CN1764446A - Pharmaceutical preparations containing long-chain and medium-chain triglycerides - Google Patents

Abstract

Pharmaceutical formulations containing emulsifiers and medium and long chain triglycerides are described. In preferred embodiments, the long chain triglycerides eliminate or reduce the deleterious central nervous system effects caused by medium chain triglycerides.

Description

Pharmaceutical preparations containing long-chain and medium-chain triglycerides

related application

This application claims priority to the following and is hereby incorporated by reference in its entirety: U.S. Provisional Patent Application 60/2003, filed July 29, 2003, entitled "Ansamycin Formulations and Methods of Making and Using the Same" 491,050; U.S. Provisional Patent Application No. 60/478,430, filed June 12, 2003, entitled "Phospholipid-Based Formulations and Methods of Making and Using the Same," Ulm et al., filed March 13, 2003, and entitled " HSP90 Inhibitor Formulations and Data," U.S. Provisional Patent Application No. 60/454,812; and PCT Application PCT/US03/, filed April 4, 2003 by Ulm et al., entitled "Novel Ansamycin Formulations and Methods of Making and Using the Same" 10533, which claims priority to likewise titled US Provisional Patent Application 60/371,668, filed April 10, 2002.

technical field

The present invention relates generally to pharmaceutical formulations and methods, and in a more particular embodiment to emulsified formulations of ansamycins, such as 17-AAG.

Background technique

Included in the following description is information that may be helpful in understanding the present invention. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication specifically or impliedly mentioned is prior art.

17-allylamino-geldanamycin (17-AAG) is a synthetic analog of geldanamycin (GDM). Both molecules belong to a large class of antibiotic molecules known as ansamycins. GDMs, first isolated from the microorganism Streptomyceshygroscopicus and initially identified as potent inhibitors of certain kinases, were later shown to act by promoting kinase degradation, specifically targeting "molecular chaperones" such as heat Shock protein 90 (HSP90). Later, various other ansamycins also showed more or less this activity, among which 17-AAG is one of the most promising and intensive clinical studies currently being carried out by the National Cancer Institute (NCI) have Use it as a research object. See, e.g., Federal Register, 66(129):35443-35444; Erlichman et al., Proc. AACR (2001), 42, abstract 4474.

HSP90 is a ubiquitous chaperone protein that is involved in the folding, activation and assembly of many proteins, including key proteins involved in signal transduction, cell cycle control and transcriptional regulation. Researchers have reported that HSP90 chaperones are associated with important signaling proteins such as steroid hormone receptors and protein kinases, including, for example, Raf-1, EGFR, V-Src family kinases, Cdk4, and ErbB-2 (Buchner J., 1999, TIBS, 24: 136-141; Stepanova, L. et al., 1996, Genes Dev. 10: 1491-502; Dai, K. et al., 1996, J. Biol. Chem. 271: 22030-4 ). Studies further suggest that certain cochaperones, such as Hsp70, p60/Hop/Stil, Hip, Bagl, HSP40/Hdj2/Hsj1, immunophilins, p23, and p50, contribute to HSP90 function (see, e.g., Caplan, A., Trends in Cell Biol., 9:262-68 (1999)).

Ansamycin antibiotics, such as herbimycin A (HA), geldanamycin (GM), and 17-AAG, are thought to bind tightly to the N-terminal ATP-binding pocket of HSP90 exert its anticancer effect (Stebbins, C. et al., 1997, Cell, 89: 239-250). This pocket is highly conserved and has weak homology with the ATP-binding site of DNA gyrase (Stebbins, C. et al., supra; Grenert, J.P. et al., 1997, J. Biol. Chem ., 272:23843-50). In addition, it has been shown that both ATP and ADP bind to the pocket with low affinity and have weak ATPase activity (Proromou, C. et al., 1997, Cell, 90:65-75; Panaretou, B. et al. ., 1998, EMBO J., 17:4829-36). In vitro and in vivo studies have shown that occupation of the N-terminal pocket by ansamycin and other HSP90 inhibitors alters HSP90 function and inhibits protein folding. At high concentrations, ansamycin and other HSP90 inhibitors can prevent the binding of protein substrates to HSP90 (Scheibel, T.H. et al., 1999, Proc. Natl. Acad. Sci. USA 96:1297-302; Schulte, T.W. et al., 1995, J.Biol.Chem.270:24585-8; Whitesell, L., et al., 1994, Proc.Natl.Acad.Sci.USA 91:8324-8328). It has also been shown that ansamycin can inhibit the ATP-dependent release of chaperone-associated protein substrates (Schneider, C.L.et al., 1996, Proc.Natl.Acad.Sci.USA, 93:14536-41; Sepp-Lorenzino et al. al., 1995, J. Biol. Chem. 270:16580-16587). In either case, the substrate is degraded by a ubiquitin-dependent process present in the proteasome (Schneider, C., L., supra; Sepp-Lorenzino, L., et al., 1995, J. Biol .Chem.270:16580-16587; Whitesell, L. et al., 1994, Proc.Natl.Acad.Sci.USA, 91:8324-8328).

The destabilization of this substrate occurs in tumors and non-transformed cells, and it is shown to be particularly effective for a subset of signal regulators, such as Raf (Schulte, T.W.et al., 1997, Biochem.Biophys.Res.Commun.239: 655-9; Schulte, T.W., et al., 1995, J.Biol.Chem.270:24585-8), nuclear steroid receptor (Segnitz, B., and U.Gehring.1997, J.Biol.Chem.272 : 18694-18701; Smith, D.F.et al., 1995, Mol.Cell.Biol.15: 6804-12), v-src (Whitesell, L., et al., 1994, Proc.Natl.Acad.Sci. USA 91:8324-8328) and certain transmembrane tyrosine kinases (Sepp-Lorenzino, L.et al., 1995, J.Biol.Chem.270:16580-16587), such as EGF receptor (EGFR) and Her2/Neu (Hartmann, F., et al., 1997, Int. J Cancer 70: 221-9; Miller, P. et al., 1994, Cancer Res. 54: 2724-2730; Mimnaugh, E.G., et al. , 1996, J.Biol.Chem.271:22796-801; Schnur, R.et al., 1995, J.Med.Chem.38:3806-3812), CDK4 and mutant p53. Erlichman et al., Proc. AACR (2001), 42, abstract 4474. Ansamycin-induced loss of these proteins results in selective disruption of certain regulatory pathways and causes growth arrest at specific phases of the cell cycle (Muise-Heimericks, R.C. et al., 1998, J. Biol. Chem. 273: 29864-72), and apoptosis and/or differentiation of treated cells (Vasilevskaya, A. et al., 1999, Cancer Res., 59:3935-40).

Recently, WO01/72779 by Nicchitta et al. (PCT/US01/09512) showed that HSP90 can be assumed to assume a different conformation after heat shock and/or binding by the fluorophore bis-ANS. Specifically, Nicchitta et al. demonstrated that this induced conformation, compared to the different forms of HSP90 that predominate in normal cells, displays a higher affinity for certain HSP90 ligands. Commonly owned application PCT/US02/39993 further supports this finding by demonstrating the use and application of cancer cell lysates as an excellent source of high affinity HSP90.

In addition to anti-cancer and anti-tumor activity, HSP90 inhibitors are also involved in a variety of other applications, including as anti-inflammatory drugs, anti-infectious drugs, drugs for the treatment of autoimmunity, drugs for the treatment of stroke, ischemia, heart disease and Drugs that help promote nerve regeneration (see, e.g., Rosen et al., WO02/09696 (PCT/US01/23640); Degranco et al., WO99/51223 (PCT/US99/07242); Gold, U.S. Patent 6,210,974 B1; DeFranco et al., US Patent 6,174,875). Overlapping some of the above, it has also been reported in the literature for the treatment of fibrogenetic diseases including but not limited to scleroderma, polymyositis, systemic lupus erythematosus, rheumatoid arthritis, cirrhosis, Keloid formation, interstitial nephritis, and pulmonary fibrosis. (Strehlow, WO02/02123; PCT/US01/20578). In addition, modulations, modulators, and applications of HSP90 have been reported in: PCT/US03/04283, PCT/US02/35938, PCT/US02/16287, PCT/US02/06518, PCT/US98/09805, PCT/US02/ US00/09512, PCT/US01/09512, PCT/US01/23640, PCT/US01/46303, PCT/US01/46304, PCT/US02/06518, PCT/US02/29715, PCT/US02/35069, PCT/US02/ 35938, PCT/US02/39993, 60/293,246, 60/371,668, 60/331,893, 60/335,391, 06/128,593, 60/337,919, 60/340,762, and 60/359,484.

At present, like many other lipophilic drugs, ansamycin is difficult to prepare into pharmaceutical preparations, especially preparations for intravenous injection. To date, attempts have been made to prepare them as lipid vesicles and oil-in-water emulsions, but these all require complex processing steps, harsh or clinically unacceptable solvents and/or cause formulation instability. See generally Vemuri, S. and Rhodes, C.T., Preparation and characterization of liposomes astherapeutic delivery systems: a review, Pharmaceutica Acta Helvetiae 70, pp.95-111 (1995); see also June 29, 2000 Publication No. PCT/US99/30631 of WO00/37050. Co-owned application PCT/US03/10533, from which this application claims priority, describes emulsions containing medium chain triglycerides. However, the medium-chain fatty acids and triglycerides that carry them may result in the metabolism of caprylate, resulting in undesirable central nervous system effects such as drowsiness, nausea, drowsiness, and EEG changes. See Cotter et al., Am. J. Clin. Nutr. 50:794-800 (1989); Miles et al., Journal of Parenteral and Enteral Nutrition 15:37-41 (1991); Traul et al., Food Chem. Toxicol. 38:79-98 (2000). To date, such side effects have been partly ameliorated by the use of long-chain fatty acids in the form of nutritional supplements, by virtue of the fact that long-chain fatty acids compete with a higher incorporation efficiency for the key octanoate pathway enzymes. However, to the applicant's knowledge so far, they have not been used in combination with medium chain triglycerides and ansamycins.

Therefore, there is a need for additional formulations and methods for their preparation, which are relatively simple to manufacture and which ameliorate one or several of the above-mentioned disadvantages generally caused by medium-chain triglycerides.

Summary of the invention

By formulating long chain triglycerides together with the compound as a component of the formulation, applicants have found that the formulations claimed herein can be used for better tolerated intravenous administration of lipophilic compounds such as ansamycins.

In a first aspect, the invention features a pharmaceutical composition comprising a pharmaceutically active compound, such as ansamycin, such as 17-AAG, and an emulsifier (such as a phospholipid, such as that found in lecithin ) in combination with oil. The oil can and preferably contains long chain triglycerides. The composition may also contain medium chain triglycerides. Emulsifiers and oils together make up the fat phase.

In certain embodiments, the lipid phase comprises 5-30% w/w, preferably 5-20% w/w of the total formulation.

In certain embodiments, the total w/w percentage of long chain triglycerides does not exceed 10%, more preferably in the range of 7% or less, more preferably in the range of 6% or less, to accommodate viscosity constraints.

In certain embodiments, the w/w ratio of medium chain triglycerides to long chain triglycerides is from 10:0.0001 to 0.0001:10, more preferably from 10:1 to 1:10.

In certain embodiments, the phospholipids comprise 3-10% w/w of the total weight.

In certain embodiments, triglycerides comprise 5-20% w/w of the total weight.

In certain embodiments, the triglycerides are, or are at least partially in, the form of naturally occurring oils, such as vegetable oils, such as soybean oil, sesame oil, safflower oil, and corn oil.

In certain embodiments, the composition further comprises one or more of water, preservatives (e.g., sodium edetate), cryoprotectants, buffers, chelating agents, and tonicifiers .

In certain embodiments, the drug is 17-AAG (17-allylaminogeldanamycin, 17-allylamino-17-demethoxy-geldanamycin) and the amount is 0.5 mg/ml-4mg/ml or 0.05%w/w-0.4%w/w of the total formulation weight.

In one embodiment, the composition comprises the following ingredients: 2 mg/ml 17-AAG, 3.3% soybean oil, 6.6% lecithin, 9.9% Miglyol 812N, 7.15% sucrose, and water.

In another embodiment, the composition comprises the following ingredients: 2 mg/ml 17-AAG, 7.5% lecithin, 15% Miglyol 812N, 10% sucrose, and water.

17-AAG is 17-allylaminogeldanamycin with the following structure:

In certain embodiments, the compositions of the present invention also comprise short chain triglycerides.

In a particularly preferred embodiment, the composition comprises a sufficient amount of long-chain triglycerides to reduce or avoid the occurrence of medium-chain triglyceride-mediated central nervous system (CNS) effects, especially in the composition Also included in the case of medium chain triglycerides. CNS effects are generally negative, undesired effects including, but not limited to, one or more of drowsiness, nausea, drowsiness, and EEG changes. However, in certain embodiments such effect(s) may be desirable in a given context such that the amount of medium chain triglycerides relative to long chain triglycerides is increased.

In certain embodiments, the composition is frozen and/or lyophilized, as outlined in PCT/US03/10533. In the lyophilized embodiment, the weight percent of each component of triglycerides, phospholipids, drug and non-volatile components must be increased and exceeded the above ranges to comply with the loss of the initially present but lost after lyophilization. After water and other volatile components their relative fractions increase.

In certain embodiments, the compositions can also be formulated and/or stored under inert gas conditions, eg, in dark and/or light-tight bottles, vials, or ampoules.

Combinations of any of the above-mentioned embodiments are also contemplated where appropriate.

In another aspect, the invention features a method of reducing central nervous system effects mediated by medium chain triglycerides in a patient comprising: providing a pharmaceutical formulation comprising a pharmaceutically active agent and long chain triglycerides, wherein The amount of said long-chain triglycerides is sufficient to reduce or avoid the occurrence of central nervous system effects mediated by medium-chain fatty acids, and the above-mentioned products are administered to patients. Embodiments of this aspect may refer to any composition of the above aspects

Embodiments and combinations thereof.

In another aspect, the invention features methods of using pharmaceutical compositions, formulations, and methods and products for treating or preventing diseases in organisms, such as mammals, by administering a pharmaceutically effective amount of the product to the organism. The disease, at least in the case of mammals, is preferably selected from the group consisting of ischemic, proliferative and neurological injuries and comprises an HSP90 inhibitor, such as one or more ansamycins, as the pharmaceutically active agent. Proliferative diseases include, but are not limited to, tumors and cancers, inflammatory diseases, fungal infections, yeast infections, and viral infections. In certain preferred embodiments, the mammal is a human. In certain preferred embodiments, the mode of administration is intravenous, however, other modes of administration are contemplated as described in more detail below.

According to specific embodiments, the advantages of the present invention include one or more of the following advantages: ease of preparation, use of clinically acceptable reagents (such as can reduce toxicity to the environment and/or patients), improved stability of the formulation, ease of shipment and storage, formulation and bedside simplicity, intravenous and systemic tolerability when administered, and avoidance of certain undesirable side effects often caused by medium chain fatty acids and triglycerides in the body. Other advantages, aspects and embodiments will be apparent from the following figures, detailed description and claims.

Brief description of the drawings

Figure 1 shows the reduction in lethargy in rats due to the addition of long chain fatty acids to formulations containing medium chain triglycerides.

Description of the preferred embodiment

The formulations of the present invention have particular advantages in water-soluble drugs suitable for intravenous and other administration to patients. The preparation method is relatively simple, usually using clinically acceptable reagents, and the prepared product has advantages over existing methods and products in terms of storage, stability and biological tolerance.

definition

The following terms have the following meanings, and terms that are not specified below also have the usual meanings in this field:

The term "pharmaceutically active compound" has the same meaning as "drug", and refers to any compound that directly or indirectly exerts a biological effect when administered to cultured cells or organisms in vivo or in vitro. The drug is preferably capable of being encapsulated in liposomes and/or emulsified, and is usually lipophilic, although not required.

The terms "evaporation" and "lyophilization" do not imply that 100% of the solvent and solution must be removed, only a lesser proportion may be removed. However, in lyophilized embodiments, substantial removal is preferred, preferably about 95% or more.

By "inert gas conditions" is meant gas conditions that are relatively less reactive than gases in standard atmospheric conditions. An example of such an inert gas condition is the use of pure or substantially pure nitrogen during formulation. Other situations are also well known to those skilled in the art.

The term "hydration" or "rehydration" refers to the addition of an aqueous solution, such as water or a physiologically compatible buffer, such as Hanks' solution, Ringer's solution, or physiological saline buffer.

The term "about" is intended to encompass a range of deviations of about 20% from the stated value. The term "comprising" when used in conjunction with "between" or "between about" is meant to include the endpoints of the stated range.

The term "ansamycin" is a broad term characterizing compounds with a "loop" structure comprising one of benzoquinone, benzohydroquinone, naphthoquinone or naphthohydroquinone moieties bridged by long chains one. Examples of naphthoquinone or naphthohydroquinone compounds are the clinically important drugs rifampicin and rifamycin, respectively. Examples of benzoquinones are geldanamycin (including its synthetic derivatives, 17-allylamino-17-demethoxygeldanamycin (17-AAG) and 17-N,N-di Methylamino-ethylamino-17-demethoxygeldanamycin (DMAG)), dihydrogeldanamycin and herbimycin. An example of a benzohydroquinone is macbecin. Ansamycins and benzoquinone ansamycins according to the invention may be synthetic, naturally occurring, or a combination of both, i.e. "semi-synthetic", and may include dimers and conjugated variants and prodrugs form. Some typical benzoquinone ansamycins and methods for their preparation useful in the present invention include, but are not limited to, those described in US Pat. Geldanamycin is also commercially available, for example, from CN Biosciences, an affiliate of Merck KGaA, Darmstadt, Germany, headquartered in San Diego, California, USA (Cat. No. 345805). Biochemical purification of geldanamycin derivative 4,5-dihydrogeldanamycin and its hydroquinone from the culture of Streptomyces hygroscopicus (ATCC55256) was published in 1993 by the applicant as Pfizer Inc and the inventor as Cullen et al. On July 22, 2009, the publication number was WO93/14215, and the international application number was PCT/US92/10189, which was described in the application; the synthesis of 4,5-dihydrogeldanamycin by the catalytic hydrogenation of geldanamycin An alternative method is also known. See, e.g., Progress in the Chemistry of Organic Natural Products, Chemistry of the Ansamycin Antibiotics, 33:278 (1976). Other ansamycins useful in various embodiments of the invention are described in the literature cited in the "Background" section above.

"Oil" includes fatty acids and glycerides containing fatty acids, such as monoglycerides, diglycerides and triglycerides known in the art. The fatty acids and glycerides used in the present invention may be saturated and/or unsaturated, natural and/or synthetic, charged or neutral. "Synthetic" can be fully synthetic or semi-synthetic, as these terms are known in the art. Oils may also be homogeneous or inhomogeneous in composition and/or origin.

Here, "medium chain triglycerides" are triglyceride compositions containing fatty acids with a length of 8-12 linear carbon atoms, more preferably 8-10 carbon atoms in length. Various embodiments of the present invention involve the use of Miglyol(R) 812 supplied by CONDEA (Cranford, NJ, USA). Miglyol(R) 812 contains approximately 50-65% caprylic acid (8 carbon atoms) and 30-45% capric acid (10 carbon atoms). Caproic acid (6 carbon atoms) can also be present, up to about 2%, as can lauric acid (12 carbon atoms). Myristic acid (14 carbon atoms) is present in smaller amounts (up to 1%). Condea also offers Miglyol® 810, 818, 829 and 840, and it is envisioned that one or more of these other Miglyol® solutions and other medium chain triglyceride solutions may also be used with more or less success in each of the present invention. aspects and implementations. As for the latter, those skilled in the art are familiar with their properties, sources and/or methods of preparation, and can obtain or prepare them without undue research or experimentation. Miglyol 812N is featured in European Pharmacopoeia as medium chain triglyceride, British Pharmacopoeia as fractionated coconut oil, and Japanese Pharmacopoeia as caprylic/capric triglyceride. Other sources of medium-chain triglycerides include coconut oil, palm kernel oil, and butter.

"Short chain triglycerides" are triglyceride compositions containing fatty acids with a length of less than 8 linear carbon atoms.

"Long chain triglycerides" are triglyceride compositions containing fatty acids with a length greater than 12 linear carbon atoms. Their common source is vegetable oil, such as soybean oil, which usually contains 55-60% linoleic acid (9,12-octadecadienoic acid), 22% oleic acid (cis-9-octadecenoic acid), and small amounts of palmitic and stearic acids.

The terms "short", "medium" and "long" may also be used for individual fatty acids, in which case their definitions include fewer than 8 straight-chain carbon atoms, 8-12 straight-chain carbon atoms and greater than 12 straight-chain carbon atoms.

"Emulsifier" is synonymous with "surfactant" and includes, but is not limited to, phospholipids such as lecithin. "Lecithin" is a naturally occurring mixture of diglycerides of stearic, palmitic and oleic acids attached to phosphorylcholine esters. The term surfactant or emulsifier also includes phosphatidylcholines, compounds of this type are well known. A preferred emulsifier for use in the present invention is soy lecithin, such as Phospholipon 90G supplied by American Lecithin Company (Oxford, CT, USA). Phospholipon 90G has been used in nutritional products for parenteral administration earlier, such as about 1.2% in Intralipid®, about 1% in Doxil® (doxorubicin), and about 1% in Ambisome® (amphotericin). About 2% in B) and about 1.2% in Propofol(R). Regarding the latter, see, eg, US Patent 6,140,374. The surfactant/emulsifier is typically present at a concentration of about 0.5-25% w/v, based on the amount of water and/or other components that dissolve the surfactant. The surfactant concentration is preferably about 0.5-10% w/v, most preferably about 1-8% w/v.

Examples of anionic surfactants include sodium lauryl sulfate, triethanolamine lauryl sulfate, sodium polyoxyethylene lauryl ether sulfate, sodium polyoxyethylene nonylphenyl ether sulfate, triethanolamine polyoxyethylene lauryl ether sulfate, Ether Sulfate, Sodium Cocoyl Sarcosinate, Sodium N-Cocoyl Methyl Taurate, Sodium Polyoxyethylene (Coconut) Alkyl Ether Sulfate, Sodium Diether Hexyl Sulfosuccinate, Alpha Olefin Sulfonate Sodium lauryl phosphate, sodium lauryl phosphate, sodium polyoxyethylene lauryl ether phosphate, perfluoroalkyl carboxylate (manufactured by Daikin Industries Ltd., trade name UNIDINE DS-101 and 102).

Examples of cationic surfactants include dialkyl (C12-C22) dimethyl ammonium chloride, alkyl (coconut) dimethyl benzyl ammonium chloride, stearylamine acetate, tetradecylamine acetate, taurine Oleyl propylenediamine imine acetate, octadecyl trimethyl ammonium chloride, alkyl (tallow) trimethyl ammonium chloride, dodecyl trimethyl ammonium chloride, alkyl ( Coconut) Trimethylammonium Chloride, Cetyltrimethylammonium Chloride, Biphenyltrimethylammonium Chloride, Alkyl (Tallow)-Imidazolium Quaternary Salt, Tetradecylmethylbenzyl Chloride Ammonium Octadecyldimethylbenzyl Ammonium Chloride, Dioleyl Dimethyl Ammonium Chloride, Polyoxyethylene Lauryl Monomethyl Ammonium Chloride, Polyoxyethylene Alkyl (C ₁₂ -C ₂₂ ) Benzyl Ammonium Chloride ammonium chloride, polyoxyethylene lauryl monomethyl ammonium chloride, 1-hydroxyethyl-2-alkyl (tallow)-imidazoline quaternary salt, and silicones containing siloxane groups as hydrophobic groups Cationic surfactant, a fluorine-containing cationic surfactant containing a fluoroalkyl group as a hydrophobic group (manufactured by Daikin Industries Ltd, trade name UNIDINE DS-202).

Examples of nonionic surfactants include polyoxyethylene lauryl ether, polyoxyethylene tridecyl ether, polyoxyethylene cetyl ether, polyoxyethylene polyoxypropylene cetyl ether, polyoxyethylene Octadecanoyl ether, polyoxyethylene oleyl ether, polyoxyethylene nonylphenyl ether, polyoxyethylene octylphenyl ether, polyoxyethylene monolauric acid, polyoxyethylene monostearic acid, polyoxyethylene mono Oleic acid, sorbitan monolaurate, sorbitan monostearate, sorbitan monopalmitate, sorbitan monostearate, sorbitan monooleate, sorbitan sesqui Oleate, sorbitan trioleate, polyoxyethylene sorbitan monolaurate, polyoxyethylene sorbitan monopalmitate, polyoxyethylene sorbitan monostearate, polyoxyethylene sorbitan monostearate Sorbitan monooleate, polyoxyethylene polyoxypropylene block polymer, polyglycerol fatty acid ester, polyether-modified silicone oil (manufactured by Toray DowCorning Silicone Co., Ltd., trade names SH3746, SH3748, SH3749 and SH3771), perfluoroalkyl ethylene oxide adducts (manufactured by Daikin Industries Ltd, trade names UNIDINE DS-401 and DS-403), fluoroalkyl ethylene oxide adducts (manufactured by Daikin Industries Ltd manufactured by Daikin Industries Ltd under the trade name UNIDINE DS-406), and perfluoroalkyl oligomers (manufactured by Daikin Industries Ltd under the trade name UNIDINE DS-451).

"Physiologically acceptable carrier" means a carrier or diluent that does not cause significant irritation to the organism and does not abolish the biological activity and properties of the administered compound.

"Excipient" refers to a substance added to a pharmaceutical composition to further facilitate the administration of the compound. Examples of excipients include, but are not limited to, calcium carbonate, calcium phosphate, various sugars and different types of starch, cellulose and cellulose derivatives, gelatin, vegetable oils, and polyethylene glycols. These may also be physiologically acceptable carriers, such as sucrose, as described above. Further falling within the definition of excipients are fillers. "Bulking agents" generally provide mechanical support to lyophilic formulations by keeping the matrix of the dry formulation in its conformation. Sugar is preferred. As used herein, sugars include, but are not limited to, monosaccharides, disaccharides, oligosaccharides, and polysaccharides. Specific examples include, but are not limited to, fructose, glucose, mannose, trehalose, sorbose, xylose, maltose, lactose, sucrose, dextrose, and dextran. Sugar also includes sugar alcohols such as mannitol, sorbitol, inositol, galactitol, xylitol and arabitol. Mixtures of sugars may also be used according to the invention. Various fillers, such as glycerol, sugars, sugar alcohols, and mono- and disaccharides can also function as isotonic agents. Preferably the filler is chemically inert to the drug and has no or very limited deleterious side effects or toxicity under the conditions of use. In addition to filler carriers, other carriers may or may not serve the purpose of fillers, including, adjuvants and diluents, as known and readily available in the art.

A preferred bulking agent of the present invention is sucrose. Without being bound by any theory, it is believed that sucrose forms an amorphous glass-like mass upon freezing and subsequent lyophilization, thereby enhancing the possible stability of the formulation by forming dispersed oil droplets containing the active ingredient within the hard glass-like mass . Stability can also be enhanced as the sugar replaces the water lost during lyophilization. Instead of water molecules, sugar molecules are hydrogen bonded to the phospholipids at the interface. Other fillers that have these characteristics and can be substituted include, but are not limited to, cellulosic products such as corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth, methylcellulose, hydroxypropylmethyl Cellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone (PVP). If desired, disintegrants such as cross-linked polyvinylpyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate may also be added.

The phrase "an amount that reduces or avoids the occurrence of central nervous system effects mediated by medium chain triglycerides" in the claims can be empirically determined without undue experimentation by one of ordinary skill using the starting points described herein.

emulsification

Emulsions comprising an oily phase and an aqueous phase are known in the art as carriers for therapeutically active ingredients or as sources of parenteral nutrition. Emulsions may be of the oil-in-water or water-in-oil type. As in the present case, if the therapeutic ingredient is particularly soluble in the oil phase, the oil-in-water form is the preferred embodiment. Simple emulsions are thermodynamically unstable systems in which the oil and water phases separate (oil droplets coalesce). The key to reducing the coalescence process to a slight level is the addition of emulsifiers to the emulsion.

To prevent or minimize oxidative degradation reactions or lipid peroxidation reactions, in addition to or as an alternative to deoxygenation (e.g., placing the formulation under inert gases such as nitrogen and argon, and/or using light-resistant containers), It is also possible to add, for example, alpha-tocopherol and butylated benzyl alcohol, and preservatives such as ethylenediaminetetraacetate.

Emulsification can be achieved by various techniques well known in the art, such as mechanical mixing, homogenization (eg, using polytron or Gaulin high energy type equipment), vortexing, and sonication. Sonication can be performed using bath-type or probe-type equipment. Microfluidizers are available, for example, from Microfluidics Corp., Newton, Mass., further described in US Patent 4,533,254, and utilize pressure assist through narrow orifices. Pressure assisted extrusion can also be used to pass through a variety of commercially available polycarbonate membranes. Low pressure devices are also available. These high and low pressure devices can be used to select and/or adjust the size of vesicles.

Sterilize by filtration. Filtration may include pre-filtration through a larger diameter filter, such as a 0.45 micron filter, and then through a smaller filter, such as a 0.2 micron filter. A preferred filter medium is cellulose acetate (Sartorius-Sartobran ^™ ).

freeze-dried

Lyophilization is the removal or substantial removal of liquid from a sample, for example by liters, as described above in the section entitled "Removal of Solvents".

Characterization and assessment of physical and chemical stability

Phospholipids and their degradation products can be extracted from the emulsion and then measured. The liquid mixture is then separated using a two-dimensional thin layer chromatography (TLC) system or a high performance liquid chromatography (HPLC) system. In TLC, spots corresponding to individual components can be taken to determine the amount of phosphorus. The total amount of phosphorus in the sample can be determined quantitatively, eg by measuring the intensity of the blue color formed relative to water at 825 nm with a spectrophotometer. In HPLC, phosphatidylcholine (PC) and phosphatidylglycerol (PG) can be separated and precisely quantified. Lipids can be detected in the 203-205 nm region. In the 200nm wavelength region of the ultraviolet spectrum, unsaturated fatty acids show higher absorbance, while saturated fatty acids show lower absorbance. As an example, Vemuri and Rhodes, supra, describe the separation of PC and PG in egg yolk in Licrosorb Diol and Licrosorb S1-60. The mobile phase used for separation was acetonitrile-methanol containing 1% hexane-water (74:16:10 v/v/v). After 8 min, the separation of PG from PC was observed. The retention times were approximately 1.1 and 3.2 minutes, respectively.

The appearance, mean droplet diameter, and size distribution of the emulsion are important parameters for observation and maintenance. These parameters can be evaluated in a number of ways. For example, dynamic light scattering and electron microscopy techniques can be employed. See, eg, Szoka and Papahadjopoulos, Annu. Rev. Biophys. Bioeng., 9:467-508 (1980). In particular, morphological characterization can be performed using freeze-fracture electron microscopy. Lower performance optical microscopes are also available.

The emulsion droplet size distribution can be measured, for example, using a particle size distribution analyzer, such as the CAPA-500 manufactured by Horiba Limited (Ann Arbor, MI, USA), the Coulter Particle Sizer (Beckman Coulter Inc., Brea, CA, USA) ), or Zetasizer (Malvern Instruments, Southborough, MA, USA).

Stability testing using HPLC

Similar to the method described above for lipid fractions in emulsions, the chemical stability of therapeutically active ingredients such as 17-AAG can also be assessed by HPLC after extraction of the emulsion. Specific assays could be developed to separate therapeutically active ansamycin from its degradation products. The extent of degradation can be assessed by a decrease in the HPLC peak signal associated with the therapeutically active ansamycin and/or by an increase in the HPLC peak signal associated with the degradation product. Ansamycin has an easily and clearly detected maximum absorbance at 345 nm compared to other components in the emulsion components.

Formulation and Administration

Although intravenous administration is preferred in various aspects and embodiments of the invention, those skilled in the art will appreciate that this method can also be modified or easily adapted to other modes of administration, such as oral, aerosol, Parenteral, subcutaneous, intramuscular, intraperitoneal, rectal, vaginal, intratumoral, or peritumoral administration. Much of what follows is known to those skilled in the art, but is nonetheless provided as background to illustrate other possibilities of the invention. It is understood that what follows will overlap with the discussion above.

Pharmaceutical compositions can be prepared by conventional methods of mixing, dissolving, granulating, sugar-coating, levigating, emulsifying, encapsulating, encapsulating or lyophilizing.

Pharmaceutically acceptable compositions can be formulated using conventional methods using one or more physiologically acceptable carriers including excipients and auxiliaries which facilitate processing of the active compounds into preparations which can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen. Some excipients and their use in the preparation of formulations have been described. Others are known in the art, such as PCT/US99/30631, Remmington's Pharmaceutical Sciences, Meade Publishing Co., Easton, PA (latest edition) and Goodman and Gilman's The Pharmaceutical Basis of Therapeutics, Pergamon Press, New York, N.Y. (latest edition).

For injection, the drug can be formulated in aqueous solution, preferably in physiologically compatible buffers such as Hanks' solution, Ringer's solution, or physiological saline buffer well known in the art. For transmucosal administration, penetrants are used in the formulation appropriate to the barrier to be penetrated. These penetrants are well known in the art.

The above-mentioned formulations of the present invention and their hydration of the lyophilized cake are well suited for immediate or near-immediate parenteral administration by injection, such as bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, eg, contained in ampoules or in multi-dose containers, with an added preservative, eg, ethylenediaminetetraacetate. As previously discussed, the pharmaceutical compositions of the present invention may be stored in an inert environment, such as ampoules or other light-resistant or oxygen-resistant packaging.

dose range

A phase I pharmacology study of 17-AAG in adult patients with advanced solid tumors, administered every three weeks for 5 days by daily 1-hour infusion, established a maximum tolerated dose (MTD) of 40 mg/m ² . Wilson et al., Am. Soc. Clin. Oncol., abstract, Phase IP Pharmacologic Study of 17-(Allylamino)-17-Demethoxygeldanamycin (AAG) in Adult Patients with Advanced Solid Tumors (2001). In this study, mean +/- SD values for terminal half-life, clearance, and steady-state volume were determined to be 2.5 +/- 0.5 hours, 41.0 +/- 13.5 L/hour, and 86.6 +/- 34.6 L/m ² . Plasma C _max levels were determined to be 1860+/-660nM and 3170+/-1310nM at doses of 40 and 56mg/ ^m2 , respectively. Using these values as a guide, useful patient doses of the active ingredient in the formulations of the invention can be expected to range from about 0.40 mg/m ² to 4000 mg/m ² . ^M2 represents the body surface area. Convert mg/ ^m2 to mg drug/kg body weight using standard algorithms.

The following examples are illustrative only, and the components and steps herein are not intended to limit the invention unless specifically stated in the claims. Examples 1-5 and 9 are borrowed from commonly-owned PCT application PCT/US03/, filed April 4, 2003, entitled "Novel Ansamycin Formulations and Methods of Making and Using the Same" from which this application claims priority. 10533.

Example

Example 1: Preparation of 17-AAG; Alternative 1

To 45.0 g (80.4 mmol) of geldanamycin dissolved in 1.45 L of dry THF in a 2 L dry flask was added dropwise 36.0 mL (470 mmol) of allylamine dissolved in 50 mL of dry THF over 30 minutes . The reaction mixture was stirred at room temperature under nitrogen for 4 hours until TLC analysis indicated that the reaction was complete [GDM: bright yellow: Rf = 0.40; (5% MeOH-95% _CHCl3 ); 17-AAG: purple: Rf = 0.42 (5 %MeOH-95% _CHCl3 )]. The solvent was removed by rotary evaporation, and the crude product was slurried in 420 mL H2O _: EtOH (90:10) at 25 °C, then filtered, and dried at 45 °C for 8 h to afford 40.9 g (66.4 mmol) of purple crystals of 17 - AAG (82.6% yield, >98% purity by HPLC at 254 nm). The melting point measured by differential scanning colorimetry (DSC) is 206-212°C. The results obtained by ¹ H NMR and HPLC were consistent with the desired product.

Embodiment 2: Preparation of low melting point type 17-AAG

An alternative method of purification is to dissolve the crude 17-AAG from Example 1 in 800 mL of 2-propanol (isopropanol) at 80 °C and then cool to room temperature. Filtration followed by drying at 45° C. for 8 hours afforded 44.6 g (72.36 mmol) of 17-AAG as purple crystals (90% yield, >99% purity by HPLC at 254 nm). The melting point measured by differential scanning colorimetry (DSC) is 147-175°C. The results obtained by ¹ H NMR and HPLC were consistent with the desired product.

Example 3: Preparation of low melting point type 17-AAG, alternative 1

An alternative method of purification was to slurry the product 17-AAG of Example 2 in 400 mL of H2O _: EtOH (90:10) at 25°C, filter, and dry at 45°C for 8 hours to yield 42.4 g (68.6 mmol) purple crystals of 17-AAG (95% yield, >99% purity by HPLC at 254 nm). The melting point is 147-175°C. The results obtained by ¹ H NMR and HPLC were consistent with the desired product.

Embodiment 4: Preparation of 17-AAG emulsion

17-AAG obtained in any one of Examples 1 to 3 above was dissolved in ethanol. The table below shows a 4000 gm batch made according to one embodiment of the invention. Those skilled in the art will appreciate that the procedure can be scaled up or down, the amounts of the individual components can be changed, etc., and that components not listed can also be added.

component %w/w Grams in a 4000g batch 17-AAG 0.2 8 Miglyol 812 15 600 Phospholipon 90G 7.5 300 Disodium EDTA, dihydrate, USP 0.005 0.2 Sucrose, NF 15 600 Water for Injection, USP QS QS 0.2N NaOH Adjust the pH to 6.0±0.2 Add in required amount

17-AAG (CNF-101) was weighed into a 5L polypropylene beaker. Add ethanol about 50 times the weight of the drug, and place the solution in a water bath for sonication to disperse the drug. Then Miglyol 812 (Sasol North America Inc; Houston, TX, USA) and Phospholipon 90G (American Lecithen Co., Oxford, CT, USA) were added to the dispersion, and the mixture was placed on a stirring plate and stirred until the solid was roughly completely dissolved. A sonicator water bath and/or heating to about 45°C can aid in the dissolution of solids. Solutions can be checked with a light microscope to ensure the desired level of dissolution.

Under vigorous stirring, a stream of dry air or nitrogen (National Drug Collection) gas is passed over the surface of the liquid to evaporate the ethanol until the ethanol content is reduced, preferably below 50% w/w of its original content, more preferably below 10% % w/w, most preferably about 5% w/w or less. Solutions are examined under a light microscope fitted with polarizing filters to ensure the desired level of dissolution, preferably complete dissolution (no crystals or precipitates).

EDTA (disodium, dihydrate, USP), sucrose, and water for injection (WFI) were weighed into a 5 L polypropylene beaker and stirred until the solids dissolved. The water phase was then added to the oil phase and mixed thoroughly using a high speed emulsifier with an emulsion head at a speed of 5000 RPM until the oil "flaked off" adhering to the surface. Then increase the shear rate to 10000RPM for 2-5 minutes to obtain a homogeneous colostrum. Laser light scattering (LLS) can be used to measure the average droplet diameter, and the solution can be further examined, such as under an optical microscope to determine the relative presence or absence of crystals and solids.

The pH of the emulsion was adjusted to 6.0±0.2 with 0.2N NaOH. Colostrum was passed through a 11OS microfluidizer (Mierofluidics Inc., Newton, MA, USA) with a 75 micron emulsion chamber (F20Y) at a static pressure of approximately 110 psi (operating pressure 60-95 psi) for 6-8 times until the average droplet diameter ≤ 190nm. Progress can be assessed with the LLS after each pass. The solution can be further examined for the presence of crystals under an optical microscope using polarized light.

The emulsion was then passed through a 0.45 micron Gelman microcapsule filter (Pall Corp., East Hills, NY, USA) followed by a 0.2 micron sterile Sartorius Sartobran P capsule filter (500 cm ) in a laminar flow hood. (Sartorius AG, Goettingen, Germany). Maintain a smooth and consistent flow with up to 60PSI of pressure. The filtrate was collected into one or more polypropylene bottles and placed immediately in a -20°C freezer. Aliquots of 1 ml can be set aside for detection by laser light scattering (LLS) and/or high performance liquid chromatography (HPLC).

Embodiment 5: the preparation method of alternative 17-AAG emulsified preparation

When ethanol is used to facilitate the dissolution of 17-AAG into the oil phase of an emulsion, it is most common to first dissolve 17-AAG in ethanol using sonication, followed by the addition of emulsifiers and medium-chain triglycerides to the solution. Dissolution of all components is then achieved using sonication and stirring.

Alternatively, 17-AAG can be added to the oil phase of the solution without ethanol by heating a pre-formed emulsifier in the triglyceride solution, such as Phospholipon in Miglyol 812, preferably to 65°C or higher, to which a drug, such as 17-AAG, is added and mixed by, for example, stirring and/or sonication. It has been found that the lower melting form of 17-AAG prepared in Example 2 more easily by crystallization of 17-AAG in isopropanol rather than in ethanol can be dissolved in Phospholipon in Miglyol solution at room temperature. See commonly-owned PCT/US01/29715, entitled "Methods of Making ... [17-AAG] and Other Ansamycins," filed September 18, 2002.

The products of Examples 5 and 6 are all light purple milky emulsions with an average diameter of oil droplets of about 200nm or less. When stored at 20°C, 2-8°C or room temperature for more than 2 months, the size of oil droplets is stable. Concentrations of up to 3 mg/ml are reached overall, while only in the oil phase, concentrations are as high as 20-30 mg/ml. If stored at 40°C, degradation of 17-AAG can be observed after about two weeks.

The mixed solvent solution of the drug was subjected to vacuum evaporation of the ethanol component to form a Miglyol solution of 17-AAG. Emulsification can be achieved by mechanical mixing, ultrasonic irradiation treatment and finally by microfluidizer, however it is understood that the terms "emulsification" and "emulsification" are not limited to these treatment methods, other emulsification techniques exist and can be selected Applied or combined with one or more of the above techniques.

Example 6: Addition of long chain triglycerides

Variations on the above methods include adding long chain triglycerides, such as in the form of soybean oil. As shown in Figure 1 for 17-AGG, a source of long-chain triglycerides (soybean oil) was mixed with Miglyol 812N (a source of medium-chain triglycerides) and an emulsifier (Phospholipon 90G (PL90G)) w/w The ratio is 16.7%:50.0%:33.3%. Homogenization was performed until PL90G was completely dissolved (approximately 20 minutes at approximately 20,000 rpm). 1% w/w 17-AAG was then added, which was homogenized/dissolved at about 20,000 rpm for about 5 minutes. The aforementioned substances constitute the "oil phase". This oil phase (1 part) was then slowly added to 3.6 parts of the aqueous phase (9.375% w/w sucrose, 0.0063% w/w EDTA dissolved in Sterile Water for Injection) homogenized at about 12,000-15,000 rpm. The resulting mixture was adjusted to pH 6.0±0.2 with sodium hydroxide and/or hydrochloric acid as necessary. This "primary" emulsion was then microfluidized through an F12Y action chamber and filter sterilized with a 0.2 μm polyethersulfone (PES) filter.

Adopt above-mentioned method, prepare following two kinds of preparations:

Component (w/w) Preparation 1 Preparation 2 17-AAG Soybean Oil Phospholipon 90G (Soybean Lecithin) Miglyol 812N Sucrose Sodium EDTA Sterile Water 2mg/ml3.3%6.6%9.9%7.5%0.005%72.5% 2mg/ml---7.5% 15% 10% 0.005% 67.3%

Embodiment 7: lyophilization

Lyophilization of the emulsions of Examples 5 and 6 can be accomplished according to a protocol similar to that in the table below. Initial temperature (°C) End temperature (℃) Pressure (mTorr) operate 25 -40 Environmental pressure Change temperature at 1°C/min -40 -40 Environmental pressure keep for 60 minutes -40 -40 50 Condenses between -60°C and -80°C -40 -28 50 Change temperature at 1°C/min -28 -28 50 Keep 7200 minutes -28 30 50 Change temperature at 1°C/min 30 30 50 keep for 300 minutes Finish Stopper the _bottle under N at approximately 0.9 atm

The stability profile of the lyophilized 17-AAG emulsion after storage at 2-8°C and reconstitution is as follows.

detection Time (week) 0 2 4 10 Determination (% initial) 100 98.5 97.7 97.0 Purity (area%) 98.8 97.2 97.8 97.8 Droplet diameter (μm) 0.187 0.200 0.197 0.190

Example 8: Long Chain Triglycerides Suppress Sleepiness

When administered rapidly, Miglyol 812N produces a sedative effect due to the metabolic release of octanoate. During intravenous infusion of 17-AAG emulsion (Miglyol 812N oil) in rats, sedation was observed at infusion rates greater than 1.1 gm total lipid/Kg/hr. See attached drawing 2. Sedation was also observed in dogs given intravenous infusions of 17-AAG emulsified formulations at rates greater than 1.13 gm total lipid/Kg/hr. To counteract this effect, soybean oil was added as above to compete with the in vivo metabolism of Miglyol 812N in order to reduce fatty acid caprylate produced during intravenous infusion. For the soybean oil/Miglyol 812NCF237 emulsion, no significant sedation was observed in rats at infusion rates up to approximately 40 gm total lipid/kg/hr. Therefore, the combination of soybean oil and Miglyol 812N greatly improved the tolerability of CF237 emulsified formulations for sedation. Similarly, no sedation nor venous irritation was observed in monkeys administered a sixfold dose of the CF237 emulsified formulation, ie 12 mL formulation/kg/hr iv infusion.

Example 9: Preparation of other ansamycins for similar formulations

Ansamycins other than 17-AAG

Essentially all ansamycins can be substituted for 17-AAG and formulated as described in the examples above. A variety of such ansamycins and their preparation are described in detail in PCT/US03/04283. The preparation of two of them is described below.

Compound 563: 17-(Benzoyl)-aminogeldanamycin. A solution of 17-aminogeldanamycin (1 mmol) _in EtOAc was treated with _Na2S2O4 (0.1M, 300ml) at room _temperature . After ₂ hours, the aqueous layer was extracted twice with EtOAc, while the combined organic layers were dried over _Na2SO4 and concentrated under low pressure to obtain 18,21-dihydro-17-aminogeldanamycin as a yellow solid. The latter was dissolved in anhydrous THF and transferred via cannula to a mixture of benzoyl chloride (1.1 mmol) and MS4 Å (1.2 g). After 2 hours, EtN(i-Pr) ₂ (2.5 mmol) was added to the reaction mixture. After stirring overnight, the reaction mixture was filtered and concentrated under reduced pressure. Water was then added to the residue, extracted three times with EtOAc, the combined organic layer was dried over _Na2SO4 and concentrated under low pressure to obtain the _crude product, which was purified by flash chromatography to give 17-(benzoyl)-aminoglycerol Dextamycin. In 80:15:5 _{CH2Cl2: EtOAc} :MeOH, Rf = 0.50. Mp = 218-220°C. 1H NMR (CDCl ₃ ) 0.94(t, 6H), 1.70(br s, 2H), 1.79(br s, 4H), 2.03(s, 3H), 2.56(dd, 1H), 2.64(dd, 1H), 2.76-2.79(m, 1H), 3.33(br s, 7H), 3.44-3.46(m, 1H), 4.325(d, 1H), 5.16(s, 1H), 5.77(d, 1H), 5.91(t , 1H), 6.57(t, 1H), 6.94(d, 1H), 7.48(s, 1H), 7.52(t, 2H), 7.62(t, 1H), 7.91(d, 2H), 8.47(s, 1H), 8.77(s, 1H).

Compound 237: a dimer. 3,3-Diamino-dipropylamine (1.32 g, 9.1 mmol) was added dropwise to geldanamycin (10 g, 17.83 mmol) in DESO (200 ml) in a flame-dried flask under _N2 solution, stirred at room temperature. After 12 hours the reaction mixture was diluted with water. The reactant formed a precipitate and was filtered to obtain crude product. The crude product was chromatographed on silica (5% _CH3OH / _CH2Cl2 ) to afford _the dimer as a purple solid. Purple pure product was obtained after purification by flash chromatography (silica gel); yield: 93%; mp 165 °C; 1H NMR (CDCl ₃ ) 0.97 (d, J = 6.6 Hz, 6H, 2CH3), 1.0 (d, J =6.6Hz, 6H, 2CH3), 1.72(m, 4H, 2CH2), 1.78(m, 4H, 2CH2), 1.80(s, 6H, 2CH3), 1.85(m, 2H, 2CH), 2.0(s, 6H , 2CH3), 2.4(dd, J=11Hz, 2H, 2CH), 2.67(d, J=15Hz, 2H, 2CH), 2.63(t, J=10Hz, 2H, 2CH), 2.78(t, J=6.5 Hz, 4H, 2CH2), 3.26(s, 6H, 2OCH3), 3.38(s, 6H, 2OCH3), 3.40(m, 2H, 2CH), 3.60(m, 4H, 2CH2), 3.75(m, 2H, 2CH ), 4.60 (d, J=10Hz, 2H, 2CH), 4.65 (Bs, 2H, 2OH), 4.80 (Bs, 4H, 2NH2), 5.19 (s, 2H, 2CH), 5.83 (t, J=15Hz, 2H, 2CH=), 5.89(d, J=10Hz, 2H, 2CH=), 6.58(t, J=15Hz, 2H, 2CH=), 6.94(d, J=10Hz, 2H, 2CH=), 7.17( m, 2H, 2NH), 7.24 (s, 2H, 2CH=), 9.20 (s, 2H, 2NH); MS (m/z) 1189 (M+H).

The corresponding hydrochloride salt was prepared by adding HCl in EtOH (5ml, 0.123N) to a solution of compound #237 (1 gm prepared above) in THF (15ml) and EtOH (50ml) at room temperature. The reaction mixture was stirred for 10 minutes. The salt precipitated out and was filtered again and washed with copious EtOH and dried in vacuo.

* * * *

Those of ordinary skill can understand that the parameters in the above examples and tables can be adjusted according to the conditions used, and according to whether the preparation method and the amount of materials used are enlarged or reduced or relative to each other and to what extent.

The above examples are not limiting, they merely represent different aspects and embodiments of the invention. All documents cited herein represent the state of the art to which this invention pertains. Although none of the documents cited is admitted to be prior art, the contents of each document cited are hereby incorporated by reference to the same extent as if each document was incorporated by reference in its entirety. In the event of a conflict between an express definition herein and a definition appearing in the prior art or in priority texts cited herein, the express definition in the present application controls.

Those skilled in the art will readily appreciate that the present invention well accomplishes the above stated objects and obtains the foregoing results, advantages, and inherent features contained therein. The methods and compositions are intended to illustrate preferred embodiments, and are exemplary rather than limiting on the scope of the invention. Certain modifications and other uses will occur to those skilled in the art and are encompassed within the scope of the invention as defined by the scope of the claims.

The reagents are either commercially available, such as from Sigma-Aldrich, or can be easily prepared by conventional methods known to those skilled in the art without undue experimentation.

It will be apparent to those skilled in the art that various substitutions and modifications can be made in the present invention without departing from the spirit and scope of the inventions. Accordingly, such other embodiments are within the scope of the invention and the following claims.

The invention illustratively described herein may suitably be practiced in the absence of any element, limitation, not specifically disclosed herein. For example, in all cases herein, the terms "comprising," "consisting essentially of," and "consisting of" can each be replaced by one of the other two terms, and each term in the patent have different meanings in law. The terms and expressions used are used as terms of description rather than limitation, and the use of these terms and expressions does not exclude any equivalent features or parts thereof described and shown in the present invention. At the same time, those skilled in the art can realize that it is completely possible to make various changes within the scope of protection of the present invention. It is therefore to be understood that while the invention has been specifically disclosed in terms of preferred embodiments and optional features, modifications and alterations to the concepts disclosed herein may be made by those skilled in the art and that such alterations and alterations are to be considered in the specification and claims within the scope of the invention as defined.

In addition, if the features or aspects of the present invention are described using Markush groups or other alternative groups, those skilled in the art may consider that the present invention may also use each member or subgroup of members of the Markush group or other groups Described and excluded if necessary by qualification of individual members.

Other embodiments are within the scope of the following claims.

Claims (32)

1. A pharmaceutical composition comprising: HSP90 inhibitor, optionally ansamycin; emulsifiers; and Oils containing both medium chain triglycerides and long chain triglycerides. 2. The pharmaceutical composition of claim 1, wherein said HSP90 inhibitor is ansamycin. 3. The pharmaceutical composition of claim 2, wherein said ansamycin is geldanamycin or a geldanamycin derivative. 4. The pharmaceutical composition of claim 3, wherein the geldanamycin derivative is selected from 17-AAG and DMAG. 5. The pharmaceutical composition of claim 1, wherein the w/w ratio of said medium chain triglycerides to said long chain triglycerides is from 10:1 to 0.01:10. 6. The pharmaceutical composition of claim 1, which is an oil-in-water emulsion having a lipid phase and an aqueous phase, wherein said lipid phase accounts for 5-30% w/w of the total weight. 7. The pharmaceutical composition of claim 1, wherein said phospholipid is present in the form of lecithin. 8. The pharmaceutical composition of claim 4, wherein the amount of lecithin is 3-10% w/w of the oil-in-water emulsion. 9. The pharmaceutical composition of claim 1 comprising 3-10% w/w phospholipid and 5-20% w/w oil. 10. The pharmaceutical composition of claim 1 comprising no more than 10% w/w long chain triglycerides. 11. The pharmaceutical composition of claim 10, further comprising less than or equal to 7% w/w long chain triglycerides. 12. The pharmaceutical composition of claim 10, wherein said long-chain triglycerides comprise fatty acid esters consisting of 16-18 linear carbon units, said triglycerides being optionally selected from the group consisting of linoleic acid, oleic acid, One or more of palmitic acid and stearic acid. 13. The pharmaceutical composition of claim 1, wherein said phospholipid is in the form of lecithin, optionally soy or egg lecithin. 14. The pharmaceutical composition of claim 4, wherein the ansamycin is a low melting point isomer of 17-AAG with a melting point not exceeding 200°C. 15. The pharmaceutical composition of claim 14, wherein said low melting point isomer is 17-AAG having a melting point of 147-175°C. 16. The pharmaceutical composition of claim 1, wherein said oil comprises one or more natural oils selected from the group consisting of soybean oil, sesame oil, safflower oil, and corn oil. 17. The pharmaceutical composition of claim 1 which is an emulsion, optionally lyophilized. 18. The pharmaceutical composition of claim 1, comprising one or more of water, preservatives, cryoprotectants, buffers, chelating agents, and tonicity regulators. 19. The pharmaceutical composition of claim 1 comprising Miglyol 812N. 20. The pharmaceutical composition of any one of claims 1-19, comprising 0.5 mg/ml-4 mg/ml of 17-AAG. 21. The pharmaceutical composition of any one of claims 1-19, comprising 0.05% w/w to 0.4% w/w of 17-AAG. 22. The pharmaceutical composition of claim 1, comprising the following components: 2 mg/ml 17-AAG, 3.3% soybean oil, 6.6% lecithin, 9.9% Miglyol 812N, 7.5% sucrose, and water. 23. The pharmaceutical composition of claim 1, comprising the following components: 2 mg/ml 17-AAG, 7.5% lecithin, 15% Miglyol 812N, 10% sucrose, and water. 24. The pharmaceutical composition of claim 22 or 23, further comprising sodium edetate. 25. The pharmaceutical composition of claim 24, wherein the sodium edetate is 0.005% w/w. 26. The pharmaceutical composition of claim 1, wherein said long chain triglyceride is present in an amount sufficient to reduce or avoid medium chain triglyceride mediated central nervous system effects. 27. The pharmaceutical composition of claim 22, wherein the central nervous system effect is selected from one or more of drowsiness, nausea, drowsiness and EEG changes. 28. The pharmaceutical composition of claim 1, further comprising short chain triglycerides. 29. A method of reducing central nervous system effects mediated by medium chain triglycerides in a patient, wherein the patient receives a pharmaceutical formulation comprising medium chain triglycerides as a component of the formulation, the method comprising: a) providing a pharmaceutical formulation comprising ansamycin and medium-chain and long-chain triglycerides in an amount sufficient to reduce or avoid medium-chain fatty acid-mediated central nervous system effects; and b) administering the product of step a) to a patient. 30. The method of claim 29, wherein the central nervous system effect is selected from one or more of drowsiness, nausea, drowsiness, and EEG changes. 31. The pharmaceutical composition of claim 1 in one or more of lyophilized, frozen, thawed or reconstituted form. 32. The pharmaceutical composition of claim 1 stored in an inert environment.

Images