WO2008058366A1

WO2008058366A1 - Oil-in-water emulsions, methods of use thereof, methods of preparation thereof and kits thereof

Info

Publication number: WO2008058366A1
Application number: PCT/CA2007/001676
Authority: WO
Inventors: Joanna Rossi; Christine Allen; Renita D'souza; Jean-Christophe Leroux
Original assignee: Université de Montréal; The Governing Council Of The University Of Toronto
Priority date: 2006-09-28
Filing date: 2007-09-19
Publication date: 2008-05-22

Abstract

An oil-in-water emulsion comprising at least one triglyceride oil, macrogol 15 hydroxystearate and a dispersing aqueous phase is described. Compositions, such as pharmaceutical compositions, and kits comprising the emulsion are also disclosed. Methods of preparing, as well as uses of, such emulsion and/or compositions are described. The emulsions and compositions of the invention are suitable for improving/increasing the stability, solubility and/or delivery of hydrophobic compounds.

Description

OIL-IN-WATER EMULSIONS, METHODS OF USE THEREOF, METHODS OF

PREPARATION THEREOF AND KITS THEREOF

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims the benefit, under 35 U. S. C. § 119(e), of U.S. provisional application Serial No. 60/827,289 filed on September 28, 2006, which is incorporated herein by reference in its entirety.

TECHNICAL FIELD

The present invention generally relates to oil-in-water emulsions, methods of use thereof, methods of preparation thereof and kits thereof.

BACKGROUND

Conventional, low-molecular weight therapeutics often exhibit poor specificity for the intended site of action in the body, which often leads to undesirable side-effects and low proportions of the administered dose reaching the target site. Moreover, many therapeutic compounds have poor solubility in aqueous media and thus are more difficult to administer.

Colloidal particulates such as oil-in-water emulsions can overcome many of the troubles encountered with drugs in the free form. For instance, incorporating the active compound in the oily internal phase of an emulsion can increase the solubility of lipophilic compounds, increase specificity for the target site, protect the encapsulated drug from premature degradation, enhance intracellular delivery of certain therapeutic compounds, and can potentially overcome efflux pumps such as P-glycoprotein (Pgp) (Prankerd, R. J. and Stella, V. J. (1990). J Parenter Sci Technol 44: 139-149; Barratt, G. (2003). Cell MoI Life Sci 60: 21- 37). One way by which emulsions achieve selectivity is a result of their large size, which restricts extravasation to locations in the body with permeable vasculature. Solid tumors and sites of infection or inflammation often have porous blood capillaries, which allow for the passage of nano-sized emulsion droplets across the endothelium and into the extravascular space. Given that the majority of the vascular endothelium is continuous with tight junctions between neighboring endothelial cells, active compounds associated with emulsion droplets are prevented from reaching the extravascular space of most tissues in the body reducing many of the adverse side effects caused by drugs in the free form (Maeda, H., et al. (2000). J Control Release 65: 271-284; Greish, K., et al. (2003). CHn Pharmacokinet 42: 1089-1105; Kurihara, A., et al. (1996). Pharm Res 13: 305-310; Wang, J., et al. (2002). J Pharm Sci 91 , 1128-1134). In addition to the selectivity imparted by size, targeting moieties that are specific for determinants found primarily or in high amounts on the membrane of target cells can be attached to the surface of the emulsion droplets to enhance specificity (Rensen, P. C. N., et al. (1995). Nat Med 1 : 221-225; Ishida, E., et al. (2004). Pharm Res 21: 932-939; Lundberg, B. B., et al. (1999). J Pharm Pharmacol 51 : 1099-1105). Depending on their size and the physicochemical properties of the surface, emulsions can be rapidly taken up by the cells of the mononuclear phagocyte system (MPS) and quickly removed from the systemic circulation (Takino, T., et al. (1994). Biol Pharm Bull 17: 121-125; Yeeprae, W., et al. (2006). J Control Release 114:193-201 ). Such systems are ideal for macrophage targeting. On the other hand, emulsions can exhibit long-circulating properties in blood and target sites in the body other than MPS tissues (Wang, J., et al. (2002). J Pharm Sc/ 91 , 1128-1134; Junping, W., et al. (2003). lnt J Pharm 251 : 13-21 ; Liu, F. and Liu, D. (1995). Pharm Res 12: 1060-1064; Lee, M.-J., et al. (1995). lnt J Pharm 113: 175-187). The uptake of the emulsions by the MPS is strongly dependent on the hydrodynamic diameter of the droplets and the physicochemical properties of the surface (Redgrave, T. G., et al. (1992). Biochim Biophys Acta 1126: 65-72; Lundberg, B. B., et al. (1996). lnt J Pharm 134: 119-127; Davis, S. S. and Hansrani, P. (1985). lnt J Pharm 23: 69-77).

Oil-in-water emulsions can be defined as mixtures of oil and water in which the oil phase is dispersed as fine droplets through a continuous aqueous medium. The preparation is stabilized by an emulsifier or surfactant which lowers the interfacial tension and increases droplet-droplet repulsion. Emulsions by definition are thermodynamically unstable systems and will eventually destabilize into separate oil and water phases. The addition of a surfactant only provides the preparation with kinetic stability which may supply stability for several years. Emulsions are cloudy and usually require a large input of energy for emulsification. Microemulsions, on the other hand, are preparations of oil, water and emulsifier(s) that are thermodynamically stable. Moreover, as opposed to emulsions, microemulsions are clear or translucent systems and require considerably less energy input for emulsification (Lawrence, M. J., and Rees, G. D. (2000). Adv Drug Deliv Rev 45: 89-121. Generally microemulsions require high amounts of surfactants and cosurfactants which may produce some toxicity.

Vegetable oils such as soybean oil, safflower oil and cottonseed oil have been widely investigated as the dispersed phase for emulsions because they are biocompatible and non-toxic (Floyd, A. G. (1999). Pharm Sci Technolo Today 4(2): 134-143). These oils are long- chain triglycerides (LCTs) whereby the number of carbons per hydrocarbon chain is between 14 and 22. Triglycerides can also be classified as medium-chain triglycerides (MCTs) or short- chain triglycerides (SCTs) in which the number of carbons per hydrocarbon chain is C₆-Ci₂ and C₂-C₄, respectively. A number of commercially available emulsions for parenteral nutrition contain triglyceride oils such as Intralipid®, Liposyn®, Soyacal®, and Travamulsion®. Triglyceride oils have also been investigated as solubilizers for lipophilic compounds for drug delivery and can be emulsified with either lipid or synthetic surfactants or a combination of the two (Floyd, A. G. (1999). Pharm Sci Technolo Today 4(2): 134-143; Kan, P., et a/. (1999). J Control Release 58: 271-278; Strickley, R. G. (2004). Pharm Res 21 : 201-230.

PEG-lipid derivatives are typically added to an emulsion to prolong the residence time of the droplet in the blood stream to target sites other than the tissues of the MPS. The longevity of PEGylated emulsions is attributed to the highly hydrated and flexible PEG chains, which reduces interactions with plasma proteins and cell surfaces (Allen, C, et al. (2002). Biosci

Rep 22: 225-250; Blume, G., and Cevc, G. (1993). Biochim Biophys Acta 1146: 157-168).

For the successful application of emulsions in the clinic, the formulation must demonstrate sufficient physical, chemical and microbial stability. Achieving this stability is often difficult because of the inherent thermodynamic instability of an emulsion. Most often, a combination of lipophilic and hydrophilic surfactants is necessary to prepare emulsions that are stable for prolonged periods of time. An additional difficulty in emulsion manufacture is the requirement of large energy inputs such as high shear forces, turbulence, cavitation and/or high-speed collisions with other droplets to reduce droplet size to the nanometer size range.

Thus, there is a need for novel emulsions and/or compositions, particularly for the formulation of hydrophobic compounds.

The present description refers to a number of documents, the content of which is herein incorporated by reference in their entirety.

SUMMARY OF THE INVENTION

In a first aspect, the present invention provides a method to manufacture submicrometer emulsions (<200 nm) for pharmaceutical use with minimal energy. By the term minimal energy is meant that the temperature of emulsification is low (e.g., about 20⁰C to about 60⁰C) and the formulation is agitated by low shear mixing such as with, but not limited to, a magnetic stirring plate.

Emulsions having a droplet mean diameter smaller than 200 nm may advantageously be sterilized with 0.2 μm filtration membrane. They are also small enough to extravasate across porous tumor blood capillaries and sites of infection or inflammation. The applications of these emulsions are numerous and include: a) solubilizer for lipophilic compounds; b) carrier that enhances the specificity towards sites with enhanced vascular permeability (e.g., tumors and sites of inflammation or infection); c) protection against premature degradation of the drug in the blood compartment; d) enhancement of the uptake of therapeutic agents by target cells; e) lowering of the toxicity of cytotoxic compounds; and f) potentially overcome efflux pumps.

The emulsions of the present invention comprise triglyceride oils and a hydrophilic surfactant, such as macrogol-15-hydroxystearate also known as poly(ethylene glycol)-15-hydroxystearate, and Solutol® HS-15. This surfactant consists of polyglycol mono- and di-esters of 12-hydroxystearic acid (lipophilic part) and of about 30% of free polyethylene glycol (hydrophilic part). Macrogol-15-hydroxystearate has a hydrophile-lipophile balance value of about 14 to about 16. The articles "a," "an" and "the" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article.

The term "including" and "comprising" are used herein to mean, and re used interchangeably with, the phrases "including but not limited to" and "comprising but not limited to". The term "such as" is used herein to mean, and is used interchangeably with, the phrase "such as but not limited to".

"Subject" in the context of the present invention relates to any mammal including a mouse, rat, pig, monkey, horse. In a specific embodiment, it refers to a human.

In a specific embodiment, the present invention may also provide prolonged circulation time of a compound/agent in vivo by the addition of a hydrophilic polymer-lipid derivative such as a poly(ethylene glycol) (PEG)-lipid derivative, a poly(vinylpyrrolidone)-lipid, a polyvinyl alcohol)-lipid or a dextran-lipid at the emulsion interface of the emulsions of the present invention. Prior to the addition of the hydrophilic polymer-lipid derivative, the emulsion interface comprises a monolayer of surfactant which surrounds the oil droplets. When the hydrophilic polymer-lipid derivative is added, its lipid portion inserts into the hydrophobic portion of the monolayer while the hydrophilic polymer portion extends into the aqueous phase, away from the oily inner core of the droplet. The total surface components of the emulsion including the hydrophilic polymer-lipid derivative are thus the emulsifier/surfactant (e.g., macrogol 15 hydroxystearate) and the hydrophilic polymer-lipid derivative. The present invention also relates to a kit for making an oil-in-water emulsion and a loaded oil-in-water emulsion. For example, a compartmentalized kit in accordance with the present invention includes any kit in which reagents are contained in separate containers. Such containers include small glass or plastic containers. Such containers allow the efficient transfer of reagents from one compartment to another compartment such that the samples and reagents are not cross-contaminated and the agents or solutions of each container can be added in a quantitative fashion from one compartment to another. In an embodiment, the above-mentioned kit further comprises at least one therapeutic agent (e.g., a lipophilic or hydrophobic therapeutic agent).

In embodiments, the above-mentioned kit may further comprise instructions, such as instructions for making an oil-in-water emulsion or a loaded oil-in-water emulsion. In an embodiment, the above-mentioned kit comprises a therapeutic agent and further comprises instructions to treat a disease or conditions. In an embodiment, the above-mentioned therapeutic agent is an antitumor agent (e.g., a taxane such as docetaxel) and the kit comprises instructions for the treatment of cancer in a subject. In an embodiment the above-mentioned cancer is breast cancer, lung cancer, colon cancer, prostate cancer, stomach cancer, head-and- neck cancer or ovarian cancer.

More specifically, in accordance with an aspect of the present invention, there is provided an oil-in-water emulsion comprising: (a) at least one triglyceride oil; (b) macrogol 15 hydroxystearate; and (c) a dispersing aqueous phase, wherein the emulsion contains dispersed oil-containing droplets with a mean diameter of less than 200 nm. In an embodiment, the above- mentioned oil-containing droplets have a mean diameter of less than 190 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 180 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 170 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 160 nm. In a further embodiment, the above- mentioned oil-containing droplets have a mean diameter of less than 150 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 140 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 130 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 120 nm. In a further embodiment, the above- mentioned oil-containing droplets have a mean diameter of less than 110 nm. In a further embodiment, the above-mentioned oil-containing droplets have a mean diameter of less than 100 nm.

In a specific embodiment of the emulsion, the above-mentioned oil-containing droplets have a polydispersity index (PDI) of less than 0.3 (e.g., as determined by dynamic light scattering). In a further embodiment, the above-mentioned oil-containing droplets have a polydispersity index (PDI) of less than 0.25. In a further embodiment, the above-mentioned oil- containing droplets have a polydispersity index (PDI) of less than 0.2. In a further embodiment, the above-mentioned oil-containing droplets have a polydispersity index (PDI) of less than 0.190. In a further embodiment, the above-mentioned oil-containing droplets have a polydispersity index (PDI) of less than 0.180. In a further embodiment, the above-mentioned oil- containing droplets have a PDI of less than 0.170. In a further embodiment, the above- mentioned oil-containing droplets have a PDI of less than 0.160. In a further embodiment, the above-mentioned oil-containing droplets have a PDI of less than 0.150.

In a specific embodiment of the emulsion, macrogol 15 hydroxystearate constitutes between about 2% to about 30% weight/volume (w/v) of the emulsion. In a further embodiment, macrogol 15 hydroxystearate constitutes between about 3% to about 25% w/v of the emulsion. In another specific embodiment, the dispersed oil-containing droplets constitute between about 5% and about 70% w/v of the emulsion. In another specific embodiment, the dispersed oil-containing droplets constitute about 50% w/v of the emulsion.

In another specific embodiment, the dispersing aqueous phase is selected from water, NaCI in water, glycerol, sorbitol, dextrose and xylitol, as well as derivatives or any combination thereof. In another specific embodiment, the dispersing aqueous phase is 0.9% w/v NaCI in water.

In accordance with another aspect of the present invention, there is provided a composition comprising the emulsion of the present invention, and further comprising at least one hydrophobic (e.g., a lipophilic) compound.

As used herein, a "hydrophobic compound" refers to a compound with limited water solubility. Examples of such compounds include organic molecules which lack groups that may support a formal charge (e.g., carboxylic acid and amino groups) or which lack polar groups such as hydroxyl groups. Such compounds may be amino acid-based (e.g., amino acids, peptides, polypeptide and proteins), wherein the amino acids are exclusively or predominantly hydrophobic (e.g., leucine, valine, etc.). Such compounds may be useful for diagnostic, therapeutic, cosmetic or other purposes, in a variety of fields such as oncology, , cardiovascular diseases, dermatology and antibiotic therapy. Examples of hydrophobic compounds useful for various medical applications are provided in Table IV below. "Lipophilic agent" or "Lipophilic compound "refers to a compound that is characterized by its favorable interaction with lipids.

In accordance with another aspect of the present invention, there is provided a pharmaceutical composition comprising the emulsion of the present invention, and further comprising at least one hydrophobic (e.g., a lipophilic) therapeutic agent. Pharmaceutical compositions of the present invention can be administered by routes such as orally, nasally, intravenously, intramuscularly, subcutaneously, sublingually, intrathecally, intraperitoneal^, intratumorally, topically or intradermally. The route of administration can depend on a variety of factors, such as the environment and therapeutic goals. Further non-limiting pharmaceutically suitable materials that may be incorporated in pharmaceutical preparations/compositions of the present invention include absorption enhancers, pH-adjusting agents and buffers, osmolarity adjusters, preservatives, stabilizers, antioxidants, surfactants, thickening agents, co-solvents, emollients, dispersing agents, flavoring agents, coloring agents and wetting agents and ligands/pilote/targeting molecules. Methods for preparing appropriate formulations are well known in the art (see e.g., Hendrickson, R. Ed. Remington: The Science and Practice of Pharmacy, 21^st ed.; Lippincott Williams & Wilkins: Baltimore MD, 2005).

In cases where parenteral administration is elected as the route of administration, preparations containing the emulsions and pharmaceutical compositions of the present invention may be provided to patients in combination with additional pharmaceutically acceptable sterile aqueous or non-aqueous solvents, suspensions or emulsions. Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, glycerol, dimethylacetamide, N-methylpyrrolidone and injectable organic esters. Aqueous solvents include water, water- alcohol solutions, including saline and buffered medical parenteral vehicles (c'est redundant avec "saline"), Ringer's dextrose solution, dextrose plus sodium chloride solution,. Intravenous vehicles may include fluid and nutrient replenishers, electrolyte replenishers, such as those based upon Ringer's dextrose, and the like.

The terms "preservative agent" as used herein are meant to refer to any ingredient capable of retarding or preventing microbial or chemical spoilage and protecting against discoloration. Without being so limited, it includes DMDM hydantoin, methylparaben, propylparaben, phenoxyethanol, ethylparaben, butylparaben, imidazolidinyl urea, diazolidinyl urea, quatemium-8, quatemium-14, quaternium-15, propylene glycol, dehydroacetic acid, methylchloroisothiazolinone, methylisothiazolinone and germaben.

The terms "antioxidant agent" as used herein are meant to refer to any ingredient capable of eliminating or reducing oxidation of the drug, oil and emulsifier/surfactant. Without being so limited, it includes plant extracts, algae extracts, fruit extracts, vegetable extracts, leguminous plant extracts, ferments, proteolytic hydrolysates, peptides, yeast extracts and its derivatives, microorganism extracts, animal derivative extracts and synthetic compounds. More particularly, such agents include furfuryladenine, panthenol, lipoic acid, ubiquinone, niacinamide, melatonin, catalase, glutathione, superoxide dismutase, polyphenols, cysteine, allantoin, kinetin, ascorbic acid and its derivatives (ascorbyl palmitate, magnesuim ascorbyl phosphate, sodium ascorbyl phosphate), vitamin E and its derivatives (e.g. α-tocopherol, δ- tocopherol, γ-tocopherol), grape seed extract and camellia sinensis extract.

The terms "thickening agent" as used herein are meant to refer to any ingredient that increases the viscosity of the external phase of the emulsions. Without being so limited, it includes glycerol, poly(ethylene glycol) (PEG), propylene glycol, polyvinylpyrolidone, and dextran.

The terms "isotonic agent" as used herein is meant to refer to ingredients capable of adjusting osmolarity. Without being so limited, it includes glycerol, sorbitol, xylitol and NaCI. As used herein the terms "emulsifier" and "surfactant" are used interchangeably. As used herein the terms "continuous" and "dispersing" when referring to the aqueous medium or phase are used interchangeably.

In an embodiment, the above-mentioned therapeutic agent is one or more of the agent(s)/compound(s) set forth in Table IV below, or prodrugs, metabolites or derivatives thereof. In an embodiment, the above-mentioned derivative is characterized by the presence of a lipid or lipophilic group/moiety (e.g., lauroyl, dilauroyl). In a further embodiment, the above- mentioned therapeutic agent is an antitumor agent. In an embodiment, the above-mentioned therapeutic or antitumor agent is a taxane. "Taxane" refers to the class of antineoplastic agents having a mechanism of microtubule action and having a structure that includes the unusual taxane ring structure and a stereospecific side chain that is required for cytostatic activity. Taxane further refers to a variety of known taxane derivatives, including both hydrophilic derivatives, and hydrophobic derivatives. Taxane derivatives include, but are not limited to, galactose and mannose derivatives described in International Patent Application No. WO 99/18113; piperazino and other derivatives described in WO 99/14209; taxane derivatives are described in WO99/09021 , WO 98/22451 , and U.S. Pat. No. 5,869,680; 6-thio derivatives described in WO 98/28288; sulfenamide derivatives described in U.S. Pat. No. 5,821 ,263; and paclitaxel derivatives described in U.S. Pat. No. 5,415,869. In a further embodiment, the above- mentioned taxane is paclitaxel, docetaxel, or prodrugs, metabolites or derivatives thereof. Chemical modification may be used as a means to alter the properties of the taxane such that the derivative has enhanced solubility and retention in the emulsion droplets. For example, the taxane may be modified by reaction with acyl halides (e.g. caproyl, C6; lauroyl, C12; myrisoyl, C14; or steroyl, C16) under specific conditions to produce mono (2'-modified docetaxel or 2'-modified paclitaxel), di (2',7-modified-docetaxel or 2',7-modified paclitaxel) or trisubstituted (2',7,10-modified docetaxel) compounds. For example the 2',7-dilauroyl-docetaxel derivative was incorporated in the nanoemulsions in Example 8. In an embodiment, the docetaxel derivative is an acyl halide mono-, di- or trisubstituted docetaxel.

In an embodiment, the above-mentioned composition comprises more than one therapeutic agent. In a specific embodiment of the pharmaceutical composition, the dispersed oil- containing droplets constitute between about 10% and about 20% w/v of the emulsion loaded with the hydrophobic therapeutic agent. In a specific embodiment of the pharmaceutical composition, the dispersed oil-containing droplets constitute about 17% w/v of the emulsion loaded with the hydrophobic therapeutic agent. In another specific embodiment, the dispersed oil-containing droplets constitute between about 40% and about 60% w/v of the emulsion loaded with the hydrophobic therapeutic agent. In another specific embodiment, the dispersed oil-containing droplets constitute about 50% w/v of the emulsion loaded with the hydrophobic therapeutic agent. In another specific embodiment, the therapeutic agent is an antitumor agent. In a further embodiment, the above-mentioned antitumor agent is taxane (e.g., paclitaxel, docetaxel).

In an embodiment, the above-mentioned therapeutic agent is present in an amount of about 0.05% to about 5% w/v in the composition. In a further embodiment, the above-mentioned therapeutic agent is present in an amount of about 0.1% to about 2.5% w/v in the composition.

In another specific embodiment, the pharmaceutical composition further comprises a lipid solubilizer for said therapeutic agent. In another specific embodiment, the lipid solubilizer is tributyrin or a prodrug, derivative, or metabolite thereof. In another specific embodiment, the pharmaceutical composition further comprises a thickening agent selected from glycerol, poly(ethylene glycol) (PEG), propylene glycol, polyvinylpyrolidone, dextran, and derivatives or any combination thereof. In another specific embodiment, the pharmaceutical composition further comprises a co-solvent selected from poly(ethylene glycol) (PEG), ethanol, dimethylacetamide, N-methylpyrrolidone and derivatives or any combination thereof. In another specific embodiment, the pharmaceutical composition further comprises an isotonic agent selected from glycerol, sorbitol, xylitol, NaCI and derivatives or any combination thereof. In another specific embodiment, the pharmaceutical composition further comprises an antioxidant selected from the group consisting of α-tocopherol, deferoxamine mesylate, ascorbic acid and derivatives or any combination thereof. In another specific embodiment, the pharmaceutical composition further comprises a preservative. In another specific embodiment, the pharmaceutical composition further comprises a pH-adjusting agent. In an embodiment, the above-mentioned pH-adjusting agent is HCI or NaOH. In another specific embodiment, the pharmaceutical composition is for intravenous, subcutaneous, intraperitoneal, intramuscular, intratumoral, oral, nasal, sublingual, intrathecal or topical administration. In another specific embodiment, the pharmaceutical composition is for intravenous administration, and further comprises aqueous NaCI as a pharmaceutical carrier/excipient. In another specific embodiment, the pharmaceutical composition further comprises a hydrophilic polymer-lipid derivative. In another specific embodiment, the hydrophilic polymer-lipid derivative is a poly(ethylene glycol) (PEG)-lipid derivative. In another specific embodiment, the PEG-lipid derivative constitutes about 1 to about 15 mol% of the total surface components of the emulsion. In another specific embodiment, the PEG-lipid derivative is selected from 1 ,2-dimyristoyl-sn-glycero-3- phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], 1 ,2-dipalmitoyl-sn-glycero-3- phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], 1 ,2-distearoyl-sn-glycero-3- phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], 1 ,2-dioleoyl-sn-glycero-3- phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], N-octanoyl-sphingosine-1- [succinyl(methoxyPEG)-2000 or 5000], and any combination thereof. In another specific embodiment, the PEG-lipid derivative is 1 ,2-distearoyl-sn-gly∞ro-3-phosphatidylethanolamine- N-methoxy-[PEG2000].

The compositions of the present invention may be used in a variety of in vitro and in vivo applications. For example, the above-mentioned compositions may be used in a method for delivering a hydrophobic compound to cells in vitro. Such a method may be useful to determine what affect, if any, a particular hydrophobic compound has upon a particular cell type, or to introduce a hydrophobic compound in order to confer specific properties to, or to alter one or more function(s) of, a cell. The compositions of the present invention may also be used in a method for delivering a hydrophobic compound to cells in vivo. For example, the composition of the present invention may be administered to a warm-blooded animal (e.g., a mammal such as a human), under proper conditions to permit delivery of the compound. It will be appreciated by those of ordinary skill in the art that the amount of a composition administered will depend upon several factors such as the gender, age and weight of the subject receiving the composition, as well as the particular hydrophobic compound utilized and the desired result/effect.

In accordance with another aspect of the present invention, there is provided a method of making an oil-in-water emulsion comprising: (a) mixing a triglyceride oil with macrogol 15 hydroxystearate at a temperature between about 35°C and about 70⁰C to melt macrogol 15 hydroxystearate so as to form a first mixture; (b) emulsifying the first mixture with a dispersing aqueous phase to obtain droplets with a mean diameter of less than 200 nm, whereby the droplets constitute about 5% to about 70% w/v of the emulsion.

In accordance with another aspect of the present invention, there is provided a method of making a lipophilic therapeutic agent loaded oil-in-water emulsion comprising: (a) mixing a triglyceride oil with the therapeutic agent so as to dissolve the therapeutic agent and form a first mixture; (b) mixing the first mixture with macrogol 15 hydroxystearate at a temperature sufficient to melt macrogol 15 hydroxystearate so as to form a second mixture; and (c) emulsifying the second mixture with a dispersing aqueous phase to obtain an emulsion containing droplets with a mean diameter of less than 200 nm, whereby the droplets constitute about 5% to about 70% w/v of the emulsion. In a specific embodiment of the method, the above-mentioned therapeutic agent is an antitumor agent. In a further embodiment, the above-mentioned antitumor agent is a taxane (e.g., docetaxel or docetaxel derivative).

In another specific embodiment of the method, the docetaxel derivative is an acyl halide mono-, di- or trisubstituted docetaxel. In another embodiment of the method, the above- mentioned docetaxel derivative is 2',7-dilauroyl-docetaxel.

In accordance with another aspect of the present invention, there is provided a method of making a lipophilic therapeutic agent loaded oil-in-water emulsion comprising: (a) mixing a triglyceride oil with the therapeutic agent and macrogol 15 hydroxystearate so as to dissolve the therapeutic agent and melt macrogol 15 hydroxystearate and form a first mixture; and (b) emulsifying the first mixture with a dispersing aqueous phase to obtain an emulsion containing droplets with mean diameter of less than 200 nm, whereby the droplets constitute about 5% to about 70% w/v of the emulsion.

In a specific embodiment of the methods of the present invention, the mixing in (a) is conducted at a temperature of between about 35°C and about 70⁰C. In another specific embodiment, emulsifying is conducted at a temperature of between about 20°C and about 6O⁰C. In another specific embodiment, mixing in (a) and emulsifying are conducted at a temperature of between about 40⁰C and about 50⁰C, and wherein the therapeutic agent is docetaxel. In another specific embodiment, mixing step (a) is conducted for about 15 minutes to about 90 minutes. In another specific embodiment, mixing step (a) is conducted at a velocity of about 400 to about 800 rpm.

In an embodiment, the above-mentioned triglyceride oil is one or more of the triglyceride oil(s) set forth in Table I below. In another specific embodiment, the triglyceride oil is a medium-chain triglyceride (MCT). In another specific embodiment, the triglyceride oil comprises saturated fatty acid. In a further embodiment, the above-mentioned saturated fatty acid is one or more of the fatty acid set forth in Table Il below.

In another specific embodiment, the dispersing aqueous phase is 0.9% w/v NaCI in water. In another specific embodiment, macrogol 15 hydroxystearate constitutes about 2% to about 30% w/v of the emulsion. In another specific embodiment, the droplets constitute about 17% w/v of the emulsion. In another specific embodiment, the droplets constitute about 50% w/v of the emulsion.

In another specific embodiment, the method further comprises homogenizing the emulsion to reduce the droplets size. In a further embodiment, the step of homogenizing is performed with a high-pressure homogenizer or a microfluidizer. Homogenization in accordance with the present invention may be performed on the emulsion to further reduce the droplet mean diameter. This homogenization may be performed with well-known techniques or devices such as, without being so limited, propeller, turbine mixers, homogenizers, colloid mills, ultrasonic mixers and microfluidizers. In an embodiment, the homogenization is performed using a high- pressure homogenizer or a microfluidizer.

In another specific embodiment, the method further comprises a step of attaching a hydrophilic polymer-lipid derivative to the droplet interface of the emulsion. In another specific embodiment, the step of attaching a hydrophilic polymer-lipid derivative is performed by adding an aqueous solution of the hydrophilic polymer-lipid derivative to the emulsion to form a hydrophilic polymerated emulsion and incubating the hydrophilic polymerated emulsion at a temperature of about 20°C to about 60°C. In another specific embodiment, the step of attaching a hydrophilic polymer-lipid derivative is performed by dissolving the hydrophilic polymer-lipid derivative in the dispersing aqueous phase prior to emulsifying, whereby emulsifying forms a hydrophilic polymerated emulsion and incubating the hydrophilic polymerated emulsion at a temperature of about 40⁰C to about 60⁰C. In another specific embodiment, the step of attaching a hydrophilic polymer-lipid derivative is performed by dissolving the hydrophilic polymer-lipid derivative in the first mixture, whereby emulsifying forms a hydrophilic polymerated emulsion and incubating the hydrophilic polymerated emulsion at a temperature of about 40°C to 6O⁰C. In another specific embodiment, the hydrophilic polymer-lipid derivative constitutes about 1 to about 15 mol% of the total surface components of the emulsion. In another specific embodiment, the hydrophilic polymer-lipid derivative is poly(ethylene glycol) (PEG)-lipid derivative. In another specific embodiment, the PEG-lipid derivative is selected from 1 ,2- dimyristoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], 1 ,2- dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], 1 ,2- distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], 1 ,2- dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000, 3000 or 5000], and N- octanoyl-sphingosine-1-[succinyl(methoxyPEG)-2000 or 5000], or any combination thereof. In another specific embodiment, the PEG-lipid derivative is 1 ,2-distearoyl-sn-glycero-3- phosphatidylethanolamine-N-methoxy-[PEG2000]. In another specific embodiment, no organic solvent is used. In accordance with another aspect of the present invention, there is provided a use of the emulsion of the present invention, in the making or manufacture of a medicament.

In accordance with another aspect of the present invention, there is provided a use of the pharmaceutical composition of the present invention, in the making or manufacture of a medicament. In accordance with another aspect of the present invention, there is provided a use of the above-mentioned pharmaceutical composition for the treatment of cancer.

In accordance with another aspect of the present invention, there is provided a use of the above-mentioned pharmaceutical composition, in the making or manufacture of a medicament for the treatment of cancer. In an embodiment the above-mentioned cancer is breast cancer, lung cancer, colon cancer, prostate cancer, stomach cancer, head-and-neck cancer or ovarian cancer.

In accordance with another aspect of the present invention, there is provided a method of improving the solubility of a lipophilic therapeutic compound comprising formulating the compound into the dispersed oil-containing droplets of the emulsion of the present invention.

In accordance with another aspect of the present invention, there is provided a method of increasing circulation time in vivo of a lipophilic compound comprising formulating the compound into the dispersed oil-containing droplets of the pharmaceutical composition of the present invention.

In accordance with another aspect of the present invention, there is provided a kit comprising (a) at least one hydrophobic therapeutic agent; (b) at least one triglyceride oil; and (c) macrogol 15 hydroxystearate. In an embodiment, the above-mentioned kit further comprises instructions for the making of a pharmaceutical composition (e.g. a medicament) using the above-mentioned at least one hydrophobic therapeutic agent, at least one triglyceride oil and macrogol 15 hydroxystearate. In another embodiment, the above-mentioned at least one hydrophobic therapeutic agent is an antitumor agent (e.g., docetaxel or docetaxel derivative). In another embodiment, the above-mentioned docetaxel derivative is a acyl halide mono-, di- or trisubstituted docetaxel.

In another embodiment, the above-mentioned docetaxel derivative is 2', 7- dilauroyl-docetaxel. In yet another embodiment, the above-mentioned kit further comprises instructions for the treatment of cancer in a subject. Other objects, advantages and features of the present invention will become more apparent upon reading of the following non-restrictive description of specific embodiments thereof, given by way of example only with reference to the accompanying drawings.

BRIEF DESCRIPTION OF DRAWINGS In the appended drawings:

Figure 1 illustrates the elimination profile of a docetaxel-loaded PEGylated emulsion from blood after intravenous (i.v.) injection in mice inoculated with C26 colon adenocarcinoma. Mean ± SD (n = 3 to 5 mice/group);

Figure 2 depicts the distribution of the docetaxel-loaded PEGylated emulsion into subcutaneously implanted C26 colon adenocarcinoma (tumor volume ~ 20 mm³) (") and muscle (•) after i.v. injection in mice. Significant selectivity for the tumor over the muscle tissue is demonstrated. Mean ± SD (n = 4 to 5 mice/group);

Figure 3 shows the blood concentration-time profiles of docetaxel encapsulated in a PEGylated emulsion versus the commercial formulation (Taxotere®) after i.v. injection in mice inoculated with C26 colon adenocarcinoma. The dose of docetaxel administered to each mouse was 5 mg kg-1. The nano-emulsions were loaded with 0.61 % w/w docetaxel. Mean ± SD

(n = 3 to 5 mice/group); and

Figure 4 presents the tissue distribution of PEGylated emulsions after i.v. injection in mice bearing C26 colon adenocarcinoma. Mean ± SD (n = 4 to 5 mice/group). DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS

Process of Preparation of Unloaded emulsion

Unloaded emulsions are prepared by first mixing the internal phase (oil) with a surfactant, namely macrogol 15 hydroxystearate. Typically, the mixture is heated between about 40 and about 70⁰C under agitation for a time and at a velocity sufficient to promote dissolution of all components and to melt macrogol 15 hydroxystearate, which is semi-solid at room temperature. Triglyceride oils (LCTs, MCTs or SCTs) are an appropriate internal phase for the present invention. The amount of surfactant added may represent between 2% to 30% w/v of the formulation. The aqueous dispersing phase (e.g., 0.9% w/v NaCI in water) is then added to the mixture under gentle agitation at a temperature of 20-60⁰C. Without being so limited, the aqueous dispersing phase may be water, saline, glycerol, sorbitol, dextrose and xylitol, as well as any derivative or combination thereof. Pure water is appropriate for administration routes other than intravenous administration. After the addition of saline, the dispersed phase can represent between 5% and 70% w/v of the emulsion. The concentration of the dispersed phase may advantageously be increased to lower the volume of formulation to be administered for a specific dosage. This composition and process produces nanosized emulsions with droplets having a mean size of less than 200 nm in diameter with a polydispersity index (PDI) of less than 0.3 as determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK). Without being so limited, oils useful for the present invention include natural oils such as safflower, soybean, corn, olive and peanut oils or semi-synthetic commercially available triglyceride oils such as those presented in Table I below. Useful triglyceride oils may also be prepared with fatty acids including, without being so limited, those presented in Table Il below. Any combination of such triglycerides may also be used in the emulsions and pharmaceutical compositions of the present invention.

Table I: Examples of commercially available triglyceride oils

Table II: Examples of Fatty Acids

To further reduce droplet size, the formulation can be homogenized with a device such as a high-pressure homogenizer (e.g. Emulsiflex™-C3™, Avestin, ON, Canada) or a microfluidizer at a pressure and duration adapted to achieve the desired mean droplet diameter. The desired mean droplet diameter may vary depending on the specific drug delivery application for which the emulsion of the present invention is used. For instance, a size of 50- 200 nm is particularly desired for drug targeting applications. The pressure, temperature and duration of homogenization necessary to achieve a desired mean droplet diameter may change depending on the concentration of the dispersed phase, the composition of the formulation and the sensitivity of the therapeutic agent(s)/excipient(s) to heat. The methods of preparation of the emulsions and formulations of the present invention are thus not limited to the pressure, temperature and durations specified in Examples presented herein.

Hydrophilic polymer-lipid derivatives such as PEG-lipid derivatives, poly(vinylpyrrolidone)-lipid, polyvinyl alcohol)-lipid and dextran-lipid may be attached to the droplet interface of the emulsions. It is believed that these derivatives confer the emulsions with circulation longevity in vivo. For instance, this may be achieved by adding an aqueous solution of a hydrophilic polymer-lipid derivative such as a PEG-lipid to the oil or water phase prior to the emulsification step or to the preformed emulsions, followed by an incubation period at a temperature of about 40⁰C to about 60⁰C for a duration of about 15 min to about 2 h. To achieve prolonged circulation time in vivo, the amount of PEG-lipid added to the emulsion is desirably between about 1 mol% and about 15 mol% of the total surface components (excluding the internal phase and taking an average molecular weight of 960 g/mol for macrogol 15 hydroxystearate). Many different PEG-lipid derivatives can be used with the present invention. In an embodiment, 1 ^-distearoyl-sn-glycero-S-phosphatidylethanolamine-N-methoxy- [PEG2000] (DSPE-PEG2000) is used. Without being so limited, Table III below provides a list of various PEG-lipid derivatives that can be used with the present invention and that are commercially available (e.g. Northern Lipids Inc., Vancouver, BC, Canada and Avanti Polar Lipids Inc., Alabaster, Ala). After PEGylation, the mean droplet diameter typically increases by about 5 nm to about 10 nm. The emulsions are preferably stored at ambient conditions and protected from light.

Table III: Examples of PEG-lipid derivatives 1 ,2-dimyristoyl-s/?-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)- 2000, 3000 or 5000]

1 ,2-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)- 2000, 3000 or 5000]

1 ,2-distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)- 2000, 3000 or 5000]

1 ,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)- 2000, 3000 or 5000] N-octanoyl-sphingosine-HsuccinyKmethoxyPEG^OOO or 5000]

The emulsions could be further decorated with a targeting ligand/pilot molecule or moiety (e.g., vitamin, dye, peptide, antibody, antibody fragment, sugar, etc.). These ligands may be useful, for example, for site-specific compound/drug delivery. This pilot molecule could be for instance linked to the extremity of the hydrophilic polymer (e.g. PEG).

A targeting ligand/moiety may be covalently or noncovalently associated with the emulsion or composition of the present invention. For example, activated lipids are commercially available (e.g., Avanti Polar Lipids, Inc.) which possess chemically reactive groups for covalently joining a targeting moiety. Typically, a targeting moiety is reacted via one of the nucleophilic groups (e.g., amino or sulfhydryl) with a chemically reactive group on an activated lipid. Chemically reactive groups include, for example, iodoacetyl, maleimidylbenzoyl, maleimidylphenyl, and pyridyldithio groups. Methodologies for the preparation of targeting moieties associated with lipids are well known to those of ordinary skill in the art (see, e.g., Liposome Technology, Vol. Ill, G. Gregoriadis, ed., CRC Press, Inc., Boca Raton, FIa.; Loughrey et al. 1987. Biochim. Biophys. Acta 901 :157; Loughrey et al., 1990. J. Immunol. Methods 132:25). In addition, certain lipids linked to a targeting moiety are commercially available.

The present invention may permit the loading of lipophilic compounds/drugs in the oily internal phase of the emulsion. To incorporate a compound (e.g., a therapeutic agent) into the emulsion, the compound is first dissolved in the oil phase with or without heat (heat typically increases dissolution rate) with or without the surfactant macrogol 15 hydroxystearate under gentle agitation. Certain compounds being susceptible to degradation when excessively heated in the presence of macrogol 15 hydroxystearate, they may be mixed and heated with the oil prior to addition of macrogol 15 hydroxystearate where the mixture is again mixed and heated to between about 35°C and about 50 ⁰C and stirred for a time sufficient to melt the surfactant (typically about 10 min to about 30 min). The dispersing phase (e.g., 0.9% w/v NaCI in water) is then added to the mixture and heated/stirred at a temperature between about 2O⁰C and about 60⁰C depending on the heat sensitivity of the drug in the presence of macrogol 15 hydroxystearate and water. This premix can be further homogenized with a high-pressure homogenizer for instance (Emulsiflex™-C3, Avestin, ON, Canada) at a pressure and duration sufficient to further decrease droplet size and/or polydispersity.

These emulsions may be used to enhance the solubility of hydrophobic compounds/drugs, protect the encapsulated drug against hydrolysis and enzymatic degradation in the blood compartment, lower the toxicity of cytotoxic compounds, increase selectivity towards target tissues, improve intracellular delivery of certain compounds, and potentially overcome efflux pumps such as P-glycoprotein (Pgp).

Several hydrophobic compounds can be incorporated into the emulsion. Without being so limited, Table IV below provides a list of therapeutic compounds. Any derivative, prodrug, metabolite or analogue of the compounds listed below may also be used in the formulations of the present invention:

Table IV: Examples of Hydrophobic therapeutic compounds

Therapeutic application Hydrophobic compounds

Anesthetics Propanidid,

Propofol,

Alphadione

Antibiotic Echinomycin

Antifungal Miconazole nitrate

Antineoplastic Taxanes (also known as taxines or taxoids) such as paclitaxel and docetaxel,

Podophyllotoxins,

Camptothecins such as camptothecin, 9- aminocamptothecin, 9-nitrocamptothecin, camptothecin-11 ("Irinotecan"), Topotecan,

Vinca alkaloids and their analogs

(vincristine, vinorelbine, vindesine, vintripol, vinxaltine, ancitabine), Lipophilic anthracyclines,

Decarbazine,

Lonidamine,

Piroxantrone,

Anthrapyrazoles,

Etoposide,

Bleomycin,

6-aminochrysene,

Navelbine

Tributyrin

Teniposide

Platinum-based agents

Chemoprophylactic Praziquantel Immunosuppressant Cyclosporin A,

18-hydroxydeoxycorticosterone,

Rapamycin

Glucocorticoid Prednisolone Nutritional supplements Vitamin A,

Vitamin E

Photosensitizers Purpurin,

Tin etiopurpurin,

Porphyrins Sunscreens Paraaminobenzoic acid

Tranquilizer Diazepam,

Delta 9-tetrahydrocannabinol Vaccines BBB-MDP

Vasodilator Verapamil,

Nifedipine

In order to increase the affinity for the emulsion droplets, the compounds may be modified with one or more lipid(s) or lipid derivative(s). For example, the compounds/drugs may be conjugated with monoglycerides, diglycerides, fatty acids, phospholipids, sterols, and the like.

In an embodiment, the present emulsion and method of making the emulsion advantageously comprises only one surfactant/emulsifier. In other embodiments, the present method of making an emulsion does not produce a gel at any step of the process and does not need pH adjustment. In other embodiments, it further does not comprise diethylene glycol monoethylether, hydroxyalkane, a dihydroxyalkane, a polyethylene glycol having an average molecular weight of at most 1000, an organic solvent, a HDL apolipoprotein, a cationic lipid, a lipophilic oily fatty alcohol, a phospholipid, ethanol, polyoxyethylene glycol trioleate, a lipophilic surfactant, a lipid emulsifier, an emulsion-stabilizing surface active drug, a HSP90 inhibitor, an emulsifier with a hydrophile-lipophile balance value of less than 6, a metallic oxide or cholesterol, and does not require that the therapeutic agent possess at least one ionizable functional group and an ionizable agent capable of ionizing the ionizable functional group. In another embodiment, the emulsions of the present invention do not form nano-capsules.

MODE(S) FOR CARRYING OUT THE INVENTION The present invention is illustrated in further details by the following non-limiting examples.

EXAMPLE 1 : PREPARATION OF EMULSIONS WITHOUT DRUG Labrafac™ CC/Macrogol 15 hvdroxystearate (emulsion 1 )

Ingredients Amount (mg) Concentration (%

Labrafac™ CC 785 5.23

Macrogol 15 hydroxystearate ^₀₇ ~ _c-i (Solutol™ HS-15) _

Total 1312 8.75 Labrafac™ CC (785 mg) and macrogol 15 hydroxystearate (527 mg) were weighed in a 20-ml scintillation vial. The mixture was heated at 4O⁰C for 15 min with agitation (630 rpm) and then cooled down to room temperature for 5 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the addition of saline, the dispersed phase represented about 9% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 134 nm, with PDI of 0.107. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 1 min 45 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 118 nm and 0.108, respectively. The emulsions are stored at ambient conditions and protected from light.

Labrafac™ CC/Macrogol 15 hvdroxystearate (emulsion 2)

Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 6487 30.89 Macrogol 15 hydroxystearate 4013 19.11 (Solutor HS-15)

Total 10500 50.00

Labrafac™ CC (6487 mg) and macrogol 15 hydroxystearate (4013 mg) were weighed in a vial. The components were heated/mixed at 4O⁰C for 15 min with agitation (630 rpm). The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 21 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the inclusion of saline, the dispersed phase represented 50% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 157 nm, with PDI of 0.159. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for various durations to reduce droplet size further. The change in mean droplet diameter size with homogenization time is presented in Table V. The emulsions are stored at ambient conditions and protected from light.

Table V: Changes in mean droplet diameter size and PDI with homogenization time

(emulsion 2) Homogenization time Di_amet_er (nm) PDI

Before homogenization 157 0.159

2 127 0.121

3 121 0.110

4 115 0.109 5 109 0.090

6 107 0.079

7 102 0.072

8 100 0.117

9 97 0.074

10 99 0.077

11 99 0.089

12 98 0.080

13 100 0.084

14 97 0.099

15 99 0.059

EXAMPLE 2: PREPARATION OF EMULSIONS WITH TRIBUTYRIN

Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate (emulsion 3)

Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 800 5.33 Tributyrin 800 5.33 Macrogol 15 hydroxystearate

990 6.60 (Solutol® HS-15)

Total 2590 17.27

Labrafac™ CC (800 mg) and tributyrin (800 mg) were weighed in a 20-ml scintillation vial. The solution was heated at 7O⁰C for 30 min with agitation (630 rpm) and then cooled down to room temperature for 5 min. Macrogol 15 hydroxystearate (990 mg) was added and the mixture was heated/mixed at 4O⁰C for 15 min (630 rpm). The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the inclusion of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 148 nm, with PDI of 0.282. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 1 min 45 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 95 nm and 0.198, respectively. The emulsions are stored at ambient conditions and protected from light.

Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate (emulsion 4) Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 984 6.56

Tributyrin 657 4.38

Macrogol 15 hydroxystearate _{onΛ β cπ} (Solutol* HS-15) _ ^{6 6}° Total 2631 17.54

Labrafac™ CC (984 mg) and tributyrin (657 mg) were weighed in a 20-ml scintillation vial. The solution was heated at 7O⁰C for 30 min with agitation (630 rpm) and then cooled down to room temperature for 5 min. Macrogol 15 hydroxystearate (990 mg) was added and the mixture was heated/mixed at 4O⁰C for 15 min (630 rpm). The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the inclusion of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 138 nm, with PDI of 0.179. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 1 min 45 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 98 nm and 0.139, respectively. The emulsions are stored at ambient conditions and protected from light.

Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate (emulsion 5)

Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 1067 7.11 Tributyrin 534 3.56 Macrogol 15 hydroxystearate

990 6.60 (Solutol^® HS-15)

Total 2591 17.27

Labrafac™ CC (1067 mg) and tributyrin (534 mg) were weighed in a 20-ml scintillation vial. The solution was heated at 7O⁰C for 30 min with agitation (630 rpm) and then cooled down to room temperature for 5 min. Macrogol 15 hydroxystearate (990 mg) was added and the mixture was heated/mixed at 4O⁰C for 15 min (630 rpm). The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the inclusion of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering

(Malvern Instruments Ltd, Malvern, UK) to be 133 nm, with PDI of 0.153. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at

10,000 psi for 1 min 45 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 102 nm and 0.163, respectively. The emulsions are stored at ambient conditions and protected from light. Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate (emulsion 6)

Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 1200 8.00 Tributyrin 400 2.67 Macrogol 15 hydroxystearate

990 6.60 (Solutor HS-15)

Total 2590 17.27

Labrafac™ CC (1200 mg) and tributyrin (400 mg) were weighed in a 20-ml scintillation vial. The solution was heated at 7O⁰C for 30 min with agitation (630 rpm) and then cooled down to room temperature for 5 min. Macrogol 15 hydroxystearate (990 mg) was added and the mixture was heated/mixed at 4O⁰C for 15 min (630 rpm). The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the inclusion of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 132 nm, with PDI of 0.127. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 1 min 45 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 106 nm and 0.126, respectively. The emulsions are stored at ambient conditions and protected from light.

Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate (emulsion 7)

Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 1280 8.53 Tributyrin 320 2.13 Macrogol 15 hydroxystearate

990 6.60 (Solutor HS-15)

Total 2590 17.27

Labrafac™ CC (1280 mg), tributyrin (320 mg) and macrogol 15 hydroxystearate (990 mg) were weighed in a 20-ml scintillation vial. The mixture was heated at 4O⁰C for 30 min with agitation (630 rpm) and then cooled down to room temperature for 5 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 5O⁰C for an additional

10 min (840 rpm). After the addition of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 130 nm, with PDI of 0.113. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON,

Canada) at 10,000 psi for 1 min 30 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 103 nm and 0.132, respectively. The emulsions are stored at ambient conditions and protected from light.

Labrafac™ CC/Tributyrin/Macrogol 15 hvdroxystearate (emulsion 8) Ingredients Amount (mg) Concentration (% w/v)

Labrafac™ CC 3866 18.41

Tributyrin 2577 12.27

Macrogol 15 hvdroxystearate _QQQ« _{Λ a aΛ} (Solutol* HS-15) _ _ Total 10436 49.70

Labrafac™ CC (3866 mg), tributyrin (2577 mg) and macrogol 15 hydroxystearate (3993 mg) were weighed in a vial. The components were heated/mixed at 4O⁰C for 15 min with agitation (630 rpm). The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 21 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the inclusion of saline, the dispersed phase represented about 50% w/v of the emulsion. Using a larger percentage of dispersed phase in the emulsion allows the administration of a smaller volume of emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 158 nm, with PDI of 0.278. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 8,000 psi for various durations to reduce droplet size further. The change in mean droplet diameter size with homogenization time is presented in Table Vl. The emulsions are stored at ambient conditions and protected from light.

Table Vl: Changes in mean droplet diameter size and PDI with homogenization time (emulsion 8)

Homogenization time Diameter (nm) PDI (mm)

Before homogenization 158 0.278

2 124 0.139

3 115 0.141

4 1 10 0.143

5 99 0.125

6 92 0.137

7 92 0.110

8 89 0.137

9 87 0.142

10 84 0.100

11 84 0.144

12 82 0.072

13 83 0.077

14 81 0.066

15 81 0.080 EXAMPLE 3: PREPARATION OF EMULSIONS LOADED WITH DOCETAXEL

Docetaxel/Labrafac™ CC/Macroqol 15 hydroxystearate (emulsion 9) Ingredients Amount (mg) Concentration (% w/v)

Docetaxel 62 0.31

Labrafac™ CC 6145 30.73

Macrogol 15 hydroxystearate ,_7M , _{o n7} (Solutol* HS-15) ™ Total 10000 50.00

The emulsion was prepared by first dissolving docetaxel (62 mg) in Labrafac™ CC (6145 mg) at 7O⁰C for 30 min with stirring (630 rpm). The solution was heated to increase the dissolution rate of docetaxel in the oil phase. The mixture was then cooled down to room temperature and 3793 mg of macrogol 15 hydroxystearate was added. The mixture was heated to 4O⁰C and stirred (630 rpm) for 15 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 20 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the addition of saline, the dispersed phase represented 50% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 153 nm, with PDI of 0.195. This premix was then homogenized with a high- pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 13 min to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 98 nm and 0.083, respectively. The emulsion was then sterilized by filtration through 0.45 μm nylon filters (25 mm in diameter, sterile). The formulation was stored at ambient conditions and protected from light. This emulsion was used for the stability experiment in Example 7.

Docetaxel/Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate (emulsion 10)

Docetaxel/Labrafac™ CC/Tributyrin/Macrogol 15 hydroxystearate. Tributyrin was used in this emulsion as a lipid solubilizer for docetaxel, which increases the amount of docetaxel that can be loaded in the formulation. Docetaxel is indeed more soluble in tributyrin than in Labrafac™ CC.

Ingredients Amount (mg) Concentration (% w/v)

Docetaxel 16 0.1 1

Labrafac™ CC 1280 8.53

Tributyrin 320 2.13

Macrogol 15 hydroxystearate

990

(Solutor HS-15) 6.60

Total 2606 17.37 Docetaxel (16 mg) was dissolved in Labrafac™ CC (1280 mg) and tributyrin (320 mg) at 7O⁰C for 30 min with stirring (630 rpm). The solution was heated to dissolve any nano- crystals that may be present in the solution. The mixture was then cooled down to room temperature and 990 mg of macrogol 15 hydroxystearate was added. The mixture was heated to 4O⁰C and stirred (630 rpm) for 15 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After the addition of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix was determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) to be 132 nm, with PDI of 0.126. This premix was then homogenized with a high- pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 1 min 45 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 104 nm and 0.116, respectively.

EXAMPLE 4: PREPARATION OF PEGYLATED EMULSIONS LOADED WITH DOCETAXEL

Five milliliters of emulsion 10 was incubated with 1536 μl of a micelle solution of DSPE-PEG₂₀₀₀ (70 mg/ml) for 1 h at 4O⁰C. This was done to coat the surface of the droplets with PEG₂₀₀₀ in order to confer the emulsion with long-circulating properties in blood. The amount of DSPE-PEG_2Ooo added to the emulsion corresponded to 10 mol% of the total surface components. After PEGylation, the mean droplet diameter was 110 nm (PDI: 0.116). The emulsions were stored at ambient conditions and protected from light.

EXAMPLE 5:

BlODISTRIBUTION OF NANO-EMULSIONS

PEGylated nano-emulsions were prepared as described in Example 4 and were diluted with NaCI (0.9% w/v) for injection. [³H]-Cholesteryl hexadecyl ether ([³H]-CHE) and [¹⁴C]- docetaxel were incorporated during emulsification to track the distribution of the emulsion droplets and drug, respectively. Each mouse received 0.8 and 0.5 μCi of [³H]-CHE and [¹⁴C]- docetaxel, respectively. Taxotere^® (Aventis Pharma Ltd., Dagenham, UK) was labelled with

[¹⁴C]-docetaxel only (0.5 μCi ¹⁴C/mouse). The emulsion and Taxotere^® were administered to Balb/C mice bearing C26 colon adenocarcinoma at a dosage of 5 mg kg^"1 via the subclavian vein in a 110 μl injection volume. The PEGylated emulsion demonstrated long-circulating properties in blood and accumulated into subcutaneously implanted C26 colon adenocarcinoma (tumor volume ~ 20 mm³) (Figures 1 and 2). Significant selectivity for the tumor over the muscle tissue is also shown in Figure 2. The pharmacokinetic profile of docetaxel encapsulated in the emulsion was similar to that of Taxotere^® (Figure 3). Uptake of the emulsion by different organs post i.v. injection is presented in Figure 4. The emulsion distributed mainly to the organs of the mononuclear phagocyte system (MPS), with the majority of the formulation accumulating in the liver. Distribution into the lungs, heart and kidneys was low.

EXAMPLE 6:

PHYSICAL STABILITY OVER TIME OF EMULSION (17% w/v DISPERSED PHASE)

The physical stability of a PEGylated emulsion (17% w/v dispersed phase) loaded with docetaxel was monitored over time. The formulation was observed for signs of phase separation and droplet diameter measurements were made over time. The formulation was prepared as described in Examples 3 (emulsion 10) and 4. The formulation maintained approximately the same size and size distribution after 13 weeks on stability at ambient conditions (Table VII).

Table VII: Changes in mean droplet diameter size with time (17% w/v dispersed size) Emulsion Diameter (nm) PDI

Before homogenization (Day 0) 132 0.126

After homogenization (Day 0) 104 0.116

After incubation with DSPE-PEG₂₀OQ (Day 0) 110 0.116

After incubation with DSPE-PEG₂₀Q₀ (Day 1 ) 113 0.123

After incubation with DSPE-PEG₂Q₀Q (13 weeks) 125 0.109

EXAMPLE 7: PHYSICAL STABILITY OVER TIME OF EMULSION (50% w/v DISPERSED PHASE)

The physical stability of the sterilized docetaxel loaded emulsion (emulsion 9) with 50% w/v dispersed phase was monitored. The formulation was observed for signs of phase separation and droplet diameter measurements were made over time. The formulation maintained approximately the same size and size distribution after 1 month on stability at ambient conditions (Table VIII). Table VIII: Changes in mean droplet diameter size with time (50% w/v dispersed size) Emulsion Diameter (nm) PDI

Before homogenization (Day 0) 153 0.195

After homogenization (Day 0) 98 0.083 After homogenization (1 month) 112 0.170 EXAMPLE 8: PREPARATION OF EMULSIONS LOADED WITH 2',7-DILAUROYL-DOCETAXEL

2',7-dilauroyl-docetaxel/Labrafac™ CC/Macrogol 15 hydroxystearate (emulsion 11 )

Ingredients Amount (mg) Concentration (% w/v)

2',7-dilauroyl-docetaxel 45 0.30 Labrafac™ CC 1455 9.70 Macrogol 15 hydroxystearate

990 6.60 (Solutol* HS-15)

Total 2490 16.60

2',7-Dilauroyl-docetaxel was dissolved in ethyl alcohol to a concentration of 25 mg/ml at room temperature. A volume of 1800 μl of the solution was transferred to a 20-ml scintillation vial to evaporate ethyl alcohol under vacuum. The 2',7-dilauroyl-docetaxel (45 mg) was dissolved in Labrafac™ CC at 4O⁰C with stirring (630 rpm) for 30 min. The solution was cooled down at room temperature before adding macrogol 15 hydroxystearate (990 mg). The mixture was stirred (630 rpm) at 4O⁰C for 15 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After inclusion of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix as determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) was 141 nm, with PDI of 0.170. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 4 min to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 110 nm and 0.106 respectively.

2',7-dilauroyl-docetaxel/Labrafac™ CC//Macrogol 15 hydroxystearate (emulsion 12)

Ingredients Amount (mg) Concentration (% w/v)

2',7-dilauroyl-docetaxel 15 0.10 Labrafac™ CC 1485 9.90 Macrogol 15 hydroxystearate

990 6.60 (Solutol* HS-15)

Total 2490 16.60

2',7-Dilauroyl-docetaxel was dissolved in ethyl alcohol to a concentration of 25 mg/ml at room temperature. A volume of 600 μl of the solution was then transferred to a 20-ml scintillation vial to evaporate ethyl alcohol under vacuum. The 2',7-dilauroyl-docetaxel (15 mg) was dissolved in Labrafac™ CC at 4O⁰C with stirring (630 rpm) for 30 min. The solution was cooled down at room temperature before adding macrogol 15 hydroxystearate (990 mg). The mixture was stirred (630 rpm) at 4O⁰C for 15 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 15 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After inclusion of saline, the dispersed phase represented about 17% w/v of the emulsion. The diameter of the droplets in the premix as determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) was 142 nm, with PDI of 0.161. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 4 min to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 109 nm and 0.119 respectively.

2',7-dilauroyl-docetaxel/Labrafac™ CC/Macrogol 15 hvdroxystearate (emulsion 13)

Ingredients Amount (mg) Concentration (% w/v)

2',7-dilauroyl-docetaxel 35 0.27 Labrafac™ CC 650 5.00 Macrogol 15 hydroxystearate

429 3.30 (Solute/ HS-15)

Total 11 14 8.57

2',7-Dilauroyl-docetaxel was dissolved in ethyl alcohol to a concentration of 25 mg/ml at room temperature. A volume of 1400 μl of the solution was transferred to a 20-ml scintillation vial to evaporate ethyl alcohol under vacuum. The 2',7-dilauroyl-docetaxel (35 mg) was dissolved in Labrafac™ CC at 4O⁰C with stirring (630 rpm) for 30 min. The solution was cooled down at room temperature before adding macrogol 15 hydroxystearate (429 mg). The mixture was stirred (630 rpm) at 4O⁰C for 15 min. The dispersing phase (0.9% w/v NaCI in water) was then added to the mixture under agitation (630 rpm) until a final volume of 13 ml was reached. The formulation was heated/mixed at 4O⁰C for an additional 10 min (840 rpm). After inclusion of saline, the dispersed phase represented about 9 % w/v of the emulsion. The diameter of the droplets in the premix as determined by dynamic light scattering (Malvern Instruments Ltd, Malvern, UK) was 130 nm, with PDI of 0.144. This premix was then homogenized with a high-pressure homogenizer (Emulsiflex™-C3, Avestin, ON, Canada) at 10,000 psi for 3 min 30 s to reduce droplet size further. After homogenization, mean droplet diameter and PDI was 95 nm and 0.156 respectively. With this derivative higher concentrations of drugs in the oil phase can be achieved: in the present example it represents 15 mg out of a 1114 mg drug, lipid and surfactant dispersed phase.

Although the present invention has been described hereinabove by way of specific embodiments thereof, it can be modified, without departing from the spirit and nature of the subject invention as defined in the appended claims.

Claims

WHAT IS CLAIMED IS:

1. An oil-in-water emulsion comprising:

(a) at least one triglyceride oil;

(b) macrogol 15 hydroxystearate; and

(c) a dispersing aqueous phase, wherein the emulsion contains dispersed oil-containing droplets with a mean diameter of less than 200 nm.

2. The emulsion of claim 1 , wherein macrogol 15 hydroxystearate constitutes between about 2% to about 30% w/v of the emulsion.

3. The emulsion of any one of claims 1 and 2, wherein the dispersed oil-containing droplets constitute between about 5% and about 70% w/v of the emulsion.

4. The emulsion of any one of claims 1-3, wherein the dispersed oil-containing droplets constitute about 9% w/v of the emulsion.

5. The emulsion of any one of claims 1-4, wherein the triglyceride oil is a medium-chain triglyceride (MCT).

6. The emulsion of any one of claims 1-5, wherein the triglyceride oil comprises saturated fatty acids.

7. The emulsion of any one of claims 1-6, wherein the dispersing aqueous phase is: (a) water; (b) NaCI in water; (c) glycerol; (d) sorbitol; (e) dextrose; (f) xylitol; or (g) any combination of (a) -

(f).

8. The emulsion of any one of claims 1-6, wherein the dispersing aqueous phase is 0.9% w/v NaCI in water.

9. A pharmaceutical composition comprising the emulsion of any one of claims 1-8, and further comprising at least one hydrophobic therapeutic agent.

10. The pharmaceutical composition of claim 9, wherein the dispersed oil-containing droplets constitute about 17% w/v of the emulsion loaded with the hydrophobic therapeutic agent.

11. The pharmaceutical composition of claim 9, wherein the dispersed oil-containing droplets constitute about 50% w/v of the emulsion loaded with the hydrophobic therapeutic agent.

12. The pharmaceutical composition of any one of claims 9-11 , wherein the therapeutic agent is a taxane.

13. The pharmaceutical composition of claim 12, wherein the taxane is docetaxel or a derivative thereof.

14. The pharmaceutical composition of claim 12, wherein the taxane is docetaxel.

15. The pharmaceutical composition of claim 13, wherein the docetaxel derivative is an acyl halide mono-, di- or trisubstituted docetaxel.

16. The pharmaceutical composition of claim 15, wherein the docetaxel derivative is a 2', 7- dilauroyl-docetaxel.

17. The pharmaceutical composition of any one of claims 14 to 16, further comprising a lipid solubilizer for docetaxel or the docetaxel derivative.

18. The pharmaceutical composition of any one of claims 9-17, further comprising a thickening agent, wherein said thickening agent is: (a) glycerol; (b) poly(ethylene glycol) (PEG); (c) propylene glycol; (d) polyvinylpyrolidone; (e) dextran; or (f) any combination of (a) - (e).

19. The pharmaceutical composition of any one of claims 9-18, further comprising a co-solvent, wherein said co-solvent is: (a) poly(ethylene glycol) (PEG); (b) ethanol; (c) dimethylacetamide; (d) N-methylpyrrolidone; or (e) any combination of (a) - (d).

20. The pharmaceutical composition of any one of claims 9-19, further comprising an isotonic agent, wherein said isotonic agent is: (a) glycerol; (b) sorbitol; (c) xylitol; (d) NaCI; or (e) any combination of (a) - (d).

21. The pharmaceutical composition of any one of claims 9-20, further comprising an antioxidant, wherein said antioxidant is: (a) α-tocopherol; (b) deferoxamine mesylate; (c) ascorbic acid; or (d) any combination of (a) - (c).

22. The pharmaceutical composition of any one of claims 9-21 , further comprising a preservative.

23. The pharmaceutical composition of any one of claims 9-22, further comprising a pH- adjusting agent selected from the group consisting HCI and NaOH.

24. The pharmaceutical composition of any one of claims 9-23, for intravenous, subcutaneous, intraperitoneal, intramuscular, intratumoral, oral, nasal, sublingual, intrathecal or topical administration.

25. The pharmaceutical composition of claim 24, which is for intravenous administration, and which further comprises aqueous NaCI as a pharmaceutical carrier.

26. The pharmaceutical composition of any one of claims 9-25, further comprising a hydrophilic polymer-lipid derivative.

27. The pharmaceutical composition of claim 26, wherein the hydrophilic polymer-lipid derivative is a poly(ethylene glycol) (PEG)-lipid derivative.

28. The pharmaceutical composition of claim 27, wherein the PEG-lipid derivative constitutes about 1 to about 15 mol% of the total surface components of the emulsion.

29. The pharmaceutical composition of claim 27 or 28, wherein the PEG-lipid derivative is:

(a) 1 ,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000],

(b) 1 ,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-3000],

(c) 1 ,2-dimyristoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-5000],

(d) 1 ,2-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000],

(e) 1 ,2-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-3000],

(f) 1 ,2-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-5000],

(g) 1 ,2-distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000], (h) 1 ,2-distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-3000], (i) 1 ,2-distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-5000], 0) 1 ,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000], (k) 1 ,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-3000], (I) 1 ,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-5000], (m) N-octanoyl-sphingosine-1-[succinyl(methoxyPEG)-2000 or 5000]; or

(n) any combination of (a) - (m).

30. The pharmaceutical composition of claim 29, wherein the PEG-lipid derivative is 1 ,2- distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000].

31. A method of making an oil-in-water emulsion comprising:

(a) mixing a triglyceride oil with macrogol 15 hydroxystearate at a temperature between about 35°C and about 70°C to melt macrogol 15 hydroxystearate so as to form a first mixture;

(b) emulsifying the first mixture with a dispersing aqueous phase to obtain droplets with a mean diameter of less than 200 nm, whereby the droplets constitute about 5% to about 70% w/v of the emulsion.

32. A method of making a lipophilic therapeutic agent loaded oil-in-water emulsion comprising:

(a) mixing a triglyceride oil with the therapeutic agent so as to dissolve the therapeutic agent and form a first mixture;

(b) mixing the first mixture with macrogol 15 hydroxystearate at a temperature sufficient to melt macrogol 15 hydroxystearate so as to form a second mixture; and (c) emulsifying the second mixture with a dispersing aqueous phase to obtain an emulsion containing droplets with a mean diameter of less than 200 nm, whereby the droplets constitute about 5% to about 70% w/v of the emulsion.

33. The method of claim 32, wherein the therapeutic agent is a taxane.

34. The method of claim 33, wherein the taxane is docetaxel or a derivative thereof.

35. The method of claim 33, wherein the taxane is docetaxel.

36. The method of claim 34, wherein the docetaxel derivative is an acyl halide mono-, di- or trisubstituted docetaxel.

37. The method of claim 36, wherein the docetaxel derivative is 2',7-dilauroyl-docetaxel.

38. A method of making a lipophilic therapeutic agent loaded oil-in-water emulsion comprising:

(a) mixing a triglyceride oil with the therapeutic agent and macrogol 15 hydroxystearate so as to dissolve the therapeutic agent and melt macrogol 15 hydroxystearate and form a first mixture;

(b) emulsifying the first mixture with a dispersing aqueous phase to obtain an emulsion containing droplets with mean diameter of less than 200nm, whereby the droplets constitute about 5% to about 70% w/v of the emulsion.

39. The method of any one of claims 32-38, wherein the mixing in (a) is conducted at a temperature of between about 35⁰C and about 70⁰C.

40. The method of any one of claims 31-39, wherein the emulsifying is conducted at a temperature of between about 20⁰C and about 60°C.

41. The method of any one of claims 32-38, wherein the mixing in (a) and emulsifying are conducted at a temperature of between about 40⁰C and about 50⁰C, and wherein the therapeutic agent is docetaxel or a derivative thereof.

42. The method of claim 41 , wherein the therapeutic agent is docetaxel.

43. The method of claim 41 , wherein the docetaxel derivative is an acyl halide mono-, di- or trisubstituted docetaxel.

44. The method of claim 43, wherein the docetaxel derivative is 2',7-dilauroyl-docetaxel.

45. The method of any one of claims 31-44, wherein the mixing in (a) is conducted for about 15 minutes to about 90 minutes.

46. The method of any one of claims 31-45, wherein mixing in (a) is conducted at a velocity of about 400 rpm to about 800 rpm.

47. The method of any one of claims 31-46, wherein the triglyceride oil is a medium-chain triglyceride (MCT).

48. The method of any one of claims 31-45, wherein the triglyceride oil contains saturated fatty acids.

49. The method of any one of claims 31-48, wherein the dispersing aqueous phase is 0.9% w/v NaCI in water.

50. The method of any one of claims 31-49, wherein macrogol 15 hydroxystearate constitutes about 2% to about 30% w/v of the emulsion.

51. The method of any one of claims 31-50, wherein the droplets constitute about 17% w/v of the emulsion.

52. The method of any one of claims 31-50, wherein the droplets constitute about 50% w/v of the emulsion.

53. The method of any one of claims 31-52, further comprising homogenizing the emulsion to reduce the droplets size.

54. The method of claim 53, wherein the homogenizing is performed with a high-pressure homogenizer or a microfluidizer.

55. The method of any one of claims 32-54, further comprising a step of attaching a hydrophilic polymer-lipid derivative to the droplet interface of the emulsion.

56. The method of claim 55, wherein the step of attaching a hydrophilic polymer-lipid derivative is performed by adding an aqueous solution of the hydrophilic polymer-lipid derivative to the emulsion to form a hydrophilic polymerated emulsion, and incubating the hydrophilic polymerated emulsion at a temperature of about 20⁰C to about 60⁰C.

57. The method of claim 55, wherein the step of attaching a hydrophilic polymer-lipid derivative is performed by dissolving the hydrophilic polymer-lipid derivative in the dispersing aqueous phase prior to emulsifying, whereby emulsifying forms a hydrophilic polymerated emulsion, and incubating the hydrophilic polymerated emulsion at a temperature of about 40⁰C to about 60⁰C.

58. The method of claim 55, wherein the step of attaching a hydrophilic polymer-lipid derivative is performed by dissolving the hydrophilic polymer-lipid derivative in the first mixture, whereby emulsifying forms a hydrophilic polymerated emulsion, and incubating the hydrophilic polymerated emulsion at a temperature of about 40⁰C to about 60⁰C.

59. The method of any one of claims55-58, wherein the hydrophilic polymer-lipid derivative constitutes about 1 mol% to about 15 mol% of the total surface components of the emulsion.

60. The method of any one of claims 55-59, wherein the hydrophilic polymer-lipid derivative is poly(ethylene glycol) (PEG)-lipid derivative.

61. The method of claim 60, wherein the PEG-lipid derivative is:

(c) i ^-dimyristoyl-sn-glycero-S-phosphatidylethanolamine-N-tmethoxyCPEGJ-SOOO],

(e) i ^-dipalmitoyl-sn-glycero-S-phosphatidylethanolamine-N-tmethoxyCPEGVSOOO],

(f) 1,2-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-5000],

(g) 1 ,2-distearoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000], (h) i ^-distearoyl-sn-glycero-S-phosphatidylethanolamine-N-ImethoxyCPEGVSOOO], (i) i ^-distearoyl-sn-glycero-S-phosphatidylethanolamine-N-ImethoxyCPEGVδOOO], G) 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-2000], (k) 1 ,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-3000], (I) 1 ,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine-N-[methoxy(PEG)-5000], (m) N-octanoyl-sphingosine-HsuccinyKmethoxyPEG^OOO or 5000]; or

(n) any combination of (a) - (m).

62. The method of claim 61 , wherein the PEG-lipid derivative is 1 ,2-distearoyl-sn-glycero-3- phosphatidylethanolamine-N-[methoxy(PEG)-2000].

63. The method of any one of claims 31-62, wherein no organic solvent is used.

64. Use of the emulsion of any one of claims 1-8, in the making of a medicament.

65. Use of the pharmaceutical composition of any one of claims 9-30, in the making of a medicament.

66. Use of the pharmaceutical composition of any one of claims 12 -17 for the treatment of cancer in a subject.

67. Use of the pharmaceutical composition of any one of claims 12-17 for the preparation of a medicament for the treatment of cancer in a subject.

68. A method of improving the solubility of a lipophilic therapeutic compound comprising formulating the compound into the dispersed oil-containing droplets of the emulsion of any one of claims 1-8.

69. A method of increasing the circulation time of a lipophilic compound in vivo, comprising formulating the compound into the dispersed oil-containing droplets of the pharmaceutical composition of any one of claims 26-30.

70. A kit comprising:

(a) at least one hydrophobic therapeutic agent;

(b) at least one triglyceride oil; and

(c) macrogol 15 hydroxystearate.

71. The kit of claim 70, further comprising instructions for making a pharmaceutical composition.

72. The kit of claim 70 or 71 , wherein said hydrophobic therapeutic agent is a taxane.

73. The kit of claim 72, wherein the taxane is docetaxel or a derivative thereof.

74. The kit of claim 73, wherein the taxane is docetaxel.

75. The kit of claim 73, wherein the docetaxel derivative is a acyl halide mono-, di- or trisubstituted docetaxel.

76. The kit of claim 75, wherein the taxane is 2',7-dilauroyl-docetaxel.

77. The kit of any one of claims 70-73, further comprising instructions for treating cancer in a subject.