Therapeutic nanoparticles comprising a therapeutic agent and methods of making and using same

Abstract

The present disclosure generally relates to nanoparticles comprising an endo-lysosomal escape agent, a nucleic acid, and a polymer. Other aspects include methods of making and using such nanoparticles.

1 . A pharmaceutically acceptable nanoparticle comprising:
 a nucleic acid and a hydrophobic counter ion agent; and
 about 50 to about 99.75 weight percent of a diblock poly(lactic) acid-poly(ethylene)glycol copolymer or a diblock poly(lactic acid-co-glycolic acid)-poly(ethylene)glycol copolymer. 
   
     
     
         2 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the hydrophobic counter ion agent is an endosomal and/or a lysosomal disrupting agent. 
     
     
         3 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the nucleic acid molecule is selected from the group comprising an antisense compound, mRNA, short interfering nucleic acid (siNA), short interfering RNA (siRNA), double stranded RNA (dsRNA), micro-RNA (miRNA), small nucleolar RNA (sno-RNA), Piwi-interacting RNA (piRNA), and short hairpin RNA (shRNA) molecules. 
     
     
         4 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the nucleic acid molecule is selected from the group comprising an oligonucleotide, an aptamer, a vector, a threose nucleic acid, a glycol nucleic acid (GNAs), and a locked nucleic acid (LNAs). 
     
     
         5 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the nucleic acid comprises a modification. 
     
     
         6 . The pharmaceutically acceptable nanoparticle of  claim 5 , wherein the modification is at least one of: a 2′O-methyl (2′OMe) modification, a 2′Fluoro (2′F) modification, a 2′Methoxyethyl (2′-O-MOE) modification, a 2′Fluorarabino (FANA) modification, a 2′-H modification, a 2′-Thiouracil modification, a locked nucleic acid (LNA) modification, a bridged nucleic acid (BNA) modification, a ethylene-bridged nucleic acid (ENA) modification, a hexitol nucleic acid (HNA) modification, an altritol nucleic acid (ANA) modification, a cyclohexene nucleic acid (CeNA) modification, an unlocked nucleic acid (UNA) modification, a 4′Thio (4′-S) modification, a thiol linkage modification, and a 3′ inverted abasic end cap modification. 
     
     
         7 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the hydrophobic counter ion agent is an endo-lysosomal disrupting agent. 
     
     
         8 . The pharmaceutically acceptable nanoparticle of  claim 1 , further comprising a poly(lactic) acid-poly(ethylene)glycol copolymer functionalized with a ligand. 
     
     
         9 . The pharmaceutically acceptable nanoparticle of  claim 8 , wherein at least one ligand is a targeting ligand. 
     
     
         10 . The pharmaceutically acceptable nanoparticle of  claim 9 , wherein the at least one targeting ligand is selected from the group consisting of a nucleic acid, an antibody, a small molecule, an aptamer, a polypeptide, a protein, a carbohydrate, a lipid and an oligonucleotide. 
     
     
         11 . The pharmaceutically acceptable nanoparticle of  claim 10 , wherein the antibody targets a target selected from the group consisting of EpCAM (CD326), IGF-R, Mesothelin, Lewis-Y antigen (CD174), CanAg (MUC1, PEM, CA242, CD205), NCAM (CD56), Cripto, Melanotransferrin (P97), Glycoprotein NMB (CG56972), CD70 (CD27 Ligand), 5T4 (trophoblast glycoprotein), CD57, CD44, Carcinoembryonic antigen (CEA), GD2, CD40, Fibronectin ED-B, Endoglin (CD105), Tenascin C, Phosphatidylserine (PS), HER3, CD30, CD33, CD40, CD52, CD74, CD138, CS1 (CD319, CRACC), TAG-72, CD2, CD64, ROBO4, DLL4, Tie2, and B7-H3. 
     
     
         12 . The pharmaceutically acceptable nanoparticle of  claim 10 , wherein the polypeptide targets a target selected from the group consisting of TRAIL R2, c-Met, EphA2, EphB2, Aminopeptidase N (CD13), VLA-4 (α4β1 integrin), CXCR4, Melanocortin receptor (MC1R), Somatostatin receptor, Cholecystokinin Receptor, GnRH Receptor, GLP1-receptor, E-Selectin, IL-11 receptor, Thrombospondin-1 receptor, Endostatin, CD79, and CD74. 
     
     
         13 . The pharmaceutically acceptable nanoparticle of  claim 9 , wherein the targeting ligand targets a receptor site for endocytosis. 
     
     
         14 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the hydrophobic counter ion is selected from the group consisting of chlorpromazine, tamoxifen, clomiphene, raloxifene, tamoxifen citrate, toremifene, clomiphene citrate, toremifene citrate, verapamil, dilitiazem, amlodipine, nifedpine, verapamil hydrochloride, nimodipine, felodipine, nicardpine, nisoldipine clevidipine, isradipine, trandolapril/verapamil, desipramine, imipramine, amitriptyline, nortiptyline, clomipramine, doxepin, amoxapine, trimipramine, protriptyline, amiodarone, lidocaine, sotalol, dronedarone, flecainide, procainamide, propafenone, quinidine, dofetilide, mexiletine, ibutilide, disopyramide, paroxetine, fluoxetine, sertraline, escitalopram, citalopram, fluvoxamine, paroxetine hydrochloride, nefazodone, citalopram hydrobromide, escitalopram oxalate, olanzapine/fluoxetine, chlorcyclizine, amodiaquine, thioridazine, chloroquine, quinine, atovaquone/proguanil, atovaquone, fluoxetine, mefloquine, primaquine, quinacrine, quinidine, halofantrine, Chloroquine, monensin, chlorcyclizine, Antrafenine, Aripiprazole, Bifeprunox, Brexpiprazole, Cariprazine, Ciprofloxacin, Dapiprazole, Dropropizine, Elopiprazole, Etoperidone, Itraconazole, Ketoconazole, Levodropropizine, Mepiprazole, Mianserin, Naftopidil, Nefazodone, Niaprazine, Oxypertine, Posaconazole, Trazodone, Umespirone, Urapidil, Vesnarinone, Lubazodone, Acaprazine, Batoprazine, Bifeprunox, Vortioxetine, Vilazodone, Tolpiprazole, Sonepiprazole, Pardoprunox, Naphthylpiperazine, Naluzotan, Lorpiprazole, Flesinoxan, Fluprazine, Flibanserin, Ensaculin, Enpiprazole, Eltoprazine, Elopiprazole, UNC 7938, sphingosine, dodecylimidazole, bafilomycin A1, quinolones, Omeprazole, Esmoprazole, pantoprazole, lansoprazole, rabeprazole, dexiansoprazole, Brefeldin A, Golgicide, Dyasore, Pitstop, amodiaquine, EGA (4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone), fluoxetine, paroxetine, sertraline, thioridazine, phenothiazine, promethazine, prochlorperzaine, trifluoperazine, fluphenazine, prochlorperzine, fluphenazine decanoate, promethazine hydrochloride, quinine, Calcimycin, mefloquine, aprindine, disopyramide, flecainide, lidocaine, mexiletine, pentisomide, propafenone, cyproheptadine azatadine, ketotifen, loratadine, pizotifen, amitriptyline, propranolol, rupatadine, deptropine, amisulpride, nortriptyline, cyclobenzaprine, octriptyline, butriptyline, iprindole, trimipramine, flavoxate, cinnarizine chlomipramine, fluoxetine, promazine, imipramine, sertraline, carbamazepine, ay9944, Clomipramine, clozapine, flecainide, haloperidol, ketoconazole, ofloxacin, perhexiline, sotalol, temoxiphen, zimelidine, Cyproheptadine, toremifene, fluphenazine, trifluoperazine, pizotyline, CGS 12066B, Prochlorperazine, Doxepin, ketotifen, lacidipine, sb 205607, lofepramine, mifepristone, clobenpropit, salmeterol, azelastine, azelastine, epinastine, desloratadine, am-251, indatraline, nelfinavir, haloperidol, benproperine, M-paroxetine, carvedilol, calcipotriol, perphenazine, phenothiazine, chlorprothixene, desipramine, tetracaine, ifenprodil, U18666A, and diphenhydramine. 
     
     
         15 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the hydrophobic counter-ion agent has a log P of about 2 or more. 
     
     
         16 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the hydrophobic counter-ion agent has a log P of between about 2 and about 3. 
     
     
         17 . The pharmaceutically acceptable nanoparticle of  claim 1 , comprising about 5 to about 20 weight percent of the nucleic acid. 
     
     
         18 . The pharmaceutically acceptable nanoparticle of  claim 1 , comprising about 5 to about 20 weight percent of the hydrophobic counterion agent. 
     
     
         19 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the zeta potential of the nanoparticle is about −10 to about 10 mV. 
     
     
         20 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the nucleic acid is substantially stable in a Dnase assay. 
     
     
         21 . The pharmaceutically acceptable nanoparticle of  claim 1 , wherein the poly(lactic) acid-poly(ethylene)glycol copolymer has a number average molecular weight of about 10 kDa to about 35 kDa poly(lactic acid) and a number average molecular weight of about 4 kDa to about 6 kDa poly(ethylene)glycol. 
     
     
         22 . A pharmaceutically acceptable nanoparticle comprising:
 a nucleic acid and an endo-lysosomal disrupting agent, wherein the amount of endolysomal disrupting agent present is at least sufficient to form a hydrophobic counter ion with the nucleic acid; and   about 50 to about 99.75 weight percent of a diblock poly(lactic) acid-poly(ethylene)glycol copolymer or a diblock poly(lactic acid-co-glycolic acid)-poly(ethylene)glycol copolymer.   
     
     
         23 . The pharmaceutically acceptable nanoparticle of  claim 22 , wherein the nucleic acid molecule is selected from the group comprising an antisense compound, mRNA, short interfering nucleic acid (siNA), short interfering RNA (siRNA), double stranded RNA (dsRNA), micro-RNA (miRNA), small nucleolar RNA (sno-RNA), Piwi-interacting RNA (piRNA), and short hairpin RNA (shRNA) molecules. 
     
     
         24 . The pharmaceutically acceptable nanoparticle of  claim 22 , wherein the nucleic acid molecule is selected from the group comprising an oligonucleotide, an aptamer, a vector, a threose nucleic acid, a glycol nucleic acid (GNAs), and a locked nucleic acid (LNAs). 
     
     
         25 . The pharmaceutically acceptable nanoparticle of  claim 22 , wherein the nucleic acid comprises a modification. 
     
     
         26 . The pharmaceutically acceptable nanoparticle of  claim 25 , wherein the modification is at least one of: a 2′O-methyl (2′OMe) modification, a 2′Fluoro (2′F) modification, a 2′Methoxyethyl (2′-O-MOE) modification, a 2′Fluorarabino (FANA) modification, a 2′-H modification, a 2′-Thiouracil modification, a locked nucleic acid (LNA) modification, a bridged nucleic acid (BNA) modification, a ethylene-bridged nucleic acid (ENA) modification, a hexitol nucleic acid (HNA) modification, an altritol nucleic acid (ANA) modification, a cyclohexene nucleic acid (CeNA) modification, an unlocked nucleic acid (UNA) modification, a 4′Thio (4′-S) modification, a thiol linkage modification, and a 3′ inverted abasic end cap modification. 
     
     
         27 . The pharmaceutically acceptable nanoparticle of  claim 22 , further comprising a poly(lactic) acid-poly(ethylene)glycol copolymer functionalized with a ligand. 
     
     
         28 . The pharmaceutically acceptable nanoparticle of  claim 27 , wherein at least one ligand is a targeting ligand. 
     
     
         29 . The pharmaceutically acceptable nanoparticle of  claim 28 , wherein the at least one targeting ligand is selected from the group consisting of a nucleic acid, an antibody, a small molecule, an aptamer, a polypeptide, a protein, a carbohydrate, a lipid and an oligonucleotide. 
     
     
         30 . The pharmaceutically acceptable nanoparticle of  claim 29 , wherein the antibody targets a target selected from the group consisting of EpCAM (CD326), IGF-R, Mesothelin, Lewis-Y antigen (CD174), CanAg (MUC1, PEM, CA242, CD205), NCAM (CD56), Cripto, Melanotransferrin (P97), Glycoprotein NMB (CG56972), CD70 (CD27 Ligand), 5T4 (trophoblast glycoprotein), CD57, CD44, Carcinoembryonic antigen (CEA), GD2, CD40, Fibronectin ED-B, Endoglin (CD105), Tenascin C, Phosphatidylserine (PS), HER3, CD30, CD33, CD40, CD52, CD74, CD138, CS1 (CD319, CRACC), TAG-72, CD2, CD64, ROBO4, DLL4, Tie2, and B7-H3. 
     
     
         31 . The pharmaceutically acceptable nanoparticle of  claim 29  wherein the polypeptide targets a target selected from the group consisting of TRAIL R2, c-Met, EphA2, EphB2, Aminopeptidase N (CD13), VLA-4 (α4β1 integrin), CXCR4, Melanocortin receptor (MC1R), Somatostatin receptor, Cholecystokinin Receptor, GnRH Receptor, GLP1-receptor, E-Selectin, IL-11 receptor, Thrombospondin-1 receptor, Endostatin, CD79, and CD74. 
     
     
         32 . The pharmaceutically acceptable nanoparticle of  claim 28 , wherein the targeting ligand targets a receptor site for endocytosis. 
     
     
         33 . The pharmaceutically acceptable nanoparticle of  claim 22 , wherein the zeta potential of the nanoparticle is about −10 to about 10 mV. 
     
     
         34 . The pharmaceutically acceptable nanoparticle of  claim 22 , wherein the nucleic acid is substantially stable in a Dnase assay. 
     
     
         35 . The pharmaceutically acceptable nanoparticle of  claim 22 , wherein the poly(lactic) acid-poly(ethylene)glycol copolymer has a number average molecular weight of about 10 kDa to about 35 kDa poly(lactic acid) and a number average molecular weight of about 4 kDa to about 6 kDa poly(ethylene)glycol.