AU2003275779B2 - Topical parasiticide formulations and methods of treatment - Google Patents

Description

WO 2004/043445 PCT/AU2003/001490 1 Topical Parasiticide Formulations And Methods Of Treatment Technical Field This invention relates to formulations for administration of benzimidazoles or salicylanilides with macrocyclic lactones to livestock for the control of endoand/or ecto-parasites, methods for dosing livestock with such formulations, and methods for controlling and/or preventing diseases or parasite infection in livestock.

Background Art A number of formulations containing active components, such as therapeutic, prophylactic and/or bioactive substances, for the treatment and/or prophylaxis of diseases or parasite infection in livestock, are known. Such formulations include tablets and solutions for oral administration, injectable solutions, treated collars and ear-tags, and topical means, including pour-on and spot-on formulations.

Many of the early such formulations were intended for topical treatment/prophylaxis of ectoparasite-related conditions, designed to spread the active component over the skin and/or hair surfaces of the animal, not to administer the active component(s) to the bloodstream of the animal being treated. More 'recently, endoparasiticide pour-on formulations for delivery of particular active agents, including macrocyclic lactones, to the bloodstream of domestic animals, such as sheep and cattle, have been developed, and these have the advantage over other administration forms, such as oral drenches and injection, of being easily applied to animals in a relatively-accurate amount.

Known pour-on and spot-on formulations for endoparasiticide treatment generally utilise a non-aqueous delivery system for administering active components to animals, since the active ingredients of interest were substantially water-insoluble (particularly macrocyclic lactones, levamisole base, benzimidazoles), and it was believed that dissolution of the parasiticide was necessary in order for the parasiticide to become systemically absorbed.

Commercial ectoparasiticide products are available as both solvent-based and aqueous-based formulations. Water-insoluble actives have been formulated as aqueous suspension pour-on formulations, deltamethrin (a synthetic pyrethroid) for the treatment of lice on sheep (Clout Schering-Plough) and WO 2004/043445 PCT/AU2003/001490 2 cattle (Coopers® Easy Dose, Schering-Plough), and diflubenzuron (insect growth regulator, or IGR) for lice on sheep (Magnum IGR®, Schering-Plough). These treatments are characterised by low levels of actives found in tissues following treatment, reflecting little penetration of active through the skin layer. Solventbased formulations containing the water-insoluble IGR, triflumuron Zapp®, Bayer) for lice control on sheep are also available. At an equivalent dose rate to the aqueous-based formulations, these solvent-based formulations lead to higher tissue residues immediately after treatment. This supports the assertion that a water-insoluble active will be more easily systemically absorbed if it is solubilized in the formulation.

By 'water-insoluble', it is meant that the water solubility is insufficient for an effective amount of an endoparasiticide to be dissolved in a commercially-viable dose of a water-based pour-on formulation. Practically, a dose of pour-on formulation should not be much more than 1.0mL/10kg bodyweight (for ease of application and to prevent runoff). At this rate, a 500kg beast would receive a dose, therefore, a 2.0mL/10kg dose is not practical, as many animals weigh much more than 500 kg.

Benzimidazoles and macrocyclic lactones are important classes of agents for the treatment or prevention of a number of important endoparasites of livestock, including acute or chronic liver fluke disease, best recognized in sheep and cattle, caused by the liver parasite Fasciola hepatica, and nematodes such as the Cooperia, Ostertagia, and Trichostrongylus species.

Triclabendazole is a particularly effective benzimidazole, and is the most effective drug currently available against all stages of Fasciola hepatica, destroying the early immature and immature fluke migrating through the liver, as well as the adult fluke in the bile duct.

Salicylanilide compounds form another important class of agents for control of endoparasites, particularly Fasciola hepatica, and nematodes, such as Haemonchus species. The salicylanilide oxyclozanide is effective against adult liver fluke (Fasciola hepatica) and immature paramphistones migrating in the intestine of cattle and the young flukes in the rumen and reticulum. Oxyclozanide is highly insoluble in water and is administered to animals in an aqueous suspension formulation by oral dosing.

WO 2004/043445 PCT/AU2003/001490 3 Commercial endectocide pour-on products containing the avermectins, ivermectin (Paramax®, Schering-Plough, Ivomec® Cattle Pour-On, Merial), moxidectin (Cydectin®, Fort Dodge) and doramectin (Dectomax®, Pfizer), are currently available for treatment of cattle for the control or prophylaxis of a number of endo- and ectoparasites, such as lice, flies and ticks. These formulations, however, require significantly higher administration rates of the active component, as compared to oral drenching techniques, typically at least two times the oral drenching rates, in order to achieve effective blood concentrations of the active ingredient in the animal, and to achieve the same efficacy of treatment. For example, ivermectin oral solution for cattle (Ivomec® Oral Solution for Cattle, Merial, registered in New Zealand) has a dose rate of 200 micrograms ivermectin/kg bodyweight, whereas Ivomec® Cattle Pour-On has a dose rate of 500 micrograms ivermectin /kg bodyweight.

Treatment of liver fluke in cattle with anthelmintics, such as triclabendazole, is generally carried out by oral drenching with a commercial product, for example Fasinex® 120 (120 g/L triclabendazole, Novartis), as well as by injection (Ivomec® Plus Antiparasitic Injection for cattle, Merial, which also controls adult liver fluke).

Pour-on or spot-on formulations of salicylanilide derivatives are not currently available, usually being administered to livestock by oral drench.

It would be highly desirable, in order to provide broad-spectrum protection against endoparasites and ectoparasites, through efficient delivery of waterinsoluble compounds, such as benzimidazoles or salicylanilides, in combination with macrocyclic lactones to the bloodstream of animals by a single, convenient topical application, rather than by oral administration.

By "efficient delivery", it is meant that the active agent is administered at a rate approximating oral dosage rates, up to about twice normal oral dosage rates, to give effective blood concentrations and equivalent efficacy.

International Publication No. WO00/61068 (PCT/NZ00/00053) discloses triclabendazole, optionally in combination with a macrocyclic lactone, dissolved in at least one solvent, preferably administered as a pour-on formulation for control of liver fluke. Efficacy data supplied (based on a low natural infection fluke challenge, mean of 20), however, shows that the formulation was applied at times the dose of a standard oral drench rate to give equivalent efficacy. Also, two of the solvents described, xylene and toluene, are highly flammable. The reported triclabendazole content of the formulation, after 345 days storage at ambient temperature, is 7.5% lower than the initial assay, although there is no WO 2004/043445 PCT/AU2003/001490 4 decrease in the abamectin content. Solvent-based formulations of ivermectin can break down rapidly unless suitably stabilized.

A solvent-based, topically-administered formulation of the salicylanilide closantel with the macrocyclic lactone ivermectin, for the control of parasites, has been described in U.S. Patent No. 6,340,672. The maximum concentration of active agents described in the examples of this document is 0.5%w/v for ivermectin and for closantel. At these concentrations, unacceptably large volumes of the formulations (from a practical viewpoint) would need to be poured onto the animals in order to achieve effective blood concentrations of the active agents.

WO 00/74489 (PCT/NZ00/00087) discloses biocidal compositions, including pour-on formulations which are water-in-oil (soyabean) emulsions stabilized with an emulsifying agent. The formulations comprise the water-soluble anthelmintic, levamisole (as the hydrochloride salt), and a macrocyclic lactone (abamectin or ivermectin), optionally in combination with a benzimidazole (oxfendazole). Only low levels of benzimidazole are present in the formulations disclosed in this document (up to 5%w/v oxfendazole in an oral drench formulation), and only one pour-on formulation comprising a benzimidazole (2.26%w/v oxfendazole) and a macrocyclic lactone (0.1%w/v abamectin) is disclosed. Whilst this pour-on formulation is described as delivering the levamisole to the bloodstream of cattle with efficiency similar to oral drench administration, the macrocyclic lactones and benzimidazoles were delivered with low efficiency and a commercially-unpractical volume of this formulation would be required to be applied to animals in order to achieve effective blood concentrations of these actives.

Objects of the Invention It is an object of this invention to provide a topical formulation capable of efficient delivery of a benzimidazole or salicylanilide, in combination with a macrocyclic lactone, to the bloodstream of an animal for broad-spectrum control of endoparasites, such as liver fluke and nematodes, in animals, such as sheep and cattle, with a single, easily-applied topical formulation.

Summary of the Invention It has now been surprisingly found that a benzimidazole or a salicylanilide, in combination with a macrocyclic lactone, may be formulated into a stable aqueous WO 2004/043445 PCT/AU2003/001490 micellar composition which, when applied topically to an animal, efficiently delivers the desired active constituents to the bloodstream of the animal, and provides effective protection against infestation by endoparasites such as liver fluke and nematodes.

Thus, the present invention provides an aqueous micellar formulation comprising a first active agent selected from benzimidazoles, salicylanilides and active derivatives or salts thereof, in combination with a second active agent selected from macrocyclic lactones or active derivatives or salts thereof, said formulation being for topical application to animals for the control of internal parasites and also comprising, per litre of formulation: from about 100g to about 400g veterinary-acceptable surfactant(s); from about 200g to about 750g veterinary-acceptable water-miscible solvent(s); and from about 50g to about 350g water.

Surprisingly, it has also been found that the stability of aqueous micellar formulations of the invention may be improved by inclusion of a stabilizer selected from anionic surfactants, such as sodium dodecyl sulphate (SDS), and/or buffering agents, such as soluble phosphates and/or dibasic phosphates.

Thus, in a preferred aspect of the invention, the aqueous micellar formulation comprises a stabilizer selected from anionic surfactants or buffering agents, or mixtures thereof. Preferably the stabilizer is a linear alkyl sulphate, such as sodium dodecyl sulphate, or one or more phosphates/dibasic phosphates, or mixtures thereof.

In a preferred embodiment, there is provided an aqueous micellar formulation comprising a benzimidazole in combination with a macrocyclic lactone, said formulation being for topical application to animals for the control of internal parasites and also comprising, per litre of formulation: about 100g to about 300g polyoxyalkylene sorbitan fatty acid ester surfactant; about 300g to about 650g alkylene glycol ether selected from alkylene or dialkylene glycol monoalkyl ethers or combinations thereof; about 10g to about 100g polyethylene glycol; about 5g to about 50g stabilizer; and WO 2004/043445 PCT/AU2003/001490 6 about 50g to about 350g water.

In a particularly preferred aspect of this embodiment, the formulation comprises, per litre formulation: about 180g to about 240g benzimidazole; about 7.5g to about 20g macrocyclic lactone or an active derivative or salt thereof; about 150g to about 250g polyoxyethylene (20) sorbitan monolaurate; about 450g to about 550g diethylene glycol monobutyl ether; about 20g to about 50g PEG 200; about 20g sodium dodecyl sulphate; and about 100g to about 200g water.

The invention also provides a method of treating or preventing a diseased or parasite-infested state in a mammal, comprising topically administering to said mammal a micellar formulation according to the instant invention.

Typically, the diseased or infested state is related to liver fluke, such as caused by Fasciola hepatica, and nematodes, such as Cooperia, Ostertagia, Trichostrongylus and Haemonchus species, or combinations thereof.

Even more typically, the diseased or infested state to be treated or prevented is a disease or infested state of cattle or sheep, more typically cattle.

Surprisingly, it was found that the location and size of the region of topical administration of the formulations was important for efficiency of permeation of the active agents across the skin into the bloodstream.

Thus, in a preferred aspect of the methods of treatment, the formulation is applied in a band along the lower portion of the back of the mammal.

Preferably, so as to maximise efficiency of delivery of the active agents to the bloodstream of the animal, the formulation is applied to the animal over as small a region as possible while avoiding run-off of the formulation, so as to maximise the concentration of active agents per cm 2 of animal surface.

In another preferred aspect of the methods of treatment, the formulation is sprayed onto the back of the animal.

WO 2004/043445 PCT/AU2003/001490 7 Where the animals to be treated are cattle, the formulation is preferably applied to the flat part of the back, typically the last third of the animal, and most typically starting from the thoracic vertebrae and proceeding towards the rump of the animal. Typically, about 24mg benzimidazole/salicylanilide and about macrocyclic lactone are applied per kilogram of animal. Typically, the band of formulation applied will be from about 5cm to about 15cm wide and, depending on the size of animal, about 20cm -to 40cm long, and even more typically, the formulation is sprayed onto the back of the animal and the height of the source of spray relative to the back of the animal is maintained at about 5cm to As used herein, the term "treating or preventing", refers to any and all uses which remedy or prevent a diseased or infested state or symptoms, or otherwise prevent, hinder, retard, or reverse the progression of disease/infestation or other undesirable symptoms in any way whatsoever. "Infestation" and corresponding derived terms relate to infestation by endo- and/or ecto-parasites.

An "effective amount", as referred to herein, includes a non-toxic therapeutic or prophylactic amount of an active agent adequate to provide the desired effect.

The "effective amount" will vary from subject-to-subject, depending on one or more of a number of factors amongst, for example, the particular agent being administered, the type and/or severity of a condition being treated, the species being treated, the weight, age and general condition of the subject and the mode of administration. For any given case, an appropriate "effective amount" may be determined by one of ordinary skill in the art using only routine experimentation.

Also, extensive literature is available for many known active agents through, for example, manufacturers' catalogues, the Internet, scientific journals and patent literature, including effective amounts for administration to target animals.

Typically, "effective amount" refers to an amount of active agent sufficient to result in one or more or the following: recession/reduction in the extent of a disease/infestation; inhibition of disease/infestation growth or progression; cessation of disease/infestation growth or progression; prevention of disease/infestation; relief of disease/infestation-imposed discomfort; or prolongation of life of the animal having the disease.

As used herein, the term "about", in the context of concentrations of components of the formulations, typically means 5% of the stated value, more typically 4% of the stated value, more typically 3% of the stated value, more typically, 2% of the stated value, even more typically 1% of the stated value, and even more typically 0.5% of the stated value.

WO 2004/043445 PCT/AU2003/001490 8 As used herein, the term "comprising" means "including principally, but not necessarily solely". Variations of the word "comprising", such as "comprise" and "comprises", have correspondingly similar meanings.

Detailed Description of the Invention Aqueous Micellar formulations The present invention is based on the finding that hydrophobic active agents, such as benzimidazoles and salicylanilides, may be provided in a formulation for topical administration along with therapeutic amounts of a macrocyclic lactone for efficient delivery of both the benzimidazole/salicylanilide and the macrocyclic lactone to the bloodstream of the animal for effective control of endoparasites such as liver fluke and nematodes. It has also been found by the present investigations that efficiency of delivery of the active agents to the bloodstream of a mammal is affected by the topical location of application of the formulation, minimising the area of the skin to which the active agents are applied and/or use of formulations having elevated concentrations of the active agents. The formulations of the present invention surprisingly allow for elevated concentrations of benzimidazole(s) or salicylanilide(s), in combination with one or more macrocyclic lactones, to be provided in a single composition for efficient delivery of the active agents to the bloodstream of a mammal by topical administration.

The formulations are aqueous micellar compositions, comprising elevated levels of the active agents and, per litre of formulation: from about 100g to 400g veterinary-acceptable surfactant(s); from 200g to 750g veterinary-acceptable water-miscible solvent(s); and from 50g to 350g water.

Advantageously, the surfactant is non-ionic and selected from sorbitan esters, polyoxyalkylated sorbitan esters, polyoxyalkylated alkyl ethers, polyoxyalkylated fatty alcohols, polyoxyalkylated fatty acids, polyalkylene glycol esters, polyoxyalkylated derivatives of castor oil, polyglycerol esters, copolymers of ethylene oxide and propylene oxide; amine ethoxylates; alkyl phenol ethoxylates; alkyl polysaccharides; or combinations thereof, although the surfactant may also be, or include, anionic surfactants selected from linear alkylbenzene sulphonates; C12-to-C16 alcohol sulphates; C12 WO 2004/043445 PCT/AU2003/001490 9 alkoxypolyethanoxy sulphates; alkyl phosphates and phosphonates or combinations thereof.

Preferred surfactants are selected from polyoxyalkylated fatty alcohols and polyoxyethylene sorbitan- or sorbitol- fatty acid esters or combinations thereof, and particularly preferred are polyoxyethylene sorbitan- or sorbitol- fatty acid esters.

Generally, the polyoxyalkylene sorbitan- or sorbitol- fatty acid esters are polyoxyethylene sorbitan fatty acid esters. Polyoxyethylene sorbitan fatty acid esters such as those of the Ecoteric® series (Huntsman) are preferred.

Especially preferred polyoxyethylene sorbitan fatty acid ester surfactants are polyoxyethylene (20) sorbitan monolaurate (Ecoteric® T 20) and polyoxyethylene sorbitan monooleate (Ecoteric® T Typically the polyoxylated fatty alcohols are polyalkylene oxide derivatives of natural or synthetic alcohols, and those of synthetic alcohols, such as provided by the Teric® series (Huntsman) are preferred. Especially preferred is Teric® BL8.

Generally, the amount of surfactant used in the formulation ranges from about 100g/L to about 400g/L, typically about 100g/L to about 300g/L, more typically about 150g/L to about 300g/L, even more typically about 150g/L to about 250g surfactant, and even more typically about 175g/L to about 225g/L, preferably about 200g/L, based on the total amount of formulation.

The water-miscible solvent(s) may be selected from: ethanol; isopropanol; benzyl alcohol; glycol ethers; liquid polyoxyethylene glycols; or a mixture of at least two of these solvents.

Particularly-preferred water-miscible solvents are the glycol ethers, and particularly in combination with a liquid polyethylene glycol. A particularlypreferred polyethylene glycol is PEG 200.

Generally, the glycol ethers are alkylene glycol alkyl ethers, including ethylene glycol monoethyl ether, ethylene glycol monomethyl ether, propylene glycol monomethyl ether (Glysolv PM®, Huntsman), dipropylene glycol monomethyl ether, diethylene glycol monoethyl ether (Ethyl di Glysolv®, Huntsman), WO 2004/043445 PCT/AU2003/001490 diethylene glycol monobutyl ether (Butyl di Glysolv® or Butyl Digol®, Huntsman), and diethylene glycol diethyl ether and the like. Particularly preferred glycol ethers are diethylene glycol monoethyl ether (Ethyl di Glysolv®) and/or diethylene glycol monobutyl ether (Butyl di Glysolv® or Butyl Digol®).

Generally, the amount of water-miscible solvent(s) used in the formulation ranges from about 200g/L to about 750g/L, typically about 300g/L to about 650g/L, more typically about 300g/L to about 550g/L and even more typically about 400g/L to about 550g/L, preferably about 450g/L to about 550g/L, based on the total amount of formulation, but will vary depending on the particular solvent(s) used and the amount of active agents to be included in the micellar formulation.

Where, according to a preferred aspect of the invention, the formulation comprises both a glycol ether and a liquid polyethylene glycol, the amount of glycol ether used in the formulation typically ranges from about 350g/L to about 650g/L, more typically about 400g/L to about 600g/L and even more typically about 450gIL to about 550g/L, preferably about 450g/L to about 500g/L, based on the total amount of formulation. The amount of liquid polyethylene glycol used in the formulation typically ranges from about 10g/L to about 100g/L, more typically from about 20g/L to about 70g/L, even more typically from about to about 50g/L, preferably about 30g/L, based on the total amount of formulation.

Generally, the amount of water used in the formulation ranges from about to about 350g/L, typically about 100g/L to about 300g/L, more typically about 100g/L to about 250g/L, and even more typically about 150g/L to about 200g/L, preferably about 150g/L, based on the total amount of formulation.

Examples of suitable benzimidazoles include: 2-(4-thiazolyl)-1H-benzimidazole, known as thiabendazole; [5-(propylthio)-1H-benzimidazol-2-yl]carbamic acid methyl ester, known as albendazole; [5-(propylsulfinyl)-1H-benzimidazol-2yl]carbamic acid methyl ester known as albendazole sulfoxide or albendazole oxide; [2-(4-thiazolyl)-lH-benzimidazol-5-yl]carbamic acid 1-methylethyl ester, known as cambendazole; [5-(phenylthio)-1H-benzimidazol-2-yl]carbamic acid methyl ester, known as fenbendazole; (5-benzoyl-lH-benzimidazol-2-yl)carbamic acid methyl ester, known as mebendazole; [5-(phenylsulfinyl)-1 H-benzimidazol-2yl]carbamic acid methyl ester, known as is oxfendazole; (5-propoxy-1Hbenzimidazol-2-yl)carbamic acid methyl ester, known as oxibendazole; WO 2004/043445 PCT/AU2003/001490 11 butyl)-lH-benzimidazol-2-yl]carbamic acid methyl ester known as parbendazole; methyl 5-cyclopropylcarbonyl-1 H -benzi mid azol-2-ylca rba mate known as cyclobendazole; methyl 5-(4-fluorobenzoyl)-1 H-benzimidazol-2-ylcarbamate known as flubendlazole; 5-chloro-6-(2,3-d ichlorophenoxy)-2-(methylth io)benzimidazole known as tricla bend azole; and [5-(4-fluoro-phenylsu lfonyloxy)-1 Hbenzi mid azol-2-yl]carbamic acid methyl ester known as luxabendlazole.

The benzimidazole antiparasitic agents are active against one or more of Haemonchus, Ostertagia, Trichostrongylus, Nematodirus, Coo peria, Bunostomum, Strongyloides, Trichuris, Qesophagostomum, Chabertia, Dictyocaulus, Moniezia and Fasciola, in sheep and against Haemonchus, Ostertagia, Trichostrongylus, Nematodirus, Coo peria, Bunostomum, Capillaria, Strongyloides, Trichuris, Oesophagostomum, Chaberia, Dictyocaulus, Moniezia and Fasciola in cattle.

Particularly preferred as benzimidazole is triclabendazole.

Examples of suitable salicylanilide compounds for use in the control of Fasciola and Haemonchus species in livestock include oxyclozanide pentachloro-2'-hyd roxysalicylanil ide), closantel (5'-chloro-4'-(4-chloro-alphaid ide), rafoxa n ide (3'-chloro-4'-(4and niclosamide (2',5-dichloro-4'nitrosalicylanilide), as well as clioxanide, brotianidle and bromoxanide.

Salicylanilide derivatives, and their use for control of endoparasites in livestock, has been described in, for example, U.S. Patent numbers 3,914,418; 3,927,071; 3,989,826; 4,005,218; and 4,025,647, "Veterinary Anthelmintics", by J.H.

Arundel, University of Sydney, Post Graduate Foundation in Veterinary Science, and the Merck Veterinary Manual (http://www~merckvetmanual.com/mvm/index.jsp?cfile=htm/bc/1 91415.htm).

Oxyclozanidle is a particularly preferred salicylanilide for use in formulations according to the invention.

Typically, the macrocyclic lactone(s) is/are selected from the group consisting of ivermectin (22,23-dihydroavermectin B 1 described in EP 295117), abamectin, avermectin Aia, avermectin Alb, avermectin A 2 a, avermectin A2b, avermectin B 12 avermectin Bib, avermectin 13 2 1, and avermectin B3 2 b. Also typically, the WO 2004/043445 PCT/AU2003/001490 12 macrocyclic lactone may be selected from active derivatives of the naturally occurring avermectins, such as derivatives which have a group at the substituent other than the isopropyl or (S)-sec-butyl groups, as set out in European patent applications 0214731, 0284176, 0308145, 0317148, 0335541 and 0340832. Also, typically, the macrocyclic lactone of the first aspect of the invention can include moxidectin (and derivatives disclosed in European patent publication No. 259779A), doramectin and its analogues (described in European patent publication No. 0214731B), selamectin, eprinomectin, milbemycin including milbemycin oxime, milbemycin D (Antibiotic B41D) and its analogues (described in U.S. Patent No. 3,950,360) and nemadectins (described in European patent publication No. 170006A).

The macrocyclic lactone antiparasitic agents are active against one or more of Haemonchus, Ostertagia, Trichostrongylus, Nematodirus, Cooperia, Strongyloides, Trichuris, Oesophagostomum, Chabertia and Dictyocaulus in is sheep and against Haemonchus, Ostertagia, Trichostrongylus, Nematodirus, Cooperia, Oesophagostomum and Dictyocaulus in cattle.

Particularly preferred as a macrocyclic lactone is ivermectin.

Generally, where present, the amount of benzimidazole used in the formulation ranges from about 90g/L to about 360g/L, typically about 90g/L to about 300g/L, more typically about 150g/L to about 300g/L, even more typically about 180g/L to about 270g/L, and even more typically about 180g/L to about 240g/L, preferably about 240g/L, based on the total amount of formulation. Generally about 9mg to about 36mg, typically about 9mg to about 30mg, more typically about 15mg to about 30mg, even more typically about 18mg to about 27mg, and even more typically 18mg to about 24mg, preferably about 24mg of benzimidazole per kg bodyweight are applied topically to a mammal in a single dosage.

Generally, where present, the amount of salicylanilide used in the formulation ranges from about 125g/L to about 500g/L, typically about 160g/L to about 375g/L, more typically about 200g/L to about 350g/L, even more typically about 250g/L to about 350g/L, and even more typically about 300g/L to about 330g/L, preferably about 330g/L based on the total amount of formulation. Generally, about 12.5mg to about 50mg of oxyclozanide, typically about 16mg to about 37.5mg, more typically about 20mg to about 35mg, even more typically about WO 2004/043445 PCT/AU2003/001490 13 to about 35mg, and even more typically about 30mg to about preferably about 33mg of salicylanilide per kg bodyweight is applied topically to a mammal in a single dosage.

Generally the amount of macrocyclic lactone used in the formulation ranges from about 2.5g/L to about 25g/L, typically about 4g/L to about 20g/L, more typically about 7.5g/L to about 20g/L and even more typically about 7.5g/L to about preferably about 15g/L, based on the total amount of formulation. Generally about 0.25mg to about 2.5mg, typically about 0.4 to about 2.0mg, more typically about 0.75mg to about 2.0mg, even more typically about 0.75mg to about preferably about 1.5mg of macrocyclic lactone per kg bodyweight are applied topically to a mammal in a single dosage.

Advantageously, the aqueous micellar formulations according to the invention also comprise a stabilizer. Preferably the stabilizer is selected from anionic surfactants such as linear alkyl sulphates (for example, sodium dodecyl sulphate), linear alkyl benzene sulphonates (such as calcium dodecyl benzene sulphonate) and buffering agents, typically selected from soluble monobasic and/or dibasic phosphates.

Sodium dodecyl sulphate is typically used as a stabilizer in the formulation in the range of from about 10g/L to about 30g/L, more typically from about 10g/L to about 20g/L, based on the total amount of formulation; phosphates are typically used in the formulation in the range of from about 1g/L to about 10g/L, more typically from about 1giL to about 5g/L, and more typically from about 1g/L to 2g/L, based on the total amount of formulation.

The aqueous micellar formulations may also include one or more further veterinary excipients, provided these do not destabilise the micellar formulation.

Veterinary acceptable excipients for use in preparing the formulations may include, for example: further solvents such as, for example, water immiscible solvents including glycol ether esters; viscosity modifiers/suspending agents, for example, gelatin, vegetable gums such as xanthan gum, cellulose derivatives microcrystalline cellulose, anionic or non-ionic cellulose ethers, such as carboxymethylcellulose), fumed silica (colloidal silicon dioxide), or polyvinylpyrrolidone polymers, and magnesium aluminium silicates such as VEEGUM® Vanderbilt), and mixtures of these.

WO 2004/043445 PCT/AU2003/001490 14 Examples of suitable veterinary acceptable adjuvants include dyes.

Dyes enable the treated mammals to be distinguished from the untreated. The dyestuff may be dissolved, suspended or dispersed in the carrier. The nature of the colouring agent is unimportant and a wide variety of suitable dyes and pigments will be known to the skilled person. The colouring agent may be soluble or insoluble in water. Generally, however, the dyestuff will be biodegradable so as to fade and not permanently mark the skin or fleece. Some examples of suitable dye agents include: FD&C Brilliant Blue No. 1 (Brilliant Blue FCF, Hexacol Brilliant Blue), and Fast Scarlet Pigment 3610.

Processes for the preparation of micellar formulations of the invention The micellar formulations according to the invention may be prepared by methods and techniques known to those of skill in the art.

Typically the formulations may be made using a simple process: Step 1. Charge 80% of the total volume of water-miscible (non flammable) solvent and the surfactant to a manufacturing vessel. Heat to 400C 750C (flammable solvents such as ethanol and isopropanol, whether added as major water-miscible solvent or as a minor component should be used at ambient temperature).

Step 2. Add the benzimidazole or salicylanilide incrementally with continued stirring and heating until dissolved.

Step 3. Add sequentially the water, and optionally stabilizers and dye, stirring until dissolved.

Step 4. Cool to room temperature with continued stirring.

Step 5. Add the macrocyclic lactone incrementally with stirring until dissolved (also, if flammable solvents such as ethanol or isopropanol are to be added as co-solvents, they should be added here).

Step 6. Add the remaining solvent to volume.

Methods of Treatment andlor prevention of diseases or infestations The formulations according to the invention may be used for the treatment and/or prevention of diseases or infestations by endoparasites in mammals, typically in WO 2004/043445 PCT/AU2003/001490 livestock such as sheep or cattle, by applying the formulation(s) to the back of the mammal. Important diseases/infestations which may be controlled include liver fluke, nematodes and lice in sheep and cattle and buffalo fly and ticks on cattle.

It was found that optimal uptake of the active agents into the bloodstream of treated mammals occurred when the formulations were applied to a region starting from the flat part of an animals back approximately at the location of the thoracic vertebrae and working towards the rump of the animal, effectively resulting in application of the formulation to the last third of the mammal's back.

This mode of application was found to be significantly more effective than application starting at the neck.

Efficiency of delivery of the active agents to the bloodstream of a mammal was also found to be greatest where the surface area to which the formulation is applied was minimised, while avoiding run-off of the formulation, so as to maximise the concentration of active agents per cm 2 of animal surface, typically covering an area of about 100cm 2 to about 400cm 2 for cattle and about 100cm 2 for sheep.

Typically the formulation is applied by spray onto the mammal's back, preferably from a constant height relative to the mammal's back.

For cattle, the band of formulation is typically applied starting from the thoracic vertebrae and proceeding towards the rump of the animal. Typically, from about 18mg to about 24mg benzimidazole and from about 0.75mg to about macrocyclic lactone are applied per kilogram animal. More typically, where triclabendazole and ivermectin are the active agents comprised in the formulation from about 18mg to about 24mg, preferably about 24mg triclabendazole and from about 0.75mg to about 2.0mg, preferably about 1.5mg ivermectin are applied per kilogram of animal. Preferably this amount of active agents is applied to the mammal in about 0.05mL to about 0.1mL per kg animal, and in a band from about 5cm to about 15cm wide. In weaned calves typically weighing from about 100cm to about 180kg per head, good results were obtained by spraying about 10mL to about 18mL formulation onto the backs of the animals, starting from the thoracic vertebrae and working towards the animals' rumps, from a WO 2004/043445 PCT/AU2003/001490 16 constant height of about 15cm relative the backs of the animals, resulting in an applied band of formulation about 10cm to about 15cm wide and about 20cm long.

Preferred forms of the present invention will now be described, by way of example only, with reference to the following examples, including comparative data, and which are not to be taken to be limiting to the scope or spirit of the invention in any way.

Examples Example 1 Aqueous micellar formulations, and processes for preparing them 1.1 Formulation A Component g/L Triclabendazole 240 Ivermectin Polyoxyethylene (20) sorbitan monolaurate (Ecoteric® T 20) 200 Polyethylene glycol 200 (PEG 200) Water 150 Sodium dodecyl sulphate Brilliant Blue FCF 0.16 Diethylene glycol monobutyl ether to 1 L 1.2 Formulation B Component g/L Triclabendazole 240 Ivermectin Polyoxyethylene (20) sorbitan monolaurate (Ecoteric® T 20) 200 Polyethylene glycol 200 (PEG 200) Water 250 Sodium dodecyl sulphate Brilliant Blue FCF 0.16 Diethylene glycol monobutyl ether to 1L WO 2004/043445 PCT/AU2003/001490 17 1.3 Formulation C Component g/L Triclabendazole 120 Ivermectin s Polyalkylene oxide derivative of synthetic alcohol (Teric® BL8) 200 Benzyl alcohol Water 150 Dihydrogen sodium phosphate 7.84 Disodium hydrogen phosphate 0.91 Brilliant Blue FCF 0.16 Diethylene glycol monobutyl ether to 1L 1.4 Formulation D Component g/L Triclabendazole 120 Ivermectin Polyoxyethylene (20) sorbitan monooleate (Ecoteric® T 80) 200 Benzyl alcohol Water 250 Disodium hydrogen phosphate 0.91 Dihydrogen sodium phosphate 7.84 Brilliant Blue FCF 0.16 Propylene glycol monomethyl ether (Glysolv PM®) to 1L Formulation E Component g/L Oxyclozanide 350 Ivermectin Polyoxyethylene (20) sorbitan monooleate (Ecoteric® T 80) 200 Water 150 Sodium dodecyl sulphate Brilliant Blue FCF 0.16 Diethylene glycol monobutyl ether to 1L WO 2004/043445 PCT/AU2003/001490 18 1.6 Formulation F Component g/L Triclabendazole 240 Ivermectin 10.0 Polyoxyethylene (20) sorbitan monolaurate (Ecoteric T 20) 200 Polyethylene glycol 200 (PEG 200) Water 150 Sodium dodecyl sulphate Brilliant Blue FCF 0.16 Diethylene glycol monobutyl ether to 1L 1.7 Formulation G Component g/L Triclabendazole 240 Ivermectin 15.0 Polyoxyethylene (20) sorbitan monolaurate (Ecoteric T 20) 200 Polyethylene glycol 200 (PEG 200) Water 150 Sodium dodecyl sulphate Brilliant Blue FCF 0.16 Diethylene glycol monobutyl ether to 1L Other stable aqueous micellar formulations according to the invention are described in Examples 2 and 3.

The formulations were prepared by the following procedure: Step 1. Charge 80% of the total volume of water-miscible solvent and the surfactant to a manufacturing vessel. Heat to 40 75cC with stirring.

Step 2. Add the benzimidazole or salicylanilide incrementally with continued stirring and heating until dissolved.

Step3. Add sequentially the water, and optionally stabilizers and dye, stirring until dissolved.

WO 2004/043445 PCT/AU2003/001490 19 Step4. Cool to room temperature with continued stirring.

Step 5. Add the macrocyclic lactone incrementally with stirring until dissolved.

Step 6. Add the remaining solvent to volume.

Example 2 Pharmacokinetic studies Materials and methods Formulations according to the invention were tested for their efficacy in delivering benzimidazoles and macrocyclic lactones to the bloodstream of mammals (cattle), and compared to the efficacy in delivering these agents to animals' bloodstreams by standard commercially available drench (Fasinex 120®), and an experimental solvent-based triclabendazole/ ivermectin pour-on formulation.

Cattle (typically Hereford or Hereford cross) with either natural or artificially infected burdens of fluke and nematodes were used in pen and field trials.

Within a given trial animals were allotted into treatment groups, each having similar mean weights and fluke and nematode burdens. Experimental treatments were applied along the backline using a commercially available backliner gun fitted with a plastic shroud to ensure correct delivery of the formulation according to the protocol.

Blood samples (plasma) were taken by venipuncture of the jugular vein at the designated time intervals. Analysis for triclabendazole and ivermectin residues in the plasma was carried out and reported by commercial contract laboratories Ivermectin was extracted from the plasma using acetonitrile and concentrated by evaporation. The sample was cleaned up by solid phase extraction (SPE) chromatography and the ivermectin determined as the N-methyl imidazole derivative using reverse phase HPLC with fluorescence detection.

The triclabendazole was extracted from the plasma using ethyl acetate. Following concentration and SPE clean up, the triclabendazole and its sulphone and sulphoxide metabolites were analysed by reverse phase HPLC using UV detection.

WO 2004/043445 PCT/AU2003/001490 Results Initial feasibility studies for development of an efficient flukicide product were based on the pharmacokinetic profile of triclabendazole only. Although noting that the bioavailability of the active agents is always delayed after application as a pour-on formulation compared to a drench treatment, blood plasma levels for the experimental formulations were targeted at the maximum triclabendazole plasma levels (Cmax) produced by the currently available flukicide, Fasinex® 120 (triclabendazole Cmax 16.5pig/mL after 2 days) when applied at a rate of 12mg/kg bodyweight.

Having reference to Table 1, the following results were obtained.

In a first feasibility trial (Hereford male weaner cattle, average weight of approximately 200kg, 2 animals per group), a solvent-based formulation (Nmethyl pyrrolidone/ Butyl diGlysolv®, Formulation triclabendazole was applied at 50mg/kg to achieve similar plasma levels as per the currently available flukicide, Fasinex® 120 (15.7g/mL after 7 days). Such a dose rate is not commercially viable.

In a second feasibility trial (Hereford male and female weaner cattle, average weight of approximately 160kg, 3 animals per group) the triclabendazole dose rate was reduced to a more commercially acceptable level (12mg/kg). A surfactant (Teric® BL8) was added to Formulation 1 to improve the formulation's hide wettability to produce Formulation 2 (non aqueous micelle), and N-methyl pyrrolidine solvent was removed. Triclabendazole Cmax (total metabolite) plasma levels achieved were low Addition of 15 water to Formulation 1 produced Formulation 3 (Formulation C described in Example 1.3 above, an aqueous micelle), and this increased the triclabendazole Cmax achieved to 4.8|pg/mL.

WO 2004/043445 WO 204/03445PCT/A1J20031001490 21 TABLE 1 Formulation Formulation Details 1g or mL Dose Rate Plasma Cmax Tmax and Type mg/kg 1days 1 Triclabendazole 250g 50 15.7 ig/mL 7 Solvent based N- Methyl 400mL pyrrolidone IButyl di Glysolv® 575m]L Control Fasinex 1 20 g/L TCBZ 12 l6.5gig/mL 2 120 2 Triclabendazole 120g 12 -2.0Otg/mL 7 ___________Iverrnectin 5.0O Non-aqueous TenicO 131- 200g Micelle Benzyl alcohol 30g Butyl di Glysolv® 650mL 3 Triclabendazole 120g 12 4.8liig/rnL 7 Ivermectin Aqueous Teric® BL8 200g micelle Water 150g Benzy]. alcohol Butyl di Glysolv® 520mL 4 Triclabendazole 120g 12 8.7pg/mL 7- Ivermectin 5.Og 0.5 1.3ng/mL Aqueous Teric®D BL8 200g micelle Water 250g Benzyl alcohol Glysolv PM® 420inL Dihydrogen Sodium 7.84g Phosphate Disodium Hydrogen 0.91g phosphate Triclabendazole 12g12 8.7 pg/mL 7 Ivermectin 5.Og 0.5 2.6ng/mL 2 Aqueous Teric® BL8 micelle Water 150g Benzyl alcohol Glysolv PM® 520mL Dihydrogen sodium 7.84g Phosphate Disodium hydrogen 0.

9 1g phosphate

I_

WO 2004/043445 PCTiAU20031001490 22 TABLE 1 (continued) Formulation jFormulation Details Tg or mL 1Dose Rate 1Plasma Tmax1 ID and Type litre mg/kg a das 6 Triclabendazole 120g 12 15.9jig/niL 7 Ivermectin 5 Og 0.5 2.8ng/mL Aqueous Ecoteric® T20 200g micelle Water 250g Benzyl alcohol Glysolv PM8 42OrnL Dihydrogen Sodium 7.84g Phosphate Disodium Hydrogen 0.91 g 7 Triclabendazole 120g 12 12.9ptg/mL 7 ________Ivermectin 5.Og 0.5 3.0ng/ml- 7 Aqueous Ecoteric®l T80 200g micelle Water 250g Benzvl alcohol Glysolv PM® 420ml- Dihydrogen Sodium 7 .8 4 g Phosphate Disodium Hydrogen 0.91g phosphate

I_

In a further feasibility trial (Hereford female cattle, average weight of approximately 235kg, 3 animals per group), the water content in the formulation was increased to 25% and Butyl di Glysolv® was replaced with Glysolv PM®I.

The resulting Formulation 4, provided an increased triclabendazole Cmax of 8.74tg/mL almost double that achieved with Formulation 3. The ivermectin Cmx achieved was 1 .3ng/mL at 5 days.

A similar formulation, Formulation 5, had a water content of 15 Although the Cmx for triclabendazole was almost the same, 8.6j.1g/mL, the C~a for ivermectin was 2.6ng/mL at 2 days.

Replacing Teric® BL18 in Formulation 4 with Ecoteric® T20 resulted in Formulation 6 (with a water content of 25%) this formulation achieved substantially the same plasma levels as Fasinex® 120 drench (triclabendazole Omax Of 15.9gf/mL versus 16.Sgtg/ml-) applied at the equivalent dose rate of 12mg/kg. The COmax achieved for ivermectin was 2.8ng/mL at 5 days.

WO 2004/043445 PCT/AU20031001490 23 Formulation 7 again showed increased bioavailability of triclabendazole when TericV BL8 was replaced with Ecoteric®) T80. The Omax achieved for triclabendlazole was 12.9ltg/mL and the C achieved for ivermectin was 3.Ong/mL at 2 days.

Having reference to Table 2, in a further feasibility trial (Hereford female weaner cattle, average weight of approximately 200kg, 3 animals per group) reduction of the water content of the formulations to 150g/L, and reverting to Ecoteric® T20 in place of Ecoteric® T80, increased the efficiency of delivery of ivermectin, the ivermectin plasma Cmzx values for the formulations ranging from 8ng/mL to l3ng/mL.

Table 2 Fruain g or mL 1Dose Rate [AUG Mean Plasma ax C pnets Iper litre mg/kg [plasma jCMUjA days__ Triclabendazole 90g 9.0 72 1 ig.d/mL 3.6pjg/lL 9jig/ml- Ivermectin 10.Og 1.0 88ng.d/mL 4.4ng/mL 8ngi,'mL EcotericO 120 200g Water 150g Benzyl alcohol Triethanolamine 5 .Og Glysolv PM® 608mL Triclabendazole 120g 12 85gtg.d/ni pgrL 12j tg/mL Ivermectin 5.Og 0.5 52ng.d/mL 2.5ng/l- 8ng/rmL Ecoteric®D T20 200g Water 150g I Benzyl alcohol 30g Triethanolamine 5 .Og GlysolV PM® 588MrL Triclabendazole 180g 18 139g.g.d/mL 6.8ktg/mL 184~g/mL Ivermectin 7.5 g 0,75 79ng.d/mL 4.Ing/ntL lUng/mL Ecoteric® T20 200g Water 150g Benzyl alcohol Triethanolamine 5 Glysolv PM® 553mL From the results provided in Tables 1 and 2, it is apparent that the pharmacokinetics of the active agents can be altered as desired by manipulating the water content, and the type and content of the surfactant and/or the cosolvent used in micellar formulations according to the invention.

WO 2004/043445 PCT/AU2003/001490 24 Manipulation of the solvent and co-solvent type has also been found during the course of these experiments to affect the physical stability of the micellar formulations, use of a combination of Butyl diGlysolv® and PEG 200 providing the best cold storage stability and highest maximum concentration for triclabendazole of the formulations tested, thereby providing a more rugged product suitable for application to animals in the cooler months of late autumn or early spring although there is no published data, it has been reported that greater amounts of active components need to be applied to animals in colder months to get the required efficacy, and these months are typically the most important in liver fluke control.

Example 3 Dosing studies Example 3.1 Concentration effect (constant volume) Having reference to Table 2, it can be seen that altering the concentration of triclabendazole and/or ivermectin in the aqueous micellar formulations of the invention provides a corresponding change in AUC, when applied to the animal in the same volume of formulation (1mL applied/ 10kg animal).

Example 3.2 Concentration effect (constant dose) Having reference to Table 3, in a critical slaughter efficacy trial of formulations according to the invention (methods as per Example 2; mixed sex Hereford weaner cattle, average weight of approximately 200kg, 5 animals per group), an aqueous micellar formulation according to the invention comprising triclabendazole at 240g/L, but varying ivermectin concentration was applied at a constant ivermectin dosage rate (0.5mg/kg), but varying triclabendazole dosage rate (12 to 36mg/kg).

The results show that application of a more concentrated ivermectin dose in a smaller volume (same final ivermectin dose rate), resulted in improved pharmacokinetic results, including greater Cmax and/or greater bioavailability (AUC) of the ivermectin.

WO 2004/043445 PCT/AU20031001490 Table 3 Formulation g or mL Dose Dose AUC Plasma Components per litre Rate Rate Cmnax days mg/kg mLlkg Triclabendazole 240g 12 1lm1/20 73ktg.d/m-L U.3~g/mL Ivermectin log 0.5 lml!20 lO4ng.d/nilL l0.4ng/mL 7 Ecoteric® T20 200g PEG 200 Water 150g Triethanolamine 0.74g Brilliant Blue FCF 0. 16g Butyl di Glysolv®) 491mL Triclabendazole 240 24 lml/1O 129jug.d/ml, 15. 1gg/mL Ivermectin 5 0.5 Imil/lO 84ng.d/rmL 9.5ng/m-L Ecoteric® T20 200 PEG 200 Water 150 Sodium dodecyl sulphate Brilliant Blue FCF 0.16 Butyl di GlysolvO 480 mL Triclabendazole 240g 36 lml/6.6 177pgg.d/mL 18.6pg/mL 7 Ivermectin 3.33g 0.5 1lml/6.6 82ng.d/mL 7.Sng/ml- 7 7 Ecoteric®D T20 200g PEG 200 30g Water 150g Triethanolamine 1. 12g Brilliant Blue FCF 0. 16g Butyl di GlysolvO 498m'L In another trial (also carried out as described in Example a formulation according to the invention having 180 giL triclabendazole and 7.5g/L ivermectin, and a formulation having 240g/L triclabendazole and l0g/L ivermectin, were applied to animals over different area sizes on the backs of the animals (from the middle of the back towards the rump), while maintaining the same dose rate for the active constituents. The results, shown in Table 4, show that application of the ivermectin and triclabendazole in a higher concentration formulation applied over a smaller area makes the active agents more bioavailable.

WO 2004/043445 PCT/AU2003/001490 26 TABLE 4 Formulation g or mL Dose rate Mean Mean AUC details per litre (mg/kg) Treatment plasma Area (cm 2 cone" Triclabendazole 180 12.0 110 3.3tg/mL 654g.d/mL Ivermectin 7.5 0.5 1.7ng/mL Ecoteric T 20® 200 (lmL/ 15 kg) PEG 200 Water 150 Triethanolamine 0.15 Brilliant Blue 0.16

FCF

Butyl diGlysolv® 536mL Triclabendazole 240 12.0 76 5.1 tg/mL 170pg.d/mL Ivermectin 10.0 0.5 2.2ng/mL 43ng.d/mL (l1mL/20kg) Ecoteric T 20® 200 PEG 200 Water 150 Triethanolamine 0.3 Brilliant Blue 0.16

FCF

Butyl diGlysolv® 500mL Example 4 Stability studies Samples of formulation A, the composition and preparation of which is described in Example 1, which contains sodium dodecyl sulphate, were stored at 4, 30 and 400C in 250 mL high density polyethylene bottles sealed with screw caps, sampled at 1, 2, 3, 6 and 12 months, and tested for ivermectin and triclabendazole content. Triclabendazole and ivermectin content of the formulations was determined using validated stability indicating methods based on reversed phase HPLC with UV detection. The results, provided in Table demonstrate the chemical stability of the formulation at accelerated storage conditions effectively no degradation of the active components occurred even after 6 months storage at 400C. After 12 months storage at 300C there was still no measured degradation of the triclabendazole and ivermectin components.

After 12 months at 40°C there was less than 5% breakdown of the ivermectin component.

WO 2004/043445 PCT/AU2003/001490 27 TABLE Storage Triclabendazole Content after Ivermectin Content after Temp. storage time (months): storage time (months): 1 2 3 6 12 1 2 3 6 12 4 0 C 250 248 247 241 247 7.55 7.53 7.84 7.53 7.44 0 C 247 248 247 240 248 7.47 7.52 7.77 7.49 7.43 0 C 247 249 241 242 246 7.45 7.55 7.71 7.41 7.25 Samples of formulation G, the composition and preparation of which is described in Example 1, which contains sodium dodecyl sulphate, were stored at 4, 30 and in 250 mL high density polyethylene bottles sealed with screw caps, sampled at 1, 2 and 3 months, and tested for ivermectin and triclabendazole content. Triclabendazole and ivermectin content of the formulations was determined using validated stability indicating methods based on reversed phase HPLC with UV detection. The results, provided in Table 6, demonstrate the chemical stability of the formulation at accelerated storage conditions effectively no degradation of the active components occurred even after 2 months storage at 30 or 400C.

TABLE 6 Storage Triclabendazole Content after Ivermectin Content after Temp. storage time (months): storage time (months): 1 2 3 1 2 3 4 0 C 243 241 238 14.7 14.8 14.7 0 C 241 239 236 14.5 14.5 14.6 0 C 237 239 237 14.5 14.5 14.5 In another stability trial a number of substances were tested for their potential as a stabilizer for the formulations, ivermectin being unstable in inadequately stabilised formulations. The substances were each tested at a concentration of 10.0 g/L, except phosphate buffers, in a formulation otherwise having the following composition (per Litre): Triclabendazole Ivermectin Teric BL 8® 120g 200g WO 2004/043445 PCT/AU2003/001490 Benzyl alcohol Water Brilliant Blue FCF Butyl Di Glysolv® 150g 0.16g approximately 485 mL (to volume) The samples were stored at 50 0 C in 250 mL high density polyethylene bottles sealed with screw caps, and sampled at 3 months, and tested for ivermectin and triclabendazole content. Triclabendazole and ivermectin content of the formulations was determined using validated stability indicating methods based on reversed phase HPLC with UV detection. The data, provided in Table 7, illustrate the difficulty of stabilising the ivermectin component of the formulation.

From the stability data it was concluded that inclusion of anionic surfactants such as the linear alkyl sulphate sodium dodecyl sulphate, or buffering agents such as one or more monobasic/ dibasic phosphates, or mixtures thereof, in the formulations of the invention significantly improve the stability of the ivermectin component.

Table 7 Triclabendazole Ivermectin Content after Content storage time: after storage time: Candidate Stabilizer g/L Initial 3 months Initial 3 months Ivermectin 50C 50 0 C Breakdown S 124.1 122.0 4.96 4.33 12.7 Butylated hydroxy 10.0 123.8 122.5 4.92 4.36 11.4 toluene (BHT) Epoxidised Resin 10.0 123.2 123.2 4.89 3.45 29.4 (ERL 4221) Vitamin E Acetate 10.0 123.1 122.2 4.87 4.36 10.5 Triethanolamine 10.0 121.7 122.4 4.70 1.88 60.0 Disodium hydrogen 0.18 110.0 109.5 4.39 4.24 3.4 phosphate Dihydrogen sodium 7 phosphate_ WO 2004/043445 PCT/AU2003/001490 29 Example 5 Efficacy studies Materials and Methods Cattle (typically Hereford or Hereford cross breed) with either natural or artificially infected burdens of fluke and nematodes were used in pen and field trials. They s were allotted into treatment groups, each having similar mean weights and fluke and nematode burdens. Experimental treatments were applied along the backline from the middle of the back towards the rump, using a commercially available backliner gun fitted with a plastic shroud to ensure correct delivery of the formulation according to the protocol.

ic Efficacy was measured by either decrease in faecal egg counts over time or total parasite counts from gastrointestinal tracts and livers recovered after slaughter.

The reported data are based on group arithmetic and/or group geometric means.

Efficacy based on faecal worm egg counts were calculated as follows: Efficacy 100 1 (T 2 C1/ T 1 C2)] where T, C, 1 and 2 refer to treated, control, pre-treatment and post treatment mean worm egg counts respectively.

All other Efficacy data were calculated using the formula: %Efficacy 100(C-T/C) where T and C refer to treated and control mean total worm counts respectively.

For critical slaughter nematode efficacy studies, the animals were slaughtered at 14 or 21 days post treatment.

For critical slaughter efficacy studies against all stages of the liver fluke (artificially infested), the animals were slaughtered 100 days after treatment.

Results Example 5.1 A critical slaughter pen efficacy trial (naturally acquired fluke and nematodes) involved mixed sex Hereford and Hereford/Angus cross weaned calves selected from 2 large commercial herds. The animals were randomly allocated to groups of 5 animals such that each group had a similar mean and range of Fasciola hepatica egg counts and body weights. Prior to treatment, animals were moved WO 2004/043445 PCT/AU2003/001490 to a research feedlot to avoid further infection. At treatment the animals were weighed and treated with formulations of the triclabendazole ivermectin pour on administered at different dose volumes and active concentrations. One group of animals remained as untreated negative control.

All animals were slaughtered 19 to 21 days post treatment, gastrointestinal tracts and livers recovered, and total worm and fluke numbers determined.

Treatment formulations involving different concentrations of active components and/or different excipients were tested, these formulations being as follows: Group 1 Triclabendazole Ivermectin Ecoteric T20® PEG 200 Water Triethanolamine Brilliant Blue FCF Butyl diGlysolv® Group 2 Triclabendazole Ivermectin Ecoteric T20® PEG 200 Water Triethanolamine Brilliant Blue FCF Butyl diGlysolv® Group 3 Triclabendazole Ivermectin Ecoteric T20® PEG 200 Water Triethanolamine Brilliant Blue FCF Butyl diGlysolv® q or mL/L 240g 10.0g 200g 150g 0.74g 0.16g 491mL q or mL/L 240g 5.0g 200g 150g 1.27g 0.16g 494mL 240g 3.33g 200g 150g 1.12g 0.16g 498mL Dosage rate (mg/kg) 12 Dosage rate (mg/kq) 24 36 Group 4 Triclabendazole Ivermectin Ecoteric T20® PEG 200 Water Brilliant Blue FCF Sodium dodecyl sulphate Butyl diGlysolv® 240g 180g 150g 0.16g 480mL WO 2004/043445 PCT/AU2003/001490 31 Group 5 q or mLIL Dosage rate (mglkg) Triclabendazole 240g 24 Ivermectin 5.0g Ecoteric T20® 200g PEG 200 Water 150g Brilliant Blue FCF 0.16g Sodium dodecyl sulphate Butyl diGlysolv® 480mL Group 6 Triclabendazole 240g 24 Ivermectin 5.0g Ecoteric T20® 200g PEG 200 Water 150g Brilliant Blue FCF 0.16g Sodium dodecyl sulphate Butyl diGlysolv® 316mL Ethylene glycoldiacetate 155mL The results, provided in Table 8, show that effective control of flukes and nematodes is achievable using a practical volume of an aqueous micellar pouron formulation of the present invention.

The product was 100 effective against adult Fasciola hepatica at dose rates of 12, 24 and 36 mg/kg triclabendazole and effective against nematodes at a dose rate of 0.5 mg/kg ivermectin. In this trial, an effective treatment of animals for endoparasites was achieved using 1mL/ 20kg of a formulation including 240g/L triclabendazole and 10.0g/L ivermectin (12mg/kg triclabendazole and 0.5 mg/kg ivermectin).

WO 2004/043445 PCT/AU2003/001490 32 TABLE 8 Treatment efficacy against parasites (values based on the geometric mean of total worm count are given in brackets where different to those based on the arithmetic mean) Liver Abomasum Group No. F. hepatica (adult) H.contortus (adult) Ostertagia spp (adult) T axei (adult) 1 100 >99.9 >99.9 >99.9 2 100 >99.9 98.2 (96.4) >99.9 3 100 >99.9 95.8 (86.6) >99.9 4 100 >99.9 89.1 (81.8) >99.9 100 >99.9 >99.9 >99.9 6 100 >99.9 69.2 (91.9) >99.9 Small intestine Trichostrongylus Cooperia spp Cooperia spp Cooperia spp Nematodirus Group No. spp (adult) (adult) (immature) L4 spp (adult) 1 94.4 88.5 (96.7) >99.9 92.3 (85.9) negative 2 54.9 (negative) 56.1 (66.4) >99.9 >99.9 negative 3 85.9 (84.9) 91.4(88.3) >99.9 >99.9 50 (18.5) 4 57.7 (93.8) 80.2 (84.3) >99.9 >99.9 25 (8) 92.5 (96.1) 89.8 (98.7) >99.9 >99.9 >99.9 6 91.5 (88.3) 36.3 (83.6) >99.9 53.8 (75.8) >99.9 Large intestine Group No. Oesophagostonman (adult) Trichuris (adult) 1 >99.9 99.9 (>99.9) 2 >99.9 14.3 (negative) 3 >99.9 99.9 (>99.9) 4 >99.9 99.9 (>99.9) >99.9 85.7 (71.2) 6 >99.9 85.7 (71.2) Example 5.2 Two critical slaughter studies were designed to compare the efficacy of a formulation according to the invention (see below) against immature and adult stages of the liver fluke Fasciola hepatica, and naturally acquired roundworm infections in cattle. The efficacy of the triclabendazole ivermectin pour-on against immature and mature stages of Fasciola hepatica based on arithmetic WO 2004/043445 PCT/AU2003/001490 33 mean was 70.5% and 99.2% respectively. Control of gastrointestinal strongyles by the test formulation (Group 5, Example 5.1, Table 8) as assessed using total worm counts at slaughter was 86% to 99.9% (arithmetic mean) for nematodes found in the abomasum, small and large intestines.

Test formulation described in Example 1.1, Formulation A Component g or mL/L Dose Rate (mg/kg) Triclabendazole 240g 24.0 Ivermectin 7.5 g 0.75 Ecoteric T20® 200g PEG 200 Water 150g Brilliant Blue FCF 0.16g Sodium dodecyl sulphate Butyl diGlysolv® approximately 475 mL (to volume) Example 5.3 Three field trials (faecal egg count reduction tests) were designed to determine the efficacy of the formulation described in Example 5.2 under field conditions.

Sixty cattle were split into groups of 15, one of the groups remaining as an untreated control. Good efficacy of the formulation against Fasciola hepatica as assessed by a reduction in faecal egg counts as compared to the untreated controls of >90% (AM) was reported in all trials 14 days post treatment.

Example 5.4 A field trial was designed to determine the efficacy of the following formulation against a mixed natural infection of adult and immature liver flukes and adult and immature nematode species.

Component g IL Dose Rate (mglkg) Triclabendazole 240g 24.0 Ivermectin 7.5 g 0.75 Ecoteric T20® 200g PEG 200 WO 2004/043445 PCT/AU2003/001490 34 Water 150g Brilliant Blue FCF 0.16g Sodium dodecyl sulphate Butyl diGlysolv® approximately 450 mL (to volume) Thirty (30) Angus cross and Limousin cross weaners, between 5 and 6 months of age, and weighing 112-242 kg, were selected from a larger commercial herd running at Armidale, New South Wales, Australia, on the basis of pre trial individual strongyle egg counts. The cattle grazed in open paddocks on a mixture of native and improved pasture with supplementary feed (buckwheat) provided on a daily basis. Over the treatment period at the Armidale Saleyards cattle had ad-lib access to Lucerne hay. The cattle had not been exposed to any anthelmintic treatments for a period of three months prior to the trial start date.

Prior to treatment cattle were ranked from highest to lowest on individual pre trial liver strongyle faecal egg counts (Day split into females and castrated males, blocked and randomly allocated to two treatment groups such that the groups had a similar mean and range of strongyle faecal egg counts within the group.

On day zero all trial cattle were weighed and vaccinated with UltraVac 7 in 1 Vaccine (CSL Limited). The animals of Group 1 were left untreated, serving as negative controls. Group 2 was treated with the triclabendazole (240g/L) ivermectin (7.5g/L) pour on formulation applied topically from the middle of the back to the base of the tail at a dose volume of 1mL/lOkg. A prototype applicator which ensured the formulation was applied as a wide band was used for treatment.

Faecal samples were collected from all trial cattle on day zero and on days seven fourteen (14) twenty one (21) and twenty eight (28) of the trial.

Strongyle and liver fluke faecal egg counts and group bulk coprocultures for larval differentiation were performed on samples collected. Raw strongyle and fluke faecal egg counts were collated by treatment group and arithmetic means calculated. Geometric means were also calculated using transformed individual egg counts. Treatment efficacy, based on both arithmetic and geometric group means were calculated as follows: Efficacy (control group mean treatment group mean)/control group mean x 100 WO 2004/043445 PCT/AU2003/001490 Pre treatment Fasciola and strongyle faecal egg counts were high, with a mean Strongyle faecal egg count of 802.7 e.p.g. (range 160-6120) and a mean Fasciola faecal egg count of 46 e.p.g. (range 0-1525) pre trial. Five genera of helminths were identified from group bulk coprocultures including: Haemonchus spp., Trichostrongylus spp., Ostertagia spp., Cooperia spp and Oesophagostomum spp.. Cooperia spp made up on average 70% of the bulk coproculture for the untreated controls from day 0 to day 28. Group arithmetic and geometric mean Fasciola faecal egg counts over the duration of the trial are presented in Table 9. Good control efficacy arithmetic mean, >97% efficacy geometric mean) of Fasciola hepatica was achieved with the triclabendazole ivermectin pour on, 7, 14, 21 and 28 days post treatment.

Treatment efficacies based on arithmetic and geometric mean fluke faecal egg counts are presented in Table TABLE 9 Fasciola faecal egg counts (e.p.g eggs per gam; Arithmetic mean AM; Geometric mean GM) Da 0 Day 7 Day 14 Day 21 Day 28 Group No. AM GM AM GM AM GM AM GM AM GM 1 (control) 58.4 44.3 86.7 44.6 86.3 55 49.1 28.6 64.2 23.8 2 159 47.2 1.4 0.4 6.8 1.3 2.6 0.3 3.3 0.6 TABLE Treatment Efficacy using arithmetic mean (AM) and geometric mean (GM) fluke faecal egg counts (percent reduction from untreated controls) Day 7 Day 14 Day 21 Day 28 AM GM AM GM AM GM AM GM 98.4 99.2 92.2 97.7 94.6 99.0 94.8 97.5 Group arithmetic and geometric mean strongyle faecal egg counts over the duration of the trial are presented in Table 11. Efficacy of the triclabendazole ivermectin pour on against strongyles was greater than 93% (geometric means) 7 and 28 days post treatment, and 89.8% and 83.5% 14 and 21 days post treatment. Efficacy based on arithmetic and geometric faecal egg counts are presented in Table 12.

WO 2004/043445 PCT/AU2003/001490 36 TABLE 11 Strongyle faecal egg counts (e.p.g eggs per gram; Arithmetic mean AM; Geometric mean GM) Day 0 Day 7 Day 14 Day 21 Day 28 Group No. AM GM AM GM AM GM AM GM AM GM 1 (control) 501 601 333 137.3 163 93.6 136 64.9 173 138.4 2 747 891 112 4.5 90.0 9.6 54.3 10.7 60.0 0.6 TABLE 12 Treatment Efficacy using arithmetic mean (AM) and geometric mean (GM) strongyle faecal egg counts (percent reduction from untreated controls) Day 7 Day 14 Day 21 Day 28 AM GM AM GM AM GM AM GM 66.4 96.7 44.7 89.8 60.1 83.5 65.4 93.4 Example A further field trial was designed to determine the efficacy of the formulation described in Example 5.4 against a mixed natural infection of adult and immature liver flukes and adult and immature nematode species.

Thirty (30) Angus and Angus cross heifers, between 12 and 14 months of age, and weighing 126-284 kg, were selected from a larger commercial herd running at Walcha, New South Wales, Australia, on the basis of pre trial individual strongyle egg counts. The cattle grazed in open paddocks on a mixture of native and improved pasture with ad-lib access to water. The cattle had not been exposed to any anthelmintic treatments for a period of three months prior to the trial start date.

Prior to treatment cattle were ranked from highest to lowest on individual pre trial liver strongyle faecal egg counts (Day blocked and randomly allocated to two treatment groups such that the groups had a similar mean and range of strongyle faecal egg counts within the group. On day zero all trial cattle were weighed. The animals of Group 1 were left untreated, serving as negative controls. Group 2 was treated with the triclabendazole (240g/L) ivermectin pour on formulation applied topically from the middle of the back to the WO 2004/043445 PCT/AU2003/001490 37 base of the tail at a dose volume of lmL/10kg. A prototype applicator which ensured the formulation was applied as a wide band was used for treatment.

Faecal samples were collected from all trial cattle on day zero and on days seven fourteen (14) twenty one (21) and twenty nine (29) of the trial.

Strongyle faecal egg counts and group bulk coprocultures for larval differentiation were performed on samples collected. Raw strongyle egg counts were collated by treatment group and arithmetic means calculated. Geometric means were also calculated using transformed individual egg counts. Treatment efficacy, based on both arithmetic and geometric group means were calculated as follows: Efficacy (control group mean treatment group mean)/control group mean x 100 Pre treatment strongyle faecal egg counts were high, with a mean Strongyle faecal egg count of 288 e.p.g. (range 40-1320). Four genera of helminths were identified from group bulk coprocultures at day zero including: Haemonchus spp., Ostertagia spp., Cooperia spp and Oesophagostomum spp.. Cooperia spp made up on average 70-80% of the bulk coproculture for the untreated controls from day 0 to day 29. Group arithmetic and geometric mean strongyle faecal egg counts over the duration of the trial are presented in Table 13. Efficacy of the triclabendazole ivermectin pour on against strongyles reached a maximum 84% reduction in egg counts (arithmetic means) 7 days post treatment, and 78%, 59% and 63% 14, 21 and 29 days post treatment. Treatment efficacies based on arithmetic and geometric strongyle egg counts are presented in Table 14.

TABLE 13 Strongyle faecal egg counts (e.p.g eggs per gam; Arithmetic mean AM; Geometric mean GM) Day 0 Day 7 Day 14 Day 21 Day 28 Group No. AM GM AM GM AM GM AM GM AM GM 1 (control) 344 262 203 95 267 175 216 129 208 116 2 379 273 32 2 59 22 88 21 77 26 WO 2004/043445 PCT/AU2003/001490 38 TABLE 14 Treatment Efficacy using arithmetic mean (AM) and geometric mean (GM) strongyle faecal egg counts (percent reduction from untreated controls) Day 7 Day 14 Day 21 Day 28 AM GM AM GM AM GM AM GM 84.2 98.2 78 87.5 59.3 83.6 62.8 77.4 Example 5.6 A dose evaluation critical slaughter study was designed to compare the pharmacokinetics and efficacy of the developmental topical triclabendazole ivermectin formulation described in Example 5.4 (240g/L triclabendazole and g/L ivermectin), and the developmental topical triclabendazole ivermectin formulations of formulae F (240 g/L triclabendazole and 10 g/L ivermectin) and G (240 g/L triclabendazole and 15 g/L ivermectin) described in Examples 1.6 and 1.7 respectively against a mixed natural infection of gastrointestinal strongyles, so as to determine the optimum concentration of ivermectin in the formulation for effective control of Cooperia spp as well as the other nematodes.

Fifty (50) Hereford and Angus cross steers, aged between five to six (5-6) months and weighing between 102-164kg at treatment, were selected from a larger mob at Casino on the North Coast of NSW, Australia on the basis of pre trial individual strongyle faecal egg counts. The cattle were relocated to "Kirby", Armidale NSW, Australia twenty days prior to treatment and grazed in open paddocks on a mixture of native and improved pastures. Trial cattle were fed Lucerne hay while they were held in the Armidale Saleyards (day 0 through to day The cattle had not been exposed to triclabendazole or ivermectin for a period of three months prior to the trial start date and had no known resistance by gastrointestinal strongyles to macrocyclic lactones.

Five days prior to treatment faecal samples were collected from each animal for individual faecal egg counts and bulk coproculture. Triplicate blood samples were collected for triclabendazole and ivermectin plasma analysis. One day prior to treatment Twenty five (25) trial cattle were re-located to the Armidale Saleyards, ranked from highest to lowest according to individual egg counts (day sequentially blocked and allocated at random to five groups of five animals, such that each group had a similar mean and range of strongyle faecal WO 2004/043445 PCT/AU2003/001490 39 egg counts. The animals of Group 1 were left untreated, serving as negative controls. Group 2 was treated with the 240g/L triclabendazole ivermectin pour on formulation. Group 3 was treated with the 240g/L triclabendazole 10.0g/L ivermectin pour on formulation. Group 4 was treated with the 240g/L triclabendazole 15.0g/L ivermectin pour on formulation. All formulations were applied topically from the middle of the back to the base of the tail at a dose volume of 1 mL/10kg (according to a dose break table). A prototype applicator which ensured the formulation was applied as a wide band was used for treatment. Two day after treatment all cattle were re-located from the Armidale Saleyards to the Kirby feedlot for the remainder of the trial.

Faecal samples were collected from each individual animal in all groups five days prior to treatment then nine for individual faecal egg counts and coprocultures pre and post treatment. All trial cattle were sacrificed 13, 14 and days post treatment. Faecal samples, abomassa, small intestine and large intestine were collected from each animal for faecal egg counts, group coprocultures and total worm counts (adult and immature). Treatment efficacy was assessed by comparison of group arithmetic and geometric mean total worm counts (as described in Examples 5.4 and 5.5) by nematode species and strongyle faecal egg counts following sacrifice and organ recovery.

Pre treatment egg counts were generally high ranging from 480-1480 eggs per gram of faeces.

At 13 15 days post treatment, animals treated with the pour-on formulations produced a reduction in egg counts when compared to the untreated controls of between 73% (240 g/L triclabendazole plus 7.5 g/L ivermectin) to 98% (240 g/L triclabendazole plus 15.0 g/L ivermectin) (arithmetic means) and between 94 and 99 respectively (geometric means). (Table WO 2004/043445 PCT/AU2003/001490 Table Treatment efficacies at Days 9 and 13, 14, 15, as assessed using arithmetic and geometric group mean faecal egg counts.

Group Treatment EPG Day 9 EPG Days 13-15 Arithmetic data 2 IVM 7.5mg/mL TCBZ 240mg/mL 82.8% 72.8% 3 IVM 10mg/mL TCBZ 240mg/mL 95.4% 89.1% 4 IVM 15mg/mL TCBZ 240mg/mL 97.7% 97.5% Geometric data 2 IVM 7.5mg/mL TCBZ 240mg/mL 96.7% 93.9% 3 IVM 10mg/mL TCBZ 240mg/mL 99.3% 98.3% 4 IVM 15mg/mL TCBZ 240mg/mL 99.5% 99.7% IVM ivermectin, TCBZ triclabendazole, epg eggs per gram At necropsy, seven genera of helminths were recovered from the gastrointestinal tract of the control cattle approximately 80% of which consisted of adult, immature and L4 stages of Cooperia spp. Other gastrointestinal nematodes identified include Trichuris spp, Nematodirus spp, Oesophagosomum spp. Trichostrongylus spp, Haemonchus spp and Ostertagia spp which each made up approximately 5% or less of the total count.

Total worm count data indicated that the small intestinal worms, Cooperia spp.

and adult Nematodirus spp., were the most difficult species to remove following treatment. Efficacy increased with increasing concentration of ivermectin in the formulation.

The 240 g/L triclabendazole plus 15.0 g/L ivermectin formulation efficacy against adult and immature stages of small intestinal nematodes (Trichostrongylus spp, Cooperia spp) was greater than 90% (arithmetic and geometric means) and greater than 99% (geometric means) with the exception of adult Nematodirus [49.1% (arithmetic means) and 93 (geometric means)].

Greater than 95% efficacy (geometric and arithmetic) was achieved against adult and immature stages of abomasal nematodes (Haemonchus spp, Ostertagia ostertagia, Trichostrongylus axei) and large intestinal nematodes (Oesphagostomum spp, Trichuris spp).

Greater than 95% efficacy (arithmetic and geometric means) was achieved by the 240g/L triclabendazole plus 7.5g/L ivermectin and the 240g/L triclabendazole WO 2004/043445 PCTIAU20031001490 41 plus l0g/L ivermectin formulations against abomasal nematodes (with the exception of fourth stage Ostertagia larvae in cattle treated with the 240gIL triclabendazole plus l0gIL ivermectin formulation Efficacy against small intestinal nematodes increased from 57.7% to greater than 99.9% with increased concentration of ivermectin.

Table 16 Arithmetic/Geometric mean andi percentage removal of the number of helminths recovered at necropsy from l5gIL ivermectin 240 gIL triclabendazole treated Ivermectin +I 240g/L Triclabendazole Helminth species Removal I Removal (GM -Adult I mmature IL4 AdutI immature L4 Abomasal Haemonchus spp. 96 >99.9 99.1 >99.9 Ostertagia spp. >99.9 >99.9 >99.9 >99.9 >99,9 >99.9 Trichostrongylus axei >99.9 >99.9 Small Intestine Trichostrongylus spp. >999 >99.9 Cooperia spp. 90.7 94.6 >99,9 99.5 99.5 >99.9 Nematodirus spp. 44.9 93.3 Large Intestine eshaosommspp. >99.9 >99.9 >99.9 >99.9 Trichuris sp >=99.9 Triplicate blood samples were also collected five days prior to treatment then 1, 3, 5 and 7 days post treatment from animals in groups 2, 3, 4 and 5 for triclabendazole and ivermectin analysis. Plasma ivermectin Cmax and AUG values increased relative to the concentration in the formulation Table 17.

WO 2004/043445 PCT/AU2003/001490 42 Table 17 Summary (mean SD) disposition of ivermectin by treatment group Treatment group (ng/mL) Tm, (day) AUC (ng.d/mL) Grp 2: IVM 7.5mg/mL 3.75 2.22 3.4 1.7 13.39 5.88 TCBZ 240mg/mL Grp3: IVM 10mg/mL+ 9.00 7.74 3.8 1.1 26.65 ±22.56 TCBZ 240mg/mL Grp4: IVM 15mg/mL+ 6.95 2.87 3.8 ±1.1 31.87 ±17.13 TCBZ 240mg/mL

I

I V IVI ivermectl.;iIII, I Lrncaoenaazoie Summary: For a given dose volume (1mL per 10 kg bodyweight), increasing the concentration of ivermectin in the formulation increased the plasma concentration and efficacy. Nematode efficacy of the 240 g/L triclabendazole plus 15.0 g/L ivermectin was higher and more consistent than the corresponding formulations containing 7.5 and 10.0 g/L ivermectin, especially against the hard to control small intestinal worms, Cooperia spp and Nematodirus spp.

Industrial Applicability The formulations of the invention can be readily used to treat, control or prevent disease caused by, and/or infestations of, endo-parasites such as liver fluke and nematodes as well as ecto-parasites, particularly in treating, controlling and/or preventing liver fluke and nematode infestations in sheep or cattle, particularly cattle.

It will be appreciated that, although specific embodiments of the invention have been described herein for the purpose of illustration, various modifications may be made without deviating from the spirit and scope of the invention as defined in the following claims.

Claims (14)

1. 2. A formulation according to Claim 1, wherein said surfactant is selected from polyoxyethylene sorbitan fatty acid esters or combinations thereof.
2. 3. A formulation according to Claim 2, wherein said surfactant is polyoxyethylene (20) sorbitan monolaurate.
3. 4. A formulation according to any one of Claims 1 to 3, wherein said water-miscible solvent is selected from ethanol, isopropanol, benzyl alcohol, glycol ethers, liquid polyoxyethylene glycols, or a mixture of at least two of these solvents.
4. 5. A formulation according to Claim 4, comprising a glycol ether and a liquid polyethylene glycol as water-miscible solvents.
5. 6. A formulation according to Claim 5, wherein said one or more of glycol ethers are selected from propylene glycol monomethyl ether, diethylene glycol monoethyl ether, and diethylene glycol monobutyl ether and the polyethylene glycol is PEG 200.
6. 7. A formulation according to any one of Claims 1 to 6, comprising about 450g to about 550g glycol ether(s) selected from alkylene or dialkylene glycol monoalkyl ethers, and about 20g to about 50g of a liquid polyethylene glycol as the one or more water-miscible solvents per litre of formulation.
7. 8. A formulation according to any one of Claims 1 to 7, further comprising from about 5g to about 50g per litre of formulation of a stabilizer selected from linear anionic surfactants, buffering agents and mixtures thereof.
8. 9. A formulation according to Claim 8, wherein said stabilizer is sodium dodecyl sulphate. 00 10. A formulation according to any one of Claims 1 to 9, comprising from Sabout 120g to about 300g benzimidazole, or a derivative thereof, per litre of o formulation. Q 11. A formulation according to any one of Claims 1 to 10, wherein said first n 5 active agent is triclabendazole.
9. 12. A formulation according to any one of Claims 1 to 11, comprising from about 7.5g to about 20g macrocyclic lactone per litre of formulation.
10. 13. A formulation according to any one of Claims 1 to 12, wherein said macrocyclic lactone is ivermectin. Cc 10 14. A formulation according to any one of Claims 1 to 13, comprising, per Slitre of formulation: about 180g to about 240g benzimidazole; about 7.5g to about 20g macrocyclic lactone or an active derivative or salt thereof; about 150g to about 250g polyoxyethylene (20) sorbitan monolaurate; about 450g to about 550g diethylene glycol monobutyl ether; about 20g to about 50g PEG 200; about 10g to about 30g sodium dodecyl sulphate; and about 100g to about 200g of water.
11. 15. The formulation of Claim 14 which comprises about 240g triclabendazole and about 15g ivermectin per litre.
12. 16. An aqueous micellar formulation comprising a benzimidazole or active derivative or salt thereof, in combination with a macrocyclic lactone or active derivative or salt thereof and also comprising, per litre of formulation: from about 100g to about 400g veterinary-acceptable surfactant(s); from about 200g to about 750g veterinary-acceptable water-miscible solvent(s); and from about 50g to about 350g of water substantially as hereinbefore described with reference to any one of the examples.
13. 17. A method of treating or preventing a diseased or parasite-infested state in a mammal, comprising topically administering to said mammal a micellar formulation according to any one of Claims 1 to 16, wherein said disease or parasite-infested state comprises a liver fluke infection or infestation, a nematode 0 infection or infestation, or both a liver fluke and a nematode infection or Sinfestation in a mammal. 0 18. A method according to Claim 17, wherein the formulation is applied to Sthe mammal over as small a region as possible, while avoiding run-off of the 5 formulation so as to maximise the concentration of active agents per cm 2 of animal surface.
14. 19. A method according to Claim 18, wherein the band of formulation is applied starting from the thoracic vertebrae and proceeding towards the rump of the animal, and from about 18mg to about 24mg triclabendazole and from about C 10 0.75mg to about 2mg ivermectin are applied per kilogram animal. A method of treating or preventing a diseased or parasite-infested state in a mammal, comprising topically administering to said mammal a micellar formulation according to any one of Claims 1 to 12, substantially as hereinbefore described with reference to any one of the examples. Dated 10 December, 2008 Schering-Plough Pty Limited Patent Attorneys for the Applicant/Nominated Person SPRUSON FERGUSON