Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/10—Dispersions; Emulsions
  • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
  • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
  • A61K47/22—Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/0012—Galenical forms characterised by the site of application
  • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/10—Dispersions; Emulsions
  • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
  • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers

Definitions

• This invention is in the field of pharmaceutical agents.
• this invention relates to pharmaceutical agents wherein one or more tocols is used as a primary solvent.
• a few examples of therapeutic substances in these categories are ibuprofen, diazepam, griseofulvin, cyclosporin, cortisone, proleukin, etoposide and paclitaxel.
• Kagkadis, KA et al. (1996) PDA J Pharm Sci Tech 50(5):317- 323; Dardel, O. 1976. Anaesth Scand 20:221-24. Sweetana, S and MJU Akers. (1996) PDA J Pharm Sci Tech 50(5):330-342.
• oils typically used for pharmaceutical emulsions include saponifiable oils from the family of triglycerides, for example, soybean oil, sesame seed oil, cottonseed oil, safflower oil and the like. Hansrani. PK et al., (1983) J. Parenter Sci. Technol 37:145-150.
• One or more surfactants are used to stabilize the emulsion, and excipients are added to render the emulsion more biocompatible, stable and less toxic.
• Lecithin from egg yolks or soybeans is a commonly used surfactant. Sterile manufacturing can be accomplished by absolute sterilization of all the components before manufacture, followed by absolutely aseptic technique in all stages of manufacture. However, improved ease of manufacture and assurance of sterility is obtained by terminal sterilization following sanitary manufacture, either by heat or by filtration. Unfortunately, not all emulsions are suitable for heat or filtration treatments.
• Stability has been shown to be influenced by the size and homogeneity of the emulsion.
• the preferred emulsion consists of a suspension of sub-micron particles, with a mean droplet diameter of no greater than 200 nanometers.
• a stable dispersion in this size range is not easily achieved, but has the benefit that it is expected to circulate longer in the bloodstream. Further, less of the stable dispersion in this size range is phagocytized non-specifically by the reticuloendothelial system. As a result the drug is more likely to reach its therapeutic target.
• a preferred drug emulsion will be designed to be actively taken up by the target cell or organ, and is targeted away from the RES.
• the use of vitamin E in emulsions is known.
• mice an injectable form of vitamin E was prepared by Kato and coworkers. Kato Y., et al. (1993) Chem Pharm Bull 41(3):599-604. Micellar solutions were formulated with Tween 80, Brij 58 and HCO-60. Isopropanol was used as a co-solvent, and was then removed by vacuum evaporation; the residual oil glass was then taken up in water with vortexing as a micellar suspension. An emulsion was also prepared by dissolving vitamin E with soy phosphatidycholine (lecithin) and soybean oil. Water was added and the emulsion prepared with sonication.
• soy phosphatidycholine lecithin
• U.S. Patent 5,689,846 discloses various alcohol solutions of paclitaxel.
• U.S. Patent 5,573,781 discloses the dissolution of paclitaxel in ethanol, butanol and hexanol and an increase in the antitumor activity of paclitaxel when delivered in butanol and hexanol as compared to ethanol.
• Alcohol-containing solutions can be administered with care, but are typically given by intravenous drip to avoid the pain, vascular irritation and toxicity associated with bolus injection of these solutions.
• patent 4,439,432 discloses preparing high concentration solutions of progesterone in tocopherol. Emulsions can be prepared from these solutions, for use as skin treatments for systemic progesterone deficiency, for treating local skin conditions such as psoriasis or for vaginal application. The solutions may also be encapsulated for oral administration
• EP application 001,851 discloses highly concentrated solutions of steroids of the oestrane. androstane, and (19-nor-) pregnane series which include tocol and tocol derivatives that are liquid at normal temperature.
• PCT publication WO 95/21217 discloses that tocopherols can be used as solvents and/or emulsifiers of drugs that are substantially insoluble in water, in particular for the preparation of topical formulations.
• the use of vitamin E-TPGS as an emulsifier in formulations containing high levels of ⁇ -tocopherol is mentioned in the specification (pages 7-8 and 12). Examples
• formulations for topical administration comprising a lipid layer ( ⁇ -tocopherol), the drug and Vitamin E-TPGS, in quantities of less than 25% w/w of the formulation, as an emulsifier.
• PCT Publication WO 97/03651 discloses lipid drug delivery compositions that contain at least five ingredients: a therapeutic drug, vitamin E, an oil in which the drug and vitamin E are dissolved, a stabilizer (either phospholipid, a lecithin, or a poloxamer which is a polyoxyethylene-polyoxypropylene copolymer) and water.
• the therapeutic drugs disclosed are itraconazole and paclitaxel.
• the "therapeutic emulsion" compositions require two oils in the dispersed phase where the therapeutic drug resides, vitamin E and another oil, typically a triglyceride such as soybean oil.
• the only working example with paclitaxel, Example 16 also contains both vitamin E and soybean oil.
• WO 97/03651 also discloses, incidentally, that tocol derivatives and tocotrienols that have Vitamin E activity are considered to be within the definition of "Vitamin E" as used in that publication.
• PCT publication WO 97/22358 discloses microemulsion preconcentrates of cyclosporin dissolved in a solvent system that can include a hydrophilic component selected from tocol, tocopherols, tocotrienols, and their derivatives.
• a hydrophilic component selected from tocol, tocopherols, tocotrienols, and their derivatives.
• the publication also mentions ⁇ -, ⁇ - and ⁇ - tocopherols, and ⁇ -, ⁇ -, ⁇ and ⁇ -tocotrienols or mixtures of them.
• These compositions also include a hydrophilic solvent, preferably propylene carbonate or polyethylene glycols having an average molecular weight of less than 1000.
• a second PCT publication of Sherman discloses microemulsion preconcentrates of cyclosporins in which the solvent system may comprise two hydrophobic solvents, one of which is selected from tocol, tocopherols, tocotrienols and their derivatives.
• NMP N-methyl-2-pyrrolidone
• PharmosolveTM can be used to improve the solubility of poorly soluble drugs in pharmaceutical formulations and has appeared in recent literature for use in veterinary medicine with forthcoming application in humans.
• polyvinylpyrrolidone under the trade name PovidoneTM with a molecular weight between 2,500 to 100,000 at a concentration of 1 to 5 percent (w/v) of the aqueous injectable base can be used as a co-solubilizer along with NMP.
• PVP polyvinylpyrrolidone
• U.S. Patent 5,726,181 discloses antitumor compositions and suspensions comprising NMP and highly lipophilic camptothecin derivatives.
• Polyethylene glycols (PEGs) and PVP are examples of two water- soluble polymers frequently used to modify the solubility behavior of drugs, including paclitaxel.
• solubility of paclitaxel in both solvents is relatively high, in dilute aqueous solutions that are suitable for parenteral administration the solubility of the drug is low and the potential for drug precipitation upon dilution is high.
• solubility of paclitaxel In admixtures of PEG 400 and water containing 50-100% PEG 400, the solubility of paclitaxel varies from 0.2 to 175 mg/ml, respectively.
• paclitaxel solubilities are quite low where larger amounts of water are used, e.g., in 35% PEG 400 and 30% PVP in water are 0.03 mg/ml and ⁇ 0.3 mg/ml, respectively.
• “Solubility of paclitaxel in Polyethylene Glycol 400/Water Mixtures” (Straubinger, R. M.
• Biopharmacuitics of paclitaxel (Taxol); Formulation, activity and pharmacokinetics, p.244 In Taxol, Science and Applications. (M. Suffness ed.), CRC Press, New York, 1995).
• the use of PEG-400 is not limited to paclitaxel and can be applied to other therapeutic agents which exhibit good solubility in polyethylene glycols (for example Etoposide).
• Derivative forms of paclitaxel including polyethylene glycol derivatives are described in U.S. Patent 5.614,549.
• the current commercial formulation for the anti-cancer drug paclitaxel for example, consists of a mixture of hydro xylated castor oil and ethanol, and rapidly extracts plasticizers such as di-(2- ethylhexyl)-phthalate from commonly used intravenous infusion tubing and bags. Adverse reactions to the plasticizers have been reported, such as respiratory distress, necessitating the use of special infusion systems at extra expense and time. Waugh, et al. (1991) Am J. Hosp. Pharmacists 48:1520.
• the ideal emulsion vehicle would be inexpensive, non-irritating or even nutritive and palliative in itself, terminally sterilizable by either heat or filtration, stable for at least 1 year under controlled storage conditions, accommodate a wide variety of water insoluble and poorly soluble drugs and be substantially ethanol-free.
• a vehicle which will stabilize, and carry in the form of an emulsion, drugs which are poorly soluble in lipids and in water.
• compositions including: one or more tocols, with or without an aqueous phase, a surfactant or mixtures of surfactants incorporating a co-solvent and a therapeutic agent.
• the compositions of the invention may be in the form of an emulsion, micellar solution or a self-emulsifying drug delivery system.
• the tocol (or tocopherol) molecule is preferably ⁇ -tocopherol.
• the compositions of the invention are generally substantially free of any monohydric alcohol.
• the co-solvent may include water-soluble polymers, preferably polyethylene glycols or polyvinylpyrrolidone with or without N-methyl-2- pyrrolidone.
• PEGs Polyethylene glycols with a molecular weight between 100 to 10,000 are the most preferred co-solvent. Most preferred is PEG-400 in amounts greater than 1% by weight of the formulation.
• the pharmaceutical compositions can be stabilized by the addition of various amphiphilic molecules, including anionic, nonionic, cationic, and zwitterionic surfactants. Preferably, these molecules are PEGylated surfactants and optimally PEGylated ⁇ -tocopherol.
• amphiphilic molecules further include surfactants such as ascorbyl-6 palmitate: stearylamine; sucrose fatty acid esters, pegylated phospholipids, various tocol derivatives and a polyoxypropylene-polyoxyethylene glycol nonionic block copolymer.
• useful surfactants also include poloxamers, tetronics. TPGS, glutamyl stearate, pegylated mono- and diglycerides, propylene glycol mono-/diesters, polyglyceryl esters , Solutol HS-15, phospholipids.
• lecithins pegylated phospholipids, pegylated sterols, pegylated cholesterol, and other tocol esters, sucrose esters, fatty acids, bile acids and conjugated bile acids, nonionic and anionic surfactants.
• the therapeutic agent may be a chemotherapeutic agent, an antibiotic (antiviral, antibacterial, antihelminthic. antiplasmodial, or antimycotic), an analgesic, an antidepressant, antipsychotic. a hormone, a steroid, a vascular tonic, an angiogenesis inhibitor, a cytomedine or a cytokine.
• Tocol based emulsions are especially advantageous with synergistic biological and therapeutic effects when used to deliver cardiovascular or cancer therapeutics.
• An added advantage of a particulate emulsion for the delivery of a chemotherapeutic is the widespread property of surfactants used in emulsions to overcome multidrug resistance by inhibiting P-glycoprotein. a membrane-bound drug transporter.
• the emulsions of the invention can comprise an aqueous medium when in the form of an emulsion or micellar solution.
• This medium can contain various additives to assist in stabilizing the emulsion or in rendering the formulation biocompatible.
• the invention is directed to a pharmaceutical composition comprising ⁇ -tocopherol. a chemotherapeutic selected from taxoids, taxines and taxanes, water and D- ⁇ -tocopherol polyethyleneglycol 1000 succinate.
• the invention is directed to a pharmaceutic composition comprising ⁇ -tocopherol. a co-solvent, one or more surfactants, an aqueous phase and a therapeutic agent wherein the composition is in the form of an emulsion or micellar solution and the solution is substantially free of any monohydric alcohol.
• the co-solvent may be polyethylene glycol, N- methyl-2-pyrrolidone, polyvinyl-pyrrolidone or mixtures thereof.
• the surfactant is an ⁇ -tocopherol derivative and the polyethylene glycol has a molecular weight between 100 to 10,000 most preferably from about 200 to about 1000.
• the therapeutic agent is a chemotherapeutic agent selected from taxoids, taxines and taxanes.
• compositions of the invention are typically formed by dissolving a therapeutic agent in the co-solvent to form a therapeutic agent solution; one or more tocols are then added along with one or more surfactants to the therapeutic agent solution to form an oil solution of the therapeutic agent in the hydrophilic co-solvent.
• the oil solution is then blended with an aqueous phase to form a pre-emulsion.
• the pre-emulsion is further homogenized to form a fine emulsion.
• the oil solution of the therapeutic agent in the co-solvent along with surfactants is typically encapsulated in a gelatin capsule.
• the therapeutic agent is dissolved in directly in polyethylene glycol or in a tocol oil which allows the avoidance of the use of monohydric alcohols as a solvent.
• Figure 1 A shows the particle size of a paclitaxel emulsion (QWA) at 7 °C over time;
• Figure IB shows the particle size of a paclitaxel emulsion (QWA) at 25°C over time
• Figure 2 is an HPLC chromatogram showing the integrity of a paclitaxel in an emulsion as described in Example 5;
• Figure 3 A shows the paclitaxel concentration of a paclitaxel emulsion (QWA) at 4°C over time
• Figure 3B shows the paclitaxel concentration of a paclitaxel emulsion (QWA) at 25°C over time
• Figure 4 shows the percentage of paclitaxel released over time from three different emulsions.
• the symbol • represents the percentage of paclitaxel released over time from an emulsion commercially available from Bristol Myers Squibb.
• the symbol A represents the percentage of paclitaxel released over time from an emulsion of this invention containing 6 mg/ml paclitaxel (QWA) as described in Example 6.
• the symbol 0 represents the percentage of paclitaxel released over time from an emulsion of this invention (QWB) containing 7 mg/ml paclitaxel as described in Example 7.
• Figure 5 shows the efficacy of a PEG-400/Vitamin E/paclitaxel emulsion against B 16 melanoma in mice.
• Tocopherols are a family of natural and synthetic compounds, also known by the generic names tocols or Vitamin E. ⁇ - Tocopherol, is the most abundant and active form of this class of compounds and it has the following chemical structure (Scheme I):
• the molecule contains three structural elements, a chroman head with a phenolic alcohol and a phytyl tail. Not all tocopherol isomers have three methyl groups on the chroman head.
• the simplest isomer contains no methyl groups on the chroman ring, (6-hydroxy-2-methyl-2-phytylchroman) and is sometimes simply referred to as "tocol” although we will use the term "tocols"as defined below to represent a broader class of compounds.
• Other members of this class include ⁇ -, ⁇ -, ⁇ -, and ⁇ - tocopherols and ⁇ -tocopherol derivatives such as tocopherol acetate, phosphate, succinate, nicotinate, and linoleate.
• tocopherols and their derivatives are useful as therapeutic agents.
• Tocotrienols have structures related to the tocopherols but which possess a 3, 7,
• tocopherols not all tocotrienol isomers have three methyl groups on the chroman head. There are four major tocotrienols, ⁇ -, ⁇ -, ⁇ -, and ⁇ -tocotrienols. Analogous compounds having fewer methyl groups such as desmethyltocotrienol and didesmtheyltocotrienol are included within the definition of tocotrienols.
• tocopherols can result in emulsions that have lower viscosity, and are thus easier to produce and/or process.
• Preferred members of this group include but are not limited to d- ⁇ - and d- ⁇ - tocopherols, d- ⁇ -, d- ⁇ - and d- ⁇ -tocotrienols.
• Tocols is used herein in a broad sense to indicate the family of tocopherols and tocotrienols and derivatives thereof, since all tocopherols and tocotrienols are fundamentally derivatives of the simplest tocopherol, 6- hydroxy-2-methyl-2-phytylchroman (sometimes referred to as "tocol"). Tocols also include tocopherol or tocotrienol derivatives, including those common derivatives esterified at the 6-hydroxyl on the chroman ring.
• Surfactants Surface active class of amphiphilic molecules which are manufactured by chemical processes or purified from natural sources or processes. These can be anionic. cationic. nonionic, and zwitterionic. Typical surfactants are described in Emulsions: Theory and Practice, Paul Becher, Robert E. Krieger Publishing, Malabar, Florida, 1965; Pharmaceutical Dosage Forms: Dispersed Systems Vol. I, Martin M. Rigear, Surfactants and U.S. Patent No. 5,595,723 which is assigned to the assignee of this invention, Sonus Pharmaceuticals. All of these references are hereby inco ⁇ orated by reference.
• TPGS or PEGylated vitamin E is a vitamin E derivative in which polyethylene glycol subunits are attached by a succinic acid diester at the ring hydroxyl of the vitamin E molecule.
• vitamin E TPGS Various chemical derivatives of vitamin E TPGS including ester and ether linkages of various chemical moieties are included within the definition of vitamin E TPGS.
• Polyethylene glycol is a hydrophilic. polymerized form of ethylene glycol. consisting of repeating units of the chemical structure - (CH 2 -CH 2 -O-).
• the general formula for polyethylene glycol is HOCH 2 (CH 2 OCH 2 ) n CH 2 OH or H(OCH 2 CH 2 ) n OH.
• the molecular weight ranges from 200 to 10,000. Such various forms are described as PEG- 200, PEG-400 and the like.
• N-Methyl-2-pyrroIidone N-methyl-2-pyrrolidone (NMP) is an organic molecule with the following chemical structure: (VI)
• a GMP grade of this compound is available under the name PharmasolveTM and is used to improve the solubility of poorly soluble drugs in pharmaceutical formulations.
• the enhanced solubility of certain drugs can be attributed to a complexing action with the nitrogen and carbonyl reactive centers of the molecule.
• Polwinyl pyrrolidone Polyvinyl pyrrolidone (PVP) or Povidone is a water soluble polymer, consisting of repeating units of the chemical structure:
• It's average MW can vary between 2500 and 3xl0 6 special grades of pyrogen free povidone are available for parenteral administration. Concentrations up to 5% w/v can be used as co-solvent for poorly soluble drugs.
• Poloxamers or Pluronics are synthetic block copolymers of ethylene oxide and propylene oxide having the general structure:
• a and b are commercially available from BASF Performance Chemicals (Parsippany, New Jersey) under the trade name Pluronic and which consist of the group of surfactants designated by the CTFA name of Poloxamer 108, 188, 217, 237, 238, 288. 338, 407, 101, 105, 122. 123, 124, 181, 182, 183, 184, 212, 231, 282, 331, 401. 402, 185, 215, 234, 235. 284, 333, 334, 335, and 403.
• Pluronic the trade name of Poloxamer 108, 188, 217, 237, 238, 288. 338, 407, 101, 105, 122. 123, 124, 181, 182, 183, 184, 212, 231, 282, 331, 401. 402, 185, 215, 234, 235. 284, 333, 334, 335, and 403.
• Pluronic the trade name of poloxamers 124, 188
• Solutol HS-15 is a polyethylene glycol 660 hydroxystearate manufactured by BASF (Parsippany, NJ). Apart from free polyethylene glycol and its monoesters, di-esters are also detectable. According to the manufacturer, a typical lot of Solutol HS-15 contains approximately 30% free polyethylene glycol and 70% polyethylene glycol esters.
• Other surfactants include ascorbyl-6 palmitate (Roche Vitamins. Nutley NJ), stearylamine. and sucrose fatty acid esters (Mitsubishi Chemicals).
• Custom surfactants include those compounds with polar water-loving heads and hydrophobic tails, such as a vitamin E derivative comprising a peptide bonded polyglutamate attached to the ring hydroxyl and pegylated phytosterol. Other peptides may be bonded to vitamin E as well.
• pegylated phospholipids are useful surfactants. Examples of pegylated phospholipids include PEG 2000 or PEG 5000 analogs of phosphatidylethanolamine where the fatty acyl chains contain C 6 - C 24 fatty acids which can be saturated, unsaturated, mixtures thereof.
• Hydrophile-lipophile balance An empirical formula used to index surfactants. Its value varies from 1 - 45 and in the case of non-ionic surfactants from about 1 - 20. In general for lipophilic surfactants the HLB is less than 10 and for hydrophilic ones the HLB is greater than 10.
• Biocompatible Capable of performing functions within or upon a living organism in an acceptable manner, without undue toxicity or physiological or pharmacological effects.
• Substantially free of anv monohydric alcohol A composition having a monohydric alcohol concentration less than about 1.0% (w/v) monohydric alcohol.
• monohydric alcohol is an alcohol containing one hydroxyl group, such as but not limited to ethanol. butanol, isopropanol.
• polyhydric alcohol or polyol is an alcohol containing two or more hydroxyl groups, such as but not limited to, ethylene glycol, propylene glycol or polyethylene glycol (PEG). PEG is also referred to as "polyglycof with ethylene glycol as a polymerized unit.
• Emulsion A colloidal dispersion of two immiscible liquids in the form of droplets, whose diameter, in general, are between 0J and 3.0 microns and which is typically optically opaque, unless the dispersed and continuous phases are refractive index matched.
• Such systems possess a finite stability, generally defined by the application or relevant reference system, which may be enhanced by the addition of amphiphilic molecules or viscosity enhancers.
• Microemulsion A thermodynamically stable isotropically clear dispersion of two immiscible liquids, such as oil and water, stabilized by an interfacial film of surfactant molecules.
• the microemulsion has a mean droplet diameter of less than 200 nm, in general between 10-50 nm.
• mixtures of oil(s) and non-ionic surfactant(s) form clear and isotropic solutions that are known as self-emulsifying drug delivery systems (SEDDS) and have successfully been used to improve lipophilic drug dissolution and oral abso ⁇ tion.
• SEDDS self-emulsifying drug delivery systems
• Pegylated Pegylated or ethoxylated means polyethylene glycol subunits attached to a given compound via a chemical linkage.
• Aqueous Medium A water-containing liquid which can contain pharmaceutically acceptable additives such as acidifying, alkalizing, buffering, chelating, complexing and solubilizing agents, antioxidants and antimicrobial preservatives, humectants, suspending and/or viscosity modifying agents, tonicity and wetting or other biocompatible materials.
• pharmaceutically acceptable additives such as acidifying, alkalizing, buffering, chelating, complexing and solubilizing agents, antioxidants and antimicrobial preservatives, humectants, suspending and/or viscosity modifying agents, tonicity and wetting or other biocompatible materials.
• Therapeutic Agent Any compound natural of synthetic which has a biological activity, is soluble in the oil phase and has an octanol-buffer partition coefficient (Log P) of at least 2 to ensure that the therapeutic agent is preferentially dissolved in the oil phase rather than the aqueous phase.
• This includes peptides, non-peptides and nucleotides. Hydrophobic derivatives of water soluble molecules such as fatty acid and lipid conjugates/prodrugs are within the scope of therapeutic agent.
• Chemotherapeutic Any natural or synthetic molecule which is effective against one or more forms of cancer, and particularly those molecules which are slightly or completely lipophilic or which can be modified to be lipophilic. This definition includes molecules which by their mechanism of action are cytotoxic (anti-cancer agents), those which stimulate the immune system (immune stimulators) and modulators of angiogenesis. The outcome in either case is the slowing of the growth of cancer cells.
• Chemotherapeutics include paclitaxel and related molecules collectively termed taxoids, taxines or taxanes. The structure of paclitaxel is shown in the figure below (Scheme VI).
• taxoids include various modifications and attachments to the basic ring structure (taxoid nucleus) as may be shown to be efficacious for reducing cancer cell growth and to partition into the oil (lipid phase) and which can be constructed by organic chemical techniques known to those skilled in the art. These include but are not limited to benzoate derivatives of paclitaxel such as 2-debenzoyl-2-aroyl and C-2-acetoxy-C-4- benzoate paclitaxel, 7-deocytaxol. C-4 aziridine paclitaxel, particularly the methyl carbonate derivative of paclitaxel.
• BMS-188797 also known as the BMS-188797 as well as various paclitaxel conjugates with natural and synthetic polymers, particularly with fatty acids, phospholipids, and glycerides and 1.2 - diacyloxypropane-3-amine.
• Docetaxel (Taxotere) is also a preferred taxane.
• paclitaxel also included within the scope of the present invention are natural products that share structural similarities with paclitaxel i.e. they inco ⁇ orate a common pharmacophore proposed for microtubule-stabilizing agents.
• These compounds include but are not limited to epothilone A and B. discodermolide, nonataxel and eleutherobin (Chem. Eng. News 1999, 77 (17) : 35-36)
• Chemotherapeutics include podophyllotoxins and their derivatives and analogues.
• the core ring structure of these molecules is shown in the following figure (Scheme VIII):
• chemotherapeutics useful in this invention are camptothecins, the common ring structure of which is shown in the following figure, including any derivatives and modifications to this basic structure which retain efficacy and preserve the lipophilic character of the molecule shown below (Scheme IX).
• chemotherapeutics useful in this invention are the lipophilic anthracyclines, the basic ring structure of which is shown in the following figure (Scheme X):
• Suitable lipophilic modifications of Scheme X include substitutions at the ring hydroxyl group or sugar amino group.
• chemotherapeutics are compounds which are lipophilic or can be made lipophilic by molecular chemosynthetic modifications well known to those skilled in the art, for example by combinatorial chemistry and by molecular modelling, and are drawn from the following list: Taxotere, Amonafide. Illudin S. 6-hydroxymethylacylfulvene Bryostatin 1, 26- succinylbryostatin 1 , Palmitoyl Rhizoxin. DUP 941. Mitomycin B, Mitomycin C, Penclomedine. Interferon ⁇ 2b.
• Cisplatin hydrophobic complexes such as 2-hydrazino-4.5-dihydro-lH-imidazole with platinum chloride and 5-hydrazino-3,4-dihydro-2H-pyrrole with platinum chloride, vitamin A, vitamin E and its derivatives, particularly tocopherol succinate.
• adriamycin epirubicin
• aclarubicin Bisantrene (bis(2-imidazolen-2- ylhydrazone)-9, 10-anthracenedicarboxaldehyde, mitoxantrone, methotrexate, edatrexate, muramyl tripeptide, muramyl dipeptide, lipopolysaccharides, 9-b-d- arabinofuranosyladenine (“vidarabine”) and its 2-fluoro derivative, resveratrol, retinoic acid and retinol, Carotenoids, and tamoxifen.
• bisantrene bis(2-imidazolen-2- ylhydrazone)-9, 10-anthracenedicarboxaldehyde, mitoxantrone, methotrexate, edatrexate, muramyl tripeptide, muramyl dipeptide, lipopolysaccharides, 9-b-d- arab
• Decarbazine Lonidamine, Piroxantrone, Anthrapyrazoles, Etoposide, Camptothecin, 9-aminocamptothecin, 9-nitrocamptothecin, camptothecin-11 ("Irinotecan'), Topotecan, Bleomycin, the Vinca alkaloids and their analogs [Vincristine, Vinorelbine, Vindesine, Vintripol, Vinxaltine, Ancitabine], 6- aminochrysene, and navelbine.
• microtubule targeting agents bind to a protein called tubulin and thus prevent microtubule polymerization.
• Representative microtubule binding agents include epothilones, elutherobin and discodermolide.
• valproic acid tacrolimus, rapamycin, clarithromycin, erythromycin, neomycin, bacitracin, thyrotropin, somatostatin.
• testosterone progesterone, cortisone, polyketides as a class, quinolones as a class, ciprofloxacin, benzodiazepines as a class, diazepam. calcitriol, clozapine, androcephin.
• the present invention is directed to pharmaceutical compositions in the form of emulsions, micellar solutions or self-emulsifying drug delivery systems that are substantially free of ethanol solvent.
• the therapeutic agents of the compositions of this invention can initially be solubilized in a co-solvent.
• a co-solvent In the case when ethanol is used as a processing solvent during the preparation of the oil phase, the ethanol is removed and a substantially ethanol-free composition is formed.
• the ethanol concentration is less than 1% (w/v), preferably less than 0.5%, and most preferably less than 0.3%.
• the therapeutic agents can also be solubilized in methanol. propanol, chloroform, isopropanol, butanol and pentanol. These solvents are also removed prior to use.
• the therapeutic agents of the compositions of the invention can initially be solubilized in non-volatile co-solvents such as benzyl alcohol, benzyl benzoate. dimethylsulfoxide (DMSO), dimethylamide
• NMP polyvinylpyrrolidone
• a major advantage/improvement of using PEG-400 to solubilize therapeutic agents rather than alcohols such as ethanol is that a volatile solvent does not have to be removed or diluted prior to administration of the therapeutic agent.
• the final polyethylene glycol levels in the emulsion can be varied from 1-50%, preferably from 1-25% and more preferably from 1-10% (w/w).
• Suitable polyethylene glycol solvents are those with an average molecular weight between 200 and 600 preferably between 300 and 400 (Table 1).
• high molecular weight PEGs 1,000-10,000
• Solubilization of the therapeutic agents of the invention in polyethylene glycol or other non-volatile co-solvents avoids the necessity of solubilizing the therapeutic agents of the invention in monohydric alcohols such as ethanol or other volatile solvents.
• Use of polyethylene glycol or N-methyl-2- pyrrolidone eliminates the need to remove the solvent prior to use of the emulsions therapeutically.
• the final polyethylene glycol levels in the emulsion can be varied from
• compositions of the invention contain tocopherols (preferably ⁇ - tocopherol) as a carrier for therapeutic drugs, which can be administered to animals or humans via intravascular. oral, intramuscular, cutaneous and subcutaneous routes.
• the emulsions can be given by any of the following routes, among others: intraabdominal, intraarterial, intraarticular, intracapsular, intracervical, intracranial, intraductal. intradural, intralesional, intralocular, intralumbar. intramural, intraocular, intraoperative. intraparietal, intraperitoneal, intrapleural, intrapulmonary, intraspinal, intrathoracic, intratracheal. intratympanic, intrauterine. and intraventricular or transdermal.
• the emulsions of the present invention can be nebulized using suitable aerosol propellants that are known in the art for pulmonary delivery of lipophilic compounds.
• the invention is directed to the use of tocopherols as the hydrophobic dispersed phase of emulsions containing water insoluble, poorly water soluble therapeutic agents, water soluble ones that have been modified to be less water soluble or mixtures thereof.
• ⁇ -tocopherol is employed.
• vitamin E ⁇ -tocopherol is not a typical lipid oil. It has a higher polarity than most lipid oils, particularly triglycerides. and is not saponifiable. It has practically no solubility in water.
• the invention is a tocopherol emulsion in the form of a self-emulsifying system where the system is to be used for the oral administration of water insoluble (or poorly water soluble or water soluble agents modified to be less water soluble or mixtures thereof) drugs where that is desired.
• an oil phase with surfactant and drug or drug mixture is encapsulated into a soft or hard gelatin capsule.
• Suitable solidification agents with melting points in the range of 40 to 60 °C such as high molecular weight polyethylene glycols (MW > 1000) and glycerides such as those available under the tradename Gelucire (Gattefose Co ⁇ .
• the invention comprises microemulsions containing one or more tocols, preferably ⁇ -tocopherol.
• Microemulsions refer to a subclass of emulsions where the emulsion suspension is essentially clear and indefinitely stable by virtue of the extremely small size of the oil/drug microaggregates dispersed therein.
• PEGylated vitamin E is used as a primary surfactant in emulsions of vitamin E.
• PEGylated vitamin E is utilized as a primary surfactant, a stabilizer and also as a supplementary solvent in emulsions of vitamin E.
• Polyethylene glycol (PEG) is also useful as a co- solvent in the emulsions of this invention. Of particular use is polyethylene glycol 200. 300, 400 or mixtures thereof.
• the concentration of ⁇ -tocopherol and/or other tocols in the emulsions of this invention can be from about 1 to about 20 % w/w.
• the ratio of tocopherol to TPGS is optimally from about 1 : 1 to about 20: 1.
• the emulsions of the invention may further include surfactants such as ascorbyl-6-palmitate, stearylamine. pegylated phospholipids, sucrose fatty acid esters, various tocol derivatives and nonionic, synthetic surfactant mixtures, such as polyoxyptopylene-polyoxyethylene glycol nonionic block copolymer.
• the emulsions of the invention can comprise an aqueous medium.
• the aqueous phase generally has an osmolality of approximately 300 mOsm and may include sodium chloride, sorbitol, mannitol, polyethylene glycol, propylene glycol albumin, polypep and mixtures thereof.
• This medium can also contain various additives to assist in stabilizing the emulsion or in rendering the formulation biocompatible.
• Acceptable additives include acidifying agents, alkalizing agents, antimicrobial preservatives, antioxidants. buffering agents, chelating agents, suspending and/or viscosity-increasing agents, and tonicity agents.
• agents to control the pH, tonicity, and increase viscosity are included.
• a tonicity of at least 250 mOsm is achieved with an agent which also increases viscosity, such as sorbitol or sucrose.
• the emulsions of the invention for intravenous injection have a particle size (mean droplet diameter) of 10 to 500 nm, preferably 10 to 200 nm and most preferably 10 to 100 nm.
• particle size mean droplet diameter
• the spleen and liver will eliminate particles greater than 500 mn in size.
• a preferred form of the invention includes paclitaxel, a very water- insoluble cytotoxin used in the treatment of uterine cancer and other carcinomas.
• An emulsion composition of the present invention comprises a solution of vitamin E containing paclitaxel at a concentration of up to 20 mg/mL. four times that currently available by prescription, and a biocompatible surfactant such that the emulsion microdroplets are less than 0.2 microns and are terminally sterilizable by filtration.
• Preferred injectable compositions contain: OJ-1.0 % paclitaxel (1-10 mg/ml); 1-10% PEG 400; 1-20 % Vitamin E; 1-10 % TPGS and 0.5-2.5% Poloxamer 407.
• Another preferred composition contains: 1.0 % paclitaxel (10 mg/ml), 6% PEG400. 8% Vitamin E. 5% TPGS. 1% Pluronic F127 and 80% aqueous solution.
• Preferred formulations for self-emulsifying systems are as follows: 0.1- 20% paclitaxel, 10-90% Vitamin E. 10-90% PEG 400 or N-methyl-2- pyrrolidone. 5-50% TPGS, 5-50% a secondary hydrophilic surfactant, such as
• the oil phase can optionally contain polyvinylpyrrolidone, glycerol and propylene glycol esters such as mono-/di- /triglycerides and mono-diesters of propylene glycol.
• high molecular weight PEGs 1,000-10.000 MW
• high melting point glycerol esters can be included to provide the formulation with semisolid consistency.
• a further embodiment of the invention is a method of treating carcinomas comprising the parenteral administration of a bolus dose of paclitaxel in vitamin E emulsion with or without PEGylated vitamin E by intravenous injection once daily or every second day over a therapeutic course of several weeks.
• Such method can be used for the treatment of carcinomas of the breast, lung, skin and uterus.
• Example 1 Dissolution of paclitaxel in ⁇ -tocopherol ⁇ -Tocopherol was obtained from Sigma Chemical Company (St Louis MO) in the form of a synthetic dl- ⁇ -tocopherol of 95% purity prepared from phytol. The oil was amber in color and very viscous. Paclitaxel was purchased from Hauser Chemical Research (Boulder CO), and was 99.9% purity by HPLC. Paclitaxel 200 mg was dissolved in 6 mL of dry absolute ethanol (Spectrum Chemical Manufacturing Co ⁇ . Gardenia CA) and added to 1 gm ⁇ - tocopherol. The ethanol was then removed by vacuum at 42OC until the residue was brought to constant weight.
• Sigma Chemical Company Sigma Chemical Company (St Louis MO) in the form of a synthetic dl- ⁇ -tocopherol of 95% purity prepared from phytol. The oil was amber in color and very viscous. Paclitaxel was purchased from Hauser Chemical Research (Boulder CO), and was 99.9% purity by HPLC. Paclitaxe
• Paclitaxel 2 gm in 10 gm of ⁇ -tocopherol, prepared as described in Example 1 was emulsified with ascorbyl palmitate as the triethanolamine salt by the following method.
• a solution consisting of ascorbic acid 20 mM was buffered to pH 6.8 with triethanolamine as the free base to form 2x buffer.
• 50 mL of the 2x buffer was placed in a Waring blender.
• 0.5 gm of ascorbyl-6- palmitate (Roche Vitamins and Fine Chemicals, Nutley NJ), an anionic surfactant, was added and the solution blended at high speed for 2 min at 40°C under argon.
• the ⁇ -tocopherol containing paclitaxel was then added into the blender with the surfactant and buffer. Mixing was continued under argon until a coarse, milky, pre-emulsion was obtained, approximately after 1 min at 40°C. Water for injection was then added, bringing the final volume to 100 mL.
• the pre-emulsion was transferred to the feed vessel of a Microfluidizer Model HOY (Microfluidics Inc, Newton MA).
• the unit was immersed in a bath to maintain a process temperature of approximately 60°C during homogenization, and was flushed with argon before use.
• the emulsion was passed through the homogenizer in continuous re-cycle for 10 minutes at a pressure gradient of about 18 kpsi across the interaction head.
• the flow rate was about 300 mL/min, indicating that about 25 passes through the homogenizer resulted.
• the resultant paclitaxel emulsion in an ⁇ -tocopherol vehicle was bottled in amber vials under argon and stored with refrigeration at 7°C and 25°C. Samples were taken at discrete time intervals for particle sizing and chemical analysis.
• TPGS vitamin-E polyoxyethyleneglycol-lOOO-succinate, obtained from Eastman Chemical Co., Kingsport TN
• water PEGylated vitamin E
• TPGS PEGylated vitamin E
• ⁇ -tocopherol PEGylated vitamin E
• ⁇ -tocopherol PEGylated vitamin E
• Mixtures were miscible at all concentrations. Water was then added to each mixture in such a way that the final water concentration was increased stepwise from zero to 97.5%.
• observations were made of the phase behavior of the mixture. As appropriate, mixing was performed by vortexing and sonication. and the mixture was heated or centrifuged to assess its phase composition.
• a broad area of biphasic o/w emulsions suitable for parenteral administration was found at water concentrations above 80%.
• the emulsions formed were milky white, free flowing liquids that contained disperse ⁇ -tocopherol microparticles stabilized by non-ionic surfactant.
• microemulsions potentially suitable as drug carriers were observed at TPGS to oil ratios above about 1 :1.
• a broad area containing transparent gels (reverse emulsions) was noted. Separating the two areas (high and low water content) is an area composed of opaque, soap-like liquid crystals.
• Phase diagrams of ⁇ -tocopherol with surfactant combinations for example TPGS with a nonionic. anionic or cationic co-surfactant (for example glutamyl stearate, ascorbyl palmitate or Pluronic F-68), or drug can be prepared in a similar manner.
• a nonionic. anionic or cationic co-surfactant for example glutamyl stearate, ascorbyl palmitate or Pluronic F-68
• drug can be prepared in a similar manner.
• paclitaxel 1.0 gm% ⁇ -tocopherol 3.0 gm%
• Pre-warmed aqueous solution containing a biocompatible osmolyte and buffer were added with gentle mixing and a white milk formed immediately. This mixture was further improved by gentle rotation for 10 minutes with continuous warming at 40-45 °C. This pre-mixture at about pH 7 was then further emulsified as described below.
• the pre-mixture at 40-45°C was homogenized in an Avestin C5 homogenizer (Avestin, Ottawa Canada) at 26 Kpsi for 12 minutes at 44°C.
• the resultant mixture contained microparticles of ⁇ -tocopherol with a mean size of about 200 nm. Further pH adjustment was made with an alkaline 1 M solution of triethanolamine (Spectrum Quality Products).
• TPGS Triethanolamine
• all operations were performed above 40°C and care was taken to avoid exposure of the solutions to cold air by covering all vessels containing the mixture.
• less than 2% TPGS should generally be dissolved in ⁇ - tocopherol oil before pre-emulsification.
• the solution gels at concentrations of TPGS higher than 2%.
• Example 4 After emulsification, the formulation of Example 4 was analyzed for paclitaxel on a Phenosphere CN column (5 microns, 150 x 4.6 mm). The mobile phase consisted of a methanol/water gradient, with a flow rate of 1.0 mL/min. A UV detector set at 230 nm was used to detect and quantitate paclitaxel. A single peak was detected ( Figure 2), which had a retention time and mass spectrogram consistent with native reference paclitaxel obtained from Hauser Chemical (Boulder CO). Chemical stability of the emulsion of example 4 was examined by
• paclitaxel 10 mg/ml for intravenous drug delivery having the following composition, was prepared as described in Example 4.
• paclitaxel 1.0 gm% ⁇ -tocopherol 3.0 gm%
• Paclitaxel Emulsion Formulation QWB A second emulsion of paclitaxel 10 mg/ml for intravenous drug delivery, having the following composition, was prepared as described in Example 4. paclitaxel 1.0 gm% ⁇ -tocopherol 3.0 gm%
• Solutol HS-15 is a product of BASF Co ⁇ , Mount Olive NJ
• a third emulsion formulation of paclitaxel 10 mg/ml was prepared as follows using Poloxamer 407 (BASF Co ⁇ , Parsippany NJ) as a co-surfactant.
• paclitaxel 1.0 gm% ⁇ -tocopherol 6.0 gm%
• Poloxamer 407 1.0 gm% Sorbitol 4.0 gm%
• 1.0 gm Poloxamer 407 and 1.0 gm paclitaxel were dissolved in 6.0 gm ⁇ -tocopherol with ethanol 10 volumes and gentle heating.
• an aqueous buffer was prepared by dissolving 3.0 gm TPGS and 4.0 gm sorbitol in a final volume of 90 mL water for injection. Both oil and water solutions were warmed to 45 °C and mixed with sonication to make a pre-emulsion. A vacuum was used to remove excess air from the pre-emulsion before homogenization.
• Homogenization was performed in an Avestin C5 as already described.
• the pressure differential across the homogenization valve was 25 kpsi and the temperature of the feed was 42°-45°C.
• a chiller was used to ensure that the product exiting the homogenizer did not exceed a temperature of 50°C.
• Flow rates of 50 mL/min were obtained during homogenization. After about 20 passes in a recycling mode, the emulsion became more translucent. Homogenization was continued for 20 min. Samples were collected and sealed in vials as described before. A fine ⁇ -tocopherol emulsion for intravenous delivery of paclitaxel was obtained. The mean particle diameter of the emulsion was 77 nm. Following sterile filtration through a 0.22 micron Durapore filter
• a fifth emulsion of ⁇ -tocopherol for intravenous administration of paclitaxel was prepared as follows: paclitaxel 0.5 gm% ⁇ -tocopherol 6.0 gm%
• Poloxamer 407 1.5 gm%
• Synthetic ⁇ -tocopherol USP-FCC obtained from Roche Vitamins (Nutley, NJ) was used in this formation.
• Polyethyleneglycol 200 (PEG-200) was obtained from Sigma Chemical Co.
• mice were prepared as follows. A stock solution of paclitaxel in TPGS was made up by dissolving 90 mg paclitaxel in 1.0 gm TPGS at 45°C with ethanol. which was then removed under vacuum. Serial dilutions were then prepared by diluting the paclitaxel stock with additional TPGS to obtain paclitaxel in TPGS at concentrations of 0J, 1, 5, 10, 25, 50, 75 and 90 mg/mL. Using fresh test tubes, 100 mg of each paclitaxel concentration in TPGS was dissolved in 0.9 mL water.
• micellar solutions in water were obtained corresponding to final paclitaxel concentrations of 0.01 , 0.1, 0.5, 1.0, 2.5, 5.0, 7.5 and 9.0 mg/mL.
• micellar solutions of paclitaxel in TPGS containing up to 2.5 mg/mL paclitaxel were stable for at least 24 hr whereas those at 5.0, 7.5 and 9.0 mg/mL were unstable and drug crystals formed rapidly and irreversibly. These observations imply that paclitaxel remains solubilized only in the presence of an ⁇ -tocopherol-rich domain within the emulsion particles. Thus, an optimum ratio of ⁇ -tocopherol to TPGS is needed in order to produce emulsions in which higher concentrations of paclitaxel can be stabilized. When adjusted to the proper tonicity and pH, micellar solutions have utility for slow IV drip administration of paclitaxel to cancer patients, although the AUC is expected to be low.
• TPGS in ⁇ -tocopherol emulsions
• TPGS has its own affinity for paclitaxel, probably by virtue of the ⁇ -tocopherol that makes up the hydrophobic portion of its molecular structure.
• interfacial tension of TPGS in water with ⁇ - tocopherol is about 10 dynes/cm, sufficient to emulsify free ⁇ -tocopherol, especially when used with a co-surfactant.
• polyoxyethylated surfactants such as TPGS
• TPGS polyoxyethylated surfactants
• TPGS polyoxyethylated surfactants
• micellar solutions When adjusted to the proper tonicity and pH, micellar solutions have utility for slow IV drip administration of paclitaxel to cancer patients, although the AUC is expected to be low.
• pegylated surfactants for example Triton X-100, PEG 25 propylene glycol stearate, Brij 35 (Sigma Chemical Co), Myrj 45, 52 and
• Tween 80 and Myrj 52 were characterized as coarse, containing rod-shaped particles up to several microns in length, consistent with crystals of paclitaxel. Unlike the TPGS emulsion, which passed readily through a 0.22 micron filter with less than 1 % loss of drug, the Tween and Myrj emulsions were unfilterable because of the presence of this crystalline drug material.
• the drug has good solubility in TPGS, up to about 100 mg/mL. Most likely it is the strength of the affinity of paclitaxel benzyl side chains with the planar structure of the ⁇ - tocopherol phenolic ring in the TPGS molecule that stabilizes the complex of drug and carrier.
• the succinate linker between the ⁇ -tocopherol and PEG tail is a novel feature of this molecule that distinguishes its structure from other PEGvlated surfactants tested.
• Poloxamer 407 2.5 gm% Ascorbyl Palmitate 0.3 gm%
• ⁇ -tocopherol emulsion was prepared using Poloxamer 407 (BASF) as the primary surfactant.
• BASF Poloxamer 407
• the white milky pre-mixture was homogenized with continuous recycling for 10 minutes at 25 Kpsi in a C5 homogenizer (Avestin, Ottawa Canada) with a feed temperature of 45°C and a chiller loop for the product out set at 15°C.
• a fine, sterile filterable emulsion of ⁇ -tocopherol microparticles resulted.
• this formulation was made with paclitaxel. precipitation of the paclitaxel was noted following overnight storage in the refrigerator, again underlying the superior utility of TPGS as the principle surfactant.
• Maltrin Ml 00 (Grain Processing Co ⁇ oration, Muscatine IA) was added as a 2x stock in water to the emulsion of Example 14. Aliquots were then frozen in a shell freezer and lyophilized under vacuum. On reconstitution with water, a fine emulsion was recovered.
• Lyophilized formulations have utility where the indefinite shelf life of a lyophilized preparation is preferred. Lyophilizable formulations containing other saccharides. such as mannitol, albumin or PolyPep from Sigma Chemicals. St. Louis, Mo. can also be prepared.
• Emulsions were prepared having paclitaxel concentrations of 6 mg/mL (QWA) and 7 mg/mL (QWB).
• Taxol contains 6 mg/ml of paclitaxel dissolved in ethanokcremophore EL 1:1
• TTD maximal tolerable dose
• Table 2 The FDA-approved formulation of Taxol ® causes death in mice at bolus intravenous doses of 10 mg/kg, a finding repeated in our hands. In the rat, Taxol ® was uniformly fatal at all dilutions and dose regimes we tested. In contrast, the composition of Example 6 was well tolerated in rats, and is even improved over Taxotere, a less toxic paclitaxel analogue commercially marketed by Rhone-Poulenc Rorer.
• the emulsion is behaving as a slow-release depot for the drug as suggested from the in vitro release data in Example 16.
• Example 6 The paclitaxel emulsion of Example 6 was also evaluated for efficacy against staged B 16 melanoma tumors in nude mice and the data is shown in Table 3. Once again, the marketed product Taxol* was used as a reference formulation. Tumor cells were administered subcutaneously and therapy started by a tail vein injection at day 4 post-tumor administration at the indicated dosing schedule. Efficacy was expressed as percent increase in life-span (% ILS).
• Taxol ® was again used as a reference formulation. Methods essentially identical to those of Example 18 were used. The data from this study is summarized in Table 3. Efficacy was expressed as: a) percent tumor growth inhibition (% T/C, where T and C stand for treated and control animals, respectively); b) tumor growth delay value (T-C), and c) log cell kill which is defined as the ratio of the
• the latter parameter for this particular tumor model was calculated to be 1.75 days.
• all measures of efficacy tumor growth inhibition, tumor growth delay value and log cell kill demonstrate superior efficacy of ⁇ - tocopherol emulsions as a drug deliver)' vehicle over Taxol*, particularly when the emulsions were dosed every four days at 70 mg/kg. As explained in Example 16, this increased efficacy is likely a result of improved drug biocompatibility and/or sustained release.
• C mean survival of control according to the NCI standards an ILS value greater than 50% indicates significant anti-tumor activity.
• Paclitaxel 50 mg/ml was prepared in ⁇ -tocopherol by the method described in Example 1. Tagat TO 20%> (w/w) was added. The resultant mixture was clear, viscous and amber in color. A 100 mg quantity of the oily mixture was transferred to a test tube. On addition of 1 mL of water, with vortex mixing, a fine emulsion resulted.
• Example 22 Self-emulsifying formulation of paclitaxel
• Paclitaxel 50 mg/ml was prepared in ⁇ -tocopherol by the method described in Example 1. After removal of the ethanol under vacuum, 20% TPGS and 10% polyoxyethyleneglycol 200 (Sigma Chemical Co) were added by weight. A demonstration of the self-emulsification ability of this system was then performed by adding 20 mL of deionized water to 100 mg of the oily mixture at 37°C. Upon gentle mixing, a white, thin emulsion formed, consisting of fine emulsion particles demonstrated with the Malvern Mastersizer (Malvern Instruments. Worcester MA) to have a mean size of 2 microns, and a cumulative distribution 90%> of which was less than 10 microns. Example 23.
• Malvern Mastersizer Malvern Mastersizer
• Etoposide 4 mg (Sigma Chemical Co) was dissolved in the following surfactant-oil mixture: Etoposide 4 mg ⁇ -tocopherol 300 mg
• a pre-emulsion was formed by adding 4.5 mL of water containing 4% sorbitol and 100 mg TPGS at 45°C with sonication.
• the particle size was further reduced by processing in an Emulsiflex 1000 (Avestin, Ottawa Canada).
• the body of the Emulsiflex 1000 was fitted with a pair of 5 mL syringes and the entire apparatus heated to 45°C before use.
• the 5 mL of emulsion was then passed through it by hand approximately 10 times. A free flowing, practical emulsion of etoposide in an ⁇ -tocopherol vehicle resulted.
• solubilized form of etoposide in ⁇ -tocopherol can also be used as an oral dosage form by adaption of the methods of the preceding examples.
• Ibuprofen Dissolution of Ibuprofen or Griseofulvin in ⁇ -tocopherol Ibuprofen is a pain-killer, and may be administered by injection when required if there is danger that the drug will irritate the stomach.
• the following solution of ibuprofen in ⁇ -tocopherol may be emulsified for intravenous administration.
• Ibuprofen (Sigma Chemicals). 12 mg. crystalline, dissolved without solvent in ⁇ -tocopherol, 120 mg, by gentle heating.
• the resultant 10 % solution of ibuprofen in vitamin E can be emulsified by the method s described in Examples 4, 6, 7, 8 or 22.
• An antifungal compound, griseofulvin, 12 mg was first dissolved in 3 mL of anhydrous ethanol; ⁇ -tocopherol was then added. 180 mg, and the ethanol was removed with gentle heating under vacuum.
• the resultant solution of griseofulvin in ⁇ -tocopherol is clear and can be emulsified by the methods described in Examples 4, 6, 7, 8 or 22.
• Vitamin E succinate has been suggested as a therapeutic for the treatment of lymphomas and leukemias and for the chemoprevention of cancer.
• Sucrose ester SI 170 is a product of Mitsubishi Kagaku Foods Co ⁇ , Tokyo Japan.
• Emulsions inco ⁇ orating other surfactants such as pluronics, and TPGS along with ⁇ - tocopherol and ⁇ -tocopherol succinate can be prepared in a similar manner with and without a therapeutic agent.
• ⁇ -Tocopherol 8 gm and vitamin E succinate 0.8 gm were dissolved together in ethanol in a round bottom flask.
• the alkaline buffer consisting of 2% glycerol, 10 mM triethanolamine. and 0.5 gm % sucrose ester SI 170.
• the pressure differential across the interaction head was 25 to 26 kpsi.
• pH was carefully monitored, and adjusted as required to pH 7.0. Care was taken to exclude oxygen during the process. A fine white emulsion resulted.
• ⁇ -tocopherol in commercially available esters tocopherol- acetate. -succinate, -nicotinate. -phosphate and TPGS were either provided by the vendor or determined by HPLC.
• the concentration of free ⁇ -tocopherol in these solutions is less than 1.0%. generally less than 0.5%.
• Example 27 Resveratrol emulsion formulation
• Resveratrol is a cancer chemopreventative first discovered as an extract of grape skins. It has been proposed as a dietary supplement.
• Resveratrol was obtained from Sigma Chemical Co. While it dissolved poorly in ethanol, upon addition of 10 mg resveratrol, 100 mg of ⁇ -tocopherol, 100 mg TPGS and ethanol, a clear solution formed rapidly. Upon removal of the ethanol, a clear amber oil remained.
• the oily solution of resveratrol can be formulated as a self-emulsifying system for oral delivery by the various methods of the preceding examples.
• Muramyl dipeptides are derived from mycobacteria and are potent immunostimulants representative of the class of muramyl peptides, mycolic acid and lipopolysaccharides. They have use, for example, in the treatment of cancer, by stimulating the immune system to target and remove the cancer, particularly in connection with anti-cancer vaccines. More recently, muroctasin, a synthetic analog, has been proposed to reduce non-specific side effects of the bacterial wall extracts.
• N-acetylmuramyl-6-O-steroyl- 1 -alanyl-d-isoglutamine was purchased from Sigma Chemical Co. and 10 mg was dissolved in 100 mg ⁇ -tocopherol and 80 mg TPGS. Ethanol was used as a co-solvent to aid in dissolution of the dipeptide. but was removed by evaporation under vacuum, leaving a clear solution in ⁇ -tocopherol and surfactant.
• This oil solution of the drug can be emulsified for parenteral administration by the various methods of the preceding examples.
• Example 29 Alcohol-containing emulsion
• paclitaxel was dissolved in a ⁇ -tocopherol and TPGS with ethanol, which was then removed under vacuum. By dry weight, residual ethanol was less than 3 mg (0.3% w/w). Fresh anhydrous ethanol 0J25 gm was then added back to the formulation. After mixing, the suitability of the formulation for oral administration, as in a gelatin capsule, was simulated by the following experiment. An aliquot of 100 mg of the free-flowing oil was added to 20 mL of water at 37°C and mixed gently with a vortex mixer. A fine emulsion resulted. But after twenty minutes, microscopy revealed the growth of large numbers of crystals in rosettes, characteristic of paclitaxel precipitation.
• paclitaxel was dissolved in ⁇ -tocopherol and TPGS with ethanol, which was then removed under vacuum. By dry weight, residual ethanol was less than 2 mg (0.5% w/w). Fresh anhydrous ethanol 0J00 gm and n-butanol 0.500 gm was then added back to the formulation. A clear oil resulted.
• the injection concentrate was tested for biocompatibility in administration by standard pharmaceutical practice of admixture with saline. About 200 mg of the oil was placed into 20 mL of saline and mixed. Large flakes of insoluble material developed immediately and the greatest amount of material formed dense deposits on the walls of the test tube.
• Lecithin is not in this class, although it could be used as a co-surfactant.
• typical o/w emulsions of triglycerides are made with surfactants of HLB between 7 and 12, demonstrating that ⁇ -tocopherol emulsions are a unique class by virtue of the polarity and extreme hydrophobicity of the ⁇ -tocopherol. factors that also favor the solubility of lipophilic and slightly polar lipophilic drugs in ⁇ - tocopherol. See Emulsions: Theory and Practice, 2nd Ed. p.248 (1985).
• Example 31 Various formulations useful in the invention (Table 5) are prepared as follows:
• step 1 The aqueous phase (step 1) was heated to 45°C and mixed at medium shear (laboratory mixing motor) while 45°C oil phase (step 2 + 3) was poured in over 1 minute. Mixing was continued 2 minutes more to form a crude emulsion.
• Vitamin E 17.176 g was added and mixed (low shear) at 45-50°C until the mixture was visibly homogeneous.
• Step 4 21.8 g of the oil phase produced in Steps 1-4 was added over 1 minute to 79.5 g water while mixing at medium shear (laboratory mixing motor).
• Emulsion was homogenized in an Avestin C5 in continuous recycle mode for 30 minutes at 22 K psi peak stroke pressures From a processing perspective it is advantageous to have all of the surfactants in the oil phase. Both the dispersion of the pre-emulsion and subsequent homogenization are facilitated and potential gellation of high melting point surfactants, such as TPGS. can be avoided.
• a vitamin E emulsion (6.0% vitamin E. 3.5% TPGS, 6.0%, PEG400, 8% Pluronic F- 127) and inco ⁇ orating 2 mg/ml of Etoposide was prepared as follows:
• Etoposide was first dissolved in PEG-300 (10 min at 72°C). TPGS and Vitamin E were then added to the drug solution. Aqueous phase (WFI containing Poloxamer 407) was degassed by boiling prior to use. Pre-emulsion was prepared by adding 5 g of the oil phase to 45 g of water at 45°C. After a 3-min mixing the pre-emulsion was homogenized at 25 Kpsi for 30 min to produce a fine emulsion. The final composition of the emulsion is shown below: Component Composition (%. w/w)
• Poloxamer 407 1.0
• Example 34 Additional paclitaxel emulsions for injection are presented in Table 6.
• compositions of various self-emulsifying emulsions useful in this invention are shown in Table 7.
• Table 7. Self-Emulsifying Emulsions
• Paclitaxel and PEG 400 were heated together at 60-67°C and sti ⁇ ed until the drug was dissolved in PEG (15 min). Then TPGS and Pluronic F127 were added and sti ⁇ ed at 70 °C for 10-15 min to dissolve the surfactant. Finally, Vitamin E ( ⁇ -tocopherol) was added and mixed for 5-10 min at 55°C until the mixture was clear and homogeneous. Upon dilution with an aqueous phase a fine emulsion can be obtained.
• Paclitaxel and PEG 400 were first sti ⁇ ed at 65-75°C for 45 min there TPGS was added and sti ⁇ ing was continued for another 30 min to completely dissolved all three components and produced a clear solution. Finally Solutol HS-15 and Vitamin E were added and mixed for about 5 min at 55°C to obtain a clear homogeneous liquid. Upon dilution with an aqueous phase a fine emulsion can be obtained.
• Example 36 Additional compositions of self-emulsifying emulsions of paclitaxel are shown in Table 8.
• SEFP-3 and SEFP-4 were prepared by first dissolving paclitaxel in solutol HS-
• the particle size of the emulsions upon dilution of SEFP-3 and SEFP-4 was determined as follows: 0.2 ml of SEFP-3 or SEFP-4 was diluted in 100 ml of Phosphate-buffered Saline at 37°C by low shear mixing with a stir bar for 5 minutes. An emulsion was quickly formed, the particle size of which was measured by the Malvern Mastersizer. The volume mean diameter of SEFP-3 and SEFP-4 was found to be, 2.49 and 1.55 ⁇ m, respectively.
• the mean droplet diameter of the resulting emulsion should be less than 10 ⁇ m and preferably less than 5 ⁇ m.
• DMPE-PEG 2000 (Dimyristoyl Phosphatidyl Ethanolamine - Polyethylene Glycol 2000) inco ⁇ orating emulsions were prepared (Table 9).
• Paclitaxel when present, was first dissolved in PEG 400 by low shear mixing at 75°C. The other ingredients were added and briefly mixed (after melting TPGS, and in the case of DMPEG-2, the P 407) to form a clear solution. A vacuum was applied to degas the oil phase prior to emulsification, and the oil phase was brought to 45°C. Water was boiled for 15 minutes to degas, then brought to 45°C also.
• the two phases were mixed at 45°C at low to medium shear to form a pre- emulsion.
• DMPE-PEG-P2 DMPE-PEG-P3 and DMPE- PEGP-4. This was accomplished by simply adding the warm water to the oil phase and swirling by hand with sonication.
• the pre-emulsion for DMPE-PEG2 was prepared by pouring oil phase into water while sti ⁇ ing with a laboratory mixing motor. Pre-emulsions were immediately homogenized in the Avestin C5 homogenizer at 20-22K psi peak stroke pressure to produce fine emulsions with a mean droplet diameters and 99% cumulative distributions of less than 200 nm.
• Table 9 Paclitaxel Emulsions Incorporating a Pegylated Phospholipid
• Formulation D (Table 6) was evaluated for efficacy against B16 melanoma in mice as described in Examples 18 and 19 and the data is summarized in Figure 5. Comparative efficacy data is presented in Table 10.
• T/C Tumor Growth Inhibition (median tumor wt. of treated/median tumor wt. control) x 100
• T-C Tumor Growth Delay value (median time for treatment group (T) and control group tumors (C) to reach a predetermined size (>750 mg)
• Log cell kill (T-C value)/(3.32 x tumor doubling time) tumor doubling time calculated to be 1.75 days.
• Particle size was measured using the Nicomp 370 sub-micron particle size analyzer. As can be seen from the data in Table 11 no significant changes were observed in either the mean droplet diameter or the 99% cumulative distribution of the particles. The latter parameter is often used as an indicator of particle aggregation and growth. In addition, no precipitation or other gross changes were observed during storage. Long term stability is ongoing.
• Emulsions Containing PEG 300 or NMP ⁇ -Tocopherol emulsions containing PEG 300 or NMP (N-Methyl-2- py ⁇ olidone) and inco ⁇ orating 10 mg/ml paclitaxel are shown in Table 13.
• paclitaxel was first dissolved in the solvent (PEG 300 or NMP) with low shear mixing. Heating to 60°C was used with the PEG 300 to speed the dissolution while with the NMP formulation a few minutes at room temperature was sufficient to dissolve the drug. The remaining ingredients (except water) were then added and the mixtures were heated to 60-65 °C with low shear mixing to melt the solid surfactant and produce homogeneous, clear solutions. The solutions were brought to 45°C, then 45°C water was added to them. The resulting mixtures were processed under medium shear to produce a thick, white crude emulsion, very similar in appearance to the pre-emulsion of formulation D (Table 6). These emulsions can further be homogenized at high pressure to produce fine emulsions.
• solvent PEG 300 or NMP
• formulation D (Table 6) was manufactured at a large scale in 2 x 2L sub-lots having the following composition (the shaded area represents the oil and aqueous phase content of the emulsion):
• a large scale (2.5 L) of formulation D in the absence of paclitaxel was prepared as described in Example 41 having the following composition.
• Table 16 shows long-term stability of the scaled up formulation of Example 41 upon a 9-month storage at 4°C or 25°C. It is evident that at least during this storage time, both the mean droplet diameter and the 99% cumulative distribution did not significantly changed from their initial values of about 65 and 150 nm, respectively, and the emulsion remains within specifications.
• Example 41 was evaluated for efficacy against B16 melanoma as described in Examples 18. 19 and 37 and the results are summarized in Table 18.
• QW8184 exhibited superior antitumor activity in mice at doses that included or well exceeded the MTD of Taxol ® but which were well tolerated. Such effects have not been reported with previous injectable emulsions of paclitaxel. MTD is the maximum tolerated dose that is determined from acute toxicity studies.
• the antitumor activity of QW81 84 (Example 41). against the human ovarian tumor xenograft IGROV- 1 using the marketed product Taxol ® as a reference formulation.
• Nude mice were implanted subcutaneously by trocar with fragments of IGROV- 1 human ovarian carcinomas harvested from subcutaneously growing tumors in nude mice hosts. When tumors were approximately 5 x 5 mm in size, the animals were paired matched into treatment and control groups contained 9 ear-tagged tumor-bearing mice per group.
• QW8184 was administered i.v. on a q3dx5, q4dx5, and qdx5 schedule at 20, 40 and 60 mg/kg.
• Taxol ® was administered i.v.
• mice were weighed twice weekly, and tumor measurements were taken by calipers starting Day 1 and converted to mg tumor weight. The experiment was terminated when the control tumors reached approximately lgr and tumors were excised and weighed and the mean tumor weight per group was calculated. The data is summarized in Table 20.
• QW8184 at 20. 40 and 60 mg/kg on a q3dx5 or q4dx5 schedule resulted in nearly 100% tumor growth inhibition at all doses with 2, 8, and 7 and 3, 9, and 8 complete tumor responses, respectively. In comparison, administration of Taxol ® resulted in 3 complete tumor responses on both schedules. On a qdx5 schedule, the antitumor activities of QW8184 and Taxol® were similar. QW8184, however, was better tolerated with no toxic deaths whereas six toxic deaths were noted with Taxol®. QW8184 was highly active against the IGROV-1 human ovarian xenograft model in a dose-dependent fashion, regardless of the dosing schedule and it was better tolerated than
• Taxol ® .
• Example 47 Pharmacokinetic Study The pharmacokinetics of the formulation of Example 41 (Q W8184), in the rat upon a single 10 mg/kg i.v. administration was determined using Taxol ® as a reference formulation. The drug was administered i.v. to male or female rats either as a 3-hr infusion (Taxol ® ) or as a bolus dose (QW8184). Blood samples were collected from 0-72 hrs after dose administration, plasma was prepared by centrifugation and analyzed for paclitaxel concentration using a high performance liquid chromatography (HPLC) method with LC/MS/MS detection. Pharmacokinetic analysis was performed on the mean composite plasma concentration-time profiles using a model independent method. The derived pharmacokinetic parameters are shown in Table 21. The pharmacokinetic parameters determined were as follows:
• T maford time required to reach peak plasma levels (C m-proof)
• AUC 0 . t non-extrapolated area under the plasma concentration-time curve from time zero to time t which is the end of the plasma sample collection
• AUC 0 . K extrapolated area under the plasma concentration-time curve from time zero to infinite
• V H volume of distribution
• V volume of distribution at steadv state
• the following tocotrienol emulsion was prepared containing 5 mg/mL paclitaxel and is suitable for intravenous cancer therapy in mice.
• Gold Tri-E ® tocotrienol concentrate was obtained from Golden Jomalina Food Industries. (Kuala Lumpur. Malaysia). An HPLC assay of the reddish brown oil revealed four major peaks. Our estimate of the ⁇ TE for the oil is -0.3, due in most part to a residual d- ⁇ -tocopherol content of about 20%. It was diluted with 2 parts ⁇ -tocopherol (Sigma Chemicals) to adjust the ⁇ TE to 0J. A drug oil solution was prepared by first dissolving the paclitaxel in PEG-400 with heat and ultrasound. TPGS and the tocotrienol oil were then added. Finally, the poloxamer was added and melted at 72 °C to yield a homogeneous, clear amber oil. The mixture was degassed under vacuum in a rotevap and held at 45 °C until use.
• the aqueous phase consisting of 40 mL of 5 mM citrate TEA buffer, pH 6.8, was brought to 45 °C before addition.
• the resultant mixture was mixed vigorously to loosen any adherent oil on the walls of the flask.
• This suspension was then placed in a feed vessel and processed in a C5 homogenizer (Avestin. Ottawa CA) for 10 min with continuous recycling. Processing conditions were 45 °C feed temperature. 20 kpsi processing pressure, 120 mL/min flow rate.
• a heat exchanger set at 22 °C was placed at the exit port to remove excess heat generated in the homogenizer.
• the temperature in the feed vessel was measured at 44 °C during steady state homogenization. Following processing, the product was collected and cooled to room temperature.
• the emulsion was then terminally sterilized by filtration through a 0.2 um filter and had a mean particle size of -70 nm when measured on a Nicomp 370 photon co ⁇ elation spectrophotometer. For convenience in handling. Gentamycin 15 ug/mL was added as a preservative.
• the use of the tocotrienol rich fraction rendered the emulsion substantially more easy to process than a comparable emulsion made with d.l- ⁇ -tocopherol (Roche Vitamins) as the oil phase.
• Example 49 ⁇ -Tocopherol emulsion of paclitaxel for intravenous administration
• This oil-drug concentrate was then cooled to 45 °C and 850 ⁇ L of warm water was added. Using a microtip sonication horn, a coarse pre-emulsion was prepared. Microscopic examination revealed a dense suspension of particles substantially less than 10 ⁇ m diameter which with further processing in a homogenizer, adjustment of pH and osmotic strength, and terminal sterile filtration, is suitable for parenteral injection.
• Example 50 Emulsion formulations of the methyl carbonate derivative of paclitaxel
• BMS-188797 was first dissolved in ethanol and then a-tocopherol, Myvacet 9- 45 (if present), TPGS, poloxamer 407. and PEG400 were added and mixed at high temperature (about 60 °C). Then ethanol was removed under vacuum at high temperature to yield a clear oil phase inco ⁇ orating the drug. It was subsequently mixed with water for injection at 45 °C to prepare the pre- emulsion.
• the final emulsion was prepared by homogenizing the pre-emulsion in the Avestin C5 homogenizer for 10-15 min at 19-20 Kpsi with the temperature being maintained between 35 and 45 °C.
• the composition of some representative emulsions are shown in the table below. The mean droplet diameter of these emulsions and 99%) cumulative distribution were determined to be less than 0.1 and 0.2 ⁇ m, respectively. These emulsions are filtered sterilizable. stable at room temperature, well tolerated in animals and are efficacious.
• Clarithromycin a macrolide antibiotic, was obtained as the free base from Wockhardt (Delhi, India). Vitamin E Succinate (VESA) was obtained from Eastman (Freeport TN). Capmul MCM was obtained from Abitec (Janesville WI); Poloxamer F127 from BASF (Parsippany NJ); PEG-400 from Spectrum Chemicals (Gardenia CA), and d- ⁇ -tocopherol from Sigma Chemicals (St Louis MO). An oil phase consisting of d- ⁇ -tocopherol and Capmul MCM was prepared using 2 parts ⁇ -tocopherol. The ⁇ TE of the oil phase is 0.0. Surfactant and clarithromycin were then added as shown in the table below. Dry ethanol was used to dissolve the components at 70 °C and the ethanol was then removed under vacuum.
• the aqueous phase consisting of 40 mL of 5 mM citrate TEA buffer, pH 6.8, was brought to 45 °C before addition.
• the resultant mixture was mixed vigorously to loosen any adherent oil on the walls of the flask.
• This suspension was then placed in a feed vessel and processed in a C5 homogenizer (Avestin. Ottawa CA) for 3 min with continuous recycling. Processing conditions were 45 °C feed temperature. 20 kpsi processing pressure, 120 mL/min flow rate.
• a heat exchanger set at 22 °C was placed at the exit port to remove excess heat generated in the homogenizer.
• the temperature in the feed vessel was measured at 44 °C during steady state homogenization.
• the product was collected and cooled to room temperature.
• the emulsion was then terminally sterilized by filtration through a 0.2 um filter and had a mean particle size of less than 52 nm when measured on a Nicomp 370 photon co ⁇ elation spectrophotometer.

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