are compositions in which the lipid carrier-RNA complex is formed through non-covalent interactions or through reversible covalent interactions. Further provided herein are compositions in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein are compositions that are stable at 2-8 degrees Celsius. Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein are compositions in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein are compositions in the form of a suspension. Further provided herein are compositions that are lyophilized.

[0148] Provided herein is a vaccine comprising: (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles, and one or more lipids; and (b) at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding an antigen, the antigen being a cancer-associated protein. Provided herein further is a vaccine, the nucleic acid being RNA. Provided herein further is a vaccine, the vaccine comprising an additional nucleic acid comprising a sequence encoding a nucleic acid polymerase, or a sequence capable of expressing a nucleic acid polymerase. Provided herein further is a vaccine, the vaccine comprising an additional nucleic acid comprising a sequence encoding an RNA polymerase, or a sequence capable of expressing an RNA polymerase. Provided herein further is a vaccine, the cancer-associated protein is associated with melanoma. Provided herein further is a composition, the cancer-associated protein sequence comprises a sequence listed in Table 1. Provided herein further is a vaccine, the cancer-associated protein is MAGE-A1. Provided herein further is a vaccine, the cancer-associated protein is MAGE-A3. Further provided herein is a vaccine, wherein the cancer-associated protein is TYRP-1 or TRP-1. Further provided herein is a vaccine, wherein the hydrophobic core comprises an oil. Further provided herein is a vaccine, wherein the oil comprises at least one of alpha-tocopherol, lauroyl polyoxyl glyceride, monoacyl glycerol, propolis, squalene, mineral oil, grape seed oil, olive oil, paraffin oil, peanut oil, soybean oil, sunflower oil, soy lecithin, triglyceride, and vitamin E, and medium chain triglyceride. Further provided herein is a vaccine, wherein the one or more inorganic nanoparticles are selected from the group consisting of metal salts, metal oxides, metal hydroxides, metal phosphates, and any combination thereof. Further provided herein is a vaccine, wherein the one or more lipids are selected from the group consisting of cationic lipids, anionic lipids, neutral lipids, and any combination thereof. Further provided herein is a vaccine, wherein the one or more lipids are cationic lipids.Cationic lipids include 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP); dipalmitoyl (C16:0) trimethylammonium propane (DPTAP); distearoyltrimethylammonium propane (DSTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N trimethylammonium chloride (DOTMA); N,N-dioleoyl-N Further provided herein is a vaccine in which the lipid carrier is selected from the group consisting of: 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DODAC); 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA); 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol)(C12-200), and any combination thereof. Further provided herein is a vaccine in which the lipid carrier optionally comprises one or more surfactants. Further provided herein is a vaccine in which the one or more surfactants are selected from the group consisting of hydrophobic surfactants, hydrophilic surfactants, and any combination thereof. Further provided herein is a vaccine, wherein the hydrophobic surfactant comprises a sorbitan ester selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate; and the hydrophilic surfactant comprises a polysorbate. Further provided herein is a vaccine, wherein the lipid carrier has a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm, with an average polydispersity index in the range of about 0.1 to about 0.4. Further provided herein is a vaccine, wherein one or more nucleic acids are incorporated into or complexed with the lipid carrier to form a lipid carrier-nucleic acid complex.Further provided herein is a vaccine in which the lipid carrier-RNA complex is formed through a non-covalent interaction or through a reversible covalent interaction. Further provided herein is a vaccine in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by the nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a vaccine in which the composition is stable at 2-8 degrees Celsius. Further provided herein is a vaccine in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein is a vaccine in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a vaccine in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a vaccine in the form of a suspension. Further provided herein is a vaccine that is lyophilized.

[0149] Provided herein is a composition for immune protection of a subject, comprising a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; and at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding an antigen capable of expressing, the antigen being a cancer-associated protein. Provided herein further is a composition, the cancer-associated protein sequence comprising a sequence listed in Table 1. Provided herein further is a vaccine, the hydrophobic core comprising an oil. Provided herein further is a vaccine, the oil comprising at least one of alpha-tocopherol, lauroyl polyoxyl glyceride, monoacyl glycerol, propolis, squalene, mineral oil, grape seed oil, olive oil, paraffin oil, peanut oil, soybean oil, sunflower oil, soybean lecithin, triglyceride, and vitamin E, and medium-chain triglyceride. Further provided herein is a vaccine, wherein the one or more inorganic nanoparticles are selected from the group consisting of metal salts, metal oxides, metal hydroxides, metal phosphates, and any combination thereof. Further provided herein is a vaccine, wherein the one or more lipids are selected from the group consisting of cationic lipids, anionic lipids, neutral lipids, and any combination thereof. Further provided herein is a vaccine, wherein the one or more lipids are cationic lipids.Cationic lipids include 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP); dipalmitoyl (C16:0) trimethylammonium propane (DPTAP); distearoyltrimethylammonium propane (DSTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N trimethylammonium chloride (DOTMA); N,N-dioleoyl-N Further provided herein is a vaccine in which the lipid carrier is selected from the group consisting of: 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DODAC); 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA); 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol)(C12-200), and any combination thereof. Further provided herein is a vaccine in which the lipid carrier optionally comprises one or more surfactants. Further provided herein is a vaccine in which the one or more surfactants are selected from the group consisting of hydrophobic surfactants, hydrophilic surfactants, and any combination thereof. Further provided herein is a vaccine, wherein the hydrophobic surfactant comprises a sorbitan ester selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate; and the hydrophilic surfactant comprises a polysorbate. Further provided herein is a composition, wherein the molar ratio of lipid carrier, characterized by the nitrogen to phosphate (N:P) molar ratio, to one or more nucleic acids ranges from about 1:1 to about 150:1.Further provided herein is a composition in which the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a composition in the form of a suspension. Further provided herein is a composition that is lyophilized.

[0150] Provided herein is a dry composition comprising the composition provided herein; and at least one cryoprotectant. Further provided herein is a dry composition, wherein the composition is freeze-dried. Further provided herein is a dry composition, wherein the composition is spray-dried. Further provided herein is a dry composition, wherein the composition is thermally stable. Further provided herein is a dry composition, wherein the composition is thermally stable at about 25°C. Further provided herein is a dry composition, wherein the composition is thermally stable at about 45°C. Further provided herein is a dry composition, wherein the composition is thermally stable at about -20°C. Further provided herein is a dry composition, wherein the composition is thermally stable at about 2°C to about 8°C. Further provided herein is a dry composition, wherein the composition is thermally stable for at least one week, at least two weeks, and / or at least one month. Further provided herein is a dry composition, wherein the hydrophobic core comprises an oil. Further provided herein is a dry composition, wherein the oil comprises at least one of α-tocopherol, lauroyl polyoxyl glyceride, monoacyl glycerol, propolis, squalene, mineral oil, grape seed oil, olive oil, paraffin oil, peanut oil, soybean oil, sunflower oil, soy lecithin, triglyceride, and vitamin E, and medium chain triglyceride. Further provided herein is a dry composition, wherein the one or more inorganic nanoparticles are selected from the group consisting of metal salts, metal oxides, metal hydroxides, metal phosphates, and any combination thereof. Further provided herein is a dry composition, wherein the one or more lipids are selected from the group consisting of cationic lipids, anionic lipids, neutral lipids, and any combination thereof. Further provided herein is a dry composition, wherein the one or more lipids are cationic lipids.Cationic lipids include 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP); dipalmitoyl (C16:0) trimethylammonium propane (DPTAP); distearoyltrimethylammonium propane (DSTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N trimethylammonium chloride (DOTMA); N,N-dioleoyl-N Further provided herein is a vaccine in which the lipid carrier is selected from the group consisting of 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA); 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol)(C12-200), and any combination thereof. Further provided herein is a dry composition in which the lipid carrier optionally comprises one or more surfactants. Further provided herein is a dry composition in which the one or more surfactants are selected from the group consisting of hydrophobic surfactants, hydrophilic surfactants, and any combination thereof. Further provided herein is a dry composition, wherein the hydrophobic surfactant comprises a sorbitan ester selected from the group consisting of sorbitan monostearate, sorbitan monooleate, and sorbitan trioleate; and the hydrophilic surfactant comprises a polysorbate. Further provided herein is a dry composition, wherein the molar ratio of lipid carrier to one or more nucleic acids, characterized by a nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1.Further provided herein is a dry composition, wherein the lipid carrier comprises a z-average hydrodynamic diameter ranging from about 40 nm to about 150 nm, with an average polydispersity index ranging from about 0.1 to 0.4. Further provided herein is a dry composition, wherein the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein is a dry composition, wherein the at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a dry composition, wherein the at least one nucleic acid sequence is present in an amount of up to about 25 μg.

[0151] Provided herein is a composition for the prevention of cancer, comprising a sorbitan fatty acid ester, an ethoxylated sorbitan ester, a cationic lipid, an immunostimulant, and at least one RNA or functional fragment thereof encoding an antigen sequence, wherein the sorbitan fatty acid ester is sorbitan monostearate. Further provided herein is a composition wherein the ethoxylated sorbitan ester is polysorbate 80. Further provided herein is a composition wherein the cationic lipid is DOTAP. Further provided herein is a composition wherein the immunostimulant is squalene. Further provided herein is a composition wherein the RNA encodes a cancer-associated protein. Further provided herein is a composition wherein the immunostimulant reduces the total amount of protein produced but increases the immune response to a vaccine. Further provided herein is a composition wherein the immunostimulant increases the total amount of protein produced but decreases the immune response to a vaccine. Further provided herein is a composition wherein the immunostimulant is Miglyol 810 or Miglyol 812. Further provided herein is a composition comprising squalene and not comprising solid particles. Further provided herein is a composition, wherein the ratio of esters results in a hydrophilic-lipophilic balance of 8 to 11. Further provided herein is a composition, wherein the particle size is 30 to 200 nanometers. Further provided herein is a composition, wherein the ratio of N to P is 5 to 35. Further provided herein is a composition, wherein the antigen sequence comprises a sequence listed in Table 1. Further provided herein is a composition, which is in the form of a suspension. Further provided herein is a composition, which is lyophilized.

[0152] Provided herein is a composition for the prevention of cancer, comprising sorbitan monostearate (e.g., SPAN® 60), polysorbate 80 (e.g., TWEEN® 80), DOTAP, an immunostimulant, and at least one RNA or functional fragment thereof encoding an antigen sequence. Further provided herein is a composition, wherein the immunostimulant reduces the total amount of protein produced, but increases the immune response to a vaccine. Further provided herein is a composition, wherein the immunostimulant increases the total amount of protein produced, but decreases the immune response to a vaccine. Further provided herein is a composition, wherein the immunostimulant is Miglyol 810 or Miglyol 812. Further provided herein is a composition, comprising squalene, and not comprising solid particles. Further provided herein is a composition, wherein the ratio of esters produces a hydrophilic-lipophilic balance of 8-11. Further provided herein is a composition, wherein the particle size is 30-200 nanometers. Further provided herein is a composition, wherein the ratio of N to P is 5-35. Further provided herein is a composition, wherein the antigen sequence comprises a sequence listed in Table 1. Further provided herein is a composition, wherein the composition is in the form of a suspension. Further provided herein is a composition, wherein the composition is lyophilized. Further provided herein is a pharmaceutical composition, comprising the composition provided herein; and a pharma- ceutically acceptable excipient. Further provided herein is a pharmaceutical composition, comprising: (a) a lipid carrier, wherein the lipid carrier is a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles; and one or more lipids; and (b) at least one nucleic acid sequence, wherein the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, and wherein the antigen is a cancer-associated protein.

[0153] Provided herein are kits that include the compositions provided herein.

[0154] Provided herein is a method of generating an immune response in a subject, comprising administering to the subject a composition provided herein, thereby generating an immune response against a cancer-associated protein. Provided herein is a method of generating an immune response in a subject, comprising administering to the subject (a) a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles, and one or more lipids; and (b) at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding an antigen capable of expressing, the antigen being a cancer-associated protein. Provided herein is a method in which the composition is administered to the subject in two doses. Provided herein is a method in which the second dose is administered about 28 days after the first dose. Provided herein is a method in which the composition further comprises administering to the subject a third dose. Provided herein is a method in which 5 μg of the composition is administered to the subject. Provided herein is a method in which 10 μg of the composition is administered to the subject. Further provided herein is a method in which 25 μg of the composition is administered to the subject. Further provided herein is a method in which the composition is administered via intramuscular, intranasal, oral, subcutaneous, intratumoral, intrathecal, or intravenous injection. Further provided herein is a method in which the subject is a human. Further provided herein is a method in which the subject has, is at risk of, or is suspected of having cancer. Further provided herein is a method in which the subject has a solid tumor or a blood cancer. Further provided herein is a method in which the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a method in which the blood cancer is a lymphoma or leukemia. Further provided herein is a method in which the subject has, is at risk of, or is suspected of having a skin cancer. Further provided herein is a method in which the skin cancer is a basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma.

[0155] Further provided herein is a method of prophylactically immunizing a subject against cancer, comprising administering to the subject a composition provided herein, thereby immunizing the subject against a cancer expressing a cancer-associated protein. Further provided herein is a method in which the subject is at risk of developing skin cancer. Further provided herein is a method in which 5 μg of the composition is administered to the subject. Further provided herein is a method in which 10 μg of the composition is administered to the subject. Further provided herein is a method in which 25 μg of the composition is administered to the subject. Further provided herein is a method in which the composition is administered via intramuscular, intranasal, oral, subcutaneous, intratumoral, intrathecal, or intravenous injection. Further provided herein is a method in which the subject is a human. Further provided herein is a method in which the skin cancer is basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a method, wherein the cancer expresses a cancer associated protein, and the cancer associated protein is a TRP-1 protein.

[0156] Further provided herein is a method of reducing the severity of cancer, the method comprising administering a composition comprising a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, optionally one or more inorganic nanoparticles, and one or more lipids, and at least one nucleic acid sequence, where the nucleic acid sequence comprises a sequence encoding a sequence capable of expressing an antigen, and the antigen is a cancer-associated protein.

[0157] Further provided herein is a method of immunoprotecting a subject, the method comprising administering to the subject a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids, at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding a sequence capable of expressing an antigen, wherein the antigen is a cancer-associated protein.

[0158] Provided herein is a method of generating an immune response in a subject, comprising administering to said subject a composition comprising a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids, and at least one nucleic acid sequence, the nucleic acid sequence comprising a sequence encoding a sequence capable of expressing an antigen, the antigen being a cancer-associated protein. Further provided herein is a composition in which the molar ratio of lipid carrier to one or more nucleic acids, characterized by a nitrogen to phosphate (N:P) molar ratio, ranges from about 1:1 to about 150:1. Further provided herein is a composition in which the lipid carrier comprises a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm, with an average polydispersity index in the range of about 0.1 to 0.4. Further provided herein is a composition in which the at least one nucleic acid sequence is present in an amount of up to about 100 micrograms (μg). Further provided herein is a composition, wherein at least one nucleic acid sequence is present in an amount of up to about 5, about 10, about 25, about 50, or about 100 micrograms (μg). Further provided herein is a composition, wherein at least one nucleic acid sequence is present in an amount of up to about 25 μg. Further provided herein is a composition in the form of a suspension. Further provided herein is a composition that is lyophilized.

[0159] Further provided herein is a method for preparing a lyophilized composition, the method comprising the steps of obtaining a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, where the nucleic acid has a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; and lyophilizing the formulation to form a lyophilized composition.

[0160] Further provided herein is a method for preparing a spray-dried composition, the method comprising the steps of obtaining a lipid carrier, where the lipid carrier is a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, where the nucleic acid has a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, 76, 88, or 89; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; and spray drying the formulation to form the spray-dried composition.

[0161] The present invention also relates to a method for reconstituting a freeze-dried composition, comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, the nucleic acid having a sequence comprising an antigen sequence as set forth in one of SEQ ID NOs: 1-2, 75, or 76; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; freeze-drying the formulation to form a freeze-dried composition; and reconstituting the freeze-dried composition in a suitable diluent. Further provided herein is a method in which the diluent is aqueous. Further provided herein is a method in which the diluent is water. Further provided herein is a method in which the freeze-dried composition is thermally stable. Further provided herein is a method in which the freeze-dried composition is thermally stable at a temperature of up to about 25° C. Further provided herein is a method in which the freeze-dried composition is thermally stable at a temperature of up to about 45° C. Further provided herein is a method, wherein the freeze-dried composition is thermally stable at a temperature of at least about -20°C. Further provided herein is a method, wherein the freeze-dried composition is thermally stable at a temperature ranging from about 2°C to about 8°C. Further provided herein is a method, wherein the freeze-dried composition is thermally stable for at least one week, at least two weeks, and / or at least one month. Further provided herein is a method, wherein the hydrophobic core comprises an oil. Further provided herein is a method, wherein the oil comprises at least one of α-tocopherol, lauroyl polyoxylglyceride, monoacylglycerol, propolis, squalene, mineral oil, grape seed oil, olive oil, paraffin oil, peanut oil, soybean oil, sunflower oil, soybean lecithin, triglyceride, vitamin E, medium chain triglyceride, dihydroisosqualene (DHIS), farnesene, and squalane. Further provided herein is a method, wherein the one or more inorganic nanoparticles are selected from the group consisting of metal salts, metal oxides, metal hydroxides, metal phosphates, metalloids, and any combination thereof.Further provided herein is a method, wherein the one or more lipids are selected from the group consisting of cationic lipids, anionic lipids, neutral lipids, and any combination thereof. Further provided herein is a method, wherein the one or more lipids are cationic lipids. Cationic lipids include 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP); dipalmitoyl (C16:0) trimethylammonium propane (DPTAP); distearoyltrimethylammonium propane (DSTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N trimethylammonium chloride (DOTMA); N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC); 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium chloride (DSTAP); and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLin-DMA); 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE); N-decyl-N,N-dimethyldecane-1-aminium bromide (DDAB); 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate (DOSPA); ethylphosphatidylcholine (ePC); and any combination thereof. Further provided herein is a method, wherein the lipid carrier optionally comprises one or more surfactants. Further provided herein is a method, wherein the one or more surfactants are selected from the group consisting of hydrophobic surfactants, hydrophilic surfactants, and any combination thereof.Further provided herein is a method, wherein the hydrophobic surfactant comprises a sorbitan ester selected from the group consisting of SPAN® 20, SPAN® 40, SPAN® 60, SPAN® 65, SPAN® 80 and SPAN® 85; and the hydrophilic surfactant comprises a polysorbate. Further provided herein is a method, wherein the lipid carrier has a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to about 0.4. Further provided herein is a method, wherein the one or more nucleic acids are RNA. Further provided herein is a method, wherein the RNA is a self-replicating RNA. Further provided herein is a method, wherein the one or more nucleic acids comprise a sequence encoding an antigen, the antigen being derived from a virus. Further provided herein is a method, wherein the virus is an oncovirus. Further provided herein is a method, wherein the RNA encodes an amino acid sequence that is at least 80% identical to an amino acid sequence of MAGE-A1 or MAGE-A3. Further provided herein is a method, wherein the RNA encodes an amino acid sequence that is at least 80% identical to the amino acid sequence of TRYP-1 or TRP-1. Further provided herein is a method, wherein one or more nucleic acids are incorporated into or complexed with a lipid carrier to form a lipid carrier-nucleic acid complex. Further provided herein is a method, wherein the lipid carrier-nucleic acid complex is formed via a non-covalent interaction or via a reversible covalent interaction. Further provided herein is a method, wherein the at least one cryoprotectant is selected from the group consisting of sucrose, maltose, trehalose, mannitol, glucose, and any combination thereof. Further provided herein is a method, wherein the at least one cryoprotectant is sucrose. Further provided herein is a method, wherein the at least one cryoprotectant is about 1% w / v to about 20% w / v. Further provided herein is a method, wherein the at least one cryoprotectant is about 10% w / v to about 20% w / v. Further provided herein is a method, wherein the at least one cryoprotectant is about 10% w / v.

[0162] Further provided herein is a method for reconstituting a spray-dried composition, the method comprising the steps of obtaining a lipid carrier, the lipid carrier being a nanoemulsion comprising a hydrophobic core, one or more inorganic nanoparticles, and one or more lipids; incorporating at least one nucleic acid into the lipid carrier to form a lipid carrier-nucleic acid complex, the nucleic acid having a sequence comprising an antigen sequence set forth in one of SEQ ID NOs: 1-2, 75, or 76; adding at least one cryoprotectant to the lipid carrier-nucleic acid complex to form a formulation; spray drying the formulation to form a spray-dried composition; and reconstituting the spray-dried composition in a suitable diluent. Further provided herein is a method in which the diluent is aqueous. Further provided herein is a method in which the diluent is water. Further provided herein is a method in which the spray-dried composition is thermally stable. Further provided herein is a method in which the spray-dried composition is thermally stable at a temperature of up to about 25° C. Further provided herein is a method in which the spray-dried composition is thermally stable at a temperature of up to about 45° C. Further provided herein is a method, wherein the spray-dried composition is thermally stable at a temperature of at least about -20°C. Further provided herein is a method, wherein the spray-dried composition is thermally stable at a temperature ranging from about 2°C to about 8°C. Further provided herein is a method, wherein the spray-dried composition is thermally stable for at least one week, at least two weeks, and / or at least one month. Further provided herein is a method, wherein the hydrophobic core comprises an oil. Further provided herein is a method, wherein the oil comprises at least one of α-tocopherol, lauroyl polyoxylglyceride, monoacylglycerol, propolis, squalene, mineral oil, grape seed oil, olive oil, paraffin oil, peanut oil, soybean oil, sunflower oil, soybean lecithin, triglyceride, vitamin E, medium chain triglyceride, dihydroisosqualene (DHIS), farnesene, and squalane. Further provided herein is a method, wherein the one or more inorganic nanoparticles are selected from the group consisting of metal salts, metal oxides, metal hydroxides, metal phosphates, metalloids, and any combination thereof.Further provided herein is a method, wherein the one or more lipids are selected from the group consisting of cationic lipids, anionic lipids, neutral lipids, and any combination thereof. Further provided herein is a method, wherein the one or more lipids are cationic lipids. Cationic lipids include 1,2-dioleoyloxy-3-(trimethylammonium)propane (DOTAP); 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol (DC cholesterol); dimethyldioctadecylammonium (DDA); 1,2-dimyristoyl-3-trimethylammonium propane (DMTAP); dipalmitoyl (C16:0) trimethylammonium propane (DPTAP); distearoyltrimethylammonium propane (DSTAP); N-[1-(2,3-dioleyloxy)propyl]-N,N,N trimethylammonium chloride (DOTMA); N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC); 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DOEPC); 1,2-dioleoyl-3-dimethylammonium chloride (DSTAP); and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLin-DMA); 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE); N-decyl-N,N-dimethyldecane-1-aminium bromide (DDAB); 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate (DOSPA); ethylphosphatidylcholine (ePC); and any combination thereof. Further provided herein is a method, wherein the lipid carrier optionally comprises one or more surfactants. Further provided herein is a method, wherein the one or more surfactants are selected from the group consisting of hydrophobic surfactants, hydrophilic surfactants, and any combination thereof.Further provided herein is a method, wherein the hydrophobic surfactant comprises a sorbitan ester selected from the group consisting of SPAN® 20, SPAN® 40, SPAN® 60, SPAN® 65, SPAN® 80 and SPAN® 85; and the hydrophilic surfactant comprises a polysorbate. Further provided herein is a method, wherein the lipid carrier has a z-average hydrodynamic diameter in the range of about 40 nm to about 150 nm with an average polydispersity index in the range of about 0.1 to about 0.4. Further provided herein is a method, wherein the one or more nucleic acids are RNA. Further provided herein is a method, wherein the RNA is a self-replicating RNA. Further provided herein is a method, wherein the one or more nucleic acids comprise a sequence encoding an antigen, the antigen being derived from a virus. Further provided herein is a method, wherein the virus is an oncovirus. Further provided herein is a method, wherein the RNA encodes an amino acid sequence that is at least 80% identical to an amino acid sequence of MAGE-A1 or MAGE-A3. Further provided herein is a method, wherein the RNA encodes an amino acid sequence that is at least 80% identical to the amino acid sequence of TRYP-1 or TRP-1. Further provided herein is a method, wherein one or more nucleic acids are incorporated into or complexed with a lipid carrier to form a lipid carrier-nucleic acid complex. Further provided herein is a method, wherein the lipid carrier-nucleic acid complex is formed via a non-covalent interaction or via a reversible covalent interaction. Further provided herein is a method, wherein the at least one cryoprotectant is selected from the group consisting of sucrose, maltose, trehalose, mannitol, glucose, and any combination thereof. Further provided herein is a method, wherein the at least one cryoprotectant is sucrose. Further provided herein is a method, wherein the at least one cryoprotectant is about 1% w / v to about 20% w / v. Further provided herein is a method, wherein the at least one cryoprotectant is about 10% w / v to about 20% w / v. Further provided herein is a method, wherein the at least one cryoprotectant is about 10% w / v.

[0163] Provided herein is a composition comprising at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a plurality of cancer-associated proteins or functional fragments thereof; and an RNA polymerase complex region; and a plurality of nanoparticles, each nanoparticle comprising a cationic surface, and at least one nucleic acid being complexed to the cationic surface. Further provided herein is a composition, the at least one nucleic acid is RNA or DNA. Further provided herein is a composition, the plurality of cancer-associated proteins are expressed by one or more cancer cells of the subject. Further provided herein is a composition, the subject has a solid tumor or a blood cancer. Further provided herein is a composition, the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a composition, the blood cancer is a lymphoma or leukemia. Further provided herein is a composition, the subject has a skin cancer. Further provided herein is a composition, the skin cancer is a basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a composition in which the subject has lung cancer. Further provided herein is a composition in which the lung cancer is non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC). Further provided herein is a composition in which the NSCLC is adenocarcinoma, squamous cell carcinoma, large cell carcinoma, adenosquamous cell carcinoma, or sarcomatoid carcinoma. Further provided herein is a composition in which the subject has pancreatic cancer. Further provided herein is a composition in which the pancreatic cancer is pancreatic adenocarcinoma, pancreatic exocrine carcinoma, pancreatic neuroendocrine carcinoma, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. Further provided herein is a composition in which the subject has metastatic cancer. At least one nucleic acid encodes two or more cancer-associated proteins, the two or more cancer-associated proteins being: (i) epidermal growth factor receptor (EGFR); (ii) vascular endothelial growth factor (VEGF); (iii) Wilms tumor 1 (WT1); (iv) preferentially expressed antigen in melanoma (PRAME); (v) PR1; (vi) proteinase 3; (vii) elastase; (viii) cathepsin G; (ix) survivin;(x) New York Esophagus 1 (NY-Eso-1); (xi) Melanoma Associated Antigen (MAGE); (xii) Tyrosinase; (xiii) Glycoprotein 100 (gp100); (xiv) Carcinoembryonic Antigen (CEA); (xv) Mucin; (xvi) Fibroblast Growth Factor (FGF); (xvii) Programmed Cell Death Protein (PD-1); (xviii) Metastatic Tumor Antigen (MTA); (xix) Human Epidermal Growth Factor Receptor 2 (Her2); (xx) Mammaglobin A (SCGB2A2); (xxi) Alpha (xxii) cyclin D1 (CCND1); (xxiii) folate receptor 1 (FOLR1); (xxiv) telomerase (TERT); (xxv) RecQ protein-like (DNA helicase Q1-like) (RECQL); (xxvi) leptin receptor (LEPR); (xxvii) ERBB receptor feedback inhibitor 1 (ERRFI1); (xxviii) lysosomal protein transmembrane 4 alpha (LAPTM4A); (xxix) Karsten rat sarcoma virus (K-Ras); (xxx) S100 protein; (xxxi) cluster of differentiation (CD) family protein; (xxxii) alpha fetoprotein (AFP); (xxxiii) epithelial tumor antigen (ETA); (xxxiv) tumor protein p53; (xxxv) ephrin receptor; (xxxvi) transferrin receptor; (xxxvii) neoglycoprotein; (xxxviii) tumor necrosis factor (TNF)-alpha (α) receptor; (xxxvix) human papilloma Virus-E6;(xl) human papillomavirus-E7;(xli) cytokeratin;(xlii) beta-catenin;(xliii) carboxypeptidase M;(xliv) EP4 receptor;(xlv) human milk fat glomerular antigen;(xlvi) tumor necrosis factor (TNF)-beta (β) receptor;(xlvii) B7-1 protein;(xlviii) B7-2 protein;(xlix) TNF receptor-associated factor 2;(l) melanoma-associated antigen recognized by T cells 1 (MART-1);Further provided herein is a composition selected from the group consisting of MAGE-A1, MAGE-A3, MART-1 / Melan-A, MAGE-A, MAGE-B, MAGE-C, MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A5, MAGE-A6, MAGE-A7, MAGE-A8, MAGE-A9, MAGE-A10, MAGE-A11, or MAGE-A12. Further provided herein is a composition wherein the cluster of differentiation family protein is CD5, CD19, CD20, CD22, CD23, CD25, CD27, CD30, CD33, CD36, CD46, CD52, CD79a, CD79b, CD123, or CD317. Further provided herein is a composition, wherein the RNA polymerase complex region is downstream of a subgenomic promoter derived from an alphavirus. Further provided herein is a composition, wherein the RNA polymerase complex region encodes an RNA-dependent RNA polymerase. Further provided herein is a composition, wherein the RNA-dependent RNA polymerase is Venezuelan equine encephalitis virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the cationic surface comprises a cationic lipid. The cationic lipid is 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N′,N′-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA);1,2-Dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyl trimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DO EPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'' '-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-hexyldecanoate);ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide;β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol;BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl) (oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-diamino Leoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9''''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl) Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate);Further provided herein is a composition, wherein the nanoparticle is PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition, wherein each nanoparticle further comprises a hydrophobic core. Further provided herein is a composition, wherein the hydrophobic core comprises an oil. Further provided herein is a composition, wherein the oil is in a liquid phase. Further provided herein is a composition in which the oil comprises alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxylglyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition in which the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerin myristate. Further provided herein is a composition in which each nanoparticle comprises a cationic lipid and an oil. Further provided herein is a composition in which each nanoparticle further comprises a surfactant. Further provided herein is a composition in which the surfactant is a polysorbate, a phosphorus-terminated surfactant, a carboxylate-terminated surfactant, a sulfate-terminated surfactant, an amine-terminated surfactant, a trioctylphosphatase; Further provided herein is a composition that is sphingosine oxide (TOPO) or distearyl phosphatidic acid (DSPA). Further provided herein is a composition that is lyophilized. Further provided herein is a composition that is in the form of a liquid, semi-liquid, solution, spray, or powder. Further provided herein is a composition that is formulated as a suspension.

[0164] Provided herein is a composition comprising a plurality of nucleic acids, the plurality of nucleic acids comprising sequences separately encoding a plurality of cancer-associated proteins; and an RNA polymerase complex region; and a plurality of nanoparticles, each nanoparticle comprising a cationic surface, and at least one nucleic acid being complexed to the cationic surface. Further provided herein is a composition, in which at least one nucleic acid is RNA or DNA. Further provided herein is a composition, in which the plurality of cancer-associated proteins are expressed by one or more cancer cells of the subject. Further provided herein is a composition, in which the subject has a solid tumor or a blood cancer. Further provided herein is a composition, in which the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a composition, in which the blood cancer is a lymphoma or leukemia. Further provided herein is a composition, in which the subject has a skin cancer. Further provided herein is a composition, in which the skin cancer is a basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a composition in which the subject has lung cancer. Further provided herein is a composition in which the lung cancer is non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC). Further provided herein is a composition in which the NSCLC is adenocarcinoma, squamous cell carcinoma, large cell carcinoma, adenosquamous cell carcinoma, or sarcomatoid carcinoma. Further provided herein is a composition in which the subject has pancreatic cancer. Further provided herein is a composition in which the pancreatic cancer is pancreatic adenocarcinoma, pancreatic exocrine carcinoma, pancreatic neuroendocrine carcinoma, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. Further provided herein is a composition in which the subject has metastatic cancer. At least one nucleic acid encodes two or more cancer-associated proteins, the two or more cancer-associated proteins being: (i) epidermal growth factor receptor (EGFR); (ii) vascular endothelial growth factor (VEGF); (iii) Wilms' tumor 1 (WT1); (iv) preferentially expressed antigen in melanoma (PRAME); (v) PR1; (vi) proteinase 3; (vii) elastase; (viii) cathepsin G; (ix) survivin; (x) New York esophagus 1 (NY-Eso-1);(xi) melanoma associated antigen (MAGE); (xii) tyrosinase; (xiii) glycoprotein 100 (gp100); (xiv) carcinoembryonic antigen (CEA); (xv) mucin; (xvi) fibroblast growth factor (FGF); (xvii) programmed cell death protein (PD-1); (xviii) metastatic tumor antigen (MTA); (xix) human epidermal growth factor receptor 2 (Her2); (xx) mammaglobin A (SCGB2A2); (xxi) alpha-lactalbumin (LALBA) );(xxii) cyclin D1 (CCND1);(xxiii) folate receptor 1 (FOLR1);(xxiv) telomerase (TERT);(xxv) RecQ protein-like (DNA helicase Q1-like) (RECQL);(xxvi) leptin receptor (LEPR);(xxvii) ERBB receptor feedback inhibitor 1 (ERRFI1);(xxviii) lysosomal protein transmembrane 4 alpha (LAPTM4A);(xxix) Kirsten rat sarcoma virus (K-Ras);(xxx) S100 proteins;(xxxi) cluster of differentiation (CD) family proteins;(xxxii) alpha fetoprotein (AFP);(xxxiii) epithelial tumor antigen (ETA);(xxxiv) tumor protein p53;(xxxv) ephrin receptor;(xxxvi) transferrin receptor;(xxxvii) neoglycoprotein;(xxxviii) tumor necrosis factor (TNF)-alpha (α) receptor;(xxxvix) human papillomavirus- E6;(xl) human papillomavirus-E7;(xli) cytokeratin;(xlii) beta-catenin;(xliii) carboxypeptidase M;(xliv) EP4 receptor;(xlv) human milk fat glomerular antigen;(xlvi) tumor necrosis factor (TNF)-beta (β) receptor;(xlvii) B7-1 protein;(xlviii) B7-2 protein;(xlix) TNF receptor-associated factor 2;(l) melanoma-associated antigen recognized by T cells 1 (MART-1);Further provided herein is a composition selected from the group consisting of MAGE-A1, MAGE-A3, MART-1 / Melan-A, MAGE-A, MAGE-B, MAGE-C, MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A5, MAGE-A6, MAGE-A7, MAGE-A8, MAGE-A9, MAGE-A10, MAGE-A11, or MAGE-A12. Further provided herein is a composition wherein the cluster of differentiation family protein is CD5, CD19, CD20, CD22, CD23, CD25, CD27, CD30, CD33, CD36, CD46, CD52, CD79a, CD79b, CD123, or CD317. Further provided herein is a composition, wherein the RNA polymerase complex region is downstream of a subgenomic promoter derived from an alphavirus. Further provided herein is a composition, wherein the RNA polymerase complex region encodes an RNA-dependent RNA polymerase. Further provided herein is a composition, wherein the RNA-dependent RNA polymerase is Venezuelan equine encephalitis virus (VEEV) RNA polymerase. Further provided herein is a composition, wherein the cationic surface comprises a cationic lipid. The cationic lipid is 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N′,N′-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA);1,2-Dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyl trimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DO EPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'' '-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-hexyldecanoate);ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide;β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol;BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl) (oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-diamino Leoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9''''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl) Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate);Further provided herein is a composition, wherein the nanoparticle is PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition, wherein each nanoparticle further comprises a hydrophobic core. Further provided herein is a composition, wherein the hydrophobic core comprises an oil. Further provided herein is a composition, wherein the oil is in a liquid phase. Further provided herein is a composition in which the oil comprises alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacyl glycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition in which the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerin myristate. Further provided herein is a composition in which each nanoparticle comprises a cationic lipid and an oil. Further provided herein is a composition in which each nanoparticle further comprises a surfactant. The surfactant is a polysorbate, a phosphorus-terminated surfactant, a carboxylate-terminated surfactant, a sulfate-terminated surfactant, an amine-terminated surfactant, trioctylphosphine oxide (TOPO), or a distearate; Further provided herein is a composition in which the glycerol phosphatidic acid (DSPA) is lyophilized. ...

[0165] Choline; FTT5, hexa(octan-3-yl) 9,9',9'',9''',9'''',9'''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanoate; Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl) Further provided herein is a composition in which the lipid carrier is selected from the group consisting of tetrakis(ethane-2,1-diyl)(9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate; PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition in which each lipid carrier further comprises a hydrophobic core. Further provided herein is a composition in which the hydrophobic core comprises an oil. Further provided herein is a composition in which the oil is in a liquid phase. Further provided herein is a composition in which the oil comprises alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition in which the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerin myristate. Further provided herein is a composition in which each lipid carrier comprises a cationic lipid and an oil. Further provided herein is a composition in which the hydrophilic surfactant is a polysorbate.Further provided herein is a composition, wherein the hydrophobic surfactant is a phosphorus-terminated surfactant, a carboxylate-terminated surfactant, a sulfate-terminated surfactant, an amine-terminated surfactant, trioctylphosphine oxide (TOPO), or distearyl phosphatidic acid (DSPA). Further provided herein is a composition, wherein the composition is lyophilized. Further provided herein is a composition, wherein the composition is in the form of a liquid, semi-liquid, solution, spray, or powder. Further provided herein is a composition, wherein the composition is formulated as a suspension. Further provided herein is a composition, comprising a plurality of nucleic acids, wherein the plurality of nucleic acids comprises an antibody or a functional fragment thereof; and a plurality of nucleic acids that separately encode an RNA polymerase complex region; and a plurality of nanoparticles, each nanoparticle comprising a cationic surface, and at least one nucleic acid is complexed with the cationic surface.

[0166] (Octan-3-yl) 9,9',9'',9''',9'''',9'''''-((((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanoate; Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis Further provided herein is a composition in which the lipid carrier is (azanetriyl))tetrakis(ethane-2,1-diyl)(9Z,9'Z,9''Z,9'''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate; PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a composition in which each lipid carrier further comprises a hydrophobic core. Further provided herein is a composition in which the hydrophobic core comprises an oil. Further provided herein is a composition in which the oil is in a liquid phase. Further provided herein is a composition in which the oil comprises alpha-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a composition in which the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglycerin myristate. Further provided herein is a composition in which each lipid carrier comprises a cationic lipid and an oil. Further provided herein is a composition in which the hydrophilic surfactant is a polysorbate.Further provided herein is a composition, wherein the hydrophobic surfactant is a phosphorus-terminated surfactant, a carboxylate-terminated surfactant, a sulfate-terminated surfactant, an amine-terminated surfactant, trioctylphosphine oxide (TOPO), or distearyl phosphatidic acid (DSPA). Further provided herein is a composition, wherein the composition is lyophilized. Further provided herein is a composition, wherein the composition is in the form of a liquid, semi-liquid, solution, spray, or powder. Further provided herein is a composition, wherein the composition is formulated as a suspension. Further provided herein is a composition, comprising a plurality of nucleic acids, wherein the plurality of nucleic acids comprises an antibody or a functional fragment thereof; and a plurality of nucleic acids that separately encode an RNA polymerase complex region; and a plurality of nanoparticles, each nanoparticle comprising a cationic surface, and at least one nucleic acid is complexed with the cationic surface.

[0167] Provided herein is a pharmaceutical composition comprising any one of the compositions provided herein; and a pharma- ceutically acceptable excipient.

[0168] Provided herein is a method for modulating an immune response, the method comprising administering a composition provided herein or a pharmaceutical composition provided herein to a subject having cancer.

[0169] Provided herein is a method for modulating an immune response, comprising administering a composition to a subject having cancer, the composition comprising at least one nucleic acid, the at least one nucleic acid comprising a sequence encoding a number of cancer-associated proteins, where prior to the administering step, the number of cancer-associated proteins comprises at least one nucleic acid that has an increased presence compared to non-cancer cells of the subject or comprises a sequence modification compared to non-cancer cells of the subject; and a number of nanoparticles, each nanoparticle comprising a cationic surface, and at least one nucleic acid complexed to the cationic surface. Provided herein is a method further comprising screening the cancer cells of the subject for a protein or nucleic acid that has an increased presence or sequence modification compared to non-cancer cells. Provided herein is a method further wherein the subject has a solid tumor or a blood cancer. Provided herein is a method further wherein the solid tumor is a carcinoma, melanoma, or sarcoma. Provided herein is a method further wherein the blood cancer is a lymphoma or leukemia. Provided herein is a method further wherein the subject has a metastatic cancer. Further provided herein is a method, wherein at least one cancer-associated protein of the plurality of cancer-associated proteins is a protein expressed by melanoma cells. The plurality of cancer-associated proteins is selected from the group consisting of (i) epidermal growth factor receptor (EGFR); (ii) vascular endothelial growth factor (VEGF); (iii) Wilms' tumor 1 (WT1); (iv) preferentially expressed antigen in melanoma (PRAME); (v) PR1; (vi) proteinase 3; (vii) elastase; (viii) cathepsin G; (ix) survivin; (x) New York Esophagus 1 (NY-Eso-1);(xi) melanoma associated antigen (MAGE);(xii) tyrosinase;(xiii) glycoprotein 100 (gp100);(xiv) carcinoembryonic antigen (CEA);(xv) mucin;(xvi) fibroblast growth factor (FGF);(xvii) programmed cell death protein (PD-1);(xviii) metastatic tumor antigen (MTA);(xix) human epidermal growth factor receptor 2 (Her2);(xx) mammaglobin A (SCGB2A2);(xxi) alpha-lactalbumin (LALBA);(xxii) cyclin D1 (CCND1); (xxiii) folate receptor 1 (FOLR1); (xxiv) telomerase (TERT); (xxv) RecQ protein-like (DNA helicase Q1-like) (RECQL); (xxvi) leptin receptor (LEPR); (xxvii) ERBB receptor feedback inhibitor 1 (ERRFI1); (xxviii) lysosomal protein transmembrane 4 alpha (LAPTM4A); (xxix) Kirsten rat sarcoma virus (K-Ras); (xxx) S100 proteins; (xxxi) cluster of differentiation (CD) family proteins; (xxxii) alpha fetoprotein (AFP); (xxxiii) epithelial tumor antigen (ETA); (xxxiv) tumor protein p 53;(xxxv) ephrin receptor;(xxxvi) transferrin receptor;(xxxvii) neoglycoprotein;(xxxviii) tumor necrosis factor (TNF)-alpha (α) receptor;(xxxvix) human papillomavirus-E6;(xl) human papillomavirus-E7;(xli) cytokeratin;(xlii) beta-catenin;(xliii) carboxypeptidase M;(xliv) EP4 receptor;(xlv) human milk fat glomerular antigen;(xlvi) tumor necrosis factor (TNF)-beta (β) receptor;(xlvii) B7-1 protein;(xlviii) B7-2 protein;(xlix) TNF receptor-associated factor 2;(l) melanoma-associated antigen recognized by T cells 1 (MART-1);and functional fragments thereof. Further provided herein is a method in which the MAGE is MAGE-A1, MAGE-A3, MART-1 / Melan-A, MAGE-A, MAGE-B, MAGE-C, MAGE-A1, MAGE-A2, MAGE-A3, MAGE-A4, MAGE-A5, MAGE-A6, MAGE-A7, MAGE-A8, MAGE-A9, MAGE-A10, MAGE-A11, or MAGE-A12. Further provided herein is a method in which the cluster of differentiation family protein is CD5, CD19, CD20, CD22, CD23, CD25, CD27, CD30, CD33, CD36, CD46, CD52, CD79a, CD79b, CD123, or CD317. Further provided herein is a method, wherein the plurality of nucleic acids comprises a sequence encoding any one of SEQ ID NOs: 3-7, 72, 74, 77, 78, 87. Further provided herein is a method, wherein the sequence comprises one or more of SEQ ID NOs: 1, 2, 71, 75, 76, 80-86, 88, 89. Further provided herein is a method, wherein at least one nucleic acid further comprises an RNA polymerase complex region. Further provided herein is a method, wherein the RNA polymerase complex region is downstream of a subgenomic promoter from an alphavirus. Further provided herein is a method, wherein the RNA polymerase complex region encodes an RNA-dependent RNA polymerase. Further provided herein is a method, wherein the RNA-dependent RNA polymerase is Venezuelan Equine Encephalitis Virus (VEEV) RNA polymerase. Further provided herein is a method, wherein the cationic surface comprises a cationic lipid. The cationic lipids were 1,2-dioleoyloxy-3(trimethylammonium)propane (DOTAP), 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol (DC cholesterol), dimethyldioctadecylammonium (DDA);1,2-Dimyristoyl 3-trimethylammonium propane (DMTAP), dipalmitoyl (C16:0) trimethylammonium propane (DPTAP), distearoyl trimethylammonium propane (DSTAP), N-[1-(2,3-dioleyloxy)propyl]N,N,N trimethylammonium, chloride (DOTMA), N,N-dioleoyl-N,N-dimethylammonium chloride (DODAC), 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (DO EPC), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dilinoleyloxy-3-dimethylaminopropane (DLinDMA), 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol) (C12-200), 306Oi10, tetrakis(8-methylnonyl)3,3',3'',3'' '-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetrapropionate, 9A1P9, decyl(2-(dioctylammonio)ethyl)phosphate; A2-Iso5-2DC18, ethyl 5,5-di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidin-1-yl)propyl)-2,5-dihydro-1H-imidazole-2-carboxylate; ALC-0315, ((4-hydroxybutyl)azanediyl)bis(hexane-6, 1-diyl)bis(2-hexyldecanoate);ALC-0159, 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide;β-sitosterol, (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol;BAME-O16B, bis(2-(dodecyldisulfanyl)ethyl) 3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl) dipropionate; BHEM-cholesterol, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl) (oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide; cKK-E12, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione; DC-cholesterol, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol; DLin-MC3-DMA, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate; DOPE, 1,2-diamino Leoyl-sn-glycero-3-phosphoethanolamine; DOSPA, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate; DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; ePC, ethylphosphatidylcholine; FTT5, hexa(octan-3-yl)9,9',9'',9''',9'''',9''''''-(((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl) Lipid H (SM-102), heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate; OF-Deg-Lin, (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetrakis(ethane-2,1-diyl) (9Z,9'Z,9''Z,9''Z,12Z,12'Z,12''Z,12''Z)-tetrakis(octadeca-9,12-dienoate);Further provided herein is a method, wherein the nanoparticle is PEG2000-DMG, (R)-2,3-bis(myristoyloxy)propyl-1-(methoxypoly(ethylene glycol)2000)carbamate; TT3, or N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide. Further provided herein is a method, wherein each nanoparticle further comprises a hydrophobic core. Further provided herein is a method, wherein the hydrophobic core comprises an oil. Further provided herein is a method, wherein the oil is in a liquid phase. Further provided herein is a method, wherein the oil comprises α-tocopherol, coconut oil, grape seed oil, lauroyl polyoxyl glyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soy lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E. Further provided herein is a method, wherein the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglyceride, triglyceride ester, or triglyceryl myristate. Further provided herein is a method, wherein each nanoparticle comprises a cationic lipid and an oil. Further provided herein is a method, wherein each nanoparticle further comprises a surfactant. Further provided herein is a method, wherein the surfactant is a polysorbate, a phosphorus-terminated surfactant, a carboxylate-terminated surfactant, a sulfate-terminated surfactant, an amine-terminated surfactant, trioctylphosphine oxide (TOPO), or distearylphosphatidic acid (DSPA). The administration is local or systemic; Further provided herein is a method in which the composition is administered to a subject. Further provided herein is a method in which the administration is via intratumoral, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, or intrathecal routes. Further provided herein is a method in which the composition is administered with a cancer therapeutic antibody or a composition provided herein. Further provided herein is a method in which the administration results in a reduction in tumor size or a reduction in tumor volume in the subject. Further provided herein is a method in which the administration results in a reduction in cancer recurrence. Further provided herein is a method in which the administration results in a reduction in tumor metastasis.

[0170] Provided herein is a method for prophylactically immunizing a subject against cancer, comprising administering to the subject a composition provided herein or a pharmaceutical composition provided herein, thereby immunizing the subject against cancer. Provided herein is a method in which the administration is via intratumoral, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, or intrathecal route. Provided herein is a method in which the subject is at risk of developing skin cancer. Provided herein is a method in which the skin cancer is basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma.

[0171] Provided herein is a method for treating cancer in a subject, comprising administering a composition provided herein or a pharmaceutical composition provided herein to the subject, thereby treating the cancer in the subject. Further provided herein is a method, wherein the administration is via intratumoral, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, or intrathecal route. Further provided herein is a method, wherein the composition is administered together with a cancer therapeutic antibody or a composition provided herein. Further provided herein is a method, wherein the subject has a solid tumor or a blood cancer. Further provided herein is a method, wherein the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a method, wherein the blood cancer is a lymphoma or leukemia. Further provided herein is a method, wherein the subject has a skin cancer. Further provided herein is a composition, wherein the skin cancer is a basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a method, wherein the subject has lung cancer. Further provided herein is a method, wherein the lung cancer is non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC). Further provided herein is a composition, wherein the NSCLC is adenocarcinoma, squamous cell carcinoma, large cell carcinoma, adenosquamous carcinoma, or sarcomatoid carcinoma. Further provided herein is a method, wherein the subject has pancreatic cancer. Further provided herein is a method, wherein the pancreatic cancer is pancreatic adenocarcinoma, pancreatic exocrine carcinoma, pancreatic neuroendocrine carcinoma, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. Further provided herein is a method, wherein the subject has metastatic cancer. Further provided herein is a method, wherein the administration results in a reduction in tumor size or a reduction in tumor volume in the subject. Further provided herein is a method, wherein the administration results in a reduction in cancer recurrence. Further provided herein is a method, wherein the administration results in a reduction in metastasis of tumors.

[0172] Provided herein is a method for personalized treatment of cancer in a subject, comprising: (a) receiving the results of an assay indicating that the subject has a tumor, the tumor comprising a cancer associated protein; (b) administering to the subject a composition as provided herein, the composition comprising at least one nucleic acid encoding the cancer associated protein in (a), thereby treating the cancer in the subject. Further provided herein is a method in which the administration is via intratumoral, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, or intrathecal routes. Further provided herein is a method in which the composition is administered together with a cancer therapeutic antibody or a composition as provided herein. Further provided herein is a method in which the subject has a solid tumor or a blood cancer. Further provided herein is a method in which the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a method in which the blood cancer is a lymphoma or leukemia. Further provided herein is a method in which the subject has a skin cancer. Further provided herein is a composition, wherein the skin cancer is basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a method, wherein the subject has lung cancer. Further provided herein is a method, wherein the lung cancer is non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC). Further provided herein is a composition, wherein the NSCLC is adenocarcinoma, squamous cell carcinoma, large cell carcinoma, adenosquamous carcinoma, or sarcomatoid carcinoma. Further provided herein is a method, wherein the subject has pancreatic cancer. Further provided herein is a method, wherein the pancreatic cancer is pancreatic adenocarcinoma, pancreatic exocrine carcinoma, pancreatic neuroendocrine carcinoma, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. Further provided herein is a method, wherein the subject has metastatic cancer. Further provided herein is a method, wherein the administration results in a reduction in tumor size or a reduction in tumor volume in the subject. Further provided herein is a method, wherein the administration results in a reduction in cancer recurrence. Further provided herein is a method, wherein the administration results in a reduction in metastasis of a tumor.

[0173] Provided herein is a method for personalized treatment of cancer in a subject, comprising: (a) receiving the results of an assay indicating that the subject has a tumor, the tumor comprising a cancer associated protein; (b) administering to the subject a composition provided herein, the composition comprising at least one nucleic acid encoding an antibody that specifically binds to the cancer associated protein in (a), thereby treating the cancer in the subject. Further provided herein is a method in which the administration is via intratumoral, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, or intrathecal routes. Further provided herein is a method in which the composition is administered with at least one additional cancer therapeutic agent. Further provided herein is a method in which the subject has a solid tumor or a blood cancer. Further provided herein is a method in which the solid tumor is a carcinoma, melanoma, or sarcoma. Further provided herein is a method in which the blood cancer is a lymphoma or leukemia. Further provided herein is a method in which the subject has a skin cancer. Further provided herein is a composition, wherein the skin cancer is basal cell carcinoma, melanoma, Merkel cell carcinoma, squamous cell carcinoma, cutaneous lymphoma, Kaposi's sarcoma, or skin adnexal carcinoma. Further provided herein is a method, wherein the subject has lung cancer. Further provided herein is a method, wherein the lung cancer is non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC). Further provided herein is a composition, wherein the NSCLC is adenocarcinoma, squamous cell carcinoma, large cell carcinoma, adenosquamous carcinoma, or sarcomatoid carcinoma. Further provided herein is a method, wherein the subject has pancreatic cancer. Further provided herein is a method, wherein the pancreatic cancer is pancreatic adenocarcinoma, pancreatic exocrine carcinoma, pancreatic neuroendocrine carcinoma, pancreatic islet cell carcinoma, or pancreatic endocrine carcinoma. Further provided herein is a method, wherein the subject has metastatic cancer. Further provided herein is a method, wherein the administration results in a reduction in tumor size or a reduction in tumor volume in the subject. Further provided herein is a method, wherein the administration results in a reduction in cancer recurrence. Further provided herein is a method, wherein the administration results in a reduction in metastasis of a tumor.

[0174] The following examples are presented to more clearly illustrate to those skilled in the art the principles and practice of the embodiments disclosed herein, and should not be construed as limiting the scope of any claimed embodiments. Unless otherwise stated, all parts and percentages are by weight. EXAMPLES

[0175] Example 1 Techniques and materials for the production of lipid nanoparticles The following materials were used in the manufacture of lipid-inorganic nanoparticles (i.e., lipid nanoparticles): The compositions, kits and methods described herein are not limited to the techniques or materials described herein.

[0176] Iron oxide nanoparticles of various mean diameters (5, 10, 15, 20, 25 and 30 nm) at 25 mg Fe / ml in chloroform were purchased from Ocean Nanotech (San Diego, CA, USA). Squalene and SPAN® 60 (sorbitan monostearate) were purchased from Millipore Sigma. TWEEN® 80 (polyethylene glycol sorbitan monooleate) and sodium citrate dihydrate were purchased from Fisher Chemical. The cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane chloride salt (DOTAP chloride) was purchased from Corden Pharma. Ultrapure water (resistivity of 18.2 megaohm-centimeter (MOhm-cm)) was obtained from a Milli-Q water purification system (Millipore Sigma).

[0177] Lipid carriers include squalene, sorbitan monostearate (e.g., SPAN® 60), polysorbate 80 (e.g., TWEEN® 80), DOTAP chloride, iron oxide nanoparticles, and sodium citrate dihydrate. Typically, chloroform was added to iron oxide nanoparticles having a number-weight average diameter of 5 nm. The chloroform was allowed to evaporate in a fume hood, leaving a dry coating of iron oxide nanoparticles. SPAN® 60, squalene, and DOTAP chloride were added to the iron oxide nanoparticles to prepare the "oil" phase.

[0178] The oil phase was sonicated for 30 minutes in a water bath preheated to 60° C. Separately, in a 1 liter glass bottle, the “water” phase was prepared by adding TWEEN® 80 to a sodium citrate dihydrate solution prepared with Milli-Q water.

[0179] The aqueous phase was stirred for 30 minutes to completely dissolve the TWEEN® 80. After complete dissolution of the TWEEN® 80, the aqueous phase was transferred to a beaker and incubated in a water bath preheated to 60° C. The preheated aqueous phase was added to the heated oil phase.

[0180] The mixture was immediately emulsified using a VWR® 200 homogenizer (VWR International) until a homogenous colloid with a milky appearance was produced. The colloid was then processed by passing the fluid through the Y-shaped interaction chamber of an LM10 microfluidizer at 20,000 psi.

[0181] Fluid was passed until the z-average hydrodynamic diameter, as measured by dynamic light scattering (Malvern Zetasizer Nano S), was 59 nm with a polydispersity index of 0.2.

[0182] The microfluidized lipid carrier samples were finally filtered through a 200 nm pore size polyethersulfone (PES) syringe filter. Example 2 Exemplary Techniques and Materials for Producing Lipid Nanoparticles

[0183] The following materials were used in the manufacture of lipid-inorganic nanoparticles (i.e., lipid nanoparticles): The compositions, kits and methods described herein are not limited to the techniques or materials described herein.

[0184] Iron oxide nanoparticles of various mean diameters (5, 10, 15, 20, 25 and 30 nm) at 25 mg Fe / ml in chloroform were purchased from Ocean Nanotech (San Diego, CA). Squalene and SPAN® 60 (sorbitan monostearate) were purchased from Millipore Sigma. TWEEN® 80 (polyethylene glycol sorbitan monooleate) and sodium citrate dihydrate were purchased from Fisher Chemical. The cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane chloride salt (DOTAP chloride) was purchased from Corden Pharma. Ultrapure water (resistivity of 18.2 MOhm-cm) was obtained from a Milli-Q water purification system (Millipore Sigma).

[0185] A lipid carrier was prepared containing 37.5 mg / ml squalene, 37 mg / ml SPAN® 60, 37 mg / ml TWEEN® 80, 30 mg / ml DOTAP chloride, 0.1 mg / ml 10 nm iron oxide nanoparticles and 10 mM sodium citrate dihydrate.

[0186] The lipid carriers were prepared using the following procedure: In a 200 ml beaker, 25 mg Fe / ml of iron oxide nanoparticles with a number-weight average diameter of 10 nm in 0.4 ml of chloroform were added.

[0187] The chloroform was allowed to evaporate in a fume hood leaving a dry coating of iron oxide nanoparticles. An "oil" phase was prepared by adding 3.7 grams of SPAN® 60, 3.75 grams of squalene, and 3 grams of DOTAP chloride to the iron oxide nanoparticles.

[0188] The oil phase was sonicated for 30 minutes in a water bath preheated to 60° C. Separately, in a 1 liter glass bottle, a "water" phase was prepared by adding 39 grams of TWEEN® 80 to 1,000 ml of a 10 mM sodium citrate dihydrate solution prepared with Milli-Q water.

[0189] The aqueous phase was stirred for 30 minutes to allow complete dissolution of TWEEN® 80. After complete dissolution of TWEEN® 80, 96 ml of the aqueous phase was transferred to a 200 ml beaker and incubated in a water bath preheated to 60° C. To the heated oil phase, 96 ml of preheated aqueous phase was added. The mixture was immediately emulsified using a VWR® 200 homogenizer (VWR International) until a homogenous colloid with a milky appearance was produced. The colloid was then processed by passing the fluid through a Y-shaped interaction chamber of an LM10 microfluidizer at 20,000 psi. The fluid was passed until the z-average hydrodynamic diameter was 54 nm with a polydispersity index of 0.2 as measured by dynamic light scattering (Malvern Zetasizer Nano S). The microfluidized lipid carrier sample was finally filtered through a polyethersulfone (PES) syringe filter with a pore size of 200 nm. Example 3 Exemplary Techniques and Materials for Producing the Nanoparticles Described Herein

[0190] A lipid carrier was prepared containing 37.5 mg / ml squalene, 37 mg / ml SPAN® 60, 37 mg / ml TWEEN® 80, 30 mg / ml DOTAP chloride, 0.2 mg / ml 15 nm iron oxide nanoparticles, and 10 M sodium citrate dihydrate. The lipid carrier of Example 9 was manufactured using the following procedure.

[0191] In a 200 ml beaker, 25 mg Fe / ml of iron oxide nanoparticles in 0.8 ml of chloroform, with a number-weight average diameter of 15 nm, were added. The chloroform was allowed to evaporate in a fume hood, leaving a dry coating of iron oxide nanoparticles. To the iron oxide nanoparticles were added 3.7 grams of SPAN® 60, 3.75 grams of squalene, and 3 grams of DOTAP chloride to prepare the "oil" phase.

[0192] The oil phase was sonicated for 30 minutes in a water bath preheated to 60° C. Separately, in a 1 liter glass bottle, the “water” phase was prepared by adding 39 grams of TWEEN® 80 to 1,000 ml of 10 mM sodium citrate dihydrate solution prepared with Milli-Q water. The water phase was stirred for 30 minutes to completely dissolve the TWEEN® 80.

[0193] After complete dissolution of the TWEEN® 80, 96 ml of the aqueous phase was transferred to a 200 ml beaker and incubated in a water bath preheated to 60° C. To the heated oil phase, 96 ml of preheated aqueous phase was added. The mixture was immediately emulsified using a VWR® 200 homogenizer (VWR International) until a homogenous colloid with a milky appearance was produced. The colloid was then processed by passing the fluid through the Y-shaped interaction chamber of an LM10 microfluidizer at 20,000 psi.

[0194] Fluid was passed until the z-average hydrodynamic diameter was 52 nm with a polydispersity index of 0.2 as measured by dynamic light scattering (Malvern Zetasizer Nano S). The microfluidized lipid carrier sample was finally filtered through a 200 nm pore size polyethersulfone (PES) syringe filter. Example 4 Compositions prepared using the construct of SEQ ID NO: 75 or SEQ ID NO: 76 for use as cancer vaccines

[0195] Lipid carrier-RNA complexes are prepared and aliquoted for lyophilization. Samples are lyophilized and then collected and selected for reconstitution. All lyophilized cakes are then reconstituted in 0.7ml of milliQ® water. Table 4 discloses exemplary materials used in the preparation of lipid carrier-RNA complexes. [Table 4]

[0196] Exemplary conditions for lyophilization are shown below in Tables 5-7. [Table 5] [Table 6] [Table 7]

[0197] Diluent preparation: Diluents containing sugar and citrate were prepared as outlined in Table 8. Each sugar was weighed into a 50ml RNase-free conical tube. Approximately 35-40ml of nuclease-free water was added to dissolve the sugar, using slight heating and sonication if necessary. Pipette in 0.5ml of 1M Na-citrate, pH=6 solution. After all sugar was dissolved and the solution was clear, QS in the conical tube with nuclease-free water to the 50ml mark. Diluents were filtered through a 0.22μm STERIFLIP® and aseptically capped to maintain sterility. [Table 8]

[0198] Preparation of precomplex formulations: Lipid carrier "DS" is 30 mg DOTAP / ml bulk solution and refers to the Fe-lipid carrier formulation, the preparation of which is described in Example 2. The 10x lipid carrier "DS" (30mg DOTAP / ml) was diluted in the respective diluent to make 3mg DOTAP / ml lipid carrier "DP" except for the 5x 50% sucrose composition lipid carrier which was diluted to make 2x 6mg DOTAP / ml lipid carrier "DP". The target RNA concentration in the liquid formulation was 50ng / μl and complexed with the lipid carrier with an N:P of 15. This simulates an RNA dose of 25μg per vial. Table 9 discloses the preparation of the pre-complexed lipid carrier complex. Unused lipid carriers were stored at 2-8 degrees Celsius. [Table 9]

[0199] Table 10 discloses the preparation of pre-complexed nanostructured lipid carrier (NLC) complexes. NLC was used as a control. Unused NLC was stored at 2-8 degrees Celsius for fresh complex control. [Table 10-1]

[0200] The preparation of RNA pre-complex is disclosed in Table 11. RNA stocks were prepared, divided into approximately 7.5 ml or 0.63 ml for 50% sucrose per aliquot and stored at -80 degrees Celsius. [Table 11-1]

[0201] The preparation of lipid carrier-RNA complexes is disclosed in Table 11. RNA stocks were prepared. The volume of RNA diluted per complexation per lyo cycle was (+5%) and the lipid carrier diluted was (+5%).

[0202] Lipid carrier+RNA or NLC+RNA complexes were prepared by mixing 1:1 volume of each of the dilution formulations listed in Tables 10 and 11 above with the corresponding "Composition ID" diluted RNA disclosed in Table 12. The complexes were equilibrated at room temperature for 30 minutes before being subjected to a lyophilization cycle or long-term storage conditions. [Table 12-1]

[0203] The RNA is a construct having a nucleic acid sequence as shown in either SEQ ID NO:75 or SEQ ID NO:76, each of which includes a VEEV RNA sequence backbone and an RNA sequence encoding a cancer-associated protein antigen. Example 5 Macrophage immune response.

[0204] Various formulations of lipid carriers and repRNA were prepared and analyzed to assay the innate immune response of lipid carriers in macrophages. Stimulation of protein expression and TNF production in THP-1 macrophages was studied.

[0205] First, THP-1 monocytes were differentiated into macrophages using phorbol 12-myristate 13-acetate (PMA).The cells were then transfected with various formulations carrying Nano Luciferase-encoding replicon RNA (SEQ ID NO: 71).The cell culture medium was then evaluated for NanoLuc and TNF expression.

[0206] The formulations used in this assay and their characteristics such as particle size and PDI are listed in Table 13. The concentration of repRNA-encoded NanoLuc was 909 ng / μl and maintained at −80 degrees Celsius. MIGLYOL® 812 N, a triglyceride ester of saturated coconut / palm kernel oil-derived caprylic and capric fatty acids and vegetable-derived glycerol, was used in this assay. [Table 13] Example 6 Fe-lipid carrier formulation-NP-1 (prepared at 100 ml scale).

[0207] The Fe-lipid carrier formulation contained 37.5 mg / ml squalene (SEPPIC), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 0.2 mg Fe / ml 12 nm oleic acid coated iron oxide nanoparticles (Imagion Biosystems, San Diego, CA, USA) and 10 mM sodium citrate dihydrate (Fisher Chemical). 20 mg Fe / ml 12 nm diameter oleic acid coated iron oxide nanoparticles in 1 ml chloroform (Imagion Biosystems, lot number 95-127) were washed three times by magnetic separation in a 4:1 acetone:chloroform (v / v) solvent mixture. After the third wash, the volatile solvents (acetone and chloroform) were allowed to evaporate completely in a fume hood, leaving a coating of dry iron oxide oleate nanoparticles. To this iron oxide coating, 3.75 grams of squalene, 3.7 grams of SPAN® 60, and 3 grams of DOTAP were added to produce an oil phase. The oil phase was sonicated in a 65° Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65° Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase.The mixture was emulsified using a VWR® 200 homogenizer (VWR International, Radnor, PA, USA) and the resulting crude emulsion was processed by passing it through an M110P microfluidizer (Microfluidics, Westwood, MA, USA) equipped with a F12Y 75 μm diamond interaction chamber and an auxiliary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter, measured by dynamic light scattering (Malvern Zetasizer Nano S), reached 40-80 nm with a polydispersity index (PDI) of 0.1-0.25 (Figure 6). The microfluidized NP-1 formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. Iron concentrations were determined by inductively coupled plasma-optical emission spectroscopy (ICP-OES). DOTAP and squalene concentrations were measured by reversed-phase high performance liquid chromatography (RP-HPLC). Example 7 High Fe-lipid carrier formulation NP-2 (prepared at 100 ml scale).

[0208] The high Fe-lipid carrier formulation contained 37.5 mg / ml squalene (SEPPIC), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 1 mg Fe / ml 15 nm oleic acid coated iron oxide nanoparticles (Imagion Biosystems) and 10 mM sodium citrate dihydrate (Fisher Chemical). 20 mg Fe / ml 15 nm diameter oleic acid coated iron oxide nanoparticles in 5 ml chloroform (Imagion Biosystems, lot number 95-133) were washed three times by magnetic separation in a 4:1 acetone:chloroform (v / v) solvent mixture. After the third wash, the volatile solvents (acetone and chloroform) were allowed to evaporate completely in a fume hood leaving a coating of dry oleic acid iron oxide nanoparticles. To this iron oxide coating, 3.75 grams of squalene, 3.7 grams of SPAN® 60, and 3 grams of DOTAP were added to produce an oil phase. The oil phase was sonicated in a 65 degree Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65 degrees Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase. The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting coarse emulsion was processed by passing it through an M110P Microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and a secondary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter, as measured by dynamic light scattering (Malvern Zetasizer Nano S), reached 40-80 nm with a polydispersity index (PDI) of 0.1-0.3.The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. Iron concentrations were determined by ICP-OES. DOTAP and squalene concentrations were measured by RP-HPLC. Example 8 Fe-lipid carrier miglyol formulation NP-3 (prepared at 100 ml scale).

[0209] The Fe-lipid carrier miglyol formulation contained 37.5 mg / ml Miglyol 812 N (IOI Oleo GmbH), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 0.2 mg Fe / ml 15 nm oleic acid coated iron oxide nanoparticles (Imagion Biosystems) and 10 mM sodium citrate dihydrate (Fisher Chemical). 20 mg Fe / ml 15 nm diameter oleic acid coated iron oxide nanoparticles in 1 ml chloroform (Imagion Biosystems, lot number 95-127) were washed three times by magnetic separation in a 4:1 acetone:chloroform (v / v) solvent mixture. After the third wash, the volatile solvents (acetone and chloroform) were allowed to evaporate completely in a fume hood, leaving a coating of dry iron oxide oleate nanoparticles. To this iron oxide coating, 3.75 grams of squalene, 3.7 grams of SPAN® 60, and 3 grams of DOTAP were added to produce an oil phase. The oil phase was sonicated in a 65° Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65° Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase. The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting coarse emulsion was processed by passing it through an M110P Microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and a secondary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter, as measured by dynamic light scattering (Malvern Zetasizer Nano S), reached 40-80 nm with a polydispersity index (PDI) of 0.1-0.3.The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. Iron concentrations were determined by ICP-OES. DOTAP concentrations were measured by RP-HPLC. Example 9 High Fe-lipid carrier Miglyol formulation NP-4 (prepared at 100 ml scale).

[0210] The high Fe-lipid carrier Miglyol formulation contains 37.5 mg / ml Miglyol 812 N (IOI Oleo GmbH), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 1 mg / ml 15 nm oleic acid coated iron oxide nanoparticles (ImagionBio) and 10 mM sodium citrate dihydrate (Fisher Chemical). 20 mg Fe / ml 15 nm diameter oleic acid coated iron oxide nanoparticles in 5 ml chloroform (ImagionBio, lot number 95-127) were washed three times by magnetic separation in a 4:1 acetone:chloroform (v / v) solvent mixture. After the third wash, the volatile solvents (acetone and chloroform) were allowed to evaporate completely in a fume hood, leaving a coating of dry iron oxide oleate nanoparticles. To this iron oxide coating, 3.75 grams of squalene, 3.7 grams of SPAN® 60, and 3 grams of DOTAP were added to produce an oil phase. The oil phase was sonicated in a 65° Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65° Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase. The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting coarse emulsion was processed by passing it through an M110P Microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and a secondary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter, as measured by dynamic light scattering (Malvern Zetasizer Nano S), reached 40-80 nm with a polydispersity index (PDI) of 0.1-0.3.The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. Iron concentrations were determined by ICP-OES. DOTAP concentrations were measured by RP-HPLC. Example 10 Alum-lipid carrier formulation NP-5 (prepared at 100 ml scale).

[0211] The alum-lipid carrier formulation contained 37.5 mg / ml squalene (SEPPIC), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 1 mg Al / ml TOPO-coated Alhydrogel® (aluminum oxyhydroxide) particles (Croda) and 10 mM sodium citrate. 10 ml of Alhydrogel was washed three times in methanol by centrifugation at 1000 rpm for 20 minutes. After the third wash, the Alhydrogel was dispersed in 10 ml of methanol, and 1 ml of 250 mg / ml trioctylphosphine oxide (TOPO) was added to the dispersion and incubated overnight at 37° C. on an orbital shaker. Excess TOPO was removed by an additional methanol wash and then dispersed in 11 ml of methanol. The methanol was allowed to evaporate overnight in a fume hood leaving a dry layer of TOPO-Alhydrogel. To this dry TOPO-Alhydrogel layer, 3.75 grams of squalene, 3.7 grams of SPAN® 60, and 3 grams of DOTAP were added to produce an oil phase. The oil phase was sonicated in a 65° Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65° Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase.The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting crude emulsion was processed by passing it through an M110P microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and an auxiliary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter reached 40-80 nm with a polydispersity index (PDI) of 0.1-0.3 as measured by dynamic light scattering (Malvern Zetasizer Nano S). The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. Aluminum concentrations were determined by ICP-OES. DOTAP and squalene concentrations were measured by RP-HPLC. Example 11 Fe-lipid carrier Solanesol formulation NP-6 (prepared at 100 ml scale).

[0212] The Fe-lipid carrier solanesol formulation (NP-6) contains 37.5 mg / ml solanesol (Cayman chemicals), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 0.2 mg Fe / ml oleic acid coated iron oxide nanoparticles (ImagionBio) and 10 mM sodium citrate. 20 mg Fe / ml oleic acid coated iron oxide nanoparticles with a diameter of 15 nm in 1 ml chloroform (ImagionBio, lot number 95-133) were washed three times by magnetic separation in a 4:1 acetone:chloroform (v / v) solvent mixture. After the third wash, the volatile solvents (acetone and chloroform) were allowed to evaporate completely in a fume hood leaving a coating of dry oleic acid iron oxide nanoparticles. To this iron oxide coating, 3.75 grams of Solanesol, 3.7 grams of SPAN® 60, and 3 grams of DOTAP were added to produce an oil phase. The oil phase was sonicated in a 65 degree Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65 degrees Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase. The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting crude emulsion was processed by passing it through a M110P microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and an auxiliary H30Z-200 μm ceramic interaction chamber at 30,000 psi. The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. Iron concentrations were determined by ICP-OES. DOTAP and solanesol concentrations were measured by RP-HPLC. Example 12 NP-7 formulation (prepared at 100 ml scale).

[0213] The NP-7 formulation contains 37.5 mg / ml squalene (SEPPIC), 37 mg / ml SPAN® 60 (Millipore Sigma), 37 mg / ml TWEEN® 80 (Fisher Chemical), 30 mg / ml DOTAP chloride (LIPOID), 2.4 mg / ml Dynasan 114 (IOI Oleo GmbH) and 10 mM sodium citrate. To a 200 ml beaker, 3.75 grams of squalene, 3.7 grams of SPAN® 60, 0.24 grams of Dynasan 114 and 3 grams of DOTAP were added to generate an oil phase. The oil phase was sonicated in a 65 degree Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 19.5 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 92 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65 degrees Celsius for 30 minutes. The oil phase was mixed with 92 ml of the aqueous phase by adding the warm oil phase to the warm aqueous phase. The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting crude emulsion was processed by passing it through an M110P microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and an auxiliary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter reached 40-80 nm with a polydispersity index (PDI) of 0.1-0.3 as measured by dynamic light scattering (Malvern Zetasizer Nano S). The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. DOTAP and squalene concentrations were measured by RP-HPLC. Example 13 NP-8 formulation (prepared at 100 ml scale).

[0214] The NP-8 formulation contains 43 mg / ml squalene (SEPPIC), 5 mg / ml SPAN® 85 (Millipore Sigma), 5 mg / ml TWEEN® 80 (Fisher Chemical), 4 mg / ml DOTAP chloride (LIPOID), and 10 mM sodium citrate. In a 200 ml beaker, 4.3 grams of squalene, 0.5 grams of SPAN® 85, and 0.4 grams of DOTAP were added to generate an oil phase. The oil phase was sonicated in a 65 degrees Celsius water bath for 45 minutes. Separately, an aqueous phase was prepared by dissolving 2.6 grams of TWEEN® 80 in 500 ml of 10 mM sodium citrate buffer prepared in nuclease-free water. 95 ml of the aqueous phase was transferred to a separate glass bottle and heated to 65 degrees Celsius for 30 minutes. The oil phase was mixed with 95 ml of the water phase by adding the warm oil phase to the warm water phase. The mixture was emulsified using a VWR® 200 homogenizer (VWR International) and the resulting crude emulsion was processed by passing it through a M110P microfluidizer (Microfluidics) equipped with a F12Y 75 μm diamond interaction chamber and a supplementary H30Z-200 μm ceramic interaction chamber at 30,000 psi until the z-average hydrodynamic diameter reached 100 ± 10 nm with a polydispersity index (PDI) of 0.05-0.1 as measured by dynamic light scattering (Malvern Zetasizer Nano S). The microfluidized formulation was finally filtered through a 200 nm pore size polyethersulfone (PES) filter and stored at 2-8 degrees Celsius. DOTAP and squalene concentrations were measured by RP-HPLC. Example 14 Cell-based assays to assess lipid nanoparticle and protein production.

[0215] Treatment groups were prepared. Eight of these groups were NanoLuc repRNA groups, with 600ng dose per well prepared using Fe-lipid carrier, high Fe-lipid carrier, Fe-lipid carrier miglyol, high Fe-lipid carrier miglyol, alum-lipid carrier, Fe-lipid carrier solanesol (SLN), NLC, and CNE formulations. The untreated group had no NanoLuc. The various formulations were prepared by diluting NanoLuc repRNA to 8ng / μL in 2.2mL of RNAse-free water. The lipid carrier formulation and RNA master mix were complexed by adding 250μL of each diluted formulation to 250μL of diluted RNA and mixed by pipetting up and down.

[0216] Cell transfections were performed at 7 × 10 cells per well in 24-well plates. 5 The transfection was performed by seeding 100 THP-1s. 80 micromolar (μM) PMA was added to each well and incubated at 37 degrees Celsius. The following day, 1 hour before transfection, the PMA-containing medium was removed and replaced with complete RPMI (cRPMI) medium. Samples were then serially diluted in Opti-MEM™ (Thermo Fisher Scientific, Waltham, MA USA) to generate a 10-point 1.5-fold dilution series starting at 0.45 ng / μL. Culture medium was then removed from the plate by pipetting. 450 μL of Opti-MEM™ and 150 μL of complexed formulation were added to the plate in duplicate. Empty wells received 450 μL of Opti-MEM™ only. After 4 hours, samples were removed from the plate by pipetting and replaced with 500 μL of growth medium. The plates were then incubated overnight at 37 degrees C. Growth medium was collected the next day and stored at -80 degrees C. Downstream assays were performed and described below.

[0217] Luciferase assays were performed by first diluting Nano-Glo® Luciferase Assay Reagent 1:50 in buffer. 25 μL of supernatant was removed and mixed with 25 μL of Nano-Glo® Reagent in a 96-well plate. This was incubated at room temperature for 3 minutes. Luminescence was read using a luminometer.

[0218] An ELISA assay was then performed to assess TNF-alpha (α) protein levels in cell culture media using Human TNF-α DUOSET™ ELISA (R&D Systems) according to the manufacturer's protocol. A 96-well microplate was coated with anti-TNFα capture antibody. The plate was blocked, and then medium samples were added directly without dilution. After addition of biotinylated detection antibody SA-HRP and substrate, absorbance was read at 450 nm in a SPECTRAMAX® i3 (Molecular Devices) plate reader.

[0219] All studies in this example were performed in duplicate. The results from the duplicates are represented as the first experiment and the second experiment, respectively. The formulations containing lipid carriers and miglyol induced higher protein production from the replicon, as shown in the first experiment in Figure 2A and the second experiment in Figure 2B. A reduction in the innate immune response was detected, as measured by TNF-α secretion (Figures 3A-3B).

[0220] A correlation between enhanced protein production and low TNF-alpha stimulation was observed with the miglyol lipid carrier formulation, as shown in the first experiment in Figure 4A and the second experiment in Figure 4B. The solanesol lipid carrier formulation induced slightly lower protein production but higher TNFα production (Figures 4A-4B). The correlation data from the first assay shown in Figure 4A is derived from the data represented in Figures 2A and 3A. The correlation data from the second assay is shown in Figure 4B, which is derived from the data represented in Figures 2B and 3B. Example 15 Exemplary techniques and materials for producing the nanoparticles described herein.

[0221] In the mouse assay, C57BL / 6 mice were inoculated as described in Table 6 below, after which secreted embryonic alkaline phosphatase (SEAP) levels were measured in serum. A summary of the materials used in this example is provided in Table 14. [Table 14] [Table 15]

[0222] Seven different formulations were prepared and administered intramuscularly across seven treatment groups (Groups 1-7). DNA-SEAP or RNA-SEAP was diluted according to the volumes set out in Table 16 to prepare the formulations for Groups 1-7. [Table 16]

[0223] The concentrations of the diluted DNA or RNA before complexing with lipid carriers were as follows (measured by NanoDrop spec): groups 1, 4 and 5 contained approximately 820 μg / ml of DNA; groups 2 and 3 contained approximately 480 μg / ml of DNA; groups 6 and 7 contained approximately 43 μg / ml of RNA.

[0224] The formulations for Groups 1-7 were diluted in 100 mM citrate as shown in Table 17 below. [Table 17]

[0225] The above formulations were complexed by adding 250μl of diluted lipid carrier to 250μl of diluted DNA or RNA. The resulting complexed formulation was incubated on ice for at least 30 minutes. Table 18 shows the experimental schedule for the assay. [Table 18]

[0226] Mice were bled at regular intervals and serum was immediately prepared and stored at -80 degrees Celsius until analysis for SEAP activity.

[0227] To assess SEAP levels in serum, all serum samples were thawed at the same time and SEAP detection was performed. Figures 5A-5F illustrate SEAP levels in BALB / c mice intramuscularly injected with various replicates of lipid carrier-formulated DNA SEAP. Mice were bled at regular intervals and serum was prepared and stored until analysis by SEAP assay. Data are presented as mean and SE (n=5 per group).

[0228] As can be seen from Figures 5A-5F, the lipid carrier formulation supports more targeted protein production than delivery of DNA alone, especially after day 6 post-injection. In addition, the data show that the inclusion of miglyol enhances protein production from the RNA replicon more than lipid carrier formulations lacking miglyol. Example 16 Self-replicating mRNA construct.

[0229] A plasmid encoding the T7 promoter followed by the 5' and 3' UTRs and nonstructural genes of Venezuelan Equine Encephalitis Virus (VEEV) strain TC-83 was generated using standard DNA synthesis and cloning methods. The VEEV replicon mRNA backbone is shown in SEQ ID NO:71. (Example 17) Additional nanoparticle formulations.

[0230] Additional nanoparticle formulations are produced according to the following tables (Table 19 and Table 10): The mRNA contains sequences encoding the TRP-1 tumor-associated antigen with a VEEV replicon mRNA backbone (SEQ ID NO: 71). [Table 19] [Table 10-2] (Example 18) The TRP-1 replicon prevents B16F0 tumor growth.

[0231] The B16 subcutaneous melanoma mouse model was used in the assays provided herein. Upon subcutaneous injection, B16 animals form palpable tumors in approximately 5-10 days, which grow to approximately 1x1x1-cm tumors in approximately 14-21 days.

[0232] A lipid carrier and a TRP-1 RNA replicon were generated (SEQ ID NO: 76). The amino acid sequence of TRP-1 is SEQ ID NO: 78.

[0233] Female C57BL / 6 mice were immunized once or twice by intramuscular injection of repRNA-TRP1 at a dose of 0.2 mcg or 1 mcg. Table 11 provides the assay conditions used. [Table 11-2]

[0234] Blood was collected 2 weeks after completion of the immunization regimen, serum was prepared for each animal, and antibodies binding to TRP1 were assessed by ELISA. n=5 per group. Results are shown as tumor growth (FIGS. 7A-7E) and survival (FIG. 7F) for each individual animal. Immunization with a TRP1-expressing replicon limits the proliferation of B16 in a dose-dependent manner.

[0235] Next, the therapeutic efficacy of TRP-1 repRNA vaccination was examined at various doses and treatment regimens, as shown in Table 12. [Table 12-2]

[0236] Female C57BL / 6 mice were immunized by intramuscular injection of repRNA-TRP1 at a dose of 1 mcg or 5 mcg starting on day 3 (prepalpable) or day 7 (postpalpable) after tumor cell implantation. Immunizations were repeated weekly thereafter. n=5 per group. Results are shown as the mean tumor growth for each group (Figure 8) and for each individual animal (Figure 9). The results further confirm that immunization with a TRP1-expressing replicon limits B16 proliferation. (Example 19) The MAGE-A1 replicon induces antigen-specific T cells.

[0237] A lipid nanoparticle carrier and a MAGE-A1 RNA replicon were generated (SEQ ID NO: 75). The amino acid sequence of MAGE-A1 is SEQ ID NO: 77.

[0238] Female C57BL / 6 mice were immunized by intramuscular injection of repRNA-MAGE-A1 at a dose of 0.2 or 1 mcg. Immunizations were performed on days 0, 14 and 80 (three times), or on days 14 and 80 (two times), and then spleens were harvested on day 91 and single cell suspensions were prepared. Cells were then incubated with MAGE-A1 or HPV E6 (non-specific) peptide pools and subjected to flow cytometry. CD8 T cells expressing Tbet and producing IFN-gamma (γ) (Figures 10A-10B) and CD4 T cells producing IFNγ or IL-2 (Figures 10C-10D) are shown. Immunization with MAGE-expressing replicons induces antigen-specific T cells. (Example 20) TRP-1 and MAGE-A3 combination therapy reduces tumor volume.

[0239] The lipid nanoparticle carrier was combined with a MAGE-A3 RNA replicon and a TRP-1 RNA replicon. The amino acid sequence of TRP-1 is SEQ ID NO: 78. The amino acid sequence of MAGE-A3 is SEQ ID NO: 87.

[0240] The B16 subcutaneous melanoma mouse model was used in the assay provided herein by Example 17. The therapeutic efficacy of TRP-1 and MAGE-A3 repRNA vaccination was examined at various doses and treatment regimens as shown in Table 12.

[0241] Tumor volumes were quantified for control, NP-1+TRP-1+MAGE-A3 prepalpable animals, and NP-1+TRP-1+MAGE-A3 (FIG. 11A). The probability of survival was increased in animals treated with lipid carriers and repRNA encoding TRP-1 and MAGE-A3 compared to untreated animals (FIG. 11B). Example 21 Self-replicating RNA prostein constructs.

[0242] The lipid nanoparticle carrier was combined with a prostein-encoding RNA replicon. The RNA encoding the prostein protein encodes the amino acid sequence of SEQ ID NO: 90. The VEEV replicon mRNA backbone sequence is shown in SEQ ID NO: 71. [ka] [ka] [ka] [ka] [ka] [ka] [ka]

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Claims

1. 1. A composition comprising:

1. A lipid nanoparticle, characterized as having a z-average diameter particle size measurement of about 20 nm to about 60 nm as measured using dynamic light scattering, wherein the lipid nanoparticle comprises: cationic lipids; a hydrophilic surfactant; and Hydrophobic surfactants lipid nanoparticles comprising: at least one nucleic acid, said at least one nucleic acid comprising: A sequence encoding an RNA-dependent polymerase; and At least one nucleic acid comprising a sequence encoding a transmembrane binding domain, an outer cell membrane contacting domain, and an inner cell membrane contacting domain of a prosteine ​​protein. A composition comprising:

2. The composition of claim 1 , wherein the prostein protein comprises an amino acid sequence that is at least 80% identical to SEQ ID NO:

90.

3. The composition of claim 1 , wherein the prostein protein comprises the amino acid sequence of SEQ ID NO:

90.

4. The composition of claim 1 , wherein the prostein protein is expressed by a solid cancer cell.

5. 5. The composition of claim 4, wherein the solid cancer cells comprise melanoma cancer cells, prostate cancer cells, colon cancer cells, ovarian cancer cells, breast cancer cells, or pancreatic cancer cells.

6. 10. The composition of claim 1, wherein the at least one nucleic acid is in complex with the lipid nanoparticle to form a nucleic acid-lipid nanoparticle complex.

7. 2. The composition of claim 1, wherein the RNA-dependent polymerase is Venezuelan equine encephalitis virus (VEEV) RNA polymerase.

8. The composition of claim 1 , wherein the sequence encoding the RNA-dependent polymerase comprises SEQ ID NO:

71.

9. The composition of claim 1 , wherein the lipid nanoparticle comprises a hydrophobic core.

10. The composition of claim 9, wherein the lipids present in the hydrophobic core are in a liquid phase at 25 degrees Celsius.

11. The composition of claim 9 , wherein the hydrophobic core comprises a liquid oil.

12. 12. The composition of claim 11, wherein the liquid oil is α-tocopherol, coconut oil, grape seed oil, lauroyl polyoxylglyceride, mineral oil, monoacylglycerol, palm kernel oil, olive oil, paraffin oil, peanut oil, propolis, squalene, squalane, soybean lecithin, soybean oil, sunflower oil, triglyceride, or vitamin E.

13. 13. The composition of claim 12, wherein the triglyceride is capric triglyceride, caprylic triglyceride, caprylic and capric triglycerides, triglyceride esters, or triglyceryl myristate.

14. The cationic lipid may be 1,2-dioleoyloxy-3(trimethylammonium)propane, 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol, dimethyldioctadecylammonium, 1,2-dimyristoyl-3-trimethylammoniumpropane, dipalmitoyl (C16:0)trimethylammoniumpropane, distearoyltrimethylammoniumpropane, N-[1-(2,3-dioleyloxy)propyl]N,N,N-trimethylammonium chloride, N,N-dioleoyl-N,N-dimethylammonium chloride, or the like. ammonium chloride, 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine, 1,2-dioleoyl-3-dimethylammonium-propane, 3β-[N-(N',N'-dimethylaminoethane)carbamoyl]cholesterol, 1,1'-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazin-1-yl)ethyl)azanediyl)bis(dodecan-2-ol), tetrakis(8-methylnonyl)3,3',3'',3'''-(((methylazanediyl) (4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate), 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide, (( ... 3S,8S,9S,10R,13R,14S,17R)-17-((2R,5R)-5-ethyl-6-methylheptan-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-ol, bis(2-(dodecyldisulfanyl)ethyl)3,3'-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexacosyl)azanediyl)dipropionate, 2-((((3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-((R)-6-methylheptan-2-yl)-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-3-yl)oxy)carbonyl)amino)-N,N-bis(2-hydroxyethyl)-N-methylethane-1-aminium bromide, 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2, 5-dione, 3β-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol, (6Z,9Z,28Z,31Z)-heptatriaconta-6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, 2,3-dioleyloxy-N-[2-(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate, 1 , 2-distearoyl-sn-glycero-3-phosphocholine, ethylphosphatidylcholine, hexa(octan-3-yl)9,9',9'',9''',9'''',9'''',9''''-((((benzene-1,3,5-tricarbonyl)iris(azanediyl))tris(propane-3,1-diyl))tris(azanetriyl))hexanonanoate, heptadecan-9-yl 8-((2-hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)o The composition of claim 1, wherein the tetrakis(octadeca-9,12-dienoate) is N1,N3,N5-tris(3-(didodecylamino)propyl)benzene-1,3,5-tricarboxamide, ...

15. The composition of claim 1 , wherein the lipid nanoparticles comprise inorganic particles.

16. The composition of claim 15, wherein the inorganic particles are within the hydrophobic core of the lipid nanoparticles.

17. The composition of claim 16 , wherein the inorganic particles comprise a metal.

18. 18. The composition of claim 17, wherein the metal comprises a metal salt, a metal oxide, a metal hydroxide, or a metal phosphate.

19. 20. The composition of claim 18, wherein the metal oxide comprises aluminum oxide, aluminum oxyhydroxide, iron oxide, titanium dioxide, or silicon dioxide.

20. The composition of claim 1 , wherein the hydrophobic surfactant is sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, or sorbitan trioleate.

21. The composition of claim 1 , wherein the hydrophilic surfactant is a polysorbate.

22. The composition of claim 9, wherein the hydrophobic core comprises squalene, the hydrophilic surfactant is polysorbate 80, the hydrophobic surfactant is sorbitan monolaurate, the cationic lipid is 1,2-dioleoyloxy-3(trimethylammonium)propane, and optionally further comprising sodium citrate.

23. The composition of claim 22, wherein the hydrophobic core further comprises oleic acid coated iron oxide nanoparticles.

24. The composition of claim 1, wherein the at least one nucleic acid comprises DNA or RNA.

25. The composition described in claim 24, wherein the RNA comprises mRNA.

26. A pharmaceutical composition comprising the composition of claim 1 and a pharmaceutically acceptable excipient.

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