Biologically active taxane analogs and methods of treatment by oral administration

Abstract

The present invention relates to a novel chemical compound for use in the treatment of cancer, to compositions containing said compound, methods of manufacture and combinations with other therapeutic agents. ##STR00001##

Claims

1. A method of treating cancer, the method comprising administering to a subject a composition comprising a compound having a structure represented by the following formula (1): ##STR00031## or a pharmaceutically acceptable salt, solvate or hydrate thereof.

2. The method of claim 1, wherein a therapeutically effective amount of the compound of formula (1) is administered by week or by 21 days for a total dose selected from the group consisting of from about 7 mg/m.sup.2 to about 555 mg/m.sup.2, from about 7 mg/m.sup.2 to about 240 mg/m.sup.2, from about 7 mg/m.sup.2 to about 185 mg/m.sup.2 during a 21 day period.

3. The method of claim 1, wherein the composition is administered by oral, pulmonary, intraperitoneal (ip), intravenous (iv), intramuscular (im), subcutaneous (sc), transdermal, buccal, nasal, sublingual, ocular, rectal or vaginal administration.

4. The method of claim 3, wherein the parenteral administration is selected from intravenous injection and intravenous infusion.

5. The method of claim 3, wherein the parenteral administration is for about 60 minutes or less.

6. The method of claim 4, wherein the said intravenous infusion is once every 7 days for 3 weeks, followed by a 7 day rest period, in a 28 day cycle, for at least one cycle.

7. The method of claim 4, wherein the intravenous infusion is once every 21 days, in a 21 day cycle, for at least one cycle.

8. The method of claim 1, wherein the composition comprises the compound of formula (1) at a dose of 240 mg/m.sup.2 or less, or 185 mg/m.sup.2 or less.

9. The method of claim 1, wherein the composition comprises the compound of formula (1) at a dose of 185 mg/m.sup.2 or less, or 160 mg/m.sup.2 or less.

10. The method of claim 1, wherein the composition comprises the compound of formula (1) in a unit dose selected from the group consisting of about 200 mg to 400 mg.

11. The method of claim 10, wherein the composition comprises the compound of formula (1) in a unit dose selected from the group consisting of about 250 mg, 300 mg and 350 mg.

12. The method of claim 1, wherein the compound formula (1) is a compound having the structure of formula S-(1): ##STR00032##

13. The method of claim 1, wherein the composition is administered in combination with one or more pharmaceutically acceptable ingredient selected from the group consisting of phomopsin, dolastatin, bevacizumab (Avastin™) steganacin, paclitaxel, taxotere, vinblastine, vincristine, vindesine, vinorelbine, navelbine, colchicine, maytansine, ansamitocin, Iressa, Tarceva, Herceptin, lapatinib, vandetanib, Sorafenib, BAY-57-9006, bevacizumab, cetuximab, gemtuzumab, panitumumab, rituximab, tositumomab, trastuzumab, apolizumab, oregovomab, mitumomab, alembuzumab, ibritumomab, vitaxin, SU-6668, semaxanib, sunitinib malate, SU-14813, vandetanib, Recentin, CP-547632, CEP-7055, AG-013736, pazopanib, combretastatin, squalamine, combrestatin A4 phosphate, TNP-470, neovastat, dasatinib, imatinib, nilotinib, sorafenib, sunitinib, triethylenethiophosphoramine, alitretinoin, altretamine, arsenic trioxide, asparaginase, bexarotene, denileukin diftitox, hydroxycarbamide, masoprocol, mitotane, pegaspargase, tretinoin, raltitrexed, IL-10, IL-12, bortezomib, leuprolide, interferon β, pegylated interferons, atrasentan, melphalan, cyclophosphamide, chlormethine, chlorambucil, trofosfamide, ifosfamide, nitromin, busulfan, thiotepa, chlorambucil, CC-1065, temozolomide, pipobroman, dacarbazine, mechlorethamine, procarbazine, uramustine, RSU-1069, CB-1954, hexamethylmelamine, cisplatin, carboplatin, oxaliplatin, BBR3464, satraplatin, tetraplatin, iproplatin, amsacrine, netropsin, pibenzimol, mitomycin, duocarmycin, dactinomycin, distamycin, mithramycin, chromomycin, olivomycin, anthramycin, bleomycin, liblomycin, rifamycin, actinomycin, adramycin, trichostatin A, propamidine, stilbamidine, rhizoxin, nitroacridine, geldamycin, 17-AAG, 17-DMAG, plicamycin, deoxycoformycin, levamisole, daunorubicin, doxorubicin, epirubicin, idarubicin, mitroxantrone, valrubicin, carmustine, fotemustine, lomustine, streptozocin, gemcitabine, 5-fluorouracil (5-FU), fludarabine, cytarabine, capecitabine, mercaptopurine, cladribine, clofarabine, thioguanine, pentostatin, floxuridine, pentostatin, aminopterin, methotrexate, pemetrexed, camptothecin, irinotecan, topotecan, epipodophyllotoxin, etoposide, teniposide, aminogluthetimide, anastrozole, exemestane formestane, letrozole, fadrozole, aminoglutethimide, leuprorelin, buserelin, goserelin, triptorelin, abarelix, estramustine, megestrol, flutamide, casodex, anandron, cyproterone acetate, finasteride, bicalutamide, tamoxifen or its citrate salt, droloxifene, trioxifene, raloxifene or zindoxifene, a derivative of 17-β-estradiol such as ICI 164, ICI 384, ICI 182, ICI 780, testolactone, fulvestrant, toremifene, testosterone, fluoxymesterone, dexamethasone, triamcinolone, dromostanolone propionate, megestrol acetate, methyltestosterone, chlorotrianisene, hydroxyprogesterone, medroxyprogesterone acetate, reloxafine, etanercept, thalidomide, revimid (CC-5013), aziridoquinones, misonidazole, NLA-1, RB-6145, misonidazole, nimorazole, RSU-1069, SR-4233, porfimer, photofrin, verteporfin, merocyanin 540, tin etiopurpurin, PUVA, aminolevulinic acid, methyl aminolevulinate, minodronate, zoledronic acid, ibandronate sodium hydrate or clodronate disodium, misonidazole, misonidazole, amifostene, oblimersen, TIMP-1 or TIMP-2, marimastat, TLK-286 and mixtures thereof.

14. The method of claim 1, wherein the composition is administered in combination with an anti-mitotic agent, an anti-microtubule agent or both.

15. The method of claim 1, wherein the composition is formulated as a solid formulation.

16. The method of claim 15, wherein the solid formulation is formulated as immediate and/or modified release.

17. The method of claim 16, wherein the modified release formulation is selected from the group consisting of delayed-, sustained-, pulsed-, controlled-, targeted and programmed release.

18. The method of claim 15, wherein the solid formulation is in a form selected from the group consisting of tablets, capsules containing particulates, liquids or powders, lozenges (including liquid-filled), chews, multi- and nano-particulates, gels, solid solution, liposome, films, and ovules.

19. The method of claim 15, wherein the solid formulation is in the form of tablets.

20. The method of claim 15, wherein, the solid formulation contains a disintegrant selected from the group consisting of sodium starch glycolate, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, croscarmellose sodium, crospovidone, polyvinylpyrrolidone, methyl cellulose, microcrystalline cellulose, lower alkyl-substituted hydroxypropyl cellulose, starch, pregelatinised starch and sodium alginate.

21. The method of claim 19, wherein the tablets comprise a binder, the binder selected from the group consisting of microcrystalline cellulose, gelatin, sugars, polyethylene glycol, natural and synthetic gums, polyvinylpyrrolidone, pregelatinised starch, hydroxypropyl cellulose and hydroxypropyl methylcellulose.

22. The method of claim 1, wherein the cancer is selected from the group consisting of pancreatic, breast, colorectal cancer, prostate, cervical, ovarian, gastric, pancreatic, bladder endometrial, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), esophageal, glioblastoma, melanoma, renal cell cancer, hepatocellular carcinoma, and brain disease, wherein said brain disease is a cancer that has metastasized to the brain, sarcoma, leukemia, lymphoma, bone marrow dyscrasias, neuroblastoma, glioblastoma, thyroid, lung, small-cell lung, non-small cell lung, gastric, head and neck and melanoma.

Description

BRIEF DESCRIPTION OF THE FIGURES

(1) FIG. 1. S-(1) crosses rat blood-brain barrier.

(2) FIG. 2. Mouse Plasma and Brain Levels of S-(1) after single IV dose.

(3) FIG. 3. Summary of U251 Orthotopic Intracranial Xenograft Study Data.

(4) FIGS. 4A, 4B, and 4C. S-(1) Oral efficacy in Mice.

(5) FIGS. 5A and 5B. Oral Efficacy of S-(1) in Neuroblastoma Xenograft.

(6) FIGS. 6A and 6B. S-(1) Oral Efficacy in Ovarian Tumor Xenograft.

(7) FIGS. 7A and 7B. S-(1) Oral efficacy in Glioblastoma Xenograft.

(8) FIGS. 8A and 8B. IV efficacy of S-(1) in Glioblastoma Xenograft Model.

(9) FIGS. 9A and 9B. Efficacy Comparison in Prostate Tumor Xenograft Model.

(10) FIGS. 10A and 10B. Efficacy Comparison in Lung Tumor Xenograft Model

(11) FIGS. 11A and 11B. Efficacy Comparison in Breast Tumor Xenograft Model

(12) FIGS. 12A and 12B. Efficacy Comparison in HT 29 Colon Xenograft

(13) FIGS. 13A and 13B. Oral S-(1)+/−Avastin

(14) FIGS. 14A and 14B. Combination Efficacy of S-(1)+/−Avastin

(15) The compound of formula S-(1) may be administered by any conventional route of administration including, but not limited to, oral, pulmonary, intraperitoneal (ip), intravenous (iv), intramuscular (im), subcutaneous (sc), transdermal, buccal, nasal, sublingual, ocular, rectal and vaginal. It will be readily apparent to those skilled in the art that any dosage or frequency of administration that provides the desired therapeutic effect is suitable for use in the present application.

(16) The therapeutically effective amount of the compound of formula S-(1) may be administered by week or by 21 days for a total dose of about 7-555 mg/m.sup.2, or 7-240 mg/m.sup.2, or 7-185 mg/m.sup.2 during a 21 day period. The dosages, however, may be varied depending on the requirements of the subject to be treated, including sex, age, weight, diet, etc. The precise amount of the compound of formula S-(1) required to be administered depends on the judgment of the practitioner and is peculiar to each individual.

(17) Another embodiment of the present application is a pharmaceutical composition for treating brain tumor, comprising a therapeutically effective amount of the compound of formula S-(1) and a pharmaceutically acceptable carrier.

(18) Another embodiment of the present application is a pharmaceutical composition for oral administration of the compound being patient friendly. The solubility properties of the compound of formula S-(1) are such that high concentration of the compound of formula S-(1) can be achieved in pharmaceutically acceptable carriers, between 75 and 200 mg/ml, that can be filled in hard and soft gelatin capsules with an appropriate chemical and physical stability profile.

(19) To prepare a pharmaceutical composition of the present application, the compound of formula S-(1) is admixed with a pharmaceutically acceptable carrier according to conventional pharmaceutical compounding techniques, wherein the carrier may take a wide variety of forms depending on the form of preparation desired for administration. Suitable pharmaceutically acceptable carriers are well known in the art. Descriptions of some pharmaceutically acceptable carriers may be found in The Handbook of Pharmaceutical Excipients Eds. Rowe et al., American Pharmaceutical Association and the Pharmaceutical Society of Great Britain.

(20) The pharmaceutical composition of the present application may be, depending on the route of administration, in the form of a tablet, pill, capsule, granule, powder, ointment, gel, solution, sterile parenteral solution or suspension, metered aerosol or liquid spray, or suppository that may contain micro or nano particles, a colloid, liposome or a biocompatible or biodegradable medium.

(21) As a solid dosage form, the pharmaceutical composition of the present application may comprise, in addition to the compound of formula S-(1), at least one diluent, binder, adhesive, disintegrant, lubricant, antiadherent, and/or glidant. Additionally, sweeteners, flavorants, colorants and/or coatings may be added for specific purposes.

(22) As a liquid dosage form, the pharmaceutical composition of the present application may comprise, in addition to the compound of formula S-(1) and a liquid vehicle, at least one wetting agent, dispersant, flocculation agent, thickener, buffer, emulsifier, amphiphilic, osmotic agent, coloring agent, anti-oxidant, flavor, fragrance, and/or preservative.

(23) Such a possible liquid formulation includes a sterile solution containing 10 mg/mL of the compound of formula S-(1) in a 15:85 or 50:50 (w/v) polyoxyl 35 castor oil/dehydrated alcohol solution. An appropriate pharmaceutical grade polyoxyl 35 castor oil is Cremophor EL-P, which is a non-ionic solubilizer made by reacting castor oil with ethylene oxide in a molar ratio of 1:35, followed by a purification process (BASF Pharma).

(24) Other suitable agents for inclusion in such formulations are as described in WO 99/45918. Such additional carriers include vitamin E TPGS, oleic acid, Tweens such as Tween 80, lipid carriers used for parenteral nutrition, solutol and the like. As shown in Table 10 below, these agents and the composition of the final formulation might have an impact of the pharmacokinetics of the compound and its pharmacodymanics and thus its tissue distribution. As previously mentioned, the compound of the invention surprisingly may be administered orally as further exemplified below. Oral administration may involve swallowing, so that the compound enters the gastrointestinal tract, or buccal or sublingual administration may be employed by which the compound enters the blood stream directly from the mouth.

(25) The therapeutically effective amount of the compound of formula S-(1) may be administered by the parenteral route using a weekly or 21 days schedule for a total dose of about 7-555 mg/m.sup.2, or 7-240 mg/m.sup.2, or 7-185 mg/m.sup.2 during a 21 day period. The dosages, however, may be varied depending on the requirements of the subject to be treated, including sex, age, weight, diet, etc. The precise amount of the compound of formula S-(1) required to be administered depends on the judgment of the practitioner and is peculiar to each individual. The compound of formula S-(1) may be administered by the oral route using a daily, weekly or 21 days schedule, and any schedule comprised within this range, for a total dose of about 5-600 mg/m.sup.2, or 25-400 mg/m.sup.2. The dosages, however, may be varied depending on the requirements of the subject to be treated, including sex, age, weight, diet, etc. The precise amount of the compound of formula S-(1) required to be administered depends on the judgment of the practitioner and is peculiar to each individual.

(26) It is believed that the compounds and hence compositions of this invention possess an enhanced safety profile (for example, on the immune system or the blood) in comparison to marketed taxanes which offers the potential for enhanced or longer dosing schedules under the direction of the skilled physician than marketed taxanes.

(27) The compositions may for oral administration contain at least 15 mg of the compound of the invention per dose, more aptly at least 50, 80, 100, 150, 200 mg and more aptly less than 250 mg per dose. However, the dosage may be varied as directed by the physician in view of the individual patient's response. A liquid composition will normally contain about 0.1 mg/ml to about 300 mg/ml for example about, 1, 5 10, 50, 100 or 150 mg/ml of the compound of the invention. A non-liquid composition may contain a proportion of the compound of the invention, for example 5% to 50%, such as 10, 20, 25 or 30% by weight. In the case of a liquid composition for parenteral administration, the composition will normally contain between 1 mg/ml and 50 mg/ml while the composition will normally contain between 40 mg/ml to 200 mg/ml if administered orally. WO 1999/45918 and the international patent applications and US patent applications referred to above discloses compositions that may be considered for use with compounds of the invention.

(28) Formulations suitable for oral administration include solid formulations such as tablets, capsules containing particulates, liquids, or powders, lozenges (including liquid-filled), chews, multi- and nano-particulates, gels, solid solution, liposome, films, ovules, sprays and liquid formulations. Liquid formulations include suspensions, solutions, syrups and elixirs. Such formulations may be employed as fillers in soft or hard capsules and typically comprise a carrier, for example, ethanol, polyethylene glycol, propylene glycol, methylcellulose, or a suitable oil, a lipohilic component with a high HLB value, an amphiphilic solvent, and one or more emulsifying agents and/or suspending agents. Liquid formulations may also be prepared by the reconstitution of a solid, for example, from a sachet. The compound of the invention may also be used in fast-dissolving, fast-disintegrating dosage forms. Tablets generally contain a disintegrant. Examples of disintegrants include sodium starch glycolate, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, croscarmellose sodium, crospovidone, polyvinylpyrrolidone, methyl cellulose, microcrystalline cellulose, lower alkyl-substituted hydroxypropyl cellulose, starch, pregelatinised starch and sodium alginate. Binders are generally used to impart cohesive qualities to a tablet formulation. Suitable binders include microcrystalline cellulose, gelatin, sugars, polyethylene glycol, natural and synthetic gums, polyvinylpyrrolidone, pregelatinised starch, hydroxypropyl cellulose and hydroxypropyl methylcellulose. Tablets may also contain diluents, such as lactose (monohydrate, spray-dried monohydrate, anhydrous and the like), mannitol, xylitol, dextrose, sucrose, sorbitol, microcrystalline cellulose, starch and dibasic calcium phosphate dihydrate. Tablets may also optionally comprise surface active agents, such as sodium lauryl sulfate and polysorbate 80, and glidants such as silicon dioxide and talc. Tablets may also generally contain lubricants such as magnesium stearate, calcium stearate, zinc stearate, sodium stearyl fumarate, and mixtures of magnesium stearate with sodium lauryl sulphate. Other possible ingredients include anti-oxidants, colourants, flavouring agents, preservatives and taste-masking agents. Tablet blends may be compressed directly or by roller to form tablets. Tablet blends or portions of blends may alternatively be wet-, dry-, or melt-granulated, melt congealed, or extruded before tabletting. The final formulation may comprise one or more layers and may be coated or uncoated; it may even be encapsulated.

(29) Consumable oral films are typically pliable water-soluble or water-swellable thin film dosage forms which may be rapidly dissolving or mucoadhesive and typically comprise the compound of formula S-(1), a film-forming polymer, a binder, a solvent, a humectant, a plasticiser, a stabiliser or emulsifier, a viscosity-modifying agent and a solvent. Some components of the formulation may perform more than one function. The film-forming polymer may be selected from natural polysaccharides, proteins, or synthetic hydrocolloids and is typically present in the range 0.01 to 99 weight %, more typically in the range 30 to 80 weight %.

(30) Other possible ingredients include anti-oxidants, colorants, flavourings and flavour enhancers, preservatives, emulsifiers, salivary stimulating agents, cooling agents, co-solvents (including oils), emollients, bulking agents, anti-foaming agents, surfactants and taste-masking agents.

(31) Solid formulations for oral administration may be formulated to be immediate and/or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted and programmed release.

BIOLOGICAL EXAMPLES

In Vitro ED50 MT Polymerization Study

(32) In this tubulin binding assay, microtubule protein (MTP) is used as a substrate. The assay contains bovine tubulin plus microtubule associated proteins (MAP). MTP is polymerized into microtubules in the presence of DAPI (4′,6′-diamidino-2-phenylindole), a fluorescent compound. DAPI binds to tubulin; when microtubules are formed and there is an enhancement of fluorescence. The microtubule formation is measured as a function of time, using a fluorescence plate reader. The ED.sub.50 values obtained with this method are in good agreement with older sedimentation techniques. The more current assay, using DAPI, is faster and uses less protein. The method used is based on the procedure published by Donna M. Barron, et al, “Fluorescence-based high-throughput assay for antimicrotuble drugs” Analytical Biochemistry, 315: 49-56, 2003, which is incorporated by reference in its entirety. The excitation wavelength, in that assay, was set at 370 nm and the emission wavelength was set at 450 nm for the DAPI experiments.

(33) A Bio-Tek FL 600 microplate Fluorescence Reader was used to measure the relative level of fluorescence in the DAPI assay.

(34) Assays were conducted in 96-well plates. Each well contained a total volume of 0.1 mL consisting of PEM buffer (0.1 M Pipes, 1 mM EGTA, 1 mM MgCl.sub.2, pH 6.9), 0.2 mg bovine microtubule protein, and 10 μg of DAPI. Compounds having paclitaxel-like activity of varying concentrations dissolved in DMSO were added last. The final DMSO concentration was 4%. The plates were incubated at 37° C. for 30 minutes and read in a fluorescence plate reader using an excitation wavelength of 360 nm and an emission wavelength of 460 nm. Fluorescence values were corrected for the sample without compound. Results were expressed as a percent of maximum assembly, with maximum assembly taken to be that obtained at 25 μM paclitaxel.

(35) Experiments were done twice in triplicate. Results were subsequently combined and fit to a non-linear regression program. The results from these studies summarized in Table 2 indicate that the S-(1) diastereoisomer has an ED.sub.50 potency that is equal to or greater than that determined for other tubulin binding agents such as paclitaxel, docetaxel and Epothilone B.

(36) TABLE-US-00002 TABLE 2 Summary of Tubulin Polymerization Assays, Comparison of S-(1) to paclitaxel, docetaxel, and epothilone B. Compound ED50, μM ED50 Compound/ED50 Paclitaxel S-(1) 1.58 ± 0.46 0.53 Paclitaxel 2.97 ± 0.50 1.00 Docetaxel 3.18 ± 0.45 1.07 Epothilone B 3.31 ± 0.51 1.11 MTS Proliferation Assay (Promega)

(37) Day 1: Cells were plated in appropriate growth medium at 5×10.sup.3 per well in 100 ul in 96 well tissue culture plates, Falcon, one for each drug to be tested. Col 1 was blank; it contained no cells, just medium. The plates were incubated overnight at 37° C., 5% CO.sub.2 to allow attachment.

(38) Day 2: Added 120 ul growth medium in wells of 96-well “dilution plates” (one for each drug) and let sit in 37° C. incubator for about 1 hr.

(39) Thawed DMSO drug stocks (usually at 10 mM). Each drug was diluted 6 ul into a tube with 3 ml growth medium, to 20 μM.

(40) Aspirated medium from col 12 of a dilution plate; added 200-300 ul of 20 uM drug to wells of col 12. Made serial dilution down this 96-well plate: for a 1:5 dilution pattern, moved 60 ul from col 12 to col 11, mixed 4-5 times (using 8 place multi-pipettor), moved 60 ul to col 10, etc. stopping at col 3.

(41) Moved 100 ul of medium+drug from dilution plate to a cell plate, i.e. col 1 from drug plate (blank=no cells) to col 1 of cell plate, etc. up to col 12. Col 2 contained cells with no drug. Col 3 had the lowest concentration of drug (0.005 nM) and col 12 had the highest drug concentration (10 μM).

(42) Day 4 or 5: Terminated the assay 48 to 72 hrs after drug addition. Thawed MTS reagent; made up enough medium+MTS to cover all plates at 115 ul per well (100 ul medium+15 ul MTS). Aspirated medium+drugs from cell plate; replaced with medium+MTS mix and incubated 1-6 hrs (37° C., 5% CO.sub.2), depending on cell type. When the color turned dark in control wells (col 2), and was still light in col 12, the absorbance at 490 nm was read on a plate reader; the results were used to calculate IC.sub.50.

(43) The effects of the compound of formula S-(1) measured in in vitro and xenograft animal models of various brain cancers were evaluated in the following experimental examples, which are intended to be a way of illustrating but not limiting the present application.

(44) S-(1) Pharmacokinetic Data in Mouse Model:

(45) S-(1) was formulated in 7% Ethanol: 3% Cremophor EL: 90% D5W (5% Dextrose in water). The formulated drug was administered as a single IV bolus via tail vein injection. The details are set out below:

(46) Animal Information:

(47) TABLE-US-00003 Wt. Total Food Spe- Range No. Re- cies Strain Sex (g) Required Source Fasted turned Mice CD-1 Male 20-30 36 CRL No NA
Study Design

(48) TABLE-US-00004 No. of Test Dose Conc. Dose Group Ani- Arti- Volume (mg/ (mg/ No. mals cle Vehicle Route (mL/kg) mL) kg) 1 36 S-(1) 7% Eth- IV 10 2 20 anol: 3% (Tail Cremophor Vein) EL: 90% D5W

(49) Sample Collection timing: Terminal blood (plasma) and brain specimens were collected at 9 times post-dose, with 4 mice per sample time. Sample collection times: 5, 15 min., 2, 8, 24, 32, 48, 72 and 96 hr post-dose (4 mice per sample time).

(50) Blood collection: A terminal blood sample was collected from each mouse via cardiac puncture. The whole blood was collected on wet ice, spun down immediately (at 4° C.), and plasma stored frozen at −20° C. until analysis. The anticoagulant used was K.sub.2EDTA. Brain Sample Collection Whole brains were removed by dissection. Brains were collected on wet ice, weighed, and stored at −20° C.

(51) TABLE-US-00005 TABLE 3 Mouse Plasma and Brain Levels of S-(1) after Single iv Dose Brain Plasma Cmax ng/ml 3726.6 Cmax ng/ml 1550.2 AUC(0-t) ng-hr/ml 64984.9 AUC (0-t) ng-hr/ml 16870.2 AUC (0-∞) ng-hr/ml 88742.9 AUC (0-∞) ng-hr/ml 16951.2 MRT (expo)hr 42.5 MRT (expo)hr 17.6 CL ml/hr 0.006 CL ml/hr 0.024
The results indicate that S-(1) is not impeded by the blood-brain barrier in the mouse model. See FIG. 2.
Antitumor Efficacy of S-(1) Against Human U251 CNS Tumor Cells Implanted in Mouse Brain:

(52) Additional studies were conducted in animal models to determine the efficacy of S-(1) against brain tumors. In one study, the anti-tumor activity of S-(1) was evaluated both when administered alone and in combination with temozolomide, against intracerebrally (ic) implanted human U251 (glioblastoma) CNS tumor cells in male athymic nude mice.

(53) Tumor Model: Each animal was implanted with one million U251 human CNS tumor cells from an in vitro cell line by ic injection with a 25 gauge needle. The day of tumor implantation (May 24, 2006) was designated as day 0. A sufficient number of mice were implanted so that animals with body weights in a range as narrow as possible were selected for the trial on the day of treatment initiation (day 1 after tumor implantation). Animals were randomly assigned to the treatment groups and individually identified by earmark codes.

(54) Drug Formulation: On each day of treatment, the appropriate amount of S-(1) was formulated in 3% cremophor EL/7% ethanol/90% D5W at a concentration of 2 mg/mL. A portion of this solution was then diluted with the complete vehicle to achieve the lower dosing concentration of 1.2 mg/mL. Both concentrations of S-(1) were then kept at 37° C. and injected within 30 minutes of formulation on the basis of exact body weight using a volume of 0.1 mL/10 g of body weight. Temozolomide (Temodar, Schering Corporation) was prepared on each day of injection in Klucel+tween 80 at a concentration of 4 mg/mL. Temozolomide was administered within 5 minutes of formulation on the basis of exact body weight using a volume of 0.2 mL/10 g of body weight.

(55) Data Collection Animal health surveillance was conducted and mortality data were collected daily. The animals were weighed twice weekly starting with the first day of treatment.

(56) Study Duration: The study was terminated 100 days after tumor implantation. Any animal found moribund or whose body weight dropped below 14 g was euthanized prior to study termination.

(57) Parameters Evaluated Number of 100-day survivors, median day of death, and the increase in lifespan based on median day of death and expressed as a percentage (% ILS), median survival time and the % ILS based on median survival time.

(58) Results:

(59) The vehicle-treated control group had a median day of death and a median survival time of 10 days. All animals died or were euthanized due to moribundity between days 9 and 11. The maximum loss in mean body eight was 32% (7 g).

(60) Due to the dehydrating effect of treatment with S-(1), animals in groups 2, 3, 5, and 6 that received treatment with S-(1) at a dosage of 20 or 12 mg/kg/dose were given 5% dextrose in lactated Ringer's solution to lessen the debilitating effects of the treatment. Administration of the Ringer's solution was initiated when animal body weights dropped by greater than 10% and continued until the animal recovered, died, or was euthanized.

(61) Intravenous treatment with S-(1), administered at dosages of 20 and 12 mg/kg/dose on a q4d×3 schedule, resulted in a 60% and 45% ILS, respectively, whether the calculation was based on day of death or survival time. The corresponding median days of death and the median survival times were 16 and 14.5 days for the S-(1) dosages of 20 and 12 mg/kg/dose, respectively. One death occurred on day 2 in the group receiving treatment with the dosage of 20 mg/kg/dose. This death may have been treatment-related or may have been a delayed effect of the anesthesia used at the time of the ic tumor implant.

(62) Treatment with temozolomide administered at a dosage of 80 mg/kg/dose given po q4d×3 was quite effective against the growth of the U251 CNS tumor cells with an ILS of 380% when calculated based on median day of death (48 days) and 400% when calculated based on median survival time (50 days). There was one survivor in this group at the time of study termination on day 100. Treatment with temozolomide was tolerated without treatment-related deaths and with only a minimal loss (8%, 2 g) in mean body weight.

(63) Administration of S-(1) given iv at a dosage of 20 mg/kg/dose in combination with temozolomide given po at a dosage of 80 mg/kg/dose resulted in an ILS of 575%, with the median day of death and the median survival time both being 67.5 days. The maximum loss in mean body weight loss observed in this treatment group was 22% (5 g). The loss was recovered following cessation of treatment. One death occurred on day 5, two deaths occurred on day 9, and one death occurred on day 16, possibly from toxicity of S-(1). The remaining six animals in the group did respond to therapy with deaths occurring between days 64 and 90.

(64) The group receiving treatment with S-(1) at a dosage of 12 mg/kg/dose in combination with temozolomide at 80 mg/kg/dose responded more favorably than the group receiving the higher dosage of S-(1). The median day of death was day 73, with an ILS of 630%, and the median survival time was 75 days with an ILS of 650%. Results are shown in FIG. 3.

(65) TABLE-US-00006 TABLE 4 Summary of U251 Orthotopic Intracranial Xenograft Study S-(1) 20 mg/kg, ip. 60% ILS (survivors day 100 = 0) QD1, 5, 9 Group 2 Group 2 vs. 2, P = 0.000 S-(1) 12 mg/kg, ip. 45% ILS (survivors day 100 = 0) QD1, 5, 9 Group 3 Group 1 vs. 3 P = 0.000 TMZ 80 mg/kg, po. 380% ILS (survivors day 100 = 1) QD1, 5, 9 Group 4 Group 2 vs 3, P = 0.450 TMZ 80 mg/kg, po. + 575% ILS (survivors day 100 = 1 S-(1) 20 mg/ip. Group 4 vs 5, P = 0.683 QD1, 5, 9 Group 5 TMZ 80 mg/kg, po. + 650% ILS (survivors day 100 = 3) S-(1) 12 mg/kg ip. Group 4 vs. 6, P = 0.046 QD1, 5, 9 Group 6 Group 5 vs. 6, P = 0.108

(66) Monotherapy with temozolomide produced an excellent effect against the U251 human CNS tumor cells, while S-(1), administered alone, produced a moderate effect. Combination treatment with S-(1) and temozolomide was quite effective at both dosages of S-(1). The better response was seen in the combination group receiving the lower S-(1) dosage, possibly because of toxicity of S-(1) at the dosage of 20 mg/kg/dose.

(67) Toxicology. Central Nervous System (CNS) Safety Study in Rats:

(68) The objective of this study was to evaluate the acute pharmacological effects of S-(1) on the central nervous system following intravenous administration in the male albino rat. This study was conducted at ClinTrials BioResearch (CTBR, Montreal, Quebec, Canada) as a GLP study.

(69) Results:

(70) All animals were observed twice daily for signs of ill health or reaction to treatment, except on day of arrival and necropsy. A Functional Observation Battery (FOB) assessment, along with grip strength, hind limb splay and body temperature measurements were performed for all animals once pre-dose and at approximately 15 minutes, 1 hour, 4 hours, 8 hours and 24 hours post-dose. After the last observation, all animals were euthanized without further examination. There were no deaths or treatment-related clinical signs.

(71) A single intravenous administration of S-(1) at 6.25, 12.5 or 25 mg/kg had no biological effect on central nervous system, when measured by qualitative assessment of the functional observational battery and quantitative assessment of grip strength, hind limb splay, and body temperature at approximately 15 minutes, 1, 4, 8 and 24 hours post-dose.

(72) TABLE-US-00007 TABLE 5 Results of CNS Safety Study Dose No. of Group/ Dose Level Concentration Dose Volume Animals Identification (mg/kg) (mg/kg) (mL/kg) Males 1. Control 0 0 3 8 2. S-(1) 6.25 2.08 3 8 3. S-(1) 12.5 4.17 3 8 4. S-(1) 25 8.33 3 8

(73) Conclusions: Dose levels of 6.25, 12.5 and 25 mg/kg had no biological effects on the central nervous system of male Sprague Dawley rats following administration by intravenous injection. The no adverse effect level for this study is 25 mg/kg.

(74) Clinical Studies of S-(1)

(75) Two Phase 1 trials examining the safety and pharmacokinetics of multiple ascending doses of single agent, intravenous S-(1), when administered in two different treatment schedules, are ongoing. Both trials have enrolled patients with advanced solid tumors, non-Hodgkin's lymphoma or Hodgkin's lymphoma that have recurred or progressed following at least standard therapy; patients with malignancies for which there is no standard therapy; patients who are not candidates for standard therapy; or patients who have chosen not to pursue standard therapy. In Study S-(1)-01, intravenous S-(1) is administered weekly for 3 consecutive weeks followed by one week of no therapy; treatment cycles are repeated every 28 days in patients who remain eligible for continued treatment. Study S-(1)-01 is currently being conducted in the United States. In Study S-(1)-02, intravenous S-(1) is administered every 3 weeks; treatment cycles are repeated every 21 days in patients who remain eligible for continued treatment. Study S-(1)-02 is currently being conducted in the United States and in Israel.

(76) TABLE-US-00008 TABLE 6 Clinical Studies of S-(1) Study No. Patients Enrolled Phase Dose Group Protocol Regimen S-(1)-01 27 S-(1) i.v. in a 50% cremaphor, Phase 1  7 mg/m.sup.2: 4 pts 50% ethanol formulation over 1  14 mg/m.sup.2: 4 pts hour or less weekly x 3 followed  28 mg/m.sup.2: 4 pts by 1 week of no therapy (28-day  56 mg/m.sup.2: 4 pts treatment cycle) 127.5 mg/m.sup.2: 7 pts.sup.  185 mg/m.sup.2: 4 pts S-(1)-02 20 S-(1) i.v. in a 15% cremphor, Phase 1  56 mg/m.sup.2: 2 pts 85% ethanol formulation over .sup. 84 mg/m.sup.2: 1 pt 1 hour or less every 3 weeks 126 mg/m.sup.2: 3 pts (21-day treatment cycle) 185 mg/m.sup.2: 6 pts 160 mg/m.sup.2: 8 pts
Clinical Safety—Adverse Events:

(77) Dose Limiting Toxicity: Dose limiting toxicity has been observed in one patient in Study S-(1)-01 who received 185 mg/m.sup.2 of S-(1). The dose limiting toxicity was Grade 3 sensory neuropathy. Dose limiting toxicity has been observed in 2 patients in Study S-(1)-02 at a dose of 185 mg/m.sup.2. In both patients, the dose limiting toxicity was Grade 3 sensory neuropathy.

(78) Clinical Pharmacokinetics:

(79) Pharmacokinetic data for the Phase 1 studies S-(1)-01 and S-(1)-02 are summarized in Table 7. Plasma pharmacokinetics of S-(1) were assessed following the first dose of S-(1) in both studies. Plasma samples were analyzed for S-(1) using a validated LC/MS-MS method. Two different formulations of S-(1) were administered in the two different Phase 1 studies: a 50:50 (w/v) formulation of Cremophor EL-P/ethanol, administered in the S-(1)-01 study, and a 15:85 (w/v) formulation of Cremophor EL-P/ethanol, administered in the S-(1)-02 study. Analysis of the PK data revealed that the plasma AUC of S-(1) appeared to be relatively dose proportional, with a moderate level of interpatient variability. Plasma t.sub.1/2 values ranged from 3.45-8.4 hours for the S-(1)-01 study, and from 3.43-8.96 for the S-(1)-02 study. Clearance did not appear to be dose-dependent in either study. There did not appear to be any difference in pharmacokinetic parameters, including V.sub.ss, for the two different formulations of S-(1).

(80) TABLE-US-00009 TABLE 7 Plasma Pharmacokinetic Parameters for S-(1) Dose AUC.sub.0−24 Cl Vss t.sub.1/2 (mg/m.sup.2) (ng/ml*hr) (L/hr/m.sup.2) (L/m.sup.2) (hr) Study S-(1)-01 (50:50 Cremophor EL-P/ethanol formulation) 14 457 ± 297 35.5 ± 29.7 23.2 ± 74.5  8.4 ± 11.5 28 682 ± 322 43.5 ± 14.5 309 ± 161 3.45 ± 1.09 56 2070 ± 888  28.6 ± 13.2 216 ± 70  5.66 ± 3.63 85 5970 ± 3900 16.6 ± 9.26  142 ± 75.6 5.52 ± 5.26 127.5 4460 ± 2140 30.2 ± 11.9 239 ± 116 6.59 ± 2.60 Study S-(1)-02 (15:85 Cremophor EL-P/ethanol formulation) 56 2530 ± 847  18.9 ± 9.53  232 ± 93.6 8.96 ± 7.09 84 3710 15.4 300 — 126 6110 ± 3030 22.1 ± 11.2 166 ± 105 6.27 ± 3.64 185 7260 ± 2620 24.7 ± 12.2 215 ± 116 3.43 ± 7.31
Summary of a Study of S-(1): Administered Weekly in Patients with Advanced Cancer:

(81) In preclinical studies, S-(1) demonstrated antitumor activity against multiple human tumor xenografts in nude mice, including xenografts that expressed mdr-1 and that were resistant to other taxanes. The safety and tolerability of S-(1) when administered weekly for 60 minutes for 3 weeks followed by a 1 week rest (4 week cycle) was examined in this Phase 1 dose escalation study in patients (pts) with advanced neoplasms.

(82) Treatment cohorts consisted of 3 pts and were expanded to 6 pts in the face of dose-limiting toxicity (DLT); pts could remain on study until the development of progressive disease or an intolerable adverse event. DLT was defined as Gr 4 heme toxicity lasting 7 days; febrile neutropenia, Gr 3 thrombocytopenia with bleeding, Gr 3 elevation of transaminases lasting 7 days or any other Gr 3/4 toxicity other than nausea or vomiting.

(83) Results: 25 pts (M:F 16:9, median age 60, range 24-86) were enrolled in 7 dose levels ranging from 7-185 mg/m.sup.2. Pts' cancers included colorectal (6 pts); NSCLC (2); prostate (2); squamous cell carcinoma (2) and 1 pt each with cervical, breast, ovarian, gastric, pancreatic, bladder endometrial, NSCLC, SCLC, glioblastoma, melanoma, renal cell and hepatocellular carcinoma. All pts but 1 had received prior chemotherapy (median no. prior treatments: 3 (range, 1-7). Drug related adverse events included nausea, vomiting, diarrhea, fatigue, anorexia, rash, anemia and peripheral neuropathy. DLT of Grade 3 peripheral neuropathy has been observed. PK data to date reveal that AUC is generally dose linear. At a dose of 127.5 mg/m.sup.2, clearance was 30.2±11.9 L/hr/m.sup.2 and t.sub.1/2 8.6±1.3 hrs. Anti-neoplastic activity was seen in a patient with pancreatic cancer.

(84) In conclusion, S-(1) can be safely administered in doses of up to 185 mg/m.sup.2 weekly×3 in heavily pre-treated patients. It is predicted that in chemo-naïve patients, or patients with more limited exposure to chemotherapy, that the dose could be escalated even further. S-(1) has activity in pancreatic cancer. PK is dose linear and predictable.

(85) Summary of a Phase 1 Study of S-(1): Administered Every 21 Days in Patients with Advanced Cancer:

(86) In preclinical studies, S-(1) suppressed the growth of multiple human tumor xenografts in nude mice, including xenografts that expressed mdr-1 and that were resistant to other taxanes. The safety and tolerability of S-(1) when administered every 21 days was examined in this Phase 1 dose escalation study in patients (pts) with advanced neoplasms.

(87) S-(1) was administered over 1 hour every 21 days in ascending doses to groups of 3 pts. Treatment cohorts were expanded to 6 pts in the face of dose-limiting toxicity (DLT); pts could remain on study until the development of progressive disease or an intolerable adverse event. DLT was defined as Gr 4 heme toxicity lasting 7 days; febrile neutropenia, Gr 3 thrombocytopenia with bleeding, Gr 3 elevation of transaminases lasting 7 days or any other Gr 3/4 toxicity other than nausea or vomiting.

(88) Results: 14 patients (M:F 5:9, median age 58.5, range 49-77) were enrolled in 5 dose levels ranging from 56-185 mg/m.sup.2. Patients' cancers included colorectal (5 pts), esophageal (2), pancreatic (2), NSCLC (2), breast (2) and ovarian (1). All patients had received prior chemotherapy (median no. prior treatments: 3 (range, 2-10)). Drug related adverse events included mucositis, vomiting, diarrhea, neutropenia, thrombocytopenia, myalgias and peripheral neuropathy. Only 1 pt. experienced Gr 4 neutropenia. DLT of Gr 3 peripheral sensory neuropathy was observed at a dose of 185 mg/m.sup.2. At a dose of 160 mg/m.sup.2 no DLT was observed. 1 pt with pancreatic cancer had a confirmed response to S-(1). PK data reveal that AUC is generally dose linear. At a dose of 126 mg/m.sup.2, clearance was 24.7±12.2 L/hr/m.sup.2 and t.sub.1/2 was 10.6±7.1 hrs.

(89) It was determined that S-(1) can be safely administered in a dose of 160 mg/m.sup.2 every 21 days in heavily pre-treated patients. It is predicted that in chemo-naïve patients, that the dose could be escalated even further. The dose limiting toxicity was Gr 3 peripheral neuropathy. S-(1) appears to have activity in pancreatic cancer. PK is dose linear and predictable.

(90) Oral Dosing of the Compound of Formula S-(1):

(91) The compound of formula S-(1) when dosed either oral or iv shows efficacy in mouse xenografts. From these efficacy studies and MTD studies, the “apparent oral bioavailability” of the compound of formula S-(1) in nude mice would be in the 40 to 80% range.

(92) TABLE-US-00010 TABLE 8 Formula S-(1) Injection Formulation Administered Orally: Mouse PK Parameters S-(1) - IV 10/mg/kg S-(1) - Oral 10 mg/kg C.sub.max (nM) na C.sub.max (nM) 460 t.sub.max (hr) na t.sub.max (hr) 6.0 t ½ (hr) 6.1 t ½ (hr) 5.1 AUC.sub.last (nM .Math. hr) 6298 AUC.sub.last (nM .Math. hr) 3434 AUC.sub.inf (nM .Math. hr) 7738 AUC.sub.inf (nM .Math. hr) 3609 CL (L/hr/kg) 1.49 F (%) 47 Vdss (L/kg) 13.35
Efficacy of Orally Dosed S-(1) in Mouse Neuroblastoma Xenografts:

(93) Summary of Experimental Protocol: Human neuroblastoma tumor cells were implanted via subcutaneous injection of 1-10×10.sup.6 cells in nude mice. Tumors were allowed to grow to 200 mg+/−50 mg in size. Drug dosing was initiated, and tumor volume and body weight were recorded twice weekly.

(94) Data Analysis:

(95) The mean value for % body weight change and % tumor volume increase for each experimental group was plotted including error bars for the standard error of the mean. Other metrics for assessing antitumor effects include: % T/C values as calculated by the following formula:

(96) $\frac{\%\mathrm{Mean}\mathrm{Tumor}\mathrm{Volume}\mathrm{of}\mathrm{Treated}\mathrm{Group}}{\%\mathrm{Mean}\mathrm{Tumor}\mathrm{Volume}\mathrm{of}\mathrm{Control}\mathrm{Group}}$$\mathrm{Log}\mathrm{kill}=\left(T-C\right)/3.32\times \left(\mathrm{Td}\right)$ T is the time in days for the median tumor volume to reach 1 gram in treated group C is the time in days for the median tumor volume to reach 1 gram in control group Td is the tumor doubling time in days Cures are excluded from T-C calculations

(97) Results are shown in FIGS. 4 to 9.

(98) S-(1) Rat Oral Bioavailability:

(99) IV formulation (Cremophor E1 P/alcohol) and a suspension of the compound of formula S-(1) in aqueous medium.

(100) Several studies were performed to investigate specific parameters. It was found that there is a gender difference but it does not influence bioavailability (F) overall, the formulation dilution does not impact F, there is saturation of the absorption that seems formulation dependent, and the data is reproducible between experiments.

(101) From the solubility and compatibility studies between the compound of formula S-(1) and excipients that could be used, a reasonable list of formulations were evaluated in vivo. Pharmacokinetics results for 9 formulations administered to female rats by oral gavage are listed in Table 9 below.

(102) TABLE-US-00011 TABLE 9 AUC of Selected Evaluated Oral Formulations AUC last AUCinf/Dose (hr*ng/ml) (hr*kg*ng/mL/mg) Cremophor:EtOH 50:50/dil. 1:9 2220 426 Tween 80:EtOH:Labrafil:Labrasol 5740 427 25:25:35:35/dil. 1:1 Solutol:EtOH 60:40/dil. 1:1 6590 503 Vit E-TPGS:Cre:EtOH:D5W 1450 266 50:17:17:16/dil. 1:3 Solutol:Cre:EtOH:D5W 2370 431 50:17:17:16/dil. 1:3 Lutrol:Cre:EtOH:D5W 2380 436 40:20:20:20/dil. 1:3 Oleic Acid:Cre:Captex:Capmul:D5W 2610 555 8:25:33:17:17/dil. 1:3 Solutol:EtOH:Labrafil:labrasol 2850 475 50:10:25:15/dil. 1:1 Gelucire:Labrafil:PEG300 3120 520 60:20:20/dil. 1:1

(103) In addition, when contemplating oral administration, one typically tries to evaluate the oral bioavailability of the compound. Using the 50:50 Cremophor:Ethanol formulation as a case study given orally (AUC last: 2540; norm AUCinf: 477), F was calculated using two different IV formulations as reference, the 50:50 Cremophor:Ethanol and the 15:85 Cremophor:Ethanol As shown in Table 10 below, the calculated F value is significantly impacted by the PK values of the IV reference.

(104) TABLE-US-00012 TABLE 10 Calculated F AUC last AUCinf/Dose Calculated F (hr*ng/ml) (hr*kg*ng/mL/mg) (%) Cremophor:EtoH 50:50/ 14600 2040 ~23% dil. 1:9 dosed IV Cremophor:EtoH 15:85/ 4470 970 ~49% dil. 1:9 dosed IV

(105) Accordingly, the invention comprises a polyethoxylated castor oil formulation comprising the compound of formula S-(1). Preferably, the polyethoxylated castor oil formulation comprises about 50% ethanol and about 50% polyethoxylated castor oil or about 85% ethanol and about 15% polyethoxylated castor oil.