Use of deserelin in chemical castration of non-human mammals associated with PK / PD interactions

Abstract

The present invention relates to deserelin in a drug or non-drug for chemical castration of a non-human mammal within 1 month after administration wherein after administration, the deserelin has a Cmax (about 1 to 2 hours after administration) of more than 7000 pg / mL and a concentration of deserelin in plasma of the non-human mammal of at least 10 pg / mL. The invention also relates to a sustained release drug delivery system for implementing such uses and other embodiments of the invention. The present invention also provides a method for chemical castration of a non-human animal, comprising the steps of: administering an effective amount of deserelin such that the Cmax concentration of deserelin or an equivalent amount of a pharmaceutically acceptable salt thereof in the resulting plasma within the first 24 hours after injection is 4000 pg / mL to 4000 pg / mL; and thereafter, (b) administering a sustaining dose such that the average concentration of deserelin or an equivalent amount of a pharmaceutically acceptable salt thereof in the plasma is higher than 10 pg / ml.

Description

Technical Field

[0001] The present disclosure generally relates to pharmaceutical compositions of deserorelin for chemical castration of non-human mammals, preferably companion animals. Background Art

[0002] Deserorelin is a gonadotropin analog of gonadotropin-releasing hormone (GnRH) that acts by suppressing pituitary-gonadal axis function when administered continuously at low doses.

[0003] GnRH is a hypothalamic decapeptide that acts at the apex of a cascade that coordinates the function of the hypothalamic-pituitary-gonadal axis. GnRH binds to the GnRH receptor (a 7-transmembrane receptor), causing the pituitary to produce and release two key gonadotropins, follicle-stimulating hormone (FSH) and luteinizing hormone (LH), which in turn control gonadal function:

[0004] - LH (luteinizing hormone), which, in addition to its role in gametogenesis, promotes the production of sex steroids (androgens, estrogens and progesterone);

[0005] - FSH (Follicle Stimulating Hormone) stimulates the production of gametes (sperm and eggs).

[0006] When GnRH binds to the receptor, it increases the synthesis of its own receptor mRNA and increases the sensitivity of the pituitary to GnRH. Continued secretion of GnRH has the opposite effect (a decrease in LH and FSH secretion is observed: this can be called desensitization). (Finch AR et al., Agonist-induced internalization and downregulation of gonadotropin-releasing hormone receptors. Am. J. Physiol. Cell Physiol. 2009 Sep., 297(3)).

[0007] Internalization of the GnRH receptor (it is no longer bound to the cell surface) and inactivation of the intracellular signaling cascade triggers desensitization of the pituitary to GnRH. Once pituitary cells begin to desensitize to GnRH, LH concentrations drop to undetectable levels, making it unable to support testosterone and sperm production.

[0008] To solve the contraceptive problem in animals, some people came up with the idea of ​​using GnRH agonists as contraceptives for a long time. This is where the idea of ​​designing a deserorelin implant that continuously diffuses a GnRH agonist (deserorelin) came from.

[0009] This type of implant can be used in two different ways:

[0010] - Fertility induction (short-term implantation): Ovuplant

[0011] - Inducing infertility through long-term action; Suprelorin

[0012] It is known in the prior art that these agonist-based implants have two phases of action. During the injection of the implant, there is a release of deserorelin, which induces a strong stimulation of the LH and FSH production cascade, a step known as the "burst". In male dogs, this induces the production of LH and FSH, leading to a strong synthesis of testosterone. Males then gain super fertility (improved sperm quality, increased motility).

[0013] After this "burst" period, an infertile period is established with adverse effects on all aspects of sperm quality and testosterone concentrations (Goericke-Pesch S., Long-term effects of GnRH agonists on fertility and behavior., Reprod Domest Anim., 2017, 52 Suppl 2:336-347).

[0014] In addition to the trials in dogs, studies have also been conducted in cats and kittens (Fontaine C., Long-term contraception in a small implant: A review of Suprelorin (deslorelin) studies in cats., J Feline Med Surg., 2015, 17(9), pp. 766-771). This showed that the implant is effective as a contraceptive in both males and females. During the period of fertility suppression, steroid hormone concentrations of less than 1 ng / ml progesterone and 10 pg / ml estradiol were detected in female cats; in male cats, plasma testosterone concentrations remained at basal levels (<0.1 ng / ml). The duration of action of the 4.7 mg implant in male cats was 78.8 weeks ± 12.9 weeks, with a range of 61.7 weeks (approximately 15 months) to 100.7 weeks (approximately 25 months).

[0015] Other studies have been conducted in leopards, rats, coyotes, and other animals.

[0016] According to the prior art, deserorelin has a contraceptive effect in females, although this effect varies according to the species.

[0017] As far as is known, deslorelin is an effective long-term contraceptive in bitches, but it can cause adverse effects of estrus in the short term. The contraceptive effect can be followed by analyzing whether the progesterone concentration is above 24 ng / ml during the luteal phase (BrändliSp et al., Long-term effect of repeated deslorelin acetate treatment in bitchesfor reproduction control., Theriogenology. 2021 Oct 1, 173, pp. 73-82).

[0018] In order to solve the problem of estrus induction, the test implanted this implant into prepubertal female dogs. The effectiveness of the implant was measured by detecting 17-β estradiol (E2) and progesterone (P4) by luciferase assay. The implant reduced these hormones (basal level exceeded 10 ng / ml): P4 concentration was 0.3 ng / ml to 1.1 ng / ml (0.6 ng / ml ± 0.2 ng / ml), and E2 concentration was 8 pg / ml to 138 pg / ml. Therefore, it is known that deslorelin is an early effective contraceptive for prepubertal female dogs, which can delay puberty for at least 4.5 months, but the effect needs to be maintained by repeated treatment (G Marino et al., deslorelin implants in pre-pubertal female dogs: short- and long-term effects on the genital tract. Reprod. Domest. Anim., 2014), 49 (2), 297-301 pages).

[0019] It is also known to have a contraceptive effect in cats, for which ovarian activity was assessed by monitoring fecal E2 (estradiol) concentrations. The implant suppressed estrus behavior and reduced estradiol secretion in all cats (mean 128.48 ng / g; control group: 283.26 ng / g) for approximately 668 days (23 months) (TSF Toydemir et al., Effects of the GnRHanalogue deslorelin implants on reproduction in female domestic cats. Theriogenology. 2012 Feb, 77(3), pp. 662-674; and Fontaine C. 2015 cited above.).

[0020] In ferrets, the 4.7 mg deserorelin implant also showed contraceptive efficacy, with the same side effect throughout: estrus. Thus, ferrets showed strong signs of estrus within 4 days after implantation, accompanied by swelling of the vulva. Two weeks after implantation, estrus ceased spontaneously. However, fecal progesterone concentrations in these animals remained low throughout the 8.5-month sampling period, indicating that they did not ovulate. The suppressive effect of the deserorelin implant on ovarian function lasted for 698 (± 122) days (A Proháczik et al., Comparison of four treatments to suppress ovarian activity in ferrets (Mustela putorius furo)., Vet. Rec., 2010 Jan 16, 166(3), pp. 74-78).

[0021] This implant has even been tested in large mammals, such as female tigers. In fact, this implant makes it possible to reversibly regulate the reproduction of female tigers in certain cases by injecting several implants. The average time to conception was 50.7 months (4.7 mg implant) and 51.9 months (9.4 mg implant) (A Guthrie et al., The past, present and future of hormonal contraceptive use in managed captive female tiger populations with a focus on the current use of deslorelin acetate., ZooBiol., 2021, 40(4) 306-319 pages).

[0022] Basic principle: So far, the known data do not show a PK / PD (pharmacokinetic / pharmacodynamic) relationship, and the efficacy of the product can be explained by a huge "burst" shortly after implantation and a long duration of GnRH receptor saturation. Current data in dogs show that deserorelin exists for 2.5 months, and this effect is observed to last at least 6 months (source: EMA website: https: / / www.ema.europa.eu / en / documents / product-information / suprelorin-epar-product-information_en.pdf).

[0023] Now, new data have been generated that show that deserorelin can be detected as long as this effect is observed in male and female dogs and cats.

[0024] Thus, it has now been demonstrated that a "burst" (also referred to herein as an "attack") effect is necessary to induce castration after a period of superfertility in non-human animals that have not been chemically castrated previously. And that maintaining very low concentrations of deserorelin (above a certain threshold) for a long period of time is necessary to maintain the chemical castration effect. Contrary to the prior art and the prevailing belief about the mode of action of deserorelin, a PK / PD relationship between the in vivo levels of deserorelin and the PD effect has also been clearly demonstrated, allowing us to determine a threshold for efficacy.

[0025] Thus, it has been demonstrated that in chemically castrated non-human animals, a "burst" (also referred to herein as an "attack") effect is not necessary, but that maintaining a long-term concentration of deserorelin above a certain threshold is necessary. Furthermore, it has been shown that maintaining a concentration range of deserorelin in the blood is necessary to obtain and maintain a castration effect. Contrary to the prior art and the prevailing belief about the mode of action of deserorelin, a PK / PD relationship between the in vivo levels of deserorelin and the PD effect has also been clearly demonstrated, allowing us to determine a threshold for efficacy. Summary of the invention

[0026] The present invention relates to deserorelin in a drug or non-drug for chemical castration of a non-human mammal within 1 month after administration, wherein after administration, the Cmax of deserorelin (preferably about 1 hour to 2 hours after administration) exceeds 7000 pg / mL, and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL. In a preferred embodiment, deserorelin is administered subcutaneously or intramuscularly to a non-human mammal in need thereof, at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a plasma concentration of less than 400 pg / ml from 8 days after administration.

[0027] In a particular embodiment of use, the deserorelin plasma concentration is 10 pg / mL to 400 pg / ml from 8 days to at least 6 months after administration.

[0028] In another embodiment, deserorelin is used to induce temporary infertility in healthy male animals over a period of several months, preferably for a period of at least 6 months, comprising administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a plasma concentration of less than 400 pg / ml from 8 days after administration. In a preferred embodiment, the deserorelin plasma concentration is less than 400 pg / ml from 8 days to at least 6 months after administration. Preferably, deserorelin is substantially continuously released at a plasma concentration of less than 400 pg / mL from 8 days to at least 6 months after administration.

[0029] In another embodiment, the present invention relates to a sustained-release drug delivery system (DDS) containing deserorelin for the use described above, which is administered to a non-human mammal by a subcutaneous or intramuscular route, wherein the sustained-release DDS releases more than 15%, preferably more than 20%, preferably more than 25%, preferably more than 30%, preferably more than 35%, preferably more than 40%, preferably more than 45%, and even preferably more than 50% of the deserorelin contained in the DDS within 2 days, and the remaining effective dose of deserorelin is released within 3 months to 6 months or more than 6 months. In a preferred embodiment, the DDS is a subcutaneous implant or intramuscular implant comprising deserorelin and a biodegradable polymer, which is used for chemical castration of a non-human mammal, wherein the implant releases deserorelin at a deserorelin Cmax of more than 7000 pg / mL after administration (preferably about 1 hour or 2 hours after administration), and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL and is maintained for at least 6 months. In a preferred embodiment, deserorelin is administered to a non-human mammal in need thereof at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a plasma concentration below 400 pg / ml starting from 8 days after administration, and preferably for 8 days to at least 6 months after administration.

[0030] In another embodiment, the present invention relates to a subcutaneous or intramuscular implant comprising deserorelin and a biodegradable polymer for inducing temporary infertility in healthy male animals for a period of at least 6 months, which comprises administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, at a dose effective to induce a plasma concentration of deserorelin of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a plasma concentration of less than 400 pg / ml from 8 days after administration.

[0031] In another embodiment, the invention relates to deserorelin for chemical castration of a non-human mammal, wherein after administration, deserorelin reduces the testosterone level in the plasma of the non-human mammal to below 0.4 ng / ml and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / ml, preferably above 15 pg / ml.

[0032] According to another embodiment, the present invention provides a method for chemical castration of a non-human animal comprising the steps of:

[0033] (a) administering an effective amount of deserorelin such that the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is between 7000 pg / mL and 40000 pg / mL within the first 24 hours after injection; and thereafter,

[0034] (b) administering a maintenance dose such that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0035] Preferably, in step (a) and / or step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In a specific embodiment, in step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). In a preferred embodiment, the sustained-release drug delivery system (DDS) of step (a) is a subcutaneous implant or an intramuscular implant. Preferably, the implant is a biodegradable polymer.

[0036] Another embodiment of the present invention relates to a method for inducing temporary infertility in a healthy, sound, sexually mature male animal, preferably a dog, comprising the steps of:

[0037] (a) administering an effective amount of deserorelin such that the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is between 7000 pg / mL and 40000 pg / mL within the first 24 hours after injection; and thereafter

[0038] (b) administering a maintenance dose such that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0039] Preferably, in step (a) and / or step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In a specific embodiment, in step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). In a preferred embodiment, the sustained-release drug delivery system (DDS) of step (a) is a subcutaneous implant or an intramuscular implant. Preferably, the implant is a biodegradable polymer.

[0040] Another embodiment of the present invention relates to a method for inducing temporary infertility in a sound and healthy sexually immature female animal, preferably a dog, to delay the first estrus and estrus signs and prevent pregnancy at an early age, comprising the steps of:

[0041] (a) administering an effective amount of deserorelin such that the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is between 7000 pg / mL and 40000 pg / mL within the first 24 hours after injection; and thereafter

[0042] (b) administering a maintenance dose such that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0043] Preferably, in step (a) and / or step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In a specific embodiment, in step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). In a preferred embodiment, the sustained-release drug delivery system (DDS) of step (a) is a subcutaneous implant or an intramuscular implant. Preferably, the implant is a biodegradable polymer.

[0044] Another embodiment of the present invention relates to a method of inducing temporary sterility, suppressing urine odor, and suppressing sexual behaviors such as libido, vocalization, urine marking, and aggression in healthy male animals, preferably cats, comprising the steps of:

[0045] (a) administering an effective amount of deserorelin such that the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is between 7000 pg / mL and 40000 pg / mL within the first 24 hours after injection; and thereafter

[0046] (b) administering a maintenance dose such that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0047] Preferably, in step (a) and / or step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In a specific embodiment, in step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). In a preferred embodiment, the sustained-release drug delivery system (DDS) of step (a) is a subcutaneous implant or an intramuscular implant. Preferably, the implant is a biodegradable polymer.

[0048] According to yet another embodiment, the present invention provides a method for diagnosing behavioral disorders (aggression) in non-human mammals treated with a deserorelin sustained-release implant, comprising the steps of:

[0049] a) collecting a blood sample from the non-human mammal treated with deserorelin,

[0050] b) measuring the testosterone plasma level in said blood sample,

[0051] c) determining the testosterone level in said plasma, wherein said testosterone plasma level in said blood sample is above 0.2 ng / mL, preferably above 0.4 ng / mL,

[0052] d) finally, determining a dose of deserorelin for further administration to said non-human mammal for resolving the behavioral disorder.

[0053] In another embodiment, the invention relates to the use of deserorelin for inducing temporary infertility in healthy male animals for a period of at least 6 months, comprising administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, wherein after administration, the dose is effective to induce a deserorelin Cmax of more than 7000 pg / mL 1 hour after administration and a deserorelin concentration in the plasma of the non-human mammal of at least 10 pg / mL.

[0054] Another embodiment of the present invention relates to a method of contraception for non-human mammals, comprising administering an implant comprising deserorelin and a biodegradable polymer subcutaneously or intramuscularly to the non-human mammal, wherein the implant releases deserorelin at a deserorelin Cmax of more than 7000 pg / mL 1 hour after administration, and wherein the deserorelin concentration in the plasma of the non-human mammal is at least 10 pg / mL and is maintained for at least 6 months. According to this embodiment, deserorelin is administered subcutaneously or intramuscularly to a non-human mammal in need thereof at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a plasma concentration of less than 400 pg / ml from 8 days after administration. Preferably, the deserorelin plasma concentration is less than 400 pg / mL from 8 days to at least 6 months after administration.

[0055] According to another embodiment, the present invention provides a maintenance regimen for inhibiting or reducing fertility in an animal, comprising:

[0056] (a) first administering a therapeutically effective amount of a deserorelin product to an animal in need thereof to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL for a first predetermined period of time;

[0057] (b) subsequently administering to the animal in need thereof a therapeutically effective amount of deserorelin in a sustained release system over a second predetermined period of time such that the plasma concentration of deserorelin is at least 10 pg / ml.

[0058] In another embodiment, the invention relates to deserorelin for use in preventing behavioral disorders, including aggression, in a non-human mammal, wherein the deserorelin is administered subcutaneously or intramuscularly via a sustained release implant, and wherein the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL for at least 6 months.

[0059] Another embodiment of the invention is directed to deserorelin for use in preventing impaired FSH / LH levels in a non-human mammal, wherein deserorelin is administered subcutaneously or intramuscularly via a sustained release implant, and wherein the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL for at least 6 months.

[0060] Finally, the present invention also generally relates to the use of deserorelin or a sustained-release drug delivery system comprising deserorelin in the preparation of a medicament for:

[0061] - inducing chemical castration in a non-human mammal, or

[0062] - Inducing temporary sterility in healthy male animals, preferably dogs, for a period of several months, or

[0063] - Inducing temporary infertility in healthy and sexually immature female animals (preferably dogs) to delay the onset of first estrus and signs of estrus and to prevent pregnancy at an early age, or

[0064] - Inducing temporary sterility and suppressing urine odor and sexual behaviors such as libido, vocalization, urine marking, and aggression in healthy male animals (preferably cats), or

[0065] - for the purpose of contraception or inhibition or reduction of fertility in non-human mammals, or

[0066] - prevention of behavioural disorders (including aggression) in non-human mammals, or

[0067] - Preventing impaired FSH / LH levels in non-human mammals.

[0068] According to the present invention, all embodiments of the various uses of deserelin and the various methods and compositions related to the use of deserelin described in this specification are also applicable to the use of deserelin for preparing the above-mentioned medicament. BRIEF DESCRIPTION OF THE DRAWINGS

[0069] In addition to the exemplary aspects and embodiments described above, further aspects and embodiments of the present invention will become apparent by reference to the drawings and study of the following descriptions.

[0070] Figure 1 The ratio of testosterone after GnRH stimulation / before GnRH stimulation is shown. In male cats, the ratio of the control group (saline solution) and the treatment group (Suppress® 4.7) was measured at baseline and on day 14. The figure corresponds to the results obtained according to Example 2.

[0071] Figure 2 The effect of deserorelin on sperm concentration in male cats is shown. According to Example 3, the sperm concentration ( / μL) in cats treated with Suyiqing® was measured over time.

[0072] Figure 3 A direct correlation between the injected deserorelin concentration and the testosterone concentration is shown. According to Example 4, the apparent relationship between the plasma deserorelin concentration and PD parameters over time was evaluated graphically by the testosterone concentration and deserorelin concentration obtained from 16 healthy male cats treated with 4.7 mg of Supremo®.

[0073] Figure 4 The results of the mean testicular volume change over time of male cats (control group n=5 cats and treatment group n=10 cats) treated with 4.7 mg of Suyiqing® according to Example 7 are shown. In this figure, the mean testicular volume (right testicle) (in mm 3 The mean diameter (in weeks) and its 95% confidence interval (CI) are presented as a function of time (in weeks).

[0074] Figure 5 Shown are the mean ratios (and their 95% CI) of testosterone post-GnRH stimulation / pre-GnRH stimulation in treated dogs (n=6) tested in a study of approximately 15 months according to Example 9. In this figure, the mean ratios are expressed as a function of time (in days).

[0075] Figure 6 Results are shown for the mean ratio of LH post-GnRH stimulation / pre-GnRH stimulation (and its 95% CI) in treated dogs (n=6) tested in a study of approximately 15 months according to Example 10. In this figure, the mean ratio is expressed as a function of time (in days).

[0076] Figure 7The mean semi-log plasma concentrations of deserorelin (pg / mL) and the linear serum concentrations of hormones (testosterone, LH and FSH) before GnRH stimulation (pg / mL) according to Example 11 are shown as a function of time (days) throughout the study (n=6 dogs treated).

[0077] Figure 8 A. Figure 8 B and Figure 8 C shows the results of the average therapeutic percentage according to Example 12 ( Figure 8 A) Detailed levels of testosterone concentrations in Study 1 (pg / mL) Figure 8 B) and detailed levels of testosterone concentration (pg / mL) in Study 2 ( Figure 8 C).

[0078] Fig. 9 A and Fig. 9 B shows the mean values ​​of the total scores of sexual behavior in Study 1 according to Example 12 ( Fig. 9 A) and the mean of the total sexual behavior scores in Study 2 ( Fig. 9 B).

[0079] Fig.10 A and Fig.10 B shows the average phonation changes of Study 1 according to Example 12 ( Fig.10 A) and mean vocalization changes in Study 2 ( Fig.10 B).

[0080] Fig.11 A and Fig.11 B shows the percentage change from baseline of urine markers in Study 1 according to Example 12 ( Fig.11 A) and percent change from baseline in urine markers in Study 2 ( Fig.11 B).

[0081] Fig.12 A and Fig.12 B shows the change / reduction in urine odor measured as a change from baseline according to Study 1 of Example 12 ( Fig.12 A) and Study 2 measured change / reduction in urine odor relative to baseline ( Fig.12 B).

[0082] Fig.13 A and Fig.13 B shows the percentage change in testicular volume according to Study 1 of Example 12 ( Fig.13 A) and the percentage change in testicular volume in Study 2 ( Fig.13 B).

[0083] Fig.14 A and Fig.14 B shows the percentage change in penile thorn appearance according to Study 1 of Example 12 ( Fig.14 A) and the percentage change in penile thorn appearance in Study 2 ( Fig.14 B).

[0084] Fig.15 The results of the PK / PD preclinical study in healthy male cats (kittens) described in Example 13 are shown. The PK study results are as follows Fig.15 Testosterone levels were recorded as Fig.15 B; records of penis pricking and behavior are as follows Fig.15 and Fig.15 As shown in D. DETAILED DESCRIPTION

[0085] Deserorelin is available as Deserorelin acetate in commercial implants for dogs named Suserolin® 4.7 mg implant or Suserolin® 9.4 mg implant. It is prescribed to induce temporary infertility in healthy, able-bodied, sexually mature male dogs. Deserorelin is a GnRH agonist that works by suppressing pituitary-gonadal axis function when administered continuously at low doses. This suppression results in the inability of the treated animal to synthesize and / or release follicle-stimulating hormone (FSH) and luteinizing hormone (LH), two hormones responsible for maintaining fertility.

[0086] Continuous low-dose deserorelin reduces male reproductive organ function, libido, and spermatogenesis, and reduces plasma testosterone levels, starting 4 to 6 weeks after implantation. A transient increase in plasma testosterone can be seen immediately after implantation. Measurements of testosterone plasma concentrations indicate that deserorelin persists in the circulation for at least 12 months after administration of the veterinary medicinal product, with a persistent pharmacological effect.

[0087] As used herein, "behavioral disorders in non-human mammals" include, but are not limited to, urine odor, sexual desire, vocalization, urine marking, and aggression in healthy male animals, preferably cats.

[0088] As used herein, the term "chemical castration" refers to the use of chemicals or drugs to stop the production of sex hormones in order to reduce or inhibit reproductive and sexual activity. Chemical castration is sometimes called medical castration or hormone therapy. Chemical castration is reversible, meaning that hormone production resumes when the chemicals or drugs are stopped. Unlike surgical castration, which removes the gonads through an incision in the body, chemical castration does not remove organs and is not a form of sterilization.

[0089] As used herein, the term "sexual activity" specifically refers to and includes sexual behaviors such as libido, vocalization, urine marking or scent marking, and aggression in both healthy male and healthy female animals. As examples of sexual activities in healthy male and healthy female animals, courtship, mating, and scent marking are also included.

[0090] As used herein, the term "Cmax" of deserorelin refers to the maximum plasma concentration of deserorelin following administration of deserorelin.

[0091] As used herein, the term "healthy animal" refers to an animal that has no contraindications to temporary infertility / sterility induction, surgical castration, or chemical castration.

[0092] As used herein, the term "sound animal" refers to an animal that has not been surgically castrated and is not currently undergoing temporary infertility / sterility treatment.

[0093] As used herein, the term "non-human animal" may refer to any organism of the animal kingdom except humans (Homo sapiens). Examples of the term "animal" as used herein include, but are not limited to, companion animals such as dogs, cats, and horses; and livestock animals such as cattle, goats, sheep, and pigs.

[0094] As used herein, the term "pharmaceutical salts thereof" in relation to deserorelin refers to pharmaceutically acceptable salts of deserorelin. An example of a pharmaceutically acceptable salt thereof is deserorelin acetate.

[0095] As used herein, the term "sexually mature female animal" refers to a female animal that has seasonal estrus and signs of estrus.

[0096] In contrast, a "sexually immature female animal" is a female animal that is not yet capable of estrus and does not show any signs of estrus.

[0097] As used herein, the term "transient" in relation to infertility means that the infertility persists for a period of time, ie, the infertility is reversible.

[0098] As used herein, the term "temporary infertility" is related to the chemicals that are able to induce it and refers to the inability to become pregnant. Some chemotherapy treatments can cause infertility during treatment. But after the treatment is over, fertility sometimes returns. This will depend on the drug, but also on other factors such as age and whether the patient is male or female. "Temporary infertility" in the meaning of the present invention can therefore be equated with the result of contraception or a method of birth control.

[0099] As used herein, the term "Tmax" refers to the time at which the maximum plasma concentration (Cmax) of deserorelin is reached or observed following administration of deserorelin.

[0100] Pharmaceutical composition

[0101] One aspect of the present disclosure provides a composition for inducing chemical castration in a non-human animal in need thereof as described in the claims, comprising a therapeutically effective amount of deserorelin and an excipient.

[0102] In a preferred embodiment of the present invention, deserorelin is formulated as a solid implant formulation.

[0103] In a preferred embodiment, the implant formulation comprises glyceryl stearate (also known as hydrogenated palm oil) and lecithin.

[0104] According to a specific embodiment of the present invention, the solid implant formulation preferably comprises about 2 (w / w)% to 12 (w / w)% of deserorelin (calculated as active ingredient), about 0.5 (w / w)% to 2.5 (w / w)% of lecithin and about 85 (w / w)% to 97.5 (w / w)% of glyceryl stearate.

[0105] More preferably, the solid implant formulation comprises about 4 (w / w)% to 10 (w / w)% of deserorelin (based on the active ingredient), about 0.5 (w / w)% to 1.5% (w / w) of lecithin and about 85 (w / w)% to 94 (w / w)% of glyceryl stearate.

[0106] Particularly preferred solid implant formulations are selected from the group consisting of formulations comprising 85 (w / w)% to 89 (w / w)% of glyceryl stearate, 9 (w / w)% to 10 (w / w)% of deserorelin (calculated as active ingredient) and 0.8 (w / w)% to 1.2 (w / w)% of lecithin.

[0107] In a further preferred embodiment, the solid implant formulation may further comprise anhydrous sodium acetate.

[0108] Glyceryl stearate is partially hydrogenated palm oil. Its main fatty acids are C16:0 (45%) and C18:0 (53%). The melting point is about 55°C.

[0109] Lecithin is a phosphatidylcholine. It is a mixture of diglycerides of stearic, palmitic and oleic acids linked to a choline ester of phosphoric acid.

[0110] In another preferred embodiment, deserorelin is formulated as a composition in liquid form.

[0111] Deserorelin can be administered by any suitable means, for example by an implant which is preferably biodegradable over time, by a sustained release drug delivery system (DDS), by a biocompatible polymer-solvent system comprising a biodegradable polymer compatible with subcutaneous, transdermal or intramuscular administration.

[0112] According to the present invention, "DDS" refers to any system capable of delivering drugs in vivo. Preferred DDS according to the present invention include any type of implant; in particular, the term includes liquid implants and solid implants, which have at least polymeric materials or non-polymeric materials.

[0113] According to the present invention, "slow release" when referring to DDS means that deserorelin is not released in the form of a single administration of the total dose of deserorelin. Instead, as disclosed in the present application, the slow release DDS is capable of releasing deserorelin over a certain amount of time.

[0114] In some embodiments, the implant is preferably biodegradable over time. It is selected from a biodegradable material and a bioerodible dosage form.

[0115] As used herein, the term "biocompatible" may mean "not harmful to living tissue." This term includes biodegradable and bioerodible.

[0116] As used herein, "degradable" means that the polymer will degrade or erode in vivo into smaller chemical species, where degradation can be accomplished by, for example, enzymatic, chemical ("biodegradable" polymers), and / or physical ("bioerodible" polymers) processes.

[0117] As used herein, the term "biodegradable" may refer to any water-insoluble material that can be converted into one or more than one water-soluble material under physiological conditions without regard to any particular degradation mechanism or process.

[0118] In some embodiments, the biodegradable material is a biodegradable polymer (or biopolymer), which is preferably selected from polylactic acid, polyglycolic acid, polylactide, polyglycolide, polycaprolactone, polyanhydride, polyamide, polyurethane, polyesteramide, polyorthoester, polydioxanone, polyacetal, polyketal, polycarbonate, polyphosphazene, polyhydroxybutyrate, polyhydroxyvalerate, polyalkylene oxalate, polyalkylene succinate, polymalic acid, polyethylene glycol, hyaluronic acid, chitin and chitosan, copolymers thereof, terpolymers thereof, and any combination thereof.

[0119] In some embodiments, the biodegradable polymer comprises a monomer selected from the group consisting of lactide, glycolide, caprolactone, p-dioxanone, trimethylene carbonate, 1,5-dioxepan-2-one, 1,4-dioxepan-2-one, ethylene oxide, propylene oxide, sebacic anhydride, diketene acetal / diol, lactic acid, and combinations thereof.

[0120] As used herein, the term "implant" may be understood as a bioerodible dosage form, including devices such as microspheres; "liquid polymer reservoir" formulations (or in situ formed implants); and solid polymer implants, such as pellets or pellets. The implant comprises deserorelin and a bioerodible polymer as an excipient.

[0121] As used herein, the term "bioerodible" may refer to any water-insoluble material that can be mechanically eroded by a biological process that dissolves at least a portion of the material.

[0122] Once eroded, the bioerodible polymer will release deserorelin in body fluids including blood and lymph or tissue. Illustrative examples of bioerodible polymers according to the present invention include polyhydroxy acids, such as polylactide, polyglycolide, polylactide-co-glycolide, polylactic acid, polyglycolic acid and polylactic acid-co-glycolic acid, polyanhydrides, polyorthoesters, polyetheresters, polyethylene glycol, poly-ε-caprolactone, polyesteramides, polyphosphazenes, polycarbonates, polyamides, and copolymers and blends thereof, fats, waxes and long chain C 12 To C 22 Triglycerides of fatty acids, such as stearate, palmitate, laurate, myristate, arachidate and behenate, and mixtures thereof, having a melting point above 50° C., preferably glyceryl tristearate. Preferably, the polymer is selected from poly-ε-caprolactone (PCL), polylactide, polyglycolide, and copolymers or blends thereof, such as polylactide-co-glycolide and glyceryl tristearate.

[0123] Injectable implants can also include antioxidants. Antioxidants suitable for injectable implants are known in the art. A preferred antioxidant is BHT (butylated hydroxytoluene). Such implants, microspheres and in situ formed implants can be prepared using methods known in the art, such as those described in patent applications EP3349726, WO90 / 03768, WO2006 / 063794, WO2009 / 091737, WO98 / 07423, EP1197207, EP0525307, WO98 / 08533, WO2020 / 222399, WO2020 / 130585 and WO2013 / 082373, and deserorelin is not dissolved during the preparation process. In a preferred embodiment, the present invention relates to an implant as defined above, comprising deserorelin, a bioerodible polymer selected from poly-ε-caprolactone (PCL), polylactide, polyglycolide, polylactide-co-glycolide, copolymers or blends thereof, and glyceryl tristearate, and optionally an antioxidant.

[0124] In some embodiments, deserorelin is present in the composition in liquid form in an effective dose for greater than one month, more preferably greater than two months, most preferably greater than three months or more. In a preferred embodiment, deserorelin is present in the composition in an effective dose for greater than four months, more preferably greater than five months, most preferably greater than six months or more.

[0125] As used herein, the term "liquid" may refer to the ability of a composition to deform under shear stress, regardless of the presence of a non-aqueous solvent.

[0126] The liquid compositions according to the present disclosure have a liquid physical state at ambient and body temperatures, and remain liquid in vivo, ie, in a largely aqueous environment.

[0127] The deserorelin compositions are formulated for sustained release, preferably in a uniform manner via a slow delivery system. For this purpose, the viscosity of the liquid composition of deserorelin at ambient temperature (i.e., about 20° C.) is greater than about 0.5 Pa.s, greater than about 1.0 Pa.s, greater than about 2.0 Pa.s, greater than about 3.0 Pa.s, greater than about 4.0 Pa.s, greater than about 5.0 Pa.s, greater than about 6.0 Pa.s, greater than about 7.0 Pa.s, greater than about 8.0 Pa.s, greater than about 9.0 Pa.s, greater than about 10.0 Pa.s, greater than about 11.0 Pa.s, greater than about 12.0 Pa.s, greater than about 13.0 Pa.s, greater than about 14.0 Pa.s, greater than about 15.0 Pa.s, greater than about 16.0 Pa.s, greater than about 17.0 Pa.s, greater than about 18.0 Pa.s, greater than about 19.0 Pa.s, or greater than about 20.0 Pa.s. In other embodiments, the viscosity of the substantially homogeneous composition can be greater than about 30.0 Pa.s, greater than about 40.0 Pa.s, greater than about 50.0 Pa.s, greater than about 60.0 Pa.s, greater than about 70.0 Pa.s, greater than about 80.0 Pa.s, greater than about 90.0 Pa.s, greater than about 100.0 Pa.s, greater than about 110.0 Pa.s, greater than about 120.0 Pa.s, greater than about 130.0 Pa.s, greater than about 140.0 Pa.s, greater than about 150.0 Pa.s, greater than about 160.0 Pa.s, greater than about 170.0 Pa.s, greater than about 180.0 Pa.s, greater than about 190.0 Pa.s, or even greater than about 200.0 Pa.s. Alternatively, the viscosity of the substantially homogeneous composition can be greater than any integer from about 0.50 Pa.s to about 200.0 Pa.s.

[0128] The viscosity can be measured using a suitable viscometer in a setting compatible with the desorelin liquid composition. For example, the viscosity can be measured using a viscometer, such as but not limited to a Brookfield viscometer or an Anton Paar Rheoplus viscometer with an appropriate setting. Whenever a viscosity value is mentioned / defined herein, it relates to the viscosity measured using a Brookfield. The viscometer is used at 20°C.

[0129] Various excipients commonly used in formulations, preferably pharmaceutical formulations, can be selected according to several criteria, such as the desired dosage form and the release profile properties of the dosage form. Non-limiting examples of suitable excipients include agents selected from the group comprising binders, fillers, preservatives, diluents, lubricants, dispersants, pH adjusters, stabilizers, and any combination of these agents.

[0130] Methods of using the deserorelin compositions

[0131] Another aspect of the present disclosure includes a method of inducing chemical castration in a non-human animal in need thereof. The method includes administering a composition comprising an effective amount of deserorelin to the non-human animal.

[0132] In one embodiment, the effective amount of desorelin is in doses of 4.7 mg and 9.4 mg of the SUPERIOR® implant.

[0133] A first object of the invention is deserorelin for chemical castration of a non-human mammal within 1 month after administration, wherein at the time of administration, the Cmax of deserorelin after administration exceeds 7000 pg / mL and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL.

[0134] Depending on the animal species, administration of deserorelin to an animal achieves a deserorelin Cmax of about 7000 pg / mL to about 200000 pg / mL at a time (Tmax) of about 0.4 hours to about 3.4 hours, such as a deserorelin Cmax of about 7500 pg / mL to about 170000 pg / mL. In a specific embodiment, the deserorelin Cmax exceeds 7429 pg / mL at 1 hour after administration.

[0135] In other embodiments, the Cmax of deserorelin may also be achieved at a Tmax of about 0.7 hours to about 2 hours, such as a Tmax of about 0.5 hours to about 1.7 hours. In some embodiments, Cmax may be different in different non-human animals, so that Cmax may be even higher. For example, the Cmax can be about 7200 pg / mL, about 7400 pg / mL, about 8000 pg / mL, about 9000 pg / mL, about 10000 pg / mL, about 11000 pg / mL, about 12000 pg / mL, about 13000 pg / mL, about 15000 pg / mL, about 20000 pg / mL, about 25000 pg / mL, about 30000 pg / mL, about 35000 pg / mL, about 40000 pg / mL, about 45000 pg / mL, about 50000 pg / mL, about 55000 pg / mL, about 60000 pg / mL, about 65000 pg / mL, about 70000 pg / mL, about 75000 pg / mL, about 80000 pg / mL, about In some embodiments, the Cmax of deserelin may be greater than 500 pg / mL. In other embodiments, the Cmax of deserelin may be less than 300000 pg / mL.

[0136] In other embodiments, Tmax can occur at intervals of 30 minutes, 1 hour or 2 hours in non-human animal individuals, and the time range of reaching Tmax is about 30 minutes to 2 hours. Tmax can be about 0.4 hours, about 0.5 hours, about 0.6 hours, about 0.7 hours, about 0.8 hours, about 0.9 hours, about 1.0 hours, about 1.1 hours, about 1.2 hours, about 1.3 hours, about 1.4 hours, about 1.5 hours, about 1.6 hours, about 1.7 hours, about 1.8 hours, about 1.9 hours, about 2.0 hours, about 2.2 hours, about 2.4 hours, about 2.6 hours, about 2.8 hours, about 3.0 hours, about 3.2 hours, or about 3.4 hours. In one embodiment, Tmax can be 0.4 hours to 4 hours. In other exemplary embodiments, Tmax can be 0.5 hours to 1.0 hours. In some embodiments, Tmax can be greater than 0.4 hours. In other embodiments, Tmax can be less than 3.4 hours. In exemplary embodiments, Tmax may be from 0.03 days to 1.33 days.

[0137] The area under the curve (AUC) can range from about 10,000 days*pg / mL to about 110,000 days*pg / mL. In exemplary embodiments, the AUC can range from about 11,000 days*pg / mL to about 15,000 days*pg / mL, about 15,000 days*pg / mL to about 20,000 days*pg / mL, about 20,000 days*pg / mL to about 25,000 days*pg / mL, about 25,000 days*pg / mL to about 30,000 days*pg / mL, about 30,000 days*pg / mL to about 350,000 days*pg / mL, 00 days∗pg / mL, about 35000 days∗pg / mL to about 40000 days∗pg / mL, about 40000 days∗pg / mL to about 45000 days∗pg / mL, about 45000 days∗pg / mL to about 50000 days∗pg / mL, about 50000 days∗pg / mL to about 55000 days∗pg / mL, about 55000 days∗pg / mL to about 60000 days∗pg / mL.

[0138] According to the first object of the present invention, deserorelin is administered subcutaneously or intramuscularly to a non-human mammal in need thereof, at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, preferably a deserorelin plasma concentration of 7000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a deserorelin plasma concentration of less than 400 pg / ml from 8 days after administration. Preferably, the deserorelin plasma concentration is 10 pg / mL to 400 pg / ml from 8 days to at least 6 months after administration.

[0139] According to a particular embodiment of the first object of the invention, deserorelin is used for inducing temporary infertility.

[0140] Therefore, the present invention also relates to deserorelin for inducing temporary infertility in healthy male animals for a period of at least 6 months, comprising administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, in an amount effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, preferably a deserorelin plasma concentration of 7000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a deserorelin plasma concentration of less than 400 pg / ml from 8 days after administration. Preferably, the deserorelin plasma concentration is less than 400 pg / ml from 8 days after administration to 6 months. According to this specific embodiment, after administration, deserorelin is substantially continuously released at a plasma concentration level of less than 400 pg / mL from 8 days to at least 6 months after administration.

[0141] In addition, the present invention also relates to deserorelin for inducing temporary infertility in healthy and sexually immature female animals to delay the first estrus and estrus signs and prevent them from becoming pregnant in childhood, which comprises the following steps:

[0142] (a) administering an effective amount of deserorelin such that the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 4000 pg / mL to 40000 pg / ml within the first 24 hours after injection, preferably the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 7000 pg / mL to 40000 pg / ml within the first 24 hours after injection; and thereafter

[0143] (b) administering a maintenance dose of deserorelin so that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0144] Preferably, according to the first object of the present invention, when deserorelin is used to induce chemical castration or temporary infertility, deserorelin is formulated in the form of a sustained release drug delivery system (DDS).

[0145] Therefore, a second object of the present invention is a sustained-release drug delivery system (DDS) containing deserorelin for chemical castration of non-human mammals according to the first object of the present invention, wherein after administration, the DDS releases deserorelin at a deserorelin Cmax of more than 7000 pg / mL, and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL and is maintained for at least 6 months. Preferably, the DDS is a subcutaneous implant or an intramuscular implant comprising deserorelin and a biodegradable material such as a biodegradable polymer.

[0146] According to a particular embodiment of the second object, after administration, deserorelin reduces the testosterone level in the plasma of said non-human mammal to below 0.4 ng / ml and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL, preferably above 15 pg / mL.

[0147] A third object of the present invention is a sustained-release drug delivery system (DDS) containing deserorelin for inducing temporary infertility in healthy male animals for a period of at least 6 months, which comprises administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, preferably a deserorelin plasma concentration of 7000 pg / mL to 40000 pg / ml within the first 24 hours after injection, and a deserorelin plasma concentration of less than 400 pg / ml from 8 days after administration.

[0148] A fourth object of the present invention is also a method for chemically castrating a non-human mammal, comprising the following steps:

[0149] (a) administering an effective amount of deserorelin such that the Cmax concentration in plasma is at least 7000 pg / mL within the first 24 hours after injection, preferably the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 7000 pg / mL to 40000 pg / mL within the first 24 hours after injection; and thereafter

[0150] (b) administering a maintenance dose so that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0151] According to the specific embodiment of the 4th object of the present invention, in step (a) and / or step (b), subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. The sustained-release DDS of step (a) is preferably a subcutaneous implant or an intramuscular implant. According to a preferred embodiment, in step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). The implant preferably comprises a biodegradable material, such as a biodegradable polymer.

[0152] A fifth object of the present invention is a method for inducing temporary infertility in a healthy, sound, sexually mature male animal, preferably a dog, comprising the steps of:

[0153] (a) administering an effective amount of deserorelin such that the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 4000 pg / mL to 40000 pg / ml, preferably within the first 24 hours after injection, the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 7000 pg / mL to 40000 pg / ml; and thereafter

[0154] (b) administering a maintenance dose so that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0155] According to the specific embodiment of the fifth object of the present invention, in step (a) and / or step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In this case, DDS preferably releases deserorelin with a deserorelin Cmax exceeding 7000 pg / mL 1 hour after administration, and the deserorelin concentration in non-human mammal plasma is at least 10pg / mL, and maintains at least 6 months. More preferably, the deserorelin plasma concentration effectively induced by the administration dose of deserorelin in 1 hour to 24 hours is 4000 pg / mL to 40000 pg / mL, and the deserorelin plasma concentration after administration for 8 days, preferably to at least six months is lower than 400 pg / ml. According to a preferred embodiment, in step (b), a subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). The implant preferably comprises a biodegradable material, such as a biodegradable polymer.

[0156] The present invention also contemplates the use of deserorelin for inducing temporary infertility in healthy male animals for a period of at least 6 months, comprising administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, wherein after administration, the dose is effective to induce a deserorelin plasma Cmax concentration of more than 7000 pg / mL at 1 hour after administration and a deserorelin concentration in the plasma of the non-human mammal of at least 10 pg / mL.

[0157] A sixth object of the present invention is a method for inducing temporary infertility in healthy and sexually immature female animals, preferably dogs, to delay the first estrus and estrus signs and prevent pregnancy in early childhood, comprising the following steps:

[0158] (a) administering an effective amount of deserorelin such that within the first 24 hours after injection, the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 4000 pg / mL to 40000 pg / ml, preferably within the first 24 hours after injection, the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 7000 pg / mL to 40000 pg / ml; and thereafter

[0159] (b) administering a maintenance dose so that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0160] According to the specific embodiment of the sixth object of the present invention, in step (a) and / or step (b), subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In this case, DDS should preferably be used at 12 weeks to 16 weeks. DDS comprises deserorelin and biodegradable materials such as biodegradable polymers, and DDS preferably releases deserorelin with a deserorelin Cmax exceeding 7000 pg / mL after administration in 1 hour, and the concentration of deserorelin in non-human mammal blood plasma is at least 10 pg / mL and maintains at least 6 months. In addition, even more preferably, the administration dose of deserorelin effectively induces a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL in 1 hour to 24 hours, and the deserorelin plasma concentration of 8 days after administration, preferably to at least 6 months is lower than 400 pg / ml. According to a preferred embodiment, in step (b), subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). The implant preferably comprises a biodegradable material, such as a biodegradable polymer.

[0161] A seventh object of the present invention is a method for inducing temporary infertility and suppressing urine odor and sexual behaviors such as libido, vocalization, urine marking and aggression in healthy male animals (preferably cats), comprising the following steps:

[0162] (a) administering an effective amount of deserorelin such that within the first 24 hours after injection, the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 4000 pg / mL to 40000 pg / ml, preferably within the first 24 hours after injection, the Cmax concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof in plasma is 7000 pg / mL to 40000 pg / ml, and thereafter

[0163] (b) administering a maintenance dose so that the mean plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is greater than 10 pg / ml.

[0164] According to the seventh object of the present invention, the healthy male animal is preferably a cat, and even more preferably a cat over 3 months old.

[0165] According to the specific embodiment of the 7th purpose of the present invention, in step (a) and / or step (b), subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected. In this case, DDS preferably comprises deserorelin and biodegradable materials such as biodegradable polymers, and DDS releases deserorelin with the deserorelin Cmax exceeding 7000 pg / mL after administration for 1 hour, and the concentration of deserorelin in non-human mammal blood plasma is at least 10 pg / mL and maintains at least 6 months. More preferably, the administration dose of deserorelin effectively induces the deserorelin plasma concentration in 1 hour to 24 hours to be 4000pg / mL to 40000 pg / mL, and the deserorelin plasma concentration of 8 days after administration, preferably to at least 6 months is lower than 400pg / ml. According to a preferred mode, in step (b), subcutaneous or intramuscular sustained-release drug delivery system (DDS) is injected while implanting in step (a). The implant preferably comprises biodegradable materials, such as biodegradable polymers.

[0166] An eighth object of the present invention is a method for diagnosing behavioral disorders (aggression) in non-human mammals treated with a deserorelin sustained-release implant, comprising the following steps:

[0167] a) detecting the testosterone plasma level in a blood sample of said non-human mammal treated with said deserorelin extended-release implant,

[0168] b) determining the testosterone level in said plasma, wherein the testosterone plasma level in said blood sample is above 0.2 ng / mL, preferably above 0.4 ng / mL,

[0169] c) finally, determining a dose of deserorelin for further administration to said non-human mammal to resolve the behavioral disorder.

[0170] A ninth object of the present invention is a method of contraception for a non-human mammal, comprising administering an implant comprising deserorelin and a biodegradable material such as a biodegradable polymer subcutaneously or intramuscularly to the non-human mammal, wherein the implant releases deserorelin at a Cmax of more than 7000 pg / mL 1 hour after administration, and wherein the deserorelin concentration in the plasma of the non-human mammal is at least 10 pg / mL and is maintained for at least 6 months.

[0171] According to the ninth object, deserorelin is administered to a non-human mammal in need thereof at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, preferably a deserorelin plasma concentration of 7000 pg / mL to 40000 pg / mL within 1 hour to 24 hours, and a deserorelin plasma concentration of less than 400 pg / ml starting from 8 days after administration. Preferably, the plasma concentration is less than 400 pg / ml starting from 8 days after administration and maintained for at least 6 months.

[0172] A tenth object of the present invention is a maintenance regimen for inhibiting or reducing fertility in an animal, comprising:

[0173] (a) first administering a therapeutically effective amount of a deserorelin product to an animal in need thereof to induce a plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof within a first predetermined period of time of 4000 pg / mL to 40000 pg / mL, preferably 7000 pg / mL to 40000 pg / mL;

[0174] (b) subsequently administering to the animal in need thereof a therapeutically effective amount of a sustained release system dose of deserorelin over a second predetermined period of time such that the plasma concentration of deserorelin or an equivalent amount of a pharmaceutically acceptable salt thereof is at least 10 pg / ml.

[0175] An eleventh object of the present invention is deserorelin for use in preventing behavioral disorders in non-human mammals, wherein deserorelin is administered subcutaneously or intramuscularly via a sustained release drug delivery system, and wherein the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL (for at least 6 months).

[0176] One of the embodiments involves the administration of deserorelin to induce changes in the hormone pathway. In fact, through the new data generated, it is now clear that deserorelin affects LH concentrations. In fact, Example 10 shows changes in the LH hormone through the results of the mean (95% CI) ratio of LH after GnRH stimulation / before GnRH stimulation in the treated group dogs (n=6) tested in the study for about 16 months, as shown in Figure 6 shown.

[0177] Furthermore, in Example 11, the correlation between the concentration of injected deserorelin and the concentration of the hormones LH, FSH and testosterone in male cats treated with Suyiqing® is shown. The study shows that deserorelin is negatively correlated with testosterone / LH and FSH. In fact, as soon as the concentration of deserorelin becomes low (12 pg / mL), the concentration of the other hormones begins to increase.

[0178] Therefore, a twelfth object of the present invention is deserorelin for chemical castration of non-human mammals, wherein after administration, deserorelin reduces the testosterone level in the plasma of the non-human mammal to below 0.4 ng / ml, and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL, preferably higher than 15 pg / mL.

[0179] The thirteenth object of the present invention is deserorelin for use in preventing impairment of follicle stimulating hormone (FSH) and luteinizing hormone (LH) in non-human animals, wherein deserorelin is administered subcutaneously or intramuscularly via a sustained-release implant, and wherein the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL (for at least 6 months).

[0180] In fact, it has been shown that Desorelin affects hormonal pathways. However, this relevance goes beyond the hormonal range. Hormonal pathways also affect the metabolic range by changing / modifying or altering several metabolic pathways, such as:

[0181] ∙ Sperm concentration : As in Example 3 and Figure 2 As shown, deserorelin has an effect on sperm concentration.

[0182] ∙ Testicular weight : As in Example 6 and Figure 4 It has been shown that deserorelin has an effect on testicular weight. Indeed, it has been shown that injection of deserorelin implants reduces testicular volume over time when the deserorelin effect is present.

[0183] ∙ Changes in fat and carbohydrates : Low testosterone levels can lead to weight gain

[0184] ∙ Improved / reduced urine odor :

[0185] Urine marking is the normal and intentional deposition of urine as a result of:

[0186] • Pheromone signals to other animals; especially unneutered male cats

[0187] • Territorial Signals

[0188] • Signs of stress or excitement related to the social or physical environment.

[0189] In this case, the urine smell is pungent, so it is desirable to remove or mask this odor. After washing the area with a mild detergent, several commercially available enzymatic urine odor neutralizers should be applied to the surface. These neutralizers can be used to remove the odor, but they are difficult to treat. Castration or spaying has been shown to be an effective treatment, especially for cats marked with reproductive notices.

[0190] One of the reasons for surgical castration is to reduce the strong and persistent urine odor in male animals, especially cats and ferrets. Urine odor is known to improve within a few days after surgical castration.

[0191] One of the purposes of the present application is to demonstrate that administration of deserorelin (also known as serotonin®) can reduce the odor of urine and / or feces. Surprisingly, the inventors have demonstrated that the odor reduction is mediated by at least two pathways. In fact, one of these pathways is a testosterone-mediated sex hormone pathway. The inventors have also demonstrated another pathway that is not related to sex hormones. Therefore, compared to the reduction in urine odor that occurs in surgically castrated male animals, especially cats, the reduction in urine odor in healthy male animals receiving deserorelin according to the present invention is improved.

[0192] One aspect of the present invention is to reduce urine odor in animals such as livestock, particularly cats, and preferably intact male cats.

[0193] One of the objectives of this application is to demonstrate that administration of Desorelin / Sulfamethoxazole® reduces urine odor by reducing the levels of testosterone / felin and its metabolites.

[0194] It is known from the prior art (Hendriks WH et al., Testosterone increases urinaryfree felinine, N-acetylfelinine and methylbutanolglutathione excretion incats (Felis catus). J Anim Physiol Anim Nutr (Berl). 2008 Feb; 92(1):53-62) that testosterone increases the free excretion of felinine, N-acetylfelinine and 3-methylbutanolglutathione in castrated adult male cats and healthy female cats, while estradiol does not modulate this effect.

[0195] Cauxin is a carboxylesterase excreted as a major urinary component that regulates the production of feline urinary amino acids. It is also known that the secretion of cauxin is sex-dependent. In mature cats, the excretion of cauxin is higher in healthy male cats than in castrated male cats or healthy female cats or spayed female cats. The daily excretion of cauxin decreases immediately after castration. Immunohistochemistry confirmed that the expression of cauxin in the proximal straight tubules of the kidneys of healthy males is higher than that of castrated males. These results indicate that: 1) the excretion of cauxin is regulated by sex hormones such as testosterone, 2) cauxin functions as an esterase in the urine rather than in renal cells, and 3) the breakdown products of cauxin are secreted in a species-dependent, sex-dependent, and age-dependent manner, as is cauxin itself.

[0196] After castration, cauxin secretion decreases immediately, and the reason is confirmed to be the decrease in cauxin expression levels in the proximal straight tubules. It is reported that at 6 months, 12 months and 16 months after birth, the plasma testosterone concentration of healthy male cats increases with age to 1.5pmol / mL, 6.8pmol / mL and 12.6pmol / mL, respectively, and drops to zero after castration (Miyazaki et al., Chemical Signals in Vertebrates, 1998, 11, pp 51-60). Therefore, it is speculated that the transcriptional activity of cauxin is regulated by sex hormones such as testosterone.

[0197] Cauxin regulates the production of feline urinary amino acid. In vitro enzyme assays have shown that cauxin hydrolyzes the feline urinary amino acid precursor 3-methylbutanolcysteinylglycine into feline urinary amino acid and glycine.

[0198] Therefore, chemical castration induced by deserorelin release also affects the production of cauxine and felinine, which are responsible for urine odor. Since felinine and N-acetylfelinine were detected in cat bile and fecal samples, it was suggested that felinine is excreted from the liver into the feces via bile (LC-MS / MS quantification of felininemetabolites in tissues, fluids, and excretions from the domestic cat (Feliscatus), Futsuta et al., J Chromatogr B Analyt Technol Biomed Life Sci., 2018 Jan1;1072:94-99).

[0199] In addition to the sex hormone pathway mediated by testosterone that causes urine and / or feces odor, the inventors have also discovered another pathway that is unrelated to sex-dependent sex hormones. In fact, as shown in the examples, the inventors have identified new molecules that cause urine and / or feces odor. These molecules are secreted outside the sex hormone pathway. In fact, these molecules are not derived from the testosterone cascade.

[0200] Thus, one aspect of the invention is the use of deserorelin to adjust or alter or downregulate or upregulate the level of at least one molecule selected from the group consisting of butenoic acid; indole; p-methylphenol; cadaverine and its derivatives, such as acetylcadaverine, glutathionylamidopropylcadaverine; putrescine and its derivatives, such as N-acetylputrescine, p-coumaroylputrescine; thiol derivatives, such as 2-mercapto-3-butanone, 2-mercaptoethanol, 3-mercaptohexylbutyrate, mercaptopyruvic acid, 3-mercaptopropionic acid, 3-mercapto-3-methylbutan-1-ol, 7-mercaptoheptanoylthreonine, 1α,5α-dimercaptoandrostane-3α,17β-diol, mercaptopyruvic acid; taurine; isovalthine and isobuteine.

[0201] Therefore, one of the objects of the present invention is to reduce the odor of urine and / or feces by administering deserorelin to non-human animals.

[0202] In another aspect of the invention, administration of deserorelin affects fertility on the one hand and animal behavior, such as sexual and marking behavior, on the other hand. In fact, as shown in Example 13, administration of deserorelin affects penile pricking and behavioral recording, such as Fig.14 C and Fig.14 D, and showed that the results for penile pricking were statistically significant from 7 weeks after injection to 67 weeks. Regarding sexual behavior, the results were statistically significant at 10, 12, 34, 36, 38, 46, 52, 58, and 66 weeks after injection. Finally, regarding reproductive behavior, no statistical differences between the two groups were noted. The personality traits of cats can influence sexual desire. Trends in decreased vocalization, aggression, urine marking, and typical male urine odor were also noted.

[0203] The present invention provides several advantages including but not limited to the following:

[0204] First, by acting on both synthetic pathways for urine or fecal odor, it is clear that the odor reduction result is at least additive for both pathways and is certainly superior to an effect based solely on urine / fecal odor synthesis mediated solely by the sex hormone pathway.

[0205] It has now been shown that molecules that cause urine or feces odor are quantitatively and qualitatively attenuated or reduced in animals chemically sterilized by administration of desorelin, compared to surgically sterilized animals.

[0206] Furthermore, it is demonstrated through the present disclosure that the method of using deserorelin as described herein is a reliable, reversible, non-surgical sterilization solution to promote the well-being of pet families while keeping options open.

[0207] The effect on the testosterone pathway, particularly the reduction of its blood levels, may help prevent kidney disease in the animals, as these types of diseases are known to be associated with the accumulation of molecules such as trimethylamine.

[0208] It should also be mentioned that, according to safety studies in cats, administration of high doses of deserorelin showed neither serious adverse events nor life-threatening events. Furthermore, according to several tolerability studies conducted in male and female species, it was concluded that deserorelin, especially when administered as an implant (i.e., Susorelin®), is well tolerated.

[0209] Another object of the present invention is the use of deserorelin for inducing temporary infertility in healthy female animals, preferably prepubertal animals such as female cats or female dogs, for a period of at least 6 months, which comprises administering deserorelin subcutaneously or intramuscularly to an animal in need thereof. Indeed, as shown in Example 14, deserorelin is effectively administered to prepubertal female dogs to delay the onset of puberty by using at least one deserorelin implant. Example

[0210] Example 1: Determination of the changes in plasma concentrations of desorelin over time in cats

[0211] The aim of this study was to determine the time-dependent plasma concentrations of deserorelin in the plasma of healthy male cats after a single subcutaneous (SC) administration of a 4.7 mg implant containing hydrogenated palm oil, lecithin and anhydrous sodium acetate as excipients.

[0212] 1.1 Materials and Methods

[0213] • Research phase: D0 to D560 (18 months)

[0214] • n=20 adult, sexually mature, healthy male cats

[0215] • Blind, randomized

[0216] • Suyiqing® group: n=16 => 4.7 mg Suyiqing® SC, neck

[0217] • Control group: n=4 => 0.5 ml saline SC, neck

[0218] Blood samples were collected from 16 healthy able-bodied male cats that received the desorelin implant. Samples were analyzed using a validated liquid chromatography-tandem mass spectrometry (LC-MS / MS) method after solid extraction using Oasis WCX μElution plates. The calibration range was 4 pg / mL ± 2000 pg / mL. Measured concentrations below the lower limit of quantitation (LLOQ) of 4 pg / mL were reported as BLQ (below the limit of quantitation).

[0219] Noncompartmental pharmacokinetic analysis of individual deserorelin concentrations in cat plasma was performed using Phoenix software (WinNonlin 8.0, NLME 1.6, Certara LP, Pharsight, St. Louis, MO, USA) to estimate the pharmacokinetic parameters of deserorelin.

[0220] 1.2 Results:

[0221] Table 1 below shows a set of data on the changes in plasma deserorelin concentration (pg / ml) over time in cats treated with Suyiqing®:

[0222] Table 1:

[0223]

[0224] From this example, it can be determined that deserorelin is present in the blood at very high concentrations at D0, which then continue to decline over time. In some cats, deserorelin remains in the blood for up to 560 days.

[0225] Example 2 :Determination of testosterone plasma concentrations in cats

[0226] The aim of this study was to determine plasma testosterone concentrations to monitor the efficacy of the desorelin implant.

[0227] 2.1 Materials and Methods

[0228] Samples were collected from 16 healthy, sound male cats that received deserorelin implants (4.7 mg). Four healthy, sound male cats in the control group were treated with saline solution. As previously described, testosterone was analyzed using a recognized and effective radioimmunoassay (RIA) (Basal testosterone concentrations after the application of a slow-release GnRH agonist implant are associated with a loss of response to buserelin, a short-term GnRH agonist, in the tom cat, Goericke-Pesch et al., Theriogenology. 2013 Jul 1; 80 (1): 65-9.). The detection limit was 0.05 ng / mL. The intra-batch coefficient and inter-batch coefficient were 3.7% and 7.6%, respectively.

[0229] 2.2 Results:

[0230] The results are as follows Figure 1 , where the geometric mean and its 95% confidence interval (95% CI) of the ratio of testosterone after GnRH stimulation / before GnRH stimulation are expressed as a function of time (in days).

[0231] At baseline, the ratio of post-GnRH stimulation / pre-GnRH stimulation was 9.39 for the control group and 5.65 for the treatment group. There was no significant difference between the two groups. The ratio of the treatment group / control group and its 95% confidence interval were 0.60 [0.18; 1.96] and p = 0.3785.

[0232] On day 14, the ratio post-GnRH stimulation / pre-GnRH stimulation was significantly higher in the control group, with the ratio of the treatment group / control group equal to 0.07 (p<0.0001), and these differences persisted until day 252, since on day 280 the ratio of the treatment group / control group was 1.37, p=0.7004. From day 308 to day 364, the superiority of the control group was again established (p<0.01). On day 392 and beyond, no statistically significant differences were observed.

[0233] Conclusion: This example shows that testosterone concentrations increased during the desorelin implant period (burst period) and then decreased from the second week to day 252. From this day on, testosterone concentrations increased slowly and returned to physiological concentrations.

[0234] Embodiment 3: Effect of Desorelin on Sperm Concentration in Male Cats

[0235] The aim of this study was to investigate whether deserorelin has an effect on sperm concentration in male cats.

[0236] 3.1 Materials and Methods

[0237] The test was carried out in the treatment group male cats of Example 2 and in the control group male cats (see point 2.1 of Example 2).

[0238] Place 1 µL of semen in 199 µL of citrate formalin to calculate sperm concentration in the sample and assess sperm morphology - when sufficient volume is available. Determine sperm concentration by standard operating procedures. Dilution ratio is 1:200.

[0239] 3.2 Results:

[0240] The results are shown in Table 2 and Figure 2 Shown are sperm concentrations ( / μL) and their interquartile ranges as a function of time (in days).

[0241] Although lower sperm concentrations were observed in the treatment group after day 56 and reached a median of 0 between days 112 and 364, the distribution of changes from baseline between the two treatment groups was not statistically significant except at day 252 (p=0.0262). The changes in sperm concentration ( / μL) over time were measured in the treatment group cats (SpeedyQ®). Details are shown in Table 2 below.

[0242] Table 2:

[0243]

[0244] Example 4: Determination of a direct correlation between injected desorelin concentrations and testosterone concentrations

[0245] The aim of this study was to investigate whether there is a direct correlation between injected deserorelin concentrations and testosterone concentrations.

[0246] 4.1 Materials and Methods

[0247] The significant relationship between time-varying plasma deserorelin concentrations and PD parameters was evaluated graphically using testosterone and deserorelin concentrations obtained from 16 healthy male cats treated with Sulfamethoxazole 4.7 mg.

[0248] Serum concentrations of testosterone were measured according to the protocol detailed in Example 2, point 2.1.

[0249] Desserelin plasma concentrations were measured according to the protocol detailed in Example 1, point 1.1.

[0250] 4.2 Results:

[0251] Figure 3 The results of the mean deserorelin plasma concentration and the mean testosterone (before and after GnRH stimulation) serum concentration over time are shown (treatment group cat n = 16). In this figure, the mean deserorelin plasma concentration (black solid circle) (in pg / mL), the mean testosterone serum concentration before GnRH stimulation (black solid square) (in pg / mL) and the mean testosterone serum concentration after GnRH stimulation (black solid triangle) (in pg / mL) are expressed as a function of time (days).

[0252] Conclusion: This study shows an inverse correlation between desorelin and testosterone. In fact, once desorelin concentrations become low (below 12 pg / mL), testosterone concentrations begin to increase.

[0253] Embodiment 5: Determination of Deserelin Plasma Concentrations over Time and Validation of the LC-MS / MS Method

[0254] The objectives were to determine the plasma concentrations of deserorelin over time and to validate the LC-MS / MS method for the determination of deserorelin in plasma of healthy male cats following a single subcutaneous administration of the 4.7 mg implant of Suyiqing®.

[0255] 5.1 Materials and Methods

[0256] Samples were collected from 10 healthy male cats treated with the Suyiqing® 4.7 mg implant. The samples (based on a plasma volume of 150 μL, treated with K2-EDTA as anticoagulant) were analyzed by LC-MS / MS method after a solid phase extraction stage using μElution Oasis® WCX plates. The compound (13C,15N)-Leu(7)-deserelin was used as standard. The calibration range was 4 pg / mL to 2000 pg / mL.

[0257] 5.2 Results:

[0258] The results of the measurements of deserorelin plasma concentrations over time in cats treated with the deserorelin implant (i.e., Suzerain® 4.7 mg) are shown in Table 3 below.

[0259] Table 3:

[0260]

[0261] Conclusion: This example shows that deserorelin is present in the blood at very high concentrations at D0, and then the concentration decreases over time. In some cats, deserorelin remains in the blood for up to 497 days.

[0262] Embodiment 6: Determination of serum testosterone concentration by radioimmunoassay (RIA)

[0263] 6.1 Purpose:

[0264] Following a single SC administration of the 4.7 mg Suyiqing® implant in healthy male cats, serum testosterone concentrations were collected to provide information on the effect of the implant on baseline testosterone levels and on the ability to produce testosterone after stimulation.

[0265] 6.2 Materials and Methods

[0266] Samples were collected from 10 healthy male cats treated with Suyiqing®. Samples were analyzed using a radioimmunoassay (RIA) method with a validated extraction method and an LLOQ of 0.01 ng / mL. Samples were analyzed in duplicate with a volume of 350 μL to allow for repeated analysis. Only results between 0.01 ng / mL and 2 ng / mL with a coefficient of variation (CV%) less than or equal to 20% are reported.

[0267] 6.3 Results:

[0268] The results of the measurement of serum testosterone concentrations over time following a single SC administration of the 4.7 mg Suyiqing® implant to healthy male cats are shown in Table 4 below.

[0269] Table 4

[0270]

[0271] Embodiment 7: Effect of Desorelin on Testis Size

[0272] 7.1 Purpose:

[0273] The aim was to see whether desmorelin had an effect on testicular size.

[0274] 7.2 Materials and Methods

[0275] To conduct this study, two groups of cats were set up:

[0276] - Control group (5 cats, healthy male cats)

[0277] - Suyiqing® 4.7 mg treatment group (10 cats, healthy male cats)

[0278] Testicular size (length, width, and depth of each testicle in mm) was measured in male cats using calipers and recorded to the nearest tenth of a millimeter.

[0279] 7.3 Results:

[0280] Results of the curve of mean testicular volume over time (n=5 cats in the control group and n=10 cats in the treatment group) are shown in Figure 4 shown.

[0281] At baseline, the mean testicular volume was similar in the control and treatment groups, at 323.5 mm 3 and 340.8 mm 3 , and the median values ​​were 318 mm 3 and 324.5 mm 3 The mean testicular volume of the control cats increased dramatically over time, reaching 3661.5 mm at 71 weeks / 18 months. 3 (a 10-fold increase from baseline), while the mean testicular volume of the treated cats fluctuated steadily, with a final value of 1394.8 mm 3 (3-fold increase over baseline).

[0282] As early as 4 weeks / 1 month, there was a significant difference in mean testicular volume between the two groups, with the mean testicular volume of the control cats (mm 3 ) and 95% CI was 664.0 [465.7; 862.3], and the mean testicular volume of the treated cats was 466.4 [388.3; 544.5]. The observed difference and 95% CI between the treated and control cats was -197.3 mm 3 [-347.7; -47.5]. The difference between the two treatments remained significant until the end of the study (-2266.7 mm 3 [-3313.7; -1219.6]), mean testicular volume of control cats at 71 weeks / 18 months (mm 3 ) and 95% CI was 3661.5 [3006.4; 4316.4], the mean testicular volume of cats in the treatment group (mm 3 ) and 95% CI 1394.8 [676.8; 2112.7]. When comparing the testosterone ratio after GnRH stimulation / before GnRH stimulation, there were no significant differences between the groups starting from D364 / week 53 (one calendar year), indicating that reversibility of the effect on testicular volume occurs later.

[0283] Conclusion: This study showed that desorelin-induced testicular volume reduction was observed in cats.

[0284] Example 8:Analysis of pharmacokinetic parameters of deserorelin in dogs

[0285] The aim of this study was to perform a descriptive analysis of the pharmacokinetic parameters of deserelin in dogs.

[0286] 8.1 Materials and Methods:

[0287] The study was conducted in 6 healthy male dogs implanted with Deserelin® (4.7 mg), from which blood was collected during the study period (approximately 15 months). The aim of this bioanalytical study was to validate the LC-MS / MS method for the determination of Deserelin in dog plasma (based on a plasma volume of 150 µL and K2-EDTA as anticoagulant). The lower LOQ (LLOQ) in dog plasma was determined to be 4 pg / mL.

[0288] 8.2 Results:

[0289] The results of the time course of deserorelin plasma concentrations (pg / ml) measured in dogs treated with Suyiqing® are shown in Table 5 below.

[0290] Table 5:

[0291]

[0292] Conclusion: This example establishes that deserorelin is present in the blood at very high concentrations within the first few hours of administration, and then the concentrations decline over time. By the end of the study (D441), plasma deserorelin concentrations were quantifiable in some dogs, with a mean concentration of 2 pg / mL.

[0293] Embodiment 9: Effect of Desorelin on Testosterone Concentration

[0294] 9.1 Purpose:

[0295] The aim of this study was to investigate whether the desorelin implant has an effect on testosterone concentrations.

[0296] 9.2 Materials and Methods:

[0297] The study was conducted in 6 healthy male dogs implanted with Suppressant® (4.7 mg) and blood was collected from the dogs during the study period (approximately 15 months).

[0298] Before testing, testosterone was extracted from serum. Serum samples were thawed at room temperature, vortexed and processed for steroid extraction. Briefly, 500 μL of serum was transferred to a clean glass tube and 2.5 mL of ethyl acetate was added. The mixture was vortexed for 3 minutes and after the layers were separated, the upper phase was transferred to a clean test tube. This process was repeated twice. The combined extracts were then evaporated by heating to 30°C under a gentle stream of compressed air. The dried extracts had been stored at -20°C for 18 hours before analysis.

[0299] Testosterone levels in dog plasma were measured using the Cayman Chemical Testosterone ELISA kit (ref 582701, Ann Arbor, USA).

[0300] 9.3 Results:

[0301] The mean ratios of testosterone after GnRH stimulation / before GnRH stimulation (and their 95% CIs) for the treated dogs (n=6) measured over the approximately 16 months of the study are shown in Table 1. Figure 5 shown.

[0302] Conclusion: This study showed that at the time of implant injection (D0), there was an initial stimulation period of testosterone (D0 to D28), followed by a suppression period from 28 days to 252 days. Therefore, the injection of the implant induces a long-term decrease in testosterone.

[0303] Embodiment 10: Effect of Desorelin on LH Concentration

[0304] 10.1 Materials and Methods:

[0305] The study was conducted in 6 healthy male dogs implanted with Suppressant® (4.7 mg) and blood was collected from the dogs during the study period (approximately 15 months).

[0306] LH was detected using the ELISA kit “LH Detect®” (ReproPharm Vet, France). The serum was diluted 1:5 and incubated with the coated capture antibody. After washing, the secondary antibody was incubated. After washing, the conjugated tertiary antibody was finally incubated. After washing again, the bound conjugated antibody was detected by using tetramethylbenzidine (TMB) as a substrate. The reaction was terminated with an acidic solution. The absorbance was read at 450 nm and the sample absorbance values ​​were compared with the calibration curve values ​​(0 ng / mL to 8 ng / mL) to determine the sample concentration. The LH values ​​are expressed in the unit of ng / mL.

[0307] 10.2 Results:

[0308] The mean ratio (95% CI) of LH after GnRH stimulation / before GnRH stimulation was measured in the treated dogs (n=6) during the approximately 15-month study period. Figure 6 shown.

[0309] At baseline, the mean ratio after GnRH stimulation / before GnRH stimulation was 15.8. From D28 to D168, the mean ratio was equal to 1. On day 252, the ratio began to increase, reaching a mean of 5.8. From then until the end of the study, the mean ratio was approximately 10.

[0310] From D28 to D168, all dogs had ratios below 2. From D196 to D224, only one dog had a ratio greater than or equal to 2. After D252, all dogs had ratios ≥ 2.

[0311] Example 11: Correlation between the concentration of injected desorelin and the concentrations of the hormones LH, FSH and testosterone in male cats treated with Suyiqing®

[0312] 11.1 Purpose:

[0313] The aim of this study was to investigate whether there is a direct correlation between the concentration of injected deserorelin and the concentrations of the hormones LH, FSH and testosterone in male cats treated with Suzerine® (4.7 mg).

[0314] 11.2 Materials and Methods:

[0315] The study was conducted in 6 healthy male dogs implanted with Sucre® (4.7 mg) from which blood was collected during the study period (approximately 15 months). The apparent relationship between plasma desorelin concentrations and PD parameters over time was evaluated graphically using testosterone concentrations / LH concentrations / FSH concentrations and desorelin concentrations obtained from 6 healthy male cats treated with 4.7 mg Sucre®.

[0316] 11.3 Results:

[0317] The results are as follows Figure 7 shown.

[0318] Figure 7 An initial peak is shown, followed by a sharp drop in mean LH concentration, followed by a sharp drop in testosterone concentration (and to a lesser extent FSH), and over time, deserorelin plasma concentrations gradually decrease relative to testosterone concentrations, LH concentrations, and FSH concentrations before GnRH stimulation, this decrease is interrupted at D224 when deserorelin concentrations are 12 pg / mL ± 10.4 pg / mL, and all hormone levels increase.

[0319] Conclusion: This study shows that deserorelin is inversely correlated with testosterone / LH and FSH. In fact, once deserorelin concentrations become low (below 12 pg / mL), the other hormone concentrations begin to increase.

[0320] Embodiment 12: Cat Field Research

[0321] The purpose of this study was to demonstrate the durability of fertility suppression and sterility in male cats.

[0322] Materials and methods

[0323] This example presents data collected from 2 combined studies (double-blind placebo-controlled randomized) that were conducted sequentially according to the following schedule:

[0324] Study 1: Study 1 was conducted in a group of 205 male cats divided into two groups, a control group (n=51-administered subcutaneously) that received 1 mL of saline solution and a treatment group (n=154) that received the 4.7 mg implant of Sulfamethoxazole (subcutaneous implant-deserelin acetate).

[0325] Study 2: At the end of the 12-month first period, the SuYeXing® implant was removed from all male cats belonging to the treatment group. Then, saline solution (1 mL subcutaneously) was administered to 12 male cats from the treatment group of Study 1 (Saline Group), and 22 cats from the treatment group of Study 1 were re-implanted with new SuYeXing® 4.7 mg implants for a 12-month second period (Twice Treatment Group). The efficacy and safety of the re-implantation were followed up for this additional 12 months, for a total of 24 months.

[0326] The observation schedule is as follows:

[0327] Study 1: V1: Day -14 to Day -7; 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;

[0328] 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.

[0329] Regarding the inclusion criteria, the criteria for both studies were as follows:

[0330] - House cat

[0331] - Age > 3 months or > 15 months in Study 2

[0332] - Healthy indoor male

[0333] - Client Owned (including shelter cats / cats rehomed from foster homes)

[0334] The additional criterion for Study 2 was success in Study 1.

[0335] In these studies, the following primary endpoints were measured:

[0336] - For Study 1: Suppression of fertility (testosterone ≤ 0.10 ng / mL *from V4 to V7* – 3 months to 12 months)

[0337] - Study 2: Suppression of fertility (testosterone ≤ 0.10 ng / mL *from V8 to V13* – 12 months to 24 months) and time to reversal of effect.

[0338] In addition to safety parameters (hematology and blood biochemistry, urinalysis, local reactions at the implantation site, changes in body weight, appetite, assessment of abnormal testicles, adverse events), the following secondary endpoints were reported:

[0339] - Sexual Behavior Total Score

[0340] - Aggression

[0341] - Vocalization

[0342] - Urine markers

[0343] - Urine odor / smell

[0344] - Penis piercing

[0345] - Testicular volume

[0346] 12.2 Results:

[0347] - Testosterone :

[0348] Figure 8 Mean percentage efficacy results are shown. Overall, the success rates were very similar in both studies for either 1 or 2 implants, with efficacy >87% at all time points in Study 2.

[0349] Figure 8 B and Figure 8 C shows the detailed levels of testosterone for Study 1 and Study 2, respectively.

[0350] - Reversibility :

[0351] The reversibility of the effects of deserelin was investigated. In fact, n=12 subjects in the deserelin-treated group of Study 1 were enrolled in the saline group of Study 2, as described above in point 12.1.

[0352] Results showed that cats from the treated group in Study 1 who were enrolled in the saline group in Study 2 had 100% reversal by D279 following enrollment in Study 2. The median time to reversibility based on serum testosterone concentrations was D379 ± 178.

[0353] - Secondary End Points :

[0354] Efficacy results for secondary endpoints are reported as follows:

[0355] · Fig. 9 A and Fig. 9 B shows the mean of the total sexual behavior scores in Study 1 and Study 2, respectively.

[0356] · Fig.10 A and Fig.10 B shows the mean phonation changes in Study 1 and Study 2, respectively.

[0357] · Fig.11 A and Fig.11 B shows the % change in urine markers from baseline in Study 1 and Study 2, respectively.

[0358] · Fig.12 A and Fig.12 B shows the change / reduction in urine odor measured as change from baseline in Study 1 and the change / reduction in urine odor measured as change from baseline in Study 2, respectively.

[0359] · Fig.13 A and Fig.13 B shows the % change in testicular volume in Study 1 and the % change in testicular volume in Study 2, respectively.

[0360] · Fig.14 A and Fig.14 B shows the % change in penile thorn appearance in Study 1 and the % change in penile thorn appearance in Study 2, respectively.

[0361] During the study, there were no safety issues and the desmorelin implant was well tolerated.

[0362] In summary, these combined studies demonstrate that:

[0363] Deserorelin was effective in chemical castration of subjects for at least 12 months. This effect has been shown to be reversible at the end of the treatment course. These studies also demonstrated that sexual behavior had been reduced and that the onset of action in reducing behavior was rapid (approximately 7 to 31 days). In addition, no safety issues were seen with either single or repeated administration.

[0364] Therefore, it can be concluded that extended-release administration of deserorelin, especially via implant, is a good alternative to surgical castration and a safe, nonsurgical method of controlling reproduction in male cats.

[0365] The method of using desorelin as described herein is a reliable, reversible, nonsurgical sterilization solution to promote the well-being of pet families while keeping options open.

[0366] Embodiment 13: PK / PD Preclinical Study in Healthy Male Cats (Healthy Male Cats)

[0367] The objectives of this study were to demonstrate inhibition of fertility, determine deserorelin levels using PK data, assess reversibility using PD analysis, and document systemic and local tolerability.

[0368] This example provides data collected from a PK / PD preclinical study in healthy male cats according to the following study design:

[0369] - Product: Suyiqing® 4.7 mg versus placebo (sodium chloride solution)

[0370] - Subjects: Cats treated at 15 weeks of age = 15 + 3 healthy male cats (15 weeks of age) were further divided into two groups; one treated with the 4.7 mg implant of Suppressant® (Group 2, n=13) and the other treated with saline (placebo) (Group 1, n=5). On D14, the extra three cats were removed from the study. 5 + 3 female cats (Group 3) who were pubertal at the start of the study were included in the study only for the evaluation of the reproductive behavior of the male cats and were not treated with the implant.

[0371] - Duration: 18 study months post-injection (PI) (D502 / week 72, i.e. 16.5 calendar months)

[0372] Table 6 below gives the details of the study:

[0373] Table 6

[0374]

[0375] In this table, BW stands for body weight

[0376] Research findings:

[0377] - PK study results such as Fig.15 A. Overall, it should be noted that at 2 hours Tmax, there was an average decrease of 92% in the first 24 hours post-injection (PI) followed by a steady and slow decrease. The first cat had undetectable concentrations 1 year after injection (52 weeks post-injection) and this was also the case in 5 / 10 male cats at the end of the study.

[0378] - Testosterone levels recorded as Fig.15 Figure 2 is shown in Figure B and shows that testosterone was low in the treated cats until one year after injection (53 weeks after injection) and then increased (except for 3 / 10 cats). At the end of the study, mean testosterone levels in the treated cats remained low (1.59 ng / mL and 2.16 ng / mL, respectively, in the treated / control groups before GnRH stimulation and 2.34 ng / mL and 6.44 ng / mL, respectively, in the treated / control groups after GnRH stimulation).

[0379] It is noteworthy that the ratio of post-GnRH stimulation / pre-GnRH stimulation remained statistically significant from 2 weeks to 1 year after injection (p=0.0001).

[0380] - Fig.15 C and Fig.15 D shows the recordings of penile pricking and behavior, showing that the results of penile pricking were statistically significant from 7 weeks after injection to 67 weeks. Regarding sexual behavior, the results were statistically significant after 10 weeks, 12 weeks, 34 weeks, 36 weeks, 38 weeks, 46 weeks, 52 weeks, 58 weeks, and 66 weeks after injection. Finally, regarding reproductive behavior, no statistical differences were noted, with one control male mating 35 weeks after injection (at 1 year of age) and one cat from the treatment group mating 65 weeks after injection. Personality traits in cats can affect sexual desire. It was also noted that Fig.15 Trends for decreased vocalization, aggression, urine marking, and typical male urine odor shown in D. It can be clearly seen from the figure that cats from the control group (animals C1 to C5) displayed more sexual activity or reproductive behaviors, including vocalization, aggression, urine marking, and typical male urine odor, compared to animals treated with deserorelin implants (T1 to T10).

[0381] - In 5 healthy control cats, puberty occurred at 5.5 months of age. The onset of puberty in cats treated with desmorelin was delayed and persisted for at least 1 year.

[0382] At the end of the study (72 weeks after injection), it was effective in completely reversing male reproductive and hormonal parameters in 5 / 10 of the treated cats.

[0383] Embodiment 14: Use of Desorelin in Prepubertal Female Dogs

[0384] The aim of this study was to evaluate the delay in the onset of puberty in prepubertal female dogs. It is known that puberty starts with the establishment of several parameters:

[0385] - Increased number of receptors in the central nervous system (CNS) and gonads

[0386] - Hypothalamic / pituitary desensitization to estrogen

[0387] - Maturation of gonads and secondary sex organs

[0388] - Follicle maturation, spermatogenesis

[0389] The results of this study suggest that delaying puberty is possible:

[0390] Implantation 1: No physical development disorder, normal vulva size, but delayed epiphyseal closure. There may be mild flare symptoms (vulva swelling)

[0391] Implantation 3 times, every 4.5 months: noted that the vulva remained immature, with an effect comparable to prepubertal castration.

[0392] Of note, desrelin concentrations were not associated with the duration of efficacy, and implant removal was not necessary.

[0393] This study allowed the definition of criteria for the administration of deserorelin to prepubertal female dogs:

[0394] - The age for implant placement should be between 4 months and 5 months.

[0395] - The onset of puberty must be excluded, which is determined by clinical examination, vaginal examination, vaginal cytology, E2, DHEA measurement (sexual activity).

Claims

1. Deserorelin for use in chemical castration of a non-human mammal within 1 month after administration, wherein at the time of administration, the Cmax of deserorelin after administration exceeds 7000 pg / mL and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL.

2. Deserorelin for use according to claim 1, wherein deserorelin is administered subcutaneously or intramuscularly to a non-human mammal in need thereof at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 to 24 hours and a plasma concentration of less than 400 pg / mL starting from 8 days after administration.

3. Deserorelin for use according to claim 2, wherein the plasma concentration of deserorelin is 10 pg / mL to 400 pg / ml from 8 days to at least 6 months after administration.

4. Deserorelin for inducing temporary infertility in healthy male animals for a period of at least 6 months, comprising administering deserorelin subcutaneously or intramuscularly to a mammal in need thereof, in an amount effective to induce a plasma concentration of deserorelin of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours and a plasma concentration of less than 400 pg / mL from 8 days after administration.

5. Deserorelin for use according to claim 4, wherein after administration, deserorelin is released substantially continuously at a plasma concentration of less than 400 pg / mL from 8 days to at least 6 months after administration.

6. A sustained-release drug delivery system (DDS) containing deserorelin for chemical castration of a non-human mammal, wherein the DDS releases deserorelin with a post-administration deserorelin Cmax of more than 7000 pg / mL, and the deserorelin concentration in the plasma of the non-human mammal is at least 10 pg / mL and is maintained for at least 6 months.

7. The sustained-release drug delivery system (DDS) according to claim 6, wherein the DDS is a subcutaneous implant or an intramuscular implant comprising deserorelin and a biodegradable material.

8. Deserorelin for use in chemical castration of a non-human mammal, wherein after administration, deserorelin reduces the testosterone level in the plasma of the non-human mammal to below 0.4 ng / ml and the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / ml, preferably above 15 pg / ml.

9. A method of contraception for a non-human mammal comprising administering subcutaneously or intramuscularly to the non-human mammal an implant comprising deserorelin and a biodegradable material, such as a biodegradable polymer, wherein the implant releases deserorelin at a Cmax of more than 7000 pg / mL 1 hour after administration, and wherein the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL and is maintained for at least 6 months.

10. The contraceptive method of claim 9, wherein deserorelin is administered to a non-human mammal in need thereof at a dose effective to induce a deserorelin plasma concentration of 4000 pg / mL to 40000 pg / mL within 1 hour to 24 hours and a plasma concentration below 400 pg / mL from 8 days after administration.

11. The contraceptive method according to claim 9 or 10, wherein the plasma concentration of desorelin is less than 400 pg / ml from 8 days to at least 6 months after administration.

12. Deserorelin for use in preventing behavioral disorders in a non-human mammal, wherein deserorelin is administered subcutaneously or intramuscularly via a sustained release drug delivery system, and wherein the concentration of deserorelin in the plasma of the non-human mammal is at least 10 pg / mL.

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