Selective CDK4 / 6 inhibitors for cancer treatment

Abstract

The present disclosure describes three classes of selective and potent CDK4 / 6 inhibitors that advantageously inhibit cancer growth even at low concentrations. The first class of anti-cancer CDK4 / 6 inhibitors is substituted pyrrolopyrimidine compounds bearing a fatty acid moiety. The second class includes substituted pyridopyrimidines bearing a fatty acid moiety. The third class includes substituted benzimidazole compounds bearing a fatty acid moiety. The compounds described herein can be used as pharmaceutical compounds for anti-cancer therapy and are useful for the treatment, prevention, and / or amelioration of cancer.

Description

[Technical field]

[0001] The present disclosure relates to pharmaceutical compounds for anti-cancer therapy, and more particularly to substituted pyrrolopyrimidine, pyridopyrimidine, and benzimidazole compounds that are potent CDK4 / 6 inhibitors useful in the treatment, prevention, and / or amelioration of cancer. [Background technology]

[0002] Cancer stem cells (CSCs) are tumor-initiating cells (TICs) that are resistant to conventional cancer therapies such as chemotherapy and radiotherapy. As such, CSCs are involved in both tumor recurrence and distant metastasis, which lead to treatment failure and poor clinical outcomes in cancer patients. Innovative approaches are therefore required to understand how to address the CSC problem. Mechanistically, this may relate to the ability of CSCs to survive and grow under harsh conditions and different microenvironments. As CSCs are a particularly small subpopulation of the tumor cell population, their metabolic and phenotypic properties have remained largely uncharacterized until recently.

[0003] Moreover, CSCs are extremely resilient and highly resistant to cellular stress, allowing for anchorage-independent growth, especially under low-attachment conditions. As such, CSCs form 3D spheroids that retain the properties of CSCs and stem cell precursors. In contrast, most "bulk" cancer cells die by anoikis, a type of apoptosis, when grown in suspension. Thus, clonal expansion of a single CSC results in the formation of 3D spheroids, but without cancer cell self-aggregation. Thus, the formation of 3D spheroids can functionally read out the stemness of epithelial cancer cells and enrich for a population of epithelioid cells with a stem-like phenotype. When prepared using breast cancer cells, such as MCF7, among others, these 3D spheroids are also called mammospheres.

[0004] Previously, 3D spheroids were generated from two ER(+) cell lines (MCF7, T47D) and subjected to unbiased, label-free proteomic analysis. This work has led to the characterization of the phenotypic behavior of CSCs at the molecular level. 3D spheroids were directly compared to monolayers of these cell lines and processed in parallel. This allowed us to identify proteomic signatures specific to the CSC phenotype in 3D spheroids compared to monolayers. Based on this molecular analysis, it was observed that mitochondrial proteins were significantly enriched in mammospheres. These mitochondria-associated proteins included molecules involved in β-oxidation, ketone metabolism and recycling, mitochondrial biogenesis, electron transport, ADP / ATP exchange and transport, CoQ synthesis, and ROS generation, as well as inhibition of mitophagy. Thus, increased mitochondrial protein synthesis or decreased mitophagy could lead to the accumulation of mitochondrial mass in CSCs.

[0005] Mitochondrial mass is increased in CSCs and is therefore considered as a new metabolic biomarker to purify CSCs. Using this overall approach, it has been observed that CSC activity could be significantly enhanced using only MitoTracker as a single marker in both ER(+) (MCF7) and ER(-) (MDA-MB-231) breast cancer cell lines. Notably, cells with high MitoTracker fluorescence intensity were found to be chemoresistant to paclitaxel and resistant to paclitaxel-induced DNA damage response.

[0006] However, there is a need for new pharmaceutical compounds for anti-cancer therapy that eradicate CSCs, prevent or reduce the likelihood of metastasis and / or recurrence, and reduce or eliminate cancer resistance to anti-cancer therapies such as chemotherapy. Furthermore, there is a need for therapeutic strategies and anti-cancer therapies that specifically target "optimal" CSCs and eliminate further cancer growth, such as anchorage-independent growth, tumor recurrence, and distant metastasis. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] International Publication No. 2016 / 166703 [Patent Document 2] International Publication No. 2016 / 193860 [Non-patent literature]

[0008] [Non-Patent Document 1] Comprehensive Organic Synthesis,Trost,Fleming,Pergamon:1991 [Non-Patent Document 2] Comprehensive organic Functional Group Transformations,Katritky,Meth-Cohn,Rees,Pergamon:1995 Summary of the Invention [Problem to be solved by the invention]

[0009] Currently, worldwide, cancer stem cells (CSCs) are considered one of the main causes of treatment failure in cancer patients. Mechanistically, this may be related to the ability of CSCs to survive and grow under harsh conditions and different microenvironments. We theorize that CSCs may acquire resistance to conventional therapies by utilizing elevated mitochondrial OXPHOS metabolism to "boost" ATP production. Consistent with this view, various mitochondrial inhibitors, including, among others, i) FDA-approved antibiotics (doxycycline, tigecycline, azithromycin, pyrvinium pamoate, atovaquone, and bedaquiline), ii) natural compounds (actinonin, CAPE, berberine, brutieridine, and melitidine), and iii) experimental compounds (oligomycin and AR-C155858, MCT1 / 2 inhibitors), successfully blocked the formation of 3D tumorspheres.

[0010] Cyclin-dependent kinases (CDKs) 4 and 6 are enzymes known to promote mitosis and meiosis in both normal and cancer cells. These enzymes cause the phosphorylation and subsequent inactivation of the retinoblastoma protein, which is involved in cell cycle progression from the G1 to S phase. Studies have identified abnormalities in which CDK activity is increased in cancer cells. This increased activity inactivates various tumor suppressor genes, thereby promoting rapid cancer stem cell proliferation and tumor growth. Naturally occurring CDK inhibitor proteins, such as p16 and p27, have been shown to inhibit the in vitro proliferation of lung cancer cell lines. Certain CDK inhibitors may be useful as chemoprotectants due to their ability to inhibit cell cycle progression in normal, non-transformed cells. [Means for solving the problem]

[0011] Targeted inhibition of these enzymes, alone or in combination with other therapies, is one of the promising strategies for anti-cancer treatment and therapeutic drugs. Inhibiting the CDK4 / 6 pathway prevents cells from progressing into S-phase, thereby resulting in cell death by apoptosis. Described herein are three classes of CDK inhibitors, primarily CDK4 and CDK6 ("CDK4 / 6") inhibitors, that are highly effective as cancer therapeutics. The first class of anti-cancer CDK4 / 6 inhibitors are substituted pyrrolopyrimidine compounds with a fatty acid moiety. The following formulas (n is an integer between 9 and 20, more preferably between 12 and 20) are illustrative of some embodiments of the first class of anti-cancer CDK4 / 6 inhibitors.

[0012] [ka]

[0013] The second class includes substituted pyridopyrimidines having a fatty acid moiety. The following formulas (where n is an integer from 9 to 20, more preferably 12 to 20) are illustrative embodiments of the second class of anticancer CDK4 / 6 inhibitors:

[0014] [ka]

[0015] The third class includes substituted benzimidazole compounds having a fatty acid moiety. The following formulas (wherein m is an integer of 0 to 4, more preferably 0 to 2, and n is an integer of 9 to 20, more preferably 12 to 20) are exemplary embodiments of the third class of anticancer CDK4 / 6 inhibitors:

[0016] [ka]

[0017] The first class, the second class, or the third class of compounds and their salts can be used as pharmaceutical compounds for cancer treatment.As will be understood by those skilled in the art, illustrative salts include succinate, trifluoroacetate, tartrate, and malate, among others.The present method also provides pharmaceutical preparations having a therapeutically effective amount of the first class, the second class, or the third class of compounds, or in some embodiments, one or more compounds of each class, or one or more therapeutically acceptable salts thereof, and pharmaceutically acceptable carriers, diluents, or excipients thereof.All of these forms are within the scope of the present method.Of course, pharmaceutically acceptable carriers known in the art can be used.

[0018] The compounds described herein may be used in combination with a method for treating cancer in a mammal, including a human, comprising administering to the mammal an amount of any of the first, second, or third class of compounds, or pharma- ceutically acceptable salts thereof, that is effective for treating such disease or condition.For example, this approach is useful for treating abnormal cell proliferation, such as cancer. The compounds described herein may be used to treat abnormal cell proliferation diseases and cancers, particularly those selected from the group consisting of cancers of the breast, ovary, cervix, prostate, testis, esophagus, stomach, skin, lung, bone, colon, pancreas, thyroid, biliary tract, oral vestibule and pharynx, lip, tongue, oral cavity, pharynx, small intestine, colorectal, large intestine, rectum, brain, and central nervous system, glioblastoma, neuroblastoma, keratoacanthoma, epidermoid carcinoma, large cell carcinoma, adenocarcinoma, adenocarcinoma, adenoma, adenocarcinoma, follicular carcinoma, undifferentiated carcinoma, papillary carcinoma, seminoma, melanoma, sarcoma, bladder cancer, liver cancer, renal cancer, myeloid diseases, lymphoid diseases, Hodgkin's lymphoma, hairy cell leukemia, and leukemia, by administering a therapeutically effective amount of a first class, second class, or third class compound, or a pharma- ceutically acceptable salt thereof, to a subject diagnosed with such cancer. In some embodiments, this approach may be used in combination with and / or to enhance the effectiveness of other therapies.

[0019] Some embodiments of this approach may take the form of compounds having the general formula:

[0020] [ka]

[0021] In the above formula, R 4 is hydrogen, C 1 -C 8 Alkyl, substituted C 1 -C 8 Alkyl, C 3 -C 8 Cycloalkyl, substituted C 3 -C 8Selected from the group consisting of cycloalkyl, aryl, substituted aryl, heteroaryl, and substituted heteroaryl; Z is CR z and R z is halo, hydrogen, C 1 -C 3 Alkyl, C 1 -C 3 Alkoxy, CN, C=NOH, C=NOCH 3 , C(O)H, C(O)C 1 -C 3 Alkyl, C 3 -C 8 Cycloalkyl, heterocyclyl, aryl, heteroaryl, substituted C 1 -C 3 Alkyl, substituted C 3 -C 8 Cycloalkyl, substituted heterocyclyl, substituted aryl, substituted heteroaryl, -B-NR a R b , -B-OR a , -BC(O)R a , -BC(O)OR a , -BC(O)NR a R a B is a bond, C 1 -C 3 Alkyl or branched C 1 -C 3 R is alkyl; a and R b are each independently hydrogen, C 1 -C 3 Alkyl, C 3 -C 8 selected from the group consisting of cycloalkyl, heterocyclyl, aryl, heteroaryl, substituted alkyl, substituted cycloalkyl, substituted heterocyclyl, substituted aryl, and substituted heteroaryl; and n is an integer from 9 to 20, preferably from 12 to 20. In some preferred embodiments, R 1 is cyclopentyl, R 2 is acetyl. Further, in some embodiments, n is preferably 12.

[0022] In some embodiments, the method may take the form of a pharmaceutical composition comprising a compound described herein as an active therapeutic agent, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier. For example, the composition may be a tablet having a core that, in some embodiments, comprises 35-55% by weight of the active therapeutic agent and a pharma- ceutically acceptable carrier. The pharma- ceutically acceptable carrier may be, for example, microcrystalline cellulose, crospovidone type A, low-substituted hydroxypropyl cellulose, magnesium stearate, and colloidal anhydrous silica.

[0023] The compounds and pharmaceutical compositions described herein have potency and selectivity for cancer stem cells, making them suitable for various anti-cancer therapeutic applications.For example, the method can take the form of a method for preventing or reducing the proliferation of at least one of cancer cells, cancer stem cells, and circulating tumor cells, in which a pharmacologic effective amount of the compounds or pharmaceutical compositions described herein is administered to a patient in need thereof.

[0024] The approach may take the form of a method of treating cancer, in which a pharma- ceutically effective amount of a compound or pharmaceutical composition described herein is administered to a patient in need thereof.

[0025] The approach may take the form of a method of treating or preventing metastatic disease, in which a pharma- ceutically effective amount of a compound or pharmaceutical composition described herein is administered to a patient in need thereof.

[0026] The approach may take the form of a method of treating or preventing tumor recurrence, in which a pharma- ceutically effective amount of a compound or pharmaceutical composition described herein is administered to a patient in need thereof.

[0027] The approach may take the form of a method for reducing cancer treatment resistance, such as chemotherapy resistance, comprising administering to a patient in need thereof a pharma- ceutical effective amount of a compound or pharmaceutical composition described herein.

[0028] The approach may take the form of a method for treating or preventing at least one of radiation therapy resistance, chemotherapy resistance, and hormone therapy resistance, in which a pharma- ceutical effective amount of a compound or pharmaceutical composition described herein is administered to a patient in need thereof.

[0029] Of course, one of ordinary skill in the art can determine the dosage amount, dosage form, and administration schedule for a particular embodiment, applying general methods known in the art.

[0030] The compounds of the present method may also be used in the manufacture of a medicament for a number of therapeutic applications, such as the treatment or prevention of cancer, the treatment or prevention of metastatic disease, and the treatment or prevention of tumor recurrence.

[0031] Those skilled in the art may recognize embodiments of the present approach upon review of the detailed description below. [Brief description of the drawings]

[0032] [Figure 1] FIG. 1 shows the dose-response curve of compound [1C] compared to its parent compound using a mammosphere formation assay in MCF7 cell line. [Diagram 2] FIG. 1 shows the dose response curve of compound [2C] compared to its parent compound using a mammosphere formation assay in MCF7 cell line. [Diagram 3] FIG. 1 shows the dose-response curve of the parent compound of compound [3C] using a mammosphere formation assay in MCF7 cell line. [Figure 4] FIG. 1 shows the dose response curve of compound [1C] compared to its parent compound using Hoechst staining assay in MCF7 cell line. [Diagram 5] FIG. 1 shows the dose response curve of compound [2C] compared to its parent compound using Hoechst staining assay in MCF7 cell line. [Figure 6] FIG. 1 shows the dose-response curve of the parent compound of compound [3C] using Hoechst staining assay in MCF7 cell line. [Figure 7]FIG. 1 shows the dose-response curve of compound [1C] compared to its parent compound using Hoechst staining assay in hTERT-BJ1 cell line. [Figure 8] FIG. 1 shows the dose-response curve of compound [2C] compared to its parent compound using Hoechst staining assay in hTERT-BJ1 cell line. [Figure 9] FIG. 1 shows the dose-response curve of the parent compound of compound [3C] using Hoechst staining assay in hTERT-BJ1 cell line. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0033] The following description includes currently contemplated aspects for carrying out exemplary embodiments of the present technique. The following description is not to be taken in a limiting sense, but is made merely for the purpose of illustrating the general principles of the invention.

[0034] In this approach, three classes of compounds of CDK4 / 6 inhibitors may be used as anti-cancer therapeutics. The first class includes substituted pyrrolopyrimidine compounds with a fatty acid moiety. The second class includes substituted pyridopyrimidine compounds with a fatty acid moiety. The third class includes substituted benzimidazole compounds with a fatty acid moiety. The compounds described herein have beneficial pharmaceutical properties and active ingredients. Many of these compounds exhibit significant selective CDK4 / 6 inhibitory activity and are therefore useful in the treatment of a wide variety of clinical conditions in which CDK4 / 6 kinase is abnormally elevated or in which CDK4 / 6 kinase is activated or present in normal amounts and activity, but in which inhibition of CDK is desirable to treat cell proliferation disorders. In particular, these compounds show promise as anti-cancer therapeutics. Definitions of each class of compound are provided below and are applicable to embodiments of the approach.

[0035] As used herein, the symbol C(O) represents a carbon-oxygen double bond. As used herein, the term "halo" means halogen and includes fluorine, chlorine, bromine, or iodine attached as understood in the art.

[0036] The term "alkyl" as used herein refers to saturated aliphatic groups including straight chain alkyl groups (e.g., methyl, ethyl, etc.), branched chain alkyl groups (isopropyl, tert-butyl, etc.), cycloalkyl (alicyclic) groups (cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl), alkyl substituted cycloalkyl groups, and cycloalkyl substituted alkyl groups. The term "alkyl" also includes alkenyl and alkynyl groups. The general formula is the term "C n "alkyl" (where n is an integer, for example, 1 to 20) can be used to indicate a particular alkyl group (straight or branched) of a particular range or number of carbons in the group. For example, the term C 1 -C 3 Alkyl includes, but is not limited to, methyl, ethyl, propyl, and isopropyl. 3-6 Cycloalkyl includes, but is not limited to, cyclopropyl, cyclopentyl, and cyclohexyl. Alkyl and cycloalkyl groups can be unsubstituted or substituted. Thus, the term "alkyl" includes both "unsubstituted alkyls" and "substituted alkyls," the latter of which refers to moieties having substituents replacing a hydrogen on one or more carbons of the hydrocarbon backbone.

[0037] The term "alkenyl" includes unsaturated aliphatic groups similar in length and possible substitution to the alkyls described above, but which contain at least one double bond. Alkenyl also includes "unsubstituted alkenyls" and "substituted alkenyls," the latter of which refers to moieties having substituents replacing a hydrogen on one or more carbons of the hydrocarbon backbone.

[0038] For example, the term "alkenyl" includes straight chain alkenyl groups (e.g., ethenyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, decenyl, etc.), branched chain alkenyl groups, cycloalkenyl (alicyclic) groups (cyclopropenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl), alkyl- or alkenyl-substituted cycloalkenyl groups, and cycloalkyl- or cycloalkenyl-substituted alkenyl groups. The term "alkenyl" further includes alkenyl groups which include oxygen, nitrogen, sulfur, or phosphorous atoms replacing one or more carbons of the hydrocarbon backbone. In certain embodiments, a straight chain or branched chain alkenyl group has six or fewer carbon atoms in its backbone (e.g., a straight chain is C 2 -C 6 , branched chain is C 3 -C 6 Similarly, cycloalkenyl groups may have from 3-8 carbon atoms in their ring structure, and more preferably have 5 or 6 carbon atoms in the ring structure. 2 -C 6 includes alkenyl groups having 2 to 6 carbon atoms.

[0039] The term "alkynyl" includes unsaturated aliphatic groups similar in length and possible substitution to the alkyls described above, but which contain at least one triple bond. Furthermore, the term "alkynyl" includes both "unsubstituted alkynyls" and "substituted alkynyls," the latter of which refers to alkynyl moieties having substituents replacing a hydrogen on one or more carbons of the hydrocarbon backbone.

[0040] By way of example, the term "alkynyl" includes straight chain alkynyl groups (e.g., ethynyl, propyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl, nonynyl, decynyl, etc.), branched chain alkynyl groups, and cycloalkyl or cycloalkenyl substituted alkynyl groups. The term "alkynyl" further includes alkynyl groups which include oxygen, nitrogen, sulfur or phosphorous atoms replacing one or more carbons of the hydrocarbon backbone. In certain embodiments, a straight chain or branched chain alkynyl group has 6 or fewer carbon atoms in its backbone (e.g., straight chain is C 2-C 6 , branched chain is C 3 -C 6 ). Term C 2 -C 6 includes alkynyl groups having 2 to 6 carbon atoms.

[0041] The term "substituted" is intended to denote a moiety having a substituent replacing a hydrogen on one or more atoms, such as the C, O, or N of a molecule. Such substituents include, for example, alkyl, alkoxy, alkenyl, alkynyl, halo, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, ... The radicals may include, but are not limited to, alkyl, aryl, arylcarbonyl, amino, carbamoyl, and ureido, amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, morpholino, phenol, benzyl, phenyl, piperizine, cyclopentane, cyclohexane, pyridine, 5H-tetrazole, triazole, piperidine, or aromatic or heterocyclic aromatic moieties, and combinations thereof.

[0042] The term "amine" or "amino" refers to any molecule, or moiety or functional group, as generally understood in the art, and may be primary, secondary, or tertiary. The term "amine" or "amino" includes compounds in which a nitrogen atom is covalently bonded to at least one carbon, hydrogen, or heteroatom. These terms include, but are not limited to, for example, "alkylamino," "arylamino," "diarylamino," "alkylarylamino," "alkylaminoaryl," "arylaminoalkyl," "alkaminoalkyl," "amide," "amido," and "aminocarbonyl." The term "alkylamino" includes groups and compounds in which the nitrogen is bonded to at least one additional alkyl group. The term "dialkylamino" includes groups in which the nitrogen atom is bonded to at least two additional alkyl groups. The terms "arylamino" and "diarylamino" include groups in which the nitrogen is bonded to at least one or two aryl groups, respectively. The terms "alkylarylamino," "alkylaminoaryl," or "arylaminoalkyl" refer to an amino group that is bonded to at least one alkyl group and at least one aryl group. The term "alkaminoalkyl" refers to an alkyl, alkenyl, or alkynyl group bound to a nitrogen atom which is also bound to an alkyl group.

[0043] The terms "amide", "amido", or "aminocarbonyl" include compounds or moieties which have a nitrogen atom bound to the carbon of a carbonyl or a thiocarbonyl group. These terms include "alkaminocarbonyl" or "alkylaminocarbonyl" groups which contain an alkyl, alkenyl, aryl or alkynyl group bound to an amino group which is bound to a carbonyl group. This includes arylaminocarbonyl and arylcarbonylamino groups which contain an aryl or heteroaryl moiety bound to an amino group which is bound to the carbon of a carbonyl or thiocarbonyl group. The term "amide" includes the terms "alkylaminocarbonyl", "alkenylaminocarbonyl", "alkynylaminocarbonyl", "arylaminocarbonyl", "alkylcarbonylamino", "alkenylcarbonylamino", "alkynylcarbonylamino", and "arylcarbonylamino". Amides also include urea groups (aminocarbonylamino) and carbamates (oxycarbonylamino).

[0044] The term "aryl" includes groups such as 5- and 6-membered monocyclic aromatic groups that may contain 0-4 heteroatoms, such as phenyl, pyrrole, furan, thiophene, thiazole, isothiazole, imidazole, triazole, tetrazole, pyrazole, oxazole, isoxazole, pyridine, pyrazine, pyridazine, and pyrimidine. In addition, the term "aryl" includes polycyclic, such as tricyclic, bicyclic, etc. aryl groups, such as naphthalene, benzoxazole, benzodioxazole, benzothiazole, benzimidazole, benzothiophene, methylenedioxyphenyl, quinoline, isoquinoline, anthryl, phenanthryl, naphtholidine, indole, benzofuran, purine, benzofuran, deazapurine, or indolizine. Those aryl groups having heteroatoms in the ring structure may also be referred to as "aryl heterocycles," "heterocycles," "heteroaryls," or "heteroaromatics." The aromatic ring may have at one or more ring positions such substituents as those mentioned above, e.g., alkyl, halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkylaminoacarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylthiocarbonyl, phosphate, phosphatase ... It can be substituted with suphonato, phosphinato, cyano, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an aromatic or heteroaromatic moiety.Aryl groups can also be fused or bridged with alicyclic or heterocyclic rings which are not aromatic so as to form a polycycle (eg, tetralin).

[0045] The term "heteroaryl" as used herein refers to a stable monocyclic or bicyclic ring having up to seven atoms in each ring, where at least one ring is aromatic and contains from 1 to 4 heteroatoms selected from the group consisting of O, N, and S. Heteroaryl groups within this definition include, but are not limited to, acridinyl, carbazolyl, cinnolinyl, quinoxalinyl, pyrazolyl, indolyl, benzotriazolyl, furanyl, thienyl, benzothienyl, benzofuranyl, quinolinyl, isoquinolinyl, oxazolyl, isoxazolyl, indolyl, pyrazinyl, pyridazinyl, pyridinyl, pyrimidinyl, pyrrolyl, and tetrahydroquinoline. As in the definition of heterocycle below, "heteroaryl" is also understood to include the N-oxide derivative of any nitrogen-containing heteroaryl. When a heteroaryl substituent is bicyclic and one ring is non-aromatic or does not contain a heteroatom, attachment is understood to be via the aromatic ring or the heteroatom-containing ring, respectively.

[0046] The term "heterocycle" or "heterocyclyl" as used herein is intended to mean a 5- to 10-membered aromatic or non-aromatic heterocycle containing 1-4 heteroatoms selected from the group consisting of O, N, and S, and includes bicyclic groups. Thus, "heterocyclyl" includes the heteroaryls mentioned above, as well as dihydro and tetrahydro analogs thereof. Other examples of "heterocyclyl" include benzimidazolyl, benzofuranyl, benzofurazanyl, benzopyrazolyl, benzotriazolyl, benzothiophenyl, benzoxazolyl, carbazolyl, carbolinyl, cinnolinyl, furanyl, imidazolyl, indolinyl, indolyl, indolazinyl, indazolyl, isobenzofuranyl, isoindolyl, isoquinolyl, isothiazolyl, isoxazolyl, naphthopyridinyl, oxadiazolyl, oxazolyl, oxazoline, isoxazoline, oxetanyl, pyranyl, pyrazinyl, pyrazolyl, pyridazinyl, pyridopyridinyl, pyridazinyl, pyridyl, pyrimidyl, pyrrolyl, quinazolinyl, quinolyl, quinoxalinyl, tetrahydropyranyl, tetrazolyl, tetrazolopyridyl, thiadiazolyl, thiazolyl, thienyl, triazolyl, azetidinyl, 1,4-dioxanyl, hexamethylcyclohexane ... Heterocyclyl groups include, but are not limited to, dihydroazepinyl, piperazinyl, piperidinyl, pyridin-2-onyl, pyrrolidinyl, morpholinyl, thiomorpholinyl, dihydrobenzimidazolyl, dihydrobenzofuranyl, dihydrobenzothiophenyl, dihydrobenzoxazolyl, dihydrofuranyl, dihydroimidazolyl, dihydroindolyl, dihydroisoxazolyl, dihydroisothiazolyl, dihydrooxadiazolyl, dihydrooxazolyl, dihydropyrazinyl, dihydropyrazolyl, dihydropyridinyl, dihydropyrimidinyl, dihydropyrrolyl, dihydroquinolinyl, dihydrotetrazolyl, dihydrothiadiazolyl, dihydrothiazolyl, dihydrothienyl, dihydrotriazolyl, dihydroazetidinyl, methylenedioxybenzoyl, tetrahydrofuranyl, and tetrahydrothienyl, and N-oxides thereof. The heterocyclyl substituent can be attached via a carbon atom or a heteroatom.

[0047] The term "acyl" includes compounds and moieties which contain the acyl group (CHCO-) ​​or a carbonyl group. The term "substituted acyl" includes compounds in which one or more hydrogen atoms have been replaced with, for example, an alkyl group, an alkynyl group, a halogen, a hydroxyl, an alkylcarbonyloxy, an arylcarbonyloxy, an alkoxycarbonyloxy, an aryloxycarbonyloxy, a carboxylate, an alkylcarbonyl, an arylcarbonyl, an alkoxycarbonyl, an aminocarbonyl, an alkylaminocarbonyl, a dialkylaminocarbonyl, an alkylthiocarbonyl, an alkoxyl, a phosphate, a phosphonato, a phosphinato, a cyano, an amino(alkylamino, dialkylamino) , arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or acyl groups replaced by an aromatic or heteroaromatic moiety.

[0048] The term "acylamino" includes moieties in which an acyl moiety is attached to an amino group. For example, this term includes alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido groups.

[0049] The term "alkoxy" includes substituted and unsubstituted alkyl, alkenyl, and alkynyl groups covalently linked to an oxygen atom. Examples of alkoxy groups include methoxy, ethoxy, isopropyloxy, propoxy, butoxy, and pentoxy groups, and can be cyclic groups such as cyclopentoxy. Examples of substituted alkoxy groups include halogenated alkoxy groups. Alkoxy groups can be substituted with alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, cyano, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or aromatic or heteroaromatic moieties. Examples of halogen substituted alkoxy groups include, but are not limited to, fluoromethoxy, difluoromethoxy, trifluoromethoxy, chloromethoxy, dichloromethoxy, trichloromethoxy, and the like.

[0050] The term "carbonyl" or "carboxy" includes compounds and moieties that contain a carbon atom double-bonded to an oxygen atom and their tautomeric forms. Examples of moieties that contain a carbonyl include aldehydes, ketones, carboxylic acids, amides, esters, anhydrides, and the like. The term "carboxy moiety" or "carbonyl moiety" refers to groups such as "alkylcarbonyl" groups in which an alkyl group is covalently bonded to a carbonyl group, "alkenylcarbonyl" groups in which an alkenyl group is covalently bonded to a carbonyl group, "alkynylcarbonyl" groups in which an alkynyl group is covalently bonded to a carbonyl group, and "arylcarbonyl" groups in which an aryl group is covalently bonded to a carbonyl group. Additionally, the term also refers to groups in which one or more heteroatoms are covalently bonded to the carbonyl moiety. For example, the term includes moieties such as aminocarbonyl moieties (where a nitrogen atom is bonded to the carbon of a carbonyl group, e.g., amides), aminocarbonyloxy moieties (where an oxygen atom and a nitrogen atom are both bonded to the carbon of a carbonyl group, e.g., also referred to as "carbamates"). Additionally, aminocarbonylamino groups (e.g., ureas) also include other combinations of heteroatoms (e.g., nitrogen, oxygen, sulfur, etc.) and carbonyl groups bonded to carbon atoms. Additionally, the heteroatoms can be further substituted with one or more alkyl, alkenyl, alkynyl, aryl, aralkyl, acyl, etc. moieties.

[0051] The term "thiocarbonyl" or "thiocarboxy" includes compounds and moieties which contain a carbon connected with a double bond to a sulfur atom. The term "thiocarbonyl moiety" includes moieties which are similar to a carbonyl moiety. For example, a "thiocarbonyl" moiety includes aminothiocarbonyl, where an amino group is bonded to the carbon atom of the thiocarbonyl group, and other thiocarbonyl moieties include oxythiocarbonyl (an oxygen atom bonded to a carbon atom), aminothiocarbonylamino, and the like.

[0052] The term "ether" includes compounds or moieties that contain an oxygen atom bonded to two different carbon atoms or heteroatoms. For example, this term includes "alkoxyalkyl," which refers to an alkyl, alkenyl, or alkynyl group covalently bonded to an oxygen atom that is covalently bonded to another alkyl group.

[0053] The term "ester" includes compounds and moieties which contain a carbon or heteroatom bound to an oxygen atom which is bonded to the carbon of a carbonyl group. The term "ester" includes alkoxycarboxy groups such as methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, butoxycarbonyl, pentoxycarbonyl, and the like. The alkyl, alkenyl, or alkynyl groups are as defined above.

[0054] The term "thioether" includes compounds and moieties that contain a sulfur atom bonded to two different carbon or heteroatoms. Examples of thioethers include, but are not limited to, alkthioalkyls, alkthioalkenyls, and alkthioalkynyls. The term "alkthioalkyls" includes compounds having an alkyl, alkenyl, or alkynyl group bonded to a sulfur atom that is bonded to an alkyl group. Similarly, the terms "alkthioalkenyls" and "alkthioalkynyls" refer to compounds or moieties in which an alkyl, alkenyl, or alkynyl group is bonded to a sulfur atom that is covalently bonded to an alkynyl group.

[0055] The term "hydroxy" or "hydroxyl" includes groups with an --OH or --O--.

[0056] The term "polycyclic" or "polycyclic group" includes moieties having two or more rings (e.g., cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, and / or heterocyclyl) in which two or more carbons are common to two adjacent rings, e.g., the rings are "fused rings." Rings that are connected through non-adjacent atoms are referred to as "bridged" rings. Each ring of a polycyclic compound may be substituted with, for example, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkoxycarbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, aminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, cyano, amino(alkylamino, dialkylamino, alkyl ... The alkyl group may be substituted with substituents as described above, such as alkylamino, arylamino, diarylamino, and alkylarylamino, acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl, and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkyl, alkylaryl, or aromatic or heteroaromatic moieties.

[0057] The term "heteroatom" refers to an atom of any element other than carbon or hydrogen. Preferred heteroatoms are nitrogen, oxygen, sulfur, and phosphorus.

[0058] Additionally, the phrase "any combination of these" means that any number of the listed functional groups and molecules may be combined to create a larger molecular architecture. For example, the terms "phenyl," "carbonyl" (or "=O"), "-O-," "-OH," and C 1-6 (i.e., -CH 3 and -CH 2 CH 2 CH2 -) can be combined to form a 3-methoxy-4-propoxybenzoic acid substituent. It is understood that when functional groups and molecules are combined to create larger molecular structures, hydrogens can be removed or added as necessary to satisfy the valence of each atom.

[0059] The compounds described herein include bonds between adjacent atoms and / or hydrogens, as necessary, to satisfy the valence of each atom, as would be understood by one of ordinary skill in the art. Bonds and / or hydrogen atoms are added, as necessary, to provide the following total number of bonds to each of the following types of atoms: carbon: 4 bonds; nitrogen: 3 bonds; oxygen: 2 bonds; and sulfur: 2-6 bonds.

[0060] The term "salt" of a compound relates to the corresponding salts prepared with an acid selected from the group consisting of inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, metaphosphoric acid, nitric acid, and sulfuric acid, and organic acids such as tartaric acid, acetic acid, trifluoroacetic acid, citric acid, malic acid, lactic acid, fumaric acid, benzoic acid, glycolic acid, gluconic acid, and succinic acid, and alkylsulfonic acids such as methanesulfonic acid, ethanesulfonic acid, ethane-1,2-disulfonic acid, and 2-hydroxyethanesulfonic acid, and arylsulfonic acids such as benzenesulfonic acid, 2-naphthalenesulfonic acid, p-toluenesulfonic acid, and naphthalene-1,5-disulfonic acid.

[0061] As used herein, the phrase "pharmacologically effective amount" refers to the amount required to be administered to a host, or to a cell, tissue, or organ of a host, to achieve a therapeutic result, such as the regulation, modulation, or inhibition of protein kinase activity, e.g., inhibition of protein kinase activity, or treatment of cancer. A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, a physician or veterinarian could start a dose of the compound of the present invention used in the pharmaceutical composition at a lower amount than is required to achieve the desired therapeutic effect, and gradually increase the dose until the desired effect is achieved.

[0062] The term "about" means that the value falls within the acceptable standard error of the mean value as considered by those skilled in the art. As expected, the meaning of "about" depends on the context in which it is used. In many cases, the term "about" may refer to ±5%, preferably ±2.5%, and more preferably ±1% of the value or range to which it refers. For example, in the context of weight fraction, the phrase "about 20%" may mean 20±5%, preferably 20±2.5%, and more preferably 20±1%.

[0063] The terms "treat", "treated", "treating" and "treatment" include the alleviation or alleviation of at least one symptom associated with or caused by the condition, disorder or disease being treated, particularly cancer. In certain embodiments, treatment includes alleviating and / or alleviating at least one symptom associated with or caused by the cancer being treated by the compounds of the present invention. For example, treatment can be the alleviation of one or more symptoms of cancer, or a cure for cancer.

[0064] The compounds described herein contain what this disclosure refers to as a fatty acid moiety. As used herein, a "fatty acid" is a carboxylic acid having an aliphatic chain, which may be saturated or unsaturated, although saturated chains are preferred. Examples of saturated fatty acids include lauric acid (CH 3 (CH 2 ) 10 COOH), palmitic acid (CH 3 (CH 2 ) 14 COOH), stearic acid (CH 3 (CH 2 ) 16 COOH), and myristic acid (CH 3 (CH 2 ) 12 COOH). Oleic acid (CH 3 (CH 2 ) 7 CH=CH(CH 2 ) 7COOH) is an example of a naturally occurring unsaturated fatty acid. Although reference may be made to salts or esters of fatty acids and their fatty amide moieties, for simplicity, these are included in the meaning of fatty acid moiety as used herein. For example, myristic acid may be referred to as myristate and oleic acid may be referred to as oleate. The fatty acid moiety may also be a carboacyl of a fatty acid, i.e., a group formed by the loss of a hydroxyl group from a carboxylic acid. In some embodiments, the fatty acid moiety may be attached to a therapeutic agent via an amide bond. As an example, a myristic acid conjugate may be attached to the fatty acid moiety CH 3 (CH 2 ) 12 It may have CO-NH-, with the tertiary nitrogen being attached to a therapeutic agent:

[0065] [ka]

[0066] In the above formula, n is an integer from 1 to 20, preferably from 10 to 20. This can occur when a myristate moiety is conjugated by myristoylation to give a tetradecanamide (or myristamide) group.

[0067] Substituted Pyrrolopyrimidine Compounds In some embodiments of this approach, the first class of anticancer CDK4 / 6 inhibitors are substituted pyrrolopyrimidine compounds, and pharma- ceutically acceptable salts thereof. It should be recognized that some compounds of the first class are derivatives of the parent compound 7-cyclopentyl-N,N-dimethyl-2-[(5-piperazin-1-ylpyridin-2-yl)amino]pyrrolo[2,3-d]pyrimidine-6-carboxamide, also known as ribociclib. Some embodiments of the first class have the chemical structure shown in the following general formula [1A], in which a fatty acid moiety is attached to piperazine.

[0068] [ka]

[0069] In the general formula [1A]: R 4 is hydrogen, C 1 -C 8 Alkyl, substituted C 1 -C 8 Alkyl, C 3 -C 8 Cycloalkyl, substituted C 3 -C 8 Selected from the group consisting of cycloalkyl, aryl, substituted aryl, heteroaryl, and substituted heteroaryl; Z is CR z and R z is halo, hydrogen, C 1 -C 3 Alkyl, C 1 -C 3 Alkoxy, CN, C=NOH, C=NOCH 3 , C(O)H, C(O)C 1 -C 3 Alkyl, C 3 -C 8 Cycloalkyl, heterocyclyl, aryl, heteroaryl, substituted C 1 -C 3 Alkyl, substituted C 3 -C 8 Cycloalkyl, substituted heterocyclyl, substituted aryl, substituted heteroaryl, -B-NR a R b , -B-OR a , -BC(O)R a , -BC(O)OR a , -BC(O)NR a R a B is a bond, C 1 -C 3 Alkyl or branched C 1 -C 3 R is alkyl; a and R b are each independently hydrogen, C 1 -C 3 Alkyl, C 3 -C 8selected from the group consisting of cycloalkyl, heterocyclyl, aryl, heteroaryl, substituted alkyl, substituted cycloalkyl, substituted heterocyclyl, substituted aryl, and substituted heteroaryl; and In the fatty acid portion, n represents an integer of 9-20, and preferably 12-20.

[0070] Of course, pharma- ceutically acceptable salts may also be used. As mentioned above, the salts may be prepared, for example, using an acid selected from inorganic acids, organic acids, alkylsulfonic acids, ethanesulfonic acids, and arylsulfonic acids. In some preferred embodiments of the present approach, the first class of anticancer CDK4 / 6 inhibitors are compounds having the general formula [1B] shown below. In such embodiments, R 4 is C 5 cycloalkyl, Z is dimethylcarboxamido or acetyl, and n represents an integer between 9 and 20, more preferably between 12 and 20. The parent compound of this general formula is ribociclib, an FDA approved drug used (in combination with an aromatase inhibitor) for the treatment of HR positive, HER2 negative advanced or metastatic breast cancer. However, in embodiments of the present approach according to formula [1A], the terminal piperazine is 11 -C 22 The fatty acid moiety is attached.Preferably, the fatty acid moiety is linear and saturated.In some preferred embodiments, the fatty acid moiety is one of lauric acid, myristic acid, palmitic acid, and stearic acid.The fatty acid moiety significantly improves the cellular uptake of the compound, and greatly improves the inhibition of cancer stem cell proliferation and the selectivity of the compound to tumor cells.

[0071] [ka]

[0072] An illustrative embodiment is shown below as compound [1C], where R 4 is unsubstituted C 5cycloalkyl, Z is dimethylcarboxamide, and n is 12. This results in the present embodiment having a 14 carbon fatty acid (i.e. myristate) moiety. The compound shown as compound [1C] was synthesized and compared to the known anti-cancer therapeutic ribociclib at concentrations between 1 and 100 μM in a mammosphere assay, demonstrating the incredible impact of the fatty acid moiety on the compound, with significantly improved inhibition of MCF7 cells. In preliminary laboratory evaluations, the present embodiment effectively inhibited 100% of cell proliferation at concentrations as low as 1 μM, demonstrating exceptional anti-cancer efficacy. For example, Figure 1 shows the dose-response curves for compound [1C] and its parent compound, revealing the improved CSC inhibition with the addition of the fatty acid moiety. Naturally, similar results are expected with other fatty acid moieties with as few as 11 carbons and as many as 22 carbons.

[0073] [ka]

[0074] In some embodiments of the present method, the first class of anticancer CDK4 / 6 inhibitors are substituted pyrrolopyrimidine compounds as shown in general formula [1D].Compared to general formula [1A], compounds having general formula [1D] contain fatty acid moiety at Z.Of course, pharmaceutically acceptable salts can also be used.

[0075] [ka]

[0076] In the general formula [1D]: R 4 is hydrogen, C 1 -C 8 Alkyl, substituted C 1 -C 8 Alkyl, C 3 -C 8 Cycloalkyl, substituted C 3 -C 8Selected from the group consisting of cycloalkyl, aryl, substituted aryl, heteroaryl, and substituted heteroaryl; R 3 is hydrogen, OH, C 1 -C 8 Alkyl, substituted C 1 -C 8 Alkyl, C 3 -C 8 Cycloalkyl, C(O)C 1 -C 8 Alkyl, C 1 -C 8 Haloalkyl, C 1 -C 8 Cyanoalkyl, C 1 -C 8 Alkyl-OH, SO 2 -C 1 -C 8 Alkyl, C 1 -C 8 Alkyl, C 3 -C 8 Cycloalkyl, and C 1 -C 8 alkoxy; R 3 When is not hydrogen, it may be substituted or unsubstituted; and In the fatty acid portion, n represents an integer of 9-20, and preferably 12-20.

[0077] In a preferred embodiment of the present approach, the first class of anti-cancer CDK4 / 6 inhibitors are compounds having the general formula [1E] shown below. In such an embodiment, R 4 is C 5 cycloalkyl, R 3 is hydrogen, and n is an integer of 9 to 20, more preferably 10 to 20, more preferably 10 to 16. The compound having the general formula [1E] is a derivative of ribociclib, in which a dimethylamino group is substituted for a fatty acid moiety in the carboxyl group.

[0078] [ka]

[0079] An illustrative embodiment, compound [1F], is shown below: 4 is unsubstituted C 5 cycloalkyl, R 3 is hydrogen and n is 12. This embodiment thus has a 14 carbon fatty acid (i.e., myristate) moiety. Compound [1F] is expected to show improved inhibition in the mammosphere assay.

[0080] [ka]

[0081] Table 1 below summarizes the results of various assays comparing compound [1C] with its parent compound (ribociclib), including in vitro biological data for both compounds. Compound [1C] showed a 7-fold improvement in potency (IC) compared to the parent compound in the 3D mammosphere assay. 50 0.2 μM and 1.5 μM), but retained similar activity in 2D cell viability assays (IC 50 2 μM for both compounds). This indicates that conjugation to a fatty acid moiety not only significantly improves potency in the 3D mammosphere assay. Comparing the selectivity index ("SI") for mammospheres and monolayers, it is found that conjugation also improves selectivity for mammospheres (SI of 10 vs. 1.3). Both compounds were non-toxic up to a concentration of 90 μM in the non-neoplastic cell line hTERT-BJ1 and showed high selectivity for tumor cell lines. Thus, the compounds produced by this approach have significantly improved potency and selectivity for tumor cells.

[0082] [Table 1]

[0083] "IC 50(MCF7 mammosphere)” refers to the half-maximal inhibitory concentration in a 3D mammosphere assay using MCF7, an ER+ breast cancer cell line. The term “IC 50 (MCF7 monolayer)" refers to the half-maximal inhibitory concentration in a 2D cell viability assay using MCF7, an ER+ breast cancer cell line. The term "IC 50 The term "IC (Mammosphere / Monolayer MCF7)" refers to the half-maximal inhibitory concentration in a 2D cell viability assay using hTERT-BJ1, an immortalized non-tumorous fibroblast cell line. The term "SI (Mammosphere / Monolayer MCF7)" refers to the IC comparing bioactivity in 3D and 2D assays against MCF7. 50 The term "SI (BJ1 / MCF7 monolayer)" refers to the mammosphere selection index, which is the ratio between the IC values ​​comparing the bioactivity in cell viability assays for MCF7 and hTERT-BJ1. 50 It refers to the cancer selectivity index, which is the ratio between values.

[0084] Substituted Pyridopyrimidine Compounds The second class includes substituted pyridopyrimidines with a fatty acid moiety, as shown in the following general formula [2A]. It should be recognized that some embodiments of the second class include derivatives of 6-acetyl-8-cyclopentyl-5-methyl-2-(5-piperazin-1-yl-pyridin-2-ylamino)-8H-pyrido[2,3-d]pyrimidin-7-one, also known as palbociclib. Of course, pharma- ceutically acceptable salts, such as those defined above, may also be used, as would be understood by one of ordinary skill in the art. Other examples of salts include malate, tartrate, bromide, hydrobromide dihydrate, hydrochloride, sulfate dihydrate, camsylate, napsylate, napsylate dihydrate, tosylate, citrate monohydrate, maleate, and oxalate.

[0085] [ka]

[0086] In the general formula [2A]: R 1 is hydrogen, aryl, C1 -C 8 Alkyl, C 1 -C 8 Alkoxy, C 3 -C 7 Cycloalkyl, or C 3 -C 7 is heterocyclyl; R 2 are independently hydrogen, halogen, C 1 -C 8 Alkyl, C 1 -C 8 Achill, C 3 -C 7 Cycloalkyl, C 1 -C 8 Alkoxy, C 1 -C 8 Alkoxyalkyl, C 1 -C 8 Haloalkyl, C 1 -C 8 Hydroxyalkyl, C 2 -C 8 Alkenyl, C 2 -C 8 Alkynyl, Nitrile, Nitro, OR 5 , S.R. 5 , N.R. 5 R 6 , N(O)R 5 R 6 , P(O)(OR 5 )(OR 6 ), (CR 5 R 6 ) m NR 7 R 8 , C.O.R. 5 , (CR 4 R 5 ) m C(O)R 7 , CO 2 R 5 ,CONR 5 R 6 , C(O)NR 5 SO 2 R 6 , N.R. 5 SO 2 R 6 , C(O)NR 5 OR 6, S(O) n R 5 , S.O. 2 NR 5 R 6 , P(O)(OR 5 )(OR 6 ), (CR 5 R 6 ) m P(O)(OR 7 )(OR 8 ), (CR 5 R 6 ) m -aryl, (CR 5 R 6 ) m -heteroaryl, and -CR 5 =CR 6 C(O)R 7 Selected from; R 3 are, in each case, independently hydrogen, halogen, C 1 -C 6 Alkyl, C 1 -C 6 Haloalkyl, C 1 -C 6 Hydroxyalkyl, or C 3 -C 7 is cycloalkyl; R 5 , R 6 , R 7 , and R 8 are independently hydrogen, C 1 -C 8 Alkyl, C 2 -C 8 Alkenyl, C 2 -C 8 alkynyl, arylalkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or heteroarylalkyl; m is 0 to 6; and In the fatty acid portion, n represents an integer of 9-20, and preferably 12-20.

[0087] The general formula [2B] shown below is a general formula of a second class of preferred embodiments according to this method. In the general formula [2B], R 1is unsubstituted C 5 cycloalkyl, R 2 is C 1 Acyl (acetyl) and R 3 is methyl, and n represents an integer of 9 to 20, preferably 12 to 20.

[0088] [ka]

[0089] An illustrative embodiment of the second class, compound [2C], is shown below: 1 is unsubstituted C 6 cycloalkyl, R 2 is C 1 Acyl (acetyl) and R 3 is methyl and n is 12. This results in this embodiment having a 14 carbon fatty acid (i.e. myristate) moiety. Compound [2C] was synthesized and showed significantly improved inhibition of MCF7 cells in a mammosphere assay when compared to the known anti-cancer therapeutic palbociclib at concentrations between 1 and 100 μM. These results also show that the second class of compounds from this approach has a significant beneficial effect on the anti-cancer efficacy of the compounds. For example, Figure 2 shows the dose-response curves for compound [2C] and its parent compound, revealing the improved CSC inhibition with the addition of the fatty acid moiety. Naturally, similar results would be expected with other fatty acid moieties with as few as 11 carbons and as many as 22 carbons.

[0090] [ka]

[0091] The second class also includes substituted pyridopyrimidines having a fatty acid moiety, as shown in the following general formula [2D]: As can be seen, the fatty acid moiety in formula [2D] is linked to a pyrido[2,3-d]pyrimidine, unlike the piperazine in formula [2A].

[0092] [ka]

[0093] In the general formula [2D]: R 1 is hydrogen, aryl, C 1 -C 8 Alkyl, C 1 -C 8 Alkoxy, C 3 -C 7 Cycloalkyl, or C 3 -C 7 is heterocyclyl; R 2 is hydrogen, halogen, C 1 -C 8 Alkyl, C 1 -C 8 Achill, C 3 -C 7 Cycloalkyl, C 1 -C 8 Alkoxy, C 1 -C 8 Alkoxyalkyl, C 1 -C 8 Haloalkyl, C 1 -C 8 Hydroxyalkyl, C 2 -C 8 Alkenyl, C 2 -C 8 Alkynyl, Nitrile, Nitro, OR 5 , S.R. 5 , N.R. 5 R 6 , N(O)R 5 R 6 , P(O)(OR 5 )(OR 6 ), (CR 5 R 6 ) m NR 7 R 8 , C.O.R. 5 , (CR 4 R 5 ) m C(O)R 7 , CO 2 R 5 ,CONR 5 R 6 , C(O)NR5 SO 2 R 6 , N.R. 5 SO 2 R 6 , C(O)NR 5 OR 6 , S(O) n R 5 , S.O. 2 NR 5 R 6 , P(O)(OR 5 )(OR 6 ), (CR 5 R 6 ) m P(O)(OR 7 )(OR 8 ), (CR 5 R 6 ) m -aryl, (CR 5 R 6 ) m -heteroaryl, and -CR 5 =CR 6 C(O)R 7 Selected from; R 3 are, in each case, independently hydrogen, halogen, C 1 -C 6 Alkyl, C 1 -C 6 Haloalkyl, C 1 -C 6 Hydroxyalkyl, or C 3 -C 7 is cycloalkyl; R 5 , R 6 , R 7 , and R 8 are independently hydrogen, C 1 -C 8 Alkyl, C 2 -C 8 Alkenyl, C 2 -C 8 selected from alkynyl, arylalkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or heteroarylalkyl; m is 0 to 6; and In the fatty acid portion, n represents an integer of 9-20, and preferably 12-20.

[0094] The following general formula [2E] is an example of another preferred embodiment of the second class according to this approach. In general formula [2E], R 1 is unsubstituted C 5 cycloalkyl, R 2 is H and R 3 is methyl, and n represents an integer of 9 to 20, preferably 12 to 20.

[0095] [ka]

[0096] An illustrative embodiment of the second class, compound [2F], is shown below: 1 is unsubstituted C 5 cycloalkyl (e.g., cyclopentyl), and R 2 is H and R 3 is methyl and n is 12. Thus, this embodiment has a 14 carbon fatty acid (i.e., myristate) moiety.

[0097] [ka]

[0098] Table 2 below summarizes the results of various assays comparing compound [2C] with its parent compound (palbociclib), including in vitro biological data for both compounds. The results show that compound [2C] was less potent in the 3D mammosphere assay compared to the parent compound palbociclib (IC 50 However, compound [2C] was non-toxic up to a concentration of 30 μM in the 2D cell viability assay, whereas palbociclib had an IC 50was approximately 0.1 μM. Furthermore, palbociclib was nonselective compared to 2D and 3D MCF7 assays. These results demonstrate that compound [2C] has improved compound selectivity for 3D mammospheres as opposed to normal cells. It should therefore be appreciated that the compounds of this approach may be used to selectively target cancer cells, particularly CSCs. Both compounds were nontoxic up to a concentration of 90 μM against the non-tumor cell line hTERT-BJ1 and showed high selectivity against tumor cell lines. Thus, the second class of compounds of this approach has improved selectivity for targeting cancer stem cells.

[0099] [Table 2]

[0100] Substituted benzimidazole compounds The third class includes substituted benzimidazole compounds having the general formula [3A] shown below. Of course, pharma- ceutically acceptable salts, such as those defined above, may also be used, as will be understood by those skilled in the art. It should be recognized that some embodiments of the third class include derivatives of N-[5-[(4-ethylpiperazin-1-yl)methyl]pyridin-2-yl]-5-fluoro-4-(7-fluoro-2-methyl-3-propan-2-ylbenzimidazol-5-yl)pyrimidin-2-amine), also known as abemaciclib. Similar to the first two classes of compounds, the third class of compounds is also a potent CDK4 / 6 inhibitor.

[0101] [ka]

[0102] In the general formula [3A]: R 1 is the fatty acid moiety

[0103] [ka]

[0104] In the above formula, m is an integer of 0 to 4, more preferably 0 to 2, when m is 0, the piperazine has a direct bond to the nitrogen, and n is an integer of 9 to 20, more preferably 12 to 20; R 2 is H or C 1 -C 3 is alkyl; R 3 and R 4 is H or fluorine, and R 3 and R 4 at least one of which is fluorine; R 5 is C 3 -C 5 Alkyl, C 3 -C 5 cycloalkyl, or cyclopropyl-methyl; R 6 is H or C 1 -C 3 is alkyl; and X is a bond, C 1 -C 3 Alkyl, O, or S.

[0105] The general formula [3B] shown below is a general formula of a third class of preferred embodiments according to this method. In general formula [3B], R 1 is the fatty acid moiety

[0106] [ka]

[0107] In the above formula, m is an integer of 0 to 4, more preferably 0 to 2. When m is 0, the piperazine has a direct bond to the nitrogen; n is an integer of 9 to 20, more preferably 12 to 20; R 2 is H and R 3 and R 4 is fluorine, R 5 is C 3alkyl(isobutyl), R 6 is methyl.

[0108] [ka]

[0109] Compound [3C] shown below is the formula for a third class of illustrative preferred embodiments according to this approach. In general compound [3C], R 1 is the fatty acid moiety

[0110] [ka]

[0111] where m is 0, n is 12, and R 2 is H and R 3 and R 4 is fluorine, R 5 is C 3 alkyl(isobutyl), R 6 is methyl. Consider and plan to evaluate embodiments where n is 9-20. The embodiment designated as compound [3C] is a derivative of abemaciclib, a CDK4 / 6 inhibitor approved by the FDA for the treatment of advanced and metastatic breast cancer. Compound [3C], and other compounds having formula [3A], as described herein, are expected to effectively inhibit CDK4 / 6 and thus be particularly suitable for use as anti-cancer therapeutics, selectively targeting and inhibiting cancer cells and CSCs.

[0112] [ka]

[0113] A second illustrative embodiment of the third class of compounds is shown below as formula [3D]. In this example, R 1 is the fatty acid moiety

[0114] [ka]

[0115] where m is 2, n is 12, and R 2 is H and R 3 and R 4 is fluorine, R 5 is C 3 alkyl(isobutyl), R 6 is methyl. Similar to compound [3C], this embodiment is expected to be more potent than abemaciclib with respect to CDK4 / 6 inhibition and selectivity, making it particularly suitable for use as an anti-cancer therapeutic agent as described herein.

[0116] [ka]

[0117] Table 3 below shows the in vitro biological data for abemaciclib. As can be seen, the compound exhibited excellent cellular activity in the 3D mammosphere assay (IC 50 <0.04 μM) before conjugation. Compounds having formula [3A] containing various fatty acid moieties have been evaluated. It should be recognized from Table 3 that abemaciclib is highly specific and potent against CSCs. Therefore, abemaciclib can be used as a therapeutic agent that specifically targets CSCs. Due to its selectivity, abemaciclib can be used to treat and / or prevent tumor recurrence and / or metastasis and target circulating tumor cells.

[0118] [Table 3]

[0119] Figures 1-3 are dose-response curves of various compounds described herein using a mammosphere assay. The assay was performed using MCF7 cell line. The curves are plotted as the average of two independent experiments, and the standard deviation of each point is represented by a vertical bar. The percentage of inhibition is shown as a function of the logarithm of the concentration (μM). Figure 1 shows the dose-response curve of compound [1C] compared to its parent compound, ribociclib, using a mammosphere formation assay in MCF7 cell line. Figure 4 shows the dose-response curve of compound [1C] compared to its parent compound, using a Hoechst staining assay in MCF7 cell line. As can be seen, compound [1C] is more potent against MCF7 cells at each concentration. This demonstrates that compounds having general formula [1A], and especially general formula [1B], are effective as potent anti-cancer therapeutics. Figure 7 shows the dose-response curve of compound [1C] compared to its parent compound, using a Hoechst staining assay in hTERT-BJ1 cell line. The parent compound was more potent at most concentrations tested, demonstrating that compounds having general formula [1A], and especially general formula [1B], are more selective for CSCs. Similar effects are expected for compounds having general formula [1D].

[0120] Furthermore, although the data disclosed herein concern MCF7 and hTERT-BJ1 cell lines, the compounds of this approach are also effective against other types of cancer. In previous studies, the inventors have demonstrated that mitochondrial biogenesis inhibitors successfully inhibit tumorsphere formation in a variety of cell lines from several tumor types. Table 4 below lists cancer cell lines that have been shown to be susceptible to mitochondrial biogenesis inhibitors. Given these results, this approach is effective against many types of cancer.

[0121] [Table 4]

[0122] The method describes a pharmaceutical composition comprising a therapeutically effective amount of a first class or a second class compound, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier, diluent, or excipient thereof. The compound according to the method can be used as an anti-cancer therapeutic. A pharma- ceutically effective amount of the compound in a pharma- ceutical acceptable carrier can be administered to a subject according to means known in the art. In some embodiments, the compound according to the method can be used in combination with other cancer therapies, including, but not limited to, chemotherapeutic agents, mitochondrial biogenesis inhibitors (e.g., repurposcin, such as mitoriboscin, mitoketoscin, antimitoscin), radiation therapy, phototherapy, and calorie restriction.

[0123] Of course, one of ordinary skill in the art can develop a formulation for a particular embodiment using methods commonly known in the art. In some embodiments, the pharmaceutical composition can be a tablet, capsule, or pill. The pharmaceutical composition can have a dose of 20-500 mg of the therapeutic composition. In some embodiments, the pharmaceutical composition can include a tablet having 200 mg of a therapeutic compound, such as the compound described above, such as compound [1C]. The tablet can contain at least about 35%, 40%, 45%, 50%, or 55% of the therapeutic compound, measured by weight percentage (w / w) of the therapeutic compound (as free base) of the core tablet.

[0124] The tablet may have a core formed of microcrystalline cellulose, crospovidone type A, low-substituted hydroxypropylcellulose, magnesium stearate, colloidal anhydrous silica. In a first illustrative embodiment, a tablet having 200 mg of a therapeutic compound (e.g., compound [1C]) may include an inner core having microcrystalline cellulose (67.44 mg), hydroxypropylcellulose (48.12 mg), crospovidone (29.20 mg), colloidal silicon dioxide (anhydrous) (2.12 mg), and magnesium stearate (6.36 mg), and an outer core having crospovidone (12.84 mg), colloidal silicon dioxide (anhydrous) (1.06 mg), and magnesium stearate (8.46 mg). In a second illustrative embodiment, the tablet may have about 10 to about 45% (w / w) of a therapeutic compound (e.g., compound [2C]), and preferably about 18 to about 28% of a therapeutic compound; about 4 to about 18% of a water-soluble acid; about 20 to about 75% of a diluent; about 5 to about 18% of a disintegrant; about 0.2 to about 10% of a lubricant; and, optionally, about 0 to about 5% of a glidant, and about 0 to about 15% of a binder. Of course, the pharmaceutical composition of this approach may closely resemble a pharmaceutical composition comprising the parent compound. For example, US Pat. No. 6,233,636, filed on Apr. 14, 2016, describes an example of a tablet of ribociclib, and is incorporated herein by reference in its entirety. As another example, US Pat. No. 6,233,636, filed on May 24, 2016, describes a solid formulation of palbociclib, and is incorporated herein by reference in its entirety.

[0125] The tablet may have a film coating. The film coating may include iron oxide, red iron oxide, soy lecithin, polyvinyl alcohol (partially hydrolyzed), talc, titanium dioxide, and xanthan gum. The tablet may be coated with a commercially available coating premix, depending on the desired appearance of the final tablet. For example, Opadry® (Colorcon, Harleysville, PA) is a HPMC (hydroxypropyl methylcellulose) coating material, with the composition HPMC (Pharmacoat 603) 71.4%, polyethylene glycol 7.15%, talc 7.15%, and iron oxide 14.3%.

[0126] The selective inhibition of CDK4 / 6 also indicates that the compounds described herein may be used to reduce or eliminate drug and / or therapeutic resistance in cancer. Because of their inhibitory activity against kinases such as CDKs, the compounds of this method are also useful as tools to study the mechanism of action of such kinases and may be used both in vitro and in vivo.

[0127] The treatment methods described herein are preferably carried out by administering a therapeutically effective amount of a first class or a second class of compound to a subject in need of treatment. The compounds can be readily synthesized using the reaction steps described below, although alternative reaction steps may be used. Alternative reaction steps will be readily recognized by those skilled in the art after reviewing this disclosure, and include those described in Non-Patent Documents 1 and 2. The compounds can be administered by a variety of routes, including orally and parenterally, with little or no toxicity.

[0128] In the examples of synthetic methods described below, the following abbreviations may be used: N-methylmorpholine (NMM), dichloromethane (DCM), dimethylformamide (DMF), ethyl acetate (EtOAc), sodium bicarbonate (NaHCO 3 ), Sodium sulfate (Na 2 SO 4 ), methanol (MeOH). In the following explanation, [M+H]+ refers to the protonated molecule and the value assigned is the mass of the protonated molecule. RT refers to the retention time of the solute.

[0129] Analytical LC-MS: Waters Sunfire C18 30x4.6mm column, gradient eluent: 3-97% acetonitrile / water with 0.05% formic acid. Time: 0-6 min. Preparative HPLC: LC column: Phenomenex Kinetex 5μm EVO C18 100 250x21.2mm. Gradient eluent: 40-95% acetonitrile / water with 0.1% formic acid.

[0130] In the first synthesis example, the above compound [2C], also known as 6-acetyl-8-cyclopentyl-5-methyl-2-[[5-(4-tetradecanoylpiperazin-1-yl)-2-pyridyl]amino]pyrido[2,3-d]pyrimidin-7-one, was synthesized using palbociclib (obtained from LC laboratories, Woburn, Massachusetts, USA). Tetradecanoic acid (0.104 g, 0.46 mmol) was dissolved in thionyl chloride at room temperature. The solution was refluxed for 60 minutes and concentrated under reduced pressure, after which the residue was dissolved in dry DCM (2 mL) at room temperature to obtain a 0.227 M stock solution of the acid chloride. The acid chloride stock solution (0.25 mL, 0.057 mmol) was added to a stirred mixture of 6-acetyl-8-cyclopentyl-5-methyl-2-[[5-(1-piperazinyl)-2-pyridinyl]amino]-pyrido[2,3-d]pyrimidin-7(8H)-one (0.024 g, 0.05 mmol) and NMM (18 μL, 0.16 mmol) in DCM (1 mL) and DMF (0.5 mL). The mixture was stirred at room temperature for 90 min. The solvent was evaporated under reduced pressure and the residue was dissolved in EtOAc (30 mL) and saturated NaHCO 3 (15 mL) and brine (15 mL), then Na 2 SO 4The drying agent was separated by filtration and the filtrate was concentrated under reduced pressure to give the crude product. The crude product was triturated with diethyl ether and the resulting light brown solid was collected by filtration, washed with diethyl ether and then dried under vacuum to give 6-acetyl-8-cyclopentyl-5-methyl-2-[[5-(4-tetradecanoylpiperazin-1-yl)-2-pyridyl]amino]pyrido[2,3-d]pyrimidin-7-one (0.0145 g). LC-MS: 658.2 [M+H] + , RT: 4.12 minutes.

[0131] In the second synthesis example, the above compound [1C], also known as 7-cyclopentyl-N,N-dimethyl-2-[[5-(4-tetradecanoylpiperazin-1-yl)-2-pyridyl]amino]pyrrolo[2,3-d]pyrimidine-6-carboxamide, was synthesized using the same procedure as in the first synthesis example, except that ribociclib (obtained from LC Laboratories, Woburn, Massachusetts, USA) (0.022 g, 0.05 mmol) was reacted with the acid chloride stock solution instead of palbociclib. The crude product was purified on silica gel (2-4% MeOH / DCM) to give 7-cyclopentyl-N,N-dimethyl-2-[[5-(4-tetradecanoylpiperazin-1-yl)-2-pyridyl]amino]pyrrolo[2,3-d]pyrimidine-6-carboxamide (0.0166 g) as a light brown solid. LC-MS: 645.2 [M+H] + , RT: 2.72 minutes.

[0132] In a third synthesis example, the above compound [3C], also known as 1-[4-[[6-[[5-fluoro-4-(7-fluoro-3-isopropyl-2-methyl-benzimidazol-5-yl)pyrimidin-2-yl]amino]-3-pyridyl]methyl]piperazin-1-yl]tetradecan-1-one, was synthesized from a series of intermediate compounds as follows. First, the intermediate compound [4A], shown below, known as tert-butyl-4-tetradecanoylpiperazine-1-carboxylate, was synthesized as follows. To a stirred solution of tetradecanoic acid (1.26 g, 5.5 mmol) and NMM (0.73 mL, 5.5 mmol) in dry DCM (20 mL) was added isobutyl chloroformate (0.65 mL, 5.0 mmol) at room temperature under a nitrogen atmosphere. After 4 hours, a solution of 1-Boc-piperazine (0.93 g, 5.0 mmol) in dry DCM (5 mL) was added to the mixture. The mixture was stirred for 16 hours. The solvent was removed under reduced pressure to give the crude product. The crude product was dissolved in EtOAc (75 mL) and washed with 2 M HCl (50 mL), saturated NaHCO 3 (40 mL), and brine (30 mL), followed by washing with MgSO 4 After filtration, the solvent was evaporated under reduced pressure to give tert-butyl-4-tetradecanoylpiperazine-1-carboxylate (1.76 g) as a white solid. LC-MS: 397.2 [M+H] + , RT: 4.02 minutes.

[0133] [ka]

[0134] Second, the intermediate compound [4B], shown below, known as 1-piperazin-1-yltetradecan-1-one, was synthesized as follows: A solution of tert-butyl-4-tetradecanoylpiperazine-1-carboxylate (0.51 g, 1.26 mmol) in a 1:1 mixture of dry DCM (10 mL) and trifluoroacetic acid (TFA) (10 mL) was stirred at room temperature under nitrogen atmosphere for 90 minutes, and the solvent was removed under reduced pressure to give the crude product. The crude product was dissolved in EtOAc (30 mL) and eluted with saturated NaHCO 3(15 mL) and brine (15 mL), followed by washing with MgSO 4 After filtration, the solvent was evaporated under reduced pressure to give 1-piperazin-1-yltetradecan-1-one (0.33 g) as a white waxy solid. LC-MS: 297.3 [M+H] + , RT: 1.81 minutes.

[0135] [ka]

[0136] Third, the intermediate compound [4C], known as 6-[[5-fluoro-4-(7-fluoro-3-isopropyl-2-methyl-benzimidazol-5-yl)pyrimidin-2-yl]amino]pyridine-3-carbaldehyde, shown below, was prepared as follows: 6-aminopyridine-3-carbaldehyde (0.076 g, 0.625 mmol), 6-(2-chloro-5-fluoro-pyrimidin-4-yl)-4-fluoro-1-isopropyl-2-methyl-benzimidazole (0.161 g, 0.500 mmol), Xantphos (0.0276 g, 0.0476 mmol), palladium chloride (0.0056 g, 0.0315 mmol), and K in 2-methyl-2-butanol (4 mL). 2 CO 3 A suspension of (0.069 g, 0.500 mmol) was heated in a sealed tube at +100° C. for 18 h. The reaction mixture was cooled to room temperature and diluted with EtOAc (30 mL) and water (30 mL). The precipitate with the aqueous phase was separated and collected by filtration. The light brown solid was washed with water (30 mL) and acetone (20 mL) and dried under vacuum to give 6-[[5-fluoro-4-(7-fluoro-3-isopropyl-2-methyl-benzimidazol-5-yl)pyrimidin-2-yl]amino]pyridine-3-carbaldehyde (0.125 g) as a light brown solid. LC-MS: 409.0 [M+H] + , RT: 2.07 minutes.

[0137] [ka]

[0138] Fourth, the intermediate compound [4D] shown below, known as 1-[4-[[6-[[5-fluoro-4-(7-fluoro-3-isopropyl-2-methyl-benzimidazol-5-yl)pyrimidin-2-yl]amino]-3-pyridyl]-methyl]piperazin-1-yl]tetradecan-1-one, and also mentioned above as compound [3C], was synthesized as follows. A suspension of 6-[[5-fluoro-4-(7-fluoro-3-isopropyl-2-methyl-benzimidazol-5-yl)pyrimidin-2-yl]amino]pyridine-3-carbaldehyde (0.050 g, 0.122 mmol), 1-piperazin-1-yltetradecan-1-one (0.030 g, 0.100 mmol), and sodium triacetoxyborohydride (0.212 g, 1.00 mmol) in DCE (20 mL) was heated at +60 °C for 90 min in a sealed tube. The reaction mixture was cooled to room temperature, diluted with DCM (30 mL), washed with water (10 mL) and brine (10 mL) and then diluted with MgSO 4 After filtration, the solvent was evaporated under reduced pressure and the resulting crude product (0.1564 g) was purified by preparative HPLC to give 1-[4-[[6-[[5-fluoro-4-(7-fluoro-3-isopropyl-2-methyl-benzimidazol-5-yl)pyrimidin-2-yl]amino]-3-pyridyl]methyl]piperazin-1-yl]tetradecan-1-one (0.012 mg). LC-MS: 689.2 [M+H] + , RT: 2.44 minutes.

[0139] [ka]

[0140] The following paragraphs describe the materials and methods used in connection with the data and embodiments described herein. Of course, one skilled in the art may use alternative materials and methods that are generally accepted in the art without departing from the present procedure.

[0141] For cell culture and reagents, human breast adenocarcinoma cell line (MCF-7) was obtained from the American Type Culture Collection (ATCC), and hTERT-BJ1 cells were obtained from Clontech, Inc. MCF-7 and hTERT-BJ1 cells were grown in DMEM supplemented with 10% fetal bovine serum, GlutaMAX, and 1% penicillin-streptomycin and incubated at 37°C, 5% CO 2 The cells were cultured in a humidified incubator at 37 °C for 1 h. The medium was changed two to three times a week.

[0142] Mammosphere formation assay: Single cell suspensions were prepared using enzymatic disaggregation (1x trypsin-EDTA, Sigma Aldrich, Cat. No.: T3924) and manual disaggregation (25 gauge needle). 5000 cells were seeded in 6-well plates coated with 2-hydroxyethyl methacrylate (poly-HEMA, Sigma, Cat. No. P3932) under non-adherent conditions in mammosphere medium (DMEM-F12 / B27 / 20ng / mL EGF / PenStrep). Cells were grown for 5 days and kept in a humidified incubator at 37°C, atmospheric pressure, and 5% (v / v) carbon dioxide / air. After 5 days, 3D spheroids larger than 50 μm in diameter were counted using a microscope equipped with a graticule eyepiece, and the percentage of cells that formed spheroids was calculated and normalized to 1 (1=100% MFE; mammosphere formation efficiency). Mammosphere assays were performed in triplicate and repeated three independent times.

[0143] A Hoechst-based viability assay was used to characterize the selectivity of the compounds of this approach for selective targeting of cancer cells. Briefly, monolayers of MCF7 cells were treated for one day at concentrations ranging from 1 to 100 μM. Cell viability was assessed using the nucleic acid dye Hoechst 33342, which stains DNA in live cells. The viability of normal human fibroblast cells (hTERT-BJ1) treated with the compounds described herein was also assessed simultaneously. Quantification was performed using a plate reader.

[0144] The terminology used in the description of the embodiments of the present technique is for the purpose of describing particular embodiments only and is not intended to be limiting. As used in the description and the appended claims, the singular forms "a," "an," and "the" are intended to include the plural forms unless the context clearly indicates otherwise. The present technique encompasses numerous alternatives, modifications, and equivalents, as will become apparent in light of the following detailed description.

[0145] In this specification, terms such as "first", "second", "third", "a)", "b)" and "c)" may be used to describe various elements of the method, but it will be understood that the claims should not be limited by these terms. These terms are used only to distinguish one element of the method from another. Thus, a first element described below could be referred to as an aspect of the element, or similarly, as a third element, without departing from the teachings of the method. Thus, terms such as "first", "second", "third", "a)", "b)" and "c)" are not necessarily intended to impose a hierarchy, such as order, on the associated elements, but are used for identification purposes only. The order of operations (or steps) is not limited to the order set forth in the claims.

[0146] Unless otherwise defined, all terms used herein (including technical and scientific terms) have the same meaning as commonly understood by those skilled in the art. It is further understood that terms as defined in commonly used dictionaries should be interpreted to have a meaning consistent with the meaning in the context of this application and related art, and should not be interpreted in an idealized or overly formal sense unless expressly defined herein. All publications, patent applications, patents, and other references mentioned herein are incorporated herein by reference in their entirety. In the event of a discrepancy in terms, the present specification shall control.

[0147] Also, as used herein, "and / or" refers to and includes every possible combination of one or more of the associated listed items, as well as the lack of combination ("or") when interpreted in the alternative.

[0148] It is specifically contemplated that the various features of the present approach described herein can be used in any combination, unless the context indicates otherwise. Further, the present approach contemplates that in some embodiments, any feature or combination of features described in connection with the illustrative embodiments can be excluded or omitted.

[0149] As used herein, the transitional phrase "consisting essentially of" (and grammatical variations thereof) is intended to be construed to encompass the recited materials or steps "and which do not materially affect the basic and novel characteristic or characteristics of the claims." Thus, as used herein, the term "consisting essentially of" should not be construed as the equivalent of "comprising."

[0150] Thus, while particular embodiments of the present technique have been described, it should be understood that the appended claims are not limited to the specific details set forth in the above description, since many obvious variations thereof are possible without departing from the spirit or scope thereof as hereinafter claimed.

Claims

1. general formula 【Chemistry 1】 A compound represented by the formula: R ４ is hydrogen, C １ -C ８ Alkyl, C ３ -C ８ selected from the group consisting of cycloalkyl, aryl, and heteroaryl; Z is R ｚ and R ｚ is halo, hydrogen, C １ -C ３ Alkyl, C １ -C ３ Alkoxy, CN, C=NOH, C=NOCH ３ , C(O)H, C(O)C １ -C ３ Alkyl, C ３ -C ８ Cycloalkyl, heterocyclyl, aryl, heteroaryl, -B-NR ａ R ｂ , -B-OR ａ , -B-C(O)R ａ , -B-C(O)OR ａ and -B-C(O)NR ａ R ａ B is a bond, C １ -C ３ Alkyl, or branched C １ -C ３ alkyl; R ａ and R ｂ are each independently hydrogen, C １ -C ３ Alkyl, C ３ -C ８ selected from the group consisting of cycloalkyl, heterocyclyl, aryl, and heteroaryl; and n is an integer from 9 to 20; The C 1 -C 8 alkyl, the C 1 -C 3 alkyl, the C 3 -C 8 cycloalkyl and the heterocyclyl are each independently selected from the group consisting of C 1 -C 20 and optionally substituted with at least one substituent selected from the group consisting of alkyl, alkoxy, alkenyl, alkynyl, halo, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, amino, acylamino, amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, morpholino, phenol, benzyl, phenyl, piperazine, cyclopentane, cyclohexane, pyridine, 5H-tetrazole, triazole, piperidine, and combinations thereof; The compound, wherein said aryl and said heteroaryl are each independently optionally substituted with at least one substituent selected from the group consisting of C 1 -C 20 alkyl, halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylthiocarbonyl, phosphate, phosphonato, phosphinato, cyano, amino, acylamino, amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfate, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, and combinations thereof.

2. The compound according to claim 1, wherein n is an integer from 12 to 20.

3. R ４ 3. The compound of claim 2, wherein is cyclopentyl and Z is dimethylcarboxamide.

4. The compound has the general formula 【Chemistry 2】 2. The compound according to claim 1, wherein n is an integer from 9 to 20.

5. The compound according to claim 4, wherein n is an integer from 12 to 20.

6. The compound of claim 4 , wherein n is 12.

7. A pharmaceutical composition comprising a compound according to any one of claims 1 to 6, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier.

8. 8. The pharmaceutical composition of claim 7, wherein the composition is a tablet comprising a core having 35-55% by weight of a compound according to any one of claims 1 to 6 and a pharma- ceutically acceptable carrier.

9. 9. The pharmaceutical composition of claim 8, wherein the pharma- ceutically acceptable carrier comprises microcrystalline cellulose, crospovidone type A, low-substituted hydroxypropyl cellulose, magnesium stearate, and colloidal anhydrous silica.

10. A pharmaceutical composition for treating cancer, comprising a pharma- ceutically effective amount of a compound according to any one of claims 1 to 6.

11. A pharmaceutical composition for treating or preventing metastatic disease, comprising a pharma- ceutically effective amount of a compound according to any one of claims 1 to 6.

12. A pharmaceutical composition for treating or preventing tumor recurrence, comprising a pharma- ceutically effective amount of a compound according to any one of claims 1 to 6.

13. A pharmaceutical composition for reducing resistance to cancer treatment, comprising a pharma- ceutically effective amount of a compound according to any one of claims 1 to 6.

14. A pharmaceutical composition for treating or preventing at least one of radiation therapy resistance, chemotherapy resistance, and hormone therapy resistance, comprising a pharma- ceutical composition comprising a pharma- ceutical effective amount of a compound according to any one of claims 1 to 6.

15. A pharmaceutical composition for preventing or reducing proliferation of at least one of cancer cells, cancer stem cells, and circulating tumor cells, comprising a pharma- tically effective amount of a compound according to any one of claims 1 to 6.

16. 13. Use of a compound according to any one of claims 1 to 6 in the manufacture of a medicament for the treatment of cancer.

17. 10. Use of a compound according to any one of claims 1 to 6 in the manufacture of a medicament for the treatment of metastatic disease.

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