AU2006287247B2 - Method for administering a composition to an animal - Google Patents

Description

Method for administering a composition to an animal Technical Field The present invention relates to compositions and methods for administering a skin necrosis composition to an animal. The invention also relates to the administration of a composition to an animal for reducing the problem of fly strike by chemical "mulesing".

Background A well documented problem experienced by many sheep and wool producers io relates to the occurrence of fly strike in and around the breech area of sheep. Where the breech area of the sheep is not appropriately maintained it is subject to soiling by faecal matter and urine which is an attractant for insects (such as the gravid fly female) to lay eggs, resulting in an enhanced fly strike rate. This is known as breech strike.

Farmers use many methods to control breech strike including chemical application by jetting and spraying, timing of shearing and crutching to coincide with known times of high fly challenge. In addition, the control of diarrhoea by good worm control and nutritional management reduces the incidence of breech strike. However, no method is permanent and all leave sheep vulnerable to breech strike at certain unpredictable times.

Regular crutching of sheep, that is, shearing of wool from the breech region, is a partially effective preventative method in at least drier localities. However, crutching needs to be done reasonably regularly in order to be effective, and is thus laborious and adds significantly to the overall cost of wool production. Also, crutching alone, whilst assisting with dealing with fly strike, is not a complete solution to the problem because a further problem is that sheep generally have loose folds of skin in the breech region, and moisture urine and faeces caught in the folds of the skin results in dermatitis that creates fly attractant sites where fly strike is likely to occur.

Accordingly, methods to tighten the skin and permanently remove wool from the breech region of sheep in order to reduce the problem of fly breech have been investigated for many years. One such procedure, known as a mulesing operation, was developed by John Mules during the early 1930s and involves the surgical removal of folds of skin in the breech region of the sheep in order to tighten the skin to thereby reduce the folds of skin in this region, and increase the area of flat, bare skin on the

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2 perineum. In its most basic form, the operation involves cutting the fold of skin on either side of the perineal area and then the allowing the resulting wound to heal. This operation Shas been modified over the years to remove skin on or adjacent the tail region of the sheep and also in the crutch region. More radical procedures involve the removal of skin s from the tail, the so called "Radical Mules Operation" Once the wounds caused by the mulesing operation have healed, the skin around I the crutch region of the sheep is generally pulled tighter, thereby expanding the naturally 00 occurring bare area of skin around the anus and in the perineum region of the sheep, IND thereby reducing the problem of soiling in this region, and hence reducing the incidence of breech fly strike.

However, the mulesing operation is not without its problems. One of the main problems is that it is painful and animal welfare considerations have long demanded a change to this practice. In addition, the wounds resulting from the operation are subject to infection and can be associated with the transmission of arthritis and other infections is and are themselves susceptible to fly strike until they are healed. The wounds also take a significant period of time to heal which results in stress to and reduced growth of the lambs.

Accordingly, there is a need to find an alternative to the mulesing operation, which is less painful and injurious to the animal, but which will provide at least some of the advantages of the mulesing operation.

Summary Described herein is a method of administering a skin necrosis composition to an animal, the method comprising administering the composition intradermally to an area of the skin of the animal.

Also described herein is a method for reducing the incidence of fly strike in an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition to an area of the skin of the animal.

Further described herein is a method of mulesing the breech region of an animal, the method comprising intradermally administering a skin necrosis composition to the breech region of the animal.

In a first aspect, the invention provides a method of administering a skin necrosis composition comprising a quaternary ammonium compound and a non-newtonian viscosity imparting agent to an animal, the method comprising administering the composition intradermally to an area of the skin of the animal.

860694 I:LNB SIn a second aspect, the invention provides a method for reducing the incidence of fly strike in an animal, the method comprising intradermally administering an effective n, amount of a skin necrosis composition comprising a quaternary ammonium compound and a non-newtonian viscosity imparting agent to an area of the skin of the animal.

In a third aspect, the invention provides a method of mulesing the breech region of an animal, the method comprising intradermally administering a skin necrosis N composition comprising a quaternary ammonium compound and a non-newtonian 00 viscosity imparting agent to the breech region of the animal.

INDThe skin necrosis composition may comprise a quaternary ammonium O io compound.

The skin necrosis composition may further comprise a viscosity imparting agent.

The skin necrosis composition may be delivered with, or further comprise, one or more vascoconstrictors.

The necrosis composition may be delivered with, or further comprise, one or Is more anaesthetics.

In a fourth aspect, the invention provides a method of administering a skin necrosis composition to an animal, the method comprising administering the composition intradermally to an area of the skin of the animal, wherein the composition comprises one or more quaternary ammonium compounds, and a non-newtonian viscosity imparting agent, and wherein the composition is optionally delivered with, or further comprises, one or more vasoconstrictors, and wherein the composition is optionally delivered with or further comprises one or more anaesthetics.

In a fifth aspect, the invention provides a method for reducing the incidence of fly strike in an animal, the method comprising the step of intradermally administering an effective amount of a skin necrosis composition to an area of the skin of the animal, wherein the composition comprises one or more quaternary ammonium compounds, and a non-newtonian viscosity imparting agent, and wherein the composition is optionally delivered with, or further comprises one or more vasoconstrictors, and wherein the composition is optionally delivered with or further comprises one or more anaesthetics.

In a sixth aspect, the invention provides a method of mulesing the breech region of an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition to the breech region of the animal, wherein the composition comprises one or more quaternary ammonium compounds, and a non-newtonian viscosity imparting agent, and wherein the composition is optionally delivered with, or comprises 860694 I:LNB one or more vasoconstrictors, and wherein the composition is optionally delivered with or comprises one or more anaesthetics.

Described herein is a method of administering a skin necrosis composition to an animal, the method comprising administering the composition intradermally to an area of s the skin of the animal, wherein the composition comprises one or more quaternary Sammonium compounds, and optionally a viscosity imparting agent, and wherein the N composition is optionally delivered with, or further comprises, one or more 00 vasoconstrictors, and wherein the composition is optionally delivered with or comprises

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IO one or more anaesthetics.

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Described herein is a method for reducing the incidence of fly strike in an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition to an area of the skin of the animal, wherein the composition further comprises one or more quaternary ammonium compounds, and optionally a viscosity imparting agent, and wherein the composition is optionally delivered with, or comprises is one or more vasoconstrictors, and wherein the composition is optionally delivered with, or comprises one or more anaesthetics.

Described herein is a method of mulesing the breech region of an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition to the breech region of the animal, wherein the composition comprises one or more quaternary ammonium compounds, and optionally a viscosity imparting agent, and wherein the composition is optionally delivered with, or further comprises one or more vasoconstrictors, and wherein the composition is optionally delivered with or further comprises one or more anaesthetics.

The viscosity imparting agent may be Newtonian, for example glycerol, or nonnewtonian, for example a cellulose derivative or polyvinylpyrrolidone.

The vascoconstrictor may be selected from a directly acting vasoconstrictor, for example epinephrine and phenylephrine, an indirectly acting vasoconstrictor, for example amphetamine and methamphetamine, or a mixed acting vasoconstrictor, for example metraminol and ephedrine, or any combination of directly, indirectly or mixed acting vasoconstrictors.

The anaesthetic may be selected from the group consisting of: procaine, prilocaine, mepivacaine, lignocaine, bupivacaine and any combination thereof.

860694 I:LNB SIn an embodiment of the first to sixth aspects, the animal may be selected from the group consisting of: sheep, goats, camelids, for example alpacas, llamas and camels.

In an embodiment of the first, second, fourth or fifth aspects, the area of skin of the animal may be the breech region.

The compositions used in the methods of the first to sixth aspects may be rdelivered together with a pharmaceutically acceptable carriers, diluents, adjuvants and/or Ni excipients and/or dyes.

00 In the methods of the first to sixth aspects, multiple simultaneous doses of the IDcomposition and/or the vasoconstrictor and/or the anaesthetic may be administered.

to In one embodiment, the compositions used in the methods of the first to sixth aspects may be delivered by high pressure injection.

The compositions used I the methods of the first to sixth aspects may be delivered with, or further comprise an alcohol. The alcohol may be a C 1 -Clo alcohol, for example methanol, ethanol, propanol, isopropanol, t-butanol, butanol, pentanol, and the Is like, or any combination thereof.

In a seventh aspect, the invention provides a skin necrosis composition, the composition comprising a synergistic combination of an effective amount of an alcohol and an effective amount of one or more skin necrotizing agents.

In an eighth aspect, the invention provides a method of administering a skin necrosis composition to an animal, the method comprising intradermally administering an effective amount of the skin necrosis composition, the composition comprising a synergistic combination of an effective amount of an alcohol and an effective amount of one or more skin necrotizing agents, to the breech region of the animal.

In a ninth aspect, the invention provides a method for reducing the incidence of fly strike in an animal, the method comprising intradermally administering an effective amount of the skin necrosis composition, the composition comprising a synergistic combination of an effective amount of an alcohol and an effective amount of one or more skin necrotizing agents, to an area of the skin of the animal.

In a tenth aspect, the present invention provides a method of mulesing the breech region of an animal, the method comprising intradermally administering an effective 860694 I:LNB amount of a skin necrosis composition, the composition comprising a synergistic combination of an effective amount of an alcohol and an effective amount of one or more skin necrotizing agents, to the breech region of the animal.

The one or more skin necrotizing agents may be quaternary ammonium s compounds.

The quaternary ammonium compound may be cetrimide.

The alcohol may be a CI-Co 10 alcohol, for example, methanol, ethanol, propanol, isopropanol, butanol, t-butanol pentanol, hexanol or the like, or any combination thereof The skin necrosis composition of the seventh aspect may comprise a viscosity imparting agent.

The skin necrosis composition of the seventh aspect may be delivered with, or comprise, one or more vasoconstrictors.

The skin necrosis composition of the seventh aspect may be delivered with, or Is comprise, one or more anaesthetics.

The compositions used in the methods of the eighth to tenth aspects may be delivered together with pharmaceutically acceptable carriers, diluents, adjuvants and/or excipients and/or dyes.

In the methods of the eighth to tenth aspects, multiple simultaneous doses of the composition and/or the vasoconstrictor and/or the anaesthetic may be administered.

In one embodiment, the compositions used in the methods of the eighth to tenth aspects may be delivered by high pressure injection.

Definitions The following are some definitions that may be helpful in understanding the description of the present invention. These are intended as general definitions and should in no way limit the scope of the present invention to those terms alone, but are put forth for a better understanding of the following description.

Throughout this specification, unless the context requires otherwise, the word "comprise", or variations such as "comprises" or" comprising", will be understood to imply the inclusion of a stated step or element or integer or group of steps or elements or integers, but not the exclusion of any other step or element or integer or group of elements or integers. Thus, in the context of this specification, the term "comprising" means "including principally, but not necessarily solely".

As used herein, the term "skin necrosis composition" refers to a composition which has the effect of necrosing the skin.

As used herein, the term "skin necrotizing agent" refers to an agent, such as a chemical compound, which has the effect of necrosing the skin.

As used herein, the term "intradermally" refers to the zone of the skin beneath the epidermis and above the subcutaneous tissue.

As used herein, the term "breech region" refers to the area around the vulva and io anus of the animal that may collect urine and/or faeces. It includes the perineum, the backs of the upper part of the hind legs, and also the tail.

An "effective amount", as used herein, includes within its meaning a sufficient amount of the particular composition to which it is referring to provide the desired necrotizing effect.

As used herein, the term "multiple simultaneous doses" refers to separate doses of the composition that are delivered at substantially the same time, but not at the exact same location. For example, a total of 16 doses may be delivered in a grid-like fashion about 1 cm apart from each other to allow for 0.5cm diffusion of the compound once delivered beneath the surface of the skin simultaneously in a 3cm x 3cm grid. This results in treatments ofa 16cm 2 skin area (allowing for diffusion).

As used herein, the expression "delivered with" in the context of an additional agent such as a local anaesthetic and/or a vasoconstrictor, includes the situation where the additional agent is present in the skin necrosis composition that is administered, and also the situation where the additional agent is not present in the skin necrosis composition, but rather is administered separately, simultaneously or sequentially with the skin necrosis composition.

As used herein, the term "fly strike" means infestation of tissue by larvae or maggots of flies.

Detailed Description The present invention is based on the discovery by the inventors that the intradermal delivery of a skin necrosis composition beneath the surface of an area of skin in the breech region of an animal causes the skin to necrose (or die) and eventually detach from the animal. The removal of this skin results in a tightening of the remaining skin in the breech region which removes wrinkles and stretches the perineal bare skin, thereby providing at least some of the advantages of the mulesing operation.

The intradermal delivery of a skin necrosis composition facilitates the reduction of the incidence of fly strike in the breech region of the animal and also facilitates the shedding of urine, prevention of dag formation and urine stain on the breech region and the back legs of the animal, which is particularly undesirable.

The methods of the present invention are useful for the same or similar problems that exist in any animal which suffers from or is prone to flystrike, for example, animals io with excessive hair or wool about the breech region which the animal either finds difficult to groom, or neglects to groom. Such animals include sheep, llamas, goats, alpacas and other camelids. The animal may be a sheep, for example a lamb, or a weaner or the like.

Without wishing to be bound by theory or mode of action, it is believed that intradermal administration of the skin necrosis composition under the surface of the skin minimises pain in the animal that occurs in slower and labour intensive delivery methods, such as topical application or subcutaneous administration. Again, without wishing to be bound by any particular theory, it is believed that the delivery of the composition beneath the surface of the skin results in the rapid destruction of nerves in the region. Such a means of delivery also permits relatively quick administration of the compositions, if desired. As a consequence, the same or similar outcome as the mulesing operation can be achieved with a reduction in pain experienced by the animal.

Moreover, an anaesthetic may be included with the composition to further assist in this respect.

Because the compositions used in the methods of the present invention are administered intradermally, necrosis occurs rapidly resulting in minimal pain and discomfort for the animal.

The methods of the present invention may be used in conjunction with other known mulesing methods, for example the methods described in Australian patent application 2006202715 entitled "Treatment of sheep to reduce flystrike". The methods of the present invention may also be used in conjunction with surgical procedures in, for example, the breech region and cautery to the tail.

Compositions The compositions used in the methods of the invention include one or more skin necrotizing agents. In one embodiment, the skin necrotizing agent is one or more quaternary ammonium compounds.

Quatemary ammonium compounds are advantageous as they are widely used in human and veterinary medicine and dentistry, such as in washes and wound dressings, so they are readily available and are relatively cheap.

Quaternary ammonium compounds are also well known by government regulators, so a substantial body of safety and toxicity data already exists. European o1 Medicines Agency (EMEA) has classified two quaternary ammonium compounds, cetrimide and benzalkonium chloride, as not requiring the establishment of maximum residue limits (MRLs) because their use in animals would not lead to tissue residues toxicologically relevant to the consumer.

The quaternary ammonium compound(s) for use in the methods and compositions is of the present invention may be selected from the group consisting of: cetrimide (which is a mixture of laurtrimonium bromide, myristyltrimethylammonium bromide, and palmityltrimethyl ammonium bromide), alkyltrimethylammonium bromides, cetylpyridinium chloride, benzalkonium chloride, dioctyldimethylammonium chloride, octyldecyldimethylammonium chloride, didecyldimethylammonium chloride, hexadecyltrimethylammonium bromide, methylbenzethonium chloride, Alcian Blue 8GX, polypropoxy quaternary ammonium chlorides (Emcol® CC9, Emcol® CC42), polypropoxy quaternary ammonium phosphate (Emcol® CC57), trimethyl C8C10 quaternary ammonium chloride (Adogen® 464), polyquaterary ammonium chloride (Mirapol® WT), amine hydrochloride (guanidine hydrochloride), imine compound (polyethyleneimine), sodium alginate, octylphenoxypolyethoxyethanol (Triton® X- 100), choline chloride, acetylcholine chloride, carboxymethyltrimethylammonium chloride, tetraethylammonium chloride, tetrabutylammonium bromide, tetraheptylammonium bromide, phenyltriethylammonium bromide, phenyltriethylammonium iodide, tricaprylmethylammonium chloride, trioctylpropylammonium bromide, nonyltrimethylammonium bromide, octyldecyldimethylammonium chloride (Bardac 2080), didecyldimethylammonium chloride (Bardac 2280, Quatramine 2-10/80), didecylmethylpropylammonium chloride, didecyldipropylammonium chloride, dodecyltrimethylammonium chloride, decamethonium bromide, hexamethonium bromide, laurtrimonium bromide, myristyltrimethylammonium bromide, and palmityltrimethyl ammonium bromide, or any combination thereof.

In one embodiment, the quaternary ammonium compound may be selected from the group consisting of: alkyl trimethylammonium bromides, cetrimide, cetylpyridinium chloride, benzalkonium chloride, hexadecyltrimethylammonium bromide, cetyltrimethylammonium bromide and cetrimonium bromide.

In another embodiment, the quaternary ammonium compound is cetrimide and/or cetylpyridinium chloride.

Quaternary ammonium compounds are known to cause chemical burs at high concentrations and can cause irritant contact dermatitis. As such, the one or more quaternary ammonium compounds may be present in the skin necrosis composition in an amount between about 1% to about 10% or between about 1% to about 8% or between about 1.5% to about 8% or between about 1.75% to about 8% (w/w),or between about 2% to about or between about 1.75% to about 7.5% or between about 1.75 to about 7% or between about 2% to about 6.5% or between about 2% to about or between about 2% to about 5% or between about 2% to about 4.75 or between about 2% to about 4.5% or between about 2% to about 4% of the total composition. In one embodiment, the quaternary ammonium compound is present in the skin necrosis composition in an amount between about 2% or 3% or 4% The concentration of the quaternary ammonium compound and the volume given per administration in the skin necrosis composition can be adjusted based on the required total amount of chemical to be administered and the total number of doses delivered intradermally to the animal per unit of skin area. For example, if a 2% solution is used with a total of 16 doses each delivered 1 cm apart on a 3 cm x 3 cm grid, the resultant treatment is 2 mg of quaternary ammonium compound per cm 2 of skin, i.e.: 16 injections x 0.1 ml/dose x 20 mg/ml 32 mg/16cm 2 =2 mg/cm 2 It will be apparent that other combinations of volume injected, concentration of quaternary ammonium compound and spacing can be used to achieve 2 mg/cm 2 The skin necrosis composition may be viscous and/or include one or more viscosity imparting agents. The presence of the viscosity imparting agent assists in retaining the composition at the targeted site of administration, thereby minimising diffusion and damage and/or pain to the surrounding areas.

The viscosity imparting agent may be newtonian, for example glycerol, or nonnewtonian, for example a cellulose derivative or polyvinylpyrrolidone.

The amount of the viscosity imparting agent present in the composition may be between about 0.5% and about 10% or between about 0.5% and about 9% or between about 1% and about 8% or between about 1.5% and about or between about 1% and about 7% or between about 1% and about or between about 1% and about 6% or between about 1.5% and about 6% or between about 1.75% and about 6% or between about 2% and about 6% or between about 2% and about 5.5% or between about 2% and about or between about 2% and about 4.5% or between about 2% and about 4% of the total composition.

The viscosity imparting agent may result in the skin necrosis composition having a thixotropic property. That is, the composition may form a gel once delivered beneath the surface of the skin.

The skin necrosis composition may also be delivered with one or more vasoconstrictors. The vasoconstrictor acts to constrict blood vessels and thus restrain the skin necrosis composition at the site of delivery, thereby minimising damage and/or pain to surrounding areas.

The one or more vasoconstrictors may be present in the skin necrosis composition, or alternatively the vasoconstrictors may be administered separately, simultaneously or sequentially with the skin necrosis composition. When present in the skin necrosis composition, the amount of the one or more vasoconstrictors may be between about 0.0005% and 3% or between about 0.0005% and 2% or between about 0.001% and about or between about 0.005% and 1.5% or between about 0.005% and 1.25% or between about 0.005% and 1.0% or between about 0.005% and 0.75% or between about 0.005% and 0.1% of the total composition. In one embodiment, the amount of the vasoconstrictor may be about 0.01% The one or more vasoconstrictors may be selected from the directly acting group of vasoconstrictors, for example adrenaline, epinephrine and phenylephrine; or the indirectly acting group of vasoconstrictors, for example amphetamine and methamphetamine. The one or more vasoconstrictors may also be selected from the mixed acting group of vasoconstrictors, for example metraminol and ephedrine. The one or more vasoconstrictors may be synthetic molecules.

In one embodiment, where the viscosity imparting agent is polyvinylpyrrolidone, the composition may not include a vasoconstrictor, as the polyvinylpyrrolidone may act to physically restrain the treatment at the site of administration.

to The skin necrosis composition may also be delivered with one or more local anaesthetics. The one or more local anaesthetics may be present in the skin necrosis composition, or alternatively the local anaesthetics may be administered separately, simultaneously or sequentially with the skin necrosis composition. The amount of local anaesthetic administered may be between about 1 to 4% of the total composition.

Is The anaesthetic solution used may have a concentration of about 10 to 30 mg/ml, or alternatively about 20 mg/ml.

The one or more local anaesthetics act to even further prevent or reduce any acute and sub-acute pain that may occur in response to the delivery of the skin necrosis composition and resulting necrosis of skin in the area of the animal in the breech region).

The local anaesthetics may be selected from the sodium channel blocker group, for example procaine, prilocaine, mepivacaine, lignocaine, and bupivacaine. The use of other components used for other purposes, but having local anaesthetic properties are also contemplated in the methods of the invention. For example, opioid analgesics, beta-adrenoreceptor antagonists, and anti-histamines.

The skin necrosis composition may also include an alcohol, which may be absolute dried) or aqueous, for example between 80% and 99%, or between and 99%, or between 90% and 99%, or between 92% and 97%. The alcohol may be a Ci-Clo alcohol, for example methanol, ethanol, propanol, isopropanol, butanol, pentanol, hexanol etc, or any combination thereof. In another embodiment, the alcohol may be a CI-C 6 alcohol, or a Ci-C 4 alcohol. In one embodiment, the alcohol is aqueous ethanol.

The presence of an alcohol in the composition may serve to reduce foaming. The alcohol may also increase the efficacy of the composition, for example in a synergistic manner. As such, the present invention also relates to a synergistic combination comprising an effective amount of an alcohol and an effective amount of one or more skin necrotizing agents. The invention also relates to administration of such a composition intradermally.

It has been observed that where an alcohol, for example ethanol, is included in the composition, and the amount of the necrotizing agent (for example a quatemrnary ammonium compound) reduced, the time taken for skin necrotization and subsequent io wound healing is reduced as compared to a composition comprising a greater amount of the nectrotizing agent in the absence of the alcohol. In some cases where an alcohol, is present in the composition, and the amount of quaternary ammonium compound is reduced by up to about 20%, or 30%, or 40%, or 50%, or 60%, the time taken for skin necrotization and subsequent wound healing is reduced..

is The amount of alcohol present in the composition may be between about 0.5% to about 3% or between about 0.5% to 2.75% or between about 0.5% to or between about 0.5% and about 2.25% or between about 0.75% and about 2% or between about 1% and about 2% or between about 1.2 and about 1.7% or between about 1.3% and about 1.75% of the total composition.

In one embodiment, the amount of alcohol present may be about 1.5% In one embodiment, the skin necrosis composition may comprise about 1.5% to about of a quaternary ammonium compound, and about 1.0% to about 2.0% of an alcohol, which may be ethanol.

Whilst it is possible for the skin necrosis composition to be delivered alone, it is preferable that the component(s) of the composition be delivered together with one or more pharmaceutically acceptable carriers, diluents, adjuvants and/or excipients and/or dyes.

In general, pharmaceutical compositions may be prepared according to methods which are known to those of ordinary skill in the art.

Each carrier, diluent, adjuvant and/or excipient must be pharmaceutically acceptable such that it is compatible with the components of the skin necrosis composition and does not cause any undesirable adverse side effects to the animal.

Examples of pharmaceutically acceptable carriers or diluents are demineralised or distilled water, saline solution, vegetable based oils such as peanut oil, safflower oil, olive oil, cottonseed oil, maize oil, sesame oils such as peanut oil, safflower oil, olive oil, cottonseed oil, maize oil, sesame oil, arachis oil or coconut oil; silicone oils, including polysiloxanes, such as methyl polysiloxane, phenyl polysiloxane and methylphenyl polysolpoxane; volatile silicones, mineral oils such as liquid paraffin, soft paraffin or squalene, cellulose derivatives such as methyl cellulose, ethyl cellulose, carboxymethylcellulose, sodium carboxymethylcellulose or hydroxypropylmethylcellulose, lower alkanols, for example ethanol or iso-propanol; lower aralkanols, lower polyalkylene glycols or lower alkylene glycols, for example polyethylene glycol, polypropylene glycol, ethylene glycol, propylene glycol, 1,3butylene glycol or glycerine, fatty acid esters such as isopropyl palmitate, isopropyl myristate or ethyl oleate, polyvinylpyrridone, agar, carrageenan; gum tragacanth or gum acacia, and petroleum jelly. The carrier or carriers may form from between 10% to 99.9% by weight of the compositions.

For administration as an injectable solution or suspension, non-toxic diluents or carriers can include Ringer's solution, isotonic saline, phosphate buffered saline, ethanol and 1,2-propylene glycol.

Suitable adjuvants typically include emollients, emulsifiers, thickening agents, preservatives, bactericides and buffering agents.

It is contemplated that the pharmaceutically acceptable carrier may include a water-soluble substance or a water-insoluble substance.

In addition to pharmaceutically acceptable carriers, diluents, adjuvants and/or excipients and/or dyes, the compositions may also include anti-foaming agents, such as alcohols.

The skin necrosis composition may also be delivered separately, sequentially or simultaneously with one or more other suitable compositions to the animal, for example compositions comprising one or more vasoconstrictors and/or anaesthetics.

The pH of the skin necrosis composition may be between about 6.5 and about 8.2, or between about 6.7 and about 8.0, or between about 6.8 and about 7.9, or between 6.9 and 7.9 or between 7.0 and 7.8.

The compositions used in the methods of the invention may also include dyes to allow visualisation of treated areas of the skin so as to avoid double treatment.

Examples of suitable dyes include, but are not limited to, brilliant blue FCF and methylene blue.

In one embodiment, compositions that may be used in the methods of the invention include the following: a composition comprising a quaternary ammonium compound and a viscosity imparting agent; (ii) a composition comprising a quaternary ammonium compound, a vasoconstrictor and a viscosity imparting agent; (iii) a composition comprising a quaternary ammonium compound, a o0 vasoconstrictor, a viscosity imparting agent and an alcohol; (iv) a composition comprising a quaternary ammonium compound and a vasoconstrictor; a composition comprising a quaternary ammonium compound, an alcohol and a viscosity imparting agent, and (vi) a composition comprising a quaternary ammonium compound a vasoconstrictor and an alcohol.

In another embodiment, compositions that may be used in the methods of the invention include the following: a composition comprising cetrimide and polyvinylpyrrolidone; (ii) a composition comprising cetrimide, phenylephrine and polyvinylpyrrolidone; (iii) a composition comprising cetrimide, phenylephrine, polyvinylpyrrolidone and ethanol; (iv) a composition comprising phenylephrine and cetrimide; a composition comprising cetrimide, ethanol and polyvinylpyrrolidine and (vi) a composition comprising cetrimide, phenylephrine, and ethanol; The amount of cetrimide present in the compositions may be between 1.5% and 8% of the total composition, the amount of phenylephrine in the compositions may be between 0.0005% and 1% of the total composition, the amount of polyvinylpyrrolidone present in the compositions may be between 1% and 10% (w/w) of the total composition, and the amount of ethanol present in the compositions may be between 0.5 and 2.5% of the total composition.

Methods of administration Where the animal to which the skin necrosis composition is to be administered is a sheep, it is envisaged that the composition may be administered to young sheep at or around the age when a mulesing operation would traditionally be conducted on the lamb. Treatment of older sheep is now also possible as welfare objections to treating older sheep no longer apply.

The skin necrosis composition may also be intradermally administered beneath the surface of an area of the skin more than once at various time intervals, for example prime administration followed by a boost administration.

In one embodiment, multiple simultaneous doses are administered by an apparatus capable of parallel delivery.

Multiple simultaneous doses of the composition may be administered to an area of the skin of an animal, for example in the breech area. Multiple simultaneous doses may be administered between about 0.1 and 2.0 cm, or between about 0.1 and 1.5 cm or between 0.5 and 1.5 cm apart in a grid pattern on the area of the skin.

The skin necrosis composition may be a liquid that is administered in a volume of between about 0.01 ml and about 1 ml, or between about 0.1 ml and about 0.5 ml. In one embodiment, the volume of the composition delivered may be about 0. 1 ml.

The skin necrosis composition may be at any appropriate pH, such as a physiological pH, i.e between about pH 7.0 to about 7.8, and may also be isotonic.

A number of means of intradermal delivery of the composition under the surface of the skin are contemplated. For example the composition may be delivered under the surface of the skin by use of a mechanical injection means, i.e, either needle or a needleless means. A needle means may include multiple needles arranged in a predetermined manner to inject the composition at spaced apart sites. Needle-less means may include multiple means arranged in a predetermined manner for injecting at high pressure the skin necrosis composition at a sufficient velocity to rapidly penetrate the epidermis into the dermal region directly without piercing the lowest layer of skin the hypodermis.

The composition may be delivered by a needle-less means so as to avoid issues that would otherwise arise with the use of a needle mechanical injection means, such as disposal of needles. Examples of suitable needle-less means include the MIT Agro jet and the dermaject.

16 It is also contemplated that the composition may be delivered under the surface of the skin by use of a non-mechanical means that allows for the rapid delivery of the composition beneath the surface of the skin.

Delivery of the composition intradermally provides that little if any of the composition will be left on the surface of the skin once administration has occurred.

This avoids toxicity to the animal that may occur due to grooming of the breech region by the animal or another animal and any chemical burn on the surface of the skin. This also reduces the risk to an operator administering the composition, since exposure of the compound to the external environment is minimised.

to Delivery of the composition as already described may also provide that relatively lower concentrations are required as most (if not all) of the composition penetrates the skin. This also provides safer handling by the operator, and minimises toxicity to the animal.

The site of delivery of the skin necrosis composition is within the dermis of the is skin in the zone beneath the epidermis and above the subcutaneous tissue.

Although conditions such as fly strike usually exist at the rear of the animal, the area of skin where the composition is delivered is not limited to the rear of the animal.

It is contemplated that it may be necessary to deliver such a composition beneath the surface of the skin of the animal in other regions. Furthermore, the composition may be delivered beneath the surface of the skin for the purpose of treating conditions which require an area of the skin to be necrosed, eg, removal of skin cancers, branding, pizzle ringing, wigging and jowling.

It will be understood that the invention disclosed and defined in this specification extends to all alternative combinations of two or more of the individual features mentioned or evident from the text. All of these different combinations constitute various alternative aspects of the invention.

Examples The invention will now be described in more detail, by way of illustration only, with respect to the following examples. The examples are intended to serve to illustrate this invention and should not be construed as limiting the generality of the disclosure of the description throughout this specification.

Example 1 Compositions for use in the methods Examples of compositions that may be administered in the methods of the invention include the following: 1. Cetrimide (2 to 4.5% w/w) and polyvinylpyrrolidone (0.5 to 3.5% w/w), pharmaceutically acceptable carriers, diluents, adjuvants, dyes and/or excipients (up to 100%); 2. Cetrimide (2 to 4.5 polyvinylpyrrolidone (0.5 to 3.5% w/w), phenylephrine 0.005% to 0.1% pharmaceutically acceptable carriers, diluents, adjuvants, dyes and/or excipients (up to 100%); 3. Cetrimide (1.5 to 3% polyvinylpyrrolidone (0.5 to 3.5% w/w), phenylephrine (0.005% to 0.1% ethanol (1.0 to 2.0% w/w), pharmaceutically acceptable carriers, diluents, adjuvants dyes and/or excipients (up to 100%); Example 2 Use of quaternary ammonium (QA) compounds benzalkonium chloride cetrimide (CT) and cetylpyridinium chloride (CPC) as a skin necrosis agent Use of quaternary ammonium (QA) compounds benzalkonium chloride (BC), cetrimide (CT) and cetylpyridinium chloride (CPC) at various concentrations to cause necrosis of the skin in the breech region of the sheep was examined. The use of these QAs was compared to the use of phenol.

Treatment A total of 17 sheep were prepared with 15 different treatments (categorised as groups 1 to 15) outlined in Table 1 below. Each sheep received 3 different treatments on the flank region.

An aqueous injection solution with various concentrations on molecular weight basis was prepared.

Group 1 to 15 treatments were administered by 16 x 0.lml intra-dermal needle injections each injection spaced Icm apart in a 3cm x 3cm grid-formation (referred to as a grid site). A total of 45 grid sites were tested.

18 Group 16 and 17 treatments were administered by painting 0.lml of higher concentrations of phenol (at 20% and 40% w/w) onto the skin (ie applied topically). A total of 6 sites were tested.

TABLE 1 Group Active Constituents Dose Volume No. of sites 1 Aqueous blank 16x 0.1 mL injections at 1 cm centre 3 2 1% phenol 16x 0.1 mL injections at 1 cm centre 3 3 2.5% phenol 16x 0.1 mL injections at 1 cm centre 3 4 5% phenol 16x 0.1 mL injections at 1 cm centre 3 10% phenol 16x 0.1 mL injections at 1 cm centre 3 6 20% phenol 16x 0.1 mL injections at 1 cm centre 3 7 1% benzalkonium chloride (BC) 16x 0.1 mL injections at 1 cm centre 3 8 2% benzalkonium chloride (BC) 16x 0.1 mL injections at 1 cm centre 3 9 4% benzalkonium chloride (BC) 16x 0.1 mL injections at 1 cm centre 3 1% cetrimide (CT) 16x 0.1 mL injections at 1 cm centre 3 11 2% cetrimide (CT) 16x 0.1 mL injections at 1 cm centre 3 12 4% cetrimide (CT) 16x 0.1 mL injections at 1 cm centre 3 13 1% cetylpyridinium chloride (CPC) 16x 0.1 mL injections at 1 cm centre 3 14 2% cetylpyridinium chloride (CPC) 16x 0.1 mL injections at 1 cm centre 3 4% cetylpyridinium chloride (CPC) 16x 0.1 mL injections at 1 cm centre 3 16 20% phenol About 0.1 mL topically until the skin is 3 wet before run off occurs 17 40% phenol About 0.1 mL topically until the skin is 3 wet before run off occurs Assessment After administration of the QA to the breech region, the treated area swelled. A scab eventually formed at the administration or grid site. The scab dislodged at around 28 to 32 days after treatment leaving the area of skin scarred and stretched with minimal o1 wrinkles.

Swelling, Scab formation, Scar formation and Wound contraction at each administration site were measured to determine effectiveness of administration of each QA tested for skin necrosis and resulting wrinkle removal and stretch the perineal bare area.

Swelling and Scab formation were assessed by eye and scored as normal mild moderate or severely affected Scar formation was expressed as the percentage of the treated area occupied by a mature scar.

Wound Contraction was measured in cm and facilitated by placing parallel lines (usually tattooed) about 2cm away from the lateral edge of each administration or grid site.

Each parameter was measured during necrosis and healing at the region of application as follows: 1. Swelling was measured at days 1 or 5 after treatment.

2. Scab formation (Scab): was measured at days 21 or 32 after treatment.

3. Scar formation (Scar): was measured at day 32 after treatment.

4. Wound contraction (Wound): was measured at day 32 after treatment.

Observations Table 2 below shows the most relevant results of Swelling Scab formation (Scab), Scar formation (Scar) and Wound contraction (Wound) post intra-dermal administration of various concentrations of QAs compared to topical administration of phenol to the same region of the sheep.

TABLE 2 Group Active Sw Scab Scar Wound constituent (mean score)' (mean score) S w/w: 7 1% BC >1.5 0.5 to 1 1% cetrimide 11 2% cetrimide >1.5 >0.3 12 4% cetrimide >1.5 >35 0.5 to 1 13 1%CPC 4% CPC >1.5 >0.3 >35 0.5 to 1 16 20% phenol 0.3 0.5 to 1 Conclusion From these observations, 4% cetrimide and 4% cetylpyridinium chloride (CPC) were considered to be the most effective of the QA treatment investigated in this example.

EXAMPLE 3: Testing various concentrations, spacings and volumes of cetrimide (CT) and cetylpyridinium chloride (CPC) and addition of adrenaline The effect of volume, concentration dose and spacing of multiple doses were explored with cetrimide (CT) and cetylpyridinium chloride (CPC). The effect of the addition of Adrenaline was also tested.

Treatment A total of 29 sheep were prepared with 17 different treatments (categorised as groups 1 to 17) outlined in Table 3 below. Each sheep received 3 different treatments on the flank of the sheep. Thus a total of 87 sites were tested, providing 5 replicates of 0o each group.

Each administration site consisted of about 3cm x 3 cm grid with injection spacings as indicated.

The injectable solutions were prepared as outlined in Example 2.

0.1 mg/ml adrenaline was combined with the QA solution and administered in the same injection in groups 15 and 16.

TABLE 3 Treatment regime 1 cetrimide 4% 1.0 0.1 Dose confirmation 3 cetrimide 4% 1.0 0.1 Dose confirmation 2 cetrimide 2% 1.0 0.1 Dose confirmation 3 cetrimide 8% 1.0 0.1 Dose confirmation 4 cetrimide 8% 1.0 0.05 Volume cetrimide 2% 1.0 0.2 Volume 6 cetrimide 4% 1.5 0.225 Spacings 7 cetrimide 4% 0.5 0.025 Spacings 8 CPC 4% 1.0 0.1 Dose confirmation 9 CPC 2% 1.0 0.1 Dose confirmation CPC 8% 1.0 0.1 Dose confirmation 11 CPC 8% 1.0 0.05 Volume 12 CPC 2% 1.0 0.2 Volume 13 CPC 4% 1.5 0.225 Spacings 14 CPC 4% 0.5 0.025 Spacings cetrimide 0.1 4% 1.0 0.1 Adrenaline mg/ml adrenaline ____addition 16 CPC 0.1 4% 1.0 0.1 Adrenaline mg/ml adrenaline addition 17 Aqueous blank 1.0 0.1 Control Assessment Skin reactions were assessed as in Example 2.

Observations/Conclusions 4% cetrimide plus adrenaline and 4% cetylpyridinium given at 0.1 ml per injection with a 1 cm spacing were the most effective treatments investigated Example 4: Treatment of the breech area of sheep and the addition of local anaesthetic to QA as necrosis agent This example examines the effectiveness of the lead treatments to treat the breech io area and replace surgical mulesing. The addition of a local anaesthetic to the QA as a necrosis agent was also investigated.

Treatment/Assessment 4% cetrimide plus adrenaline, 4% cetrimide plus adrenaline plus lignocaine and 4% cetylpyridinium were each injected into the intradermal skin on the breeches of six sheep using a needless applicator that for each shot delivered 0.5 ml intradermally to a cm 2 area of skin. The area of treated skin was similar to the skin removed in the usual mulesing operation except that discrete areas of skin adjacent to and on top of the tail were treated rather that the whole area removed in the 'tail stripping' operation.

Observations The result was necrosis of the treated skin to give an effect similar to that following surgical mulesing with, removal of breech wrinkle and expansion of the perineal bare area. No undue pain or discomfort was observed.

Example 5: Treatment of the breech area of sheep with cetrimide-containing compositions 1. Validating the practical utility of intradermal injections of cetrimide in the breech ofMerino sheep in the field as an alternative to mulesing Field trials were conducted in four commercial flocks. At each site the breech region of 100 ewe lambs or weaners was given intradermal treatment with a 4% concentration of cetrimide and 0.01% adrenaline. A total of 35 injections were made to each side of the breech in a 1cm grid pattern using a needle-less applicator. Treated animals were assessed for perineal bare area stretch 30, 60, 90 and 180 days after treatment. A score for breech wrinkle, accumulated dag and urine stain on a scale from (minimal to severe) was also noted at each assessment point.

Results In all four field trials there was an increase in perineal bare area in treated animals. A reduction in breech wrinkle was noted. Results were similar to those following surgical mulesing.

2. Development of the optimum compositions following evaluation of the systemic io safety, site behaviour and local tissue reaction of several Cetrimide based formulations when administered into the skin ofMerino sheep 4% solutions of cetrimide plus 0.01% adrenaline reliably caused dermal necrosis following intra-dermal injection. However, adrenaline is not stable at neutral pH and the use of phenylephrine as an alternative vasoconstrictor was investigated. The effect of tonicity and pH needed to be discerned. The use of a gel forming formulations to restrict clearance and prolong efficacy was investigated as was the use of ethanol to reduce foaming. 26 female Merino sheep were treated on the mid side with 13 test formulations of 4% cetrimide (six replicates) into the dermis of the skin using a needless applicator. The formulations are shown below in Table 4.

TABLE 4 Treatment regime Grou Formiulation 1 4% cetrimide 0.01% adrenaline 2 4% cetrimide alone 3 Phenylephrine 0.005% 4% cetrimide 4 Phenylephrine 0.01% 4% cetrimide Phenylephrine 0.02% 4% cetrimide 6 10% Hypertonic 4% cetrimide 0.01% adrenaline 7 Hypotonic 4% cetrimide+ 0.01% adrenaline 8 10 x hypertonic 4% cetrimide 9 0.5% PVP 4% cetrimide 1.5% PVP 4% cetrimide 11 3% PVP 4% cetrimide 12 3% PVP 0.01% adrenaline 4% cetrimide 13 3% ethanol 4% cetrimide, 0.01% adrenaline Detailed treatment site examinations were made before treatment and at 24 and 48 hours post treatment, thence days 6 and 14 post-treatment. Each site was assessed for bruising, swelling/oedema, discolouration of the skin and over time scab formation and scar formation and contraction, i.e. measurement of gap between indelible marks.

Sheep were weighed, examined clinically at weekly intervals and observed in their pens for postural evidence of discomfort, normal or abnormal feeding and drinking and rubbing or chewing of treated areas. After 20 days the sheep were euthanasied the skin examined at necropsy the area of the effected skin measured and sections taken for histopathology.

lo Results No signs of pain or discomfort were noted. Groups 2, 6 and 11 gave the best wound contraction, 4 and 7 were also good. Group 7 gave the largest area of effected skin, 13 was average and 11, 2 and 6 were smallest.

Histopathology In nearly all cases there was profound full thickness necrosis of the epidermis, dermis and hypodermis. In group 13 there was an excessive reaction. Group 11 had low inflammation in the deeper tissues and moderate fibrosis consistent with repair.

Groups 2, 6, 11 had modest changes in deeper tissues and adequate fibrosis and inflammation. This was consistent with contraction. At the margins of the effected areas in many sections there was undergrowth of epidermis under the necrotic skin and sloughing. This was best in 11.

Phenylephrine gave equivalent reaction to adrenaline at all concentrations and 0.01% was optimum. The treated area declined and wound contraction increased as PVP concentration increased. 3% PVP gave the best efficacy. Tonicity was not significant. 3% ethanol with cetrimide gave an excessive response.

3. Development of the optimum formulation following evaluation of the systemic safety, site behaviour and local tissue reaction of several Cetrimide based compositions when administered to the skin ofMerino sheep via a needleless applicator Several injectable aqueous formulations of Cetrimide in combination with PVP and/or phenylephrine were evaluated for efficacy at causing skin necrosis and speed of wound resolution. The formulations are given below in Table 5. Twenty-seven unmulesed Merino wether weaners were treated on both mid-sides total six sites with 2 ml over an area of 20 cm 2 with one of 9 test or control formulations (18 replicates) using a needleless applicator. The treated area was observed at 4 hours thence 1, 2, 4, 7, 13, 22 and 28 days after treatment. On Day 20, 18 trial animals were sacrificed and the skin reflected and removed intact from each side. On Day 35, the remaining 9 trial animals were sacrificed and the skin removed intact from each side. The area of affected skin was measured and tissue sections collected for further histological examination.

1o TABLE 5 Treatment Regime reatment Cetrimide Phenylephrine, :'PVP Ethanol 1 4% -3% 2 4% 0.01% 3 4% 0.01% 3% 3% 4 4% 0.01% 3% 2% 0.01% 3% 3% 6 2% 0.01% 3% 7 2% 0.01% 3% 8 1% 0.01% 3% 9 1% 0.01% 3% 3% Results There was a transient rise in body temperature on Day 1. Otherwise no sheep displayed any abnormal clinical signs associated with the specific skin treatments. No sign of pain or discomfort was noted. The results are summarised in Tables 6 to 8 below.

TABLE 6 Area of Necrosis Formulation Day 20 Meaii Area (cm 2 iDay 35Mean Area.(cm 2 1 15.67 a 10.17a 2 13.67 a 13.33 a 3 12.92 a 9.67 a 4 13.42 a 12.67 a 13.04 a 9.50 a 6 13.96a 9.83 a 7 13.75 a 9.33 a 8 14.00 a 7.83 a 9 14.75 a 9.17 a *eans within the same coluim- ith different sierscripts atr sigfcapty diffdrent using One-Way AQV analysis-at p<O.5 'y 1 TABLE 7- Summary of component effects on area of necrosis (Day Cetrimide .Area cmr) Ethano ea (cm 2 PVP Area (cm 2 1 14.38 a 0 14.14a 0 13.67a 2 13.58 a 1.5 13.75 a 3 13.94 a 4 13.92 a 3 13.57 a *Means within the same column with different superscripts are significantly different using One-Way AOV analysis at p<0.05 TABLE 8 Summary of component effects on area of necrosis (Day Cetrimide:% Area (cm 2 Ethaioli% Area (cm 2 PVP'% :Area c) 1 8.50a 0 10.77 a 0 13.33 a 2 9.56 a 1.5 9.33 a 3 9.77 a 4 11.46 a 3 9.44 a_ 'Means within the,: sa" ,c61umnrwithK iffesqg ehtniteriff using OneWayAV yP V. 4 s All the formulations caused the contraction of treatment site width and height over the trial period, resulting in a maximum contraction at between Day 22 and Treatment site assessment of bruising, swelling, skin discolouration and scab formation was similar between groups. There were no statistically significant differences between the area of necrosis in any of the treatments, treatment components to or treatment site at either Day 20 or Day Histopathology Scores of from 0-5 were allocated for necrosis, inflammatory cell infiltration, fibroplasia, sloughing of the necrotic dermis and re-epithelialisation of the dermis with 0 representing no change and 5 the most severe change seen in the study. The results is are presented in Table 9.

1. Despite within group variation, there were considerable differences in the effects of the different treatments. Thus necrosis and sloughing of the necrotic dermis were most severe after treatments 1 and 3, and least severe after treatments 6, 8 and 9.

2. Fibroplasia, with dermal necrosis and sloughing, was most pronounced after treatments 1 and 3 and least severe after treatments 6, 8 and 9.

3. In some of the 35-day treatment groups there was progression to complete sloughing of the necrotic dermis and even complete re-epithelialisation.

4. In some cases, there was sloughing of the treated skin, pronounced fibroplasia of the underlying tissue and re-growth of the epidermis, leading to the formation of scar tissue, covered by epidermis which closely resembled the appearance of skin sections from sites of the Mules operation.

TABLE 9 Mean Scores for Histopathological Changes in Skin Specimens FORMULATION DAY SLOUGH RE. SUPERFICIAL DEEP DEEPER DERMIS DERMIS/ TISSUES I IHYPODERMIS NEC-' FIB NEC FIB NEC FIB 1 20 4.2 1.3 4.3 N/A 1.7 4.7 <1 3.1 4.8 2.5 5.0 N/A 0 4.2 <1 2 20 3.4 1.5 4.1 N/A 1.1 3.9 <1 2.9 3.3 1.3 3.7 <1 0 3.8 <1 2.2 3 20 4.1 1.5 4.6 N/A 2.4 3.4 1.4 3.1 4.2 3.2 4.2* <1 <1 4.5 0 1.3 4 20 3.3 2.1 3.8 N/A <1 4.0 <1 3.1 4.2 3.7 4.2* <1 <1 4.2 <1 2.3 20 3.1 2.3 4.1 N/A 1.0 4.2 <1 2.9 1.8 2.5 1.7 1.2 0 2.7 <1 2.7 6 20 2.4 1.3 3.5 N/A <1 3.6 <1 1.9 3.2 2.2 2.5 1.7 0 3.7 0 1.7 7 20 3.2 2.0 4.0 N/A 1.0 3.4 <1 1.9 3.0 2.8 3.3* 1.0 <1 4.3 0 2.7 8 20 2.1 1.9 2.8 N/A <1 3.5 <1 2.0 3.0 2.2* 2.2 0 3.7 0 9 20 <1 1.1 2.2 N/A 0 2.7 <1 2.3 1.7 2.5 <1 0 3.0 0 <1 One or more specimens with the superficial dermis completely sloughed off.

N/A Because of the presence of complete necrosis in many specimens, no mean score for fibroplasia was calculated.

Conclusions 3% alcohol was associated with necrosis that was undesirable. As the concentration of cetrimide declined the degree of pathology declined and 1% cetrimide caused inadequate skin damage. 4% cetrimide PVP and 4% cetrimide PVP phenylephrine gave a suitable reaction as did 2% cetrimide 1.5% ethanol PVP phenylephrine. This finding indicated a synergistic action between cetrimide and ethanol to cause necrosis and subsequent wound healing in a shorter time frame than the case where 4% cetrimide is used. These combinations were selected for testing on the breech to ensure efficacy as a mulesing replacement and to confirm that breech and flank skin reacted in a similar way to the treatments.

4. Evaluation of the tissue healing, site behaviour and local tissue reaction following chemical intra-dermal treatment to the breech of unmulsed Merino sheep This study documented wound healing parameters following chemical mulesing by a skilled operator delivering one of the three formulations given below in Table using a needleless injector to 3 groups of 8 unmulesed Merino ewe weaners. Sheep and the treated areas were carefully observed 4 hours thence 1, 2 and 4 days thence weekly until the 35 days. Before treatment and 21 and 35 days after treatment the dimensions of the perineal bare area were measured. 35 days after treatment the trial animals were euthanized, the skin removed from the breech and tail regions and the size of the to affected area measured. Representative sections were collected for histological examination.

TABLE 10 Treatment regime 1 4% cetrimide, 3% PVP 2 4% cetrimide, 0.01% phenylephrine, 3% PVP 3 2% cetrimide, 0.01% phenylephrine, 3% PVP, 1.5% ethanol 10 days after treatment 3 of 8 sheep treated with treatment 1 were stiff in the hind legs for 1 day. Otherwise there were no adverse effects. All three treatment were effective and gave a result similar to surgical mulesing. The wounds were closest to resolved in sheep treated with treatment 3 (see Table 11 below).

TABLE 11 Mean areas of affected skin at day Treatment :Breech LHS Breech RHS 1 14.2 17.3 2 18.3 18.8 3 14.2 13.9

Claims (47)

1. A method of administering a skin necrosis composition comprising a ==quaternary ammonium compound and a non-newtonian viscosity imparting agent to an animal, the method comprising administering the composition intradermally to an area of the skin of the animal.

2. A method for reducing the incidence of fly strike in an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition comprising a quaternary ammonium compound and a non-newtonian 4N viscosity imparting agent to an area of the skin of the animal. S lo 3. A method of mulesing the breech region of an animal, the method CN comprising intradermally administering a skin necrosis composition comprising a quaternary ammonium compound and a non-newtonian viscosity imparting agent to the breech region of the animal.

4. The method of any one of claims 1 to 3 wherein the non-newtonian is viscosity imparting agent is a cellulose derivative or polyvinylpyrrolidone. The method of any one of claims 1 to 4, wherein the skin necrosis composition is delivered with, or further comprises, one or more vasoconstrictors.

6. The method of any one of claims 1 to 5, wherein the skin necrosis composition is delivered with, or further comprises, one or more anaesthetics.

7. A method of administering a skin necrosis composition to an animal, the method comprising administering the composition intradermally to an area of the skin of the animal, wherein the composition comprises one or more quaternary ammonium compounds, and a non-newtonian viscosity imparting agent, and wherein the composition is optionally delivered with, or further comprises, one or more vasoconstrictors, and wherein the composition is optionally delivered with or further comprises one or more anaesthetics.

8. A method for reducing the incidence of fly strike in an animal, the method comprising the step of intradermally administering an effective amount of a skin necrosis composition to an area of the skin of the animal, wherein the composition comprises one or more quaternary ammonium compounds, and a non-newtonian viscosity imparting agent, and wherein the composition is optionally delivered with, or further comprises one or more vasoconstrictors, and wherein the composition is optionally delivered with or further comprises one or more anaesthetics.

9. A method of mulesing the breech region of an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition to the breech region of the animal, wherein the composition comprises one 859343 I or more quaternary ammonium compounds, and a non-newtonian viscosity imparting agent, and wherein the composition is optionally delivered with, or comprises one or more vasoconstrictors, and wherein the composition is optionally delivered with or comprises one or more anaesthetics.

10. The method of any one of claims 7 to 9 wherein the non-newtonian t viscosity imparting agent is a cellulose derivative or polyvinylpyrrolidone.

11. The method of any one claims 1 to 10 wherein the quatemary 0 ammonium compound is selected from the group consisting of: cetrimide, C cetylpyridinium chloride, benzalkonium chloride, dioctyldimethylammonium chloride, O 10 octyldecyldimethylammonium chloride, didecyldimethylammonium chloride, (C hexadecyltrimethylammonium bromide, methylbenzethonium chloride, Alcian Blue 8GX, polypropoxy quaternary ammonium chlorides (Emcol® CC9, Emcol® CC42), polypropoxy quaternary ammonium phosphate (Emcol® CC57), trimethyl C8C10 quaternary ammonium chloride (Adogen® 464), polyquaternary ammonium chloride (Mirapol® WT), Amine hydrochloride (guanidine hydrochloride), Imine compound (polyethyleneimine), sodium alginate, octylphenoxypolyethoxyethanol (Triton® X-100), choline chloride, acetylcholine chloride, carboxymethyltrimethylammonium chloride, tetraethylammonium chloride, tetrabutylammonium bromide, tetraheptylammonium bromide, phenyltriethylammonium bromide, phenyltriethylammonium iodide, tricaprylmethylammonium chloride, trioctylpropylammonium bromide, nonyltrimethylammonium bromide, octyldecyldimethylammonium chloride (Bardac 2080), didecyldimethylammonium chloride (Bardac 2280, Quatramine 2-10/80), didecylmethylpropylammonium chloride, didecyldipropylammonium chloride, dodecyltrimethylammonium chloride, decamethonium bromide, hexamethonium bromide. laurtrimonium bromide, myristyltrimethylammonium bromide, palmityltrimethyl ammonium bromide, or any combination thereof.

12. The method of claim 11, wherein the quaternary ammonium compound is selected from the group consisting of: alkyl trimethylammonium bromides, cetrimide, cetylpyridinium chloride, benzalkonium chloride, hexadecyltrimethylammonium bromide, cetyltrimethylammonium bromide and cetrimonium bromide.

13. The method of claim 11, wherein the quaternary ammonium compound is selected from the group consisting of cetrimide, cetylpyridinium chloride and benzalkonium chloride.

14. The method of any one of claims 1 to 13, wherein the amount of the quaternary ammonium compound present in the composition is between about 2% and about 8% 859343 1 The method of any one of claims 1 to 14, wherein the non-newtonian viscosity imparting agent is present in the composition in an amount between about 1% Sand about 10%

16. The method of any one of claims 1 to 15, wherein the composition is s delivered with, or further comprises an alcohol.

17. The method of claim 16, wherein the alcohol is a Ci-Clo alcohol.

18. The method of claim 17, wherein the alcohol is ethanol.

19. The method of any one of claims 16 to 18, wherein the amount of 00 N alcohol present in the composition is between about 0.5% and about 3% S 10 20. The method of any one of claims 5 to 19, wherein the vasoconstrictor is CN selected from a directly acting vasoconstrictor, an indirectly acting vasoconstrictor, a mixed acting vasoconstrictor, or a mixture thereof.

21. The method of claim 20, wherein the directly acting vasoconstrictor is selected from the group consisting of epinephrine and phenylephrine.

22. The method of claim 20, wherein the indirectly acting vasoconstrictor is selected from the group consisting of: amphetamine and methamphetamine.

23. The method of claim 20, wherein the mixed acting vasoconstrictor is selected from the group consisting of: metraminol and ephedrine.

24. The method of any one of claims 5 to 23, wherein the vasoconstrictor is present in the composition in an amount between about 0.0005% and 1% The method of any one of claims 6 to 24, wherein the anaesthetic is selected from the group consisting of: procaine, prilocaine, mepivacaine, lignocaine and bupivacaine.

26. The method of any one of claims 1, 2, 3, 7, 8, or 9, wherein the composition is selected from the group consisting of: a composition comprising a quaternary ammonium compound, a vasoconstrictor and a non-newtonian viscosity imparting agent; (ii) a composition comprising a quaternary ammonium compound, a vasoconstrictor, a non-newtonian viscosity imparting agent and an alcohol, and (iv) a composition comprising a quaternary ammonium compound, an alcohol and a non-newtonian viscosity imparting agent

27. The method of claim 26, wherein the composition is selected from the group consisting of: a composition comprising cetrimide and polyvinylpyrrolidone; (ii) a composition comprising cetrimide, phenylephrine and polyvinylpyrrolidone; 85)343 I (iii) a composition comprising cetrimide, phenylephrine, polyvinylpyrrolidone; and ethanol, and S(iv) a composition comprising cetrimide, ethanol and polyvinylpyrrolidine

28. The method of claim 27, wherein the amount of cetrimide present in the compositions is between 2.0% and 8% the amount of phenylephrine present in the compositions is between 0.0005% and 1% the amount of polyvinylpyrrolidone present in the compositions is between 1% and 10% and the amount of ethanol present in the compositions is between 0.5 and 2.5% 00 Ni 29. The method of any one of claims 1 to 28, wherein the skin necrosis O 10 composition is delivered with, or comprises a pharmaceutically acceptable carrier, Ni diluent, dye, adjuvant and/or excipient The method of any one of claims 1 to 29, wherein the skin necrosis composition is administered by high pressure injection.

31. The method of any one of claims 1 to 30, wherein the animal is a sheep.

32. The method of any one of claims 1 to 31, wherein multiple simultaneous doses of the skin necrosis composition are administered.

33. The method of any one of claims 1, 2, 3, 7, 8, or 9, wherein the skin necrosis composition comprises: Cetrimide 2 4% (w/w) Phenylephrine 0.005 to 0.05% (w/w) Polyvinylpyrrolidone 2.5 to 3.5% (w/w) Ethanol 1 to 2% (w/v) Pharmaceutically acceptable carriers, diluents, dyes, adjuvants and/or excipients (up to 100%);

34. A skin necrosis composition, the composition comprising a synergistic combination of an effective amount of an alcohol and an effective amount of one or more skin necrotizing agents. The skin necrosis composition of 34, wherein the skin necrotizing agent is a quaternary ammonium compound.

36. The skin necrosis composition of claim 35 wherein the quaternary ammonium compound is selected from the group consisting of: cetrimide, cetylpyridinium chloride, benzalkonium chloride, dioctyldimethylammonium chloride, octyldecyldimethylammonium chloride, didecyldimethylammonium chloride, hexadecyltrimethylammonium bromide, methylbenzethonium chloride, Alcian Blue 8GX, polypropoxy quaternary ammonium chlorides (Emcol® CC9, Emcol® CC42), polypropoxy quaternary ammonium phosphate (Emcol® CC57), trimethyl C8C10 851343 1 quaternary ammonium chloride (Adogen® 464), polyquatemary ammonium chloride (Mirapol® WT), Amine hydrochloride (guanidine hydrochloride), Imine compound S(polyethyleneimine), sodium alginate, octylphenoxypolyethoxyethanol (Triton® X-100), choline chloride, acetylcholine chloride, carboxymethyltrimethylammonium chloride, tetraethylammonium chloride, tetrabutylammonium bromide, tetraheptylammonium bromide, phenyltriethylammonium bromide, phenyltriethylammonium iodide, F tricaprylmethylammonium chloride, trioctylpropylammonium bromide, nonyltrimethylammonium bromide, octyldecyldimethylammonium chloride (Bardac CN 2080), didecyldimethylammonium chloride (Bardac 2280, Quatramine 2-10/80), S 0o didecylmethylpropylammonium chloride, didecyldipropylammonium chloride, CN dodecyltrimethylammonium chloride, decamethonium bromide, hexamethonium bromide, laurtrimonium bromide, myristyltrimethylammonium bromide, palmityltrimethyl ammonium bromide and any combination thereof.

37. The skin necrosis composition of claim 36, wherein the quaternary ammonium compound is selected from the group consisting of: alkyl trimethylammonium bromides, cetrimide, cetylpyridinium chloride, benzalkonium chloride, hexadecyltrimethylammonium bromide, cetyltrimethylammonium bromide and cetrimonium bromide

38. The skin necrosis composition of claim 37, wherein the quaternary ammonium compound is selected from the group consisting of cetrimide, benzalkonium chloride and cetyl pyridinium chloride.

39. The skin necrosis composition of any one of claims 34 to 38, wherein the amount of the quaternary ammonium compound present in the composition is between about 1.5% and about 2.5%

40. The skin necrosis composition of any one of claims 34 to 39, wherein the amount of alcohol present in the composition is between about 1.0 and 2.0%

41. The skin necrosis composition of any one of claims 34 to 40, wherein the alcohol is a Ci-Cio alcohol.

42. The skin necrosis composition of any one of claims 34 to 41, wherein the alcohol is ethanol.

43. The skin necrosis composition of any one of claims 34 to 42, wherein the composition comprises a viscosity imparting agent.

44. The skin necrosis composition of claim 43, wherein the viscosity imparting agent is polyvinylpyrrolidone. 859343 I The skin necrosis composition of claim 43 or claim 44, wherein the viscosity imparting agent is present in the composition in an amount between about 1% Sand about 10%

46. The skin necrosis composition of any one of claims 34 to 45, wherein the skin necrosis composition comprises one or more vasoconstrictors.

47. The method of claim 46, wherein the vasoconstrictor is present in the composition in an amount between about 0.0005% and 1%

48. The skin necrosis composition of any one of claims 34 to 47, wherein 00 N the skin necrosis composition comprises one or more anaesthetics. S lo 49. The skin necrosis composition of any one of claims 34 to 48, wherein CN the composition is selected from the group consisting of: a composition comprising a quaternary ammonium compound, a vasoconstrictor, a viscosity imparting agent and an alcohol; (ii) a composition comprising a quaternary ammonium compound, an alcohol and a viscosity imparting agent; (iii) a composition comprising a quaternary ammonium compound a vasoconstrictor and an alcohol. The method of claim 49, wherein the composition is selected from the group consisting of: a composition comprising cetrimide, phenylephrine, polyvinylpyrrolidone; and ethanol; (ii) a composition comprising cetrimide, ethanol and polyvinylpyrrolidine; and (iii) a composition comprising cetrimide, phenylephrine, and ethanol;

51. The skin necrosis composition of claim 50, wherein the amount of cetrimide present in the compositions is between 2.0% and 8% the amount of phenylephrine present in the compositions is between 0.0005% and 1% the amount of polyvinylpyrrolidone present in the compositions is between 1% and and the amount of ethanol present in the compositions is between 0.5 and

52. A method of administering a skin necrosis composition to an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition according to any one of claims 34 to 51, to an area of the skin of the animal.

53. A method for reducing the incidence of fly strike in an animal, the method comprising of intradermally administering an effective amount of a skin necrosis composition according to any one of claims 34 to 51, to an area of the skin of the animal. 85)343 I

54. A method of mulesing the breech region of an animal, the method comprising intradermally administering an effective amount of a skin necrosis composition of any one of claims 34 to 51, to the breech region of the animal. The method of any one of claims 52 to 54, wherein the skin necrosis composition is delivered with, or comprises a pharmaceutically acceptable acceptable Scarrier, diluent, dye, adjuvant and/or excipient S56. The method of any one of claims 52 to 55, wherein the skin necrosis composition is administered by high pressure injection. 00 N57. The method of any one of claims 52 to 56, wherein the animal is a S 10 sheep. S58. The method of any one of claims 52 to 57, wherein multiple simultaneous doses of the skin necrosis composition are administered.

59. The method claim 32 or claim 58, wherein multiple simultaneous doses of the skin necrosis composition are administered in a grid pattern at a separation of Is between about 0.1 cm and 2.0 cm. A skin necrosis composition substantially as hereinbefore described with reference to examples one and four.

61. A method of administering a skin necrosis composition to an animal, the method comprising intradermally administering an effective amount of the skin necrosis composition according to claim

62. A method for reducing the incidence of fly strike in an animal, the method comprising intradermally administering an effective amount of the skin necrosis composition according to claim

63. A method of mulesing the breech region of an animal, the method comprising intradermally administering an effective amount of the skin necrosis composition according to claim Dated 26 July, 2007 Australian Wool Innovation Limited Patent Attorneys for the Applicant/Nominated Person SPRUSON FERGUSON H59343 I