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WO2024205336A1 - Composition pharmaceutique présentant une activité inhibitrice de la tubuline - Google Patents

Composition pharmaceutique présentant une activité inhibitrice de la tubuline Download PDF

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WO2024205336A1
WO2024205336A1 PCT/KR2024/004107 KR2024004107W WO2024205336A1 WO 2024205336 A1 WO2024205336 A1 WO 2024205336A1 KR 2024004107 W KR2024004107 W KR 2024004107W WO 2024205336 A1 WO2024205336 A1 WO 2024205336A1
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cells
chemical formula
pharmaceutical composition
present
disease
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장규범
안치현
양영은
이준호
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에이치엘비생명과학알앤디 주식회사
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/4985—Pyrazines or piperazines ortho- or peri-condensed with heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]

Definitions

  • the present invention relates to a novel use of a compound having tubulin inhibitory activity, and more particularly, to a pharmaceutical composition for inhibiting tubulin polymerization, comprising the compound of the present invention or a pharmaceutically acceptable salt thereof as an active ingredient.
  • Microtubules are the main components of the cytoskeleton, which are composed of heterodimers of ⁇ -tubulin and ⁇ -tubulin. ⁇ -Tubulin and ⁇ -tubulin are arranged in a spiral arrangement to form a hollow tube with an outer wall diameter of about 25 nm and an inner wall diameter of about 14 nm. ⁇ -tubulin is exposed at the (+) end of the microtubule, and ⁇ -tubulin is exposed at the (-) end. Microtubules, which are dynamic polymers, recycle tubulin through continuous polymerization and depolymerization processes. This dynamics of microtubules can be regulated by binding proteins, etc.
  • Microtubules are absolutely necessary for all eukaryotic cells and perform various functions, such as maintaining the cellular framework and shape, cell movement, chromosome segregation by spindle formation during cell division, and intracellular signal transmission. Additionally, it plays a critical role in the movement of mRNA, proteins, vesicles, and organelles between the cell body and axon together with motor proteins, or forms the internal structure of cilia and flagella.
  • Microtubule-targeting therapeutics change microtubule dynamics by stabilizing or destabilizing microtubules. This has led to cytotoxic effects caused by cell division arrest and cell death, and they are being developed as anticancer agents, or as treatments for other diseases related to cell migration and metastasis, and inflammatory responses through innate immune activation.
  • microtubule-stabilizing agents include taxanes, paclitaxel (Taxol), and docetaxel, which bind to the taxane-binding site or the overlapping site of ⁇ -tubulin to prevent microtubule depolymerization and enhance polymerization.
  • microtubule-destabilizing agents include colchicine and vinca alkaloids, which can interfere with microtubule formation by binding to the colchicine-binding site or the vinca-binding site.
  • Colchicine a representative microtubule inhibitor, binds to tubulin to form a complex, thereby inhibiting the elongation of microtubule polymers, and thus exhibits an anti-inflammatory effect. Therefore, it has been used as a gout treatment since the past, and is also used to treat familial Mediterranean fever and the amyloidosis caused by it.
  • the benzimidazole compound group such as albendazole, mebendazole, triclabendazole, and thiabendazole, which bind closely to the colchicine-binding site of ⁇ -tubulin, have been used as antiparasitic agents to treat a wide variety of parasitic infections.
  • CA-4P combretastatin A-4 phosphate
  • microtubule-targeting therapeutics must overcome are systemic toxicity and acquired drug resistance.
  • nocodazole was recognized as a possible anthelmintic, but it was not used clinically due to poor bioavailability and high toxicity.
  • Colchicine has a narrow therapeutic index where the non-toxic dose, toxic dose, and lethal dose are not clearly distinguished, so serious toxicity has been observed at the approved therapeutic dose. Therefore, there is increasing interest in developing new tubulin inhibitors that can overcome the resistance mechanisms of existing drugs.
  • the present inventors have completed the present invention by confirming the excellent tubulin polymerization inhibition effect of the compound of the present invention in a situation where there is an increasing demand for the development of novel tubulin inhibitors that can reduce toxicity without lowering therapeutic efficacy or inducing resistance.
  • One object of the present invention is to provide a pharmaceutical composition for inhibiting tubulin polymerization, comprising a compound having tubulin inhibitory activity or a pharmaceutically acceptable salt thereof as an active ingredient.
  • Another object of the present invention is to provide a method for inhibiting tubulin polymerization, comprising the step of administering a therapeutically effective amount of the compound to a subject in need thereof.
  • a pharmaceutical composition comprising the compound of the present invention or a pharmaceutically acceptable salt thereof as an active ingredient exhibits tubulin inhibitory activity, and therefore can be usefully used for the purpose of inhibiting tubulin polymerization, particularly for the prevention or treatment of inflammatory diseases, acute respiratory distress syndrome, actinic keratosis, or diseases related to neovascularization.
  • Figure 1 shows the results of tubulin polymerization analysis according to treatment with HLS-22001, HLS-22002 or microtubule stabilizers (paclitaxel, nocodazole).
  • Figure 2 shows the results of tubulin polymerization analysis according to treatment with HLS-22001, HLS-22002, microtubule-stabilizing agents (paclitaxel, docetaxel), or microtubule-destabilizing agents (plinabulin, combretastatin, vincristine, nocodazole).
  • microtubule-stabilizing agents paclitaxel, docetaxel
  • microtubule-destabilizing agents plinabulin, combretastatin, vincristine, nocodazole.
  • Figure 3 shows the results of tubulin polymerization analysis according to treatment with HLS-22001, HLS-22002, nocodazole, paclitaxel, or tirvanibuline.
  • Figure 4 shows the results of confirming tubulin expression according to treatment with different concentrations of HLS-22001, HLS-22002, nocodazole, or tirvanibulin in HT297.T and A431 cells.
  • Figure 5 is a graph comparing cell viability following HLS-22001 or HLS-22002 treatment with nocodazole or tirvanibuline in HT297.T and A431 cells.
  • Figure 6 is a graph showing the results of comparing the cell cycle distribution ratio according to HLS-22001 or HLS-22002 treatment with 5-FU, nocodazole or tirvanibuline in HT297.T and A431 cells.
  • Figure 7 is a graph showing the results of comparing the rate of cell death progression of dead cells by HLS-22001 or HLS-22002 treatment with 5-FU, nocodazole or tirvanibulin in HT297.T and A431 cells.
  • Figure 8 shows the results of Western blot confirming the expression of c-PARP and c-caspase 3 after treatment of A431 cells with HLS-22001, HLS-22002, 5-FU, nocodazole, or tirvanibulin.
  • Figure 9 is a graph showing cell survival rate by HLS-22001 treatment in THP-1 cells.
  • Figure 10 is a graph comparing the levels of IL-1 ⁇ and IL-6 mRNA expression by HLS-22001 treatment in LPS-stimulated THP-1 cells.
  • Figure 11 is a graph comparing the levels of IL-1 ⁇ and TNF- ⁇ secretion by HLS-22001 treatment in LPS-stimulated THP-1 cells.
  • One embodiment of the present invention for achieving the above object is a pharmaceutical composition for inhibiting tubulin polymerization, comprising a compound represented by the following chemical formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
  • R 1 is hydrogen or halogen
  • R 2 is straight or branched chain C 1-6 alkyl
  • R 3 to R 7 are each independently hydrogen, halogen, straight-chain or branched C 1-6 alkyl, straight-chain or branched C 1-6 haloalkyl, C 1-6 alkoxy or C 1-6 haloalkoxy.
  • Another embodiment for achieving the above object is a method for inhibiting tubulin polymerization, comprising the step of administering a therapeutically effective amount of a compound represented by the chemical formula 1 or a pharmaceutically acceptable salt thereof to a subject in need thereof.
  • Another embodiment for achieving the above object is the use of a compound represented by the above chemical formula 1 or a pharmaceutically acceptable salt thereof for inhibiting tubulin polymerization.
  • Another embodiment for achieving the above object is the use of a compound represented by the above chemical formula 1 or a pharmaceutically acceptable salt thereof for preparing a medicament for inhibiting tubulin polymerization.
  • the compound represented by the chemical formula 1 may be a compound represented by the following chemical formula 2.
  • R 6 is straight-chain or branched C 1-3 alkyl, straight-chain or branched C 1-3 haloalkyl or C 1-3 alkoxy.
  • the compound represented by the chemical formula 1 may be a compound represented by the following chemical formula 3 or chemical formula 4.
  • the compound represented by the above chemical formula 3 is 4-(3-methoxy-5-methylphenyl)-N-(2-methoxythieno[3,2-b]pyrazin-3-yl)piperazine-1-carboxamide, also referred to herein as 'HLS-22001'.
  • the compound represented by the above chemical formula 4 is 4-(3,5-dimethoxyphenyl)-N-(2-methoxythieno[3,2-b]pyrazin-3-yl)piperazine-1-carboxamide, also referred to herein as 'HLS-22002'.
  • alkyl means a straight-chain or branched-chain acyclic saturated hydrocarbon, unless otherwise specified.
  • C 1-6 alkyl means alkyl having 1 to 6 carbon atoms.
  • alkyl may include, but is not limited to, methyl, ethyl, n -propyl, n -butyl, isopropyl, sec -butyl, isobutyl, or tert -butyl.
  • alkoxy means an alkyl ether group, -(O-alkyl), wherein alkyl is as defined above.
  • C 1-6 alkoxy means an alkoxy containing C 1-6 alkyl , i.e., -(OC 1-6 alkyl), and as an example, the alkoxy may include, but is not limited to, methoxy, ethoxy, n -propoxy , isopropoxy, n -butoxy, isobutoxy , sec -butoxy, or tert -butoxy.
  • halogen may be F, Cl, Br, or I.
  • haloalkyl or “haloalkoxy” refers to an alkyl group or alkoxy group in which one or more hydrogen atoms of the alkyl group or alkoxy group are replaced with halogen atoms, wherein halogen, alkyl and alkoxy are as defined above.
  • haloalkyl or haloalkoxy may be an alkyl group or alkoxy group in which one or more hydrogen atoms are replaced with homogeneous or heterogeneous halogen atoms.
  • the compound of the present invention may exist in the form of a pharmaceutically acceptable salt. Accordingly, the category of the compound of the present invention includes a pharmaceutically acceptable salt of the compound represented by the chemical formula 1.
  • pharmaceutically acceptable salt in the present invention means a salt commonly used in the pharmaceutical industry, and includes, for example, an inorganic ionic salt manufactured with calcium, potassium, sodium or magnesium; an inorganic acid salt manufactured with hydrochloric acid, nitric acid, phosphoric acid, hydrobromic acid, iodic acid, perchloric acid or sulfuric acid; an organic acid salt manufactured with acetic acid, trifluoroacetic acid, citric acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, mandelic acid, propionic acid, lactic acid, glycolic acid, gluconic acid, galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, aspartic acid, ascorbic acid, carbonic acid, vanillic acid or hydroiodic acid; a sulfonic acid salt manufactured with methanesulfonic acid, ethanesulfonic acid, benzenes
  • the compound of the present invention includes not only pharmaceutically acceptable salts, but also all salts, hydrates and solvates that can be prepared by conventional methods.
  • tubulin polymerization refers to a process in which ⁇ -tubulin and ⁇ -tubulin within a cell polymerize to form microtubules, and the compound of the present invention can inhibit tubulin polymerization in vitro , in vivo , or ex vivo , and therefore has an excellent effect as a composition for inhibiting tubulin polymerization.
  • a pharmaceutical composition comprising the compound of the present invention or a pharmaceutically acceptable salt thereof as an active ingredient can be used as a pharmaceutical composition for the prevention or treatment of diseases for which the effect can be expected by inhibition of tubulin polymerization, such as inflammatory diseases, acute respiratory distress syndrome, actinic keratosis, or diseases related to neovascularization.
  • diseases for which the effect can be expected by inhibition of tubulin polymerization such as inflammatory diseases, acute respiratory distress syndrome, actinic keratosis, or diseases related to neovascularization.
  • the term "inflammatory disease” refers to any condition that is characterized by a local or systemic biological defense response to external physical or chemical stimuli or infection by external infectious agents such as bacteria, fungi, viruses, and various allergens. This response involves a series of complex physiological reactions such as activation of various inflammatory mediators and enzymes related to immune cells (e.g., iNOS, COX-2, etc.), secretion of inflammatory mediators (e.g., secretion of NO, TNF- ⁇ , IL-6, IL-1 ⁇ , PGE2), infiltration of body fluids, cell migration, and tissue destruction, and is externally manifested by symptoms such as erythema, pain, edema, fever, and deterioration or loss of specific body functions.
  • inflammatory mediators e.g., secretion of NO, TNF- ⁇ , IL-6, IL-1 ⁇ , PGE2
  • the inflammatory disease may be acute, chronic, ulcerative, allergic, or necrotic, as long as any disease is included in the definition of the inflammatory disease as described above, it does not matter whether it is acute, chronic, ulcerative, allergic, or necrotic.
  • the compound of the present invention inhibits the expression and/or secretion of proinflammatory cytokines IL-6, IL-1 ⁇ and TNF- ⁇ in immune cells, and therefore has an excellent effect in the prevention and treatment of inflammatory diseases.
  • the inflammatory disease may be, but is not limited to, atopy, psoriasis, dermatitis, allergy, arthritis, rhinitis, otitis media, pharyngitis, tonsillitis, bronchitis, pneumonia, cystitis, nephritis, prostatitis, pyelonephritis, pelvic inflammatory disease, hepatitis, inflammatory bowel disease, Crohn's disease, ankylosing spondylitis, systemic lupus erythematosus, arteriosclerosis, coronary arteritis, Behcet's disease, asthma, edema, rheumatoid arthritis, delayed-onset allergy (type IV allergy), transplant rejection, graft-versus-host disease, autoimmune encephalomyelitis, multiple sclerosis, cystic fibrosis, diabetic retinopathy, rhinitis, ischemic-reperfusion injury, vascular restenos
  • colchicine significantly affects the deformability and motility of human neutrophils, and thus the cytoskeleton can act as a pharmacological target in the inflammatory process involving neutrophils.
  • colchicine has been reported to inhibit the activation of the NLRP3 inflammasome, block IL-1 ⁇ secretion, inhibit the expression of genes involved in cell regulation, and reduce the expression of endothelial cell adhesion proteins and IL-1-induced L-selectin, thereby reducing cytokines and neutrophil chemotaxis (Nicola Dalbeth, MD et al., Clinical Therapeutics, 36, 1465-1479, 2014). Therefore, it can be seen that the cytoskeletal changes resulting from the tubulin inhibition effect are closely related to the regulation of inflammatory responses and the treatment of diseases caused by them.
  • analogues of noscapine are known to exhibit anti-inflammatory activity by inhibiting TLR-mediated TNF- ⁇ and NO release in human and mouse macrophages. Brominated noscapine analogues induce cellular autophagy to alleviate inflammatory responses (Susu Werhaier et al., PloS One, 5(2), e9165, 2010). Anti-inflammatory effects were also confirmed in experiments in which NT-07-16 was treated on mouse macrophage RAW264.7 cells, which are widely used as an inflammation model for evaluation of anti-inflammatory effects. NT-07-16 is a microtubule depolymerizing agent.
  • acute respiratory distress syndrome in the present invention refers to a lung disease that causes respiratory distress that does not improve even when high concentrations of oxygen are supplied, and acute respiratory failure may occur due to lung damage caused by inflammation.
  • Acute respiratory distress syndrome can be caused by various causes, such as infection with pathogens, trauma, and genetic predisposition, and lung damage that occurs in acute respiratory distress syndrome is highly related to a systemic inflammatory response, which may cause multi-organ failure complications, including acute kidney damage.
  • acute respiratory distress syndrome progresses in the alveoli or pulmonary blood vessels, inflammatory cells and proteins fill the alveoli instead of air due to a severe inflammatory response, a hyaline membrane is formed, and inflammatory cell infiltration is observed in the pulmonary interstitium.
  • actinic keratosis refers to precancerous lesions that occur on sun-exposed areas of the skin. Actinic keratosis can appear as rough, dry, tan, pink or red spots (lesions) on the scalp, including the face, throat, neck, nose, forehead, ears, or lips, and may also appear on other parts of the body that have been exposed to intense sunlight for long periods of time, such as the hands, back, and other parts of the trunk and legs.
  • Actinic keratosis as used herein includes all clinical variants, such as classic (or general), hypertrophic (or hyperkeratotic), atrophic, actinic keratosis with cutaneous horn, pigmented actinic keratosis, actinic cheilitis and Bowenoid actinic keratosis.
  • a tubulin inhibitor it is known that the amount of IL-8 released is not higher than that of 5-FU in CCD-1106 KERTr cells, that the apoptosis marker IL-1 ⁇ is increased, and that the growth of highly proliferative cells is inhibited (Todd Schlesinger et al., Clin Cosmet Investig Dermatol, 15, 2495-2506, 2022), suggesting the possibility that microtubule or tubulin inhibitors can be utilized as therapeutic agents for actinic keratosis.
  • the compound of the present invention since the compound of the present invention has superior tubulin polymerization inhibition activity than tirvanibulin, it can be more usefully used for the prevention and treatment of actinic keratosis.
  • angiogenic disease in the present invention refers to a disease, disorder or condition related to or caused by angiogenesis, and particularly, considering the efficacy of the compounds of the present invention, refers to a disease caused by excessive angiogenesis.
  • Angiogenesis-related disease may be related to the formation of blood vessels that should not be formed, or the formation of blood vessels that are abnormally localized or strong.
  • Angiogenesis is the process by which new blood vessels are formed from preexisting blood vessels, and includes the growth and/or development of new blood vessels, the dilatation of small blood vessels, excessive or long-term blood vessel growth, and the maintenance of existing vasculature.
  • Angiogenesis involves a series of sequential steps including the degradation of the basement membrane by proteases secreted by activated endothelial cells, the migration of endothelial cells, the proliferation of endothelial cells, the formation of solid endothelial cell sprouts into the stromal space, and the formation and deposition of new basement membrane tight junctions, along with the formation of vascular rings and capillaries, which processes are important in normal regeneration, embryogenesis and wound healing.
  • inappropriately regulated angiogenesis has been linked to a number of diseases, including cancer.
  • the neovascularization-related disease may be, but is not limited to, age-related macular degeneration, neovascular glaucoma, retinal vein occlusion, myopic macular degeneration, retinopathy of prematurity, proliferative diabetic retinopathy, posterior capsular opacification, pediatric hemangioma, acne rosacea, Kaposi's sarcoma, atopic keratitis, epidemic keratoconjunctivitis, bacterial ulcer, fungal ulcer, herpes simplex infection, herpes zoster infection, protozoal infection, mycobacterial infection, polyarteritis, sarcoidosis, scleritis, rosacea, Sjogren's disease, systemic lupus, acquired immunodeficiency syndrome (AIDS), syphilis, or Treponema pallidum or related parasitic infection that increases angiogenesis.
  • age-related macular degeneration neovascular glau
  • Angiogenic inhibitors such as vascular disrupting agents (VDAs)
  • VDAs vascular disrupting agents
  • CA-4P has been found to inhibit the proliferation and migration of human umbilical vascular endothelial cells (HUVECs) by inhibiting microtubule polymerization, and to induce morphological transformation of endothelial cells, thereby weakening blood vessels (Changwen Deng et al., Mol Ther, 28(1), 75-88, 2020).
  • UUVECs human umbilical vascular endothelial cells
  • 2013/0131018 describes the tubulin inhibition and cytotoxic effects of various microtubule inhibitors, as well as the therapeutic use and method of isoquinolines by inhibiting tubulin polymerization or inhibiting unwanted neovascularization, which can be seen in the above content, the association between tubulin polymerization and neovascularization-related diseases.
  • the compound of the present invention exhibits an effect of inhibiting tubulin polymerization by binding to tubulin.
  • the compound of the present invention may specifically bind to ⁇ -tubulin, and specifically may inhibit tubulin polymerization by binding to a colchicine portion of ⁇ -tubulin (e.g., a region including cysteine 239 and cysteine 354) to form a complex, but is not limited thereto.
  • the compound of the present invention exhibits an effect of inhibiting microtubule formation.
  • the compound of the present invention exhibits cytotoxicity in actinic keratosis and squamous cell carcinoma cells.
  • the compound of the present invention exhibits a G2/M cell cycle arrest effect in actinic keratosis and squamous cell carcinoma cells.
  • the compound of the present invention exhibits an apoptotic effect in actinic keratosis and squamous cell carcinoma cells.
  • the compound of the present invention exhibits little cytotoxicity in cells that are not in a pathological state, so that a reduction in side effects can be expected compared to other drugs.
  • the compound of the present invention exhibits an effect of inhibiting the expression and/or secretion of proinflammatory cytokines such as IL-1 ⁇ , IL-6 mRNA, and TNF- ⁇ in immune cells.
  • proinflammatory cytokines such as IL-1 ⁇ , IL-6 mRNA, and TNF- ⁇ in immune cells.
  • treatment refers to clinical intervention to alter the natural course of an individual or cell to be treated.
  • the desired therapeutic effect includes alleviating the symptoms of a disease, reducing all direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, alleviating or temporarily alleviating the disease state, curing or improving the prognosis.
  • the present invention includes all acts that improve the course of a disease by administering the pharmaceutical composition of the present invention.
  • prevention in the present invention means all acts that inhibit or delay the onset or recurrence of the disease by a composition comprising a compound having tubulin inhibitory activity according to the present invention.
  • the pharmaceutical composition of the present invention can be used in the form of a general pharmaceutical preparation, and may additionally contain a suitable carrier, excipient, diluent, or a combination thereof commonly used in the manufacture of a pharmaceutical composition.
  • a suitable carrier such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants commonly used.
  • the pharmaceutically acceptable carriers include, but are not limited to, those commonly used in the art, such as lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, mineral oil, and the like.
  • Solid preparations for oral administration may include tablets, pills, powders, granules, capsules, etc., and these solid preparations may be formulated by including one or more compounds and at least one excipient, such as starch, calcium carbonate, sucrose or lactose, gelatin, etc. In addition to simple excipients, lubricants such as magnesium stearate, talc, etc. may also be used.
  • Liquid preparations for oral administration include suspensions, solutions, emulsions, syrups, etc., and in addition to commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, flavoring agents, preservatives, etc. may be included.
  • Preparations for parenteral administration may include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories.
  • Non-aqueous solvents and suspending agents can include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate.
  • Suppository bases can include witepsol, macrogol, Tween 61, cacao butter, laurin butter, and glycerogelatin.
  • the pharmaceutical composition of the present invention may be used alone, or in combination with one or more other effective ingredients or auxiliary ingredients, for the purpose of increasing efficacy, reducing side effects, etc., of the compound represented by the chemical formula 1 or a pharmaceutically acceptable salt thereof.
  • the pharmaceutical composition of the present invention may be administered in single or multiple doses in a therapeutically effective amount.
  • the pharmaceutical composition of the present invention can be administered to a subject in need of tubulin polymerization inhibition in a therapeutically effective amount.
  • subject as used herein means any animal, including mammals such as humans, that has developed or may develop a tubulin polymerization-related disease, such as an inflammatory disease, acute respiratory distress syndrome, actinic keratosis, or an angiogenesis-related disease, and may typically be an animal that can exhibit a beneficial effect by treatment with the compound of the present invention or a pharmaceutically acceptable salt thereof.
  • any subject that has symptoms of a tubulin polymerization-related disease or is likely to have such symptoms is included in the scope of the present invention without limitation.
  • administration means introducing a given substance into a human or animal by any appropriate method, and the route of administration of the composition of the present invention may be oral or parenteral administration through any general route as long as it can reach the target tissue.
  • the composition of the present invention may be administered by any device through which the active ingredient can move to the target cell.
  • terapéuticaally effective amount means an amount sufficient to prevent or treat a disease at a reasonable benefit/risk ratio applicable to medical prevention or treatment, and the effective dosage level can be determined according to the severity of the disease, the activity of the drug, the patient's age, weight, health, sex, the patient's sensitivity to the drug, the time of administration of the composition used, the route of administration and the excretion rate, the treatment period, the drug used in combination with or concurrently with the composition of the present invention, and other factors well known in the medical field.
  • the compound represented by Chemical Formula 1 according to the present invention or a pharmaceutically acceptable salt thereof can be administered at 0.0001 to 100 mg/kg per day, and the administration can be administered once a day or in several divided doses.
  • the pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, and humans by various routes, for example, oral administration, intrathecal, intra-auricular, intraperitoneal or intravenous, intramuscular, subcutaneous, intrauterine, sublingual, or intracerebrovascular injection, but is not limited thereto.
  • the pharmaceutical composition of the present invention may contain 0.01 to 95 wt%, preferably 1 to 80 wt%, of the compound represented by chemical formula 1 or a pharmaceutically acceptable salt thereof based on the total weight of the composition.
  • the pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents.
  • the pharmaceutical composition of the present invention may be administered singly or in multiple doses. Considering all of the above factors, it is important to administer an amount that can achieve the maximum effect with the minimum amount without side effects, and this can be easily determined by those skilled in the art.
  • 6-Chloropyrazin-2-amine (10.4 g, 80 mmol) was dissolved in methanol (300 ml), and N-bromosuccinimide (15.6 g, 88 mmol) was added while stirring at room temperature. The mixture was stirred for further 60 minutes, concentrated under reduced pressure, and water was added. The mixture was extracted three times with ethyl acetate. The organic layer was collected, dried over magnesium sulfate, concentrated under reduced pressure, and the residue was purified by chromatography on a silica gel column. The residue was eluted with a mixed solvent of hexane and ethyl acetate (3:1 v/v) to obtain the title compound (12.0 g, 72%).
  • 5-Bromo-6-chloropyrazin-2-amine (4.2 g, 20 mmol) obtained from the above step 1) and copper iodide (190 mg, 1 mmol), triethylamine (16.7 ml, 120 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) complex dichloromethane (1.3 g, 1.6 mmol) and tetrahydrofuran (100 ml) were added and nitrogen gas was bubbled. Trimethylsilylacetylene (3.6 ml, 26 mmol) was added to the mixture and reacted at 80 °C for 2 hours.
  • the reaction mass was concentrated under reduced pressure, water was added, and the mixture was extracted three times with ethyl acetate.
  • the organic layer was recovered, dried over magnesium sulfate, concentrated under reduced pressure, and the residue was purified by chromatography on a silica gel column.
  • the title compound (3.16 g, 70%) was obtained by eluting with a mixed solvent of hexane and ethyl acetate (5:1 v/v).
  • 6-Chloro-5-((trimethylsilyl)ethynyl)pyrazin-2-amine obtained from the above step 2) was dissolved in dimethylformamide (5.0 g, 22.2 mmol), sodium sulfide pentahydrate (14.9 g, 88.8 mmol) was added, and the mixture was stirred at 90 °C for 2 hours. Water was added, and extraction was performed three times with ethyl acetate. The organic layer was recovered, dried over magnesium sulfate, concentrated under reduced pressure, and the residue was purified by chromatography on a silica gel column. The title compound (3.9 g, 69%) was obtained by eluting with a mixed solvent of hexane and ethyl acetate (1:1 v/v).
  • 2,6-Dichlorothieno[3,2-b]pyrazin-3-amine (819 mg, 3.8 mmol) obtained from the above step 4) was dissolved in methanol (15 ml), sodium methoxide (30 wt%; 7.06 ml, 38 mmol) was added, and the mixture was stirred at 90 °C for 1 hour. Methanol was removed, water was added, and the mixture was extracted three times with ethyl acetate. The organic layer was recovered, dried over magnesium sulfate, concentrated under reduced pressure, and the residue was purified by chromatography on a silica gel column. The title compound (492 mg, 60%) was obtained by eluting with a mixed solvent of hexane and ethyl acetate (5:1 v/v).
  • 6-Chloro-2-methoxythieno[3,2-b]pyrazin-3-amine (431 mg, 2 mmol) obtained from the above step 5) was dissolved in ethanol (15 ml), and palladium charcoal (851 mg, 2 mmol) and ammonium formate (1.51 g, 24 mmol) were added, and the mixture was reacted at 100 °C for 30 minutes using a microwave reactor.
  • the reaction solution was filtered through celite, and the solvent was removed. Water was added, and the mixture was extracted with ethyl acetate.
  • the organic layer was recovered, dried over magnesium sulfate, and concentrated under reduced pressure, and the residue was purified by chromatography on a silica gel column.
  • the title compound (315 mg, 87%) was obtained by eluting with a mixed solvent of hexane and ethyl acetate (5:1 v/v).
  • Paclitaxel, Docetaxel, Plinabulin, Combretastatin, Vincristine, Nocodazole, Vinblastine and Tirvanibulin shown in Table 1 below were used as comparative compounds.
  • Tubulin Polymerization Assay Kit (Cytoskeleton, #BK006P) was used. This is an analysis method that utilizes the principle that absorbance is measured according to the degree of light scattering in proportion to the concentration of microtubules formed by the binding between ⁇ -tubulin and ⁇ -tubulin recombinant proteins.
  • tubulin powder was dissolved in 1.1 ml of the mixed solution to prepare a 10 mg/ml tubulin stock solution, which was then stored at -70 °C.
  • Working solutions were prepared by diluting the stock solution for each tubulin inhibitor with general tubulin buffer. 10 ⁇ l of general tubulin buffer (control) and the tubulin inhibitor working solutions of the indicated concentrations were added to each well of the plate, and incubated at 37 °C for 2 minutes.
  • tubulin stock solution which had been stored frozen to prevent the polymerization reaction from proceeding, was thawed on ice, and diluted with TP buffer (tubulin polymerization buffer) to prepare a 3 mg/ml tubulin working solution.
  • TP buffer tubulin polymerization buffer
  • 100 ⁇ l of the tubulin working solution was added to each well of the plate at a time using a multi-pipette to ensure that the polymerization reaction proceeds simultaneously.
  • the absorbance change over time was measured at 340 nm using a SpectraMax iD3 Multi-Mode Microplate Reader (Molecular Devices).
  • Figure 1 shows the results of a comparative experiment to determine whether the compound of the present invention inhibits tubulin polymerization using paclitaxel and nocodazole.
  • Paclitaxel was used as a negative control as a microtubule stabilizer
  • nocodazole was used as a positive control as a microtubule destabilizer.
  • 1 ⁇ M HLS-22001 and HLS-22002 exhibited tubulin polymerization inhibition activity equivalent to that of 10 ⁇ M nocodazole, and treatment with 10 ⁇ M HLS-22001 and HLS-22002 exhibited significantly superior tubulin polymerization inhibition activity.
  • Figure 2 shows the results of a comparative experiment on the inhibition of tubulin polymerization using the compounds of the present invention with microtubule stabilizers (paclitaxel, docetaxel) and microtubule destabilizers (plinavulin, combretastatin, vincristine, nocodazole).
  • 10 ⁇ M HLS-22001 and HLS-22002 showed tubulin polymerization inhibition activity that was significantly superior to most comparative compounds, and at a similar level to combretastatin and vincristine.
  • Figure 3 shows the results of a comparative experiment of the compounds of the present invention with nocodazole, paclitaxel, and tirvanibuline.
  • 2 ⁇ M HLS-22001 and HLS-22002 showed better tubulin polymerization inhibition activity than 5 ⁇ M nocodazole and 2 ⁇ M tirvanibuline.
  • HT297.T cells are skin cells derived from actinic keratosis patients, and A431 cells are squamous cell carcinoma cells used to evaluate the efficacy of actinic keratosis.
  • nicodazole, and tirvanibuline After treating each cell line with the compound of the present invention, nicodazole, and tirvanibuline, the expression of microtubules was confirmed using immunofluorescence.
  • HT297.T cells (1 x 10 4 cells/well; ATCC #CRL-7782) and A431 cells (2 x 10 4 cells/well; KCB #80005 Passage 12) were seeded in 24-well Flat Bottom Microplate with Lid (Greiner, #662892), respectively, and cultured in a 37 °C, 5% CO 2 incubator for 16 to 24 h.
  • the compound working solution diluted with DMSO and the culture medium (HT297.T (Wellgene, DMEM; LM001-05), A431 (Wellgene, RPMI; LM011-51) containing 10% FBS (Wellgene, S001-01), 1% penicillin-streptomycin (Wellgene, LS202-02)) were mixed to prepare the mixtures of the indicated concentrations, added to each well containing the cultured cells, and incubated for 6 h in a 37 °C, 5% CO2 incubator. After washing twice or more with DPBS, 500 ⁇ l of 100% methanol was added to each well and exposed for 10 min at 30 rpm on a shaker.
  • Methanol was removed from the plate and washed three or more times with DPBS.
  • PBS containing 0.1% BSA was added to each well (500 ⁇ l) and exposed for 60 minutes at 30 rpm in a shaker.
  • Antibodies binding to the antigen ( ⁇ -tubulin) were diluted in PBS at the manufacturer's recommended ratio, added to each well (500 ⁇ l), and exposed overnight at 4°C in a shaker.
  • secondary antibodies (fluorescent) matching the host of the primary antibody were diluted in PBS, added to each well (500 ⁇ l), and reacted for 1 to 2 hours in a shaker.
  • HLS-22001 and HLS-22002 showed a concentration-dependent decrease in the tubulin region compared to the control group, and this microtubule formation inhibition effect was superior to the nocodazole or tirvanibulin treatment group at the same concentration.
  • both HLS-22001 and HLS-22002 showed a marked tubulin expression reduction effect.
  • a WST-8 assay was performed using HT297.T cells and A431 cells.
  • HT297.T cells (5 x 10 3 cells/well) or A431 cells (3 x 10 3 cells/well) and 90 ⁇ l of culture medium (HT297.T (Wellgene, DMEM; LM001-05), A431 (Wellgene, RPMI; LM011-51) containing 10% FBS (Wellgene, S001-01), 1% penicillin-streptomycin (Wellgene, LS202-02)) were added to each well of a 96-well Clear Flat Bottom TC-treated Culture microplate, and cultured for 16 to 24 h in a 37 °C, 5% CO 2 incubator.
  • the working solutions (2 ⁇ l) of compounds at various concentrations prepared by 1/3 serial dilution with DMSO and the culture medium (98 ⁇ l) were mixed to prepare the mixtures of the indicated concentrations, 10 ⁇ l was added to each well containing cells cultured the previous day, and the mixture was incubated in an incubator at 37 °C, 5% CO2 for 72 h.
  • the Chromo-CK reaction reagent Chromogen, Korea, #CH-10000
  • a WST-8 assay solution was added to each well, 10 ⁇ l, and the mixture was reacted in an incubator at 37 °C, 5% CO2 for 1 h.
  • the absorbance was measured at 450 nm using an ABS plus Microplate reader with Nanodrop 500 (Molecular devices, Califormia, USA).
  • the IC 50 values in HT297.T cells were 7.05 ⁇ M for 5-FU, 0.96 ⁇ M for nocodazole, and 0.38 ⁇ M for tirvanibulin, whereas HLS-22001 and HLS-22002 showed much lower values of 0.14 ⁇ M and 0.02 ⁇ M, respectively.
  • the IC 50 values of the compounds of the present invention were 0.05 ⁇ M for HLS-22001 and 0.02 ⁇ M for HLS-22002, which were significantly lower than those of other comparative compounds.
  • tubulin inhibitors inhibit cell division and exhibit the characteristics of G2/M cycle arrest during the cell cycle
  • flow cytometry analysis was performed using PI (Propidium iodide), a fluorescent substance, to confirm the cell cycle arrest effect of the compound of the present invention.
  • PI Propidium iodide
  • HT297.T cells (3 ⁇ 10 5 ) were seeded in 60 mm dishes and A431 cells (2 ⁇ 10 5 ) were seeded in 6-well plates, respectively, and cultured in an incubator at 37 °C, 5% CO 2 for 16 to 24 hours.
  • a mixture of 2 ⁇ l of working solutions of compounds at various concentrations diluted with DMSO and 1 ml of culture medium (HT297.T (Welgene, DMEM; LM001-05), A431 (Welgene, RPMI; LM011-51) containing 10% FBS (Welgene, S001-01), 1% penicillin-streptomycin (Welgene, LS202-02)) was prepared.
  • the mixture at the indicated concentration was added to each well and cultured for 48 hours (HT297.T cells) or 6 hours (A431 cells) in a 37°C, 5% CO2 incubator.
  • the cells were diluted with culture medium to a concentration of 1 x 10 5 to 1 x 10 7 cells/ml, and the cells were lysed into single cells by pipetting. 1 ml of ethanol was added to 1 x 10 6 cells, fixed at -20°C for 3 hours or more, and then the cells were washed with DPBS and centrifuged.
  • the ratio of G2/M phase cells accounted for about 20% of the total, but in the case of HLS-22001 and HLS-22002, it was about 40%, showing a similar pattern to the positive control group, nocodazole and tirvanibuline.
  • the G2/M percentage was different from that of HT297.T cells, it showed a similar pattern to the positive control group, nocodazole and tirvanibuline, when treated with HLS-22001 and HLS-22002.
  • cell distribution was analyzed using Annexin V protein, which specifically binds to PS (Phosphatidylserine) exposed to the outside of the cell membrane, which is a morphological characteristic of cell death.
  • PS Phosphatidylserine
  • HT297.T cells (3 ⁇ 10 5 ) were seeded in 60 mm dishes and A431 cells (2 ⁇ 10 5 ) were seeded in 6-well plates, respectively, and cultured in an incubator at 37 °C, 5% CO 2 for 16 to 24 hours.
  • a mixture of 2 ⁇ l of compound working solution diluted with DMSO and 1 ml of culture medium (HT297.T (Welgene, DMEM; LM001-05), A431 (Welgene, RPMI; LM011-51) containing 10% FBS (Welgene, S001-01), 1% penicillin-streptomycin (Welgene, LS202-02)) was prepared.
  • the mixture at the indicated concentration was added to each well and cultured in a 37°C, 5% CO 2 incubator for 72 hours (HT297.T cells) or 24 hours (A431 cells).
  • the cells were diluted with culture medium to a concentration of 1 x 10 5 to 1 x 10 7 cells/ml, and the cells were lysed into single cells by pipetting.
  • 1 x 10 4 to 1 x 10 6 cells and 100 ⁇ l of Muse TM Annexin V & Dead Cell Reaction Reagent (Millipore, #MCH100105) were reacted in a dark room at room temperature for 30 minutes.
  • Samples of stained cells were analyzed using a MUSE analyzer (Millipore, Burlington, #MUSE 0500-3115B), and the four quadrant cell populations were classified based on the presence or absence of staining in the whole cells into living cells (lower left), necrotic cells (upper left), early apoptotic cells (lower right), and late apoptotic cells (upper right).
  • MUSE analyzer Millipore, Burlington, #MUSE 0500-3115B
  • the number of cells in each quadrant was measured, and the number of apoptotic population was calculated using the following formula.
  • Apoptotic population (%) (number of cells stained with Annexin V/total number of cells) x 100
  • the rate of cell death increased in a concentration-dependent manner by HLS-22001 and HLS-22002 treatment in both HT297.T cells and A431 cells, and was confirmed to be even higher than that of nocodazole and tirvanibulin, which have the same mechanism.
  • A431 cells (2 x 10 5 ) were seeded into 60 mm dishes and cultured for 16 to 24 hours in a 37 °C, 5% CO 2 incubator.
  • a mixture of 2 ml of compound working solution diluted with DMSO and culture medium (RPMI (Wellgene, LM011-51) containing 10% FBS (Wellgene, S001-01), 1% penicillin-streptomycin (Wellgene, LS202-02) was prepared. After removing the supernatant of the cultured cells, the mixture at the indicated concentration was added to each well and cultured for 24 hours in a 37 °C, 5% CO 2 incubator.
  • cell lysis buffer (RIPA buffer, CST, #9806) was added and the reaction was performed on ice for 20 minutes. The supernatant was extracted through centrifugation, and the amount of protein was measured using the BCA quantitative method. The measured lysate was mixed with protein loading buffer, and 20 ⁇ g per sample was boiled at 95 °C for 5 minutes, and then the proteins were separated by size using SDS-PAGE. The separated proteins were transferred to a nitrocellulose membrane in a Trans-Blot Turbo Transfer System at 25 V, 2.5 mA for 8 minutes, placed in 5% blocking buffer, and reacted for 1 hour in a RT rocker.
  • the primary antibody diluted 1:1000 was reacted overnight in a 4 °C rocker.
  • the membrane reacted with the primary antibody was washed using TBS-T, and then the secondary antibody diluted 1:3000 in 1% blocking buffer was reacted for 1 hour in a RT rocker.
  • Clarity Western Peroxide Reagent and Clarity Western Luminol Reagent were mixed 1:1 and reacted on the membrane for 5 minutes.
  • the amount of specific proteins was confirmed by chemiluminescence using ChemiDoc Imaging System. (Cleaved PARP, cleaved caspase-3: Apoptosis marker / ⁇ -actin: housekeeping gene)
  • the amount of cleaved PAPP and cleaved caspase-3 proteins increased in a concentration-dependent manner by HLS-22001 and HLS-22002 treatment in A431 cells, and was confirmed at a higher rate than nocodazole and tirvanibulin, which have the same mechanism.
  • THP-1 cells are human monocytes derived from patients with acute monocytic leukemia, and can induce macrophage differentiation by PMA (Phorbol 12-Myristate 13-Acetate) treatment, and are therefore widely used in the study of monocyte and macrophage functions.
  • PMA Phorbol 12-Myristate 13-Acetate
  • Each well of a 96-well Clear Flat Bottom TC-treated Culture microplate was filled with 90 ⁇ l of culture medium (RPMI medium (Wellgene; LM011-51) containing 10% FBS (Wellgene, S001-01), 1% penicillin-streptomycin (Wellgene, LS202-02)) containing THP-1 cells (ATCC #TIB-202) at 2.5 x 10 4 cells/well, treated with PMA (10 ng/ml or 50 ng/ml), and stimulated for 16 to 24 h in a 37 °C, 5% CO 2 incubator.
  • RPMI medium Wellgene; LM011-51
  • FBS Wellgene, S001-01
  • penicillin-streptomycin Wellgene, LS202-02
  • THP-1 cells ATCC #TIB-202
  • HLS-22001 working solutions (2 ⁇ l) diluted with DMSO were mixed with culture medium (98 ⁇ l) to prepare a mixture of 5 nM to 1000 nM, and added to each well containing cultured cells.
  • 10 ⁇ l was added to each well and incubated for 48 hours in a 37 °C, 5% CO 2 incubator.
  • 10 ⁇ l of Chromo-CK reaction reagent Chromogen, Korea, #CH-10000
  • WST-8 assay solution was added to each well and incubated for 1 hour in a 37 °C, 5% CO 2 incubator. Afterwards, the absorbance was measured at 450 nm using an ABS plus Microplate Reader with Nanodrop 500 (Molecular devices, California, USA).
  • IL-1 ⁇ proinflammatory cytokines
  • IL-6 proinflammatory cytokines
  • TNF- ⁇ proinflammatory cytokines
  • IL-1 ⁇ is one of the proinflammatory cytokines that promotes inflammatory responses, and is expressed at high levels in various inflammatory diseases, causing inflammatory responses by inducing NF- ⁇ B signaling pathway activity.
  • IL-6 is also a proinflammatory cytokine that transmits signals to the JAK/STAT pathway associated with various chronic inflammatory diseases, and excessive IL-6 production is associated with inflammatory immune-mediated diseases.
  • TNF- ⁇ is a proinflammatory cytokine involved in abnormal immune responses, and performs a wide range of immune-regulating functions, and is actively used as a target for numerous disease therapeutics associated with inflammatory responses.
  • THP-1 cells were first seeded in 60 mm dishes at a density of 2 ⁇ 10 5 cells/ml and treated with 10 ng/ml PMA, and then stimulated and cultured in a 37 °C, 5% CO 2 incubator for 24 h. After treatment with 0.5 ⁇ g/ml LPS and HLS-22001 (5 nM or 10 nM) to induce an inflammatory response for 3 h, cells were harvested. RNA was extracted from the harvested cells using an RNeasy mini kit (Quiagen, Hilden, NRW, Germany, #74104), and cDNA was synthesized using AccuPower Cyclescript RT master mix (Bioneer, Daejeon, Korea, #K-2051).
  • mRNA expression levels of IL-1 ⁇ and IL-6 were measured from the CT values obtained by performing real-time RT-PCR using Advanced Universal SYBR Green Supermix (BIORAD, Hercules, CA, USA, #1725271) and a CFX96 Touch Real-Time PCR Detection System (BIORAD, Hercules, CA, USA).
  • the compound of the present invention has an anti-inflammatory effect by inhibiting the mRNA expression of pro-inflammatory cytokines.
  • THP-1 cells were seeded at a density of 2 ⁇ 10 5 cells/ml in 60 mm dishes and treated with 10 ng/ml PMA, and then stimulated and cultured in a 37 °C, 5% CO 2 incubator for 24 h. After treatment with 0.5 ⁇ g/ml LPS and HLS-22001 (5 nM or 10 nM), which induce an inflammatory response, for 24 h, the cell supernatant was collected.
  • the levels of IL-1 ⁇ and TNF- ⁇ secreted in the supernatant were quantified by measuring the absorbance at 450 nm using Human IL-1 beta Duoset ELISA (RnDSystems, Minneapolis, USA, #DY201) and Human TNF-alpha DuoSet ELISA (#DY210) kits, respectively, with an ABS plus Microplate Reader with Nanodrop 500 (Molecular devices, California, USA).
  • the compound of the present invention has an anti-inflammatory effect by effectively inhibiting the secretion of proinflammatory cytokines.

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Abstract

La présente invention concerne une composition pharmaceutique comprenant comme principe actif un composé présentant une activité inhibitrice de la tubuline ou un sel pharmaceutiquement acceptable de celui-ci. La composition pharmaceutique de la présente invention inhibe la polymérisation de la tubuline et peut ainsi être utilisée très efficacement, en particulier pour prévenir ou traiter des maladies inflammatoires, le syndrome de détresse respiratoire aiguë, la kératose actinique ou des maladies associées à l'angiogenèse.
PCT/KR2024/004107 2023-03-30 2024-03-29 Composition pharmaceutique présentant une activité inhibitrice de la tubuline WO2024205336A1 (fr)

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Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2014051698A1 (fr) * 2012-09-28 2014-04-03 Vanderbilt University Composés hétérocycliques condensés comme inhibiteurs sélectifs de bmp
KR20160070005A (ko) * 2014-12-08 2016-06-17 동국대학교 산학협력단 신규한 4-(아릴)-N-(2-알콕시티에노[3,2-b]피라진-3-일)-피페라진-1-카복스아미드 유도체 및 이의 항증식 효과
WO2017139779A1 (fr) * 2016-02-12 2017-08-17 Forma Therapeutics, Inc. Thiénopyrazine carboxamides en tant qu'inhibiteurs de protéase spécifique de l'ubiquitine
WO2018136634A1 (fr) * 2017-01-18 2018-07-26 Vanderbilt University Composés hétérocycliques fusionnés en tant qu'inhibiteurs sélectifs de protéine morphogénétique osseuse (bmp)
WO2018234342A1 (fr) * 2017-06-21 2018-12-27 F. Hoffmann-La Roche Ag Dérivés d'isoindolinone utilisés en tant que modulateurs d'irak4

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2014051698A1 (fr) * 2012-09-28 2014-04-03 Vanderbilt University Composés hétérocycliques condensés comme inhibiteurs sélectifs de bmp
KR20160070005A (ko) * 2014-12-08 2016-06-17 동국대학교 산학협력단 신규한 4-(아릴)-N-(2-알콕시티에노[3,2-b]피라진-3-일)-피페라진-1-카복스아미드 유도체 및 이의 항증식 효과
WO2017139779A1 (fr) * 2016-02-12 2017-08-17 Forma Therapeutics, Inc. Thiénopyrazine carboxamides en tant qu'inhibiteurs de protéase spécifique de l'ubiquitine
WO2018136634A1 (fr) * 2017-01-18 2018-07-26 Vanderbilt University Composés hétérocycliques fusionnés en tant qu'inhibiteurs sélectifs de protéine morphogénétique osseuse (bmp)
WO2018234342A1 (fr) * 2017-06-21 2018-12-27 F. Hoffmann-La Roche Ag Dérivés d'isoindolinone utilisés en tant que modulateurs d'irak4

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