Deslorelin for reducing the odor of animal urine

Abstract

The present invention relates to deslorelin for use in reducing the odor of urine and feces in non-human animals, preferably mammals, and preferentially companion animals.

Description

Technical Field

[0001] The present disclosure generally relates to pharmaceutical compositions of deslorelin for use in reducing the odor of urine and feces in non-human animals, preferably mammals, and preferentially companion animals.

Background Art

[0002] Deslorelin is a gonadotropin-releasing hormone (GnRH) gonadotropin analogue that acts by suppressing the function of the gonadotropic pituitary axis when administered continuously at low doses.

[0003] GnRH is a hypothalamic decapeptide that acts at the top of the cascade that regulates the function of the hypothalamic-pituitary-gonadal axis. GnRH binds to GnRH receptors (seven transmembrane receptors) and causes the pituitary to produce and release two important gonadotropins, follicle-stimulating hormone (FSH) and luteinizing hormone (LH), which then control gonadal function. - LH (Luteinizing Hormone): In addition to its role in gametogenesis, it promotes the production of sex steroids (androgens, estrogens and progesterone); - FSH (Follicle-Stimulating Hormone): Stimulates the production of gametes (sperm and eggs).

[0004] When GnRH binds to the receptor, it increases the synthesis of the receptor's mRNA and enhances the pituitary's sensitivity to GnRH. Continuous secretion of GnRH has the opposite effect (a breakdown in LH and FSH secretion is observed, which can be called desensitization) (Finch A.R. et al., Agonist-induced internalization and downregulation of gonadotropin-releasing hormone receptors. Am. J. Physiol. Cell Physiol. 2009 Sep., 297(3)).

[0005] Desensitization of the pituitary gland to GnRH is caused by internalization of the GnRH receptor (no longer present and not binding to the cell surface) and inactivation of the intracellular signaling cascade. As soon as desensitization of pituitary cells to GnRH begins, LH levels drop to undetectable levels and thus cannot support testosterone and sperm production.

[0006] To solve the problem of contraception in animals, there are people who consider using GnRH agonists as contraceptives continuously over a long period of time, and thus the idea of designing a deslorelin implant that continuously diffuses this GnRH agonist (deslorelin) has emerged.

[0007] This type of implant is used in two different ways. - Fertility induction (short-term implant): Ovuplant - Induction of infertility by long-term action: Suprelorin

[0008] These agonist-based implants are known in the state of the art to have a two-stage action during implantation of the implant, releasing deslorelin and inducing a strong stimulation of the LH and FSH production cascades. This step is called the "flare-up". In male dogs, this induces the production of LH and FSH and leads to strong synthesis of testosterone. Subsequently, the male has strong fertility (improvement of sperm quality, improvement of motility).

[0009] After this "flare-up" period, a period of infertility with harmful effects on all situations of sperm quality and testosterone concentration is established (Goericke-Pesch S., Long-term effects of GnRH agonists on fertility and behaviour. Reprod. Domest. Anim., 2017, 52 Suppl2: 336 - 347).

[0010] In addition to testing in dogs, studies have been conducted in cats and pre-pubescent cats (Fontaine C., Long-term contraception in a small implant: A review of Suprelorin (deslorelin) studies in cats., J. Feline Med. Surg., 2015, 17(9), pages 766-771). This has shown the efficacy of the implant as a contraceptive in both males and females. During the reproductive suppression period, steroid hormone concentrations of less than 1 ng / ml progesterone and 10 pg / ml estradiol were measured in females, and plasma testosterone concentration in male cats was at basal levels (<0.1 ng / ml). In males, the duration of action using a 4.7 mg implant was 78.8 ± 12.9 weeks, varying from 61.7 weeks (about 15 months) to 100.7 weeks (about 25 months).

[0011] To date, no PK / PD (pharmacokinetic / pharmacodynamic) relationship has been shown in the known data, and the efficacy of the product has been explained by a very large initial "burst" immediately after implantation and a very long-lasting saturation of the GnRH receptor. Current RCP data in dogs show the presence of deslorelin for 2.5 months, but its effects are observed for at least 6 months (Source: EMA website: https: / / www.ema.europa.eu / en / documents / product-information / suprelorin-epar-product-information_en.pdf).

[0012] New data have been generated, showing that deslorelin can be measured if its effects are observed in male and female dogs and cats.

[0013] In addition to this, maintaining the concentration range of deslorelin in the bloodstream to obtain and retain the castration effect has been shown to be related to the PK / PD interaction. Contrary to the prior art and common knowledge regarding the mode of action of deslorelin, the PK / PD relationship between the in vivo level of deslorelin and the PD effect has been further clearly demonstrated, allowing us to identify the efficacy threshold.

[0014] In addition to the contraceptive effect of deslorelin, other effects have been found as reported in the examples of the present invention regarding sexual behavior and urine marking and / or the smell of urine.

[0015] Urine marking is · A pheromone signal to other animals, especially intact male cats · A territorial signal · An indication of stress or sexual excitement related to the social or physical environment the normal and intentional deposition of urine as.

[0016] In such situations, the urine may have a strong smell, and it is particularly desirable for pet owners to remove or mask this smell and / or reduce or decrease it. Some commercially available enzyme-based urine odor neutralizers can be used on the surface after cleaning the area with a mild detergent and can be used to remove the smell, but these neutralizers require great care. Castration or ovariectomy has been proven as a treatment for cats that mark, especially for breeding or territorial activities. One of the reasons for surgical castration is to alleviate the strong and persistent urine smell of male animals, especially cats and ferrets. After surgical castration, it is known that the urine smell improves after a few days.

[0017] However, using surgery to solve this problem is very drastic considering the cause of this problem and can cause side effects.

[0018] One of the goals of the present application is to show that deslorelin administration (e.g., when administered with an implant such as Suprelorin®) reduces the odor of urine and / or feces. Surprisingly, the inventors have shown that the odor reduction is brought about via at least two pathways. In fact, one such pathway is the sex hormone pathway mediated by testosterone. The inventors have also demonstrated at least one other pathway not related to sex hormones, as described in the present application, showing that the reduction of the unpleasant odor of urine and / or feces is improved.

[0019] Therefore, when inducing chemical castration of the administered non-human animals, this problem is solved by using deslorelin, which reduces the odor of urine and / or feces.

[0020] A recent study conducted by Chumki Banik et al. (Simultaneous Chemical and Sensory Analysis of Domestic Cat Urine and Feces with Headspace Solid-Phase Microextraction and GC-MS-Olfactometry Separations 2021,8,15pp1of19) aimed to identify volatile organic chemicals (VOCs) and related odors in cat urine and feces using gas chromatography-mass spectrometry of fresh and old samples, as well as simultaneous sensory analysis. The purpose of this study was different from that of the present invention.

Prior Art Documents

Non-Patent Documents

[0021]

Non-Patent Document 1

[0022] The present invention relates to deslorelin for use in reducing or decreasing the odor of urine and / or feces, particularly unpleasant odor, in non-human animals, wherein after administration to the non-human animal, the deslorelin plasma concentration in the non-human animal is higher than 10 pg / mL. In a preferred embodiment, the deslorelin plasma concentration is from 10 pg / mL to 400 pg / ml from 8 days after administration up to at least 6 months after administration. In another preferred embodiment, deslorelin is continuously administered to the non-human animal or is continuously released after administration. For example, deslorelin is substantially continuously released at a plasma concentration level of less than 400 pg / mL from 8 days after administration up to at least 6 months after administration.

[0023] In another preferred embodiment, after administration, deslorelin modulates, i.e., downregulates or upregulates, molecules that cause the odor of urine and / or feces, and the molecules are selected from the group consisting of felinine derivative compounds, volatile organic compounds (VOCs), sulfur compounds, and testosterone-induced compounds.

[0024] In certain embodiments, deslorelin is - sulfur compounds selected from the group consisting of 4-methyl-4-sulfanyl-pentan-2-one; 3-methyl-3-sulfanyl-butan-1-ol; 3-methyl-3-sulfanyl-1-butyl formate; - VOCs: 2,5-dimethylpyrazine; 1,2-dichloro-propane; 4,4-dimethyl-2-pentanone; and 3-methyl-2-buten-1-ol; - compounds selected from the group consisting of 2-(methylthio)-1-ethanol; 2-methyl-3-furanthiol; oxathiolane; 4-(methylthio)-2-butanone; 3-methylbutyl-3-sulfanyl formate; and 3-methyl-3-(methylthio)-1-butanol; - p-cresol; - felinine and its derivatives such as 3-mercapto-3-methyl-1-butanol; 3-mercapto-3-methylbutyl formate; 3-methyl-3-methylthio-1-butanol; and 3-methyl-3-(2-methyldisulfanyl)-1-butanol. - Ferinin precursors such as N-acetylferinin, γ-glutamylferinylglycine, and 3-methylbutanol cysteinylglycine; - Coccine; - 2-Phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isovaltin; - Isobutain; - Trimethylamine; - Indole; - Cadaverine and cadaverine derivatives such as N-acetylcadaverine (also known as acetylcadaverine) and glutathione aminopropylcadaverine, - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid; - Crotonic acid; - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvic acid, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoyl threonine, 1alpha,5alpha-dimercaptoandrostane-3alpha,17beta-diol, mercaptopyruvic acid, and 2-mercapto-2-methylpentan-4-one; and - Taurine - Valine regulates the molecules that cause the odor of urine and / or feces, that is, down-regulates or up-regulates them.

[0025] In a more preferred embodiment, the molecules that cause the odor of urine and / or feces and that deslorelin has an effect on are - Sulfur compounds selected from the group consisting of 4-methyl-4-sulfanyl-pentan-2-one; 3-methyl-3-sulfanyl-butan-1-ol; 3-methyl-3-sulfanyl-1-butyl formate; - VOC: 2,5-dimethylpyrazine; 1,2-dichloro-propane; 4,4-dimethyl-2-pentanone; and 3-methyl-2-buten-1-ol; - Compounds selected from the group consisting of 2-(methylthio)-1-ethanol; 2-methyl-3-furanthiol; oxathiolane; 4-(methylthio)-2-butanone; 3-methylbutyl 3-sulfanylformate; and 3-methyl-3-(methylthio)-1-butanol; - p-Cresol - Ferinen and derivatives: 3-mercapto-3-methyl-1-butanol; 3-mercapto-3-methylbutyl formate; 3-methyl-3-methylthio-1-butanol; and 3-methyl-3-(2-methyldisulfanyl)-1-butanol; - Ferinen precursors: N-acetylferinen, γ-glutamylferinylglycine and 3-methylbutanol cysteinylglycine; - Coxine; - 2-Phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isovaltin; - Isobutain; - Trimethylamine, - Indole, - Cadaverine and cadaverine derivatives such as N-acetylcadaverine and glutathionylaminopropylcadaverine; - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid and pentanoic acid; - Crotonic acid; and - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2 - mercapto - 3 - butanone, 2 - mercaptoethanol, 3 - mercaptohexyl butyrate, mercaptopyruvic acid, 3 - mercaptopropionic acid, 3 - mercapto - 3 - methylbutan - 1 - ol, 7 - mercaptoheptanoyl threonine, 1alpha,5alpha - dimercaptoandrostane - 3alpha,17beta - diol, mercaptopyruvic acid; - Taurine is selected from.

[0026] In a more preferred embodiment, the molecule is - Trimethylamine, - 3 - Methylbutanoic acid - Indole - Cadaverine and acetylcadaverine, derivatives such as glutathionylaminopropylcadaverine, - Short - chain free fatty acids: acetic acid, propanoic acid, 2 - methylpropanoic acid, butanoic acid, 3 - methylbutanoic acid and pentanoic acid - Crotonic acid - Putrescine and N - acetylputrescine, derivatives such as p - coumaroylputrescine; - Mercapto derivatives such as 2 - mercapto - 3 - butanone, 2 - mercaptoethanol, 3 - mercaptohexyl butyrate, mercaptopyruvic acid, 3 - mercaptopropionic acid, 3 - mercapto - 3 - methylbutan - 1 - ol, 7 - mercaptoheptanoyl threonine, 1alpha,5alpha - dimercaptoandrostane - 3alpha,17beta - diol, mercaptopyruvic acid; - Taurine is selected from the list of.

[0027] According to another preferred embodiment, deslorelin down - regulates the molecules that cause the odor of urine and / or feces, and the molecules are - Sulfur compounds selected from the group consisting of 4 - methyl - 4 - sulfanyl - pentan - 2 - one; 3 - methyl - 3 - sulfanyl - butan - 1 - ol; 3 - methyl - 3 - sulfanyl - 1 - butyl formate; - VOC: 2,5-dimethylpyrazine; 1,2-dichloro-propane; 4,4-dimethyl-2-pentanone; and 3-methyl-2-buten-1-ol; - A compound selected from the group consisting of 2-(methylthio)-1-ethanol; 2-methyl-3-furanthiol; oxathiolane; 4-(methylthio)-2-butanone; 3-methylbutyl 3-sulfanylformate; and 3-methyl-3-(methylthio)-1-butanol; - p-Cresol - Ferulinin and its derivatives such as 3-mercapto-3-methyl-1-butanol; 3-mercapto-3-methylbutyl formate; 3-methyl-3-methylthio-1-butanol; and 3-methyl-3-(2-methyldisulfanyl)-1-butanol; - Ferulinin precursors such as N-acetylferulinin, γ-glutamylferulylglycine, and 3-methylbutanol cysteinylglycine; - Cochicine; - 2-Phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isovaltin; - Isobutain; - Trimethylamine; - Indole; - Cadaverine and its derivatives such as N-acetylcadaverine and glutathionylaminopropylcadaverine; - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid; - Crotonic acid; - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2 - mercapto - 3 - butanone, 2 - mercaptoethanol, 3 - mercaptohexyl butyrate, mercaptopyruvic acid, 3 - mercaptopropionic acid, 3 - mercapto - 3 - methylbutan - 1 - ol, 7 - mercaptoheptanoyl threonine, 1alpha,5alpha - dimercaptoandrostane - 3alpha,17beta - diol, mercaptopyruvic acid; 2 - mercapto - 2 - methylpentan - 4 - one; and - Taurine is selected from the group consisting of.

[0028] According to another preferred embodiment, deslorelin up - regulates molecules that cause the odor of urine and / or feces, and the molecules are selected from the group consisting of 2 - mercapto - 2 - methylpentan - 4 - one and valine.

[0029] According to another embodiment, the present invention relates to the non - pharmaceutical or non - therapeutic use of deslorelin for regulating, that is, down - regulating or up - regulating, the level of molecules in the urine and / or feces of non - human animals, and the molecules are - Sulfur compounds selected from the group consisting of 4 - methyl - 4 - sulfanyl - pentan - 2 - one; 3 - methyl - 3 - sulfanyl - butan - 1 - ol; 3 - methyl - 3 - sulfanyl - 1 - butyl formate; - VOCs: 2,5 - dimethylpyrazine; 1,2 - dichloro - propane; 4,4 - dimethyl - 2 - pentanone; and 3 - methyl - 2 - buten - 1 - ol; - Compounds selected from the group consisting of 2 - (methylthio) - 1 - ethanol; 2 - methyl - 3 - furanthiol; oxathiolane; 4 - (methylthio) - 2 - butanone; 3 - methylbutyl - 3 - sulfanyl formate; and 3 - methyl - 3 - (methylthio) - 1 - butanol; - p - Cresol; - Ferinen and derivatives: 3 - mercapto - 3 - methyl - 1 - butanol; 3 - mercapto - 3 - methylbutyl formate; 3 - methyl - 3 - methylthio - 1 - butanol; 3 - methyl - 3 - (2 - methyldisulfanyl) - 1 - butanol and 3 - mercapto - 3 - methyl - 1 - butanol, - Ferenin precursors such as N-acetylferenin, γ-glutamylferenylglycine, and 3-methylbutanol cysteinylglycine; - Coccin; - 2-Phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isovaltin; - Isobutein; - Trimethylamine; - Indole; - Cadaverine and cadaverine derivatives such as acetylcadaverine and glutathionylaminopropylcadaverine; - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid; - Crotonic acid - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvic acid, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoyl threonine, 1alpha,5alpha-dimercaptoandrostane-3alpha,17beta-diol, mercaptopyruvic acid, and 2-mercapto-2-methylpentan-4-one, - Taurine, and - Valine is selected from the group consisting of.

[0030] In a preferred embodiment, the non-pharmaceutical or non-therapeutic use of deslorelin is to adjust, i.e., down-regulate or up-regulate, the level of a molecule in a non-human animal, and the molecule is - Trimethylamine, - 3-Methylbutanoic acid, - Indole, - Cadaverine and cadaverine derivatives such as acetylcadaverine and glutathionylaminopropylcadaverine, - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid, - Crotonic acid, - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvic acid, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoyl threonine, and 1alpha,5alpha-dimercaptoandrostane-3alpha,17beta-diol, mercaptopyruvic acid and 2-mercapto-2-methylpentan-4-one; and - Taurine is selected from the group consisting of.

[0031] In a further preferred embodiment, deslorelin modulates the level of molecules in urine, i.e., down-regulates or up-regulates.

[0032] In a further preferred embodiment, deslorelin modulates the level of molecules in feces, i.e., down-regulates or up-regulates.

[0033] In a further preferred embodiment, the present invention relates to the non-therapeutic use of deslorelin for down-regulating the level of molecules in the urine and / or feces of non-human animals, said molecules being - Sulfur compounds selected from the group consisting of 4-methyl-4-sulfanyl-pentan-2-one; 3-methyl-3-sulfanyl-butan-1-ol; 3-methyl-3-sulfanyl-1-butyl formate; - VOCs: 2,5-dimethylpyrazine; 1,2-dichloropropane; 4,4-dimethyl-2-pentanone; and 3-methyl-2-buten-1-ol; - A compound selected from the group consisting of 2-(methylthio)-1-ethanol; 2-methyl-3-furanthiol; oxathiolane; 4-(methylthio)-2-butanone; 3-methylbutyl 3-sulfanylformate; and 3-methyl-3-(methylthio)-1-butanol; - p-Cresol; - Ferulinin and its derivatives such as 3-mercapto-3-methyl-1-butanol; 3-mercapto-3-methylbutyl formate; 3-methyl-3-methylthio-1-butanol; and 3-methyl-3-(2-methyldisulfanyl)-1-butanol; - Ferulinin precursors such as N-acetylferulinin, γ-glutamylferulylglycine, and 3-methylbutanol cysteinylglycine; - Coccygin; - 2-Phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isovaltin; - Isobutain; - Trimethylamine; - Indole; - Cadaverine and its derivatives such as N-acetylcadaverine and glutathionylaminopropylcadaverine; - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid; - Crotonic acid; - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvic acid, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoyl threonine, 1alpha,5alpha-dimercaptoandrostane-3alpha,17beta-diol, and mercaptopyruvic acid; and - Taurine is selected from the group consisting of.

[0034] In another preferred embodiment, the present invention relates to the non-therapeutic use of deslorelin for upregulating the levels of molecules in the urine and / or feces of non-human animals, said molecules being selected from the group comprising 2-mercapto 2-methylpentan-4-one and valine.

[0035] The present invention also relates to a method for reducing or decreasing the odor of urine and / or feces in non-human animals, said method comprising administering to the non-human animal an amount of deslorelin such that after administration to the non-human animal, the plasma concentration of deslorelin in the non-human animal is higher than 10 pg / mL.

[0036] Another object of the present invention is also a method for reducing or decreasing the odor of urine and / or feces in non-human animals, said method comprising administering to the non-human animal an amount of deslorelin that induces regulation, i.e., downregulation or upregulation, of the levels of odor molecules in the urine and / or feces. Preferably, the non-human animal is an intact male cat. According to this embodiment, deslorelin induces regulation, i.e., downregulation or upregulation, of the levels of odor molecules in the urine and / or feces via the sex hormone synthesis pathway.

[0037] In another embodiment, the present invention also relates to a method for improving the reduction or decrease of the odor of urine and / or feces in non-human intact male animals compared to castrated male animals, said method comprising administering to the non-human intact male animal an amount of deslorelin that induces regulation, i.e., downregulation or upregulation, of the levels of odor molecules in the urine and / or feces. According to this embodiment, deslorelin induces regulation, i.e., downregulation or upregulation, of the levels of odor molecules in the urine and / or feces via a non-sex hormone-dependent pathway.

[0038] The present invention also encompasses deslorelin for use in improving the reduction or decrease of the odor of urine and / or feces in non-human animals.

[0039] Finally, the present invention also generally Regarding the use of deslorelin for producing an agent that reduces or decreases the odor of urine and / or feces in non-human animals, after administration to the non-human animals, the deslorelin plasma concentration of the non-human animals is higher than 10 pg / mL.

[0040] The use of deslorelin in cats or dogs, male or female, modulates, i.e., downregulates or upregulates, the level of a certain molecule in urine, and at the same time, it has been shown that the level of the derivative of that molecule increases or decreases in urine.

[0041] The use of deslorelin in cats or dogs, male or female, modulates, i.e., downregulates or upregulates, the level of a certain molecule in feces, and at the same time, it has been found that the level of the derivative of that molecule increases or decreases in feces.

[0042] The use of deslorelin in cats or dogs, male or female, modulates, i.e., downregulates or upregulates, the level of a certain molecule in urine, and at the same time, it has also been found that the level of the same molecule or its derivative level in feces increases or decreases.

[0043] The use of deslorelin in cats or dogs, male or female, modulates, i.e., downregulates or upregulates, the level of a certain molecule in feces, and at the same time, it has also been found that the level of the same molecule or its derivative level in urine increases or decreases.

[0044] Also, several metabolomics molecules have been identified in feces. In fact, from the data, the following molecules are selected from the following molecules: trimethylamine, 3-methylbutanoic acid, and crotonic acid.

[0045] In fact, cat feces have trimethylamine, which has the smell of rotten fish and could not be identified by GC-MS in a fresh fecal sample. However, the fishy odor has been recorded by odor experts, and it has been confirmed that trace levels of concentration can be detected by the sensory organs of the living body. Therefore, a reduction in the odor induced by trimethylamine is required.

[0046] The present invention has several advantages including but not limited to the following. First of all, it has an effect on at least two pathways that synthesize the odor of urine or feces. The results regarding odor reduction are the combination of the results regarding at least these two pathways, and it is clear that they are definitely superior to the action alone based on the synthesis of urine / feces odor by the sex hormone pathway.

[0047] It is currently shown that in animals chemically castrated by deslorelin administration compared to surgically castrated animals, the molecules that cause the odor of urine or feces change quantitatively and qualitatively, and some are reduced or decreased.

[0048] Also, according to the present disclosure, the method of using deslorelin as described herein is a reversible, non-surgical, reliable contraceptive measure, and it has been demonstrated that it promotes the health of the pet family while leaving options.

[0049] The deslorelin action, at least via the testosterone pathway, especially when reducing its blood level, can contribute to the prevention of kidney diseases in the animal. This is because these types of diseases are known to be related to the accumulation of molecules such as trimethylamine.

[0050] Furthermore, it should also be mentioned that for the safety study of cats, high-dose deslorelin administration has not revealed any serious adverse events or life-threatening events. Also, according to several tolerance studies of male and female species, deslorelin administered as an implant (i.e., Suprelorin (registered trademark)) has been concluded to show good tolerance.

[0051] In addition to the above typical aspects and embodiments, further aspects and embodiments of the present invention will become apparent by referring to the drawings and studying the following description.

Brief Description of the Drawings

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Mode for Carrying Out the Invention

[0053] One of the goals of the present application is to show that deslorelin administration (for example, when administered with an implant such as Suprelorin®) reduces the odor of urine and / or feces, particularly unpleasant urine and / or fecal odor. Surprisingly, the inventors have shown that the odor reduction is brought about via at least two pathways. Indeed, one such pathway is the sex hormone pathway mediated by "classical" testosterone. The inventors have also demonstrated at least one other pathway not related to sex hormones, as described in the present application.

[0054] Therefore, the object of reducing the odor of urine and / or feces in animals is solved by using deslorelin. Deslorelin reduces or decreases the unpleasant odor of urine and / or feces when inducing chemical castration of the administered non-human animal.

[0055] Indeed, the inventors have shown that deslorelin administration affects various molecules involved in the excretion of urine and / or feces. Depending on the breakdown or synthesis of these molecules, they can also cause the odor of urine or feces. Therefore, one object of the present invention is to intervene in the composition of the odor of urine and / or feces by using deslorelin to increase, decrease or change the concentration of specific molecules in urine and / or feces, and in particular to down-regulate or up-regulate them. Examples of deslorelin action are further developed below.

[0056] The term "regulate" here refers to an increase or decrease in molecular weight in response to deslorelin administration. Regulation includes up-regulation and down-regulation.

[0057] The term "down-regulate" here refers to a decrease in the amount of a molecule in response to deslorelin administration compared to the amount of the molecule in urine and / or feces before the start of deslorelin administration.

[0058] The term "up-regulate" here refers to an increase in the amount of a molecule in response to deslorelin administration compared to the amount of the molecule in urine and / or feces before the start of deslorelin administration.

[0059] The terms "continuously" and "substantially continuously" in relation to the administration of deslorelin mean that deslorelin is administered without interruption or substantially without interruption, for example, by implantation of a sustained-release drug delivery system.

[0060] The molecule can be measured by methods known to those skilled in the art, including tandem mass spectrometry (also known as MS / MS or MS2), two-dimensional gas chromatography-mass spectrometry (also known as the GCxGC-MS method, as described, for example, in C. Suzuki et al., Journal of Chemical Ecology, 45, 579-587 (2019)), and liquid chromatography MS / MS.

[0061] According to a preferred embodiment of the present invention, deslorelin downregulates the urinary concentration of molecules selected from the group consisting of para-cresol, ferenin, ferenin precursors such as 3-methylbutanol cysteinylglycine (MBCG), ferenin derivatives such as N-acetylferenin, and derivatives thereof such as 3-mercapto-3-methylbutan-1-ol, isovaltin, isobutain, indole, cadaverine, and N-acetylcadaverine, putrescine, and derivatives thereof such as N-acetylputrescine and p-coumaroylputrescine, mercaptopyruvic acid, 3-mercaptopropionic acid, mercapto compounds such as 7-mercaptoheptanoyl threonine, and taurine.

[0062] According to the most preferred embodiment, deslorelin downregulates the urinary concentration of molecules selected from the group consisting of ferenin and its ferenin precursors such as 3-methylbutanol cysteinylglycine, derivatives thereof such as indole, cadaverine, and N-acetylcadaverine, putrescine, and derivatives thereof such as N-acetylputrescine and p-coumaroylputrescine, mercaptopyruvic acid, 3-mercaptopropionic acid, 7-mercaptoheptanoyl threonine, and taurine.

[0063] According to another preferred embodiment of the present invention, deslorelin up-regulates the urinary concentration of molecules selected from the group including 2-mercapto-2-methylpentan-4-one (a very preferred important food odorant of onions) and valine (a preferred fragrance of vanilla and maple syrup).

[0064] According to the most preferred embodiment of the present invention, deslorelin up-regulates the urinary concentration of valine.

[0065] According to another preferred embodiment of the present invention, deslorelin down-regulates the fecal concentration of molecules selected from the group including crotonic acid and trimethylamine.

[0066] As will be described hereinafter, the down-regulation and up-regulation of these various molecules can be affected by two different synthetic pathways.

[0067] Synthetic pathways of urinary or fecal odors by sex hormones It is known from the prior art (Hendriks WH et al., Testosterone increases urinary free felinine, N-acetylfelinine and methylbutanolglutathione excretion in cats (Felis catus)., J Anim Physiol Anim Nutr (Berl)., 2008 Feb; 92(1):53-62) that testosterone increases the secretion of free felinine, N-acetylfelinine and 3-methylbutanol glutathione in castrated adult male cats and non-castrated female cats, while estradiol does not modulate this effect.

[0068] Felinine and N-acetylfelinine were detected in various tissues. Furthermore, all compounds were detected in cat bile and fecal samples, indicating that felinine is secreted from the liver through bile into feces (LC-MS / MS quantification of felinine metabolites in tissues, fluids, and excretions from the domestic cat (Felis catus), Futsuta et al, J Chromatogr B Analyt Technol Biomed Life Sci., 2018 Jan 1;1072:94-99).

[0069] Therefore, a decrease in felinine levels and / or its metabolite levels and other molecules affects the odor of urine or feces.

[0070] It is also known from the prior art that coixin secretion is sex-dependent (M. Miyazaki et al., Comp Biochem Physiol B Biochem Mol Biol, 2006 Nov-Dec;145(3-4):270-7). In mature cats, coixin secretion is higher in intact male cats than in castrated male cats, or intact or ovariectomized female cats. Coixin secretion per day decreased immediately after castration. Immunohistochemistry confirmed that coixin expression in the renal proximal straight tubules is higher in intact male cats than in castrated male cats.

[0071] These results suggest that coixin secretion is regulated by sex hormones such as testosterone, and that the degradation products by coixin are secreted in a species-, sex- and age-dependent manner like coixin itself.

[0072] Also, coixin is a carboxylesterase secreted as a major urinary component and regulates felinine production. In vitro enzyme assays showed that coixin hydrolyzes 3-methylbutanol cysteinylglycine, a felinine precursor, into felinine and glycine.

[0073] Surprisingly, the inventors of the present invention have found and demonstrated that chemically induced castration by deslorelin affects the production of coprostanol and felinine. Felinine, as well as derivative molecules such as N-acetylfelinine, 3-methylbutanol cysteinylglycine (MBCG) and 3-mercapto-3-methylbutan-1-ol, also cause the smell of urine.

[0074] One object of the present invention is to reduce or decrease the smell of urine by the pathway of synthesizing the smell of urine or feces via sex hormones by using deslorelin in non-human animals, and after administration to the non-human animals, the deslorelin plasma concentration of the non-human animals is higher than 10 pg / mL.

[0075] Accordingly, according to the first object, the present invention relates to deslorelin for use in reducing or decreasing the smell of urine and / or feces in non-human animals, and after administration to the non-human animals, the deslorelin plasma concentration of the non-human animals is higher than 10 pg / mL.

[0076] More specifically, after administration to the non-human animals, the deslorelin plasma concentration is from 10 pg / mL to 400 pg / ml from 8 days to at least 6 months after administration. Preferably, deslorelin is administered continuously to the non-human animals or is continuously released after administration.

[0077] Also, a second object of the present invention is a method for reducing or decreasing the smell of urine and / or feces in non-human animals, which method comprises administering to the non-human animals an amount of deslorelin such that after administration to the non-human animals, the deslorelin plasma concentration of the non-human animals is higher than 10 pg / mL.

[0078] According to another embodiment, the present invention relates to a method for reducing or decreasing the smell of urine and / or feces in non-human animals, which method comprises administering to the non-human animals an amount of deslorelin that induces regulation, i.e., down-regulation or up-regulation, of the level of odor molecules in urine and / or feces.

[0079] Preferably, according to the present embodiment, the non-human animal is a non-castrated male animal, preferably a canine or feline, more preferably a dog or a cat, and most preferably a cat.

[0080] Preferably, according to the present embodiment, deslorelin regulates, or induces down-regulation or up-regulation of, the level of odor molecules in urine and / or feces via the sex hormone synthesis pathway.

[0081] As demonstrated in the examples illustrating the present invention, the concentration of felinine precursors such as felinine and 3-methylbutanol cysteinylglycine decreases after administration of deslorelin to non-castrated male adult cats.

[0082] Thus, according to a particularly preferred embodiment, the present invention relates to deslorelin for use in down-regulating the concentration of felinine and felinine derivatives in the urine of non-human animals.

[0083] Odor synthesis in urine or feces via non-sex hormone-dependent pathways In addition to the typical sex hormone pathway via testosterone that causes the odor of urine and / or feces, the inventors have discovered another pathway that is not related to sex-dependent hormones. Indeed, as shown in the examples, the inventors have identified novel molecules as molecules that cause the odor of urine and / or feces. These molecules are secreted outside the sex hormone pathway. In fact, these molecules do not originate from the testosterone cascade.

[0084] Accordingly, one aspect of the present invention is the use of deslorelin for reducing, increasing or changing the level of, or downregulating or upregulating, a molecule selected from the group consisting of crotonic acid; indole; p-cresol; cadaverine and cadaverine derivatives such as acetyl cadaverine and glutathione aminopropyl cadaverine; putrescine and putrescine derivatives such as N-acetyl putrescine and p-coumaroyl putrescine; and mercapto derivatives selected from the group consisting of 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvic acid, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoyl threonine, 1alpha,5alpha-dimercaptoandrostan-3alpha,17beta-diol, 2-mercapto-2-methylpentan-4-one, taurine, valine, isovaltin, isobutain and trimethylamine.

[0085] According to a preferred embodiment, deslorelin downregulates the urinary levels of molecules selected from the group consisting of isovaltin, isobutain, indole, cadaverine and its derivatives such as N-acetyl cadaverine, putrescine and its derivatives such as N-acetyl putrescine and p-coumaroyl putrescine, mercapto compounds such as mercaptopyruvic acid, 3-mercaptopropionic acid and 7-mercaptoheptanoyl threonine, and taurine.

[0086] According to the most preferred embodiment, deslorelin downregulates the levels of molecules selected from the group consisting of indole, cadaverine and its derivatives such as N-acetyl cadaverine, putrescine and its derivatives such as N-acetyl putrescine and p-coumaroyl putrescine, and mercaptopyruvic acid.

[0087] According to another preferred embodiment, deslorelin upregulates the urinary levels of molecules selected from the group consisting of 2-mercapto-2-methylpentan-4-one and valine.

[0088] According to another most preferred embodiment, deslorelin upregulates the valine level.

[0089] Isovalthine is a branched-chain sulfur-containing amino acid and is found in the urine of normal cats. The isovalthine concentration in the urine of healthy adult cats is about 24 - 66 micromoles / l and is not affected by the sex of the cat.

[0090] Isovalthinuria can be induced in other species (rats, rabbits, guinea pigs, humans, dogs) after administration of certain inducers such as certain cholesterol-lowering agents, bile acids, hormones or cholesterol precursors. The methods of inducing isovalthinuria have been widely studied in the 1960s and efforts have been made to understand its biosynthesis.

[0091] However, as shown in Figure 1, although the origin of the sulfur atom of isovalthine is shown to be derived from cysteine or methionine, the origin of the carbon skeleton remains unknown. Since isovalthine is thought to be found in the urine of patients with hypercholesterolemia, there is some interest in isovalthine metabolism. Thus, little is known about isovalthine.

[0092] The present inventors have now found that isovalthine is one of the molecules responsible for the smell of urine. Since urine is known to be a mixture of several molecules, isovalthine is considered to be one of the molecules actively involved in odor emission.

[0093] Little is also known about isobutein, which is also described as 2-amino-6-carboxy-4-thiaheptanoic acid. This amino acid has been isolated from the urine of healthy humans and cats, but its biological role is also unknown.

[0094] The present inventor has now found that isobutene is one of the molecules responsible for the smell of urine. Since urine is known to be a mixture of several molecules, isobutene is considered to be one of the molecules actively involved in odor emission.

[0095] In addition, several metabolomic molecules have also been identified in feces. In fact, from the data, the following molecules: trimethylamine and 3-methylbutyric acid have been identified.

[0096] Therefore, chemical castration induced by deslorelin administration also alleviates the smell of urine and / or feces over the same length of time. The inventor has surprisingly found that chemical castration with deslorelin reduces the smell of urine after several weeks. Generally, the smell is alleviated between 21 and 42 days, which means about 3 to 6 weeks.

[0097] In one aspect of the present invention, the smell of urine in non-sterilized animals, particularly male domestic cats, is reduced by administering deslorelin to the animals.

[0098] It is also recognized that some molecules are downregulated in urine but conversely upregulated in feces. This indicates a compensatory mechanism between the molecular levels removed in urine and feces at various levels compared to non-chemically castrated animals (data not shown).

[0099] Therefore, the present invention also relates to a method for improving the reduction or decrease of the smell of urine and / or feces in non-human non-castrated male animals compared to castrated males, the method comprising adjusting the level of odor molecules in urine and / or feces, or administering to the non-human non-castrated male animal an amount of deslorelin that induces downregulation or upregulation.

[0100] According to this embodiment, deslorelin induces the adjustment of the level of odor molecules in urine and / or feces, i.e., downregulation or upregulation, via a non-sex hormone-dependent pathway.

[0101] According to another embodiment, the present invention relates to deslorelin for use in improving the reduction or decrease of the odor of urine and / or feces in non-human animals.

Example

[0102] Example 1: Large-scale field evaluation of the efficacy and safety of a single 4.7 mg deslorelin implant in suppressing testosterone, sexual behavior, and urine odor in male cats The intention of this multi-site, double-blind, placebo-controlled, randomized study was to confirm that deslorelin implants could be used as an option for castration of male cats and that they could also reduce related male cat sexual behavior.

[0103] 1.1. Materials and methods Individually owned indoor male cats [3 months to 11 years old] were assigned to two groups, one received a 4.7 mg deslorelin implant (Suprelorin® 4.7 mg, Virbac, France) implanted, or the other received sodium chloride administered. The implant and saline were administered subcutaneously between the shoulder blades on day 0 (D0). Serum testosterone concentration (ng / mL) (T), sexual behavior (presence of urine marking, frequency of meowing, frequency of aggression), and the presence of the odor of urine in intact male cats (M) were evaluated up to 12 months (D372). General safety assessment, body weight, appetite, and side effects were monitored regularly.

[0104] The protocol was approved by an ethics review. The Cochran-Mantel-Haenszel test stratified by facility using RIDIT transformation and general relevant statistics were used for inter-group comparison.

[0105] Of the 205 male cats, n = 154 received Suprelorin® 4.7 mg (treatment group) and n = 51 received saline as the control group.

[0106] 1.2 Results: As shown in Figure 2, the average T value decreased in the treatment group and was significantly lower at D45 (0.13 ± 0.375 vs. 1.74 ± 1.547 ng / mL) and at M12 (0.64 ± 1.138 vs. 2.04 ± 1.708 ng / mL) compared to the control.

[0107] The sexual behavior score, testis volume, visibility of penile spines, and urine odor also decreased significantly and were significantly lower in the deslorelin group compared to the saline group throughout the study. No sexual behavior outbursts or safety concerns were observed.

[0108] 1.3 Conclusion: This study demonstrates that Suprelorin® 4.7 mg implant can be used in male cats from 3 months of age as an effective and safe castration option for at least one year, resulting in a reduction in male cat sexual behavior and urine odor.

[0109] Example 2: Evaluation of molecular level changes related to the sex hormone-dependent pathway The intent of these studies was to quantify the levels of specific molecules responsible for urine odor. The objective of this study was also to evaluate plasma levels of testosterone and metabolomics parameters present in the urine of chemically castrated male cats with deslorelin and surgically castrated male cats. Therefore, several parameters were evaluated. In particular, the dosing was performed in serum / plasma and urine.

[0110] 2.1 Materials and methods The first study A was performed on two groups of a total of 7 healthy male cats, 12 to 13 months old and weighing 4 to 6 kg. Control group: n = 3, intact males, administered sodium chloride (saline) on day 0 (D0). Chemical treatment group: n = 4, received one implant of Suprelorin® 4.7 mg on day 0 (D0).

[0111] Study A was performed for 63 days after implantation of Suprelorin® 4.7 mg.

[0112] From D-21 to D-7, clinical examinations (weight, temperature) of the cats were performed, and urine and fecal samples were collected. On D0, 4.7 mg of Suprelorin® (Virbac, France) was subcutaneously implanted in the cats of the treatment group.

[0113] From D7 to D63, clinical examinations (weight, temperature) of the cats were performed once a week, and urine and fecal samples were collected.

[0114] The samples were analyzed using a non-classical analytical technique based on tandem mass spectrometry, also known as MS / MS or MS2, to search for the molecules responsible for the odor of the urine and fecal samples. This unique technique evaluates the metabolomics present in the samples.

[0115] The plasma levels of testosterone are also being evaluated for the chemically treated group and the control group.

[0116] Metabolomics analysis was performed on urine and fecal samples collected before treatment (D-14), allowing comparison of the groups and confirmation of the same levels of molecules observed between the two groups (confirmation was performed, data not shown).

[0117] Also, to compare the molecular levels between treated and control animals and evaluate the effect of castration on the odor molecule levels, metabolomics analysis was performed on feces at D49 and urine at D63 after treatment.

[0118] A second study B was conducted on two groups of four healthy male cats, 12 to 13 months old and weighing 4 to 6 kg. Control group: n = 2, non-castrated males, administered sodium chloride (physiological saline) on day 0 (D0). Surgical castration group: n = 4 (surgical castration by orchiectomy; adult cats castrated for over 6 months), administered sodium chloride (physiological saline) on day 0 (D0).

[0119] The protocol was the same as that of the first study A.

[0120] LC-MS / MS analysis Metabolomics analysis was performed using LC-MS / MS.

[0121] Liquid chromatography analysis was carried out using a DIONEX Ultimate3000 HPLC system (Thermo Fisher Scientific). 10 μL of each sample was injected into a Synergi 4 μm Hydro-RP 80Å, 250×3.0 mm column (Phenomenex, France, Le Pecq). The mobile phase was composed of 0.1% formic acid (Thermo Fisher Scientific) water (Study A) and acetonitrile containing 0.1% formic acid (Study B). The gradient was set to maintain 0% phase B from 0 to 5 minutes at a flow rate of 0.9 mL / min, 0 - 95% B from 5 to 21 minutes, hold at 95% B until 21.5 minutes, 95 - 0% B from 21.5 to 22 minutes, and hold at 0% B until 25 minutes for column equilibration. Mass spectrometry was performed using a Q Exactive Plus Orbitrap mass spectrometer (Thermo Fisher Scientific) equipped with a heated electrospray ionization source (HESI II) operating in positive / negative ion exchange mode. Further analysis of the samples was performed in positive mode. High-resolution accurate mass full scan MS and the top 5 MS2 spectra were collected data-dependently with a resolution of 70,000 and 35,000 at m / z 400 respectively. All samples were processed continuously in the same run.

[0122] Metabolomics analysis Compound Discoverer 3.2 (Thermo Fisher Scientific) was used for post-processing of the data.

[0123] The workflow template was non-targeted metabolomics. Statistics were performed to detect unknown substances with IDs using an online database, and mzLogic.ons obtained from positive and negative ionization modes were also analyzed using MZmine (version 2.39).

[0124] Mass detector tool (mass detector: wavelet transform, MS level 1; noise level 10 5, Scale level: 5, Wavelet window size: 30%) were used to perform mass detection. ADAP chromatogram builder (MS level: 1, Minimum group size of scan number: 5, Group intensity threshold: 5x10 2 , Minimum highest intensity: 10 5 , m / z tolerance: 10 ppm) were used to detect the chromatogram. Peak extender module (M / Z tolerance: 10 ppm, Minimum height: 10 5 ) was used to separate the peaks. Retention time correction module (m / z tolerance: 10 ppm; Retention time tolerance (relative): 10%, Minimum height: 10 5 ) was used to standardize the retention time. Then, with a tolerance of m / z at 10 ppm and a retention time of 1 minute, the RANSAC aligner (Random Sample Consensus) algorithm was used to align the peaks. Then, the Human Metabolome Database (HMDB, version 3.0) was used to identify the peaks with a mass tolerance of 10 ppm. The same m / z and RT range gap filling were used to fill in the missing values with a tolerance of 10 ppm for m / z. The results obtained for each polarity were combined, and for the metabolites identified in both modes, only the mode in which the peak had the highest average intensity was considered.

[0125] 2.2 Results Ferritin: The results are reported in the attached Figure 4. In this figure, the average relative abundance peak of ferritin present in the urine samples of male cats collected on D63 after administration of saline (control group) is compared with that of cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group), and surgically castrated male cats (surgical castration group).

[0126] The average relative abundance peaks of ferritin in both control groups of Study A and Study B were the same, and the results were combined and shown in a graph (Figure 4).

[0127] These results indicate that on day 63, ferritin decreased significantly in both the chemical treatment group and the surgical castration group compared to the control group.

[0128] MBCG: Report the results in the attached FIGS. 5A and 5B. In FIG. 5A, the average relative abundance of MBCG present in urine samples from male cats collected on D63 after administration of saline (control group) is compared with that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group). In FIG. 5B, the average relative abundance of MBCG present in urine samples from male cats collected on D63 after administration of saline (control group) is compared with that in surgically castrated male cats (surgical castration group).

[0129] These results indicate that on day 63, MBCG was significantly decreased in both the chemical treatment group and the surgical castration group compared to the control group.

[0130] Example 3: Evaluation of Molecular Level Alterations Related to Non-Sex Hormone Dependent Pathways The intent of this study was to evaluate whether the use of deslorelin could modulate or alter, downregulate or upregulate the levels of molecules identified as causes of urine odor.

[0131] The intent of this study was also to demonstrate that the use of deslorelin could also modulate the level of crotonic acid in feces. Crotonic acid is one of the molecules that causes fecal odor.

[0132] 3.1 Materials and Methods: The study was the same as that conducted according to the protocol of Example 2, using the same group of cats.

[0133] As already detailed in Example 2, samples were analyzed using a non-classical analytical technique based on tandem mass spectrometry, also known as MS / MS or MS2, to search for odor-causing molecules in urine and fecal samples. This unique technique evaluates the metabolomics present in the samples.

[0134] The following metabolomics levels were measured in urine collection samples of each group during the study. N-acetylcadaverine, N-acetylputrescine, p-coumaroylputrescine, taurine, valine, indole, isovaline, and mercapto derivatives, namely mercaptopyruvic acid, mercaptopropionic acid, 3-mercaptoheptanoyl threonine, and 2-mercapto-2-methylpentan-4-one, all of which are related to non-sex hormone-dependent pathways. These molecules have been identified as one of the molecules causing the smell of urine.

[0135] During the study, crotonic acid levels were measured in fecal collection samples of each group.

[0136] Perform dissociation methods or chemical reactions to change the mass or charge of ions while performing mass spectrometry at least twice by tandem mass spectrometry (MS / MS or MS 2 )). MS / MS improves specificity beyond the separation range by molecular weight, provides insight into structure elucidation, and identifies elemental composition. The most common dissociation method is collision-induced dissociation (CID), where the first analyzer is used to isolate specific target ions (precursor ions), which then undergo a fragmentation method to produce neutral fragments and fragment ions (product ions). The second analyzer analyzes these product ions and forms a unique pattern based on the precursor ion structure and components (Richard A. McPherson, Methods in Enzymology, 2019).

[0137] MS / MS is particularly useful for analyzing complex mixtures and involves two-stage MS. In the first stage of MS / MS, a given series of m / z ions are isolated from other ions from the ion source and fragmented by chemical reactions. In the second stage, a mass spectrum is generated for the fragments. Tandem MS is generally used for the bioanalysis of drugs. Phase I and Phase II drug metabolites are identified and determined by tandem MS combined with HPLC (Glish, G.L. and Vachet, R.W., 2003. Nature Reviews Drug Discovery 2(2):140; Holcapek, M. et al., 2008., Analytical and Bioanalytical Chemistry 391(1):59-78).

[0138] 3.2 Research Results: N-acetylcadaverine: The results are reported in the attached Figure 6. In this figure, the mean relative abundance of N-acetylcadaverine present in urine samples from male cats collected on D63 after administration of physiological saline (control group) is compared to that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group) (Figure 6A), and surgically castrated male cats (surgical castration group) (Figure 6B).

[0139] These results indicate that N-acetylcadaverine, which is known to have the odor of putrid meat on day 63, was significantly decreased in the chemical treatment group compared to the control group, while the N-acetylcadaverine concentration measured in urine samples from the surgical castration group was not significantly different from that of the control group.

[0140] N-acetylputrescine: The results are reported in the attached Figure 7. In this figure, the mean relative abundance of N-acetylputrescine present in urine samples from male cats collected on D63 after administration of physiological saline (control group) is compared to that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group).

[0141] The results demonstrate that N-acetylputrescine, which is known to have the odor of putrid meat on the 63rd day, was significantly reduced in the chemical treatment group according to the present invention compared to the control group.

[0142] p-Coumaroylputrescine: The results are reported in the attached Figure 8. In this figure, the progression of the average relative abundance of p-coumaroylputrescine is represented according to time (days) for the control group (continuous curve) and the chemical treatment group (discontinuous curve).

[0143] These results indicate that the administration of deslorelin induces a decrease in the p-coumaroylputrescine concentration (chemical treatment group) compared to the control group.

[0144] Taurine: The results are reported in the attached Figure 9. In this figure, the average relative abundance of taurine present in urine samples of male cats collected on D63 after administration of physiological saline (control group) is compared with that of cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group).

[0145] The results demonstrate that on the 63rd day, taurine was significantly reduced in the chemical treatment group according to the present invention compared to the control group.

[0146] Valine: The results are reported in the attached Figure 10. In this figure, the average relative abundance of valine present in urine samples of male cats collected on D42 after administration of physiological saline (control group) is compared with that of cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group) (Figure 10A), and that of surgically castrated male cats (surgical castration group) (Figure 10B).

[0147] These results indicate that on the 42nd day, valine, which is known to have a pleasant sweet odor, was significantly increased in the chemical treatment group compared to the control group, while the valine concentration measured in urine samples of the surgical castration group showed no significant difference compared to the control group.

[0148] Indole: Report the results in the attached Figure 11. In this figure, compare the average relative abundance of indole present in urine samples from male cats collected on D63 after administration of physiological saline (control group) with that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group) (Figure 11A), and in surgically castrated male cats (surgical castration group) (Figure 11B).

[0149] These results indicate that on day 63, indole, which is known to have a strong fecal odor, was significantly reduced in the chemical treatment group compared to the control group, while the indole concentration measured in urine samples from the surgical castration group showed no significant difference compared to the control group.

[0150] Isovaline: Report the results in the attached Figure 12. In this figure, compare the average relative abundance of isovaline present in urine samples from male cats collected on D63 after administration of physiological saline (control group) with that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group).

[0151] The results demonstrate that on day 63, isovaline, which is known to have a foot / cheese odor, was reduced in the chemical treatment group according to the present invention compared to the control group.

[0152] Mercaptopyruvic acid: Report the results in the attached Figure 13. In this figure, compare the average relative abundance of mercaptopyruvic acid present in urine samples from male cats collected on D63 after administration of physiological saline (control group) with that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group) (Figure 13A), and in surgically castrated male cats (surgical castration group) (Figure 13B).

[0153] These results indicate that on day 63, mercaptopyruvic acid, which is known as an odoriferous compound, was significantly decreased in the chemically treated group compared to the control group, while there was no significant difference in the concentration of mercaptopyruvic acid measured in the urine samples of the surgically castrated group compared to the control group.

[0154] 3-Mercaptopropionic acid: The results are reported in the attached Figure 14. In this figure, the average relative abundance of 3-mercaptopropionic acid present in urine samples of male cats collected on D63 after administration of physiological saline (control group) is compared with that of cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemically treated group).

[0155] The results demonstrate that on day 63, 3-mercaptopropionic acid, which is known as an odoriferous compound, was significantly decreased in the chemically treated group according to the present invention compared to the control group.

[0156] 7-Mercaptoheptanoyl threonine: The results are reported in the attached Figure 15. In this figure, the average relative abundance of 7-mercaptoheptanoyl threonine present in urine samples of male cats collected on D63 after administration of physiological saline (control group) is compared with that of cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemically treated group) (Figure 15A), and that of surgically castrated male cats (surgically castrated group) (Figure 15B).

[0157] These results indicate that on day 63, 7-mercaptoheptanoyl threonine was significantly decreased in the chemically treated group and the surgically castrated group compared to the control group.

[0158] 2-Mercapto-2-methylpentan-4-one: Report the results in the attached Figure 16. In this figure, compare the average relative abundance of 2-mercapto-2-methylpentan-4-one present in urine samples from male cats collected on D63 after administration of physiological saline (control group) with that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group).

[0159] The results demonstrate that, on day 63, 2-mercapto-2-methylpentan-4-one, which is known to have a very pleasant onion-like odor, was significantly increased in the chemical treatment group according to the present invention compared to the control group.

[0160] Crotonic acid: Report the results in the attached Figure 17. In this figure, compare the average relative abundance of crotonic acid present in fecal samples from male cats collected on D63 after administration of physiological saline (control group) with that in cats implanted with one 4.7 mg deslorelin implant (Suprelorin®) (chemical treatment group).

[0161] The results demonstrate that, on day 63, crotonic acid, which is known to have an unpleasant odor, was significantly decreased in the chemical treatment group compared to the control group.

[0162] All results regarding the measurement of mercapto derivatives in urine samples indicate (for the first time) that the concentration of mercapto derivatives in urine is affected by physical castration and deslorelin treatment. Without wishing to be bound by any theory, the inventors of the present invention presume that these changes are not directly related to testosterone because the changes are different (e.g., mercaptopyruvic acid) in surgically castrated cats and deslorelin-castrated cats. Also, the decrease in urine odor, and thus marking, due to the decrease in mercapto derivatives appears to be more significant in deslorelin-treated cats compared to surgically castrated cats.

[0163] These results also demonstrate (shown for the first time) that other tested odor compounds in urine (unpleasant or favorable odors) are not affected by surgical castration, but are affected by castration with deslorelin treatment. Therefore, these changes do not appear to be directly related to the testosterone pathway, as the changes are different in surgically castrated cats and deslorelin-castrated cats. Also, the decrease in urine odor, and thus marking, due to the decrease of compounds other than mercapto derivatives, appears to be more important in deslorelin-treated cats compared to surgically castrated cats.

[0164] These data surprisingly demonstrated that deslorelin affects the regulation of the levels of several metabolic molecules. In particular, deslorelin downregulates the levels of the following molecules, cadaverine, indole, p-cresol, isovaleric acid, and isobutyric acid, either individually or at least two of them together.

[0165] These results indicate that deslorelin affects the molecular levels related to non-sex-hormone-dependent pathways. This data shows that the odor of urine and / or feces is also determined by molecules not related to the sex hormone pathway.

[0166] As a conclusion, it is very interesting to note that the molecules causing unpleasant odors are downregulated, while the molecules causing favorable odors are upregulated.

[0167] Example 4: Field study in cats 4.1 Research protocol The intention of these studies was to demonstrate the suppression of fertility and the persistence of infertility in male cat animals.

[0168] This example combines two studies (double-blind placebo-controlled randomized) conducted continuously according to the following schedule and provides the collected data. Study 1: Study 1 was conducted by dividing a group of 205 male cats into two groups. One was a control group that received 1 mL of physiological saline (n = 51, subcutaneous administration), and the other was a treatment group that received an implant of 4.7 mg of Suprelorin® (subcutaneous implant, deslorelin acetate) (n = 154). Study 2: At the end of the first 12-month period, all male cats in the treatment group had their Suprelorin® implants removed. Subsequently, 12 male cats from the treatment group of Study 1 were administered physiological saline (1 mL subcutaneously) (physiological saline group), and 22 cats from the treatment group of Study 1 were re-implanted with a new 4.7 mg Suprelorin® implant during the second 12-month period (treatment twice group). The effectiveness and safety of the re-implantation were followed up for these additional 12 months, that is, a total of 24 months.

[0169] The observation schedule was as follows. Study 1: V1: from -14 to -7 days; V2: day 0; V3: 1.5 months ± 3 days; V4: 3 months ± 5 days, V5: 6 months ± 5 days, V6: 9 months ± 5 days; V7: 12 months ± 5 days; Study 2: V8: 12 months ± 3 days; V9: 13.5 months ± 3 days; V10: 15 months ± 5 days; V11: 18 months ± 5 days; V12: 21 months ± 5 days; V13: 24 months ± 5 days.

[0170] Regarding the inclusion criteria, in both studies, the criteria were as follows. - Domestic short-haired cats - Age > 3 months or > 15 months in Study 2 - Intact male cats kept indoors - Cats owned by the requester (including cats in protection facilities / cats taken in by foster families) The additional criterion for Study 2 was success in Study 1.

[0171] In these studies, the following primary evaluation items were measured. - Study 1: Suppression of fertility (testosterone ≤ 0.10 ng / mL from 3 to 12 months *from V4 to V7*). - Study 2: Suppression of fertility (*testosterone ≤ 0.10 ng / mL from V8 to V13 (12 to 24 months)) and time to effect reversal.

[0172] In addition to safety parameters (hematology and blood biochemistry, urine tests, local reactions at the implant site, body weight changes, appetite, abnormal testicular evaluation, adverse events), the following secondary evaluation items are also reported. - Total sexual behavior score - Aggressiveness - Crying - Urine marking - Odor / smell of urine - Penile spine - Testicular volume

[0173] 4.2 Results - Testosterone: The results of the mean efficacy are shown in Figure 18. Overall, the success rates of both studies were very similar for 1 or 2 implants, and the efficacy was > 87% at all time points in Study 2.

[0174] The detailed levels of testosterone are shown in Figure 19A for Study 1 and Figure 19B for Study 2, respectively.

[0175] - Reversibility: The reversibility of the deslorelin effect is being studied. In fact, among the deslorelin treatment groups in Study 1, as described in 4.1 above, n = 12 subjects were recorded in the saline group of Study 2. The results showed that 100% of the cats that were in the treatment group in Study 1 and the saline group in Study 2 reversed at D279 after being recorded in Study 2. The median time for reversibility based on serum testosterone concentration measurement was D379 ± 178.

[0176] - Secondary evaluation items: The efficacy results of the secondary evaluation items are reported as follows. · The mean values of the total sexual behavior scores are shown in Figure 20A for Study 1 and Figure 20B for Study 2, respectively. ·The average vocalization variation is shown in Figure 21A for Study 1 and in 21B for Study 2. ·The percentage change in urine marking from baseline is shown in Figure 22A for Study 1 and in Figure 22B for Study 2, respectively. ·The urine odor variation / decrease measured as the change from baseline is shown in Figure 23A for Study 1 and in Figure 23B for Study 2, respectively. ·The percentage change in testis volume is shown in Figure 24A for Study 1 and in Figure 24B for Study 2, respectively. ·The percentage change in the penile spines that appeared is shown in Figure 25A for Study 1 and in Figure 25B for Study 2, respectively.

[0177] During the course of the study, there were no safety concerns, and the deslorelin implant showed good tolerance.

[0178] In short, combining these studies demonstrates the following. Deslorelin was effective for at least 12 months when chemically castrating the subjects. This effect has been proven to be reversible at the end of the treatment process. These studies also demonstrate that sexual behavior decreases due to the rapid onset of the effectiveness of behavior reduction (about 7 - 31 days). Also, there were no safety concerns regarding single or repeated administrations.

[0179] Therefore, it can be concluded that deslorelin administered in sustained release, especially by implant, is an excellent alternative to surgical castration and a safe non - surgical method for controlling the reproduction of male cats.

[0180] The method of using deslorelin as described herein is a reversible, non - surgical, reliable castration measure that promotes the health of the pet family while leaving options.

Claims

1. Deslorerin for use in reducing or decreasing the odor of urine and / or feces in non-human animals, wherein the plasma concentration of deslorerin in the non-human animals after administration is higher than 10 pg / mL.

2. The deslorerin to be used according to claim 1, wherein, after administration to the non-human animal, the plasma concentration of deslorerin is 10 pg / mL to 400 pg / mL from 8 days after administration to at least 6 months after administration.

3. The deslorerin used according to claim 2, wherein deslorerin is administered continuously to the non-human animal or released continuously after administration.

4. The deslorerin to be used according to claim 3, wherein, after administration, the deslorerin is released substantially continuously at a plasma concentration level of less than 400 pg / mL from 8 days after administration to at least 6 months after administration.

5. The deslorerin used according to claim 3, wherein, after administration, the deslorerin controls down or up molecules that cause odor in urine and / or feces, and the molecules are selected from the group including ferrinine derivative compounds, volatile organic compounds (VOCs), sulfur compounds, and testosterone-inducing compounds.

6. After administration, deslorerin reduces the molecules that cause the odor of urine and / or feces, and these molecules - Sulfur compounds selected from the group including 4-methyl-4-sulfanyl-pentan-2-one; 3-methyl-3-sulfanyl-butan-1-ol; and 3-methyl-3-sulfanyl-1-butyl formate; - VOCs: 2,5-dimethylpyrazine; 1,2-dichloropropane; 4,4-dimethyl-2-pentanone; and 3-methyl-2-buten-1-ol; - Compounds selected from the group including 2-(methylthio)-1-ethanol; 2-methyl-3-frantiol; oxathiolane; 4-(methylthio)-2-butanone; 3-methylbutyl-3-sulfanyl formate; and 3-methyl-3-(methylthio)-1-butanol; - Paracresol - Felinine and its derivatives such as 3-mercapto-3-methyl-1-butanol; 3-mercapto-3-methylbutyl formate; 3-methyl-3-methylthio-1-butanol; and 3-methyl-3-(2-methyldisulfanyl)-1-butanol; - Felinine precursors such as N-acetylfelinine, γ-glutamylfelinylglycine, and 3-methylbutanolcysteinylglycine; - Corkin; - 2-phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isobaltin; - Isobutein; - Trimethylamine; - Indole; - Cadaverine and derivatives such as N-acetylcadaverine and glutathioneylaminopropylcadaverine; - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid; - Crotonic acid; - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvate, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoylthreonine, 1-alpha,5-alpha-dimercaptoandrostan-3-alpha,17-beta-diol, mercaptopyruvate, 2-mercapto-2-methylpentan-4-one; and - Taurine Deslorerin to be used according to claim 5, selected from the group including the group.

7. Deslorerin to be used according to claim 6, wherein the molecule is selected from the group comprising indole, cadaverine and its derivatives such as N-acetylcadaverine, putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine, and mercaptopyruvic acid.

8. The deslorerin to be used according to claim 5, wherein, after administration, the deslorerin upregulates molecules that cause odor in urine and / or feces, and the molecules are selected from the group comprising 2-mercapto-2-methylpentan-4-one and valine.

9. Non-therapeutic use of deslorerin to downcontrol the level of a molecule in the urine and / or feces of a non-human animal, wherein the molecule is - Sulfur compounds selected from the group including 4-methyl-4-sulfanyl-pentan-2-one; 3-methyl-3-sulfanyl-butan-1-ol; and 3-methyl-3-sulfanyl-1-butyl formate; - VOCs: 2,5-dimethylpyrazine; 1,2-dichloropropane; 4,4-dimethyl-2-pentanone; and 3-methyl-2-buten-1-ol; - Compounds selected from the group including 2-(methylthio)-1-ethanol; 2-methyl-3-frantiol; oxathiolane; 4-(methylthio)-2-butanone; 3-methylbutyl-3-sulfanyl formate; and 3-methyl-3-(methylthio)-1-butanol; - Paracresol; - Felinine and its derivatives such as 3-mercapto-3-methyl-1-butanol; 3-mercapto-3-methylbutyl formate; 3-methyl-3-methylthio-1-butanol; and 3-methyl-3-(2-methyldisulfanyl)-1-butanol; - Felinine precursors such as N-acetylfelinine, γ-glutamylfelinylglycine, and 3-methylbutanolcysteinylglycine; - Corkin; - 2-phenylethylamine and S-(1-hydroxy-3,7-dimethyl-6-octen-3-yl)cysteine; - Isobaltin; - Isobutein; - Trimethylamine; - Indole; - Cadaverine and cadaverine derivatives such as N-acetylcadaverine and glutathioneylaminopropylcadaverine; - Short-chain free fatty acids such as acetic acid, propanoic acid, 2-methylpropanoic acid, butanoic acid, 3-methylbutanoic acid, and pentanoic acid; - Crotonic acid; - Putrescine and its derivatives such as N-acetylputrescine and p-coumaroylputrescine; - Mercapto derivatives such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexyl butyrate, mercaptopyruvate, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoylthreonine, 1-alpha,5-alpha-dimercaptoandrostan-3-alpha,17-beta-diol, and mercapto derivatives such as mercaptopyruvate, 2-mercapto-2-methylpentan-4-one; and - Taurine Non-therapeutic use, selected from the group including [specific group].

10. Non-therapeutic use of deslorerin for up-controlling molecular levels in the urine and / or feces of non-human animals, wherein the molecular is selected from the group comprising 2-mercapto-2-methylpentan-4-one and valine.

11. A method for reducing or decreasing the odor of urine and / or feces in a non-human animal, the method comprising administering to the non-human animal an amount of deslorerin such that, after administration to the non-human animal, the plasma concentration of deslorerin in the non-human animal is greater than 10 pg / mL.

12. A method for reducing or decreasing the odor of urine and / or feces in a non-human animal, the method comprising administering to the non-human animal an amount of deslorerin that induces a downward or upward control of the level of odor molecules in the urine and / or feces.

13. The method according to claim 11 or 12, wherein the non-human animal is an uncastrated male cat.

14. The method according to claim 11 or 12, wherein deslorerin induces downward or upward control of odor receptor levels in urine and / or feces via the sex hormone synthesis pathway.

15. A method for improving the reduction or decrease of the odor of urine and / or feces in non-human uncastrated male animals compared to castrated males, the method comprising administering to the non-human uncastrated male animals an amount of deslorerin that induces downward or upward control of the level of odor molecules in the urine and / or feces.

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