Dosing regimen of heterocyclic compound as janus kinase inhibitor

Abstract

The present invention relates to: a use of a Janus kinase (JAK) inhibitor for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals that includes dogs and cats; and a prevention or treatment method using same.

Description

[0001]

Description of the Invention

[0002]

Title of Invention

[0003] Administration method of heterologous compounds as Janus kinase inhibitors

[0004]

Technology Field

[0005] The present invention relates to Janus kinase (JAK) inhibitors, and more specifically, to the use, administration method and dosage of JAK inhibitors useful for the prevention or treatment of allergic dermatitis, atopic dermatitis or pruritus, and a method using the same.

[0006]

Background Techniques

[0007] Protein kinases are enzymes that regulate various intracellular processes by phosphorylating interacting proteins to control their activity, localization, and function. Janus kinases (JAKs) are located at intracellular receptors, and when inflammatory cytokines bind to these receptors, they undergo a phosphorylation process to phosphorylate downstream proteins, such as signal transduction and transcription activating factors (STATs), thereby transmitting the inflammatory cytokine signals into the cell. Excessive activation of these inflammatory cytokine signaling pathways can lead to the onset or exacerbation of autoimmune and inflammatory diseases.

[0008] Prior art literature

[0009] Patent Literature

[0010] (Patent Document 1) Korean Published Patent No. 2019-0043437

[0011] Non-patent literature

[0012] (Non-patent literature 1) J. vet. Pharmacol. Therap. 37, 317—324.

[0013]

Description of the Invention

[0014]

Technical Challenges

[0015] The present invention provides a use and method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in animals, particularly mammals including dogs and cats that require treatment, as a Janus kinase inhibitor.

[0016]

Technical Solution

[0017] This is explained in detail as follows. Meanwhile, each description and embodiment disclosed in the present invention may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. Furthermore, the scope of the present invention should not be considered limited by the specific descriptions provided below.

[0018] The present invention provides a pharmaceutical composition comprising, as an active ingredient, a heterocyclic compound of the following formula 1 or a pharmaceutically acceptable salt thereof, for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals including dogs and cats.

[0019] [Chemical Formula 1]

[0020]

[0021] In the present invention, the pharmaceutical composition is administered orally at a concentration of 0.4 to 1.35 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropancarboxamide.

[0022] The name of the compound of Chemical Formula 1 above is N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide, (R)-N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide, (S)-N-(4-(1-(2-cyanoacetyl]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)- The invention includes 1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or racemic mixtures thereof. In the present invention, the compound of Formula 1 may preferably be (S)-N-(4-(1-(2-cyano-atyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, but is not limited thereto. The pharmaceutically acceptable salt of the heterocyclic compound in the present invention may be a hydrochloride (HC1), hydrogen bromide (HBr), methylate, phosphate, napadisylate, camsylate, or oxalate, and specifically may be a phosphate, but is not limited thereto.

[0023] In the present invention, the compound of Formula 1 or a pharmaceutically acceptable salt thereof may be a Janus kinase (hereinafter, JAK) inhibitor, specifically a JAK1 selective inhibitor.

[0024] <Pharmaceutical composition, use, or method for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus>

[0025] The present invention provides a pharmaceutical composition, use, or method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals including dogs and cats.

[0026] The present invention relates to a pharmaceutical composition comprising N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals,

[0027] The above pharmaceutical composition provides a pharmaceutical composition administered orally at a concentration of 0.4 to 1.35 mg / kg based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo[2,3- b]pyridin- 6-yl)cyclopropanecarboxamide.

[0028] Specifically, the above pharmaceutical composition comprises a pharmaceutically acceptable salt (e.g., phosphate salt) of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, and may be administered orally at a concentration of 0.4 to 1.35 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0029] More specifically, the above pharmaceutical composition comprises the phosphate salt of (S)-N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, and may be administered orally at a concentration of 0.4 to 1.35 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0030] As used herein, the term "clinical sign" means an observable or measurable pathological condition or behavior in an individual exhibiting a disease, illness, or symptom. Clinical signs may be symptoms, illnesses, or behaviors that can be measured by known or established diagnostic evaluations. For example, a definitive diagnostic evaluation of allergic dermatitis or atopic dermatitis can be measured by Visual Analog Scale (VAS) scores, clinical pathological evaluations, or established scoring systems, such as the Canine Atopic Dermatitis Degree and Severity Index (CADESI) score (e.g., CADESI-04 score).

[0031] The term "PVAS (Pruritus Visual Analogue Scale)" used at this clinic is a clinical indicator that evaluates the degree of itching (pruritus) in pets observed by guardians on a scale of 0 to 10, and "CADESI (Canine Atopic Dermatitis Extent and Severity Index)" is a clinical indicator that quantifies the extent and severity of skin lesions (erythema, redness, lichenification, abrasions, hair loss, etc.) evaluated by a veterinarian.

[0032] Examples of some clinical signs of atopic dermatitis and allergic dermatitis that may sometimes be applied in such evaluation or scoring systems include the following: severe (in companion animals, e.g., dogs, manifesting primarily as almost continuous scratching, chewing, and licking), severe (presenting itching while awake, and persistence of itching symptoms at night and / or while eating, playing, or exercising), moderate (often presenting as itching symptoms), and very mild (intermittent itching symptoms) levels of itching; presence of pustules or epidermal collarets; presence of skin lesions; pruritus; erythema; erosions, scratches, and / or spontaneous alopecia; presence of papules and / or crusts; lichenification and / or hyperpigmentation.

[0033] "Symptoms" of a disease or illness are any symptoms known to experts in the field related to the disease or illness. In atopic dermatitis, allergic dermatitis, flea allergy dermatitis, and sarcoptic mange, symptoms include, for example, pruritus, itching, and skin lesions.

[0034] In many cases, the "symptoms" of a disease or condition, such as atopic dermatitis or allergic dermatitis, are also "clinical signs."

[0035] Allergic dermatitis may be, for example, flea allergic dermatitis, i.e., FAD (also referred to as flea allergic dermatitis, flea bite dermatitis (FBD), or flea-associated dermatitis), food allergic dermatitis, contact dermatitis, or Sarcoptes scabiei (i.e., sarcoptes scabiei) associated allergic dermatitis, but is not limited to these.

[0036] In the present invention, "prevention" means any act of suppressing or delaying the onset of a disease by administering the compound of Formula 1 of the present invention or a pharmaceutically acceptable salt thereof; or a pharmaceutical composition containing the same.

[0037] In the present invention, "treatment" means any act in which the symptoms of an individual suspected of having or suffering from a disease are improved or beneficially altered by the administration of the compound of Formula 1 of the present invention or a pharmaceutically acceptable salt thereof; or a pharmaceutical composition containing the same.

[0038] The pharmaceutical composition of the present invention may be manufactured in a unit dose form by formulation using pharmaceutically acceptable carriers and excipients, or by being contained in a multi-dose container. The formulation may be manufactured by conventional methods used in the art for formulation or by the method disclosed in Remington's Pharmaceutical Science (19th ed., 1995), and may be formulated into various formulations depending on each disease or component.

[0039] The pharmaceutical composition of the present invention can produce excellent effects when used alone, but can also be used in combination with various methods such as hormone therapy and drug therapy to increase therapeutic efficiency.

[0040] In one embodiment, the pharmaceutical composition may be administered orally once a day at a concentration of 0.4 to 1.35 mg / kg (e.g., 0.4 mg / kg, 0.6 mg / kg, 0.9 mg / kg, or 1.35 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0041] In one embodiment, the pharmaceutical composition may be administered orally once a day at a concentration of 0.9 to 1.35 mg / kg (e.g., 0.9 mg / kg or 1.35 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0042] In one embodiment, the pharmaceutical composition may be administered orally once a day at a concentration of 0.4 to 0.9 mg / kg (e.g., 0.4 mg / kg, 0.6 mg / kg, or 0.9 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0043] In one embodiment, the pharmaceutical composition may be administered orally twice a day at a concentration of 0.4 to 0.9 mg / kg (e.g., 0.4 mg / kg, 0.6 mg / kg, or 0.9 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0044] In one embodiment, the pharmaceutical composition may be administered orally twice a day at a concentration of 0.45 to 0.9 mg / kg (e.g., 0.45 mg / kg, 0.6 mg / kg, 0.9 mg / kg, or 0.6 to 0.9 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0045] In one embodiment, the pharmaceutical composition may be administered orally twice a day at a concentration of 0.6 to 0.9 mg / kg (e.g., 0.6 mg / kg or 0.9 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0046] In the present invention, the pharmaceutical composition may be administered orally for a period sufficient to alleviate, improve, or eliminate one or more symptoms or clinical signs associated with allergic dermatitis, atopic dermatitis, or pruritus in mammals including dogs and cats. For example, the pharmaceutical composition may be administered for a period of 1 to 7 days, 1 to 14 days, 1 to 21 days, 1 to 28 days, 7 to 28 days, 14 to 28 days, 21 to 28 days, or 28 days or more, or until the alleviation, improvement, or elimination of symptoms, but is not limited thereto.

[0047] In one embodiment, the pharmaceutical composition may be administered for 1 to 28 days.

[0048] In one embodiment, the pharmaceutical composition may be administered orally at a dose of 0.6 to 1.35 mg / kg (e.g., 0.6 to 0.9 mg / kg, 0.9 to 1.35 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide for a period of 1 to 28 days, 1 to 14 days, 7 to 14 days, 14 days, 28 days, or 28 days or more (until the time of relief or improvement of symptoms) once a day or twice a day.

[0049] In one embodiment, the pharmaceutical composition may be administered orally once a day for 28 days at a concentration of 0.9 to 1.35 mg / kg based on N- (4- (l- (2-cyanoacetyl)-3-methyl- 1,2,3,6-tetrahydropyridin-4-yl)- 1H-pyrrolo[2,Sb]pyridin-6-yl)cyclopropanecarboxamide.

[0050] In one embodiment, the pharmaceutical composition may be administered orally at a concentration of 0.6 to 0.9 mg / kg based on N- (4- (l- (2-cyanoacetyl)-3-methyl- 1,2,3,6-tetrahydropyridin-4-yl)- 1H-pyrrolo[2,Sb]pyridin-6-yl)cyclopropanecarboxamide twice daily for 14 days.

[0051] In one embodiment, the pharmaceutical composition is used for 1 to 14 days, 7 to 14 days, or 14 days! Administered orally at a dose of 0.6 to 0.9 mg / kg (e.g., 0.6 mg / kg or 0.9 mg / kg) based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide twice daily, and thereafter at a dose of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3- b] Pyridin-6-yl) may be administered orally at 0.6 to 0.9 mg / kg (e.g., 0.6 mg / kg or 0.9 mg / kg) based on cyclopropancarboxamide.

[0052] In one embodiment, the pharmaceutical composition may be administered orally at a concentration of 0.6 to 0.9 mg / kg based on N-(4-(l-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,Sb]pyridin-6-yl)cyclopropanecarboxamide twice daily for 14 days, and then orally at a concentration of 0.6 to 0.9 mg / kg once daily for 14 days.

[0053] In one embodiment of the present invention, the mammal may be a companion animal such as a dog or a cat. In another embodiment, the mammal may not be a human. In yet another embodiment, the mammal may be a dog or a cat, specifically a dog, but is not limited thereto. The mammal may be an individual that is unresponsive to ochlasitinib or a pharmaceutically acceptable salt thereof, exhibits a chronic history of non-seasonal pruritus, has a PVAS score of 6 or higher based on baseline (Day 0, Baseline), or has an overactivated JAK1-STAT3 signaling pathway compared to a normal individual, but is not limited thereto.

[0054] In one embodiment, the pharmaceutical composition may improve, alleviate, or treat one or more symptoms selected from the group consisting of itching, pruritus, redness, lichenification, abrasions, and hair loss, but is not limited thereto.

[0055] In the present invention, Visit 1 (Day 0), Visit 2 (Day 14 ± 3 or Day 14 ± 2) and Visit 3 (Day 28 ± 3 or Day 28 ± 2) refer to the number of visits (days elapsed from the start date of the clinical trial).

[0056] In the present invention, non-response to oklasitinib or a pharmaceutically acceptable salt thereof means that the PVAS score does not improve by 2 points or more even after administering oklasitinib or a pharmaceutically acceptable salt thereof (e.g., Apoquel® tablets).

[0057] In the present invention, chronic non-seasonal pruritus refers to pruritus that occurs without being limited to a specific environment or season and persists for a certain period (e.g., lasting for 6 weeks or more). Chronic non-seasonal pruritus can be diagnosed based on taking a medical history, clinical symptoms of atopic dermatitis as presented by ICADA (International Committee for Allergic Diseases of Animals), Favort's criteria, or allergy tests (serological allergen-specific IgE test, intradermal inoculation test).

[0058] In the present invention, the pharmaceutical composition may be such that upon repeated administration (administered two or more times), no accumulation in plasma of N-(4-(1-(2-cyano-atethan]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof occurs, and / or the plasma concentration does not reach the IC5o for JAK2.

[0059] In the present invention, the pharmaceutical composition may be such that even when administered at high doses, the plasma concentration of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof does not reach an IC5o for JAK2.

[0060] Specifically, the above high dose administration may be N- (4- (1- (2-cyanoacetyl)- 3-methyl-1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo[2,3- b]pyridin- 6-yl)cyclopropancarboxamide administered at a dose of 0.4 mg / kg or more. More specifically, N- (4- (1- (2-cyano-ethyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo [2,3- b]pyridin- 6-yl)cyclopropanecarboxamide may be administered at a dose of 0.4 mg / kg or more, 0.5 mg / kg or more, 0.6 mg / kg or more, 0.7 mg / kg or more, 0.8 mg / kg or more, or 0.9 mg / kg or more, and more specifically, at a dose of 0.4 to 1.35 mg / kg, 0.4 to 0.6 mg / kg, 0.6 to 0.9 mg / kg, or 0.9 to 1.35 mg / kg.

[0061] Specifically, the high dose administration may be a high dose single administration. Specifically, the high dose single administration may include, but is not limited to, a high dose once administration, repeated administration of a high dose once a day, etc.

[0062] In the present invention, "plasma accumulation" refers to a phenomenon in which the blood drug concentration gradually increases when a drug is administered repeatedly at regular intervals, as the next dose is administered while the previously administered drug is not completely eliminated from the body. This typically corresponds to the drug exposure amount (Area Under the Curve, AUC) or peak blood concentration (C) after a single dose. m Compared to ax), it can be evaluated as the rate of increase in the corresponding value in the normal state after repeated administration.

[0063] In the present invention, "non-accumulation in plasma" refers to the AUC or C after the initial administration of the drug. mAUC or C after the next drug administration compared to ax m This means that ax does not increase by 1.2 times or less, or that the half-life (T1 / 2), which is the time it takes for the plasma drug concentration to decrease to 50% over time, does not increase (meaning there is no significant difference from the half-life after the initial administration).

[0064] In the present invention, N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same, has high JAK1 selectivity and low JAK2 inhibitory effect, so it can target a specific pathway and minimize side effects. In addition, even if the pharmaceutical composition is administered repeatedly over a certain period (e.g., once a day or twice a day for 28 days), there is little accumulation of the drug in the body, so it can safely prevent or treat allergic dermatitis, atopic dermatitis, or pruritus in mammals while minimizing side effects.

[0065] The present invention provides a method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals, comprising the step of orally administering to a mammal a compound of Formula 1, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition as defined above containing the same. The compound of Formula 1, a pharmaceutically acceptable salt thereof, dosage, administration cycle, number of administrations, target of administration, administration effect, etc., are as described above.

[0066] In the present invention, "administration" means introducing a specific substance to an individual in an appropriate manner.

[0067] The method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus according to the present invention includes administering a compound of Formula 1, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing it, thereby not only treating the disease itself before the onset of symptoms but also inhibiting or avoiding the symptoms thereof.

[0068] In addition, the method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus according to the present invention may further include administering a therapeutically effective amount of an additional active agent that aids in the prevention or treatment of the disease together with the compound of Formula 1, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same. The additional active agent may exhibit a synergistic or additive effect together with the compound of Formula 1, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same.

[0069] The present invention provides the use of a compound of Formula 1, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition as defined above comprising the same for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals.

[0070] The present invention provides the use of a compound of Formula 1, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition as defined above containing the same for the manufacture of a drug for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals. The compound of Formula 1, the pharmaceutically acceptable salt, the dosage, the administration cycle, the number of administrations, the target of administration, the administration effect, etc. are as described above.

[0071] For the manufacture of a pharmaceutical product, a pharmaceutically acceptable adjuvant, diluent, carrier, etc. may be mixed with the compound of Formula 1 of the present invention, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition as defined above containing the same, and may be manufactured into a complex formulation with other active agents to have a synergistic effect.

[0072] The present invention comprises the following items (1) to (21) relating to the above composition, the use thereof, and a treatment method including the administration thereof.

[0073] (1) The present invention relates to a pharmaceutical composition comprising N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof as an active ingredient for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals,

[0074] The above pharmaceutical composition provides a pharmaceutical composition administered orally at a concentration of 0.4 to 1.35 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0075] (2) In the above (1), the pharmaceutically acceptable salt of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropancarboxamide may be a hydrochloride salt, a hydrogen bromide salt, a mesylate salt, a phosphate salt, a napadisilate salt, a camsilate salt, or an oxalate salt.

[0076] (3) In the above (1) or (2), the N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide may be (S)-N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0077] (4) In any one of (1) to (3) above, the pharmaceutical composition may be administered orally at a dose of 0.4 to 1.35 mg / kg once daily based on N-(4-(1-(2-cyano-atehr]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0078] (5) In any one of (1) to (4) above, the pharmaceutical composition may be administered orally at a dose of 0.9 to 1.35 mg / kg once daily based on N-(4-(1-(2-cyano-atehr]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0079] (6) In any one of (1) to (5) above, the pharmaceutical composition may be administered orally at a dose of 0.4 to 0.9 mg / kg once daily based on N-(4-(1-(2-cyano-atehr]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0080] (7) In any one of (1) to (6) above, the pharmaceutical composition may be administered orally at a dose of 0.4 to 0.9 mg / kg twice daily with N-(4-(1-(2-cyano-atehr]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0081] (8) In any one of (1) to (7) above, the pharmaceutical composition may be administered orally at a dose of 0.45 to 0.9 mg / kg twice daily based on N-(4-(1-(2-cyano-atehr]tyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide.

[0082] (9) In any one of (1) to (8) above, the pharmaceutical composition may be administered until the alleviation or improvement of symptoms of allergic dermatitis, atopic dermatitis, or pruritus.

[0083] (10) In any one of (1) to (9) above, the pharmaceutical composition may be administered for a period of 1 to 7 days, 1 to 14 days, 1 to 21 days, 1 to 28 days, 7 to 28 days, 14 to 28 days, 21 to 28 days or 28 days or more.

[0084] (11) In any one of (1) to (10) above, the pharmaceutical composition may be administered orally at a dose of 0.6 to 1.35 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide once a day for a period of 1 to 28 days or more than 28 days.

[0085] (12) In any one of (1) to (11) above, the pharmaceutical composition may be administered orally at a dose of 0.9 to 1.35 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide once a day for 28 days.

[0086] (13) In any one of (1) to (12) above, the pharmaceutical composition is administered orally at a dose of 0.6 to 0.9 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide twice daily for 1 to 14 days, 7 to 14 days, or 14 days, and thereafter at a dose of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H- Pyrrolo [2,3-b]pyridin-6-yl) may be administered orally at a dose of 0.6 to 0.9 mg / kg based on cyclopropane carboxamide.

[0087] (14) In any one of (1) to (13) above, the pharmaceutical composition is used for 14 days! It may be administered orally twice daily at a dose of 0.6 to 0.9 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide, and then orally once daily for 14 days at a dose of 0.6 to 0.9 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide.

[0088] (15) In any one of (1) to (14) above, the mammal may be unresponsive to ochlasitinib or a pharmaceutically acceptable salt thereof, exhibit a chronic history of non-seasonal pruritus, have a PVAS score of 6 or higher based on the baseline (Day 0, Baseline), or have an overactivated JAK1-STAT3 signaling pathway compared to a normal individual.

[0089] (16) In any one of (1) to (15) above, the pharmaceutical composition may improve, alleviate, or treat one or more symptoms selected from the group consisting of itching, pruritus, redness, lichenification, abrasion and hair loss.

[0090] (17) In any one of (1) to (16) above, the pharmaceutical composition may not cause accumulation in plasma of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof when administered two or more times.

[0091] (18) In any one of (1) to (17) above, the pharmaceutical composition may be such that even when administered at high doses, the plasma concentration of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof does not reach an IC50 for JAK2.

[0092] (19) The present invention provides a method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals, comprising the step of orally administering to a mammal N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, a pharmaceutically acceptable salt thereof, or a composition according to any one of (1) to (18).

[0093] (20) The present invention provides for the use of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, pharmaceutically acceptable salts thereof, or compositions according to any one of (1) to (18) above for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals.

[0094] (21) The present invention provides for the use of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropancarboxamide, pharmaceutically acceptable salts thereof, or compositions according to any one of (1) to (18) above for the manufacture of a drug for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals.

[0095] The details described above regarding the compositions of the present invention may be applied substantially identically to each composition, method, and use of the present invention, provided they do not contradict one another.

[0096] The doses described in the present invention may be doses based on the free base of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide. Specifically, the doses described in the present invention may be doses based on the free base of (S)-N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide.

[0097] Embodiments of the present invention may be modified in various different forms, and the scope of the present invention is not limited to the embodiments described below. Furthermore, embodiments of the present invention are provided to more completely explain the present invention to those skilled in the art. Moreover, throughout the specification, the term "comprising" any component means that, unless specifically stated otherwise, it does not exclude other components but may include additional components.

[0098]

Effects of the Invention

[0099] The compound of the present invention has excellent JAK1 inhibitory activity and relatively low inhibitory activity against JAK2, and since no drug accumulation occurs with repeated administration, it can reduce the risk of side effects such as thrombosis, cardiovascular adverse reactions, infection, and the development of malignant tumors, which have been pointed out as problems with existing JAK inhibitors. Therefore, the compound of the present invention can provide a safer treatment option, and tolerability is expected to be improved even in long-term treatment.

[0100]

Brief Description of the Drawing

[0101] Figure ! is the NMR result for (S)—N—(4—(1—(2—cyanoacetyl)—3—methyl—1,2,3,6—tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide phosphate of Example !.

[0102] Figure 2 shows the study design of Experimental Example 1.

[0103] Figure 3 shows the CADESI-04 score in Experimental Example 1.

[0104] Figure 4 shows the results of scoring pruritus after administration of Example 1 and the control drug in Experimental Example 1.

[0105] Figure 5 shows the transepidermal water loss after 2 weeks and 4 weeks after administration of Example 1 or the control drug as a result of Experimental Example 1.

[0106] Figure 6 shows the results of measuring the number of mast cells and epidermal thickness after administration of Example 1 or the control drug in Experimental Example 1.

[0107] Figure 7 shows the study design of Experimental Example 2. Figure 8 shows the pruritus score and the duration of pruritus (itching) as the results of Experimental Example 2.

[0108] Figure 9 shows the pruritus scores over time after administration of Example 1 and the control drug in Experimental Example 2.

[0109] Figure 10 shows the study design of Experimental Example 3, which shows that a control drug or Example 1 was orally administered to normal beagle dogs at different doses (0.2, 0.3, 0.45, 0.6, 0.9 mg / kg) twice a day for 7 days.

[0110] Figure 11 shows the change in plasma concentration over time on days 1 and 7 of Example 1 (intragastric administration, IG) or oklacitinib (oral administration, PO) as a result of Experimental Example 3.

[0111] Figure 12 shows the JAK1 activity inhibitory ability as a result of measuring the positive cells among IL-6-activated pSTAT3 / CD4 in whole blood cells of Beagle dogs treated with Example 1 or Oklasitinib on days 1 and 7 using flow cytometry.

[0112] Figure 13 shows the relationship (Day 1, Day 7) between the plasma concentration-time profile of each drug and the IC5o value for PSTAT5 (inhibitory activity against JAK2) induced by GM-CSF in TF-1 cells up to 12 hours after oral administration of 0.6 mg / kg or 0.9 mg / kg of Example 1 or 0.4-0.6 mg / kg of oklasitinib twice daily to Beagle dogs as the result of Experimental Example 3.

[0113] Figure 14 shows the plasma concentration-time profiles of each drug up to 24 hours after a single oral administration of Example 1 at 0.4–0.6 mg / kg or 0.6–0.9 mg / kg to Beagle dogs, and the IC50 for GM-CSF-induced pSTAT5 in TF-1 cells. 50 It shows the relationship between the values ​​(inhibitory ability against JAK2).

[0114] Figure 15 shows the clinical trial design of Experimental Example 4.

[0115] Figure 16 shows the method of assigning animals to participate in a clinical trial.

[0116] Figure 17 shows the estimated marginal mean change of PVAS scores during the administration period. Figure 18 shows the estimated marginal mean change of CADESI-04 scores during the administration period.

[0117]

Forms for Carrying Out the Invention

[0118] Preparation Example 1: (8)-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydro

[0119]

[0120] Synthesis of pyridin-4-yl-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide. The compound of the title was prepared according to the method disclosed in Korean Patent Publication No. 2019-0043437.

[0121] iR NMR (400 MHz, DMSO-d6) 8 11.44 (s, 1H), 10.57 (s, 1H), 7.84 (d, J = 10.2 Hz, 1H), 7.34 (d, J = 3.1 Hz, 1H), 6.48 (dd, J = 1.8, 3.7 Hz, 1H), 6.17 — 6.03 (m, 1H), 4.31 — 4.01 (m, 6H), 3.96 — 3.62 (m, 2H), 3.02 (m J = 36.6 Hz, 1H), 2.02 (s, 1H), 0.88 (s, 3H), 0.84 — 0.73 (m, 4H); MS(ESI+) m / z 364 (M+H)+

[0122] Example 1. (8) Preparation of phosphate salt of -(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b)]pyridin-6-yl)cyclopropanecarboxamide

[0123] 90 mL of methanol was added to 30 g of (S)-N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide and stirred, then heated to 50°C. Subsequently, 1.1 eq. of hydrochloric acid (in MeOH) was slowly added dropwise and stirred at 50°C for 2 hours to obtain a solution. The solution was cooled to room temperature and precipitated. Subsequently, the solution was filtered, washed with methanol, and vacuum dried to obtain the title compound (yield amount 40 g, yield 85%) as a yellow powder. NMR analysis was performed to confirm the formation of the title compound. The NMR results are shown in Figure 1.

[0124] Experimental Example 1. Study on Atopic Dermatitis

[0125] The test for evaluating the efficacy of Example 1 in improving allergic dermatitis is described. In the test, atopic dermatitis was induced in Beagle dogs using house dust mite extract (Dermatophagoides farinae extract, DFE), and then Example 1 was administered at various doses to analyze the therapeutic effects (CADESI-04, TEWL, pruritus score, epidermal thickness, and mast cell count). During the test period, the results of CADESI-04, TEWL, pruritus score, epidermal thickness, and mast cell count were analyzed using GraphPad Prism software (Ver 10). The analysis was performed through comparisons between the group administered the test agent and the vehicle group, and between the group administered the test agent and the positive control group. (*: p<0.05; **: p<0.01; ***: p<0.001)

[0126] The test method and dosage are presented as follows (see Fig. 2):

[0127] • Target animals: Beagle (12 months old, male, n=43)

[0128] • Induction method: Sensitization with DFE at a concentration of 2 mg / mL twice a week for 24 weeks, followed by a 2-week rest period. Subsequently, induction with DFE at a concentration of 1 mg / mL three times a week for 2 weeks.

[0129] • Composition of administration group:

[0130] - Control group (G1): Administration of distilled water

[0131] - Control group (G2): 0.5% methylcellulose administered

[0132] - Positive control group (G3): Apoquel® 0.4-0.6 mg / kg administered (free base form reference dose)

[0133] - Test group (G4-G6): Administered Example 1 0.4 mg / kg, 0.6 mg / kg, 0.9 mg / kg (Reference dose in free base form)

[0134] • Method of administration :

[0135] - Administer 0.4, 0.6, and 0.9 mg / kg of the compound of Example 1 (reference dose in free base form) orally twice daily for 4 weeks

[0136] - Apoquel® is administered at a dose of 3.6 mg or 5.4 mg / animal based on body weight (reference dose of free base form).

[0137] - The normal group and the control group were administered the same volume of distilled water and 0.5% methylcellulose.

[0138] There were no significant changes in specific clinical symptoms or weight measurements in any of the test groups.

[0139] (1) CADESI-04 Score CADESI-04 is a clinical indicator that quantitatively scores and evaluates the severity and extent of involvement of typical lesions (erythema, lichenification, abrasions, etc.) of atopic dermatitis by site. Specifically, the following evaluations were performed to calculate the CADESI-04 score. The severity of pruritus was evaluated once a week during the sensitization and induction period, and twice a week during the administration period. The evaluations were performed according to the CADESI-04 criteria, and erythema, lichenification, abrasions, and alopecia were observed and scored as None (0), Mild (1), Moderate (2), and Severe (3). The average score of both axillary and groin areas was used as the result.

[0140] As shown in Figure 3, the CADESI-04 index was statistically significantly reduced in the Apoquel (0.4-0.6 mg / kg) and Example 1 (0.9 mg / kg) groups starting from day 10 (p<0.05), and a significant decrease compared to the vehicle group was observed in all test formulation groups starting from day 14 after administration.

[0141] The investigator's evaluation of the CADESI-04 lesion score for the dogs in the Example 1 group showed a statistically significant decrease starting from day 10 of administration compared to placebo-treated dogs (p<0.05). In addition, the 0.4 mg / kg and 0.6 mg / kg groups of Example 1 showed a decrease similar to that of the positive control group, Apoquel, while the 0.9 mg / kg group showed a greater decrease than the Apoquel group (p<0.05).

[0142] (2) Pruritic Score

[0143] The behavior of all animals was observed for 2 hours at 2 and 4 weeks after the administration of the test agent, and the frequency and duration of scratching and licking were evaluated as pruritus indicators. Frequency was determined by dividing the 2-hour recorded video into 1-minute intervals, assigning 1 point if symptoms were observed and 0 points otherwise, and summing the scores over a total of 120 minutes. Duration was determined by summing the seconds in which pruritus symptoms appeared during the 2-hour recorded period.

[0144] As shown in Figure 4, regarding the evaluation of the pruritic score, it was confirmed that at week 2 of administration, the Example 1 0.4 mg / kg administration group and the Example 1 0.6 mg / kg administration group reduced the score to a level similar to that of the Apoquel administration group, and a significant reduction was confirmed in the Example 1 0.9 mg / kg administration group compared to the control group and the Apoquel administration group (p<0.05).

[0145] (3) Transepidermal Water Loss (TEWL) Transepidermal Water Loss (TEWL) is a method of measuring the amount of water that evaporates through the skin and is an indirect indicator of dysfunction caused by a lack of the skin lipid layer. The higher the value, the more severe the dysfunction of the skin lipid layer.

[0146] TEWL was measured at 2 and 4 weeks after administration of the test agent, and was determined using a Tewameter (GP skin barrier pro-II, GP skin, Korea) at four induction sites. The average TEWL measurements from both axillaries and the groin were used as the results. As a result, as shown in Figure 5, a significant decrease in TEWL (Transepidermal Water Loss) compared to the control group was confirmed in all test groups at weeks 2 and 4 (p<0.05).

[0147] (4) Mast cells and transdermal thickness

[0148] Mast cell count and transdermal thickness are representative pathological markers of atopic dermatitis. Accordingly, to confirm the effect of allergy dermatitis improvement in a canine atopic dermatitis model, mast cell count and transdermal thickness were measured. Specifically, after anesthetizing with intravenous ketamine, skin samples were collected using a disposable biopsy punch (8 mm, KAI Industries Co., Ltd, Japan). After disinfecting the collection site, slides were prepared following standard tissue processing procedures, and H&E staining was performed. Epidermal thickness was measured using a microscope and an image analyzer (eXcope software, excope Inc.). Toludin blue staining was performed on the slides prepared through standard tissue processing procedures. A total of 3 to 5 regions were observed per slide, averaging a minimum of 2 to a maximum of 5, under a microscope at a magnification of 200x. The average value of the observed mast cell count was used as the result.

[0149] As a result of measuring epidermal thickness and mast cell count, as shown in Fig. 6, the 0.9 mg / kg administration group of Example 1 was reduced to a level similar to the normal group (p<0.05).

[0150] The administration of Example 1 showed an effect of improving allergic dermatitis in a canine atopic dermatitis model, and in particular, when Example 1 was administered at a dose of 0.9 mg / kg, it showed a therapeutic effect similar to or better than that of Apoquel.

[0151] Experimental Example 2. Study on Pruritus

[0152] This section describes a study to evaluate the antipruritic effect of Example 1 in an IL-31-induced pruritus model. The study involved Beagle dogs with pruritus induced by intravenous administration of IL-31, and the effects of orally administering Example 1 at various doses were analyzed. The test methods and dosage regimen are presented below (see Fig. 7):

[0153] • Target animals: Beagle (13–15 months old, male, n=32)

[0154] • Induction method: Pruritus was induced by intravenously administering 5.25 pg / kg of IL-31. Example 1 and the positive control drug (Apoquel) were administered, respectively, approximately 20 minutes prior to IL-31 administration. The test drug was prepared in liquid form and administered through a feeding tube, while the positive control drug was administered in tablet form along with a small amount of water.

[0155] • Composition of administration group:

[0156] - Control group (Gl): IL-31 not administered, distilled water administered

[0157] - Control group (G2): IL-31 administration, 0.5% methylcellulose administration

[0158] —Positive control group (G3): IL-31 administration, oral administration of 1 tablet of Apoquel® (0.4–0.6 mg / kg) (free base form reference dose)

[0159] - Test group (G4~G5): IL-31 administration, Example 1 0.6 mg / kg or 0.9 mg / kg oral administration (free base form reference dose)

[0160] • Method of administration :

[0161] - Orally administer Example 1 at a dose of 0.6 mg or 0.9 mg per kg of body weight (reference dose in free base form)

[0162] - Apoquel® is administered orally as one tablet (based on free base form dose).

[0163] IL-31 was administered 20 minutes after the administration of the test drug, and video surveillance was performed starting 20 minutes after IL-31 administration to observe and score the pruritic behavior of the experimental animals. A single observer evaluated pruritic behavior in real-time for a total of 360 minutes (6 hours). Whether pruritic behavior occurred was recorded as "Yes / No" at 1-minute intervals; behaviors such as licking or chewing the soles of the feet, flanks, and anal area; scratching the flanks or neck; scratching the floor; shaking the head; and dragging the buttocks on the floor were registered as "Yes." The number of "Yes / No" (1 or 0) judgments during each observation period was accumulated and provided as the pruritic score. The mean and standard deviation of the results were calculated using Microsoft® Excel® and expressed in the format Mean ± Standard Deviation (Mean±SD). The normality of the data in this study was assumed, and significant differences between the experimental groups were tested using Student's t-test. Statistical analysis was performed using Prism 9.4.1 (GraphPad Software Inc., San Diego, CA, USA), and a p-value less than 0.05 was considered statistically significant. (*: p<0.05; **: p<0.01; ***: p<0.001)

[0164] As a result of analyzing itching behavior over 360 minutes, the itching behavior score of the Vehicle control group (G2) was significantly higher than that of the Normal control group (G1) (p<0.001). The itching behavior scores of the Apoquel tablet group (G3), Example 1 0.6 mg / kg group (G4), and Example 1 0.9 mg / kg group (G5) were significantly lower than those of G2 (p<0.001). The itching behavior score of G5 was significantly lower than that of G3 (p<0.05) (see Fig. 8).

[0165] In the analysis of each 60-minute interval, the aptitude behavior scores of G2 were significantly higher than those of G1 in all intervals, and G3, G4, and G5 all showed significantly lower values ​​than G2 (p<0.001 to p<0.05) (see Fig. 9).

[0166] In particular, in the 61–120 minute interval of administration, the 0.9 mg / kg group of Example 1 (G5) showed a superior improvement effect compared to the Apoquel administration group (G3), and after 241 minutes, the Apoquel administration group (G3) showed a decrease in improvement effect compared to the control group (G2) to the extent that no significant difference was observed, but the 0.6 mg / kg group of Example 1 (G4) and the 0.9 mg / kg group (G5) showed a significant difference in improvement effect compared to the control group (G2) up to 360 minutes (see Fig. 9).

[0167] Regarding the duration of pruritus, the duration of pruritus increased in the control group (G2) compared to the normal group (G1) (p<0.001), and the duration of pruritus significantly decreased in the Example 1 0.9 mg / kg group (G5) compared to the Apoquel administration group (G3) (p<0.05) (see Fig. 8).

[0168] The administration of Example 1 showed an inhibitory effect on pruritus in an IL-31-induced pruritus model, and at doses of 0.6 mg / kg and 0.9 mg / kg, it showed a therapeutic effect similar to or better than Apoquel, and the duration of the effect also showed a superior sustained effect compared to Apoquel.

[0169] Experimental Example 3. Pharmacokinetic and Pharmacodynamic Studies

[0170] (1) Evaluation of the inhibitory effect on the wZroJAK-STAT pathway in wZroJAK-STAT pathway

[0171] Whole blood samples from naive canine dogs (Beagle dogs) were collected in K2EDTA blood collection tubes. Test compound 10 was added to a 5.0 ml microtube containing 180g of whole blood and incubated at 37°C for 45 minutes. Subsequently, human recombinant interleukin-6QL-6 10 was added to the tube at a final concentration of 50 ng / ml. The mixed sample was incubated at 37°C for 15 minutes, followed by the addition of 4 ml of Lyse / Fix buffer (BD Biosciences) and an additional incubation at 37°C for 20 minutes. The sample was then centrifuged at 800 x 10 for 5 minutes, and the supernatant was removed. The cell pellet was washed with 0.5% bovine serum albumin (BSA) / D-PBS buffer, then resuspended in 1 ml of Perm Buffer III (BD Biosciences) and incubated on ice for 30 minutes. After two washes, 250 ml of an antibody mixture containing 2.5 ml of CD4 FITC antibody (Cat #: 11-5040-42, Invitrogen, Waltham, MA, USA, RRID: AB_10596977) and 5 ml of human phosphorylated (phospho)-STAT3 PE antibody (Cat #: 651004, BioLegend, San Diego, CA, US, RRID: AB_2571892) was mixed with the cells for IL-6 induction samples and incubated overnight. After washing, analysis was performed using a MACSQuant analyzer. It was previously confirmed that human recombinant IL-6 and human phosphorylated-STAT3 PE antibodies exhibit cross-reactivity with canine blood cells. Quantification of unstimulated (NS) samples involved gating lymphocytes (or monocytes) in an FSC-SSC plot and setting the CD4+pSTAT3+ region to be between 0.5% and 0.9%. This reflected the baseline level of pSTAT3 in a normal inactive state that did not induce activation.Next, the CD4+pSTAT3+ region of the stimulus (S) sample was quantified.

[0172] To evaluate the inhibitory effect of the test compound on IL-31-induced STAT3 activation, DH82 mononuclear cell lines (Cat. # CRL-1039, ATCC, Manassas, VA, USA, RRID: CVCL_2018) were used. Cells were placed in EMEM medium (Cat. # 30—2003, ATCC) containing 15% heat-inactivated fetal calf serum and 10 ng / ml recombinant interferon-gamma (Cat. # 781-CG, R&D Systems) at a rate of 1 x 10⁶ cells per well. 5 They were seeded into 96-well plates at a density of [number] and cultured for 24 hours at 37 °C under 5% CO2 conditions. The next day, 10–3.2 U To establish six concentration gradients in the mol / L range, the test compound or sibling control was treated for 1 hour. Subsequently, the cells were treated with lyg / ml of recombinant IL-31 (Cat # CYT-1016, ProSpec, Rehovot, Israel) for 5 minutes. The medium was removed, and cytokine treatment was terminated by adding cell lysis buffer (Cat # 9803, Cell Signaling Technology, Danvers, MA, USA). STAT3 activation was detected using the PathScan Phospho-STAT3 (Tyr 705) Sandwich ELISA Kit (Cat. # 7300C, Cell Signaling Technology).

[0173] The inhibitory effects of the test compounds on IL-6-induced STAT3 activation or GM-CSF-induced STAT5 activation were evaluated using the TF-1 human erythroblast cell line (Cat. # CRL-1039, ATCC, Manassas, Virginia, USA, RRID: CVCL_0559). Cells were suspended in RPMI1640 culture medium (Cat. # 22400-089, GIBCO) containing 10% heat-inactivated FBS (fetal bovine serum) and plated at 1 x 10⁶ cells per well in 96-well plates. 5 Dispensed at a density of [number] and incubated for 24 hours at 5% CO2, 37°C.

[0174] Next, a titration curve with six points was constructed by treating with the test compound or the control for 1 hour. Cells were treated with 15 ng / mL human IL-6 (Cat. # 206-IL-050, R&D systems) for 5 minutes or with 10 ng / mL human GM-CSF (Cat. # 300-03-20 UG, PEPROTECH, Cranberry, NJ, USA) for 3 hours. The culture medium was removed, and cytokine treatment was terminated by adding cell lysis buffer (Cat # 9803, Cell Signaling Technology, Danvers, Massachusetts, USA). Activation of STAT3 or STAT5 was detected using the PathScan® Phospho- STAT3 (Tyr 705) Sandwich ELISA Kit (Cat. # 7300C, Cell Signaling Technology) or the STAT5 A / B pY694 / 699 ELISA Kit (Cat. # AB176656, ABcam), respectively.

[0175] IC5o values ​​were calculated by constructing dose-response plots based on activity by concentration using a 4-parameter logistic nonlinear regression model.

[0176] [Table 1]

[0177]

[0178]

[0179] Results of in vitro canine whole blood, DH82 analysis, and human TF-1 cell analysis regarding the IC of Example 1 for the JAK1-STAT3 pathway induced by cytokines IL-6 and IL-31 50 The values ​​were found to be 9.4 nM in canine whole blood and 8.4 nM in DH82 cells, whereas oklashinib showed 61.3 nM and 49.7 nM, respectively. Additionally, the IC5o values ​​for IL-6-induced JAK1-STAT3 pathway inhibition in human TF-1 cells were confirmed to be 15.6 nM for Example 1 and 56 nM for oklashinib. For the JAK2-STAT5 pathway activated in human TF-1 cells by the hematopoietic-associated cytokine GM-CSF, the IC5o values ​​for Example 1 and oklashinib were measured to be 749 nM and 1,214 nM, respectively (see Table 1). These results demonstrate that Example 1 inhibits the JAK1-STAT3 pathway more potently and selectively compared to oklashinib. The prominent JAK1 selectivity of Example 1 suggests that it can be an important characteristic in the development of treatments for JAK1-related diseases such as rheumatoid arthritis and atopic dermatitis.

[0180] (2) Evaluation of in vivo pharmacokinetics and pharmacodynamics

[0181] Ten-month-old male Beagles were purchased from Orient Bio (Seongnam, Gyeonggi-do, South Korea), and animal experiments were conducted according to a protocol approved by the Animal Ethics Committee of Seoul National University Hospital (Approval No. 22-0220-S1A1). Animals were housed indoors at 20–26°C and 30–70% humidity, and were orally administered experimental dog food (Purina, St. Louis, Missouri, USA) twice daily. On the day of pharmacokinetic and pharmacodynamic sampling, feeding was withheld for 2 hours after drug administration.

[0182] Apoquel® (Oclacitinib maleate) was used as a positive control in Group 1. Apoquel® was administered orally at a concentration of 5.4 mg according to the label recommended dose (Oclacitinib free base 0.4-0.6 mg / kg). Example 1 was administered gastrically at a free base mg / kg using a 22 French gauge rubber gavage tube. The experimental group for Example 1 was set up as shown in Table 2 below. [Table 2]

[0183]

[0184] All groups were administered the drug twice daily (bis in die, BID) for 7 days at 8:30 AM and 8:30 PM. Twelve animals were used per experiment, and the experiment was repeated a total of three times. In each experiment, animals were randomly assigned and rotated between groups to ensure that the same animals were not reassigned to the same group in subsequent experiments.

[0185] To evaluate changes in animal body weight (BW), body weight was measured 24 hours prior to the first administration (Day 0) and 24 hours after the final administration (Day 8). Whole blood was collected from the cephalic vein 24 hours prior to the first administration (Day 0) and 24 hours after the final administration (Day 8) for Complete Blood Count (CBC) and blood chemistry analysis. Pharmacokinetics and pharmacodynamics were evaluated on Day 1 and Day 7. Both compounds were administered twice daily (BID) for 7 days. On Day 7, blood samples were collected from dogs treated with each compound at 0, 0.25, 0.5, 1, 2, 3, 5, 7, and 12 hours after drug administration to analyze blood exposure and the inhibitory effect of IL-6 on STAT3 phosphorylation (see Fig. 10).

[0186] 1) Toxicity assessment

[0187] JAK inhibitors can cause neutropenia, anemia, thrombocytopenia, and body weight gain in humans, and ochlasitinib is known to be associated with side effects such as vomiting, anorexia, leukopenia, decreased globulin, and elevated cholesterol levels. Accordingly, to evaluate the toxicity of Example 1, the animals' total blood counts, blood chemistry, and changes in body weight were analyzed throughout the entire experimental period, from 24 hours before the first administration to 24 hours after the last administration. Administering Example 1 twice daily for 7 days did not affect body weight changes, and it was confirmed that there were no significant clinical effects hematologically or serologically. Therefore, these results suggest that daily oral administration of Example 1 to dogs at a dose of 0.9 mg / kg for 7 days is toxicologically safe.

[0188] 2) Pharmacokinetic evaluation

[0189] Example 1 To analyze pharmacokinetics, whole blood was collected from animals administered the corresponding drug at each time point on day 1 and day 7 of administration. Subsequently, the plasma concentration of each molecule was measured via LC-MS / MS.

[0190] According to the results on day 1, the plasma of the example Cmax and AUC0-12h increased significantly more than the dose-dependent increase. Despite differences in drug formulations, the Tmax values were consistently similar across all Example 1 administration groups and the oclacitinib administration group. Tmax values were observed to be between 0.5 to 1.5 hours in all groups. In addition, T1 / 2 was similar across all Example 1 administration groups (3.6 to 6.0 hours) and the oclacitinib administration group (3.6 hours).

[0191] The results on day 7 were similar to those on day 1. The plasma Cmax and AUC0-12h of Example 1 increased significantly more than the dose-dependent increase. Compared with day 1, the c of oclacitinib on day 7 max and AUC0-12h increased by 1.4-fold and 1.6-fold, respectively, while Example 1 showed small changes in Cmax and AUC0-12h, which were 0.96 to 1.07-fold and 1.02 to 1.14-fold, respectively, in the effective dose range of 0.45 to 0.9 mg / kg. The Tmax value on day 7 was 0.8 to 1.4 hours, which remained consistent with that on day 1. Similarly, the T1 / 2 of the Example 1 administration group was the same as that on day 1. However, the T1 / 2 of the oclacitinib administration group on day 7 was significantly prolonged from 3.6 ± 0.9 hours on day 1 to 4.4 ± 0.3 hours (see Table 3 and Figure 11).

[0192] [Table 3]

[0193] . 福 흐 t .

[0194]

[0195] 2 people 1 o'clock, 1 ye 1 IG 0.2 1 11.9 75.9 1.0 ± 0.5 6.0 ±2.1 2.12 17.4 4.58 1.84 19.0 0.58 5.96 0.29 (N = 6) 7 16.7 ± 112 ± 1.4 ± 0.9 4.5 ± 1.0 1.7 ± 0.9 10.6 ± 4.57 ±

[0196] 5.82 36.1 4.75 0.36 3 J 27.2

[0197] 1 IG 0.3 1 Sat 162 Sat 10.8 Sat 4.34 Sat 7.30 47.9 0.9 ± 0.6 4.4 ±0.5 1.7 ± 0.6 4.38 0.31 27.6 Sat 153 Sat 2.04 Sat 10.6 Sat 4.07 (N = 6) 7 0.8 ± 0.3 3.6 ±0.5 Sat 11.9 59.9 0.95 4.75 0.25 , , 48.7 Sat 250 Sat 1.61 Sat 9.72 Sat 4.06 Sat 4 실시예 1 IG 0.45 1 1.3 ± 1.2 4.1 ±0.5

[0198] 4.21 50.4 0.35 2.85 0.25 46.9 Sat 289 Sat 1.46 Sat 7.71 Sat 4.21 Sat (N= 5) 7 1.3 ± 0.7 3.8 ±0.6

[0199] 13.5 66.6 0.51 2.02 0.09 Person 1, 1 92.1 Sat 473 Sat 1.21 Sat 6.37 Sat 4.07 Sat 5 실시예 1 IG 0.6 1 0.9 ± 1.0 3.6 ±0.6

[0200] 22.4 126 0.36 2.40 0.36 99.0 Sat 539 Sat 1.13 Sat 5.64 Sat 3.86 Sat (N = 6) 7 1.3 ± 0.9 3.5 ±0.8

[0201] 38.5 185 0.45 2.35 0.37 167 Saturday 6 Silsiye 1 0.9 1 29.0 Dinghai River (N= 5) 7

[0202]

[0203] 85.9

[0204] 3) Pharmacodynamic evaluation

[0205] To analyze the pharmacodynamics of Example 1 and oklasitinib, whole blood was collected at each time point on day 1 and day 7 after the administration of the two drugs. The inhibitory effect on IL-6-induced STAT3 phosphorylation (pSTAT3) was evaluated by flow cytometry.

[0206] On day 1, in Group 1 (oklasitinib), IL-6-induced pSTAT3 increase showed maximum inhibition at 1 hour after administration, decreasing by -31.42 (15.84%) compared to 0 hours. In Group 1 of Example, pSTAT3 increase showed maximum inhibition at 1 hour or 3 hours after administration. The maximum inhibition effects for each group were -26.64 (27.06%) for the 0.3 mg / kg group, -31.15 (30.67%) for the 0.45 mg / kg group, -46.68 (20.43%) for the 0.6 mg / kg group, and -70.46 (23.58%) for the 0.9 mg / kg group. Exceptionally, in the 0.2 mg / kg group, the maximum pSTAT3 inhibition was -17.72 7.39% at the 12-hour mark. In addition, as the dose of Example 1 increased, the duration of the inhibitory effect also tended to increase, and the effect of Example 1 was observed to be maintained for 7 to 12 hours after administration (see Fig. 12).

[0207] The data for Day 7 also showed a similar pattern to Day 1. In the oklasitib group, IL-6-induced pSTAT3 increase showed maximum inhibition at 3 hours after administration, decreasing by -44.53 (7.42%) compared to 0 hours. In the administration group of Example 1, inhibitory effects were observed as follows: -31.14 (23.65%) for the 0.3 mg / kg group, -43.93 (24.70%) for the 0.45 mg / kg group, -56.09 (15.84%) for the 0.6 mg / kg group, and -83.39 ± 7.17% for the 0.9 mg / kg group. In the 0.2 mg / kg group, maximum pSTAT3 inhibition was -18.44 (25.12%) at 7 hours (see Fig. 12).

[0208] These results suggest that orally administered Example 1 effectively inhibits the JAK-STAT pathway and exhibits significant pharmacodynamic effects at doses similar to or higher than those of oklasitinib in the approved dose range.

[0209] 4) JAK Selectivity Evaluation

[0210] To evaluate the JAK selectivity (concern regarding JAK2 inhibition) of oklasitinib and Example 1, the in vivo pharmacokinetic data and in vitro pharmacodynamic data (TF-1 cells) shown in Fig. 11 and Table 1, respectively, were compared. As a result of the analysis, the Cmax of oklasitinib approached the IC50 for the JAK2 pathway (GM-CSF-induced pSTAT5 in TF-1 cells) on day 7, whereas the plasma concentration-time profile of Example 1 remained below the IC50 value for the JAK2 pathway even at 0.9 mg / kg (see Fig. 13).

[0211] In addition, JAK selectivity was evaluated after administering Example 1 as a single dose at 0.4 to 0.6 mg / kg or 0.6 to 0.9 mg / kg in the same manner as above. As a result of the analysis, it was confirmed that the plasma concentration-time profile of Example 1 remained above the IC50 value for the JAK1 pathway for most of the 24 hours after administration and below the IC50 value for the JAK2 pathway. This suggests that Example 1 is sufficiently effective even when administered orally once a day (see Fig. 14).

[0212] The in vivo pharmacokinetic data and in vitro pharmacodynamic data (IL-6-induced pSTAT3 in whole blood cells of Beagle dogs) shown in Figure 11 and Table 1, respectively, were further compared. Although there were some significant differences in the duration of JAK1 inhibition above IC50 between Example 1 and oklashitinib on days 1 and 7, these were clinically irrelevant. However, regarding the duration of IC90 above, it was confirmed that Example 1 (0.9 mg / kg) was significantly increased compared to oklashitinib (0.4-0.6 mg / kg). This suggests that Example 1 (0.9 mg / kg) can maintain the JAK1 inhibitory effect for a longer period. In addition, for oklashitinib, the duration of IC90 above was significantly increased on day 7 (5.9 mg / kg, 1.0 hour) compared to day 1 (2.8 mg / kg, 0.6 hours). This is interpreted as the duration of JAK1 inhibition of oklasitinib being prolonged after repeated administration. On the other hand, in the case of Example 1 (0.9 mg / kg), IC between Day 1 and Day 7 90 No significant change in duration of excess was observed, suggesting that a sufficiently high drug concentration is already maintained at the corresponding dose (see Table 4).

[0213] [Table 4]

[0214]

[0215] Plasma concentration for IL-6-induced JAK-STAT inhibition is IC 50 Alternatively, the period exceeding ICw was calculated as the duration over 12 hours after administration. Values ​​are expressed as the mean and standard deviation. *, **, ****: Student's t-test analysis results indicate significant differences at the p < 0.05, p < 0.01, and p < 0.0001 levels, respectively, compared to the oclacitinib administration group. #, ####: Student's t-test analysis results indicate significant differences at the p < 0.05 and p < 0.0001 levels, respectively, compared to Day 7.

[0216] Therefore, through the above Experimental Example 3, it was confirmed that Example 1 is a more effective pruritus treatment with minimized side effects, having higher JAK1 selectivity and lower JAK2 inhibitory effect than the positive control oklasitinib, and less accumulation with repeated administration.

[0217] Experimental Example 4. Exploratory clinical trial on the efficacy of Experiment 1 in improving pruritus in canine allergic dermatitis

[0218] 1. Purpose of Clinical Trial

[0219] This exploratory clinical trial was conducted to evaluate the safety and efficacy of the administration method and dosage of Example 1 in dogs with allergic dermatitis accompanied by pruritus.

[0220] 2. Clinical Trial Plan

[0221] 2.1 Test Design and Planning

[0222] This clinical trial was conducted using an open-label, randomized, active-drug controlled, multicenter, parallel design. If the animals were deemed to meet the inclusion / exclusion criteria, they were assigned identification numbers in the order they were registered in the eCRF system within each animal hospital and assigned to treatment groups according to the randomization table (see Fig. 15).

[0223] 2.1.1 Visit 1 (Day 0)

[0224] 1) Obtaining consent: Consent was obtained from the guardian of the animal diagnosed with allergic dermatitis.

[0225] 2) Check Selection / Exclusion Criteria

[0226] 3) Assignment of Treatment Groups

[0227] 4) Assignment of identification numbers: Identification numbers were assigned to each animal hospital in the order they were registered in the eCRF system.

[0228] 5) Basic characteristics survey: Age, weight, sex, neutering status, breed (size, information), and concomitant medications were investigated.

[0229] 6) Physiological indicator tests: body weight, heart rate, respiratory rate, body temperature

[0230] 7) Central laboratory tests: Hematological tests, blood biochemical tests

[0231] 8) Measurement of efficacy evaluation variables by the attending veterinarian: Canine Atopic Dermatitis Extent and Severity Index—04 (Canine Atopic Dermatitis Extent and Severity Index-04, hereinafter, CADESI-04)

[0232] 9) Guidelines for Target Animal Owners and Training on Pruritus Visual Analog Scale (PVAS) Assessment

[0233] For the guardian PVAS evaluation, guardians were instructed to record an evaluation of symptoms, such as the number of times the target animal scratches during the day, on a guardian evaluation sheet.

[0234] 10) Measurement of Guardian Validity Evaluation Variables: PVAS

[0235] 11) Photographing the lesion site

[0236] 12) Prescription of test drug or control drug

[0237] 2.1.2 Visit 2 (Day 14 - 3)

[0238] 1) Physiological indicator tests: body weight, heart rate, respiratory rate, body temperature

[0239] 2) Measurement of efficacy evaluation variables by the attending veterinarian: CADESI-04

[0240] 3) Measurement of Guardian Validity Evaluation Variables: PVAS

[0241] 4) Investigation of concomitant drugs

[0242] 5) Photographing the lesion site

[0243] 6) Collection of guardian PVAS assessments and verification of medication adherence

[0244] 7) Evaluation of Adverse Reactions

[0245] The recurrence of symptoms, serious adverse events (SAE), severity, causal relationship with the investigational animal drug, related measures, and corrective treatment were recorded.

[0246] 2.1.3 Visit 3 (Day 28 3)

[0247] 1) Physiological indicator tests: body weight, heart rate, respiratory rate, body temperature

[0248] 2) Central laboratory tests: Hematological tests, blood biochemical tests

[0249] 3) Measurement of validity evaluation variables by the attending veterinarian: CADESI-04

[0250] 4) Measurement of Guardian Validity Evaluation Variables: PVAS

[0251] 5) Investigation of concomitant drugs

[0252] 6) Photographing the lesion site

[0253] 7) Recall of investigational animal drugs and verification of medication adherence

[0254] 8) Evaluation of Adverse Reactions 2.2 Consideration of Test Design

[0255] The efficacy evaluation variables of PVAS and CADESI-04 established in this clinical trial are key efficacy endpoints internationally agreed upon in clinical studies of canine atopic dermatitis. PVAS can sensitively detect changes in subjective itching reported by owners, while CADESI-04 provides a standardized, objective lesion assessment performed by veterinarians. The combination of these indicators was designed to demonstrate clinical utility in real-world clinical settings by integrating owner reports, veterinarians' clinical assessment of skin lesions, and the overall treatment evaluation.

[0256] The efficacy evaluation period was set to 28 days, which is considered an internationally standardized initial evaluation period. This was also set as the primary efficacy evaluation period in the clinical trials for the product approval of Apoquel® tablets and Cytopoint® injections. Although it may be difficult to observe changes in some lesions (such as lichenification) included in CADESI-04 in trials of less than 6 weeks, this was determined to be a sufficient evaluation period to assess the onset of drug efficacy and initial safety, as well as the optimal period to ensure medication adherence.

[0257] Although the clinical trial protocol did not classify efficacy evaluation variables into primary and secondary, PVAS was described as the primary efficacy evaluation variable and CADESI-04 as the secondary efficacy evaluation variable to analyze the efficacy of improving pruritus, similar to the evaluation of general canine atopic dermatitis.

[0258] 2.3 Selection of Homologous Materials for Clinical Trials

[0259] 2.3.1 Selection Criteria

[0260] 1) Target animals aged 12 months or older and weighing 3 kg or more as of Visit 1 (Day 0)

[0261] 2) Subject animals diagnosed with allergic dermatitis via the following methods based on Visit 1 (Day 0)

[0262] — Based on medical history, clinical symptoms of atopic dermatitis as presented by ICADA (International Committee for Allergic Diseases of Animals), Favrot's criteria, or allergy tests (serological allergen-specific IgE test, intradermal inoculation test), subjects exhibiting a chronic history of non-seasonal pruritus were diagnosed with allergic dermatitis.

[0263] 3) Subject animals exhibiting moderate to severe pruritus (PVAS score 6) as of Visit 1 (Day 0)

[0264] 4) Target animals of guardians who have given written consent on the companion animal clinical research consent form

[0265] 2.3.2 Exclusion Criteria

[0266] 1) Subject animals with diseases that can induce immunosuppression, such as adrenal cortical hyperfunction, hypothyroidism, or progressive malignant tumors

[0267] 2) Pregnant, nursing, or breeding dogs

[0268] 3) Target animals under 12 months of age or weighing less than 3 kg

[0269] 4) Target animals being administered or treated with the following medications

[0270] - Oral steroids

[0271] - Topical and oral calcineurin inhibitors

[0272] - Lokivetmab (Cytopoint® Injection)

[0273] - Topical and oral antihistamines

[0274] Other drugs with anti-inflammatory or itching-relieving effects

[0275] However, participation was permitted after a wash-out period of 1 week for antihistamines, 6 weeks for lokivetmab (Cytopoint® injection), 8 weeks for long-acting steroid injections, and 2 weeks for oclacitinib (Apoquel® tablets) and other preparations including topical and oral steroids.

[0276] 5) Subject animals with skin conditions similar to allergic dermatitis, such as external parasitic infections causing itching, bacterial or fungal skin infections, or Malassezia dermatitis

[0277] 6) In addition to the above, subject animals with clinically significant findings that the attending veterinarian medically judges to be inappropriate for this study

[0278] 2.3.3 Criteria for Suspension and Dropout

[0279] Clinical trial animals were discontinued or dropped from the clinical trial according to the following criteria.

[0280] 1) Subject animals that did not complete treatment or received the wrong dosage during the study

[0281] 2) Cases where tracking is impossible due to failure to visit at the scheduled appointment 3) Cases where medication is discontinued due to worsening clinical symptoms or the occurrence of adverse reactions 4) Cases where clinical trial animals that do not meet the selection / exclusion criteria have participated in a clinical trial

[0282] 5) In the event of a serious violation of the clinical trial protocol

[0283] - In the event of discontinuation or withdrawal from a clinical trial, the attending veterinarian discontinued the administration of the investigational animal drug and recorded the date of trial termination and withdrawal, the reason for discontinuation and withdrawal, etc., along with all data obtained up to the point of discontinuation and withdrawal, in the supporting documents and eCRF. The results of the clinical trial obtained up to the point of discontinuation and withdrawal were made available for review regarding items eligible for evaluation during the final evaluation.

[0284] 2.3.4 Number of Animals for Analysis and Composition of Dosage Groups

[0285] - Approximately 40 subjects who visited the veterinary clinic with itching as the main symptom and were diagnosed with allergic dermatitis

[0286] - As shown in Table 5 below, the groups were organized into three, and at least 13 individuals were recruited for each administration group.

[0287] [Table 5]

[0288]

[0289] * BID to QD: Administer twice daily for up to 14 days, and once daily thereafter until 28 days.

[0290] 2.4 Veterinary Medicines for Clinical Trials

[0291] 2.4.1 Dosage and Administration of Veterinary Drugs for Clinical Trials, Manufacturing No. 1) Test Drug: Example 1 Tablet (Refer to Table 6)

[0292] [Table 6]

[0293]

[0294] 2) Control drug: Apoquel® tablets (see Table 7)

[0295] [Table 7]

[0296]

[0297] 3) Usage • Dosage

[0298] [G1]

[0299] • Initial dosage: Oklasitinib was administered orally twice daily at a dose of 0.4 - 0.6 mg per kg of body weight (based on free base form) for 14 days from the first day of administration.

[0300] • Maintenance dose: Oklasiti nib was administered orally once daily at the same dose from 14 days after the start of the maintenance period until 28 days. The number of tablets administered according to the body weight of the dogs is shown in Table 8 below.

[0301] [Table 8]

[0302]

[0303]

[0304] [G2]

[0305] • Initial dosage: Example 1 was orally administered twice daily at a dose of 0.6 to 0.9 mg per kg of body weight (based on the free base form) from the first day of administration until 14 days.

[0306] • Maintenance dose: Example 1 was administered orally once a day at the same dose from 14 days after the date of death until 28 days. The number of tablets administered according to the body weight of the dogs is as shown in Table 9 below.

[0307] [Table 9]

[0308]

[0309] [G3]

[0310] • Example 1 was orally administered once a day at a dose of 0.9 to 1.35 mg per kg of body weight (standard dose in free base form) from the first day of administration until 28 days. The number of tablets administered according to the body weight of the dogs is as shown in Table 10 below.

[0311] [Table 10]

[0312]

[0313]

[0314] 2.4.2 Assignment of Dosage Groups

[0315] 1) The attending veterinarian explained the participation in the clinical trial to the guardian and obtained written consent. The selection / exclusion criteria for the target animals were verified, and if suitable, the animals were enrolled in the clinical trial.

[0316] 2) To enhance inter-group comparability and ensure balanced randomization, this clinical trial assigned randomization numbers based on subgroups of the PVAS score, the primary efficacy endpoint. Randomization was performed using a stratified randomization method, allocating G1, G2, and G3 in a 1:1:1 ratio. The detailed randomization method was as follows.

[0317] Based on PVAS scores before drug administration, subgroups were divided into moderate (6 to less than 8 points) and severe (8 points or more). Randomization numbers (MXX or SXX) starting from 01 were assigned to each subgroup in the order of registration, and administration groups were assigned according to the randomization table.

[0318] - The randomization table was designed and created in advance prior to the clinical trial, applying G1, G2, and G3 of the trial sequentially starting from randomization number 1.

[0319] 3) Despite randomization, a bias was identified in the CADESI-04 score, a secondary efficacy endpoint, due to a group skew towards a specific institution caused by the small sample size, and an imbalance occurred in the number of animals analyzed due to the dropout of one institution. Consequently, it was determined that a comparison between G1, G2, and G3 was impossible, so the clinical trial protocol was revised to apply an adjusted allocation method that minimizes the imbalance between groups.

[0320] 4) The group assignment results of all registered subjects and the animals subject to final analysis were recorded and stored in accordance with the relevant SOP forms. 5) Even if the guardian of a subject participating in the study discontinued the clinical trial or dropped out midway, the number assigned to that subject animal was not reused.

[0321] 2.4.3 Basis for Capacity Setting

[0322] The dosage and administration method defined in Example 1 of this clinical trial were established based on the results of efficacy, pharmacokinetic, and pharmacodynamic studies using Beagle dogs equivalent to a Phase 2 clinical trial.

[0323] For G2, the dosage regimen was set to the 0.9 mg / kg dose of Example 1, which was confirmed to have superior efficacy compared to the group administered a competing drug (Apoquel® tablets) based on the evaluation results of a house dust mite (HDM)-induced allergic dermatitis model in Beagle dogs, and the same administration method as the competing drug (administered twice a day for the first 2 weeks, and once a day for the following 2 weeks).

[0324] In the case of G3, pharmacokinetic (PK) and pharmacodynamic (PD) tests performed in Beagle dogs showed that the maximum dose at which the maximum blood drug concentration was lower than the IC5o of JAK2 upon single administration of Example 1 was 1.35 mg / kg, and the blood drug concentration was maintained above the IC5o of JAK1 throughout the 24 hours, and no drug accumulation was observed even upon repeated administration, so the dosage regimen was set to 1.35 mg / kg and once daily administration.

[0325] As a result of the safety evaluation, in a 13-week repeated-dose toxicity study conducted in Beagles, the drug exposure at the no observed adverse effect level (NOAEL) of 3 mg / kg was Cmax 6-fold and AUC 5-fold compared to the G2 maximum dose, and C compared to the G3 maximum dosemax With 4 times and AUC 3 times, it is determined that both capacities have secured a sufficient safety margin.

[0326] 2.4.4 Evaluation of Medication Adherence

[0327] It was recommended that the veterinary drugs for the clinical trial be administered from Visit 1 to Visit 2, and from Visit 2 to Visit 3, followed by a visit at each respective time. Medication adherence was assessed by checking whether the medication was taken during the Visit 1-2 (Day 0-13) period during Visit 2, and by checking whether it was taken during the Visit 2-3 (Day 14-28) period during Visit 3.

[0328] The results of the two intervals were summed to calculate the final medication adherence, and cases where the overall medication adherence was less than 90% were excluded from the efficacy evaluation analysis. Medication adherence was evaluated using the following mathematical formula.

[0329] [Mathematical Formula 1] Medication Adherence (%) = (Actual number of doses / Number of doses required during clinical trial period) x 100

[0330] 2.5 Methods for Evaluating Efficacy and Safety

[0331] 2.5.1 Efficacy and Safety Evaluation Schedule

[0332] The schedule for the clinical trial visits and efficacy and safety evaluations was carried out according to Table 11 below.

[0333] [Table 11]

[0334]

[0335] - The first day of administration of the test drug or control drug was defined as the reference day (Day 0), and based on the previous visit date, visits were allowed within 3 days before and after the scheduled visit date.

[0336] - Central laboratory testing was conducted at the same institution, and the following items were tested.

[0337] • Hematological tests: RBC, Hb, HCT, MCV, MCH, MCHC, RDW-SD, RDW-CV, reticulocyte, RET%, NRBC, IRF, LFR, MFR, HFR, RET-He, platelet count, PDW, MPV, P-LCR, Pct, WBC, neutrophil, lymphocyte, monocyte, eosinophil, basophil

[0338] • Blood Biochemical Tests: Total protein, globulin, albumin, A / G ratio, ALT (GPT), AST (GOT), ALP, GGT, total bilirubin, total cholesterol, triglycerides, glucose, amylase, lipase, BUN, creatinine, B / C ratio, CK, Ca, P, Na, K, Cl, Na / K ratio, corrected Cl

[0339] - At every visit, at least one representative skin lesion on the animal was photographed and recorded. The photos were taken to ensure the skin lesion was clearly visible, and were taken in environments and lighting as similar as possible to allow for comparison of changes in the skin lesion between visits.

[0340] - If the subject animal exhibits a "serious adverse reaction," such as requiring emergency treatment or a side effect that is difficult to reverse, it is to be reported to the attending veterinarian and the principal investigator, regardless of whether it is related to the investigational animal drug.

[0341] - If this plan was not complied with, the test manager and the test administrator recorded the deviations and violations in the deviation record sheet and kept it in the TMF (Trial Master File).

[0342] 2.5.2 Primary Validity Evaluation Variables

[0343] 1) PVAS score: PVAS scores measured at days 0, 1, 2, 3, 4, 5, 6, 7, 14, and 28 of administration

[0344] 2) Change in PVAS score: Change in PVAS score at days 1, 2, 3, 4, 5, 6, 7, 14, and 28 compared to before administration (Day 0).

[0345] The PVAS evaluation was designed to proceed during Visit 1 after the veterinarian confirmed the owner's evaluation bias and explained the scoring method to assist in making an accurate judgment. The scoring method was based on behavioral characteristics and severity, using a 10 cm ruler to rate the complete absence of itching as "0 points" and the presence of extreme itching as "10 points."

[0346] 2.5.3 Secondary Efficacy Evaluation Variables

[0347] 1) CADESI-04 Score: CADESI-04 scores measured at days 0, 14, and 28 of administration

[0348] 2) Change in CADESI-04 score: Change in CADESI-04 score at days 14 and 28 compared to before administration (Day 0 administration)

[0349] 2.5.4 Safety Evaluation Method

[0350] The safety evaluation was analyzed for subjects that had completed all central laboratory tests among subjects that had taken the investigational animal drug at least once.

[0351] 1) Physiological indicator tests (body weight, heart rate, respiratory rate, body temperature) were measured using scales, stethoscopes, thermometers, etc., in the animal hospital during Visit 1, Visit 2, and Visit 3.

[0352] 2) Central laboratory tests (hematological tests, blood biochemical tests) were performed by collecting blood samples at Visit 1 and Visit 3 and transferring them to an external analysis laboratory (KVL). For each item, the results were evaluated by comparing the pre- and post-administration of the investigational animal drug, and any clinically significant changes were recorded as adverse reactions.

[0353] 3) All adverse reactions occurring during the clinical trial period were recorded in the eCRF by the attending veterinarian, and the severity of adverse reactions was classified according to the criteria in Table 12 below based on maximal intensity.

[0354] [Table 12]

[0355]

[0356] 4) All adverse reactions were recorded in the adverse reaction section of the eCRF, even if there was no causal relationship with the drug, and were classified according to the criteria in Table 13 below.

[0357] [Table 13]

[0358]

[0359]

[0360] 2.5.5 Statistical Analysis

[0361] 1) The experimental / observation unit of this clinical trial was the individual.

[0362] 2) The statistical analysis program used was SPSS version 20 (SPSS, Inc., Chicago, IL, USA).

[0363] 3) All statistical significance tests were performed as two-sided tests at a significance level (a) of 5%. For the efficacy and safety data obtained from the animals in this clinical trial, descriptive statistics (mean, standard deviation, etc.) were presented for continuous data for each group, while frequencies and percentages were presented for categorical data. Missing values ​​were not imputed.

[0364] 4) Statistical analysis of efficacy evaluation variables

[0365] - PVAS scores: A two-way repeated measures ANOVA (hereinafter, RM ANOVA) was performed to simultaneously analyze the main effects of time and group factors and their interaction effects on the PVAS scores obtained at each measurement time point. A one-way ANOVA was conducted to analyze the differences between groups at each time point.

[0366] - Change in PVAS score: A paired t-test was performed to compare the difference in PVAS scores for each group between before administration (Day 0 of administration) and at each measurement time point (Days 1, 2, 3, 4, 5, 6, 7, 14, and 28 of administration). The presence and frequency of adverse effects, such as specific clinical symptoms and hematological abnormalities, were evaluated after administration.

[0367] - CADESI-04 scores at each time point: To simultaneously analyze the main effects of time and group factors and their interaction effects on the CADESI-04 scores obtained at each measurement time point, RM ANOVA was performed. To analyze the differences between groups at each time point, one-way ANOVA was conducted.

[0368] - Change in CADESI-04 score: A paired t-test was performed to compare the difference in CADESI-04 scores between each group before administration (day 0 of administration) and at each measurement time point (days 14 and 28 of administration).

[0369] 5) Statistical analysis of safety evaluation variables

[0370] - Physiological indicator tests: Body weight, heart rate, respiratory rate, and body temperature obtained at each measurement time point were analyzed using RM-ANOVA to simultaneously analyze the main effects of time and group factors and their interaction effects. - Central laboratory tests (hematological tests, blood biochemical tests): Each blood test indicator obtained on day 0 and day 28 of administration was analyzed using RM-ANOVA to simultaneously analyze the main effects of time and group factors and their interaction effects.

[0371] - Adverse Reactions: The severity of adverse reactions, association with the investigational animal drug, and the number of reported cases and, if necessary, the percentages were presented by group.

[0372] 6) Statistical analysis of basic characteristics

[0373] - One-way ANOVA was performed to analyze differences between groups in age, weight, PVAS, and CADESI-04 scores.

[0374] Fisher's exact test was performed to analyze differences between groups in sex, breed size, and provisional diagnosis.

[0375] 3. During the clinical trial period

[0376] - Screening start date: January 23, 2025

[0377] - First clinical trial homologous material registration date: February 24, 2025

[0378] - Final clinical trial homologous animal visit end date: June 26, 2025

[0379] 4. Clinical trial subjects

[0380] 4.1 Status of Participation of Subject Animals in Clinical Trials

[0381] A total of 49 subjects were recruited for this clinical trial, and 48 subjects were registered after being deemed suitable for the trial based on selection and exclusion criteria. The guardian of the remaining 1 subject withdrew their consent prior to group assignment. Of the 48 registered subjects, 32 were randomly assigned to the administration group according to a pre-designed randomization table and conditions. Subsequently, the 16 registered subjects were assigned using a controlled assignment method to minimize group bias and recruitment imbalance (see Fig. 16).

[0382] 5. Validity Evaluation Results

[0383] 5.1 Basic characteristics of target animals prior to treatment

[0384] The mean ages of the 39 subjects included in the final efficacy evaluation analysis were 8.62 years, 8.38 years, and 8.54 years in the order of G1, G2, and G3, respectively, and the mean body weights were 8.93 kg, 10.58 kg, and 7.93 kg in the order of G1, G2, and G3, respectively, and no statistically significant differences were found between the administration groups in age and body weight (p = 0.986 and 0.761, respectively). The sex distribution was generally neutered males, which were similar across the groups (p = 0.361).

[0385] In terms of breed size distribution, small dogs (less than 10 kg) accounted for the majority, while medium (10–25 kg) and large dogs (over 25 kg) were relatively few (p = 1.000). Major registered breeds included Bichon Frise, Maltese, Poodle, Shih Tzu, and French Bulldog; each breed was included in G1, G2, and G3, indicating no bias toward a specific breed (see Table 14). There were also no significant differences in PVAS and CADESI-04 scores between groups at the time of registration (P = 0.119 and 0.933, respectively).

[0386] [Table 14]

[0387]

[0388] 5.2 Concomitant drugs

[0389] In this clinical trial, since all subjects did not receive any other drugs in combination with the test drug or the control drug, there were no confounding factors between groups due to the combined drugs.

[0390] 5.3 Provisional short lifespan of the target animals

[0391] As a result of analyzing the detailed provisional diagnosis of allergic dermatitis recorded at the time of animal registration for the 39 animals subject to final analysis, all animals had atopic dermatitis.

[0392] In addition, 17 subjects with findings of dietary allergy (6 in G1, 7 in G2, and 4 in G3), 8 subjects with findings of contact dermatitis (3 in G1, 2 in G2, and 3 in G3), and 1 subject with findings of flea allergy dermatitis (1 in G2) were reported as concomitant diagnoses. Furthermore, 11 subjects with findings of other allergic dermatitis (3 in G1, 2 in G2, and 6 in G3) were reported (see Table 15). No statistically significant difference was found in the detailed provisional diagnosis distribution among the three groups (p = 0.793).

[0393] [Table 15]

[0394]

[0395] 5.4 Medication Adherence

[0396] Medication adherence was calculated based on the number of doses taken in each interval during Visit 2 and Visit 3, and the final medication adherence was evaluated by summing the results of the two intervals (see Table 16). The average medication adherence for each group was 99.4–100.0% in Visit 2 and 98.6–100.0% in Visit 3, and the overall medication adherence over the entire period was 99.5–99.7%. The median for all groups was 100.0%, and there were no statistically significant differences between groups (p = 0.611, 0.378, and 0.913 for each period, respectively).

[0397] [Table 16]

[0398]

[0399]

[0400] 5.5 Presentation and Analysis of Validity Evaluation Results

[0401] For the efficacy evaluation, a total of 39 subjects were analyzed by setting the subjects who complied with the clinical trial protocol as the principal analysis group.

[0402] 5.5.1 Analysis of Daily Validity Evaluation Variables (Analysis of Changes in PVAS Scores Over Time)

[0403] PVAS was the primary efficacy endpoint repeatedly evaluated by the guardian, and changes in symptoms over time and differences between groups were analyzed, and changes according to the time of visit from Visit 1 to Visit 3 and the proportion of target animals that reached N2M were evaluated.

[0404] PVAS scores evaluated by guardians were collected over a total of 10 times (see Table 17). Before administration (Day 0 of administration), the mean PVAS scores of each group were similarly collected, and there were no statistically significant differences between the groups (p = 0.119).

[0405] PVAS scores decreased significantly in all groups from the beginning of administration compared to before administration (day 0 of administration). In all administration groups, PVAS scores gradually decreased until day 14 of administration; however, in the case of G1, PVAS scores showed an increasing trend (increasing from an average of 3.68 points to 3.88 points) on day 28 of administration (change in dosage to once a day after day 14) compared to day 14 of administration (administration twice a day), whereas in G2 and G3, PVAS scores showed a continuous decreasing trend until day 28 of administration.

[0406] At the 28th day of administration, the PVAS score of G3 was the lowest among the administration groups (3.22 2.50), and the rate of reduction in PVAS scores from before administration (0th day of administration) to 28th day of administration was 43.1%, 52.0%, and 57.6% for G1, G2, and G3, respectively.

[0407] [Table 17]

[0408]

[0409] Repeated measures analysis of variance (RM-ANOVA) was performed to evaluate the overall pattern of score change according to the date of visit (Table 18).

[0410] As a result of the analysis, the time factor had a statistically significant effect (p < 0.001), and the PVAS score in the entire group showed a tendency to decrease significantly over time.

[0411] [Table 18]

[0412]

[0413] The estimated marginal mean changes in PVAS scores by group over time showed a tendency for PVAS scores to decrease as the number of visit days increased in all groups, which was consistent with the results of repeated measures ANOVA (see Fig. 17).

[0414] To visually compare the range of change in PVAS scores before and after administration for each group, an interquartile range (IQR) analysis was performed. The gray dotted line indicates the baseline at PVAS 3.6 to confirm the normal to mild (N2M) level. For G1 and G2, most of the median and interquartile ranges were located outside or widely distributed within the respective regions at 28 days of administration. The median for G3 was located slightly below the baseline of PVAS 3.6, and the largest portion of the interquartile ranges was included within that region, indicating that the relatively largest number of subjects reached that level of treatment response.

[0415] 5.5.2 Analysis of Secondary Validity Evaluation Variables (Analysis of Changes in CADESI-04 Values ​​According to Visit Date)

[0416] CADESI-04 is a clinical indicator that quantifies the extent and severity of skin lesions as assessed by a veterinarian, and changes in symptoms were analyzed over 0, 14, and 28 days of administration.

[0417] CADESI-04 scores evaluated by veterinarians were assessed and collected a total of three times at each visit (see Table 19). The mean CADESI-04 scores of each group before administration (Day 0 of administration) did not show a statistically significant difference (p = 0.993).

[0418] For each group, there was a statistically significant decrease on days 14 and 28 of administration compared to before administration (day 0 of administration) (p < 0.05).

[0419] Analysis of changes in CADESI-04 scores by measurement time point showed that in all treatment groups, scores decreased on day 14 of treatment compared to before treatment (day 0 of treatment). For G1, the CADESI-04 score increased slightly on day 28 of treatment (change in dosage regimen to once daily after day 14) compared to day 14 of treatment (administration twice daily from the start of treatment), whereas it increased from an average of 32.54 points to 33.00 points. In contrast, G2 and G3 continued to decrease even on day 28 of treatment, with G3 showing the lowest average score and the largest decrease. The rate of decrease in CADESI-04 scores from before treatment (day 0 of treatment) to day 28 of treatment was confirmed to be 32.2%, 40.0%, and 49.1% for G1, G2, and G3, respectively.

[0420] [Table 19]

[0421]

[0422]

[0423] Repeated measures analysis of variance (RM-ANOVA) was performed to evaluate the overall pattern of change in CADESI-04 scores according to the date of visit. The analysis results showed that the time factor had a statistically significant effect (p < 0.001), and the CADESI-04 scores in the entire subject group showed a tendency to significantly decrease over time (see Table 20).

[0424] [Table 20]

[0425]

[0426] The estimated marginal mean change of CADESI-04 scores by group over time showed that in all groups, the CADESI-04 scores decreased as the visit days progressed, and a trend consistent with the results of repeated measures analysis of variance was confirmed (see Fig. 18).

[0427] Interquartile range (IQR) analysis was performed to visually compare the magnitude of change in CADESI-04 scores before and after administration for each group. The gray dotted line indicates the baseline to confirm the normal to mild (N2M) level with a CADESI-04 score of 35. For G1 and G2, the median value was located below the baseline at 28 days after administration, and the values ​​between the two groups were similar. G3 had a relatively lower median value compared to G1 and G2, indicating that a relatively larger number of subjects achieved a reduction in the extent and severity of skin lesions.

[0428] 6. Safety Assessment

[0429] 6.1 Analysis of Safety Evaluation Variables

[0430] The analysis was conducted on subjects that had completed all central laboratory tests among those that had administered the investigational veterinary drug at least once. Accordingly, a total of 42 subjects were analyzed, including 39 subjects for the final efficacy analysis and 3 subjects. 6.1.1 Analysis of Changes in Physiological Indicators

[0431] Major physiological indicators such as body weight, heart rate, respiratory rate, and body temperature were measured on each visit day to analyze the effect of administering the test drug and the control drug on safety (see Table 21).

[0432] The results of the repeated measures ANOVA analysis showed that there were no statistically significant differences in the time, group, and time X group interaction factors for all indicators other than body temperature (p > 0.05).

[0433] Body weight remained at similar levels across all groups during the visit period, with no significant changes observed. Heart rate and respiratory rate showed overall changes within the physiological normal range, without significant differences between groups or over time. Body temperature showed a significant difference only in the time factor, but the changes were within the normal range, and there were no clinically significant changes such as hypothermia.

[0434] [Table 21]

[0435]

[0436] 6.1.2 Analysis of results of central laboratory tests (hematological and blood biochemical tests) Central laboratory tests were performed on the subjects before and after administration during the clinical trial period. Among them, abnormal values ​​in hematological tests were confirmed in 3 subjects, but no clinically significant signs of infection or clinical symptoms were observed, and it was determined to be a mild physiological response of low clinical significance.

[0437] In addition, repeated measures analysis of variance (RM-ANOVA) was performed to evaluate the overall patterns of change in blood tests according to the date of visit for each group and interactions between groups. The analysis results showed that among the blood test items, reticulocytes, RET%, platelet count, Pct, amylase, and Na / K ratio showed statistically significant changes over time; however, these values ​​were within the normal range with no clinically significant changes, and no interaction between time and group was observed.

[0438] Among the liver function indicators, AST, creatinine, and CK levels showed statistically significant interactions between time and groups, but the changes were within the normal range.

[0439] 6.1.3 Analysis of Abnormal Reaction Results

[0440] There were no serious adverse events (SAEs) during the clinical trial period, and a total of 3 adverse event cases were collected through guardians and veterinarians (2 cases in G1 and 1 case in G3), all of which were mild and recovered without separate clinical treatment. In addition, in the causality analysis, all cases were classified as having an unclear or low causal relationship with the investigational drug or the control drug.

[0441] 7. Conclusion and Discussion

[0442] This clinical trial was conducted to evaluate the efficacy and safety of Example 1, a JAK1 selective inhibitor, on dogs diagnosed with allergic dermatitis who visited an animal hospital with pruritus as the main symptom, and the therapeutic response according to the dosage and administration of the test drug Example 1 tablet was compared and analyzed with the control drug Apoquel® tablet.

[0443] As a result of the efficacy evaluation, G1, G2, and G3 all showed significant therapeutic effects compared to before administration. G3 showed relatively superior efficacy, consistently demonstrating the greatest improvement across all evaluation indicators (PV AS, CADESI-04). Conversely, in the case of G1, efficacy indicators improved significantly up to day 14 of administration with twice-daily administration, but the overall improvement in efficacy tended to be halved on day 28 of administration when the dosage regimen was changed to once-daily; this phenomenon was similarly observed in clinical trials of Zenrelia™ (ilunocitinib), which was recently approved as a JAK inhibitor treatment. Due to these reasons, ochlasitinib exhibits an accumulation of blood drug exposure in the body upon repeated administration and possesses relatively low JAK2 selectivity; therefore, it is judged that when administered twice a day for the initial two weeks and then reduced to once a day thereafter, the effect is halved compared to the initial dose. In contrast, Example 1 has high selectivity for JAK2 and does not exhibit an accumulation of blood drug exposure in the body even upon repeated administration, so it is expected to demonstrate excellent efficacy without safety concerns even with a once-daily dosage regimen.

[0444] As a result of the safety evaluation, no serious adverse events (SAEs) were reported, and no serious adverse events were identified in physiological indicators or central laboratory test results in any of the treatment groups. The three reported adverse events (two in G1 and one in G3) resolved without clinical treatment and were assessed as 'unlikely related' and 'unassessable' to the investigational animal drug, confirming an overall excellent safety profile in the treatment groups.

[0445] In conclusion, Example 1 is considered to be a therapeutic candidate substance that possesses both efficacy and safety for canine allergic dermatitis. In particular, G3 showed the best results in terms of consistency and maintenance of symptom improvement and guardian satisfaction, and is expected to provide advantages in terms of guardian convenience due to once-daily administration (QD).

[0446] Based on the results described in the above experimental examples, the pharmaceutical composition of the present invention comprising the heterocyclic compound of Formula 1 or a pharmaceutically acceptable salt thereof exhibits excellent JAK1 selective inhibitory activity even with once-daily administration, and since no drug accumulation occurs with repeated administration, it may be possible to reduce the risk of side effects associated with existing JAK inhibitors. Therefore, it is expected to be an option for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals with improved safety and tolerability.

Claims

1. [Scope of Claim] 【Claim 11 A pharmaceutical composition comprising N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide or a pharmaceutically acceptable salt thereof as an active ingredient for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals, The above pharmaceutical composition is a pharmaceutical composition administered orally at a concentration of 0.4 to 1.35 mg / kg based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo[2,3- b]pyridin- 6-yl )cyclopropanecarboxamide.

2. In Article 1, The above N _(4- (1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl) The pharmaceutically acceptable salts of cyclopropanecarboxamide are hydrochloride, hydrogen bromide, mesylate, phosphate, napadisilate, camsilate, or oxalate, pharmaceutical composition. 【claim Paragraph 3] In Article 1, The above N _(4- (1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarvoxamide is (S)-N-(4-(1- (2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,Sb]pyridin-6-yl)cyclopropanecarvoxamide, pharmaceutical composition.

4. In Article 1, The above pharmaceutical composition is a pharmaceutical composition administered orally at a dose of 0.4 to 1.35 mg / kg once daily based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo[2,3- b]pyridin- 6-yl )cyclopropanecarboxamide. 【claim Paragraph 5] A pharmaceutical composition according to claim 1, wherein the pharmaceutical composition is administered orally at a dose of 0.9 to 1.35 mg / kg once daily based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl )- 1H-pyrrolo[2,3- b]pyridin- 6-yl )cyclopropanecarboxamide. 【claim Paragraph 6] In Article 1, The above pharmaceutical composition is a pharmaceutical composition administered orally at a dose of 0.4 to 0.9 mg / kg once daily based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo[2,3- b]pyridin- 6-yl )cyclopropanecarboxamide. 【claim Paragraph 71 In Article 1, The above pharmaceutical composition is a pharmaceutical composition comprising N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide administered orally at a dose of 0.4 to 0.9 mg / kg twice daily. 【Claim】 Paragraph 8] In Article 1, The above pharmaceutical composition is administered orally at a dose of 0.45 to 0.9 mg / kg twice daily based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl )- 1H-pyrrolo[2,3- b]pyridin- 6-yl )cyclopropanecarboxamide. 【claim Paragraph 91 In Article 1, The above pharmaceutical composition is a pharmaceutical composition administered until the time of alleviation or improvement of symptoms of allergic dermatitis, atopic dermatitis, or pruritus.

10. In Article 1, The above pharmaceutical composition is administered for a period of 1 to 7 days, 1 to 14 days, 1 to 21 days, 1 to 28 days, 7 to 28 days, 14 to 28 days, 21 to 28 days, or 28 days or more.

11. In Article 1, The above pharmaceutical composition is administered orally at a dose of 0.6 to 1.35 mg / kg based on N- (4- (1- (2-cyanoacetyl )-3-methyl- 1,2,3,6-tetrahydropyridin-4-yl )- 1H-pyrrolo [2,3- b]pyridin-6-yl )cyclopropanecarboxamide once daily for a period of 1 to 28 days or more than 28 days.

12. In Paragraph 11, The above pharmaceutical composition is administered orally once daily for 28 days at a concentration of 0.9 to 1.35 mg / kg based on N- (4- (1- (2-cyanoacetyl )-3-methyl- 1,2,3,6-tetrahydropyridin-4-yl )- 1H-pyrrolo [2,3- b]pyridin-6-yl )cyclopropancarboxamide.

13. In Article 1, The above pharmaceutical composition is administered orally at a dose of 0.6 to 0.9 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide twice daily for 1 to 14 days, 7 to 14 days, or 14 days, and thereafter at a dose of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl) for a period of 1 to 14 days, 7 to 14 days, 14 days, or longer than 14 days, twice daily. A pharmaceutical composition administered orally at a concentration of 0.6 to 0.9 mg / kg based on din-6-yl cyclopropanecarvoxamide.

14. In Paragraph 13, The above pharmaceutical composition is administered orally at a dose of 0.6 to 0.9 mg / kg based on N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropancarboxamide twice daily for 14 days, and thereafter for 14 days ! A pharmaceutical composition administered orally at a dose of 0.6 to 0.9 mg / kg based on N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)- 1H-pyrrolo [2,3- b]pyridin- 6-yl)cyclopropanecarboxamide.

15. In Article 1, The above-mentioned mammal is a pharmaceutical composition that is unresponsive to ochlasitinib or a pharmaceutically acceptable salt thereof, exhibits a chronic history of non-seasonal pruritus, has a PVAS score of 6 or higher based on the baseline (Day 0, Baseline), or has an overactivated JAK1-STAT3 signaling pathway compared to normal individuals.

16. In claim 1, the pharmaceutical composition is a pharmaceutical composition that improves, alleviates, or treats one or more symptoms selected from the group consisting of itching, pruritus, redness, lichenification, abrasion, and hair loss.

17. In Article 1, The above pharmaceutical composition is a pharmaceutical composition that does not cause plasma accumulation of N- (4- (1- (2-cyanoacetyl )-3-methyl- 1,2,3,6-tetrahydropyridin-4-yl )- 1H-pyrrolo [2,3- b]pyridin-6-yl ) cyclopropanecarboxamide or pharmaceutically acceptable salts thereof upon repeated administration two or more times.

18. In Article 1, The above pharmaceutical composition is a pharmaceutical composition in which, even at high doses, the plasma concentration of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropancarboxamide or a pharmaceutically acceptable salt thereof does not reach an IC50 for JAK2.

19. A method for preventing or treating allergic dermatitis, atopic dermatitis, or pruritus in mammals, comprising the step of orally administering to a mammal N- (4- (1- (2-cyanoacetyl)- 3-methyl- 1,2,3,6-tetrahydropyridin- 4-yl)-1H-pyrrolo[2,3- b]pyridin- 6-yl)cyclopropanecarboxamide, a pharmaceutically acceptable salt thereof, or the composition of Claim 1.

201. Use of N-(4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, pharmaceutically acceptable salts thereof, or the composition of claim 1 for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals.

21. For the manufacture of a drug for the prevention or treatment of allergic dermatitis, atopic dermatitis, or pruritus in mammals N _ (4-(1-(2-cyanoacetyl)-3-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1H-pyrrolo[2,3-b]pyridin-6-yl)cyclopropanecarboxamide, pharmaceutically acceptable salts thereof, or the use of the composition of claim 1.

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