WO2006129162A2

WO2006129162A2 - Vaccine compositions and methods for the treatment of urinary incontinence

Info

Publication number: WO2006129162A2
Application number: PCT/IB2006/001398
Authority: WO
Inventors: Syed Ziauddin Hashmi; Bradley John Nettleton; John Walker
Original assignee: Pfizer Products Inc.
Priority date: 2005-06-01
Filing date: 2006-05-22
Publication date: 2006-12-07
Also published as: TW200716164A; JP2008545738A; RU2007143992A; ZA200710213B; CN101189026A; AU2006253914A1; MX2007015151A; EP1890725A2; JP2009137972A; NO20075923L; BRPI0611390A2; UY29574A1; WO2006129162A3; CA2609244A1; KR20080007648A; AR054370A1

Abstract

The present invention relates to the use of compositions comprising an anti-LHRH vaccine for the control or treatment of urinary incontinence. The present invention further relates to methods of treating or controlling urinary incontinence in mammals, and in particular, in those that have been surgically neutered, e.g., spayed female dogs.

Description

VACCINE COMPOSITIONS AND METHODS

FOR THE TREATMENT OF URINARY INCONTINENCE

Field of the Invention

Background of the Invention Urinary incontinence is the involuntary loss or passing of urine consequent to bladder or urethral sphincter dysfunction. During the normal storage of urine, as the bladder fills, the urethral sphincter closes tightly to prevent leaking. Then, when urine is to be voided, the urethral sphincter relaxes and allows urine to pass through the urethra. Urinary incontinence is often the result of loss of urethral sphincter tone, which lessens its ability to tighten. In such cases, urinary incontinence occurs in an uncontrolled manner whenever the pressure within the bladder exceeds the pressure required to close the urethra. This problem is often more noticeable when the mammal is relaxed, such as during deep sleep, or when pressure on the bladder is momentarily increased, such as when the mammal coughs or sneezes.

Those familiar with the condition will appreciate that urinary incontinence is prevalent among adult women. Because of the social stigma associated with urinary incontinence, many sufferers do not even report the problem to a health care provider. As a result, this medical problem is vastly under-diagnosed and under-reported.

Urinary incontinence is also quite common in dogs. Usually, it occurs in female dogs, and most often in the spayed female dog. Male dogs are prone to this condition as well, but less commonly so than their female counterpart. Previously, it has been explained that the underlying mechanism of incontinence in spayed or neutered dogs is related to low post-surgical estrogen or testosterone concentrations following the removal of the ovaries or testes. In the case of the female dog, estrogen hormone is believed to be responsible for providing tone to the urethral sphincter muscle. Following spaying, estrogen hormone is produced in significantly lower amounts than it was prior to the removal of the ovaries. While the adrenal glands produce small amounts of estrogen hormone, it was believed that the levels produced are insufficient to provide sphincter tone. In such cases, incontinence can occur and some form of treatment is then required to support sphincter tone. In the same manner, male dogs can develop urinary incontinence following neutering, as the testes, the primary testosterone source, are removed.

Treatment for urinary incontinence has traditionally been aimed at improving the urethral closure pressure. In this regard, substances that increase the resistance of the urethra or cervix of the bladder have been prescribed. In particular, these medicaments include alpha-adrenergic substances, such as ephedrine or phenylpropanolamine ("PPA"). Treatment with the above typically involves a long-term use of the medicaments and has been shown to cause severe cardiac electrophysiological side effects in humans. These adverse side effects can include, but are not limited to, ventricular fibrillation and cardiac arrhythmias. Such potentially lethal cardiac side effects have led to the withdrawal of several medications for humans from the market, including PPA, which has recently been recalled by the U.S. Food and Drug Administration ("FDA"). Alpha-adrβnβrgic substances are also contraiπdlcated in case of glaucoma and progressive nephropathy.

As an alternative therapy, forms of estrogen or oestradiol, e.g., diethylstilbestrol, (or testosterone in the case of male dogs) replacement therapy can be used, which improve the responsiveness of the catecholamine receptors of the urethra. However, estrogen therapy is widely considered less safe than the use of alpha-adrenergic substances since it can lead to bone marrow suppression in dogs, which can be fatal.

More recently, it has been hypothesized that increased serum concentrations of the gonadotropins Follicle Stimulating Hormone ("FSH") and Luteϊnisiπg Hormone ("U-T), rather than low levels of estrogen per se, are the cause of urinary incontinence^' in adult women and in spayed female dogs. Reichler, I.M. et al., The Effect of GnRH Analogs on Urinary Incontinence after Ablation of the Ovaries in Dogs, Theriogenology, 60: pp. 1207-1216 (2003); United States Patent Application Publication No. US 2005/0043342 (collectively, hereinafter referred to as "Reichler et al.^*). Reportedly, the pituitary gland releases LH and FSH in the system in response to the production of Luteinizing Hormone Releasing Hormone ("LHRH", also commonly referred to as Gonadotropin Releasing Hormone, or "GnRH" or Gonadotrophs Releaseing Factor "GnRF"). LHRH is released - from the hypothalamus into the bloodstream in a pulsatile manner in response to a range of factors, and its control is generally regarded as complex. Its release is positively influenced by hypothalamic neuronal activity and negatively controlled by low levels of estrogen and other gonadal steroids and hormones such as inhibin A and B. Released LHRH travels via the blood to the pituitary. The receptor cells in the anterior pituitary gland respond to the rise in LHRH by releasing the gonadotropins, FSH and LH. The relative levels of FSH and LH that vary over the course of the oestrus cycle, in turn stimulates the production of estrogen by the ovaries. In the spayed female dog, following removal of the ovaries (i.e., the primary estrogen source), estrogen is measurable only in very low serum concentrations. Other hormones that usually originate in the gonads such as the inhibins that are secreted in the ovary by granulose cells selectively inhibit pituitary FSH sectretion. Consequently, as there is no longer a feedback mechanism, FSH and LH are secreted unhindered, resulting in increased concentrations of the two gonadotropins. Reichler et al. contend that, in both post-menopausal women and spayed female dogs suffering from urinary incontinence, the levels of FSH and LH are elevated above normal levels. Accordingly, Reichler et al. have proposed to use LHRH analogues, such as agonists, antagonists, or antibodies in medicaments to control or treat urinary incontinence.

There are, however, distinct disadvantages associated with this approach. In particular, typically high doses of LHRH antagonists or agonists are required to achieve the desired anttgonadotropic effects. Moreover, while the effects of using such analogues are immediate, they are of short duration and therefore, require frequent administration. In addition, in some cases, large amounts of these analogues may cause undesirable or adverse side effects. They are also very expensive since their use, and hence their availability, is usually limited to the treatment of hormone dependant tumors. Likewise, LHRH antibodies, in addition to being quite costly, have the disadvantage of having rather short half-lives, which result in short durations of treatment and the potential requirement for multiple doses. Thus, the necessary frequent and continuous administration of LHRH antibodies using this approach would be both costly and inconvenient to dog owners. Accordingly, LHRH analogs are not of practical use for veterinary applications as such.

Thus, despite the reports of some of the currently available urinary incontinence treatments, complete responses to these treatments are either infrequently observed, come with serious side effects, lack a long-lasting effect or are too costly or Inconvenient to maintain. The development of new therapeutic regimens, particularly those capable of long-lasting control and treatment of urinary incontinence in a cost-effective manner, while reducing the potentially lethal side effects of current therapeutics (such as cardiac arrhythmia or bone marrow suppression), remains necessary. The present invention meets these needs. . Summary of tha invention

An alternative and/or additional approach to urinary incontinence therapy heretofore unrecognized- is to target the immune system through the use of vaccine compositions. A potential benefit of active immunotherapy is to provide improved efficacy by enhancing the subject's own immune response to the effective and long-lasting control of unrestricted production and release of gonadotropins, FSH and LH. Accordingly, the present invention relates to anti-LHRH vaccines and the use thereof for the treatment or control of urinary incontinence in mammals. In particular, the present invention provides a method for the treatment of urinary incontinence In a mammal in need of such treatment, wherein the method comprises administering to the mammal a therapeutically effective amount of an anti-LHRH vaccine. In one aspect of the invention, the mammal in need of treatment for urinary incontinence is a female; preferably a female having elevated serum concentrations of luteinizing hormone or follicle stimulating hormone. In another aspect, the mammal in need of treatment for urinary incontinence is a female dog. In a particularly preferred embodiment, the mammal is one that has been surgically neutered, e.g., a spayed female dog.

In one aspect of the present invention, the anti-LHRH vaccine comprises an immunogenic conjugate between a carrier and LHRH. Preferably, the carrier comprises a protein selected torn the large range of known carrier proteins including, but not limited to, bacterial toxoids. Most preferably, the carrier comprises diphtheria toxoid. In another aspect the carrier may be a synthetic peptide comprising a T-cell helper epitope. In either aspect, an effective form of the LHRH peptide is joined to the carrier, either through conjugation, or during synthesis of a contiguous T-helper -LHRH peptide. In another aspect of the invention, the anti-LHRH vaccine further comprises an adjuvant. Preferably, the adjuvant comprises an ionic polysaccharide; most preferably, diethyl aminoethyl dextran ("DEAE dextran"). In a particularly preferred embodiment, the adjuvant is a DEAE dextran in combination with an immunoεtimulating complex that comprises a saponin and cholesterol.

In yet another aspect, the invention relates to a method for treating or controlling urinary incontinence in a mammal comprising- administering to the mammal a composition comprising a therapeutically effective amount of an immunogenic conjugate between a carrier and LHRH. For a protein-LHRH peptide conjugate based vaccine, the effective amount is in the range of about 100 to about 300ug, and for the T-helper-LHRH peptide based vaccine in the range of about 20 to about 200ug.

Detailed pescriPtlon Of The Invention The invention relates to immunogenic compositions and vaccines that are suitable for eliciting an immune response against Luteinizing Hormone Releasing Hormone ("LHRH", or Gonadotropin Releasing Hormone, or "GnRH"). The invention further relates to the use of such an anti-LHRH vaccine or physico- chemically stable composition in a method of immunizing a mammal against LHRH to control or treat urinary incontinence.

The term "anti-LHRH vaccine" Includes any vaccine, composition, or pharmaceutical preparation that is capable of eliciting an immune response to LHRH in a host. More specifically, the anti-LHRH vaccines of the present invention comprise at least one LHRH antigen or immunogen in a pharmaceutically- accepiable carrier useful for inducing an immune response in said host. By way of example only, anti-LHRH vaccines that can be used in connection with the present invention, and methods of producing them, include, but are not limited to, those that are described in the International Application No. PCT/AU99/01167, published on January 21, 1999 as WO 99/02180, International Application No. PCT/AU96/00532, published , on July 20, 2000 as WO 99/41720, and United States Patent No. 6,685,947, which. are incorporated by reference herein.

The term "LHRH" should be read as including reference to all forms of LHRH and derivatives thereof, including, but not limited to, all peptides having LHRH immunogenicity. The term "derivatives" include fragments, parts, portions, chemical equivalents, mutants, homologs and analogs from natural, synthetic or recombinant sources, including fusion proteins. LHRH can be obtained from any suitable source and methods for producing, isolating, and purifying native and synthetic LHRH are well known in the art. In a preferred embodiment, the LHRH comprises an LHRH C-terminal fragment of at least five amino acids. Because LHRH itself is too small to be immunogenic, the LHRH used in the context of the present invention must be brought into an immunogenic form. It is well known to those of ordinary skill in the art that there are different methods of producing an immunogenic form of a substance that is, itself, not immunogenic. One suitable method is to couple LHRH to an immunogenic carrier. Accordingly, in the context of the present invention, LHRH can be administered after conjugation to an immunogenic carrier to a mammal as an antigen to induce the formation of host antibodies to LHRH. The carrier (unctions to stimulate T-helper cells that are required for an effective antibody response. Thus, the carrier may comprise small peptides of at least 9 amino acids In length that form a T-ceH helper epitope. Such T-helper epitopes may be derived from Canine Distemper Virus (CDV), which are particularly suited to application in the dog since most dogs are already vaccinated against CDV. The antibodies induced by such vaccines will subsequently act against the body's own LHRH, thereby providing a long term antigonadotropic effect. In a preferred embodiment, LHRH is administered via vaccine after conjugation to an immunogenic carrier to a mammal to stimulate the immune system to produce. anti-LHRH antibodies that react with LHRH to effectively reduce its concentration in the body. The effects caused by the presence of LHRH, including high serum concentrations of FSH arid LH, which are believed to cause urinary incontinence, are thus reduced or eliminated. i

The immunogenic carriers used in the preparation of the LHRH conjugates described above include any substances that are capable of increasing the immunogenicity of LHRH. The immunogenic carriers used in the conjugates of the present invention can be natural or synthetic. In one aspect, the immunogenic carrier is a protein. Any one of a variety of immunogenic carrier proteins that are well known to those of ordinary skill in the art may be used in the conjugate vaccine of the present invention. Suitable classes of proteins include pili, outer membrane proteins and excreted toxins of pathogenic bacteria, nontoxic or "toxoid" forms of such excreted toxins, nontoxic proteins antigβnically similar to bacterial toxins and other proteins. Nonllmiting examples of bacterial toxoids contemplated for use in the present invention include tetanus toxin/tbxoid, diphtheria toxin/toxoid, detoxified P. aeruginosa toxin A, cholera toxin/toxoid, pertussis toxin/toxoid and Clostridium perfrlngens exotoxins/toxoid. The toxoid forms of these bacterial 5 toxins are preferred. Most preferably, the immunogenic carrier used in the LHRH conjugates of the present invention is diphtheria toxoid. Reference to diphtheria toxoid should be understood to include all forms of diphtheria toxoid and derivatives thereof. The term "derivatives" has the same meaning as hereinbefore defined. Derivatives may include, for example, molecules comprising the diphtheria toxoid T cell epitopes or T cell epitopes in isolation.

10 In another aspect of the present invention, the carrier may be a synthetic peptide comprising at least one T-cell helper epitope. The epitopes may be singular or be linked together to form a single polypeptide. It will be understood that where the epitopes are' linked together In a single polypeptide the epitopes may be contiguous or spaced apart by additional amino acids that are not themselves part of the T helper cell -epitopes. In one embodiment, the T-cell helper epitope is contained within a peptides

15 sequence that is selected from among those described in United States Patent No, 6,685,947, which is incorporated herein by reference. In a preferred embodiment, the synthetic peptide vaccine further comprises at least one B cell epitope and/or at least one CTL epitope. In a most preferred embodiment, the synthetic peptide vaccine comprises from about 3 to about 10 T-cell helper epitopes, each in linear sequence with a 4-10 form of LHRH, with the total peptide content being about 120 μg peptide/dose in

20 combination with about 80 μg of an adjuvant. Preferably, the adjuvant comprises an immunostimulating complex.

Suitable methods of coupling LHRH to an immunogenic carrier are well known to those of ordinary skill in the art. Without intending to limit the invention to any one method, LHRH may, for example, be coupled to a carrier in the manner described in Ladd et al., American J. Reproductive Immunology 22:56-63

25. (1990) or in Bonneau et al., J. Animal Science 72:14-20 (1994). The resulting conjugate can be processed to remove any remaining unbound peptide or other byproducts of conjugation. Such processing may be achieved by conventional dialysis or by ultrafiltration.

The vaccines of the present invention may be combined with or may further include an adjuvant Adjuvants are included or added to vaccines and other immunogenic formulations to increase and in

30 some cases direct the immune response. Those of ordinary skill in the art will appreciate that any well- known standard adjuvant may be used in the present invention. Suitable adjuvants for use in the vaccines of the present invention include, but are no limited to, insoluble aluminum salts, particularly the hydroxide and phosphate forms (collectively, "alum"), murabutide, Freund's incomplete adjuvant, Freund's complete adjuvant, ionic polysaccharides, such as a diethyl-aminoethyl ("DEiAE") group, saponins,

35 immunostimulating complexes, and combinations thereof. In a preferred embodiment, adjuvants, which elicit no or only mild side reactions, are used. Other suitable adjuvants that can be used in connection with the present Invention include, but are not limited to, those that are described in International Application No. PCT/AU99/01167, published on January 21, 1999 as WO 99/02180, and International Application No. PCT/AU98/00532, published on July 20, 2000 as WO 99/41720, which are incorporated by reference herein. 0 In a preferred embodiment, the adjuvant is a DEAE-dextran. In a more preferred embodiment, the adjuvant is a DEAE dextran in combination with an immunostimulating complex comprising a saponin and cholesterol.

Without limiting the present invention to any one theory or mode of action, the administration of an effective amount of the anti-LHRH vaccine of the present invention induces an immune response to LHRH, and thereby prevents the release of the hormones LH and FSH from the anterior pituitary gland. Preferably, the amount of said antr-LHRH vaccine Is effective to improve the side-effects associated with ovarectomy or symptoms associated with reproductive senescence, in particular, urinary incontinence, by lowering the serum concentration of circulating FSH and/or LH In a female mammal to whom the vaccine Is administered. It should be understood that efficacy is a functional measure and is not defined by reference to anti-LHRH antibody litre alone since the presence of circulating antibody atone Is not necessarily indicative of the capacity of said circulating antibody to control urinary incontinence. Thus, as with all immunogenic compositions for eliciting antibodies, the immunologically effective amounts of the vaccines of the present invention must be determined empirically. Factors to be considered include the choice of adjuvant, the immunogenic molecule to which LHRH is coupled, the route of administration and the number of doses to be administered. Such factors are known in the vaccine art, and one of ordinary skill in the art would be able to make such determinations without undue experimentation.

Accordingly, by the methods of the present invention, the side effects of ovarectomy or symptoms associated with reproductive senescence, in particular urinary incontinence, in female mammals, such as spayed female dogs, can be treated or controlled. In a preferred embodiment, the present invention relates to a method of treating or controlling urinary incontinence in a mammal, said method comprising administering to said mammal an effective amount of an anti-LHRH vaccine. Preferably, the anti-LHRH vaccine is in the form of an LHRH-immunogenic carrier conjugate. In a more preferred embodiment, the conjugate comprises an LHRH-diphtheria toxoid conjugate. In an alternative embodiment, the anti-LHRH vaccine is in the form of a synthetic peptide that comprises a T-cell helper epitope. Preferably, the synthetic peptide vaccine comprises from about 3 to about 10 T-cell helper epitopes, each in linear sequence with a 4-10 form of LHRH, with the total peptide content being about 120 μg peptide/dose in combination with about 80 μg of an immunostimulating complex. The preferred mammal to be treated in accordance with the vaccine of the present invention is a female; more preferably a female having elevated serum levels of FSH or LH or both. In a most preferred embodiment, the mammal is a female dog, and in particular, a female dog that has been surgically neutered or spayed. In another embodiment, the invention provides a method of preventing the onset of urinary incontinence in a spayed dog, by administering to the dog an effective amount of an anti-LHHR vaccine following spaying.

The vaccines of the present Invention may be administered by any suitable method known in the art, including, but not limited to, oral administration or by injection. The pharmaceutical forms suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. They must be stable, both physically and chemically, under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetable oils. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. The prevention of the action of microorganisms can be brought about by various antibacterial and antifungal agents, for example, parabans, chlorobutanol, phenol, eorbic acid, thimβrosal and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars or sodium chloride. Prolonged absorption or delayed release of the injectable compositions can be brought about by the use in the compositions of agents delaying absorption, for example, aluminum monostearate and gelatin.

Sterile injectable solutions are prepared by incorporating the active compounds in the required amount in the appropriate solvent with various of the other ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum drying, the freeze-drying technique and the spray-drying technique which yield a powder of the active ingredients plus any additional desired ingredient from previously sterile-filtered solution thereof. When the active Ingredients are suitably protected they may be orally administered, for example, with an inert diluent or with an assimilable edible carrier, or they may be enclosed in hard or soft shell gelatin capsule, or compressed into tablets, or incorporated directly with the food of the diet For oral administration, the active compound may be incorporated with excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. Such compositions and preparations should contain at least 1 % by weight of active compound. The percentage of the compositions and preparations may, of course, be varied and may conveniently be between about 5 to about 80% of the weight of the unit. The amount of active compound in such useful compositions is such that a suitable dosage will be obtained. Preferred composition's or preparations according to the present invention are prepared so that an oral dosage unit form contains between about 0. I μg and 2000 μg of active compound.

The tablets, troches, pills, capsules and the like may also contain the components as listed hereafter A binder such as gum, acacia, corn starch .or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; and a lubricant such as magnesium stearate. When the dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier. Various other materials may be present as coatings or to otherwise modify the physical form of the dosage unit A syrup or elixir may contain the active compound, methyl and propylparabens as preservatives,, and a dye. Of course, any material used in preparing any dosage unit form should be veterinarily pure and substantially non-toxic in the amounts employed. In addition, the active cαmpound(s) may be incorporated into sustained-release preparations and formulations. Pharmaceutically acceptable carriers and/or diluents include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. The use of such media and agents for veterinarily active substances is well known in the art Except insofar as any conventional media or agent is incompatible with the active ingredient, use thereof in the therapeutic compositions is contemplated. Supplementary active ingredients can also be incorporated into the compositions. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. For administration to livestock it is particularly advantageous to use a multi-dose container linked to a repeating vaccination gun. Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subjects to be treated; each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. The specification for the novel dosage unit forms of the invention are dictated by and directly dependent on (a) the unique characteristics of the active material and the particular therapeutic effect to be achieved, and (b) the limitations inherent in the art of compounding such an active material for the treatment of disease in Irving subjects having a diseased condition in which bodily health is impaired as herein disclosed in detail.

- The principal active ingredient is compounded for convenient and effective administration in effective amounts with a suitable pharmaceutically acceptable carrier in dosage unit form as hereinbefore disclosed, A unit dosage form can, for example, contain the principal active compound in amounts ranging from about 0.5 μg to about 2000 μg. EExpressed in proportions, the active compound is generally present in from about 0.5 μg to about 2000 μg /ml of carrier. The relative amounts of the active ingredient, the pharmaceutically acceptable carrier, and any additional ingredients in a pharmaceutical composition of the invention will vary, depending upon the identity, size, and condition of the subject treated and further depending upon the route by which the composition is to be administered.

The invention is further described in detail by reference to the following experimental examples. These examples are provided for purposes of illustration only, and are not intended to be limiting unless otherwise specified. Thus, the invention should in no way be construed as being limited to the following examples, but rather, should be construed to encompass any and all variations which become evident as a result of the teaching provided herein. ,

While the invention has been disclosed with reference to specific embodiments, it is apparent that other embodiments and variations of this invention may be devised by others skilled in the art without departing from the true spirit and scope of the invention. The appended claims are intended to be construed to include all such embodiments and equivalent variations.

[Example 1 Scope of Clinical Study:

Female dogs (bitches) that have been surgically neutered or spayed, frequently suffer urinary Incontinence. The incidence of this disease has been reported as high as 20%. Current treatments rely on daily administration of drugs such as synthetic oestrogeπs for extended periods, which is both expensive and inconvenient. The use of a long acting vaccine would be a significant improvement in the treatment of this common condition of spayed female dogs.

Considerations: • . . . - ^• .

A formulation to treat urinary incontinence in dogs includes any effective anti-LHRH vaccine.

Many such vaccines have been described in the prior art, although most of these fail to provide a high enough efficacy and safety to be commercially viable. There have been very few examples of effective vaccines described in the literature in dogs. There are several examples of LHRH vaccines currently commercially available, particularly in Australia and New Zealand. These include IMPROVAC® for the control of boar taint in male pigs, and EQUITY® for the control of oestrus related behavior in mares and fillies, both products commercially obtainable from Pfizer Animal Health. These formulations are for application in large animals, and so would be expected to be unsafe in smaller animals such as dogs, and would-be expected to require modification for safe and effective use in the canine and other companion animals. A further ^'formulation has been conditionally registered with the USDA in the USA for the treatment of Benign Prostatic Hyperplasia in adult intact male dogs. This vaccine has been demonstrated to be both safe and effective in adult male dogs and Is contemplated for use in the present invention to control of Canine Urinary Incontinence (CUI), as follows:

A study is conducted as a double blinded study, with two groups receiving either CUl vaccine or placebo, the placebo consisting of adjuvant alone. Neither the owners nor the clinical veterinarians are aware of the treatment being given.

Animals:

Bitches (up to 35 in number) that are kept as household pets by members of the community are selected from a clinical register without limitation, such as for example the register at The University of Melbourne / Veterinary Clinic. Only spayed bitches greater than 2 years of age, having long term clinical condition of urinary incontinence, and having been treated previously for this condition (with other recognized treatments, such as stilbestrol, a synthetic form of estrogen) or with an alpha-adrenergic stimulator such as PPA are selected for the study.

in one embodiment, the vaccine comprises that which is provisionally licensed by USDA to Pfizer Animal Health for the treatment of Benign Prostatic Hyperplasia, which contains 200ug/dose of conjugate of 2-10 form of LHRH peptide conjugated to diphtheria toxoid as carrier, with a combined adjuvant system of 10 mg DEAE dextran and 80 ug of an immunostlmulating complex as described in International Application No. PCT/AU99/01167. published on January 21, 1999 as WO 99/02180. In another ^f embodiment, the vaccine comprises a synthetic peptide vaccine comprising from about 3 to about 10 T-cell helper epitopes, each in linear sequence with a 4-10 form of LHRH₁ total peptide content being 120ug peptide/dose in combination with 80ug immunostimulatiπg complex.

The vaccine is administered to each dog in the study on at least two occasions at a 28-day interval.

The following parameters are measured directly following each vaccination: body temperature, demeanour, including lethargy, pain at site of injection, and swelling at site of injection. In addition, samples of serum are taken to measure levels of LH and antibody responses to LHRH. Scoring of urinary incontinence in each bitch is conducted by the individual owners, according to a common scoring system, starting 2 weeks prior to the first vaccination. In this way each individual dog acts as its own control for changes following treatment

Claims

What Ia Claimed:

1. A method for the treatment of urinary incontinence in a mammal in need of such treatment, said method comprising administering to said mammal a therapeutically effective amount of an anti-LHRH vaccine.

2. The method of claim 1 ; wherein said mammal is a female.

3. The method of claim 1. wherein said mammal is a female having elevated serum concentrations of a gonadotropin.

4. The method of claim 3, wherein said gonadotropin is selected from the group consisting of luteinizing hormone and follicle stimulating hormone.

5. The method of claim 1 , wherein said mammal is surgically neutered.

6. The method of claim 5, wherein said mammal is a spayed female dog.

7. The method of claim 1, wherein said anti-LHRH vaccine comprises an immunogenic conjugate between a carrier and LHRH.

8. The method 7, wherein said carrier comprises a protein.

9. The method of claim 8, wherein said protein comprises a bacterial toxoid.

10. The method of claim 9, wherein said bacterial toxoid is diphtheria toxoid.

11. The method of claim 7, wherein said carrier comprises a synthetic peptide comprising at least one T-cell helper epitope.

12. The method of claim 1 , wherein said vaccine further includes an adjuvant.

13. The method of claim 12, wherein said adjuvant comprises a DEAE-dextran.

14. The method of claim 13, wherein said adjuvant further comprises an immunostimulating complex, wherein said immunostimulatlng complex is comprised of a saponin and a cholesterol component.

15. The method of claim 7, wherein said immunogenic conjugate comprises an LHRH-diphtheria toxoid conjugate.

16. A method for treating or controlling urinary incontinence in a mammal comprising: administering to said mammal, a composition comprising a therapeutically effective amount of an immunogenic conjugate between a carrier and LHRH.

17. The method of daim 16, wherein said carrier comprises a protein.

1 β. The method of claim 17. wherein said,carrier comprises diphtheria toxoid.

19. The method of daim 16, wherein said carrier comprises a T-CΘII epitope.

20. The method of claim 16, wherein said composition further includes an adjuvant

21. A vaccine .for the treatment or control of urinary incontinence in a female mammal, said vaccine comprising a therapeutically effective amount of LHRH.

22. The vaccine of claim 21, further comprising an adjuvant.

23. The vaccine of daim 21, wherein the LHRH is conjugated with an immunogenic carrier.

24. The vaccine of claim 23, wherein said immunogenic carrier comprises a protein.

25. The vaccine of claim 23, wherein said immunogenic carrier comprises a T- cell helper epitope

26. The vaccine of claim 24, wherein said protein comprises diphtheria toxoid.

27. The vaccine of claim 21 , wherein said female mammal comprises a surgically spayed dog.

28. A method of enhancing the immune response of a mammal suffering from urinary incontinence due to the unrestricted production and release of a gonadotropin, said method comprising administering to said mammal a therapeutically effective amount of LHRH.

29. The method of claim 28, wherein said LHRH is administered in the form of an antigen.

30. ■ The method of claim 28, wherein said LHRH is administered in the form of a vaccine.

31. A method for treating urinary incontinence in a surgically neutered or spayed dog comprising administering to the dog a therapeutically effective amount of an aπti-LHRH vaccine.

32. The method of claim 31 , wherein the dog is a female.

33. A method of preventing the onset of urinary incontinence in a spayed female dog, said method comprising administering to said spayed female dog an effective amount of an antl-LHRH vaccine following spaying.