2H-Indazole derivatives as therapeutic agents for brain cancer and brain metastasis

Abstract

The present disclosure provides a method for treating brain metastasis of brain cancer or other cancers associated with CDK4 and / or CDK6 activity or preventing brain metastasis, wherein the method comprises administering a therapeutically effective amount of a compound of formula (I) to a subject in need thereof. Also disclosed is the use of a compound of formula (I) in the preparation of a medicament for treating brain metastasis of brain cancer or other cancers associated with CDK4 and / or CDK6 activity or preventing brain metastasis.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority under 35 USC §119(e) to U.S. Provisional Application Serial No. 62 / 798,220, filed on January 29, 2019, the disclosure of which is incorporated herein by reference in its entirety. Technical Field

[0003] The present application relates to methods for treating brain cancer and brain metastasis using 2H-indazole derivatives and compositions thereof. Background Art

[0004] Cyclin-dependent kinases are a family of protein kinases that regulate cell division and proliferation. The cell cycle process is controlled by cyclins and their associated cyclin-dependent kinases (such as CDK1-CDK4 and CDK6), while other CDKs (such as CDK7-CDK9) are essential for transcription. CDK binds to cyclins to form a heterodimer complex, phosphorylating substrates on serine and threonine residues, which in turn initiates events required for cell cycle transcription and progression (Malumbres et al., Trends Biochem. Sci. 2005, 30, 630-641). Since uncontrolled cell proliferation is a hallmark of cancer, and most cancer cells show CDKs dysregulation, inhibition of CDKs has become a potential treatment for various cancers. Inhibitors with varying degrees of selectivity for CDKs have been reported. Due to the key role of CDK4 / 6 in regulating cell proliferation and the toxic effects associated with other CDKs inhibition, selective CDK4 / 6 inhibitors are currently considered as a promising class of potential cancer therapeutic agents.

[0005] Abemaciclib, palbociclib, and ribociclib are CDK4 / 6 inhibitors that have recently been approved for the treatment of HR + / HER2 - Breast cancer.

[0006]

[0007]

[0008] However, these drugs have not shown good blood-brain barrier (BBB) ​​permeability in preclinical pharmacokinetic (PK) and efficacy models. See, for example, Raub, TJ et al., Drug Metab. Dispos. 2015, 43, 1360–1371. In addition, both palbociclib and abemaciclib are p-glycoprotein (P-gp) substrates, which is a very undesirable property for a potential CNS drug and it may hinder its development for brain diseases.

[0009] Brain metastasis (or "secondary brain tumor") occurs when cancer cells spread from the original diseased organ in the body to the brain. This can occur with any cancer, but is more common with lung cancer, breast cancer, colon cancer, kidney cancer, and melanoma. According to the literature, approximately 24-45% of cancer patients in the United States develop brain metastasis (see https: / / emedicine.medscape.com / article / 1157902-overview ), 10% to 30% of adult cancer patients develop brain metastases (see https: / / www.mayoclinic.org / diseases-conditions / brain-metastases / symptoms-causes / syc-20350136). Brain metastases put pressure on the surrounding brain tissue and can cause a variety of signs and symptoms, including severe pain. Treating brain metastases not only helps extend the life of cancer patients, but also helps relieve pain and other symptoms, thereby improving the patient's quality of life.

[0010] Therefore, developing a CDK4 / 6 inhibitor with high BBB permeability is clearly an unmet medical need. Summary of the invention

[0011] The present invention is based on the surprising discovery that indazole compounds of formula (I) are potent, selective CDK4 / 6 inhibitors with good blood-brain barrier (BBB) ​​permeability. Therefore, these compounds are effective therapeutic agents for treating or preventing brain metastases of brain cancer and various other cancers.

[0012] In one aspect, the present invention provides a method of treating brain cancer or brain metastasis in a subject, the method comprising administering a therapeutically effective amount of a compound of formula (I):

[0013]

[0014] or a pharmaceutically acceptable salt, solvate or prodrug thereof, wherein:

[0015] R 1 It is hydrogen, C 1 -C 8 Alkyl, C 3 -C 7Cycloalkyl, R 6 C(O)- or R 7 O(CO)-;

[0016] R 2 and R 3 are independently hydrogen, C 1 -C 8 Alkyl, C 3 -C 7 Cycloalkyl or C 3 -C 7 Cycloalkylmethyl;

[0017] R 4 For hydrogen, halogen, C 1 -C 8 Alkyl or C 3 -C 7 Cycloalkyl;

[0018] R 5 is hydrogen or a halogen;

[0019] R 6 is hydrogen, C 1 -C 8 Alkyl; or C 3 -C 7 Cycloalkyl; and

[0020] R 7 It is C 1 -C 8 Alkyl; or C 3 -C 7 Cycloalkyl,

[0021] wherein any of the alkyl or cycloalkyl groups are optionally substituted.

[0022] In another aspect, the present invention provides the use of a compound of formula (I) in the preparation of a medicament for treating brain cancer or brain metastasis associated with CDK4 and / or CDK6 activity.

[0023] Compound 1, N-(5-((4-ethylpiperazin-1-yl)methyl)pyridin-2-yl)-5-fluoro-4-(3-isopropyl-2-methyl-2H-indazol-5-yl)pyrimidin-2-amine, is an example of a compound of formula (I), wherein R 1 Ethyl, R 2 is isopropyl, R 3 is methyl, R 4 is hydrogen, and R 5Compound 1 is a potent CDK4 / 6 selective inhibitor that can be used to treat or prevent diseases, disorders or medical conditions mediated by certain CDKs (particularly CDK4 and CDK6), such as various types of cancer and inflammation-related conditions. Brain cancer, such as glioblastoma, represents a therapeutic area in which CDK4 / 6 inhibitors are expected to have high efficacy potential.

[0024]

[0025] In particular, the present invention provides methods for treating brain metastases of various cancers, including but not limited to breast cancer, lung cancer, especially non-small cell lung cancer (NSCLC), colorectal cancer, prostate cancer, renal cancer, melanoma, mantle cell lymphoma (MCL), chronic myeloid leukemia (CML), acute myeloid leukemia (AML), etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 The efficacy of the abemaciclib / TMZ combination was shown. Dosage: TMZ, QD X 5; 6 mg / kg + abemaciclib, PO, QDX 21, 100 mg / kg.

[0027] Figure 2 The efficacy of the compound 1 / TMZ combination was shown. Dosage: TMZ: QD X 5; 6 mg / kg + compound 1, PO, QD X21, 100 mg / kg. DETAILED DESCRIPTION

[0028] One aspect of the present invention relates to a method for treating brain cancer or brain metastasis from other cancers, comprising administering to a subject in need thereof a therapeutically effective amount of a composition comprising a compound of formula (I):

[0029]

[0030] or a pharmaceutically acceptable salt, solvate or prodrug thereof, wherein:

[0031] R 1 It is hydrogen, C 1 -C 8 Alkyl, C 3 -C 7 Cycloalkyl, R 6 C(O)- or R 7 O(CO)-;

[0032] R 2 and R 3 are independently hydrogen, C 1 -C 8 Alkyl, C 3 -C 7Cycloalkyl or C 3 -C 7 Cycloalkylmethyl;

[0033] R 4 For hydrogen, halogen, C 1 -C 8 Alkyl or C 3 -C 7 Cycloalkyl;

[0034] R 5 is hydrogen or a halogen. 1 Can be C 1 -C 6 alkyl;

[0035] R 6 is hydrogen, C 1 -C 8 Alkyl; or C 3 -C 7 Cycloalkyl; and

[0036] R 7 It is C 1 -C 8 Alkyl; or C 3 -C 7 Cycloalkyl,

[0037] wherein any of the alkyl or cycloalkyl groups are optionally substituted.

[0038] In one embodiment, R 1 is hydrogen, methyl, ethyl, propyl or isopropyl.

[0039] In another embodiment, R 2 Can be C 1 -C 6 Alkyl, C 3 -C 6 Cycloalkyl or C 3 -C 6 Cycloalkylmethyl.

[0040] In another embodiment, R 2 It is methyl, ethyl, propyl, isopropyl, cyclopropyl, cyclopentyl, cyclopropylmethyl or cyclopentylmethyl.

[0041] In another embodiment, R 3 Can be C 1 -C 6 Alkyl or C 3 -C 6 Cycloalkyl.

[0042] In another embodiment, R 3 It is methyl, ethyl, propyl, isopropyl or cyclopropyl.

[0043] In another embodiment, R 4 is hydrogen or a halogen.

[0044] In another embodiment, R 5 It is hydrogen or fluorine.

[0045] In another embodiment, it is sometimes preferred that R 1 is methyl or ethyl; R 2 is isopropyl, cyclopropyl, cyclopropylmethyl or cyclopentyl; R 3 is methyl or ethyl; R 4 is hydrogen or fluorine; and R 5 It is hydrogen or fluorine.

[0046] In another embodiment, the invention encompasses any combination of the embodiments described herein.

[0047] Preferably, the brain cancer or metastatic cancer being treated expresses CDK4 and / or CDK6. Preferably, the brain cancer is glioblastoma.

[0048] Another aspect of the present invention relates to a method of treating brain cancer or brain metastases from other cancers, comprising administering to a subject in need thereof a therapeutically effective amount of a composition comprising a compound of the formula:

[0049]

[0050] Or a pharmaceutically acceptable salt, solvate or prodrug thereof. Preferably, the brain cancer or metastatic cancer being treated expresses CDK4 and / or CDK6. Preferably, the brain cancer is glioblastoma.

[0051] Another aspect of the present invention relates to the use of a compound of formula (I) or a pharmaceutically acceptable salt, solvate or prodrug thereof in the preparation of a medicament for treating brain cancer associated with CDK4 and / or CDK6 activity,

[0052]

[0053] in:

[0054] R 1 It is hydrogen, C 1 -C 8 Alkyl or C 3 -C 7 Cycloalkyl;

[0055] R 2 and R 3 are independently hydrogen, C 1 -C 8 Alkyl, C 3 -C7 Cycloalkyl or C 3 -C 7 Cycloalkylmethyl;

[0056] R 4 For hydrogen, halogen, C 1 -C 8 Alkyl or C 3 -C 7 Cycloalkyl; and

[0057] R 5 is hydrogen or a halogen.

[0058] In some embodiments, R 1 It is C 1 -C 6 Preferably, R 1 is hydrogen, methyl, ethyl, propyl or isopropyl.

[0059] In some embodiments, R 2 It is C 1 -C 6 Alkyl, C 3 -C 6 Cycloalkyl or C 3 -C 6 Preferably, R 2 It is methyl, ethyl, propyl, isopropyl, cyclopropyl, cyclopentyl, cyclopropylmethyl or cyclopentylmethyl.

[0060] In some embodiments, R 3 It is C 1 -C 6 Alkyl or C 3 -C 6 Preferably, R 3 It is methyl, ethyl, propyl, isopropyl or cyclopropyl.

[0061] In some embodiments, R 4 is hydrogen or a halogen.

[0062] In some embodiments, R 5 It is hydrogen or fluorine.

[0063] In some embodiments, sometimes more preferably, R 1 is methyl or ethyl; R 2 is isopropyl, cyclopropyl, cyclopropylmethyl or cyclopentyl; R 3 is methyl or ethyl; R 4 is hydrogen or fluorine; and R 5 It is hydrogen or fluorine.

[0064] In some preferred embodiments, and sometimes preferably, the brain cancer associated with CDK4 and / or CDK6 activity is a brain metastasis of glioblastoma or other cancers.

[0065] Another aspect of the present invention relates to the use of a compound of the formula: or a pharmaceutically acceptable salt, solvate or prodrug thereof in the preparation of a medicament for treating brain cancer or metastatic cancer (e.g., metastatic brain cancer) associated with CDK4 and / or CDK6 activity,

[0066]

[0067] Preferably, the brain cancer is glioblastoma.

[0068] In any of the above embodiments, cancers associated with CDK4 and / or CDK6 activity and causing brain metastasis include, but are not limited to, breast cancer, lung cancer (especially non-small cell lung cancer (NSCLC)), colorectal cancer, prostate cancer, renal cancer, melanoma, mantle cell lymphoma (MCL), chronic myeloid leukemia (CML), acute myeloid leukemia (AML), etc., and the method comprises administering to a cancer patient a therapeutically effective amount of a compound according to any embodiment disclosed herein.

[0069] In a preferred embodiment, the method is for the treatment of metastatic breast cancer.

[0070] In another preferred embodiment, the method is for the treatment of metastatic lung cancer, particularly metastatic non-small cell lung cancer.

[0071] In some embodiments, the present invention provides a method of using the compounds disclosed herein in a cancer patient for preventing brain metastasis, the spread of cancer cells from the original diseased organ.

[0072] In all embodiments, preferably, the brain cancer or brain metastasis is associated with the activity of CDK, in particular the activity of CDK4 or CDK6.

[0073] The present invention includes all possible combinations of any embodiments disclosed herein.

[0074] Unless otherwise indicated, the term "alkyl" as used herein is intended to include branched and straight chain saturated aliphatic hydrocarbon groups containing 1 to 8 carbons, preferably 1 to 6, and more preferably 1 to 4 carbons. The term includes, but is not limited to, methyl, ethyl, propyl, isopropyl, butyl, tert-butyl, isobutyl, pentyl, hexyl, and the like.

[0075] As used herein, unless otherwise indicated, the term "alkylene" refers to a divalent saturated aliphatic group derived from an alkane by removing two hydrogen atoms. Examples include, but are not limited to, methylene (-CH 2 -), ethylene (-CH2 CH 2 -), propylene (-CH 2 CH 2 CH 2 -)wait.

[0076] Unless otherwise indicated, the term "cycloalkyl" as used herein alone or as part of another group includes saturated cyclic hydrocarbon groups having 3 to 8, sometimes preferably 3 to 6, carbon atoms forming the ring. Examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0077]

[0046] As used herein, "halo" or "halogen" refers to fluorine (F), chlorine (Cl), bromine (Br) and iodine (I).

[0078] Furthermore, in any embodiment disclosed herein, alkyl, alkylene, cycloalkyl and cycloalkylmethyl may each be optionally substituted with one or more, preferably one to three, sometimes preferably one to two substituents independently selected from the group consisting of halogen, C 1 -C 4 Alkyl, OH, C 1 -C 4 Alkoxy and CN.

[0079] When any group is referred to as "optionally substituted", unless specifically defined, it means that the group is or is not substituted, provided that such substitution does not violate conventional bonding principles known to those of ordinary skill in the art. When the phrase "optionally substituted" is used before a group of groups, it means that each of the listed groups can be optionally substituted.

[0080] One of ordinary skill in the art will understand that for any molecule described as containing one or more substituents, only sterically practical and / or synthetically feasible compounds are intended to be included. Unless otherwise stated in this specification, when a variable is referred to as being optionally substituted or replaced by a substituent, it is understood that such substitution occurs by replacing a hydrogen covalently bonded to the variable with one of these substituents.

[0081] The compounds of the present invention are generally regarded as organic bases, which are capable of reacting with acids, particularly pharmaceutically acceptable acids, to form pharmaceutically acceptable salts.

[0082] The term "pharmaceutically acceptable salt" as used herein refers to salts that are suitable for contact with human and lower animal tissues without excessive toxicity, irritation, allergic reaction, etc., within the scope of reasonable medical judgment, and are commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. See, for example, SM Berge et al., J. Pharm. Sci., 1977, 66, 1-19, which is incorporated herein by reference. Pharmaceutically acceptable salts of the compounds of the present invention include those derived from suitable inorganic and organic acids. Examples of pharmaceutically acceptable non-toxic acid addition salts are amino salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid, or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, dodecylsulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, trimethylacetate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, etc. Preferred pharmaceutically acceptable salts include hydrochloride.

[0083] The term "solvate" as used herein refers to the physical association of a compound of the present invention with a stoichiometric or non-stoichiometric amount of solvent molecules. For example, one molecule of a compound is associated with one or more, preferably one to three, solvent molecules. It is also possible that multiple (e.g., 1.5 or 2) molecules of a compound share one solvent molecule. This physical association may include hydrogen bonding. In some cases, the solvate will be able to be isolated as a crystalline solid. The solvent molecules in the solvate may exist in a regular arrangement and / or a disordered arrangement. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, and isopropanolates. Solvation methods are well known in the art.

[0084] Although the compounds of formula (I) disclosed herein may themselves be in the form of "prodrugs", i.e., when R 1When it is an acyl group (i.e., RC(O)-) or an ester group (i.e., ROC(O)-), these "prodrugs" can be generated in vivo from other "prodrugs" under physiological conditions. Therefore, for these disclosed compounds, the term "prodrug" as used herein refers to a derivative of a compound that can be converted in vivo to produce the parent compound, for example, by hydrolysis in the blood. Common examples of prodrugs in the present invention include, but are not limited to, amide or phosphoramide forms of active amine compounds, such as compounds of formula (II):

[0085]

[0086] Where R 6 is an acyl group (e.g., acetyl, propionyl, formyl, etc.) or a phosphoryl group [e.g., -P(=O)(OH) 2 ] group; or optionally, when R 3 When is hydrogen, the corresponding amide or phosphoramide compound can be used as a prodrug. Such amide or phosphoramide prodrug compounds can be prepared according to conventional methods known in the art.

[0087] Although the therapeutically effective amount of the compound of the present invention, or its pharmaceutically acceptable salt or solvate can be administered as a chemical raw material for treatment, the active ingredient can be presented as a pharmaceutical composition. Therefore, the present disclosure further provides a pharmaceutical composition comprising any compound of the present invention or its pharmaceutically acceptable salt or solvate, and one or more, preferably one to three pharmaceutically acceptable carriers, diluents or other excipients. The carrier, diluent or other excipient must be acceptable in the sense of being compatible with the other ingredients of the formulation and harmless to the subject being treated.

[0088] As used herein, "pharmaceutically acceptable" refers to those properties of compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of patients without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio and effective for their intended use.

[0089] Pharmaceutical preparations can exist in unit dosage form, each unit dosage containing a predetermined amount of active ingredient. Representatively, the pharmaceutical composition of the present disclosure will be administered once every 1-5 days to about 1-5 times a day, or optionally, as continuous administration. This administration can be used as chronic or acute treatment. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will depend on the condition being treated, the severity of the condition, the time of administration, the route of administration, the excretion rate of the compound used, the duration of treatment, and the age, sex, weight and condition of the patient. Preferred unit dosage formulations are those containing daily doses or sub-doses of active ingredients, as described above, or a suitable portion thereof. Typically, at the beginning of treatment, the dosage is much lower than the optimal dose of the compound. Thereafter, the dosage is increased in small increments until the best effect is achieved in this case. Generally speaking, it is ideal to apply the drug at a concentration level without causing substantial harmful or harmful side effects.

[0090] When the compositions of the present disclosure comprise a combination of a compound of the present disclosure and one or more, preferably one or two, additional therapeutic or prophylactic agents, the compound and the additional agent(s) are typically present at dosage levels of about 10-150%, and more preferably about 10-80%, of the dose normally administered in a monotherapy regimen.

[0091] The pharmaceutical preparation can be suitably administered by any appropriate route, for example, by oral (including oral or sublingual), rectal, nasal, topical (including oral, sublingual or transdermal), vaginal or parenteral (including subcutaneous, intradermal, intramuscular, intraarticular, intrasynovial, intrasternal, intrathecal, intralesional, intravenous or intradermal injection or infusion) route. Such preparations can be prepared by any method known in the pharmaceutical field, for example, by combining the active ingredient with a carrier or excipient. Oral administration or injection is preferred.

[0092] Pharmaceutical formulations suitable for oral administration may be presented in discrete unit forms such as capsules or tablets; powders or granules; solutions or suspensions in aqueous or non-aqueous liquids; edible foams or desserts; or oil-in-water liquid emulsions or water-in-oil emulsions.

[0093] For example, for oral administration in tablet or capsule form, the active pharmaceutical ingredient can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, etc. Powders are prepared by pulverizing the compound to a suitable fine size and mixing it with a similarly pulverized pharmaceutical carrier (such as an edible carbohydrate, such as starch or mannitol). Flavoring agents, preservatives, dispersants and coloring agents may also be present.

[0094] As described above, capsules are made by preparing a powder mixture and filling a formed gelatin sheath. Glidants and lubricants, such as colloidal silicon dioxide, talc, magnesium stearate, calcium stearate or solid polyethylene glycol, may be added to the powder mixture prior to the filling operation. Disintegrants or solubilizers, such as agar, calcium carbonate or sodium carbonate, may also be added to improve the availability of the drug when the capsule is ingested.

[0095] In addition, when needed or necessary, suitable binders, lubricants, disintegrants and coloring agents can also be added to the mixture. Suitable binders include starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as gum arabic, tragacanth or sodium alginate, carboxymethyl cellulose, polyethylene glycol, etc. The lubricants used in these dosage forms include sodium oleate, sodium chloride, etc. Disintegrants include but are not limited to starch, methyl cellulose, agar, bentonite, xanthan gum, etc. The preparation of tablets, for example, by preparing a powder mixture, granulating or briquetting, adding lubricants and disintegrants, and pressing into tablets. The preparation method of the powder mixture is to mix the appropriately crushed compound with the above-mentioned diluent or matrix, and optionally with a binder such as carboxymethyl cellulose, alginate, gelling agent or polyvinyl pyrrolidone, a solution retardant such as paraffin, an absorption promoter such as a quaternary ammonium salt and / or an absorbent such as bentonite, kaolin or dicalcium phosphate. The powder mixture can be granulated by wetting with a binder (such as syrup, starch paste, Acadia mucus or a solution of cellulose or polymeric material) and pressing through a screen. As an alternative to granulation, the powder mixture can be run through a tablet press, with the result that incompletely formed lumps are broken into particles. The particles can be lubricated by adding stearic acid, stearate, talcum powder or mineral oil to prevent sticking to the tablet forming die. The lubricated mixture is then pressed into tablets. The compounds of the present disclosure can also be combined with a free-flowing inert carrier and directly pressed into tablets without the need for granulation or briquetting steps. A transparent or opaque protective layer consisting of a sealing coating of shellac, a coating of sugar or polymeric material and a polishing layer of wax can be provided. Dyes can be added to these coatings to distinguish different unit doses.

[0096] Oral liquids such as solutions, syrups and elixirs can be prepared in dosage unit form so that a certain amount of the compound contains a predetermined amount. Syrups can be prepared by dissolving the compound in an aqueous solution of appropriate flavor, while elixirs are prepared by using a non-toxic carrier. Solubilizers and emulsifiers such as ethoxylated isostearyl alcohol and polyoxyethylene sorbitol ether, preservatives, flavor additives such as peppermint oil or natural sweeteners, or saccharin or other artificial sweeteners, etc. can also be added.

[0097] Where appropriate, dosage unit formulations for oral administration can be microencapsulated. The formulation can also be prepared to prolong or sustain the release, for example, by coating or embedding particulate material in polymers, waxes or the like.

[0098] It should be understood that in addition to the ingredients specifically mentioned above, the formulations may include other conventional formulations discussed in the art regarding the type of formulation, for example, formulations suitable for oral administration may include flavoring agents.

[0099] The term "subject" or "patient" includes humans and other mammals, including but not limited to horses, dogs, cats, pigs, monkeys, etc., preferably humans.

[0100] The term "therapeutically effective amount" refers to an amount of a compound or composition that, when administered to a subject to treat a disease, is sufficient to achieve such treatment of the disease. The "therapeutically effective amount" may vary depending on the compound, the disease and its severity, and the age, weight or other factors of the subject to be treated. When applied to an individual active ingredient, the term refers to the ingredient itself. When applied to a composition, the term refers to the combined amount of the active ingredients that produces a therapeutic effect, whether administered in combination, serially or simultaneously.

[0101] In some embodiments, the term "treat" or "therapy" refers to: (i) inhibiting a disease, disorder, or condition, i.e., preventing its development; (ii) alleviating a disease, disorder, or condition, i.e., causing regression of the disease, disorder, and / or condition; or (iii) preventing a disease, disorder, or condition from occurring in a subject who may be susceptible to the disease, disorder, and / or condition but has not yet been diagnosed with the disease. Thus, in some embodiments, "treat" or "therapy" refers to ameliorating a disease or condition, which may include improving one or more physical parameters, although perhaps not perceptible to the subject being treated. In some embodiments, "treat" or "therapy" includes modulating a disease or condition, which may be physical (e.g., stabilizing perceptible symptoms) or physiological (e.g., stabilizing physical parameters) or both. In yet other embodiments, "treat" or "therapy" includes delaying the onset of a disease or condition.

[0102] The efficacy and comparison of Compound 1 and abemaciclib in combination with temozolomide (TMZ) in the treatment of orthotopic U87MG-luc human glioblastoma in mice were performed. In each study, TMZ was administered at 6 mg / kg, QD X 5PO, and Compound 1 or abemaciclib was administered at 100 mg / kg PO. Tumor growth was observed by bioluminescence. The abemaciclib / TMZ combination showed a reduction in tumor volume until day 42, followed by regrowth at day 49 ( Figure 1 In contrast, the Compound 1 / TMZ combination showed significant tumor volume reduction on day 28, and continued to reduce tumor volume on day 63 ( Figure 2 ). Given the comparable in vitro potency of Compound 1 and abemaciclib, the superior in vivo efficacy of Compound 1 relative to abemaciclib in the glioblastoma model can be attributed to the more favorable BBB permeability characteristics of Compound 1 relative to abemaciclib. Broadly speaking, the significant differences between Compound 1 and abemaciclib in brain disease models can be traced back to their different molecular structures.

[0103] The main difference between compound 1 and abemaciclib in molecular structure is that compound 1 contains a 2H-indazole nucleus, while abemaciclib contains a benzimidazole nucleus:

[0104]

[0105] Compared to

[0106]

[0107] This structural difference unexpectedly leads to a significant difference in BBB permeability profile between the two compounds. Example 3 describes an in vivo study in mice where the brain concentration of Compound 1 was observed to be approximately 3-fold higher than that of Abemaciclib, with a brain / plasma (B / P) ratio of 1.43 for Compound 1 and 0.43 for Abemaciclib (see Tables 1 and 2). In addition, it is noteworthy that Compound 1 is not a P-gp substrate (see Example 2).

[0108] Table 1. Brain concentration and B / P ratio of compound 1 in mice (10 mg / kg, po)

[0109]

[0110] Table 2. Brain concentrations and B / P ratios of abemaciclib in mice (10 mg / kg, po)

[0111]

[0112]

[0113] Although not intended to be limiting, illustrative, non-limiting examples of compounds that can be used in the present invention are listed in Table 3.

[0114] Table 3. Selected examples of compounds of formula (I)

[0115]

[0116]

[0117]

[0118]

[0119]

[0120]

[0121]

[0122]

[0123]

[0124]

[0125]

[0126]

[0127]

[0128]

[0129] Example

[0130] Example 1. In vivo efficacy studies in mice

[0131] Materials and methods

[0132] D-Luciferin (lot#0000204125) was obtained from Promega as a white powder and stored at -80°C in a covered box to reduce light exposure. Physiological saline was added to the D-Luciferin powder to generate a 15 mg / ml yellow transparent solution for in vivo imaging. D-Luciferin was prepared immediately before each bioluminescent imaging and stored on wet ice in the dark during use.

[0133] Temozolomide (99.0% parent, MW 194 g / mol, FW 194 g / mol, 99% purity, C 6 H 6 N 6 O 2 , lot#S123705) was obtained from SelleckChem as a pink fine powder. Upon receipt, store at -20°C in the dark. The compound was formulated in a vehicle of sterile water. The dosing formulation was vortexed to form a colorless, transparent solution with a pH of 6.3. The dosing solution was prepared once a week and stored at 4°C in the dark between treatments.

[0134] Compound 1 (92.8% parent, MW 489 g / mol, FW 525 g / mol, 99.7% purity, C 27 H 33 FN8 ·HCl, stored in a nitrogen-rich atmosphere at 4°C, protected from light. The compound was prepared in 10% ethanol, 10% The dosing formulation was prepared by first heating all vehicle components in a water bath set to approximately 42°C. Ethanol was first added to a sterile dose bottle containing pre-weighed BPI-1178 powder. The mixture was then vortexed to ensure that all powder was completely dissolved. Next, Add to the solution and vortex to mix. Finally, add normal saline and vortex the final mixture to form a colorless, transparent solution with a pH of 5.7. The dosing solution was prepared fresh every day.

[0135] Abemaciclib (83.7% parent, MW 506 g / mol, FW 603 g / mol, 99.6% purity, C 27 H 32 F 2 N 8 -H 3 CSO 3 H was obtained from Beta Pharma as a white flaky powder. Upon receipt, it was stored in a nitrogen-rich atmosphere at 4°C, protected from light. The compound was prepared in 10% ethanol, 10% The dosing formulation is prepared by first heating all the vehicle components in a water bath set to approximately 42°C. Ethanol is first added to a sterile dose bottle containing pre-weighed abacillin powder. The mixture is then vortexed to ensure that all powder is completely dissolved. Next, Add to the solution and vortex to mix. Finally, add normal saline and vortex the final mixture to form a colorless, transparent solution with a pH of 4.0. The dosing solution was prepared fresh every day.

[0136] Animals and husbandry

[0137] Female Envigo nude mice (Hsd: Athymic Nude-Foxn1nu) were used in this study. They were 6-7 weeks old on the first day of the experiment. The animals were fed irradiated Harlan 2918.15 mouse chow and water ad libitum. The animals were housed in In the clean room All treatments, body weights, and tumor measurements were performed in the bubble environment in disposable ventilated cages lined with corn cob bedding. The environment was controlled within a temperature range of 70° ± 2°F and a humidity range of 30-70%.

[0138] All procedures complied with all laws, regulations, and guidelines of the National Institutes of Health (NIH) and were approved by the Animal Care and Use Committee of Molecular Imaging, Inc. Molecular Imaging, Inc. is an AAALAC-accredited institution.

[0139] Example 1A. Cell preparation

[0140] MG-Luc cells were obtained from ATCC. They were grown in Minimum Essential Medium (MEM) containing Earle's Salts modified with 1% 100 mM sodium pyruvate, 1% 100X NEAA (non-essential amino acids), 200 μg / mL G418, and supplemented with 10% non-heat-inactivated fetal bovine serum (FBS) and 1% 100X penicillin / streptomycin / L-glutamine (PSG). The growth environment was maintained at 37°C, 5% CO 2 atmosphere incubator. When the expansion is complete, the cells are trypsinized using a 0.25% trypsin-EDTA solution. After cell separation, trypsin is inactivated by dilution with complete growth medium, and any cell clumps are separated by pipetting. The cells are centrifuged at 200rcf for 8 minutes at 4°C, the supernatant is aspirated, and the pellet is resuspended in cold Dulbecco's phosphate-buffered saline (DPBS) by pipetting. The uniform cell suspension is diluted in a trypan blue solution and counted using a Luna automated cell counter. The cell suspension is centrifuged at 200rcf for 8 minutes at 4°C. The supernatant is aspirated and the cell pellet is resuspended in cold serum-free medium to generate a final concentration of 1.0E+08 cells / ml without trypan blue. The cell suspension is kept on wet ice during implantation. After implantation, an aliquot of the remaining cells is diluted with a trypan blue solution and counted to determine cell viability after implantation.

[0141]

[0142]

[0143] Example 1B. Intracranial Implantation

[0144] The experimental mice were implanted intracranially with 1.0E+06 cells per 10 μl on days 0, 1 and 2. For aseptic surgical implantation, mice were injected with 0.2 mg / kg buprenorphine and anesthetized with 2% isoflurane in air. The mice were then fixed in a stereotaxic frame (ASI instruments, Inc.) using non-breaking ear bars. Ophthalmic ointment was applied to the eyes of the mice to prevent dryness during surgery. During the implantation, a circulating water heating pad at 37°C was used to maintain the body temperature of the animals.

[0145] Once in the stereotaxic frame, the skull was alternately wiped with cotton swabs saturated with chlorhexidine solution and 70% ethanol to disinfect the skin surface and prepare for the incision. A 1 cm longitudinal incision was made in the center of the skull at the bregma using a No. 15 BD scalpel. The incision was retracted using small serrated spring forceps. The thin layer of connective tissue covering the skull surface was removed with a dry cotton swab under light pressure. Bleeding vessels were cauterized to prevent blood loss. A 0.9 mm drill bit was then placed in the center of the bregma, moved 2 mm to the right lateral, 1 mm anterior to the coronal suture, and the drill was lowered using the stereotaxic electrode manipulator arm to score the skull surface. The drill bit was removed from the stereotaxic frame, and the drilling through the skull to the dura mater surface was completed by hand.

[0146] The cell suspension (stored on wet ice) was mixed thoroughly and aspirated into a 50 μl gas-tight Hamilton syringe. A standard 27 g needle was filled with the cell suspension to eliminate air bubbles and the Luer tip of the syringe was inserted into the needle port. The syringe was secured to a custom syringe holder (ASI Instruments, Inc.) and attached to the stereotaxic frame manipulator arm. The syringe needle was aligned with the center of the drilled hole and lowered until the beveled tip was flush with the underside of the skull on the dura surface. The needle was then lowered 3 mm into the brain and retracted 1 mm to form a “reservoir” for depositing the cell suspension. 10 μl of the cell suspension (1x10 6 cells / mouse) were slowly injected into the brain tissue, and any minor leakage (typical of IC implants) was absorbed with a dry cotton swab.

[0147] After injection, the needle was withdrawn and the burr hole was immediately sealed with bone wax to minimize the loss of implanted cells. The skull surface was then cleaned alternately with dry cotton swabs and cotton swabs saturated with 70% ethanol to remove foreign cells and prevent the growth of extracranial tumors. The mouse was removed from the stereotaxic frame and the incision was closed using stainless steel wound clips. Once the mouse regained consciousness and dorsal position, it was returned to its cage. The mice were implanted between February 20 and 22, 2017.

[0148] Example 1C. Treatment

[0149] All mice were divided into study groups based on bioluminescent imaging (BLI) estimates of tumor burden. Mice were allocated to ensure that the mean tumor burden of all groups was within 10% of the overall mean tumor burden of the study population. Since implantation occurred over three days, day 0 was defined as the median implantation date (February 21, 2017). All groups started treatment on day 21, regardless of the initial implantation date.

[0150] Group 1: Vehicle control (10% EtOH, 10% 80% saline (0.9% NaCl), 0.2 mL / 20 g, PO, QD x 21 (Days 21-41)

[0151] Group 2: Temozolomide, 6 mg / kg, PO, QDx5 (Days 21-25)

[0152] Group 3: Compound 1, 100 mg / kg, PO, QDx21 (Days 21-41)

[0153] Group 4: Abemaciclib, 100 mg / kg, PO, QDx21 (Days 21-41)

[0154] Group 5: Temozolomide, 6 mg / kg, PO, QDx5 (Days 21-25) + Compound 1, 100 mg / kg, oral administration, QDx21 (Days 21-41)

[0155] Group 6: Temozolomide, 6 mg / kg, PO, QDx5 (Days 21-25) + Abemaciclib, 100 mg / kg, PO, QDx21 (Days 21-41)

[0156] Example 1D. In vivo bioluminescence imaging (BLI)

[0157] In vivo bioluminescence imaging (BLI) was performed using an IVIS Spectrum (Caliper Life Sciences, Hopkinton, MA). A maximum of 5 animals were imaged at a time under approximately 1-2% isoflurane gas anesthesia. Each mouse was injected subcutaneously with 150 mg / kg (15 mg / ml) D-luciferin and imaged in the prone position 10 minutes after injection. Large binning of the CCD chip was used, and the exposure time (2 seconds to 2 minutes) was adjusted to obtain at least hundreds of counts per image and avoid saturation of the CCD chip. BLI images were collected on days 21, 28, 35, 42, 49, 56, and 64.

[0158] Images were analyzed using Matlab R2015a software. Primary brain fixed volume ROIs were placed on prone images of each animal to estimate brain tumor burden. Total flux (photons / second) for all ROIs was calculated and exported to facilitate between-group analysis.

[0159] Example 1E. Evaluation of Side Effects

[0160] All animals were observed for clinical symptoms at least once a day. Animals were weighed on each day of treatment. Individual body weights were recorded 3 times a week.

[0161] Treatment-related weight loss greater than 20% is generally considered unacceptable toxicity. In this study, a dose level was described as tolerated if treatment-related weight loss (during treatment and 2 weeks post-treatment) was <20% and mortality was ≤10% during this period in the absence of a potentially lethal tumor burden.

[0162] Following death or euthanasia, all animals were dissected to provide a general assessment of potential causes of death and possible target organs of toxicity. The presence or absence of metastases was also recorded. Significant observations of clinical signs and dissecting findings were recorded, and toxicity findings were summarized for both individuals and groups.

[0163] Example 2. Cell permeability study of compound 1

[0164] Summarize

[0165] Test samples P-gp substrate classification 1 Negative

[0166] The test sample passed the fluorescein yellow monolayer integrity test standard (≤0.8×10 -6 cm / s).

[0167] Purpose

[0168] The aim of this study was to determine the P-gp substrate potential of a test sample using MDR1-MDCK monolayers.

[0169] Experimental procedures

[0170] MDR1-MDCK cell monolayers were grown to confluence on collagen-coated microporous membranes in 12-well assay plates. Detailed descriptions of the plates and their certification are provided below. The permeability assay buffer was Hanks balanced salt solution (HBSS) containing 10 mM HEPES and 15 mM glucose, pH 7.4. The buffer in the receiving compartment also contained 1% bovine serum albumin. The concentration of the dosing solution was 5 μM test sample + / - 1 μM valspodar in the assay buffer. The cells were first pre-incubated with HBSS containing + / - 1 μM valspodar for 30 minutes. The cell monolayers were dosed apically (AB) or basolaterally (BA) and incubated at 37°C and 5% CO 2 The cells were incubated in a humidified incubator at 4 °C. Samples were taken from the donor and receiver compartments at 120 min. Each assay was performed in duplicate. The flux of lucifer yellow per each monolayer was also measured after the experiment to ensure that no damage was caused to the cell monolayer during the flux. All samples were analyzed by LC-MS / MS using electrospray ionization. The analytical conditions are outlined in Appendix 1. The apparent permeability (P app ) and the recovery rate are calculated as follows:

[0171] P app =(dCr / dt)×V r / (A×C A ) (1)

[0172] Recovery rate = 100 × ((V r ×C r final )+(V d ×C d final )) / (V d ×C N ) (2)

[0173] in

[0174] dC r / dt is the slope of the cumulative concentration in the receiving compartment over time, in μM s -1 ;

[0175] V r is the volume of the receiving compartment in cm 3 ;

[0176] V d is the volume of the donor compartment in cm 3 ;

[0177] A is the area of ​​the insert (12 holes is 1.13 cm 2 );

[0178] C A is the average of the nominal dosing concentration and the measured 120-min dosing concentration, in μM;

[0179] C N is the nominal concentration of the dosing solution, in μM;

[0180] C r final is the cumulative received concentration in μM at the end of the culture period;

[0181] C d final is the donor concentration in μM at the end of the incubation period.

[0182] The outflow rate (ER) is defined as P app (BA) / P app (AB).

[0183] Cell batch quality control results

[0184]

[0185] Experimental Results

[0186]

[0187] P-gp substrate classification criteria:

[0188] ER ≥ 2.0 without valspodar and reduced by ≥ 50% with valspodar: positive

[0189] ER ≥ 2.0 without valspodar and reduced by < 50% with valspodar: negative

[0190] ER < 2.0 without and with valspodar: negative

[0191] Based on the above results, compound 1 is not a substrate of P-gp.

[0192] Analytical methods

[0193] Liquid chromatography

[0194] Column: Waters BEH phenyl column 30×2.1mm, 1.7μm

[0195] MP buffer: 25 mM ammonium formate buffer, pH 3.5

[0196] Aqueous solution container (A): 90% water, 10% buffer

[0197] Organic container (B): 90% acetonitrile, 10% buffer

[0198] Flow rate: 0.7mL / min

[0199] Gradient elution:

[0200] Time (minutes) ％A ％B 0.00 99 1 0.65 1 99 0.75 1 99 0.80 99 1 1.00 99 1

[0201] Total running time: 1.0min

[0202] Autosampler: 5 μL injection volume

[0203] Rinse 1: Water / methanol / 2-propanol: 1 / 1 / 1; containing 0.2% formic acid

[0204] Rinse 2: 0.1% formic acid in water

[0205] Mass spectrometry

[0206] Instrument: PE SCIEX API 4000

[0207] Interface: Turbine ion sprayer

[0208] Mode: Multiple Reaction Monitoring

[0209] Method: Last 1.0min

[0210] set up:

[0211] Test samples + / - Q1 Q3 DP EP CE CXP IS BPI-1178-7 + 489.4 375.3 12 10 28 12 5500

[0212] TEM: 500; CAD: 7; CUR: 30; GS1: 50; GS2: 50

[0213] Example 3. Brain concentrations and brain / plasma ratios in mice

[0214] Mice were treated with 10 mg / kg po. As shown in Tables 1 and 2, the brain concentration of Compound 1 was observed to be approximately 3 times higher than that of Abemaciclib, and the brain / plasma (B / P) ratio of Compound 1 was 1.43, while that of Abemaciclib was only 0.43.

[0215] The present disclosure provides some numerical ranges. It should be understood that, unless the context clearly specifies otherwise, each intermediate value between the upper and lower limits of the range and one-tenth of the lower limit unit is also specifically disclosed. Each smaller range between any specified value or intermediate value in the specified range and any other specified or intermediate value in the specified range is included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the range, and each range including any one, two or two limits in the smaller range is also included in the present invention, but any specific exclusion limit in the range must be observed. When the range includes one or two limits, the range excluding one or two of these included limits is also included in the present invention. The term "about" generally includes within plus or minus 10% of the indicated number. For example, "about 10%" can represent a range of 9% to 11%, and "about 20" can refer to 18 to 22. Preferably, "about" includes plus or minus 6% of the indicated value at most. Alternatively, "about" includes no more than plus or minus 5% of the indicated value. Other meanings of "about" may be apparent from the context, such as rounding, so that, for example, "about 1" may also mean 0.5 to 1.4.

[0216] For all purposes, all publications cited herein are incorporated by reference in their entirety. It should be understood that the embodiments described herein should be considered to be illustrative only, without limiting the scope of the invention. The description of the features or aspects in each embodiment should generally be considered to be other similar features or aspects that can be used in other embodiments.

[0217] While several embodiments have been described in the foregoing examples, it will be understood by those skilled in the art that various changes in form and details may be made without departing from the spirit and scope of the present disclosure as defined by the following claims.

Claims

1. Use of a compound of the following formula or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating brain metastasis of other cancers associated with CDK4 and / or CDK6 activity, or for preventing brain metastasis in subjects with other cancers, Wherein the other cancer is selected from the group consisting of breast cancer, lung cancer, colorectal cancer, prostate cancer, renal cancer, melanoma, mantle cell lymphoma, chronic myeloid leukemia and acute myeloid leukemia.

2. The use according to claim 1, wherein the compound or a pharmaceutically acceptable salt thereof is used together with a second therapeutic agent for the preparation of the medicament.

3. The use according to claim 2, wherein the second therapeutic agent is a different CDK inhibitor, HER2 inhibitor, mTOR inhibitor or EGFR inhibitor.

4. Use according to any one of claims 1 to 3, wherein the other cancer is non-small cell lung cancer.

Images