Remote loading of sparingly water-soluble drugs into liposomes

Abstract

The present invention provides liposome compositions containing sparingly soluble drugs that are used to treat life-threatening diseases. A preferred method of encapsulating a drug inside a liposome is by remote or active loading. Remote loading of a drug into liposomes containing a transmembrane electrochemical gradient is initiated by co-mixing a liposome suspension with a solution of drug, whereby the neutral form of the compound freely enters the liposome and becomes electrostatically charged thereby preventing the reverse transfer out of the liposome. There is a continuous build-up of compound within the liposome interior until the electrochemical gradient is dissipated or all the drug is encapsulated in the liposome. However, this process as described in the literature has been limited to drugs that are freely soluble in aqueous solution or solubilized as a water-soluble complex. This invention describes compositions and methods for remote loading drugs with low water solubility (<2 mg/mL). In the preferred embodiment the drug in the solubilizing agent is mixed with the liposomes in aqueous suspension so that the concentration of solubilizing agent is lowered to below its capacity to completely solubilize the drug. This results in the drug precipitating but remote loading capability is retained. The process is scalable and, in liposomes in which the lipid composition and remote loading agent are optimized, the resulting drug-loaded liposomes are characterized by a high drug-to-lipid ratios and prolonged drug retention when the liposome encapsulated drug is administered to a subject.

What is claimed is: 
     
         1 . A pharmaceutical formulation comprising a sparingly water-soluble agent encapsulated within a liposome, said formulation manufactured by a method comprising:
 contacting an aqueous suspension of said liposome with said aqueous suspension of said agent under conditions appropriate to encapsulate said sparingly water-soluble agent in said liposome, wherein   said liposome has an internal aqueous environment encapsulated by a lipid membrane and said aqueous suspension of said liposome comprises a gradient selected from a proton gradient, an ion gradient and a combination thereof across said membrane, and wherein   said conditions are appropriate for said sparingly water-soluble agent to traverse said membrane and concentrate in said internal aqueous environment, thereby forming said pharmaceutical formulation.   
     
     
         2 . The pharmaceutical formulation according to  claim 1 , wherein said aqueous suspension of said sparingly water-soluble agent is prepared by a method comprising:
 essentially completely dissolving the agent in an aprotic solvent, forming an agent solution and subsequently contacting said agent solution with said aqueous suspension of said liposome, such that said aprotic solvent is diluted, reducing solubility of said agent, thereby forming said aqueous suspension of said sparingly water-soluble agent.   
     
     
         3 . The pharmaceutical formulation of  claim 1 , wherein said liposome is selected from multilamellar vesicles (MLV), large unilamellar vesicles (LUV) and small unilamellar vesicles (SUV), oligolamellar vesicles (OLV), paucilamellar vesicles (PLV) or reverse phase evaporation vesicles (REV). 
     
     
         4 . The pharmaceutical formulation according to  claim 1 , wherein said agent is a low molecular weight therapeutic agent having a water solubility of less than or equal to about 2 mg/mL. 
     
     
         5 . The pharmaceutical formulation of  claim 1 , wherein, the agent is selected from an anthracycline compound, a camptothecin compound, a vinca alkaloid, an ellipticine compound, a taxane compound, a wortmannin compound, a geldanamycin compound, a pyrazolopyrimidine compound, a steroid compound, a peptide-based compound, a derivative of any of the foregoing, a pro-drug of any of the foregoing, and an analog of any of the foregoing. 
     
     
         6 . The pharmaceutical formulation according to  claim 1 , wherein said agent is selected from carfilzomib, voriconazole, amiodarone, ziprasidone, aripiprazole, imatinib, lapatinib, oprozomib, cyclopamine, CUR-61414, PF-05212384, PF-4691502, toceranib, PF-477736, PF-337210, sunitinib, SU14813, axitinib, AG014699, veliparib, MK-4827, ABT-263, SU11274, PHA665752, Crizotinib, XL880, PF-04217903, XR5000, AG14361, veliparib, bosutunib, PD-0332991, PF-01367338, AG14361, NVP-ADW742, NVP-AUY922, NVP-LAQ824, NVP-TAE684, NVP-LBH589, erubulin, doxorubicin, daunorubicin, mitomycin C, epirubicin, pirarubicin, rubidomycin, carcinomycin, N-acetyladriamycin, rubidazone, 5-imido daunomycin, N-acetyl daunomycin, daunoryline, mitoxanthrone, camptothecin, 9-aminocamptothecin, 7-ethylcamptothecin, 7-Ethyl-10-hydroxy-camptothecin, 10-hydroxycamptothecin, 9-nitrocamptothecin,10,11-methylenedioxycamptothecin, 9-amino-1O,11-methylenedioxycamptothecin, 9-chloro-l 0,11-methylenedioxycamptothecin, irinotecan, lurtotecan, silatecan, (7-(4-methylpiperazinomethylene)-10,ll-ethylenedioxy-20(S)-camptothecin, 7-(4-methylpiperazinomethylene)-10, II-methylenedioxy-20(S)-camptothecin, 7-(2-N-isopropylamino)ethyl)-(20S)-camptothecin, CKD-602, vincristine, vinblastine, vinorelbine, vinflunine, vinpocetine, vindesine, ellipticine, 6-3-aminopropyl-ellipticine, 2-diethylaminoethyl-ellipticinium, datelliptium, retelliptine, paclitaxel, docetaxel, diclofenac, bupivacaine, 17-allylamino-geldanamycin, 17-dimethylaminoethylamino-17-demethoxygeldanamycin, cetirizine, fexofenadine, Onx 0912, Onx 0914, PD0332991, Axitinib, Lenvatinib, PHA665752, SU11274, PF-02341066, foretinib, XL880, PX-478, GDC-0349, PD0332991, AZD4547, Golotimod, SCH900776, TG02, UNC0638, ARRY-520, Elacridar hydrochloride, golvatinib, MK-1775, PF-03758309, AT13387, BAY 80-6946, cobicistat, GDC-0068, INNO-206, MLN0905, resminostat, tariquidar, primidone and other catecholamines, epinephrine, salts, prodrugs and derivatives of these medicinal compounds and mixtures thereof. 
     
     
         7 . The pharmaceutical formulation according to  claim 1 , wherein said agent is selected from an antihistamine ethylenediamine derivative, bromphenifamine, diphenhydramine, an anti-protozoal drug, quinolone, iodoquinol, an amidine compound, pentamidine, an antihelmintic compound, pyrantel, an anti-schistosomal drug, oxaminiquine, an antifungal triazole derivative, fliconazole, itraconazole, ketoconazole, miconazole, an antimicrobial cephalosporin, thelating agents, deferoxamine, deferasirox, deferiprone, FBS0701, cefazolin, cefonicid, cefotaxime, ceftazimide, cefuoxime, an antimicrobial beta-lactam derivative, aztreopam, cefmetazole, cefoxitin, an antimicrobial of erythromycin group, erythromycin, azithromycin, clarithromycin, oleandomycin, a penicillin compound, benzylpenicillin, phenoxymethylpenicillin, cloxacillin, methicillin, nafcillin, oxacillin, carbenicillin, a tetracycline compound, novobiocin, spectinomycin, vancomycin; an antimycobacterial drug, aminosalicycic acid, capreomycin, ethambutol, isoniazid, pyrazinamide, rifabutin, rifampin, clofazimine, an antiviral adamantane compound, amantadine, rimantadine, a quinidine compound, quinine, quinacrine, chloroquine, hydroxychloroquine, primaquine, amodiaquine, mefloquine, an antimicrobial, qionolone, ciprofloxacin, enoxacin, lomefloxacin, nalidixic acid, norfloxacin, ofloxacin, a sulfonamide; a urinary tract antimicrobial, nitrofurantoin, trimetoprim; anitroimidazoles derivative, metronidazole, a cholinergic quaternary ammonium compound, ambethinium, neostigmine, physostigmine, an anti-Alzheimer aminoacridine, tacrine, an anti-parkinsonal drug, benztropine, biperiden, procyclidine, trihexylhenidyl, an anti-muscarinic agent, atropine, hyoscyamine, scopolamine, propantheline, an adrenergic compound, dopamine, serotonin, a hedgehog inhibitor, albuterol, dobutamine, ephedrine, epinephrine, norepinephrine, isoproterenol, metaproperenol, salmetrol, terbutaline, a serotonin reuptake inhibitor, an ergotamine derivative, a myorelaxant, a curare series, a central action myorelaxant, baclophen, cyclobenzepine, dentrolene, nicotine, a nicotine receptor antagonist, a beta-adrenoblocker, acebutil, amiodarone, abenzodiazepine compound, ditiazem, an antiarrhythmic drug, diisopyramide, encaidine, a local anesthetic compound, procaine, procainamide, lidocaine, flecaimide, quinidine, an ACE inhibitor, captopril, enelaprilat, Hsp90 inhibitor, fosinoprol, quinapril, ramipril; an opiate derivative, codeine, meperidine, methadone, morphine, an antilipidemic, fluvastatin, gemfibrosil, an HMG-coA inhibitor, pravastatin, a hypotensive drug, clonidine, guanabenz, prazocin, guanethidine, granadril, hydralazine, a non-coronary vasodilator, dipyridamole, an acetylcholine esterase inhibitor, pilocarpine, an alkaloid, physostigmine, neostigmine, a derivative of any of the foregoing, a pro-drug of any of the foregoing, and analog of any of the foregoing. 
     
     
         8 . The pharmaceutical formulation according to  claim 2 , wherein the aprotic solvent is selected from dimethylsulfoxide, dioxane, tetrahydrofuran, dimethylformamide, acetonitrile, dimethylacetamide, sulfolane, gamma butyrolactone, pyrrolidones, 1-methyl-2-pyrrolidinone, methylpyrroline, ethylene glycol monomethyl ether, diethylene glycol monomethyl ether, polyethylene glycol. 
     
     
         9 . The pharmaceutical formulation according to  claim 1 , wherein the liposome is prepared from one or more lipids selected from egg phosphatidylcholine (EPC), egg phosphatidylglycerol (EPG), dipalmitoylphosphatidylcholine (DPPC), sphingomyelin (SM), cholesterol (Chol), cholesterol sulfate and its salts (CS), cholesterol hemisuccinate and its salts (Chems), cholesterol phosphate and its salts (CP), cholesterol phthalate, cholesterylphosphorylcholine, 3,6,9-trioxaoctan-ol-cholesteryl-3e-ol, dimyristoylphosphatidylglycerol (DMPG), dimyristoylphosphatidylglycerol (DMPG), dimyristoylphosphatidylcholine (DMPC), distearoylphosphatidylcholine (DSPC), hydrogenated soy phosphatidylcholine (HSPC), distearoylphosphatidylglycerol (DSPG), sterol modified lipids (SML), inverse-phosphocholine lipids, cationic lipids and zwitterlipids. 
     
     
         10 . The pharmaceutical formulation according to  claim 1 , wherein the percentage of agent from said aqueous suspension of said agent encapsulated in said internal aqueous medium of said liposome ranges from about 10% to about 100% of the total agent in said aqueous suspension of said agent. 
     
     
         11 . The pharmaceutical formulation according to  claim 1 , wherein said ion gradient is caused by a difference in concentrations across said membrane of a member selected from an amine salt and a metal salt of a member selected from a carboxylate, a sulfate, a phosphonate and a phosphate. 
     
     
         12 . The pharmaceutical formulation according to  claim 11 , wherein said carboxylate is acetate 
     
     
         13 . The pharmaceutical formulation according to  claim 11 , wherein said amine salt and said metal salt are selected from a salt of a member selected from a monovalent carboxylate, a multivalent carboxylate, a sulfate, a phosphonate and a phosphate. 
     
     
         14 . The pharmaceutical formulation according to  claim 11 , wherein the cation in said salt is selected from sodium, calcium, magnesium, zinc, copper, potassium, primary, secondary, tertiary and quaternary ammonium species. 
     
     
         15 . The pharmaceutical formulation according to  claim 1 , wherein said formulation is lyophilized. 
     
     
         16 . The pharmaceutical formulation according to  claim 1 , wherein said agent is present in said internal aqueous medium as a unit dosage format. 
     
     
         17 . A method of treating a disease state in a subject in need of said treatment, said method comprising administering to said patient a therapeutically effective amount of said formulation according to  claim 1 .