LONG-ACTING SPIRO-ISOXAZOLINE ANTIPARASITIC COMPOSITIONS

Abstract

The invention describes a long-acting composition comprising a spiro-azetidine isoxazoline of Formula (1) or (2)

##STR00001##

wherein R.sup.1a, R.sup.1b, R.sup.1c and R.sup.2 are as described herein, and stereoisomers thereof. The composition is a veterinary composition and also comprises a glycol ether and at least one veterinarily acceptable solvent, and optionally, at least one precipitation inhibitor, antioxidant and additional veterinary agent, and any mixture thereof. The invention also includes a method of treating an animal with a parasitic infestation by administering the long-acting composition to the animal in need thereof.

Claims

1. A long-acting topical composition comprising: a) a spiro-azetidine isoxazoline of Formula (2) ##STR00003## wherein R.sup.1a and R.sup.1c are both chloro and R.sup.1b is fluoro; and R.sup.2 is —CH.sub.2S(O)CH.sub.3, and stereoisomers thereof; b) a glycol ether; c) at least one veterinarily acceptable solvent; d) optionally, at least one precipitation inhibitor; and e) optionally, at least one antioxidant.

2. The composition of claim 1 wherein the spiro-azetidine is (S)-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethenone.

3. The composition of claim 2 wherein in the glycol ether is a diglycol ether selected from diethylene glycol monomethylether, diethylene glycol monoethylether, diethylene glycol monobutylether, dipropyleneglycol monomethyl ether, dipropyleneglycol monomethyl ether, and diethylene glycol dimethyl ether, and further comprising a precipitation inhibitor.

4. The composition of claim 3 wherein the diglycol ether is diethylene glycol monobutylether or diethylene glycol monoethylether and wherein the solvent is selected from the group selected from dimethyl isosorbide, caprylic/capric triglyceride, isopropyl myristate, eucalyptol, benzyl alcohol, benzyl benzoate, propylene glycol laurate, ethanol, isopropanol, oleic acid, propylene glycol caprylate, caprylocaproyl macrogol-8 glyceride, and any mixture thereof, and further comprising an antioxidant.

5. A long-acting topical composition comprising: a. (S)-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; b. diethylene glycol monobutylether or diethylene glycol monoethylether; c. at least one veterinarily acceptable solvent; d. optionally, at least one precipitation inhibitor; and e. optionally, at least one antioxidant.

6. The composition of claim 5 wherein the solvent is selected from dimethyl isosorbide, caprylic/capric triglyceride, isopropyl myristate, oleic acid, eucalyptol, benzyl alcohol, benzyl benzoate, ethanol, propylene glycol caprylate, propylene glycol laurate, caprylocaproyl macrogol-8 glyceride, and isopropanol, and any mixture thereof.

7. The composition of claim 6 wherein the solvent is selected from dimethyl isosorbide, propylene glycol laurate, and caprylocaproyl macrogol-8 glyceride, or a mixture thereof, and further comprising at least one precipitation inhibitor.

8. The composition of claim 5 wherein the solvent is selected from isopropyl myristate, oleic acid, eucalyptol, benzyl alcohol, benzyl benzoate, ethanol, and isopropanol, and any mixture thereof, and further comprising at least one precipitation inhibitor.

9. The composition of claim 1, further comprising at least one additional veterinary agent.

10. The composition of claim 1 wherein said composition is a topical composition.

11. A method of treating an animal with a parasitic infestation comprising administering a long-acting topical composition comprising: a) a spiro-azetidine isoxazoline of Formula (2) ##STR00004## wherein R.sup.1a and R.sup.1c are both chloro and R.sup.1b is fluoro; and R.sup.2 is CH.sub.2S(O)CH.sub.3, and stereoisomers thereof; b) a glycol ether; c) at least one veterinarily acceptable solvent; d) optionally, at least one precipitation inhibitor; and e) optionally, at least one antioxidant.

12. The method of claim 11 wherein the spiro-azetidine isoxazoline is (S)-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone.

13. The method of claim 12 wherein the glycol ether is a diglycol ether selected from diethylene glycol monobutylether or diethylene glycol monoethylether, wherein the solvent is selected from dimethyl isosorbide, caprylic/capric triglyceride, isopropyl myristate, oleic acid, eucalyptol, benzyl alcohol, benzyl benzoate, ethanol, propylene glycol caprylate, propylene glycol laurate, caprylocaproyl macrogol-8 glyceride, and isopropanol, and any mixture thereof, and optionally, the composition further comprises at least one precipitation inhibitor, and optionally, at least one additional veterinary agent.

14. The method of claim 13 further comprising at least one antioxidant.

15. The method of claim 11 wherein the animal is a companion animal or livestock and the composition is administered topically at least once every 3 or more months.

16. The method of claim 15 wherein the animal is a companion animal.

17. The method of claim 15 wherein the animal is livestock.

18. The method of claim 16 wherein the companion animal is canine.

19. The method of claim 17 wherein the livestock is bovine.

20. The method of claim 15 wherein the composition is administered topically once every 3 months.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0030] FIG. 1. Depicts Flux Permeability.

[0031] FIG. 2. Depicts Dose Dependent Permeability Flux using Franz Cell Diffusion.

[0032] FIG. 3. Depicts Dose Constant, Butyl Digol:Dimethyl Isosorbide Flux/Kp Determination.

[0033] FIG. 4. Depicts Dose Constant Butyl Digol:Oleic Acid Flux/Kp Determination.

[0034] FIG. 5. 3-Month Canine Pharmacokinetics

DETAILED DESCRIPTION

[0035] It is to be understood by one of ordinary skill in the art that the present discussion is a description of exemplary embodiments only and is not intended as limiting the broader aspects of the present invention, which broader aspects are embodied in the exemplary construction. In fact, it will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the scope or spirit of the invention. For instance, features illustrated or described as part of one embodiment can be used in another embodiment to yield a still further embodiment. It is intended that the present invention cover such modifications and variations as come within the scope of the appended claims and their equivalents.

[0036] The spiro-azetidine compounds of the instant invention are characterized according to either Formula (1) or Formula (2) below:

##STR00002##

wherein R.sup.1a, R.sup.1b, and R.sup.1c are each independently hydrogen, chloro, bromo, fluoro, or trifluoromethyl; and R.sup.2 is ethyl, propyl, isopropyl, isobutyl, cyclopropyl, —C(OH)(CH.sub.3).sub.2, —CH.sub.2cyclopropyl, —CH.sub.2CF.sub.3, —CH.sub.2OH, —CH.sub.2SCH.sub.3, —CH.sub.2S(O)CH.sub.3, —CH.sub.2S(O).sub.2CH.sub.3, —CH.sub.2SCF.sub.3, 2,2-difluorocyclopropyl, 1,1-dioxidothietane, and —CH.sub.2-1H-pyrazole.

[0037] The spiro-azetidine isoxazoline compounds can be synthesized according to procedures described in WO2012/120399.

[0038] It is to be understood that the spiro-azetidine isoxazoline compounds of the invention contain an asymmetric carbon (chiral) atom, thus compounds of the invention can exist as two or more stereoisomers. Included within the scope of the present invention are all stereoisomers such as enantiomers (e.g. S and R enantiomers) and diasteromers, all geometric isomers and tautomeric forms of the spiro-azetidine isoxazoline compounds. The spiro-azetidine isoxazolines of the present invention can be racemates, which include the (S) and (R) enantiomers.

[0039] The present invention provides for a composition for the treatment of a parasitic infestation in an animal which comprises a veterinarily effective amount of a spiro-azetidine isoxazoline compound. The spiro-azetidine compounds of the present invention include the compounds selected from: 1-(cyclopropanecarbonyl)-5′-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro{azetidine-3,1′-isobenzofuran}-3′-one; 5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-1-propionyl-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-((trifluoromethyl)thio)ethanone; (5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)(1,1-dioxidothietan-3-yl)methanone; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfinyl)ethanone; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; (S)-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(cyclopropanecarbonyl)-5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-1-(3-methylbutanoyl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 1-(2-cyclopropylacetyl)-5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 1-butyryl-5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-1-(2-(methylthio)acetyl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-1-(2,2-difluorocyclopropanecarbonyl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-1-(4,4,4-trifluorobutanoyl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-3′-one; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-methylpropan-1-one; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-hydroxyethanone; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)propan-1-one; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-hydroxy-2-methylpropan-1-one; 2-cyclopropyl-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)methanone; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-3-methylbutan-1-one; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(1H-pyrazol-1-yl)ethanone; cyclopropyl(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-111)methanone; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)butan-1-one; 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 2-(methylsulfonyl)-1-(5′-(5-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)methanone; 1-(5′-(5-(3,5-dichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(3-chloro-5-(trifluoromethyl)phenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(3,4-dichloro-5-(trifluoromethyl)phenyl)-5-(trifluoromethyl)-4,5-dihydro-isoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)-ethanone; 1-(5′-(5-(4-bromo-3,5-dichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(3,5-bis(trifluoromethyl)phenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(3-bromo-5-chlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(4-chloro-3,5-bis(trifluoromethyl)phenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(3-chloro-5-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; 1-(5′-(5-(3-chloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone; and 2-(methylsulfonyl)-1-(5′-(5-(trifluoromethyl)-5-(3-(trifluoromethyl)phenyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)ethanone, including the stereoisomers, veterinarily acceptable salts, and the crystalline and amorphous forms thereof.

[0040] The preferred spiro-azetidine compound is 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone (i.e., Formula 2, wherein R.sup.1a and R.sup.1c are each chloro, R.sup.1b is fluoro, and R.sup.2 is —CH.sub.2S(O).sub.2CH.sub.3. The more preferred compound is the (S) enantiomer of 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone, which is also referred to herein as Compound 1.

[0041] Veterinary compositions suitable for the delivery of compounds of the present invention and methods for their preparation will be readily apparent to those skilled in the art. Such compositions and methods for their preparation may be found, for example, in ‘Remington's Pharmaceutical Sciences’, 19th Edition (Mack Publishing Company, 1995).

[0042] In the present invention, the composition comprises a glycol ether. The glycol ether includes the mono-, di-, and tri-glycol ethers. Non-exclusive examples of the mono-glycol ethers include: ethylene glycol monomethyl ether (EGMME), ethylene glycol monoethyl ether (EGMEE), ethylene glycol monopropyl ether (EGMPE), ethylene glycol monoisopropyl ether (EGMIE), propylene glycol mono-t-butyl ether (PGMBE), propylene glycol propyl ether (PGMPE), propylene glycol monomethyl ether (PGMME), propylene glycol monoethyl ether (PGMEE), and the like. Non-exclusive examples of the di-glycol ethers include: diethylene glycol monomethylether (DEGMME), diethylene glycol monoethylether (DEGMEE), diethylene glycol monobutylether (DEGMBE, butyl digol), dipropyleneglycol monomethyl ether (DPGMME, DPG), diethylene glycol dimethyl ether (DEGDME), and the like. Non-exclusive examples of the tri-glycols include: tripropylene glycol monomethyl ether (TPGMME), tripropylene glycol monoethyl ether (TPGMEE), triethylene glycol monoethyl ether (TEGMEE), triethylene glycol monomethyl ether (TEGMME), and the like. The glycol ethers also include the acetylated glycol ethers, for example, diethylene monoethylether acetate and diethylene monobutylether acetate. The preferred glycol ether is selected from the group consisting of diethylene glycol monomethylether (DEGMME), diethylene glycol monoethylether (DEGMEE), diethylene glycol monobutylether (DEGMBE, butyl digol), dipropyleneglycol monomethyl ether (DPGMME), and diethylene glycol dimethyl ether (DEGDME), and mixtures thereof. A preferred glycol ether is DEGMBE. Another preferred glycol ether is DEGMEE.

[0043] In the present invention, the composition comprises at least one veterinarily acceptable solvent. Non-limiting examples of solvents include glycols, lactones, cyclic carbonates, glyceryl acetates, alcohols, and triglycerides.

[0044] Non-limiting examples of glycols include: ethylene glycol, propylene glycol, propane-1,2-diol, butylene glycol, polyethylene glycols (PEGs, e.g., hexaethylene glycol, pentaethylene glycol, tetraethylene glycol, triethylene glycol), methoxypolyethylene glycols (MPEGs, e.g., MPEG 350 and MPEG 550), polypropylene glycols (PPGs, e.g., PPG-10, PPG-55, PPG-9, PPG-17, and the like)), polybutylene glycol (PBG), and the like. The preferred glycol is a polyethylene glycol selected from hexaethylene glycol, pentaethylene glycol, tetraethylene glycol, and triethylene glycol. The more preferred glycol is triethylene glycol.

[0045] Non-limiting examples of suitable lactones include: δ-valerolactone, γ-caprolactone, γ-hexalactone, γ-butyrolactone, δ-hexalactone, γ-dodecalactone, δ-nonalactone, δ-decalactone, γ-decalactone, γ-caprolactone, δ-valerolactone, and δ-dodecalactone and other alkyl lactones and combinations thereof. The preferred lactone is selected from γ-hexalactone, γ-butyrolactone, δ-hexalactone, γ-dodecalactone, δ-decalactone, γ-decalactone, and δ-dodecalactone. The more preferred lactone is γ-hexalactone.

[0046] Non-limiting examples of cyclic carbonates include: 4-methyl-1,3-dioxolan-2-one, 4-ethyl-1,3-dioxolan-2-one, 1,3-dioxolan-2-one, 4-propyl-1,3-dioxolan-2-one, 4,4-dimethyl-1,3-dioxolan-2-one, 4,5-dimethyl-1,3-dioxolan-2-one, 1,3-dioxan-2-one, 4-methyl-1,3-dioxan-2-one, and the like. The preferred cyclic carbonate is selected from 4-methyl-1,3-dioxolan-2-one, 4-ethyl-1,3-dioxolan-2-one, and 4-methyl-1,3-dioxan-2-one. The more preferred cyclic carbonate is 4-methyl-1,3-dioxolan-2-one.

[0047] The glyceryl acetates refer to the esters of glycerol and include the monoacetylglycerols, diacetylglycerols, and triacetylglycerol.

[0048] The alcohols refer to C.sub.1-C.sub.18 aliphatic alcohols and to C.sub.4-C.sub.6 cyclic and aromatic (as applicable) alcohols. The alcohols also include the fatty alcohols. Non-limiting examples of the aliphatic alcohols include methanol, ethanol, propanol, isopropanol, butanol, pentanol, hexanol, decanol, dodecanol, myristyl, cetyl, stearyl, oleic, octyldecyl, and the like. Non-limiting examples of cyclic and aromatic alcohols include cyclobutanol, cyclopentanol, cyclohexanol, benzyl alcohol, and the like.

[0049] The triglycerides include short chain, medium chain, and long chain triglycerides. Triglycerides also include mono- and di-esters as well as mono- and di-propylides, for example, Captex 200, Captex 300, Captex 355, and the like. The short chain triglycerides are fatty acids with aliphatic tails of fewer than six carbon atoms, for example, butyric acid and triacetin. The medium chain and long chain triglycerides are fatty acids with aliphatic tails of 6-12 carbon atoms and 13-21 carbon atoms, respectively. Some non-limiting medium chain fatty acids include: capric, caprylic, lauric, and the like. Some long-chain fatty acids (saturated and unsaturated) include: stearic, oleic, linoleic acid, myristic, and the like. Additional non-limiting examples of triglycerides include: castor oil, cottonseed oil, sesame oil, linseed oil, safflower oil, peanut oil, soybean oil, coconut oil, olive oil, corn oil, almond oil, vegetable oil, glyceryl stearates, glyceryl hexanoates, caprylic/capric glycerides, glyceryl cocoate, caprylic glycerides, glyceryl monooleate, glyceryl ricinoleate, capric glycerides, and the like. The fatty acids also include the aromatic acids like benzoic acid and the diacids, for example, succinic, adipic, azelaic, sebacic, and the like, as well as the esters isopropyl myristate, ethyl oleate, ethyl laurate, dibutyl adipate, propylene glycol monocaprylate, propylene glycol monolaurate (Lauroglycol) and the spider esters.

[0050] In the present invention, the veterinarily acceptable solvent also includes anionic, cationic, and non-ionic surfactants, and any mixture thereof. Non-limiting examples of these surfactants include: alkaline stearates (for example, sodium, potassium, or ammonium stearate, calcium stearate, and triethanolamine stearate), alkyl sulphates (for example, sodium laurel sulphate, sodium dodecyl sulphate, sodium cetyl sulphate), fatty acid sorbitan esters (for example, Span 20), polyoxyethylenated sorbitan esters (for example, polysorbate 80), polyoxyethylenated alkyl ethers, polyethylene glycol stearate, polyoxyethylenated derivatives of castor oil (for example Cremaphor EL), polyglycerol esters, caprylocaproyl macrogol-8 glyceride (Labrasol), Kolliphor HS15 (Macrogol 15 hydroxystearate or Polyoxyl 15 hydroxystearate), and the like. Additional veterinarily acceptable solvents include: terpene alkaloids (for example, limonene, eucalyptol, menthol); pyrrolidones (for example, 2-pyrrolidone, N-methyl pyrrolidone, and azone), glycerol formal, tetraglycol, tetrahydrofurfuryl alcohol, solketal, dimethyl isosorbide (Arlasolve), which is a dimethyl ether of an anhydride of a sorbitol isomer. Further, the veterinarily acceptable solvent includes spreading agents, precipitation inhibitors, and stabilizers. Non-limiting examples of spreading agents include: siloxanes (e.g., dimethyl polysiloxane), indapoles (e.g., polyisobutylene), and the like. Non-limiting examples of precipitation inhibitors include: poloxamers (e.g., pluronic F68 and pluronic F127), indapols (e.g., polyisobutylene), polyvinyl pyrrolidones (PVP's) (e.g., PVP K-15, K-18, K-20 and K-90), alginates, xanthans, and celluloses (e.g., methyl- and ethyl cellulose), and the like. Non-limiting examples of stabilizers (pH adjuster) include: citric acid, lactic acid, mono-, di- and tri-ethanolamine, meglumine, and the like.

[0051] The long-acting composition of the present invention further comprises an antioxidant. Non-limiting examples of antioxidants include: ascorbic acid, vitamin E (tocopherol), vitamin E derivatives, butylated hydroxanisole (BHA), butylated hydroxytoluene (BHT), propyl gallate, thioglycerol, citric acid, and the like.

[0052] The long-acting composition of the present invention comprises a spiro-azetidine isoxazoline, a glycol ether, at least one veterinarily acceptable solvent, or a mixture of more than one veterinarily acceptable solvents as described herein.

[0053] The long-acting composition of the present invention comprises 1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone, a glycol ether, at least one veterinarily acceptable solvent, or a mixture of more than one veterinarily acceptable solvents as described herein.

[0054] The long-acting composition of the present invention comprises (S)-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone, a glycol ether, at least one veterinarily acceptable solvent, or a mixture of more than one veterinarily acceptable solvents as described herein.

[0055] Such compositions are prepared in a conventional manner in accordance with standard medicinal or veterinary practice.

[0056] The amounts of these spiro-azetidine isoxazoline compounds are easily determined by a skilled artisan and further depend on the dose amount and dose volume of the final composition. Representative amounts of a veterinarily effective amount of a spiro-azetidine isoxazoline compound ranges from about 0.5 mg/kg to about 50 mg/kg, with a preferred range of about 5 mg/kg to about 40 mg/kg. An even more preferred dose of a spiro-azetidine isoxazoline compound is about 10 mg/kg to about 30 mg/kg. An even more preferred dose of a spiro-azetidine isoxazoline compound is about 15 mg/kg to about 25 mg/kg.

[0057] The spiro-azetidine isoxazoline compositions of the present invention are useful as parasiticides for the control and treatment of parasitic infestations in an animal. The veterinary compositions of the present invention have utility as a parasiticide, in particular, as an ectoparasitic. The preferred ectoparasites are acarines and insects. The compositions may, in particular, be used in the fields of veterinary medicine, livestock husbandry, and the maintenance of public health: against acarines and insects which are parasitic upon animals, particularly domestic animals such as dogs, cats, cattle, sheep, goats, horses, llamas, bison, and swine, more particularly cats, dogs, and cattle. Some non-limiting examples of acarine parasites include: ticks (e.g., Ixodes spp., Rhipicephalus spp., Boophilus spp., Amblyomma spp., Hyalomma spp., Haemaphysalis spp., Dermacentor spp., Ornithodorus spp., and the like); and mites (e.g., Dermanyssus spp., Sarcoptes spp., Psoroptes spp., Eutrombicula spp., Chorioptes spp., Demodex spp., and the like). Some non-limiting examples of parasitic insects include: chewing and sucking lice (e.g., Damalinia spp., Linognathus spp., and the like); fleas (e.g., Siphonaptera spp., Ctenocephalides spp., and the like); and flies, mosquitos, and midges (e.g., Order Diptera; Aedes spp., Anopheles spp., Tabanidae spp., Haematobia spp., Stomoxys spp., Dermatobia spp., Simuliidae spp., Ceratopogonidae spp., Psychodidae spp., Cochliomyia spp., Muscidae spp., Hypoderma spp., Gastrophilus spp., Simulium spp., and the like); true bugs (e.g., Order Hemiptera); cockroaches (Periplaneta spp, Blatella spp) and wasps and ants (Hymenoptera spp).

[0058] The composition of the present invention can also be used for the treatment of endoparasites, for example, heartworms, roundworms, hookworms, whipworms, tapeworms, fluke, and other cestodes and trematodes. The gastrointestinal roundworms include, for example, Ostertagia ostertagi (including inhibited larvae), O. lyrata, Haemonchus placei, H. similis, H. contortus, Toxocara canis, T. leonina, T. cati, Trichostrongylus axei, T. colubriformis, T. longispicularis, Cooperia oncophora, C. pectinata, C. punctata, C. surnabada (syn. mcmasteri), C. spatula, Ascaris suum, Hyostrongylus rubidus, Bunostomum phlebotomum, Capillaria bovis, B. trigonocephalum, Strongyloides papillosus, S. ransomi, Oesophagostomum radiatum, O. dentatum, O. columbianum, O. quadrispinulatum, Trichuris spp., and the like. Other parasites include: hookworms (e.g., Ancylostoma caninum, A. tubaeforme, A. braziliense, Uncinaria stenocephala); lungworms (e.g., Dictyocaulus viviparus and Metastrongylus spp); eyeworms (e.g., Thelazia spp.); parasitic stage grubs (e.g., Hypoderma bovis, H. lineatum, Dermatobia hominis); kidneyworms (e.g., Stephanurus dentatus); screw worm (e.g., Cochliomyia hominivorax (larvae); filarial nematodes of the super-family Filarioidea and the Onchocercidae Family.

[0059] Non-limiting examples of filarial nematodes within the Onchocercidae Family include the genus Brugia spp. (i.e., B. malayi, B. pahangi, B. timori, and the like), Wuchereria spp. (i.e., W. bancrofti, and the like), Dirofilaria spp. (D. immitis, D. repens, D. ursi, D. tenuis, D. spectans, D. lutrae, and the like), Dipetalonema spp. (i.e., D reconditum, D. repens, and the like), Onchocerca spp. (i.e., O. gibsoni, O. gutturosa, O. volvulus, and the like), Elaeophora spp. (E. bohmi, E. elaphi, E. poeli, E. sagitta, E. schneideri, and the like), Mansonella spp. (i.e., M. ozzardi, M. perstans, and the like), and Loa spp. (i.e., L. loa). In another aspect of the invention, the composition of the present invention is useful for treating endoparasiticidal infection from filarial nematodes within the genus Dirofilaria (i.e., D. immitis, D. repens, D. ursi, D. tenuis, and the like).

[0060] The following list of additional veterinary agents together with which the composition of the present invention can be used is intended to illustrate the possible combinations, but not to impose any limitation. Non-limiting examples of additional veterinary agents include: amitraz, arylpyrazoles, amino acetonitriles, anthelmintics (e.g., albendazole, cambendazole, dichlorvos, fenbendazole, flubendazole, mebendazole, octadepsipeptides, oxantel, oxfendazole, oxibendazole, paraherquamide, parbendazole, piperazines, praziquantel, epsiprantel, thiabendazole, tetramisole, triclabendazole, emodepside, levamisole, pyrantel, oxantel, morantel, monepantel, and the like), avermectins (e.g., abamectin, doramectin, emamectin, eprinomectin, ivermectin, moxidectin, selamectin, and the like), milbemycin, milbemycin oxime, DEET, demiditraz, diethylcarbamazine, fipronil, insect growth regulators (e.g., lufenuron, novaluron, hydroprene, kinoprene, methoprene, and the like), metaflumizone, niclosamide, nitenpyram, permethrin, pyrethrins, pyriproxyfen, spinosad, and the like, and mixtures thereof. In certain instances, compositions of the present invention with at least one additional veterinary agent can result in a greater-than-additive effect, for example, synergy (a synergistic effect).

[0061] The veterinary compositions of the present invention are of particular value in the control of ectoparasites which are injurious to, or spread or act as vectors of diseases in animals, for example those described herein, and more especially in the control of ticks, mites, lice, fleas, midges and biting, nuisance flies, that may cause, for example, leishmaniasis, demidicosis, Lyme, and borreliosis. They are particularly useful in controlling acarines and insects which feed on the skin or tissue or suck the blood of the animal, for which purpose they may be administered topically.

[0062] The spiro-azetidine isoxazoline compound binds tightly to ligand-gated chloride channels, in particular those gated by the neurotransmitter gamma-aminobutyric acid (GABA), thereby blocking pre- and post-synaptic transfer of chloride ions across cell membranes in insects and acarines when exposed by ingestion or contact. This mechanism of action results in lethal uncontrolled activity of the central nervous system of insects and acarines yielding highly efficacious control against said ectoparasite.

[0063] The method of treating an animal with a parasitic infestation comprises the administration of the long-acting composition comprising a therapeutically effective amount of a spiro-azetidine isoxazoline compound. Administration is contemplated as dermal administration, wherein dermal administration comprises topical administration by spot-on, pour-on, spray-on, and comb-on methods. The long-acting composition can be topically applied to the animal in need thereof, by administering an effective amount of the composition thereof to the animal at least once every 2-months, 3-months, 4-months, 5-months, 6-months, 7-months, 8-months, 9-months, 10-months, 11-months, or 12-months. The preferred dosing administration is contemplated to be at least once every 4 to 8 months, and more preferrably at least once every 3 to 6 months. Fractional dosing intervals between 2- and 12-months is also contemplated.

[0064] The veterinary compositions of the present invention also have value for the treatment and control of the various lifecycle stages of arachnids and insects, including egg, nymph, larvae, juvenile and adult stages.

[0065] The present invention also relates to a method of administering a veterinary composition of the present invention to an animal in good health comprising the application to said animal to reduce or eliminate the potential for both animal and human parasitic infestation carried by the animal and to improve the environment in which the animals and humans inhabit.

EXAMPLES

[0066] In the following composition tables, C1 refers to the spiro-azetidine isoxazoline, Compound 1, and SAI represents a different spiro-azetidine isoxazoline compound described herein. Non-limiting veterinarily acceptable compositions are shown below. The amounts are exemplified as weight/volume (w/v). These amounts can readily be converted to mg/mL and normalized weight %, and liquids as mL/mL and normalized weight %. Amounts for solutions are exemplified as volume/volume percent (v/v %) and is determined by dividing solute volume (mL) by the total volume of solution (mL) times 100.

[0067] In the formula examples and tables, the following acronyms are herein described: Captex 355 refers to the medium-chain triglyceride, caprylic/capric triglyceride. PVP-K18 is a polyvinylpyrrolidone with a designated viscosity. Capryol-90 (CP90) is propylene glycol caprylate, also known as 1,2-propanediol monocaprylate. Lauroglycol is propylene glycol laurate, also known as 1,2-propanediol monolaurate. Labrasol (LAB) is a mixture of glyceryl and polyethylene glycol esters (caprylocaproyl macrogol-8 glyceride). Triacetin is glycerol triacetin. Poloxamer F127 is also known as Pluronic F127 and is a di-block copolymer of polyoxyethylene and polyoxypropylene. BHA is butylated hydroxyanisole. BHT is butylated hydroxytoluene. DEGMBE is diethylene glycol monobutyl ether (butyl digol). DEGMEE is diethylene glycol monoethyl ether (Transcutol). Arlasolve (ARL) is dimethyl isosorbide. Butyl digol (BD) is diethylene glycol monobutyl ether (DEGMBE). Tween80 is polysorbate 80. The following acronyms include: C1 is Compound 1, SAI is a spiro-azetidine isoxazoline of Formula 1 or Formula 2, NMP (n-methyl pyrrolidone), OA (oleic acid), BnOH (benzyl alcohol), 2P is 2-pyrrolidone, Span80 is sorbitan oleate, Span20 is sorbitan laurate, C200 is Captex200, C355 is Captex355, C15 PVP is C15 polyvinylpyrrolidone, K29 PVP is K29 polyvinylpyrrolidone, K90 PVP is K90 polyvinylpyrrolidone, EtOH is ethanol, IPA is isopropyl alcohol, IPM is isopropyl myristate, DES is diethyl sebacate, TBAC is tributyl acetocitrate, THFFA is tetrahydro furfuryl alcohol, OZD is 4-decy-1,3-oxazolidin-2-one, NOP is n-octyl pyrrolidone; AZ is azone; DPG is dipropylene glycol monomethyl ether, TRC is transcutol, LG90 is lauroglycol/propylene glycol laurate, and EO is ethyl oleate. Range in the following Formula Examples (1-23) below, is depicted in percent, and in particular, the solids (C1, SAI, BHA, BHT, PVP-K18, poloxomer F127, citric acid, and PVP C15) are measured as w/v % and the remaining liquids are measured as v/v %. The following compositions are non-limiting examples, and include:

[0068] Formula 1

TABLE-US-00001 Component Range % Ideal % C1 or SAI 4-30 25 butyl digol 50-100 65 dimethylisosorbide 5-50 10

[0069] Formula 2

TABLE-US-00002 Component Range % Ideal % C1 or SAI 4-30 25 butyl digol 50-100 62.5 dimethyisosorbide 5-50 7.5 Captex 355 2-20 5.0

[0070] Formula 3

TABLE-US-00003 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.3 dimethyisosorbide 5-50 7.5 BHA 0.01-0.2  0.2

[0071] Formula 4

TABLE-US-00004 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 66.3 dimethyisosorbide 5-50 7.5 BHA 0.01-0.2  0.2 PVP K18 0.1-5.0  1.0

[0072] Formula 5

TABLE-US-00005 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 66.3 dimethyisosorbide 5-50 7.5 BHA 0.01-0.2  0.2 Poloxamer F127 0.1-5.0  1.0

[0073] Formula 6

TABLE-US-00006 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.2 dimethyisosorbide 5-50 7.5 BHA 0.01-0.2  0.2 citric Acid 0.01-1.0  0.1

[0074] Formula 7

TABLE-US-00007 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.5 isopropyl myristate 5-30 7.5

[0075] Formula 8

TABLE-US-00008 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.5 oleic acid 5-30 7.5

[0076] Formula 9

TABLE-US-00009 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.5 1,8-cineole 5-30 7.5 (eucalyptol)

[0077] Formula 10

TABLE-US-00010 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.5 benzyl alcohol 5-30 7.5

[0078] Formula 11

TABLE-US-00011 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 67.5 benzyl benzoate 5-30 7.5

[0079] Formula 12

TABLE-US-00012 Component Range % Ideal % C1 or SAI 4-30 25.0 butyl digol 50-100 65.0 ethanol 5-80 10

[0080] Formula 13

TABLE-US-00013 Component Range % Ideal % C1 or SAI  4-30 25.0 butyl digol 50-100 65.0 isopropyl alcohol  5-80 10

[0081] Formula 14

TABLE-US-00014 Component Range % Ideal % C1 or SAI  4-30 25.0 g-Hexalactone 50-100 67.5 dimethyisosorbide  5-50  7.5

[0082] Formula 15

TABLE-US-00015 Component Range % Ideal % C1 or SAI  4-30 25.0 propylene carbonate 50-100 67.5 dimethyisosorbide  5-50  7.5

[0083] Formula 16

TABLE-US-00016 Component Range % Ideal % C1 or SAI  4-30 25.0 tetraglycol 50-100 67.5 dimethyisosorbide  5-50  7.5

[0084] Formula 17

TABLE-US-00017 Component Range % Ideal % C1 or SAI  4-30 25.0 triacetin 50-100 67.5 dimethyisosorbide  5-50  7.5

[0085] Formula 18

TABLE-US-00018 Component Range % Ideal % C1 or SAI  4-30 25.0 DEGMEE 50-100 60.0 Capryol 90  5-50  7.5 Labrasol  5-50  7.5

[0086] Formula 19

TABLE-US-00019 Component Range % Ideal % C1 or SAI   4-30 25.0 Butyl digol   50-100 45.0 Dimethylisosorbide   5-50 10.0 Lauroglycol   5-50 20.0 BHT 0.01-0.2  0.02

[0087] Formula 20

TABLE-US-00020 Component Range % Ideal % C1 or SAI   4-30 25.0 Butyl digol   50-100 44.5 Dimethylisosorbide   5-50 10.0 Lauroglycol   5-50 20.0 PVP C15  0.1-5.0  0.5 BHT 0.01-0.2  0.02

[0088] Formula 21

TABLE-US-00021 Component Range % Ideal % C1 or SAI   4-30 25.0 DEGMEE   50-100 45.0 Dimethylisosorbide   5-50 10.0 Labrasol   5-50 20.0 BHT 0.01-0.2  0.02

[0089] Formula 22

TABLE-US-00022 Component Range % Ideal % C1 or SAI   4-30 25.0 DEGMEE   50-100 44.5 Dimethylisosorbide   5-50 10.0 Labrasol   5-50 20.0 PVP C15  0.1-5.0  0.5 BHT 0.01-0.2  0.02

[0090] Formula 23

TABLE-US-00023 Component Range % Ideal % C1 or SAI  4-30 25.0 Butyl Digol 50-100 45.0 Dimethylisosorbide  5-50 10.0 Captex 200  5-50 20.0

Biological

6 Month PKPD Study

[0091] The spiro-azetidine isoxazoline, (S)-1-(5′-(5-(3,5-dichloro-4-fluorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)-3′H-spiro[azetidine-3,1′-isobenzofuran]-1-yl)-2-(methylsulfonyl)ethanone (Compound 1) was used to conduct an in-vivo long-acting efficacy study.

[0092] The study assessed the efficacy of a 25 mg/kg (0.1 mL/kg) topical dose of Compound 1 versus control against fleas (Ctenocephalides felis) and ticks (Ixodes scapularis) on beagle dogs. The formulation was butyl digol:dimethyl isosorbide (90:10 v/v %), selected as it had excellent solubility for Compound 1. Eight male and eight female dogs were acclimated to the testing facility. On Day 0 of the study, each animal received a topical dose of Compound 1 or control (formulation without Compound 1). On Days 56, 89, 117, 145, and 173, animals were artificially infested with about 100 adult unfed fleas. Flea counts were conducted 24 hours post infestation. At each count, all fleas were removed from the dogs. On Days 55, 88, 116, 144, and 172, all dogs were infested with about 50 viable, unfed adult ticks. Tick counts were conducted 48 hours post infestation. At each count, all ticks were removed from the dogs. Geometric mean efficacy results are presented in Table 1, below. Further, some hair was removed from the left or right shoulder from three individual dogs from each treatment group 26 days post dose. The hair from each dog was equally divided and placed into two separate 20 mL scintillation vials. Ten female Rhipicephalus sanguineus and ten female Ixodes scapularis ticks were placed in the vials with the hair. Ticks were evaluated for viability at 2-4 hours, 24 hours, 48 hours and 72 hours post vial infestation. The number of ticks found dead in the vials was reported as the mean value for each group and is presented in Table 2, below.

TABLE-US-00024 TABLE 1 Geometric Mean Live Tick and Flea Counts and percentage Reductions Following Dosing of Compound 1 on Day 0 57 days 90 days 118 days 146 days 174 days Group LT LF LT LF LT LF LT LF LT LF Control 0.8 13.8 10.1 54.9 10.2 63.5 9.3 49.2 12.7 50.5 C1 0.0 0.2 0.0 0.0 0.1 0.9 0.3 0.8 4.5 13 % Reduction 100 98.6 100 100 99.1 98.6 97.3 98.3 64.3 74.3 LT = Live Ticks; LF = Live Fleas; C1 = Compound 1

[0093] Compound 1, administered in a topical formulation comprising butyl digol and dimethyl isosorbide at 25 mg/kg, provided >95% control of ticks (Ixodes scapularis) and fleas (Ctenocephalides felis) for 5 months, as measured by reductions in geometric mean counts compared to placebo-treated controls.

TABLE-US-00025 TABLE 2 Mean Live Tick Counts and Percentage Reductions from Tick Infested Hair Samples # Dead Ticks:Hours Post Infestation Species Group 2-4 24 48 72 I. scapularis Compound 1 0  3.7  7.3  9.3 Control 0  1.0  1.0  1.3 % Reduction 0.0  72.7  86.4  85.7 R. sanguineus Compound 1 0  9.0  9.3  9.7 Control 0  0  0  0 % Reduction 0.0 100.0 100.0 100.0

[0094] The results in Table 2 show that Compound 1 also has contact activity against ticks for at least 26-days following the topical dose.

[0095] The 6-month pk/pd study using the vehicle provided robust efficacy for at least 5-months. In view of the pk/pd data, Compound 1 (250 mg/mL) in the vehicle (BD:ARL, 90:10 v/v %) was used as a control to guide subsequent formulation development.

Formulation Development

[0096] To assess and optimize the in-vitro permeation characteristics of the spiro-azetidine isoxazoline compounds, Franz diffusion cell screening (FDCS) was employed to assess the novel formulations. FDCS yields flux rates and permeation constants for the formulations through a fixed membrane (e.g., canine-, feline-, and bovine-skin). The flux rate is the amount of drug per unit area per unit time that crosses the membrane, and the permeation constant (Kp) is the flux value normalized for the applied concentration of drug product, generally represented in the log 10 form (logKp). A higher flux rate can be correlated to higher in-vivo Cmax and AUC.

[0097] Canine skin, stored at −20° C. for a maximum of 1 year, was thawed, trimmed, and dermatomed to give a thin layer of skin about 0.8-1.5 mm in thickness. The skin was mounted onto the Franz diffusion cell and equilibrated with the receptor media, 50:50 v/v % EtOH:Milli-Q water, for 2 to 4 hours. The receptor media was selected to provide sink conditions for Compound 1. Once equilibrated, test formulations were applied to the donor side, generally 50 μL/cm.sup.2. Samples were generally obtained at 12, 24, 30, 36, 42, and 48 hours, but can be obtained at any six timepoints out to 72 hours. A control vehicle of with Compound 1 (250 mg/mL) in butyl digol:dimethyl isosorbide (90:10 v/v %) was included in every FDCS study.

[0098] Post experiment samples were stored at 4° C. until analysed for Compound 1 concentration by LCMS. Cumulative amount of Compound 1 in ng/cm.sup.2 was calculated and plotted versus time. The gradient of straight line portion of the graph yields the flux value (ng/cm.sup.2/hr) for a given formulation, FIG. 1. The flux can be divided by the applied concentration of Compound 1 to yield the permeability constant Kp (cm/hr)—often converted to a log 10 scale.

Example Study 1: Varying Drug Load

[0099] Compound 1 at varying drug loads (50 mg/mL, 150 mg/mL and 250 mg/mL) in butyl digol:dimethyl isosorbide (90:10 v/v %) was assessed in FDCS to determine Flux and Kp and the results are shown in FIG. 2, below.

[0100] As can be observed in FIG. 2, increased drug load correlates to increased flux. The permeability constant (log kp) is unaffected, as would be expected since the same vehicle was used for each group.

Example Study 2: Dimethyl Isosorbide Titration

[0101] Compound 1 at 250 mg/mL, in varying ratios of butyl digol:dimethyl isosorbide (90:10 v/v %, 80:20 v/v %, and 60:40 v/v %) was assessed in FDCS to determine flux and Kp and the results are shown in FIG. 3, below. The acronym, ARL, refers to Arlasolve, which is dimethyl isosorbide, and API is active pharmaceutical ingredient, and in this instance, Compound 1.

[0102] As can be observed in FIG. 3, increasing the amount of the solvent, dimethyl isosorbide, induced a small reduction in flux rates and permeation constants. This may indicate that the butyl digol is driving penetration rather than the dimethyl isosorbide component of the formulation.

Example Study 3: Oleic Acid Titration

[0103] Compound 1 at 250 mg/mL, in varying ratios of butyl digol:oleic acid (95:5 v/v %, 90:10 v/v %, and 80:20 v/v %) was assessed in FDCS to determine flux and Kp and the results are shown in FIG. 4, below. The acronym, ARL, refers to Arlasolve, which is dimethyl isosorbide and OA is oleic acid.

[0104] As can be observed in FIG. 4, flux rates and permeation constants increase with increasing oleic acid levels.

Results

[0105] Multiple studies were run assessing over 50 novel formulation vehicles. For each study, the Flux of our control vehicle was used to generate an average Flux value (1138.8 ng/cm2/hr). By dividing the average Flux by the experimental Flux generated for each animal skin, a normalization factor for each skin was generated. Data from these studies is presented in Table 3.

TABLE-US-00026 TABLE 3 Control Vehicle Results and Normalization Factors Flux Normalisation Study Skin (ng/cm.sup.2/hr) Factor Dog 1 441.4 2.580 2 2170.9 0.525 3 226.7 5.024 Varying Drug 1 2161.0 0.527 Load 2 2124.1 0.536 3 2373.3 0.480 Arlasolve 1 542.0 2.101 Titration 2 1148.8 0.991 3 321.3 3.545 Fatty Ester 1 298.0 3.822 2 1400.7 0.813 3 816.0 1.396 IPM Titration 1 765.7 1.487 2 1642.9 0.693 3 2911.4 0.391 Oleic Acid 1 172.5 6.601 Titration 2 473.1 2.407 3 1790.3 0.636 THFFA 1 865.6 1.316 Titration 2 469.3 2.427 3 290.9 3.916 Alcohol 1 903.8 1.260 Pyrrolidone 1 2637.8 0.432 Surfactant 1 913.3 1.247 Lipophillic 1 1277.9 0.891 Surfactant Lauroglycol 1 344.5 3.306 IPM-OA 1 1074.3 1.060 Pyrrolidone 2 1 1687.0 0.675 Glycol Ether 1 1965.5 0.579 Polymer 1 226.1 5.037 Assessment Drug Load vs 1 868 1.312 % Lauroglycol

Normalised Flux and Log Kp for Vehicles

[0106] In an effort to maximize duration of efficacy, a high and low flux profile was targeted. A “high flux” vehicle generates a higher flux through the skin than the control vehicle leading to an increased Cmax and AUC, achieving higher percent bioavailability and longer duration of action. A “low flux” vehicle generates a lower flux through the skin than the control vehicle leading to a depoting of Compound 1 in the skin. This reservoir serves to maintain efficacious levels of Compound 1 in the plasma extending duration of action.

[0107] Multiple vehicles with varying degrees of drug load were assessed to differentiate between low and high flux. The experimental flux value obtained from each vehicle was multiplied by the normalization factor obtained and reported in Table 3, so that each test system could be compared to each other. The test vehicles and flux data is shown in Table 4. In Table 4, log Kp is the permeation constant, drug load is mg/mL, and Flux (ng/cm.sup.2/hr) is the normalized average value.

TABLE-US-00027 TABLE 4 Test Formulations and Relative Flux Drug Study Load Vehicle (v/v %) Flux log Kp Control 250 BD:ARL 90:10 1138.80 −4.341 Drug Load 150 BD:ARL 90:10 742.67 −4.527  50 BD:ARL 90:10 276.22 −4.957 Arlasolve 250 BD:ARL 80:20 1196.60 −4.320 Titration 250 BD:ARL 60:40 691.71 −4.558 Fatty Ester 250 BD:IPM 90:10 1645.10 −4.182 250 BD:DES 90:10 2177.15 −4.060 250 BD:TBAC 90:10 1991.14 −4.099 IPM 250 BD:IPM 90:10 1143.18 −4.340 Titration 250 BD:IPM 80:20 1711.06 −4.165 OA Titration 250 BD:IPM 70:30 1938.14 −4.111 250 BD:OA 95:5 1445.07 −4.238 250 BD:OA 90:10 1907.31 −4.118 250 BD:OA 80:20 2822.96 −3.947 THFFA 250 BD:THFFA 90:10 1522.88 −4.215 Titration 250 BD:THFFA 80:20 945.98 −4.422 250 BD:THFFA 70:30 1303.03 −4.283 Alcohol 250 BD:ARL:EtOH 70:10:20 1039.52 −4.381 250 BD:ARL:IPA 70:10:20 939.66 −4.425 250 BD:ARL:BnOH 70:10:20 940.24 −4.425 Pyrrolidone 250 BD:2P 80:20 709.90 −4.547 250 BD:NMP 80:20 1039.47 −4.381 250 BD:OZD 80:20 1454.99 −4.235 Surfactant 250 BD:ARL:Tween80 70:10:20 324.58 −4.887 250 BD:ARL:LAB 70:10:20 350.87 −4.853 250 BD:ARL:Span20 70:10:20 1453.27 −4.236 Lipophillic 250 BD:ARL:Span 20 70:10:20 1141.82 −4.340 Surfactant 250 BD:ARL:Span 80 70:10:20 1475.63 −4.229 250 BD:ARL:CP90 70:10:20 758.09 −4.518 250 BD:ARL:LG90 70:10:20 2723.54 −3.963 250 BD:ARL:EO 70:10:20 1371.41 −4.261 250 BD:ARL:C200 70:10:20 1853.20 −4.130 250 BD:ARL:C355 70:10:20 1622.34 −4.188 Lauroglcol 250 BD:ARL:LG90 85:10:5 4276.95 −3.767 Studyy 250 BD:ARL:LG90 80:10:10 4410.91 −3.753 250 BD:ARL:LG90 70:10:20 7202.60 −3.540 250 BD:ARL:LG90:C200 4735.37 −3.723 70:10:10:10 250 BD:ARL:LG90:C355 5544.73 −3.654 70:10:10:10 250 BD:ARLIG90:IPM 4207.95 −3.774 70:10:10:10 IPM:OA 250 BD:ARL:IPM:OA 60:10:20:10 2173.37 −4.061 Combos 250 BD:ARL:IPM:OA 2125.66 −4.070 60:10:10:20 250 BD:ARL:IPM:OA 80:10:5:5 1663.77 −4.177 250 BD:ARL:IPM:OA 70:10:10:10 2513.56 −3.998 250 BD:ARL:IPM:OA 60:10:15:15 1216.64 −4.313 Pyrollidone 2 250 BD:ARL:2P 85:10:5 1888.47 −4.122 250 BD:ARL:NMP 85:10:5 928.48 −4.430 250 BD:ARL:NOP 85:10:5 2371.62 −4.023 250 BD:ARL:AZ 85:10:5 670.95 −4.571 250 BD:ARL:AZ 80:10:10 2202.41 −4.055 250 BD:ARL:AZ 70:10:20 3448.14 −3.860 Glycol Ether 250 DPG:ARL 90:10 468.51 −4.727 Study 250 TRC:ARL 90:10 207.16 −5.082 250 TRC:ARL:LG90 70:10:20 1286.15 −4.289 250 TRC:ARL:CP90 70:10:20 776.57 −4.508 250 TRC:ARL:Labrasol 70:10:20 103.34 −5.384 250 TRC:ARL:Tween 80 147.63 −5.229 70:10:20 Polymer 250 BD:ARL:LG90 70:10:20 9700.46 −3.411 Assessment 250 TRC:ARL:LAB 70:10:20 506.80 −4.693 250 BD:ARL:LG90 60:20:20 5336.85 −3.671 250 TRC:ARL:LAB 60:20:20 240.43 −5.017 250 BD:ARL:LG90 70:10:20 + 2772.98 −3.955 2% C15 PVP 250 BD:ARL:LG90 70:10:20 + 2049.75 −4.086 2% K29 PVP 250 BD:ARL:LG90 70:10:20 + 3234.00 −3.888 2% K90 PVP Drug Load + 250 BD:ARL:LG90 70:10:20 3384.04 −3.869 % 200 BD:ARL:LG90 70:10:20 3283.15 −3.882 Lauroglycol 150 BD:ARL:LG90 70:10:20 3210.18 −3.891 200 BD:LG90 80:20 2597.95 −3.983 200 BD:ARL:LG90 60:10:30 4332.67 −3.761 150 BD:LG90 80:20 3728.01 −3.826 150 BD:ARL:LG90 60:10:30 3912.73 −3.805

[0108] Overall, as can be observed in Table 4, the fatty acids, fatty acid esters, and mono esters of propylene glycol with fatty acid elicited the greatest Flux enhancement of Compound 1 through canine skin. A transcutol base vehicle and hydrophilic surfactants Tween 80 and Labrasol had the greatest retarding effect on flux rate through canine skin.

[0109] The following formulations in Table 5 were used in a 3-month canine pharmacokinetic study to determine the in-vitro in-vivo correlation (IVIVC).

TABLE-US-00028 TABLE 5 Pharmacokinetic Study Formulations Compound 1 Group Vehicle (v/v %) Flux (ng/cm.sup.2/hr) Comment T01 Butyl Digol:Arlasolve 90:10 1138.8 (n = 31) Control Vehicle T02 Butyl Digol:Arlasolve: 5752.7 (n = 4) Highest Lauroglycol 70:10:20 Fluxing T03 Butyl Digol:Arlasolve: 1853.2 (n = 1) Higher Flux, Captex 200 70:10:20 spreading T04 Transcutol:Arlasolve:  305.1 (n = 2) Lowest Labrasol 70:10:20 Fluxing

[0110] The average flux values in Table 5 were obtained from the individual flux values presented in Table 3. Three month plasma data confirmed good correlation between plasma profiles and the Flux rate values especially at the early timepoints. The plasma Compound 1 profiles for each of the vehicles tested is shown in FIG. 5.

[0111] The plasma profiles can be viewed in 3 distinct sections. During the first week after dosing both the higher fluxing vehicles (T02, T03) had higher plasma levels than the control vehicle (T01), whilst the low fluxing vehicle (T04) had much lower plasma levels. At day 28 all new vehicles (T02, T03, T04) displayed higher plasma levels than the control (T01), with the lowest flux vehicle showing the highest plasma levels indicating that a drug depot may have been formed in the skin. By the three month time point, day 84, both the high (T02) and low (T04) vehicles had about 3× higher plasma levels than the control vehicle, indicating they should perform better in efficacy studies.