Azetidinyl-acetamides as cxcr7 inhibitors and use thereof

Abstract

This invention provides compounds having formula I, Or a pharmaceutically acceptable salt, hydrate, or N-oxide thereof, which is suitable for binding to CXCR7 and for treating diseases at least partially dependent on CXCR7 activity. Therefore, the present invention provides compositions in other forms comprising one or more of the above-mentioned compounds mixed with a pharmaceutically acceptable excipient.

Description

Prior Technology

[0001] This invention relates to novel compounds and pharmaceutical compositions that inhibit the binding of SDF-1 chemokine (also known as CXCL12 chemokine) or I-TAC (also known as CXCL11) to the chemokine receptor CRCX7 (also known as ACKR3). These compounds are suitable for the prevention of tumor cell proliferation, tumor formation, tumor angiogenesis, metastasis, inflammatory diseases (including but not limited to arthritis, nephritis, and multiple sclerosis), inappropriate angiogenesis conditions (including but not limited to wound healing), treatment of HIV infection, and treatment of stem cell differentiation and mobilization disorders, acute renal failure, hemolytic uremic syndrome, ischemia / reperfusion injury, opioid addiction, and neuralgia (see also USSN 10 / 912,638, 11 / 407,729, and 11 / 050,345, which are also filed).

[0002] Chemokines belong to a small superfamily of intercytokines that induce cellular scaffold rearrangement, strong adhesion of leukocytes to endothelial cells, leukocyte degranulation and directional migration, and can also affect cell activation and proliferation. Chemokines act in a synergistic manner with cell surface proteins to guide various cell subgroups to specific anatomical sites.

[0003] Multiple early studies have indicated the role of the chemokine receptor CXCR4 in metastasis and tumor growth. Muller et al.'s "Involvement of Chemokine Receptors in Breast Cancer Metastasis," Nature, 410:50-56 (2001), demonstrated that breast cancer cells utilize chemokine-mediated mechanisms during metastasis, such as the regulation of leukocyte transport. Tumor cells exhibit a unique, non-random pattern of functionally active chemokine receptors. CXCR4 signaling mediates actin polymerization and pseudopodia formation in breast cancer cells and induces chemotactic and invasive responses. Furthermore, organs representing the primary sites of breast cancer metastasis (such as lymph nodes, bone marrow, and lungs) are the richest sources of CXCR4 receptor ligands.

[0004] Using immunodeficient mice, Muller and colleagues successfully reduced the metastasis of injected human breast cancer cells by treating mice with antibodies known to bind to CXCR4. Their findings suggest that breast cancer metastasis can be reduced by treating patients with CXCR4 antagonists.

[0005] Bertolini et al.'s study, "CXCR4 Neutralization, a Novel Therapeutic Approach for Non-Hodgkin's Lymphoma," Cancer Research, 62:3106-3112 (2002), showed that immunodeficient mice injected with human lymphoma cells treated with anti-CXCR4 antibodies experienced reduced tumor volume and prolonged survival. They explained their findings as implying that tumor volume could be reduced by treating patients with CXCR4 antagonists.

[0006] Subsequent studies have shown that another chemokine receptor, CXCR7, can also be targeted in cancer treatment. CXCR7 is preferentially expressed in transformed cells compared to normal cells and is detectable in a variety of human cancers. In vitro studies indicate that the proliferation of CXCR7-expressing cells can be inhibited by CXCR7 antagonists. In vivo studies in mice have shown that CXCR7 antagonists can inhibit tumor formation and tumor growth (reviewed in Morian, D. et al., Front Immunol (2020) 11:952-971).

[0007] Some CXCR7 antagonists can prevent cancer growth and spread, and their performance patterns indicate limited tissue distribution of CXCR7 receptors associated with tumorigenesis (Luo, Y. et al., Int. J. Cancer, (2018) 142:2163-2174).

[0008] In addition, CXCR7 can act as a co-receptor for certain genetically divergent human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV), especially for HIV-2-ROD, an X4 tropism isolate (Shimizu, N. et al., J. Virol., (2000) 74: 619-626; Balabanian, K. et al., J. Biol. Chem., (2005) 280:35760-35766; D'huys, T. et al., Heliyon, (2018) 4:e00557).

[0009] Furthermore, SDF-1 has been described as playing a role in the mobilization of hematopoietic precursor cells and stem cells, particularly those carrying the CXCR4 receptor, from specific hematopoietic tissues, including the bone marrow (Hattori, K. et al., Blood, (2000) 97:3354-3360; WO 2005 / 000333, the contents of which are incorporated herein by reference). Recent research indicates that CXCR7 may also be involved in stem cell mobilization (Melo, RD et al., Stem Cell Res Ther, (2018) 9:34-38).

[0010] In addition to its initial role in cancer cell and stem cell mobilization, the regulation of this receptor has shown promise in preventing HIV infection, Shiga toxin-associated hemolytic uremic syndrome, ischemia / reperfusion injury in cardiac tissue and transplanted kidneys, and reducing muscle sclerosis. Recent findings that CXCR7 binds to opioids suggest that its regulation can modulate pain and provide potent treatment for opioid addiction.

[0011] In view of the above, it is evident that compounds capable of specifically binding to the CXCR7 receptor are suitable for treating diseases and other biological conditions that may benefit from such interactions. This invention provides such compounds, pharmaceutical compositions, and related treatment methods. Summary of the Invention

[0012] [Statement of rights regarding inventions made under federally sponsored research and development] []

[0013] not applicable [Reference to the "Serial List," table, or computer program list appendix submitted on the CD] []

[0014] not applicable

[0015] In one embodiment, the present invention provides a compound having formula I. [] Or a pharmaceutically acceptable salt, hydrate, or N-oxide thereof. Various groups (e.g., R1, R2, R3, R4, R4a, R5, R5a, R6, R7, Ar1, Ar2, HAr, and subscripts m, n, p, and q) are described in embodiments of the invention.

[0016] The compounds provided herein are suitable for binding to CXCR7 and for treating diseases that are at least partially dependent on CXCR7 activity. Therefore, the present invention provides compositions in other forms comprising one or more of the above-described compounds mixed with a pharmaceutically acceptable excipient.

[0017] In another embodiment, the present invention provides a method for treating various diseases further discussed herein, comprising administering a therapeutically effective amount of the compound of the formula to an individual in need of such treatment for a duration sufficient to treat the disease.

[0018] In another aspect, the present invention provides a method for diagnosing a disease in an individual. In this method, the compound provided herein is administered to the individual in a labeled form, followed by diagnostic imaging to determine the presence or absence of CXCR7. In a related aspect, the method for diagnosing a disease is performed by contacting a tissue or blood sample with a labeled compound as provided herein and determining the presence, absence, or amount of CXCR7 in the sample.

[0019] In some embodiments, a certain amount of chemotherapeutic agent or radiation is administered to an individual before, after, or in combination with the compounds of the present invention. In some embodiments, when the chemotherapeutic agent or radiation is administered alone, the amount is less than a therapeutic dose. Implementation

[0020] [Cross-reference to related applications] []

[0021] This application claims priority to U.S. Provisional Application No. 63 / 176,451, filed April 19, 2022, pursuant to 35 USC § 119(e), the disclosure of which is incorporated herein by reference in its entirety. I. Abbreviations and Definitions

[0022] Unless otherwise stated, the term "alkyl" itself, or as part of another substituent, means a straight-chain or branched hydrocarbon group having a specified number of carbon atoms (i.e., C1-8 means one to eight carbons). Examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, tributyl, isobutyl, dibutyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, and similar alkyl groups. The term "alkenyl" refers to an unsaturated alkyl group having one or more double bonds. Similarly, the term "alkynyl" refers to an unsaturated alkyl group having one or more double bonds. Examples of such unsaturated alkyl groups include vinyl, 2-propenyl, crotonyl, 2-isopentenyl, 2-(butadienyl), 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1-propynyl, and 3-propynyl, 3-butynyl, and their higher homologs and isomers.

[0023] The term "cycloalkyl" refers to a saturated or partially unsaturated hydrocarbon ring (e.g., C3-6 cycloalkyl) having a specified number of ring atoms. Cycloalkyl groups can include any number of carbon atoms, such as C3-6, C4-6, C5-6, C3-8, C4-8, C5-8, C6-8, C3-9, and C3-10. Partially unsaturated cycloalkyl groups have one or more double or triple bonds in the ring, but the cycloalkyl group is not aromatic. Saturated monocyclic cycloalkyl rings include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cyclooctyl.

[0024] The term "cycloalkoxy" refers to a cycloalkyl group having an oxygen atom that connects the cycloalkyl group to the junction point: cycloalkyl-O-. Cycloalkyl groups are as defined herein.

[0025] The term "bridged cycloalkyl" or "bridged cycloalkyl group" refers to a cycloalkyl ring (having 4 to 8 ring vertices) in which two non-adjacent ring atoms are connected by a (CRR')n group, where n is 1 to 3 and each R is independently H or methyl (also referred to herein as a "bridging" group). Bridged cycloalkyl groups do not have any heteroatoms as ring vertices. Additionally, C5-8 refers to bridged cycloalkyl groups having 5 to 8 ring members. Examples include, but are not limited to, bicyclo[1.1.1]pentane, bicyclo[2.2.2]octane, bicyclo[2.2.1]heptane, and similar groups.

[0026] The term "spirocyclic" or "spirocyclic alkyl" refers to a saturated or partially unsaturated bicyclic ring having 6 to 12 ring atoms, wherein the two rings are linked by a single carbon atom (also called a spiro atom). Partially unsaturated spirocyclic alkyl groups have one or more double or triple bonds in the ring, but are not aromatic. Representative examples include, but are not limited to, spiro[3.3]heptane, spiro[4.4]nonane, spiro[3.4]octane, and similar groups.

[0027] The term "heterocyclic alkyl" refers to a saturated or partially unsaturated monocyclic ring having a specified number of ring vertices (e.g., 3- to 7-membered rings) and one to five heteroatoms selected from N, O, and S as ring vertices. A partially unsaturated heterocyclic alkyl group has one or more double or triple bonds in the ring, but the heterocyclic alkyl group is not aromatic. Heterocyclic alkyl groups may include any number of ring atoms, such as 3 to 6, 4 to 6, 5 to 6, 3 to 7, 4 to 7, or 5 to 7 ring members. Heterocyclic alkyl groups may include any suitable number of heteroatoms, such as 1, 2, 3, or 4, or 1 to 2, 1 to 3, 1 to 4, 2 to 3, 2 to 4, or 3 to 4. Non-limiting examples of heterocyclic alkyl groups include pyrrolidine, imidazoidine, pyrrolidine, butyrolactam, valproic acid, imidazolidinone, hydantoin, dioxapentane, o-phthalimide, piperidine, 1,4-dialkyl, pyrrolidine, thiopyrrolidine, thiopyrrolidine-S-oxide, thiopyrrolidine-S,S-oxide, piperazine, piperan, pyridinone, 3-pyrrolidine, thiopiperanone, piperanone, tetrahydrofuran, tetrahydrothiophene. Pyridine and similar groups. Heterocyclic alkyl groups can be attached to the rest of the molecule via a cyclic carbon or heteroatom.

[0028] The terms "bicyclic heterocyclic alkyl" or "bicyclic heterocyclic group" refer to a saturated or partially unsaturated fused bicyclic ring having a specified number of ring vertices (e.g., 6- to 12-membered rings) and one to five heteroatoms selected from N, O, and S as ring vertices. A partially unsaturated bicyclic heterocyclic alkyl group has one or more double or triple bonds in the ring, but is not aromatic. Bicyclic heterocyclic alkyl groups may include any number of ring atoms, such as 6 to 8, 6 to 9, 6 to 10, 6 to 11, or 6 to 12 ring members. Heterocyclic alkyl groups may include any suitable number of heteroatoms, such as 1, 2, 3, or 4, or 1 to 2, 1 to 3, 1 to 4, 2 to 3, 2 to 4, or 3 to 4. Non-limiting examples of bicyclic heterocyclic alkyl groups include decahydro-1,5-diphenyl ether, octahydropyrrolo[1,2-a]pyridine, and similar groups.

[0029] The terms "bridged heterocyclic group" or "bridged heterocyclic alkyl group" refer to a heterocyclic alkyl ring (having 5 to 7 ring vertices) in which two non-adjacent ring atoms are connected by a (CRR')n group, where n is 1 to 3 and each R is independently H or methyl (also referred to herein as a "bridging" group). The bridged heterocyclic group has one to five heteroatoms selected from N, O, and S as ring vertices. The heteroatom ring vertices can be in both the heterocyclic alkyl ring portion and the bridging group. When in the bridging group, the heteroatom replaces the CRR' group. Examples include, but are not limited to, 2-azabicyclo[2.2.2]octane, Pyridine, 7-bis(2.2.1)heptane and similar groups.

[0030] The term "spiroheterocyclic" or "spiroheteroalkyl" refers to a saturated or partially unsaturated bicyclic ring having 6 to 12 ring atoms, wherein the two rings are connected by a single carbon atom (also called a spiro atom). The spiroheterocyclic group has one to five heteroatoms selected from N, O, and S as ring apexes, and the nitrogen atom is, where appropriate, quaternary ammonium. Partially unsaturated spiroheteroalkyl groups have one or more double or triple bonds in the ring, but the spiroheteroalkyl group is not aromatic. Representative examples include, but are not limited to, 2,6-diazaspiro[3.3]heptane, 2,6-diazaspiro[3.4]octane, 2-azaspiro[3.4]octane, 2-azaspiro[3.5]nonane, 2,7-diazaspiro[4.4]nonane, and similar groups.

[0031] The term "alkylene" itself, or as part of another substituent, refers to a divalent group derived from an alkane, such as -CH₂CH₂CH₂CH₂-. Typically, an alkyl (or alkylene) will have 1 to 24 carbon atoms, while in this invention, groups having 10 or fewer carbon atoms are preferred. "Low-carbon alkyl" or "low-carbon alkylene" refers to a shorter-chain alkyl or alkylene group, generally having four or fewer carbon atoms. Similarly, "alkyleneyl" and "alkylynyl" refer to alkylene groups in their saturated forms, respectively, having double or triple bonds.

[0032] As used herein, the wavy lines intersecting single bonds, double bonds, or reference bonds in any chemical structure depicted herein are […]. "" indicates the connection point between a single bond, double bond, or parabond and the rest of the molecule.

[0033] The terms "alkoxy," "alkylamine," and "alkathio" (or thioalkoxy) are used in their conventional sense and refer to alkyl groups connected to the remainder of a molecule via an oxygen atom, an amino group, or a sulfur atom, respectively. Additionally, for dialkylamine groups, the alkyl motifs may be the same or different and may be combined to form 3- to 7-membered rings with their respective connected nitrogen atoms. Therefore, groups denoted as -NR aR b are intended to include piperidinyl, pyrrolidinyl, α-linyl, acridineyl, and similar groups.

[0034] Unless otherwise stated, the term "halogen" or "halogen" itself, or as part of another substituent, means a fluorine, chlorine, bromine, or iodine atom. Additionally, terms such as "haloalkyl" mean both monohaloalkyl and polyhaloalkyl. For example, the term "C1-4 haloalkyl" means trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and similar groups.

[0035] The term "hydroxyalkyl" means an alkyl group as defined above, having one or two hydroxyl groups as substituents. For example, the term "C1-6 hydroxyalkyl" means including 2-hydroxyethyl and 2,4-dihydroxybutyl.

[0036] Unless otherwise stated, the term "aryl" means a polyunsaturated, usually aromatic, hydrocarbon group, which can be a monocyclic or fused together or covalently linked multiple rings (up to three rings). The term "heteroaryl" refers to an aryl group (or ring) containing one to five heteroatoms selected from N, O, and S, wherein the nitrogen and sulfur atoms are oxidized as appropriate, and the nitrogen atom is quaternized as appropriate. Heteroaryl groups can be attached to the rest of the molecule via heteroatoms. Non-limiting examples of aryl groups include phenyl, naphthyl, and biphenyl, while non-limiting examples of heteroaryl groups include pyridyl, pyridyl, pyrimidinyl, triazolyl, quinolinyl, quinolinyl, quinazolinyl, aceolinyl, terpenoid, benzotriazolyl, purine, benzimidazolyl, benzopyrazolyl, benzotriazolyl, benzoisoazolyl, isobenzofuranyl, isoindolyl, indolyl, and others. Benzotriazolyl, thienenopyridyl, thienenopyrimidyl, pyrazolopyrimidyl, imidazolylpyridine, benzothiazolyl, benzofuranyl, benzothiaphenyl, indolyl, quinolinyl, isoquinolinyl, isothiazolyl, pyrazolyl, indazole, pteridinyl, imidazolyl, triazolyl, tetrazolyl, acezolyl, isoacezolyl, thiadiazolyl, pyrroleyl, thiazolyl, furanyl, thienyl, and similar groups. The substituents in each of the aryl and heteroaryl ring systems mentioned above are selected from the group of acceptable substituents listed below.

[0037] The term "arylalkyl" means that an aryl group is attached to an alkyl group (e.g., benzyl, phenethyl, and similar groups). Similarly, the term "heteroaryl-alkyl" means that a heteroaryl group is attached to an alkyl group (e.g., pyridylmethyl, thiazolylethyl, and similar groups).

[0038] In some embodiments, the terms above (e.g., "alkyl", "aryl", and "heteroaryl") will include substituted and unsubstituted forms of the specified group. Preferred substituents for various groups are provided below.

[0039] The substituents of alkyl groups (including those commonly referred to as alkylene, alkenyl, ynyl, and cycloalkyl groups) may be selected from a variety of groups including: -halogen, -OR', -NR'R'', -SR', -SiR'R''R''', -OC(O)R', -C(O)R', -CO 2R', -CONR'R'', -OC(O)NR'R'', -NR''C(O)R'', -NR''C(O) 2R', -NHC(NH 2)=NH, -NR'C(NH 2)=NH, -NHC(NH 2)=NR', -S(O)R', -S(O) 2R', -S(O) 2NR'R'', -NR'S(O) 2R'', -CN, and -NO 2, in numbers ranging from zero to (2 Within the range of m'+1), where m' is the total number of carbon atoms in such groups. R', R'', and R''' each independently refer to hydrogen, an unsubstituted C1-8 alkyl group, an unsubstituted aryl group, an aryl group substituted with 1 to 3 halogens, an unsubstituted C1-8 alkyl group, a C1-8 alkoxy group, or a C1-8 thioalkoxy group, or an unsubstituted aryl-C1-4 alkyl group. When R' and R'' are attached to the same nitrogen atom, they can combine with the nitrogen atom to form 3-, 4-, 5-, 6-, or 7-membered rings. For example, -NR'R'' means including 1-pyrrolidyl and 4-pyrolinyl.

[0040] Similarly, the substituents of aryl and heteroaryl groups are diverse and generally selected from: -halogen, -OR', -OC(O)R', -NR'R'', -SR', -R', -CN, -NO 2, -CO 2R', -CONR'R'', -C(O)R', -OC(O)NR'R'', -NR''C(O)R', -NR''C(O) 2R', -NR'C(O)NR''R''', -NHC(NH 2)=NH, -NR'C(NH 2)=NH, -NHC(NH 2)=NR', -S(O)R', -S(O) 2R', -S(O) 2NR'R'', -NR'S(O) 2R'', -N 3, perfluoro(C 1-C 4)alkoxy and perfluoro(C 1-C 4)alkoxy 4) Alkyl groups, in the range of zero to the total number of open valences on the aromatic ring system; wherein R', R'', and R''' are independently selected from hydrogen, C1-8 alkyl, C1-8 haloalkyl, C3-6 cycloalkyl, C2-8 alkenyl, C2-8 alkynyl, unsubstituted aryl and heteroaryl, (unsubstituted aryl)-C1-4 alkyl, and unsubstituted aryloxy-C1-4 alkyl. Other suitable substituents include each of the above aryl substituents connected to the ring atom by an alkyl chain of 1-4 carbon atoms.

[0041] Two of the substituents on adjacent atoms of the aryl or heteroaryl ring may be substituted, as appropriate, with a substituent of the formula -TC(O)-(CH2)qU-, where T and U are independently -NH-, -O-, -CH2-, or single bonds, and q is an integer from 0 to 2. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may be substituted, as appropriate, with a substituent of the formula -A-(CH2)rB-, where A and B are independently -CH2-, -O-, -NH-, -S-, -S(O)-, -S(O)2-, -S(O)2NR'-, or single bonds, and r is an integer from 1 to 3. One of the single bonds in the resulting new ring may be substituted, as appropriate, with a double bond. Alternatively, two of the substituents on adjacent atoms of the aryl or heteroaryl ring may be replaced by a substituent of the formula -(CH2)sX-(CH2)t-, where s and t are independently integers from 0 to 3, and X is -O-, -NR'-, -S-, -S(O)-, -S(O)2-, or -S(O)2NR'-. The substituent R' in -NR'- and -S(O)2NR'- is selected from hydrogen or an unsubstituted C1-6 alkyl group.

[0042] As used in this article, the term "heteroatoms" means that they include oxygen (O), nitrogen (N), sulfur (S), and silicon (Si).

[0043] The term "medically acceptable salt" means salts of active compounds prepared with relatively non-toxic acids or bases, depending on the specific substituents present on the compounds described herein. When the compounds of the present invention contain relatively acidic functional groups, base addition salts can be obtained by contacting a sufficient amount of the desired base in a solvent-free environment or in a suitable inert solvent with the neutral form of such compounds. Examples of salts derived from pharmaceutically acceptable inorganic bases include aluminum salts, ammonium salts, calcium salts, copper salts, iron salts, ferrous salts, lithium salts, magnesium salts, manganese salts, manganese salts, potassium salts, sodium salts, zinc salts, and similar salts. Salts derived from pharmaceutically acceptable organic bases include salts of primary, secondary, and tertiary amines, including substituted amines, cyclic amines, naturally occurring amines, and their analogues, such as arginine, betaine, caffeine, choline, N,N'-diphenylmethylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylpyridine, N-ethylpiperidine, reduced glucosamine, glucosamine, histidine, hyprothidine, isopropylamine, lysine, methylglucosamine, pyridine, piperidine, piperidine, polyamine resins, procaine, purine, theobromine, triethylamine, trimethylamine, tripropylamine, thiazolinone, and their analogues. When the compounds of the present invention contain relatively basic functional groups, acid addition salts can be obtained by contacting the neutral form of these compounds with a sufficient amount of the desired acid in a solvent-free environment or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, hydrocarbonic acid, phosphoric acid, phosphoric acid monohydrogen phosphate, phosphoric acid dihydrogen phosphate, sulfuric acid, hydrosulfuric acid, hydroiodic acid, or phosphorous acid and similar inorganic acids; and salts derived from relatively non-toxic organic acids such as acetic acid, propionic acid, isobutyric acid, malonic acid, benzoic acid, succinic acid, octanoic acid, fumaric acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid, and similar organic acids. Also included are salts of amino acids such as arginine and similar acids, and salts of organic acids such as glucuronic acid or galacturonic acid and similar acids (see, for example, Berge, SM, et al., "Pharmaceutical Salts", Journal of Pharmaceutical Science).

[1977] (66, 1-19). Certain specific compounds of the present invention contain both basic and acidic functional groups that allow the compound to be converted into a base addition salt or an acid addition salt.

[0044] The neutral form of a compound can be regenerated by contacting the salt with a base or acid and separating the parent compound in a conventional manner. The parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but for the purposes of this invention, in other respects, the salt is equivalent to the parent form of the compound.

[0045] In addition to salt forms, the present invention provides compounds in prodrug form. The prodrugs of the compounds described herein are those compounds that readily undergo chemical changes under physiological conditions to provide the compounds of the present invention. Furthermore, the prodrugs can be converted into the compounds of the present invention in an in vitro environment by chemical or biochemical methods. For example, when a prodrug is placed together with a suitable enzyme or chemical reagent in a percutaneous patch reservoir, it can be slowly converted into the compounds of the present invention.

[0046] Some of the compounds of this invention may exist in both unsolvated and solvated forms (including hydrated forms). Generally, the solvated form is equivalent to the unsolvated form and is intended to be included within the scope of this invention. Some of the compounds of this invention may exist in various crystalline or amorphous forms. Generally, all physical forms are equivalent for the uses covered by this invention and are intended to be within the scope of this invention.

[0047] Some of the compounds of this invention possess asymmetric carbon atoms (optical centers) or double bonds; racemic, non-mirror image isomers, geometric isomers, regioisomers, and individual isomers (e.g., independent mirror image isomers) are all intended to be covered within the scope of this invention. In some embodiments, the compounds of this invention are present in a mirror-isomerically enriched form, wherein the amount of mirror-isomeric excess of a particular mirror-isomer is calculated by known methods. The preparation of mirror-isomerically enriched forms is also well known in the art and can be achieved, for example, by palmar separation via chromatography or via palmar salt formation. Furthermore, this invention covers different configurational isomers and different rotational isomers. A configurational isomer is one that can be distinguished by rotating about one or more σ bonds. A rotational isomer is one that can be distinguished by rotating only around a single σ bond. Moreover, the compounds of this invention may also contain atomic isotopes in non-natural proportions at one or more atoms constituting such compounds. Therefore, in some embodiments, the compounds of this invention are present in an isotopically enriched form. Non-natural proportions of isotopes can be defined as those found in nature up to the amount comprising 100% of the atomic composition in question. For example, compounds may incorporate radioactive isotopes such as tritium (3H), iodine-125 (125I), or carbon-14 (14C), or non-radioactive isotopes such as deuterium (2H) or carbon-13 (13C). Such isotopic variants may provide additional utility to those described elsewhere in this application. For example, isotopic variants of the compounds of the present invention may be found to have additional utility, including but not limited to their use as diagnostic and / or imaging agents or as cytotoxic / radiotoxic therapeutic agents. Furthermore, isotopic variants of the compounds of the present invention may have altered pharmacokinetic and pharmacodynamic characteristics, which may contribute to enhanced safety, tolerability, or efficacy during treatment. All isotopic variants of the compounds of the present invention, whether or not radioactive, are intended to be included within the scope of this invention.

[0048] "CXCR7", also known as "RDC1" or "CCXCKR2", refers to a seven-transmembrane domain-preferenceed G protein-coupled receptor (GPCR). The canine ortholog of CXCR7 was first identified in 1991. See Libert et al., Science 244:569-572 (1989). The canine sequence is described in Libert et al., Nuc. Acids Res. 18(7):1917 (1990). The mouse sequence is described, for example, in Heesen et al., Immunogenetics 47:364-370 (1998). The human sequence is described, for example, in Sreedharan et al., Proc. Natl. Acad. Sci. USA 88:4986-4990 (1991), which incorrectly describes the protein as a receptor for vasoactive intestinal peptide. II. Overview

[0049] The compounds of this invention can inhibit the binding of ligands to the CXCR7 receptor and are suitable for the treatment of various diseases, including cancer, especially solid tumors and lymphoma. In recent years, it has been noted that inhibition of ligand binding to CXCR7 reduces the severity of rheumatoid arthritis in animal models.

[0050] Those skilled in the art will understand that agents modulating CCX-CKR2 activity (CXCR7 activity) can be combined in treatment regimens with other anti-angiogenic agents and / or chemotherapy agents or radiation and / or other anti-arthritis agents. In some cases, the amount of chemotherapy agent or radiation is lower than the therapeutic dose when not combined with an anti-angiogenic agent. Those skilled in the art will understand that "combination" can refer to combinations in treatment (i.e., two or more drugs can be administered in the form of a mixture, or introduced into the individual at least simultaneously or at least at different times, but such that both are simultaneously in the individual's bloodstream). Furthermore, the compositions of the present invention can be administered before or after a second treatment regimen, for example, before or after a dose of chemotherapy or irradiation. III. Embodiments of the Invention A. Compounds

[0051] In one embodiment, the present invention provides a compound having formula I. [] Or its pharmaceutically acceptable salts, hydrates, N-oxides, isotopic enrichments or mirror-image isomer enrichments or rotational isomers, wherein HAr is a five-membered aromatic ring; Ar 1 is selected from the following groups: phenyl, pyridyl, pyrimidinyl, and pyridine; Ar 2 is aryl or heteroaryl, each of which is independently a monocyclic or fused bicyclic ring; The subscript m is 0, 1, or 2; The subscript n is 0, 1, 2, or 3; The subscript p is 0, 1, 2, or 3; The subscript q is 0, 1, 2, 3, or 4; Each R1 series is independently selected from members of the following groups: halogens, CN, C1-4 alkyl, C1-4 haloalkyl, -NR aRb, -OR a, -CO 2R a, and -C(O)NR aRb; Each R2 series is independently selected from members of the following groups: halogen, CN, C1-4 alkyl, C1-4 haloalkyl, -NR aRb, -OR a, -CO 2R a, and -C(O)NR aRb; Each R3 group is selected from members of the following compositional groups: C1-4 alkyl, C1-4 haloalkyl, C1-4 hydroxyalkyl, -CO2Ra, -X-CO2Ra, -C(O)NRaRb and -XC(O)NRaRb; Each of R4a and R5a is independently selected from members of the following group: H, C1-6 alkyl, C1-6 haloalkyl, C1-6 hydroxyalkyl, -X-ORa, -CO2Ra, -X-CO2Ra, -X-NRaRb, -C(O)NRaRb and -XC(O)NRaRb; Each of R4 and R5 is independently selected from a member of the group consisting of: H, C1-6 alkyl, C1-6 haloalkyl, C1-6 hydroxyalkyl, -X-ORa, -CO2Ra, -X-CO2Ra, -X-NRaRb, -C(O)NRaRb, and -XC(O)NRaRb; or R4 and R5 are combined to form a three- to five-membered ring having 0 or 1 heteroatom ring vertices selected from O, S, or N, wherein the three- to five-membered ring is unsubstituted or substituted with 1 to 4 substituents independently selected from the group consisting of: halogen, CN, C1-4 alkyl, C1-4 haloalkyl, C1-4 alkoxy, and C1-4 haloalkoxy; Each R6 group is independently selected from members of the following groups: halogen, CN, -X-CN, C1-4 alkyl, C1-4 haloalkyl, C3-6 cycloalkyl, C1-4 hydroxyalkyl, -ORa, -CO2Ra, -X-CO2Ra, -NRaRb, -X-NRaRb, -C(O)NRaRb and -XC(O)NRaRb. R7 is a member selected from the following groups: C1-8 alkyl, C3-8 hydroxyalkyl, C1-4 alkoxy-C2-4 alkyl, -C(O)NH-C1-8 alkyl, -C(O)-C1-8 alkyl, -S(O)2-C1-8 alkyl, C3-8 cycloalkyl, -XC3-8 cycloalkyl, C6-9 spirocycloalkyl, -XC6-9 spirocycloalkyl, 4- to 7-membered heterocycloalkyl, -X-4- to 7-membered heterocycloalkyl, 7- to 11-membered spiroheterocycloalkyl and -X-7- to 11-membered spiroheterocycloalkyl, wherein each R7 is substituted with zero to four independent substituents selected from the following groups: hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy, ethoxy and cyclopropyl; Each Ra and Rb is independently selected from the group consisting of: H, C1-4 alkyl, C1-4 haloalkyl, C3-6 cycloalkyl, and C3-6 cycloalkyl-C1-4 alkyl; and Each of the X series C1-4 alkyl-linked groups, wherein any one of the methylene moieties of X is unsubstituted or substituted with one or two methyl groups. []

[0052] In one set of embodiments, compounds having Formula I are provided, wherein HAr is selected from the group consisting of: isothiazolium, isothiazole, imidazole, pyrazole, thiazole, 1,2,4-isodiazole, 1,3,4-isodiazole, 1,2,4-thiadiazole, 1,3,4-thiadiazole, 1,2,3-triazole, and 1,2,4-triazole. In another set of embodiments, HAr is selected from isothiazolium and thiadiazole.

[0053] In one set of embodiments, a compound having formula I is provided, wherein Ar 1 is a phenyl group.

[0054] In one set of embodiments, compounds having Formula I are provided, as well as the other embodiments described above, wherein the subscript q is 1, 2, or 3; and each R1 is independently selected from a group consisting of halogens, CN, C1-4 alkyl, and C1-4 haloalkyl.

[0055] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein Ar 2 is selected from the group consisting of: pyridinyl, pyrimidinyl, pyridyl, phenyl, indolyl, thiazolyl, pyrazolyl, indazole, and pyrrolopyridinyl. In other embodiments, Ar 2 is selected from pyrimidinyl, pyridinyl, and phenyl.

[0056] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein Ar 2 is selected from the group consisting of 2-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl, 2-thiazolyl, 4-pyrazolyl, phenyl, and indole; and each R 6 is independently selected from the group consisting of halogen, CN, C1-4 alkyl, C1-4 haloalkyl, C3-6 cycloalkyl, and C1-4 alkoxy.

[0057] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein -Ar2-(R6)p is selected from the group consisting of: .

[0058] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein R 7 is selected from the group consisting of: .

[0059] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein the subscript m is 0.

[0060] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein the subscript n is 0.

[0061] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein the subscript p is 0, 1 or 2.

[0062] In another set of embodiments, compounds having Formula I and the other embodiments described above are provided, wherein the subscript q is 1 or 2.

[0063] In another set of embodiments, compounds having formula I are provided, as shown in formula (Ia): , Or a pharmaceutically acceptable salt thereof, wherein the variable has the meaning provided for Formula I or any of the above embodiments. In some selected embodiments, HAr is isoflavone or thiadiazole.

[0064] In another set of embodiments, compounds having formula I are provided, as shown in formula (Ia1): , Or a pharmaceutically acceptable salt thereof, wherein the variable has the meaning provided for Formula I or any of the above embodiments. In some selected embodiments, HAr is isoflavone or thiadiazole.

[0065] In another set of embodiments, compounds having formula I are provided, as shown in formula (Ia2): , Or a pharmaceutically acceptable salt thereof, wherein the variable has the meaning provided for Formula I or any of the above embodiments. In some selected embodiments, HAr is isoflavone or thiadiazole.

[0066] In another set of embodiments, compounds having formula I are provided, such as those shown in formula (Ib), formula (Ic), or formula (Id): , Or a pharmaceutically acceptable salt thereof, wherein the variable has the meaning provided for Formula I or any of the above embodiments. In some selected embodiments, HAr is isoflavone or thiadiazole.

[0067] In another set of embodiments, compounds having formula I are provided, as shown in formula (Ib1), (Ic1), or (Id1): , Or a pharmaceutically acceptable salt thereof, wherein the variable has the meaning provided for Formula I or any of the above embodiments. In some selected embodiments, HAr is isoflavone or thiadiazole.

[0068] In another set of embodiments, compounds having Formula I and the other formulas and embodiments described above are provided, wherein R7 is a member selected from the group consisting of: C1-8 alkyl, C3-8 hydroxyalkyl, C1-4 alkoxy-C2-4 alkyl, C3-8 cycloalkyl, C6-9 spirocycloalkyl, 4- to 7-membered heterocycloalkyl and 7- to 11-membered spiroheterocycloalkyl, wherein each R7 is substituted with zero to four substituents independently selected from the group consisting of: hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy, ethoxy and cyclopropyl.

[0069] In another set of embodiments, compounds having Formula I and the other formulas and embodiments described above are provided, wherein R7 is a member selected from the group consisting of: -XC 3-8 cycloalkyl, -XC 6-9 spirocycloalkyl, -X-4 to 7 heterocycloalkyl and -X-7 to 11 spiroheterocycloalkyl, wherein each R7 is substituted with zero to four substituents independently selected from the group consisting of: hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy, ethoxy and cyclopropyl.

[0070] In another set of embodiments, compounds having Formula I and the other formulas and embodiments described above are provided, wherein R 7 is a member selected from the group consisting of cyclohexyl, cyclopentyl, piperidinyl, tetrahydropiperanyl and tetrahydrofuranyl, each of which is substituted with zero to two independent substituents selected from the group consisting of hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy and ethoxy.

[0071] In one of the selected group of embodiments, the compounds are selected from the examples below or the compounds provided in Table 1.

[0072] In other selected embodiments, the mentioned compounds may be present in a pharmaceutically acceptable salt or hydrate form.

[0073] Furthermore, regarding the non-stereochemical compounds shown above, the present invention also relates to the palmar form of each of these compounds and the mirror-isomer enriched form of the mentioned compounds. The mirror-isomer enriched form can be prepared using palmar chromatography, according to well-known methods practiced in this art, or, for example, by palmar separation using palmar salt forms. In some embodiments, the mirror-isomer enriched form has a mirror-isomer excess of at least 10%, 20%, 30%, 40%, 50%, 60%, or more. In other embodiments, a mirror-isomer enriched form of at least 70%, 80%, 90%, 95%, or higher is provided. Preparation of compounds

[0074] The compounds of the present invention can be prepared according to the methods described in the Examples section herein. Furthermore, the synthesis of certain intermediate compounds that facilitate the preparation of the compounds of the present invention is also described. B. Composition

[0075] In addition to the compounds provided above, compositions for modulating CXCR7 activity in humans and animals will typically contain pharmaceutical carriers or diluents.

[0076] As used herein, the term "composition" is intended to cover a product containing a specified amount of a specified ingredient, and any product produced directly or indirectly from a combination of a specified amount of the specified ingredient. "Pharmaceutically acceptable" means that the carrier, diluent, or excipient must be compatible with the other components of the formulation and harmless to its recipient.

[0077] Pharmaceutical compositions for administering the compounds of the present invention are preferably presented in unit dosage forms and can be prepared by any of the methods well known in pharmaceutical science and drug delivery technology. All methods include the step of binding the active ingredient with a carrier constituting one or more adjuncts. Generally, pharmaceutical compositions are prepared by uniformly and tightly binding the active ingredient with a liquid carrier or a finely powdered solid carrier, or both, and then, if necessary, shaping the product into the desired formulation. In the pharmaceutical composition, the amount of the active target compound included is sufficient to exert the desired effect on the course or condition of a disease.

[0078] Pharmaceutical compositions containing active ingredients may be in forms suitable for oral use, such as tablets, sugar-coated tablets, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions and self-emulsifiers (as described in U.S. Patent Application 2002-0012680), hard capsules or soft capsules, syrups, elixirs, solutions, oral patches, oral gels, chewable tablets, effervescent powders, and effervescent tablets. Compositions intended for oral use may be prepared according to any method known in this art for manufacturing pharmaceutical compositions, and such compositions may contain one or more agents selected from the group consisting of sweeteners, flavoring agents, coloring agents, antioxidants, and preservatives to provide a pharmaceutically refined and palatable formulation. Tablets contain the active ingredient mixed with pharmaceutically acceptable, non-toxic excipients suitable for the manufacture of tablets. These excipients may be, for example, inert diluents such as cellulose, silicon dioxide, alumina, calcium carbonate, sodium carbonate, glucose, mannitol, sorbitol, lactose, calcium phosphate, or sodium phosphate; granulating and disintegrants such as corn starch or alginate; binders such as PVP, cellulose, PEG, starch, gelatin, or gum arabic; and lubricants such as magnesium stearate, stearic acid, or talc. The tablets may be uncoated, or may be coated using known techniques (enteric-coated or otherwise) to delay disintegration and absorption in the gastrointestinal tract, thereby providing sustained action over a longer period. For example, time-delaying materials such as glyceryl monostearate or glyceryl distearate may be used. They may also be coated using techniques described in U.S. Patents 4,256,108, 4,166,452, and 4,265,874 to form osmotic therapeutic tablets for controlled release.

[0079] Formulations intended for oral use may also be presented in hard gelatin capsules, wherein the active ingredient is mixed with an inert solid diluent (e.g., calcium carbonate, calcium phosphate, or kaolin); or in soft gelatin capsules, wherein the active ingredient is mixed with water or an oily medium (e.g., peanut oil, liquid paraffin, or olive oil). Additionally, emulsions may be prepared using non-aqueous miscible components (such as oils) and stabilized using surfactants (such as glyceryl monosodium phosphate, PEG esters, and their analogues).

[0080] Aqueous suspensions contain active materials mixed with excipients suitable for manufacturing aqueous suspensions. Such excipients are suspending agents, such as sodium carboxymethyl cellulose, methyl cellulose, hydroxypropyl methyl cellulose, sodium alginate, polyvinylpyrrolidone, tragacanth gum, and gum arabic; dispersants or wetting agents may be naturally occurring phospholipids, such as lecithin, or condensation products of alkyl esters and fatty acids, such as polyoxyethylene stearate, or condensation products of ethylene oxide and long-chain aliphatic alcohols, such as heptadecanethoxylated cetyl alcohol, or condensation products of ethylene oxide and esters derived from fatty acids and hexitols (such as polyoxyethylene sorbitan monooleate), or condensation products of ethylene oxide and esters derived from fatty acids and hexitol anhydrides (such as polyvinyl sorbitan monooleate). Aqueous suspensions may also contain one or more preservatives, such as ethylparaben or n-propylparaben; one or more colorants; one or more flavorings; and one or more sweeteners, such as sucrose or saccharin.

[0081] Oily suspensions can be formulated by suspending the active ingredients in vegetable oils (such as peanut oil, olive oil, sesame oil, or coconut oil) or mineral oils (such as liquid paraffin). Oily suspensions may contain thickeners such as beeswax, hard paraffin, or cetyl alcohol. Sweeteners (such as those mentioned above) and flavoring agents may be added to provide a palatable oral formulation. These compositions may be preserved by adding antioxidants such as ascorbic acid.

[0082] Dispersants and granules suitable for preparing aqueous suspensions by adding water provide active ingredients that can be mixed with dispersants or wetting agents, suspending agents, and one or more preservatives. Suitable dispersants or wetting agents and suspending agents are exemplified by the reagents mentioned above. Other excipients, such as sweeteners, flavoring agents, and colorants, may also be present.

[0083] The pharmaceutical compositions of this invention may also be in the form of an oil-in-water emulsion. The oil phase may be a vegetable oil, such as olive oil or peanut oil; or a mineral oil, such as liquid paraffin; or a mixture thereof. Suitable emulsifiers may be naturally occurring gums, such as gum arabic or tragacanth; naturally occurring phospholipids, such as soybean or lecithin; and esters or metaesters derived from fatty acids and hexitanic anhydrides, such as sorbitan monooleate; and condensation products of such metaesters and ethylene oxide, such as polyoxyethylene sorbitan monooleate. The emulsion may also contain sweeteners and flavorings.

[0084] Syrups and elixirs can be formulated using sweeteners such as glycerin, propylene glycol, sorbitol, or sucrose. These formulations may also contain modifiers, preservatives, flavoring agents, and colorants. Oral solutions can be prepared in combination with, for example, cyclodextrin, PEG, and surfactants.

[0085] Pharmaceutical compositions may be in the form of sterile injectable aqueous solutions or oily suspensions. These suspensions may be formulated using suitable dispersants or wetting agents and suspending agents mentioned above, according to known techniques. Sterile injectable formulations may also be sterile injectable solutions or suspensions in non-toxic, non-enteric-acceptable diluents or solvents, such as solutions in 1,3-butanediol. Among acceptable mediators and solvents, water, Ringer's solution, and isotonic sodium chloride solution may be used. Additionally, sterile, non-volatile oils are commonly used as solvents or suspension media. For this purpose, any mild, non-volatile oil may be used, including synthetic monoglycerides or diglycerides. Furthermore, fatty acids such as oleic acid may be used in the preparation of injectable formulations.

[0086] The compounds of the present invention can also be used for rectal administration in suppository form. These compositions can be prepared by mixing the drug with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature and thus melts in the rectum to release the drug. Such materials include cocoa butter and polyethylene glycol. Alternatively, the compounds can be administered via ocular delivery in the form of a solution or ointment. Furthermore, percutaneous delivery of the compounds of the present invention can be achieved using iontophoresis patches and the like. For topical application, creams, ointments, gels, solutions, or suspensions containing the compounds of the present invention can be used. As used herein, topical application also means the use of mouthwash and mouthwashes.

[0087] The compounds of this invention can also be coupled to carriers of suitable polymers used as targeted drug delivery systems. Such polymers may include polyvinylpyrrolidone, piperan copolymers, polyhydroxypropyl-methacrylamide-phenol, polyhydroxyethyl-aspartamide-phenol, or polyethylene oxide-polylysine substituted with palmityl residues. Furthermore, the compounds of this invention can be coupled to carriers that are biodegradable polymers suitable for controlled drug release, such as polylactic acid, polyglycolic acid, copolymers of polylactic acid and polyglycolic acid, polycaprolactone, polyhydroxybutyric acid, polyorthoesters, polyacetals, polydihydropiperanone, polycyanoacrylates, and crosslinked or amphoteric block copolymers of hydrogels. The polymers and semi-permeable polymer matrices can be formed into molded articles such as valves, stents, catheters, prostheses, and the like. C. Usage Method

[0088] While not wishing to be bound by any particular theory, it is believed that the compounds and compositions of the present invention provide therapeutic effects by inhibiting the binding of SDF-1 and / or I-TAC to the CXCR7 receptor. Therefore, the compounds and compositions of the present invention can be used to treat or prevent diseases or conditions in mammals, wherein inhibiting the binding of SDF-1 and / or I-TAC to the CXCR7 receptor provides therapeutic effects.

[0089] In one embodiment, a preferred method for inhibiting the binding of chemokine SDF-1 and / or I-TAC to the CXCR7 receptor includes contacting one or more of the previously mentioned compounds with cells expressing the CXCR7 receptor for a duration sufficient to inhibit the binding of the chemokines to the CXCR7 receptor.

[0090] In some embodiments, the compounds and compositions of the present invention are administered to an individual suffering from cancer. In some cases, CXCR7 modulators are administered to treat cancers such as carcinomas, gliomas, mesotheliomas, melanomas, lymphomas, leukemias (including acute lymphoblastic leukemia), adenocarcinomas, breast cancers, ovarian cancers, cervical cancers, glioblastomas, leukemias, lymphomas, prostate cancers and Burkitt's lymphomas, head and neck cancers, colorectal cancers, non-small cell lung cancers, small cell lung cancers, esophageal cancers, gastric cancers, pancreatic cancers, hepatobiliary cancers, gallbladder cancers, small bowel cancers, rectal cancers, kidney cancers, renal cell carcinomas, bladder cancers, prostate cancers, penile cancers, urethral cancers, testicular cancers, cervical cancers, vaginal cancers, uterine cancers, ovarian cancers, thyroid cancers, parathyroid cancers, adrenal cancers, pancreatic endocrine carcinomas, carcinoid tumors, bone cancers, skin cancers, retinoblastomas, and Hodgkin's lymphomas. Lymphoma, non-Hodgkin's lymphoma (for other cancers see CANCER: PRINCIPLES AND PRACTICE (DeVita, VT et al., eds., 1997)); and brain and neuronal dysfunction, such as Alzheimer's disease, multiple sclerosis, and demyelinating diseases; hypertensive conditions, such as pulmonary hypertension; kidney dysfunction; renal dysfunction; rheumatoid arthritis; allogeneic transplant rejection; atherosclerosis (and elevated cholesterol); asthma; glomerulonephritis; contact dermatitis; inflammatory bowel disease; colitis; psoriasis; reperfusion injury; and other conditions and diseases described herein. In some embodiments, the individual does not have Kaposi's sarcoma, multicentric Castleman's disease, or AIDS-related primary exudative lymphoma.

[0091] This invention also covers reducing angiogenesis in any individual in need by applying the compounds and compositions of this invention. For example, reducing CXCR7 activity by contacting it with the compounds of this invention, thereby reducing angiogenesis, can be used to inhibit tumor (especially solid tumors) formation, growth, and / or metastasis. Descriptions of embodiments relating to the regulation of CXCR7 and angiogenesis are described, for example, in U.S. Patent Application No. 11 / 050,345.

[0092] Other conditions involving unwanted or problematic angiogenesis include rheumatoid arthritis; psoriasis; ocular angiogenesis disorders such as diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal transplant rejection, neovascular glaucoma, retrolental fibrosis, and iridochromia; Osler-Webber syndrome; myocardial angiogenesis; plaque neovascularization; telangiectasia; hemophilic arthritis; angiofibroma; and diseases with excessive or abnormal stimulation of endothelial cells, including intestinal adhesions, Crohn's disease, skin diseases (such as psoriasis, eczema, and scleroderma), diabetes, diabetic retinopathy, retinopathy of prematurity, age-related macular degeneration, atherosclerosis, scleroderma, wound granulation tissue, and hypertrophic scars, i.e., keloids, as well as diseases with angiogenesis as a pathological consequence, such as cat scratch disease and ulcers (Helicobacter pylori), which can also be treated with the antibodies of this invention. Angiogenesis inhibitors can be used to prevent or inhibit adhesions, especially intraperitoneal or pelvic adhesions, such as those resulting from open or laparoscopic surgery and hip contractions. Other conditions for which angiogenesis inhibitors should be advantageously treated include prevention of post-transplant scarring, cirrhosis, pulmonary fibrosis following acute respiratory distress syndrome or other pulmonary fibrosis in newborns, temporary prosthesis implantation, and adhesions following surgery between the brain and dura mater. Endometriosis, polyposis, cardiac hypertrophy, and obesity can also be treated by inhibiting angiogenesis. These conditions may involve an increase in the size or growth of other types of normal tissue, such as uterine fibroids, benign prostatic hyperplasia, and amyloidosis. The compounds and compositions of the present invention can be used to prevent or treat any of the conditions or diseases described herein.

[0093] The compounds and compositions of this invention, when used to reduce CXCR7 activity, can also be used to prevent angiogenesis for the effective treatment of a wide range of conditions. Therefore, for example, reducing angiogenesis can be used as part of the treatment of vascular conditions (e.g., hemangiomas and capillary proliferation within atherosclerotic plaques), muscle diseases (e.g., myocardial angiogenesis, myocardial infarction, or smooth muscle angiogenesis), joint conditions (e.g., arthritis, hemophilic arthritis, etc.), and other conditions associated with angiogenesis. Promoting angiogenesis can also help accelerate various physiological processes and the treatment of diseases requiring increased angiogenesis, such as wound healing, fractures and burns, inflammatory diseases, ischemic heart disease, and peripheral vascular disease. The compounds of this invention can also provide benefits in conditions where normal blood flow is restricted (such as pulmonary hypertension).

[0094] The compounds and compositions of this invention can also be used to promote wound healing. It is not intended to limit this invention to a specific mechanism of action; it is possible that CXCR7 antagonism causes endogenous ligands to bind to lower-affinity receptors, thereby triggering enhanced wound healing. For example, SDF-1 binds to both CXCR7 and CXCR4, but binds to CXCR4 with lower affinity. Similarly, I-TAC binds to CXCR3 with lower affinity than I-TAC binds to CXCR7. By preventing the binding of these ligands to CXCR7, CXCR7 antagonists allow the ligands to bind to other receptors, thereby promoting wound healing. Therefore, the antagonistic effect of CXCR7 in promoting wound healing can be mediated through a different mechanism than the mechanism by which activating CXCR7 activity promotes wound healing with agonists.

[0095] In addition to treating conditions and symptoms related to angiogenesis, inhibition of angiogenesis can be used to regulate or prevent the occurrence of normal physiological conditions associated with angiogenesis. Therefore, for example, compounds and compositions can be used for contraception. According to the present invention, reducing CXCR7 activity in the ovaries or endometrium can reduce angiogenesis associated with ovulation, embryo implantation, placental formation, etc.

[0096] Angiogenesis inhibitors have other therapeutic uses. For example, the compounds and compositions of the present invention can be used in the following: (a) Adipose tissue ablation and treatment of obesity. See, for example, Kolonin et al., Nature Medicine 10(6):625-632 (2004); (b) Treatment of preeclampsia. See, for example, Levine et al., N. Engl. J. Med. 350(7): 672-683 (2004); Maynard et al., J. Clin. Invest. 111(5): 649-658 (2003); (c) Treatment of cardiovascular diseases. See, for example, March et al., Am. J. Physiol. Heart Circ. Physiol. 287:H458-H463 (2004); Rehman et al., Circulation 109: 1292-1298 (2004); (d) Treatment / prevention of local ischemia / reperfusion injury by inhibiting hypoxic preconditioning of mesenchymal stem cells and rescuing locally ischemic tissues (including but not limited to myocardium and transplanted kidney tissue), and see, for example, Liu, H. et al., PLoS One (2012) 7:e34608 and Zhang, S. et al., Biomed Pharmacother (2020) 127:110168; (e) Treatment of Shiga toxin-associated hemolytic uremic syndrome, see, for example, Petruzziello-Pellegrini, TN et al., J Clin Invest (2012) 122:759-776; (f) Treatment of pain, anxiety, stress, opioid addiction and other CNS conditions, see, for example, Meyrath M. et al., Nat Commun (2020) 19:3033 and Zhu, Y. et al.; (g) Treatment / prevention of acute renal failure by blocking harmful migration of renal progenitor cells within the glomerulus; see, for example, Mazzinghi, B, et al., J Exp Med (2008) 205:479-490 and Romaol, S. et al., Kidney Int (2018) 94:1111-1126; (h) Treatment of neuroinflammatory conditions, including but not limited to multiple sclerosis (MS), see, for example, Williams, JL et al., Glia (2020) 68:1361-1374. Methods of treating cancer

[0097] More specifically, the present invention also provides a method for treating cancer. A preferred method for treating cancer includes administering to a cancer patient a therapeutically effective amount of one or more of the aforementioned compounds (or salts thereof) for a duration sufficient to treat the cancer.

[0098] For treatment, the compositions of the present invention can be administered orally, non-enterically (e.g., intramuscular, intraperitoneal, intravenous, ICV, intracisional injection or infusion, subcutaneous injection or implantation), by inhalation spray, nasal, vaginal, rectal, sublingual or local administration, and can be formulated alone or in combination into suitable dosage units containing known non-toxic and medically acceptable carriers, adjuvants and mediators suitable for each administration route.

[0099] In some embodiments, the CXCR7 modulator of the present invention can be administered in combination with other suitable therapeutic agents (including, for example, chemotherapy agents, radiation, etc.). It should be understood that such administration can be performed before, after, or simultaneously with a second therapeutic agent, such that the therapeutic effect of the second agent is enhanced compared to administration of the second agent in the absence of the CXCR7 modulator. The selection of suitable agents for combination therapy can be made by those skilled in the art based on known medical principles. The combination of therapeutic agents can work synergistically to treat or prevent various conditions, such as cancer, trauma, renal dysfunction, brain dysfunction, or neuronal dysfunction. Using this approach, therapeutic efficacy can be achieved with lower doses of each agent, thus reducing the likelihood of adverse side effects.

[0100] Besides primates (such as humans), various other mammals can be treated according to the method of the present invention. For example, mammals including, but not limited to, cows, sheep, goats, horses, dogs, cats, guinea pigs, rats or other bovine, sheep, equines, canines, felines, rodents or rat species can be treated. However, the method can also be practiced in other species, such as avian species (e.g., chickens).

[0101] In the treatment or prevention of conditions requiring chemokine receptor regulation, the appropriate dose concentration will generally be from about 0.001 mg to 100 mg per kilogram of patient body weight per day, which may be administered in a single or multiple doses. Preferably, the dose concentration will be from about 0.01 mg / kg to about 25 mg / kg per day; more preferably from about 0.05 mg / kg to about 10 mg / kg per day. Suitable dose concentrations may be from about 0.01 mg / kg to 25 mg / kg per day, from about 0.05 mg / kg to 10 mg / kg per day, or from about 0.1 mg / kg to 5 mg / kg per day. Within this range, the dose may be from 0.005 mg / kg to 0.05 mg / kg, from 0.05 mg / kg to 0.5 mg / kg, or from 0.5 mg / kg to 5.0 mg / kg per day. For oral administration, the composition is preferably provided in tablet form, such tablets containing 1.0 to 1000 mg of active ingredient, particularly 1.0, 5.0, 10.0, 15.0, 20.0, 25.0, 50.0, 75.0, 100.0, 150.0, 200.0, 250.0, 300.0, 400.0, 500.0, 600.0, 750.0, 800.0, 900.0 and 1000.0 mg of active ingredient, to achieve dose-modulation for the patient being treated. The compound can be administered 1 to 4 times daily, preferably once or twice daily.

[0102] However, it will be understood that the specific dose concentration and frequency of administration for any particular patient may vary and will depend on various factors including the activity of the specific compound used, the metabolic stability and duration of action of the compound, the individual's age, weight, genetic characteristics, general health status, sex and diet, as well as the mode and time of administration, excretion rate, drug combination and the severity of the individual's specific condition.

[0103] The compounds and compositions of this invention can be combined with other compounds and compositions, and have therapeutic effects in preventing and treating cancer and diseases or conditions related to CXCR7 signaling. Such other drugs can be administered simultaneously or sequentially with the compounds or compositions of this invention via their usual routes and in their usual amounts. When the compounds or compositions of this invention are used simultaneously with one or more other drugs, pharmaceutical compositions containing such other drugs besides the compounds or compositions of this invention are preferred. Therefore, pharmaceutical compositions of this invention include compositions containing one or more other active ingredients or therapeutic agents besides the compounds or compositions of this invention. Examples of other therapeutic agents that can be combined with, separately from, or added to the compounds or compositions of the present invention in the same pharmaceutical composition include, but are not limited to: cisplatin, paclitaxel, methotrexate, cyclophosphamide, ifosfamide, chlorambucil, carmustine, carboplatin, vincristine, vinblastine, thiotepa, lomustine, semustine, 5-fluorouracil, and cytarabine. The weight ratio of the compounds of the present invention to the second active ingredient may vary and will depend on the effective dose of each ingredient. Generally, the effective dose of each will be used. Therefore, for example, when the compounds of the present invention are combined with a second anticancer agent, the weight ratio of the compounds of the present invention to the second agent is generally in the range of about 1000:1 to about 1:1000, preferably about 200:1 to about 1:200. Combinations of the compounds of the present invention with other active ingredients are also generally within the aforementioned range, but in each case, the effective dose of each active ingredient should be used. Methods for treating inflammation

[0104] Furthermore, the compounds and compositions of the present invention are suitable for treating inflammation and can be combined with other compounds and compositions having therapeutic effects, which may be necessary before, after, or simultaneously with the treatment of cancer or inflammation using the compounds of the present invention. Therefore, the combination methods and compositions are also components of the present invention for the prevention and treatment of related conditions or diseases, such as inflammatory or autoimmune conditions, illnesses, and diseases, including inflammatory bowel disease, rheumatoid arthritis, osteoarthritis, psoriatic arthritis, polyarthritis, multiple sclerosis, allergic diseases, psoriasis, atopic dermatitis, and asthma, and the aforementioned lesions.

[0105] For example, in the treatment or prevention of inflammation, autoimmune disorders, or, for example, arthritis-related bone loss, the compounds and compositions of the present invention can be used in combination with the following drugs: anti-inflammatory agents or analgesics, such as opiate agonists; lipid oxygenase inhibitors, such as inhibitors of 5-lipoxygenase; cyclooxygenase inhibitors, such as cyclooxygenase-2 inhibitors; interleukin inhibitors, such as interleukin-1 inhibitors; NMDA antagonists; nitric oxide inhibitors or inhibitors of nitric oxide synthesis; nonsteroidal anti-inflammatory agents; or interleukin-inhibiting anti-inflammatory agents, such as compounds, such as acetaminophen. Phenol, aspirin, codeine, fentanyl, ibuprofen, indomethacin, ketorolac, morphine, naproxen, phenacetin, piroxicam, steroidal analgesics, sufentanyl, sunlindac, tenidap, and their analogues. Similarly, the compounds and compositions of the present invention may be administered together with the following: analgesics listed above; enhancers, such as caffeine, H2 antagonists (e.g., ranitidine), polydimethicone, aluminum hydroxide, or magnesium hydroxide; decongestants, such as phenylephrine, phenylpropanolamine, pseudoephedrine, oxymetazoline, epinephrine, naphazoline, xylometazoline, cyclohexylpropyl methylamine, or levodeoxyephedrine; antitussives, such as codeine, hydrocodone, caramiphen, carbetapentane, or dextramethorphan; diuretics; and sedative or non-sedative antihistamines.

[0106] As indicated, the compounds and compositions of the present invention can be used in combination with other medicaments for treating, preventing, inhibiting, or improving the diseases or conditions to which the compounds and compositions of the present invention are applicable. Such other medicaments can be administered simultaneously or sequentially with the compounds or compositions of the present invention via their usual route and in their usual amounts. When the compounds or compositions of the present invention are used simultaneously with one or more other medicaments, pharmaceutical compositions containing such other medicaments besides the compounds or compositions of the present invention are preferred. Therefore, pharmaceutical compositions of the present invention include compositions that, in addition to the compounds or compositions of the present invention, also contain one or more other active ingredients or therapeutic agents. Examples of other therapeutic agents that can be combined with, separately from, or administered in the same pharmaceutical composition with the compounds or compositions of the present invention include, but are not limited to: (a) VLA-4 antagonists; (b) corticosteroids, such as beclomethasone, methylprednisolone, betamethasone, prednisone, dexamethasone, fluticasone, hydrocortisone, budesonide, triamcinolone, salmeterol, salbutamol, and formeterol; and (c) immunosuppressants, such as cyclosporine (Cyclosporine A, Sandimmune®, Neoral®), tacrolirnus (FK-506, Prograf®), and rapamycin. (sirolimus, Rapamune®) and other FK-506 immunosuppressants, and mycophenolic esters, such as mycophenolic morpholine ethyl ester (CellCept®); (d) antihistamines (H1-histamine antagonists),Such as bromopheniramine, chlorpheniramine, dexchloipheniramine, triprolidine, clemastine, diphenhydramine, diphenylpyraline, tripelennamine, hydroxyzine, methdilazine, promethazine, trimeprazine, azatadine, cyproheptadine, antazoline, and pheniramine. (e) pyrilamine, astemizole, terfenadine, loratadine, cetirizine, fexofenadine, descarboethoxyloratadine and their analogues; (e) nonsteroidal antiasthmatic drugs (e.g., terbutaline, isoproterenol, fenoterol, isoetharine, albuterol, bitolterol and pirbuterol), theophylline, sodium cromoglycate, atropine, ipratropium bromide (f) Leukotriene antagonists (e.g., zafmlukast, montelukast, pranlukast, iralukast, pobilukast, and SKB-106,203), leukotriene biosynthesis inhibitors (zileuton, BAY-1005); and nonsteroidal anti-inflammatory drugs (NSAIDs), such as propionic acid derivatives (e.g., bromide), leukotriene antagonists (e.g., zafmlukast, montelukast, pranlukast, iralukast, pobilukast, and SKB-106,203), leukotriene biosynthesis inhibitors (zileuton, BAY-1005);Alminoprofen, benzoxaprofen, bucloxic acid, carprofen, fenbufen, fenoprofen, fluprofen, flurbiprofen, ibuprofen, indoprofen, ketoprofen, iriroprofen, naproxen, oxaprozin, pirprofen, pranoprofen, suprafen, tiaprofenic acid Acetic acid derivatives (e.g., indomethacin, acemetacin, alclofenac, clidanac, diclofenac, fenclofenac, fenclozic acid, fentiazac, furofenac, ibufenac, isoxepac, oxpinac, sulindac, tiopinac, tolmetin, zidometacin, and zomepirac), and fenamic acid derivatives (e.g., flufenamic acid, meclofenamic acid, mefenamic acid, niflumic acid). (g) cyclooxygenase-2 (COX-2) inhibitors, and tolfenamic acid; diphenylcarboxylic acid derivatives (e.g., diflunisal and flufenisal); oxicam (e.g., isoxicam, piroxicam, sudoxicam, and tenoxicam); salicylic acid (e.g., acetyl salicylate and sulfasalazine); and pyrazolone (e.g., apazone, bezpiperylon, feprazone, mofebutazone, oxyphenbutazone, and phenylbutazone); and cyclooxygenase-2 (COX-2) inhibitors.Such as celecoxib (Celebrex®) and rofecoxib (Vioxx®); (h) phosphodiesterase type IV (PDE IV) inhibitors; (i) gold compounds, such as auranofin and glucosinolate; (j) etanercept (Enbrel®); (k) antibody therapeutics, such as orthoclone (OKT3), daclizumab (Zenapax®), basiliximab (Simulect®), and infliximab (Remicade®); (l) other chemokine receptors, especially CCR5, CXCR2, CXCR3, CCR2, CCR3, CCR4, CCR7, and CXCR2. Antagonists of 3CR1 and CXCR6; (m) lubricants or emollients, such as petrolatum and lanolin; (n) keratolytic agents (e.g., tazarotene); (o) vitamin D3 derivatives, such as calcipotriene or calcipotriol (Dovonex®); (p) PUVA; (q) phloroglucinol (Drithrocreme®); (r) etretinate (Tegison®) and isotretinoin; and (s) multiple sclerosis treatment agents, such as interferon β-1β (Betaseron®), interferon β-1α (Avonex®), azathioprine (Imurek®, Imuran®), glatiramer acetate (Capoxone®), glucocorticoids (e.g., presbyron), and cyclophosphamide; (t) DMARDs, such as methotrexate(u); other compounds, such as 5-aminosalicylic acid and its prodrugs; hydroxychloroquine; D-penicillamine; antimetabolites, such as azathioprine, 6-mercaptopurine, and methotrexate; DNA synthesis inhibitors, such as hydroxyurea; and microtubule disruptors, such as colchicine. The weight ratio of the compound of the present invention to the second active ingredient may vary and will depend on the effective dose of each ingredient. Generally, the effective dose of each will be used. Therefore, for example, when the compound of the present invention is combined with an NSAID, the weight ratio of the compound of the present invention to the NSAID is generally in the range of about 1000:1 to about 1:1000, preferably about 200:1 to about 1:200. The combination of the compound of the present invention with other active ingredients will also generally be within the aforementioned range, but in each case, the effective dose of each active ingredient should be used. Methods for inducing the mobilization of pioneer cells / stem cells

[0107] Furthermore, the compounds and compositions of this invention are applicable to mobilizing pioneer cells / stem cells, and are therefore suitable for treating or improving conditions or illnesses where pioneer cell / stem cell mobilization is effective or necessary, as appropriate, using the compounds of this invention according to the procedures and protocols described in WO05 / 000333, which is incorporated herein by reference in its entirety for all purposes. Conditions that can be improved or otherwise benefited include, for example, hematopoietic disorders, such as aplastic anemia, leukemia, drug-induced anemia, and hematopoietic defects resulting from chemotherapy or radiotherapy. Furthermore, the compounds and compositions of this invention can be used to enhance transplant success rates during and after immunosuppressive therapy, and to achieve more effective wound healing. Furthermore, the compounds and compositions of this invention are applicable to mobilizing pioneer cells / stem cells, and are therefore suitable for treating or improving conditions or illnesses where pioneer cell / stem cell mobilization is effective or necessary, using the compounds of this invention according to the procedures and protocols described in WO05 / 000333, which is incorporated herein by reference in its entirety for all purposes. Conditions that can be improved or otherwise benefited include, for example, hematopoietic disorders such as aplastic anemia, leukemia, drug-induced anemia, and hematopoietic defects resulting from chemotherapy or radiotherapy. Furthermore, the compounds and compositions of the present invention can be used to enhance transplant success rates during and after immunosuppressive therapy, and to achieve more effective wound healing and treatment of bacterial infections. Where appropriate, after administration of the compounds of the present invention and after pioneer cell / stem cell mobilization, blood containing mobilized cells may be collected and, where appropriate, the mobilized cells may be purified and, where appropriate, expanded, and, if necessary, reintroduced into the same or a second person (e.g., a matched donor).

[0108] Multiple cell types can be mobilized as needed. In some embodiments, hematopoietic progenitor cells (HSCs) are mobilized after administration of the compounds or compositions of the present invention, and are collected and purified from other blood components as appropriate. HSC mobilization may be induced by administration of at least one of the compounds of the present invention and one or more of granulocyte colony-stimulating factor (G-CSF) or AMD3100 (1,1'-[1,4-epenylphenylbis(methylene)]bis[1,4,8,11-tetraazacyclotetradecane]octahydrobromide dihydrate) or its salts, racemates or isomers.

[0109] In some embodiments, endothelial precursor cells (EPCs) are mobilized after administration of a compound or composition of the present invention, and are collected and purified from other blood components as appropriate. EPC mobilization may be induced by administration of at least one compound of the present invention and one or more of vascular endothelial growth factor (VEGF), a VEGF agonist (including, but not limited to, VEGF agonist antibodies), or AMD3100 or its salts, racemates, or isomers.

[0110] In some embodiments, mesenchymal stem cells (MSCs) or stromal precursor cells (SPCs) are mobilized after administration of the compounds or compositions of the present invention, and are collected and purified from other blood components, as appropriate. Such mobilization may be induced by administration of at least one compound of the present invention and one or more of G-CSF, VEGF, VEGF agonists (including but not limited to VEGF agonist antibodies), AMD3100, or their salts, racemates, or isomers.

[0111] To immobilize pioneer cells or stem cells, appropriate dosage concentrations will typically range from approximately 0.001 mg to 100 mg per kilogram of patient body weight per day, which can be administered in a single or multiple doses. The compounds can be administered in a single dose, over time (e.g., intravenously or percutaneously), or in multiple doses. The compounds of this invention can also be used in ex vivo treatment protocols to prepare cell cultures, which are subsequently used to replenish the individual's blood cells. Ex vivo treatment can be performed on autologous cells collected from peripheral blood or bone marrow, or on allogeneic grafts from matched donors.

[0112] The compounds of this invention can be combined with other compounds and compositions that induce the activation, proliferation, or mobilization of pioneer cells / stem cells. In addition to those described above, these include, but are not limited to, Fms-related tyrosine kinase 3 ligands (Flt3 ligands), interleukin-3 (IL-3), interleukin-7 (IL-7), interleukin-20 (IL-20), Steel factor (SF), and granulocyte-macrophage community-stimulating factor (GM-CSF), and can provide therapeutic efficacy that may be desired or beneficial before, after, or simultaneously with pioneer cell / stem cell mobilization. Therefore, the combination methods and compositions are also components of this invention for the prevention and treatment of conditions or diseases of interest. Furthermore, the compounds of this invention can provide benefits in conditions in which dysregulation of stem cell mobilization may play a role, such as heart disease and pulmonary hypertension. Methods for diagnosing diseases and symptoms related to CXCR7

[0113] Furthermore, the compounds and compositions of the present invention are suitable for diagnosing diseases and conditions related to CXCR7. Specifically, the compounds of the present invention can be prepared in a labeled form (e.g., radiolabeled) and used for diagnosing, for example, cancer. The labeled compounds of the present invention (e.g., antagonists or agonists) in combination with CXCR7 can be used to determine the CXCR7 content in mammalian individuals. In some embodiments, CXCR7 modulators are administered to individuals with cancer. In some cases, labeled compounds are administered to detect the development of cancers such as carcinomas, gliomas, mesotheliomas, melanomas, lymphomas, leukemias, adenocarcinomas, breast cancers, ovarian cancers, cervical cancers, glioblastomas, leukemias, lymphomas, prostate cancer and Burkitt's lymphoma, head and neck cancers, colorectal cancers, colorectal cancers, non-small cell lung cancers, small cell lung cancers, esophageal cancers, stomach cancers, pancreatic cancers, hepatobiliary cancers, gallbladder cancers, small bowel cancers, rectal cancers, kidney cancers, bladder cancers, prostate cancers, penile cancers, urethral cancers, testicular cancers, cervical cancers, vaginal cancers, uterine cancers, ovarian cancers, thyroid cancers, parathyroid cancers, adrenal cancers, pancreatic endocrine cancers, carcinoids, bone cancers, skin cancers, retinoblastomas, Hodgkin's lymphomas, and non-Hodgkin's lymphomas (for other cancers, see CANCER: PRINCIPLES AND PRACTICE (DeVita, (VT et al., 1997)); and brain and neuronal dysfunction, such as Alzheimer's disease and multiple sclerosis; renal dysfunction; rheumatoid arthritis; cardiac allogeneic transplant rejection; atherosclerosis (and elevated cholesterol); asthma; glomerulonephritis; contact dermatitis; inflammatory bowel disease; colitis; psoriasis; reperfusion injury; and other conditions and diseases described herein. In some embodiments, the individual does not have Kaposi's sarcoma, multicentric Kasoxari's disease, or AIDS-related primary exudative lymphoma. Because CXCR7 is typically expressed in cancer cells but not in non-cancerous cells, CXCR7 antagonists are often administered to individuals at risk of developing cancer.

[0114] Various imaging and detection methods can be used to detect cancer. In some embodiments, direct methods can be used to assess the biodistribution of CXCR7 in the body, such as magnetic resonance imaging (“MRI”), positron emission tomography (“PET”), and single-photon emission computed tomography (“SPECT”). Each of these methods can detect the distribution in the body of a suitable labeled compound (typically such as one bound to CXCR7) if the compound contains atoms with appropriate nuclear properties. MRI detects paramagnetic nuclei; PET and SPECT detect the emission of particles from radioactive nuclei decay.

[0115] For methods involving PET, an appropriate positron-emitting radioactive nucleus must be incorporated. Relatively few positron-emitting isotopes are suitable for labeling therapeutic agents. The carbon isotope 11C has been used in PET, but has a short half-life of 20.5 minutes. Therefore, facilities for synthesis and use are typically located near cyclotrons, where precursor 11C starting materials are generated. Another suitable isotope, 18F, has a half-life of 110 minutes. This allows sufficient time for incorporation into the radiolabeled tracer for purification and administration to human or animal individuals. Other isotopes have even shorter half-lives. 13N has a half-life of 10 minutes, and 15O has even shorter half-lives of 2 minutes. However, both emit higher energies than 11C, and these isotopes have been used in PET studies (see Clinical Positron Emission Tomography, Mosby Year Book, 1992, KF Hubner et al., Chapter 2).

[0116] SPECT imaging uses isotopic tracers as gamma emitters. While the range of applicable isotopes is greater than that of PET, SPECT imaging offers lower three-dimensional resolution. However, in some cases, SPECT is used to obtain clinically important information about compound binding, localization, and clearance. One applicable isotope for SPECT imaging is 123I, a gamma emitter with a half-life of 13.3 hours. Compounds labeled with 123I can be transported up to approximately 1000 miles from the manufacturing site, or the isotope itself can be transported for in-situ synthesis. Eighty-five percent of the isotopes emit 159 keV photons, which are easily measurable with current SPECT instruments. Other halogen isotopes can be used for PET or SPECT imaging, or for labeling with conventional tracers. These include 75Br, 76Br, 77Br, and 82Br, which have usable half-lives and emission characteristics.

[0117] In view of the above, the present invention provides a method for imaging tumors, organs, or tissues, the method comprising: (a) Applying a radiolabeled or detectable form of a Formula I compound to an individual requiring such imaging; and (b) Detect the compound to determine where it is concentrated in the individual.

[0118] In addition, the present invention provides a method for detecting elevated CXCR7 levels in a sample, the method comprising: (a) Expose a sample suspected of having elevated CXCR7 levels to a radiolabeled or detectable form of a compound of formula I; (b) To determine the content of CXCR7-bound compounds present in the sample, and to determine the content of CXCR7 present in the sample; and (c) Compare the content determined in step (b) with the control sample to determine whether there is an elevated CXCR7 content in the sample.

[0119] As described in this article, the administration of labeled compounds can be carried out via any of the routes commonly used to introduce compounds into final contact with the tissue to be evaluated, and are well known to those skilled in the art. Although more than one route may be used to administer a particular composition, a particular route often provides a more direct and effective diagnosis than another. Combination therapy

[0120] Inhibitors of CXCR7 can be supplied alone or in combination with one or more other drugs. Possible combinations may include, for example, additional anti-angiogenic factors and / or chemotherapeutic agents (e.g., cytotoxic agents) or radiation, cancer vaccines, immunomodulators, anti-angiogenic agents, signal transduction inhibitors, antiproliferators, or apoptosis inducers. IV. Examples

[0121] The following examples are provided to illustrate, rather than to limit, the inventions claimed.

[0122] The reagents and solvents used below were available from commercial sources, such as Aldrich Chemical Co. (Milwaukee, Wisconsin, USA). ¹H-NMR spectra were recorded on a Varian Mercury 400 MHz NMR spectrometer. Peaks relative to TMS are listed in the following order: multiplicity (s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet) and proton number. Mass spectrometry results are reported as mass-to-charge ratios, followed by the relative abundance of each ion (in parentheses). In the example, for M+H (or MH, as described) ions containing the most common atomic isotopes, a single m / e value is reported. In all cases, the isotopic pattern corresponds to the expected pattern. Electrospray ionization (ESI) mass spectrometry analysis was performed on a Hewlett-Packard MSD electrospray mass spectrometer using an HP1100 HPLC for sample delivery. Typically, the analyte is dissolved in methanol at 0.1 mg / mL and 1 μL of the delivery solvent is injected into the mass spectrometer, which scans from 100 Daltons to 1500 Daltons. All compounds can be analyzed in positive ESI mode using acetonitrile / water (with 1% formic acid) as the delivery solvent. Alternatively, the compounds described below can be analyzed in negative ESI mode using 2 mM NH₄OAc in acetonitrile / water as the delivery system.

[0123] The following abbreviations are used in the examples and throughout this invention specification: rt, room temperature; HPLC, high-performance liquid chromatography; TFA, trifluoroacetic acid; LC-MSD, liquid chromatography / mass selectivity detector; LC-MS, liquid chromatography / mass spectrometry; Pd 2dba 3, triphenylmethyleneacetone dipalladium; THF, tetrahydrofuran; DMF, dimethylformamide or N,N-dimethylformamide; DCM, dichloromethane; DMSO, dimethyl sulfoxide; TLC, thin-layer chromatography; KHMDS, potassium hexamethyldisilazane; ES, electrospray ionization; sat., saturation.

[0124] Compounds within the scope of this invention can be synthesized using a variety of reactions known to those skilled in the art, as described below. Those skilled in the art will also recognize that alternative methods can be used to synthesize the target compounds of this invention, and that the pathways described herein are not exhaustive, but provide a wide range of applicable and practical pathways for the related compounds.

[0125] Certain molecules claimed in this patent may exist in different mirror-image and non-mirror-image isomer forms, and all such variants of such compounds are claimed.

[0126] The detailed description of the experimental procedures used to synthesize the key compounds in this paper produces physical data to identify them, as well as molecular structures described by related structural drawings.

[0127] Those skilled in this art will also recognize that acids and bases are often used during standard processing procedures in organic chemistry. During the experimental procedures described in this patent, if they possess the desired inherent acidity or alkalinity, salts of the parent compound are sometimes produced. [Example] [1] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [Pyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0128] Step a: A mixture of 15 g (88 mmol) of tributyl 3-amino-3-(2-ethoxy-2-ethoxyethyl)acryl-1-carboxylic acid, monoethyl malonate (17 g, 130 mmol), and ammonium formate (20 g, 260 mmol) in 120 mL of EtOH was stirred at 85 °C for 3 hours. Volatiles were removed under vacuum, and the residue was diluted with Et₂O and washed with saturated NaHCO₃ (aqueous solution). The organic layer was dried over MgSO₄, filtered, and subsequently concentrated under vacuum. The crude material was purified by silica gel column chromatography to give 3-amino-3-(2-ethoxy-2-ethoxyethyl)acryl-1-carboxylic acid tributyl ester. MS: (ES) m / z C₁₂H₂₃N₂O₄[M+H]⁺ Calculated value 259.2, experimental value 259.3.

[0129] Step b: Add HATU (2.8 g, 7.4 mmol) to a solution of 3-amino-3-(2-ethoxy-2-t-oxyethyl)acetidine-1-carboxylic acid tributyl ester (1.8 g, 7.1 mmol), 5-(2,4-difluorophenyl)isothiazolyl-3-carboxylic acid tributyl ester (1.6 g, 7.1 mmol), and DIPEA (1.3 mL, 7.5 mmol) in 40 mL of DCM. Stir the mixture at room temperature for 2 hours. Quench the reactants with saturated NH4Cl (aqueous solution) and separate the phases. Extract the aqueous phase with DCM, dry the combined organic layers with MgSO4, filter, and concentrate under vacuum. The crude material was purified by silicone column chromatography to obtain tributyl 3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-3-(2-ethoxy-2-sideoxyethyl)acetidine-1-carboxylic acid. MS: (ES) m / z C 22H 26F 2N 3O 6[M+H] + Calculated value 466.2, experimental value 466.2.

[0130] Step c: Stir a mixture of 3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-3-(2-ethoxy-2-sideoxyethyl)acetidine-1-carboxylic acid tributyl ester (1.2 g, 3.3 mmol) in a 4:1 DCM / TFA (20 mL) solution for 1 hour. Concentrate the contents under vacuum to give ethyl acetate 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acetidine-3-yl). MS: (ES) m / z C 17H 18F 2N 3O 4[M+H] + Calculated value 366.1, experimental value 366.1.

[0131] Step d: Add NaBH(OAc)3 (840 mg, 4.0 mmol) to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acetidin-3-yl)acetate (800 mg, 2.0 mmol), triethylamine (0.55 mL, 4.0 mmol), and 4-hydroxy-4-methylcyclohexane-1-one (510 mg, 4.0 mmol) in a 4:1 DCM / MeOH (20 mL). Stir the contents at room temperature for 1 hour. Quench the reaction mixture with saturated NaHCO3 (aqueous solution) and extract with DCM. Dry the combined organic layers with MgSO4, filter, and concentrate under vacuum. The crude material was purified by silicone column chromatography to obtain ethyl acetate (a mixture of cis / trans isomers). MS: (ES) m / z C 24H 30F 2N 3O 5 [M+H] + Calculated value 478.2, experimental value 478.5.

[0132] Step e: Add NaOH (300 mg, 7.5 mmol) to a solution of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (600 mg, 1.2 mmol) in 1:1 THF / H₂O (20 mL). Stir the mixture overnight at room temperature. Quench the reactants with 1 N HCl (10 mL) and MeCN. Concentrate the contents under vacuum. Wet mill the residue with acetone and filter through diatomaceous earth. Concentrate the filtrate to produce 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (a mixture of cis / trans isomers). MS: (ES) m / zC 22H 26F 2N 3O 5[M+H] + Calculated value 450.2, experimental value 450.2.

[0133] Step f: To a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (100 mg, 0.21 mmol) in DCM (2 mL), 2-(pyridin-2-yl)propyl-2-amine dihydrochloride (43 mg, 0.21 mmol), DIPEA (0.14 mL, 0.82 mmol), and HATU (82 mg, 0.22 mmol) were added. The mixture was stirred for 1 hour, followed by concentration under vacuum. By preparative HPLC purification of the residue, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0134] Isomer 1: ¹H NMR (400 MHz, DMSO-d⁶) δ 8.59 ̶ 8.49 (m, ¹H), 8.22 ̶ 8.12 (m, ¹H), 8.06 (d, J = 8.1 Hz, ¹H), 7.83 ̶ 7.77 (m, ¹H), 7.67 ̶ 7.58 (m, ¹H), 7.25 (dd, J = 20.8, 9.8 Hz, 2H), 7.15 ̶ 7.07 (m, ¹H), 4.58 ̶ 4.27 (m, 4H), 3.20 - 3.11 (m, 3H), 1.83 - 1.73 (m, 2H), 1.56 ̶ 1.44 (m, 9H), 1.37 ̶ 1.15 (m, 4H), 1.21 (s, 3H). MS: (ES) m / zC 30H 36F 2N 5O 4[M+H] + Calculated value 568.3, experimental value 568.3.

[0135] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.54 ̶ 8.48 (m, 1H), 8.06 (m, 2H), 7.71 (d, J = 8.0 Hz, 1H), 7.56 ̶ 7.50 (m, 1H), 7.32 - 7.17 (m, 2H), 7.11 (dd, J = 10.4, 3.5 Hz, 1H), 4.52 (d, J = 11.8 Hz, 1H), 4.49 - 4.46 (m, 2H), 4.37 (d, J = 11.7 Hz, 1H), 3.16 - 3.10 (m, 3H), 1.80 ̶ 1.66 (m, 4H), 1.66 (s, 6H), 1.62 ± 1.52 (m, 2H), 1.46 ± 1.39 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 30H 36F 2N 5O 4[M+H] + Calculated value 568.3, experimental value 568.4. [Example] [2] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-] [Phenylacetyl] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] [] []

[0136] To a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL), 2-phenylprop-2-amine (31 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) were added. The reaction mixture was stirred at room temperature for 1 hour and then quenched with 3 drops of H₂O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-phenylprop-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0137] Isomer 1: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.61 - 9.48 (m, ¹H), 8.41 - 8.36 (s, ¹H), 8.19 - 8.07 (m, ¹H), 7.67 - 7.58 (m, ¹H), 7.42 - 7.32 (m, ¹H), 7.31 - 7.18 (m, ³H), 7.16 - 7.02 (m, ³H), 4.49 - 4.27 (m, 4H), 3.31 - 3.16 (m, ¹H), 3.14 - 3.04 (m, 2H), 1.88 - 1.73 (m, 2H), 1.61 - 1.46 (m, 8H), 1.38 - 1.15 (m, 4H), 1.12 - 1.02 (m, 3H). MS: (ES) m / zC 31H 37F 2N 4O 4[M+H] + Calculated value 567.3, experimental value 567.3.

[0138] Isomer 2: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.50 (s, ¹H), 8.36 - 8.32 (m, ¹H), 8.17 - 8.07 (m, ¹H), 7.68 - 7.59 (m, ¹H), 7.41 - 7.34 (m, ¹H), 7.31 - 7.17 (m, 3H), 7.14 - 7.02 (m, 3H), 4.46 - 4.26 (m, 4H), 3.19 - 3.01 (m, 3H), 1.73 - 1.37 (m, ¹²H), 1.30 - 1.16 (m, ²H), 1.10 (s, 3H). MS: (ES) m / zC 31H 37F 2N 4O 4[M+H] + Calculated value 567.3, experimental value 567.3. [Example] [3] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Fluorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0139] To a solution of 2-(3-fluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL), 2-(3-fluorophenyl)prop-2-amine (32 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) were added. The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-3-(2-((2-(3-fluorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-(1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methamide were obtained.

[0140] Isomer 1: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.61 - 9.48 (m, ¹H), 8.41 - 8.36 (s, ¹H), 8.16 - 8.02 (m, ¹H), 7.65 - 7.56 (m, ¹H), 7.39 - 7.30 (m, ¹H), 7.27 - 7.09 (m, 2H), 7.08 - 7.01 (m, ¹H), 7.01 - 6.93 (m, ¹H), 6.92 - 6.82 (m, ¹H), 4.50 - 4.25 (m, 4H), 3.22 (s, ¹H), 3.18 - 3.08 (m, 2H), 1.78 (d, J = 8.5 Hz, 2H), 1.58 - 1.42 (m, 8H), 1.38 - 1.15 (m, 4H), 1.11 - 1.01 (m, 3H). MS: (ES) m / z C 31H 36F 3N 4O 4[M+H] + Calculated value 585.3, experimental value 585.5.

[0141] Isomer 2: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.50 (s, ¹H), 8.40 - 8.35 (m, ¹H), 8.14 - 8.04 (m, ¹H), 7.67 - 7.55 (m, ¹H), 7.35 (dd, J = 8.8, 8.8 Hz, ¹H), 7.28 - 7.08 (m, 2H), 7.08 - 7.01 (m, 1H), 7.00 - 6.82 (m, 2H), 4.48 - 4.19 (m, 4H), 3.10 - 3.01 (m, 3H), 1.72 - 1.52 (m, 4H), 1.48 (d, J = 2.4 Hz). Hz, 8H), 1.28 - 1.13 (m, 2H), 1.08 (d, J = 2.5 Hz, 3H). MS: (ES) m / zC 31H 36F 3N 4O 4[M+H] + Calculated value 585.3, experimental value 585.5. [Example] [4] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Chlorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0142] Step a: 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) solution was added dropwise to a solution of 3-chlorobenzonitrile (1.00 g, 7.30 mmol) in ether (100 mL), followed by titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The mixture was refluxed overnight under N2, then cooled to 0°C and quenched with water, followed by 10% NaOH (aqueous solution). The contents were filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na2SO4, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(3-chlorophenyl)prop-2-amine. MS: (ES) m / z C9H13ClN [M+H]+ Calculated value 170.1, experimental value 170.2.

[0143] Step b: Add 2-(3-fluorophenyl)propane-2-amine (34 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidyl-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. By preparative HPLC purification of the mixture, two isolated isomers of N-(3-(2-((2-(3-chlorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methamide were obtained.

[0144] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.58 (s, ¹H), 8.47 (s, ¹H), 8.12 - 7.96 (m, ¹H), 7.38 - 7.05 (m, 7H), 4.58 - 4.35 (m, 4H), 3.30 - 3.09 (m, 3H), 2.03 - 1.91 (m, 2H), 1.74 - 1.64 (m, 2H), 1.63 - 1.55 (m, 6H), 1.55 - 1.45 (m, 2H), 1.44 - 1.25 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 31H 36ClF 2N 4O 4[M+H] + Calculated value 601.2, experimental value 601.3.

[0145] Isomer 2: ¹H NMR (400 MHz, CD₃OD) δ 8.49 (s, ¹H), 8.47 (s, ¹H), 8.10 - 8.02 (m, ¹H), 7.32 - 7.07 (m, 7H), 4.56 - 4.33 (m, 4H), 3.30 - 3.07 (m, 3H), 1.86 - 1.70 (m, 4H), 1.65 - 1.49 (m, 8H), 1.48 - 1.35 (m, 2H), 1.20 (s, 3H). MS: (ES) m / z C₃¹H₃₆ClF₂N₄O₄[M+H]₂ + Calculated value 601.2, experimental value 601.3. [Example] [5] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-] [3] [-(] [2] [-((] [2] [-(] [2] [,] [5] [-] [Difluorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0146] Step a: 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) solution was added dropwise to a solution of 2,5-difluorobenzonitrile (1.00 g, 7.19 mmol) in ether (100 mL), followed by the addition of titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The resulting mixture was refluxed overnight under N₂, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The contents were filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na₂SO₄, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(2,5-difluorophenyl)prop-2-amine. MS: (ES) m / z C₁₈H₁₂F₂N₂[M+H]⁺ Calculated value 172.1, experimental value 172.2.

[0147] Step b: Add 2-(2,5-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (50 mg, 0.10 mmol) to a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidyl-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL) to the solution. The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. By preparative HPLC purification of the mixture, two isolated isomers of 5-(2,4-difluorophenyl)-N-3-(2-((2-(2,5-difluorophenyl)propyl-2-yl)amino)-2-sideoxyethyl)-(1-(4-hydroxy-4-methylcyclohexyl)-acetidin-3-yl)isoazol-3-methamide were obtained.

[0148] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.65 (s, ¹H), 8.55 (s, ¹H), 8.14 - 8.02 (m, ¹H), 7.32 - 7.18 (m, 2H), 7.15 - 7.02 (m, 2H), 6.98 - 6.74 (m, 2H), 4.57 - 4.33 (m, 4H), 3.30 - 3.07 (m, 3H), 1.97 (s, 2H), 1.75 - 1.56 (m, 8H), 1.55 - 1.30 (m, 4H), 1.21 (s, 3H). MS: (ES) m / zC 31H 35F 4N 4O 4[M+H] + Calculated value 603.3, experimental value 603.5.

[0149] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.57 (s, ¹H), 8.55 (s, ¹H), 8.13 - 8.03 (m, ¹H), 7.33 - 7.18 (m, 2H), 7.14 - 7.02 (m, 2H), 6.96 - 6.87 (m, ¹H), 6.86 - 6.75 (m, ¹H), 4.55 - 4.30 (m, 4H), 3.15 - 3.08 (m, 3H), 1.85 - 1.71 (m, 4H), 1.66 - 1.51 (m, 8H), 1.47 - 1.35 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 31H 35F 4N 4O 4[M+H] + Calculated value 603.3, experimental value 603.5. [Example] [6] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [,] [5] [-] [Difluorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0150] Step a: To a solution of 3,5-difluorobenzonitrile (1.00 g, 7.19 mmol) in ether (100 mL), a solution of 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) was added dropwise, followed by titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The resulting mixture was refluxed overnight under N2, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The mixture was filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na2SO4, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(3,5-difluorophenyl)prop-2-amine. MS: (ES) m / z C9H12F2N[M+H]+ Calculated value 172.1, experimental value 172.2.

[0151] Step b: Add 2-(3,5-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (50 mg, 0.10 mmol) to a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidyl-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL) to the solution. The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. Two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(3,5-difluorophenyl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methylamine were obtained by purification of the crude material by reverse-phase HPLC.

[0152] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.58 (s, ¹H), 8.49 (s, ¹H), 8.11 - 8.01 (m, ¹H), 7.32 - 7.18 (m, 2H), 7.13 - 7.06 (m, ¹H), 6.95 - 6.84 (m, 2H), 6.76 - 6.65 (m, ¹H), 4.58 - 4.35 (m, 4H), 3.30 - 3.09 (m, 3H), 2.05 - 1.92 (m, 2H), 1.75 - 1.65 (m, 2H), 1.64 - 1.46 (m, 8H), 1.45 - 1.31 (m, 2H), 1.21 (s, 3H). MS: (ES) m / z C 31H 35F 4N 4O 4[M+H] + Calculated value 603.3, experimental value 603.5.

[0153] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.51 (s, 1H), 8.48 (s, 1H), 8.11 - 8.01 (m, 1H), 7.31 - 7.18 (m, 2H), 7.10 (dd, J = 10.1, 3.5 Hz, 1H), 6.92 - 6.83 (m, 2H), 6.74 - 6.66 (m, 1H), 4.55 - 4.34 (m, 4H), 3.30 - 3.11 (m, 3H), 1.86 - 1.70 (m, 4H), 1.66 - 1.50 (m, 8H), 1.48 - 1.34 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 31H 35F 4N 4O 4[M+H] + Calculated value 603.3, experimental value 603.5. [Example] [7] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Cyanophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0154] Step a: Zn(CN)₂ (0.747 g, 6.36 mmol) was added to a solution of (1.00 g, 3.18 mmol) tributyl (2-(3-bromophenyl)prop-2-yl)aminocarbamate in 40 mL of dimethyl ether. The contents were bubbled with N₂ for 10 min, followed by the addition of Pd(dppf)Cl₂ (0.233 g, 0.318 mmol), and the mixture was heated under N₂ for 4 h. The reaction mixture was cooled to room temperature and quenched with saturated NH₄Cl (aqueous solution), followed by extraction with DCM. The organic layer was collected, dried over MgSO₄, concentrated under vacuum, and purified by silica gel column chromatography to give tributyl (2-(3-cyanophenyl)prop-2-yl)aminocarbamate. MS: (ES) m / zC 15H 21N 2O 2[M+H] + Calculated value 261.2, experimental value 261.2.

[0155] Step b: Trifluoroacetic acid was added to a solution of (0.50 g, 1.9 mmol) tributyl (2-(3-cyanophenyl)prop-2-yl)aminocarbamate in 8 mL of DCM. The reaction mixture was stirred overnight at room temperature, followed by quenching with saturated NaHCO3 (aqueous solution). The contents were extracted with DCM and the organic layer was collected, dried over MgSO4, and concentrated under vacuum to give 3-(2-aminoprop-2-yl)benzonitrile. MS: (ES) m / z C 10H 13N 2[M+H]+ Calculated value 161.1, experimental value 161.2.

[0156] Step c: Add 3-(2-aminopropyl-2-yl)benzonitrile (32 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the mixture, two isolated isomers of N-(3-(2-((2-(3-cyanophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methamide were obtained.

[0157] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.61 (s, ¹H), 8.52 (s, ¹H), 8.12 - 8.02 (m, ¹H), 7.71 - 7.59 (m, 2H), 7.56 - 7.49 (m, ¹H), 7.46 - 7.35 (m, ¹H), 7.30 - 7.17 (m, 2H), 7.14 - 7.07 (m, ¹H), 4.61 - 4.32 (m, 4H), 3.30 - 3.09 (m, 2H), 2.05 - 1.91 (m, 2H), 1.77 - 1.45 (m, 8H), 1.45 - 1.27 (m, 1H), 1.20 (s, 3H). MS: (ES) m / z C 32H 36F 2N 5O 4[M+H] + Calculated value 592.3, experimental value 592.3.

[0158] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.55 (s, ¹H), 8.52 (s, ¹H), 8.11 - 8.03 (m, ¹H), 7.68 - 7.59 (m, 2H), 7.55 - 7.49 (m, 1H), 7.45 - 7.35 (m, 1H), 7.31 - 7.17 (m, 2H), 7.13 - 7.07 (m, 1H), 4.56 - 4.33 (m, 4H), 3.30 - 3.11 (m, 3H), 1.86 - 1.70 (m, 4H), 1.66 - 1.50 (m, 8H), 1.48 - 1.33 (m, 2H), 1.20 (s, 3H). MS: (ES) m / z C 32H 36F 2N 5O 4[M+H] + Calculated value 592.3, experimental value 592.3. [Example] [8] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Methoxyphenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0159] Step a: 3.0 M methylmagnesium bromide (29.8 mL, 89.5 mmol) solution was added dropwise to a solution of 3-methoxybenzonitrile (5.0 g, 29.8 mmol) in diethyl ether (120 mL), followed by the addition of titanium isopropoxide (IV) (8.82 mL, 29.8 mmol). The mixture was refluxed overnight under N₂, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The contents were filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na₂SO₄, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(3-methoxyphenyl)prop-2-amine. MS: (ES) m / z C₁₀H₁₆NO₃[M+H]⁺ Calculated value 166.1, experimental value 166.0.

[0160] Step b: Add 2-(3-methoxyphenyl)propane-2-amine (40 mg, 0.24 mmol), HATU (100 mg, 0.26 mmol), and DIPEA (0.57 mL, 0.82 mmol) to a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamidoamino)-1-(4-hydroxy-4-methylcyclohexyl)acetidyl-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-((2-(3-methoxyphenyl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methoxyamine were obtained.

[0161] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.50 (s, 1H), 8.41 (s, 1H), 8.10 - 8.02 (m, 1H), 7.30 - 7.18 (m, 2H), 7.14 - 7.06 (m, 1H), 6.92 - 6.72 (m, 1H), 6.69 (d, J = 8.0 Hz, 1H), 4.57 - 4.35 (m, 4H), 3.70 - 3.64 (m, 3H), 3.18 - 3.08 (m, 2H), 1.95 (bs, 2H), 1.73 - 1.30 (m, 13H), 1.20 (s, 3H). MS: (ES) m / zC 32H 39F 2N 4O 5[M+H] + Calculated value 597.3, experimental value 597.5.

[0162] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.41 (s, 1H), 8.12 - 8.03 (m, 1H), 7.30 - 7.18 (m, 2H), 7.14 - 7.06 (m, 2H), 6.87 (d, J = 8.0 Hz, 1H), 6.83 (s, 1H), 6.69 (d, J = 7.6 Hz, 1H), 4.52 - 4.34 (m, 3H), 3.65 (s, 3H), 3.12 - 3.09 (m, 3H), 1.90 - 1.30 (m, 15H), 1.20 (s, 3H). MS: (ES) m / zC 32H 39F 2N 4O 5[M+H] + Calculated value 597.3, experimental value 597.5. [Example] [9] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [5] [-] [Fluoro-based] [-] [2] [-] [Methoxyphenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0163] Step a: 3.0 M methylmagnesium bromide (5.55 mL, 16.7 mmol) solution was added dropwise to a solution of 5-fluoro-2-methoxybenzonitrile (1.00 g, 5.46 mmol) in ether (100 mL), followed by the addition of titanium isopropoxide (IV) (1.65 mL, 5.46 mmol). The resulting mixture was refluxed overnight under N₂, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The contents were filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na₂SO₄, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(5-fluoro-2-methoxyphenyl)prop-2-amine. MS: (ES) m / z C₁₀H₁₅FNO₃ [M+H]⁺ Calculated value 184.1, experimental value 184.0.

[0164] Step b: Add 2-(5-fluorophenyl)isoazol-3-methoxyphenyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (50 mg, 0.10 mmol) to a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methoxyphenyl)prop-2-amine (37 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) in DMF (2 mL). The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. By preparative HPLC purification of the mixture, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-((2-(5-fluoro-2-methoxyphenyl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0165] Isomer 1: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.66 - 9.47 (m, ¹H), 8.23 ​​- 8.14 (m, ¹H), 8.12 - 8.03 (m, ¹H), 7.65 - 7.56 (m, ¹H), 7.38 - 7.30 (m, ¹H), 7.24 - 7.16 (m, ¹H), 6.93 - 6.78 (m, ³H), 4.46 - 4.22 (m, ⁴H), 3.61 (s, ³H), 3.30 - 3.22 (m, ¹H), 3.09 - 2.98 (m, ²H), 1.85 - 1.71 (m, ²H), 1.61 - 1.46 (m, 8H), 1.37 - 1.14 (m, 4H), 1.09 - 1.00 (m, 3H). MS: (ES) m / zC 32H 38F 3N 4O 5[M+H] + Calculated value 615.3, experimental value 615.3.

[0166] Isomer 2: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.54 (s, ¹H), 8.19 - 8.14 (m, ¹H), 8.13 - 8.02 (m, ¹H), 7.65 - 7.56 (m, ¹H), 7.39 - 7.30 (m, ¹H), 7.26 - 7.15 (m, ¹H), 6.94 - 6.78 (m, ³H), 4.43 - 4.21 (m, ⁴H), 3.61 (s, ³H), 3.15 - 2.96 (m, ³H), 1.71 - 1.34 (m, ¹²H), 1.27 - 1.12 (m, ²H), 1.07 (s, ⁴H). 3H). MS: (ES) m / zC 32H 38F 3N 4O 5[M+H] + Calculated value 615.3, experimental value 615.3. [Example]

[10] [:] [, N , ] [-(] [1] [-] [Cyclohexyl] [-] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Fluorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] [] []

[0167] Step a: To a solution of methyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamino)acetidin-3-yl)acetate (1.4 g, 3.6 mmol) in 27 mL of DCM, cyclohexanone (0.62 mL, 6.0 mmol) and DIPEA (2.1 mL, 12.0 mmol) were added. After 30 minutes, NaBH(OAc)3 (1.68 g, 8.0 mmol) was added and the mixture was stirred at room temperature. Upon completion, the reaction mixture was quenched with H2O and the aqueous layer was extracted with EtOAc. The organic layers were combined, dried over sodium sulfate, filtered, and concentrated. The residue was purified by silicone column chromatography to give methyl 2-(1-cyclohexyl-3-(5-(2,4-difluorophenyl)isoazol-3-methamino)acetidin-3-yl)acetate. MS: (ES) m / zC 22H 26F 2N 3O 4[M+H] + Calculated value 434.2, experimental value 434.2.

[0168] Step b: Add 1 M LiOH solution (10.7 mL, 10.7 mmol) to a solution of methyl 2-(1-cyclohexyl-3-(5-(2,4-difluorophenyl)isoazol-3-methamino)acetidin-3-yl)acetate (1.55 g, 3.9 mmol) in 10.7 mL THF. Stir the reaction mixture at room temperature, then quench with 1 N HCl. Remove the volatiles under vacuum and filter the resulting solid, wash and collect to give 2-(1-cyclohexyl-3-(5-(2,4-difluorophenyl)isoazol-3-methamino)acetidin-3-yl)acetic acid. MS: (ES) m / z C 21H 24F 2N 3O 4[M+H] + Calculated value 420.2, experimental value 420.1.

[0169] Step c: To a solution of 2-(1-cyclohexyl-3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acetidin-3-yl)acetic acid (75 mg, 0.18 mmol) in 1.2 mL of DMF, add 2-(3-fluorophenyl)prop-2-amine (41 mg, 0.30 mmol), DIPEA (0.16 mL, 0.92 mmol), and HATU (0.20 g, 0.53 mmol). The reaction mixture was stirred at room temperature for 2 hours, followed by quenching with H₂O. The aqueous layer was extracted with EtOAc, and the organic layers were combined, dried over sodium sulfate, filtered, and concentrated. The crude residue was purified by silicone column chromatography, followed by preparative HPLC to obtain N-(1-cyclohexyl-3-(2-((2-(3-fluorophenyl)prop-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methylamine. 1H NMR (400 MHz, DMSO- d 6 ) δ: 9.53 (d, J= 26.2 Hz, 1H), 8.38 (d, J= 2.9 Hz, 1H), 8.08 (dddd, J= 8.8, 8.8, 6.4, 2.5 Hz, 1H), 7.60 (ddd, J= 10.2, 10.2, 2.0 Hz 1H), 7.34 (ddd, J= 8.4, 8.4, 2.4 Hz, 1H), 7.23 (dd, J= 20.4, 3.1 Hz, 1H), 7.16 - 7.06 (m, 1H), 7.03 (d, J= 8.0 Hz, 1H), 6.95 (dddd, J = 11.0, 2.2, 2.2, 2.2 Hz, 1H), 6.86 (dddd, J = 8.3, 8.3, 5.8, 2.4 Hz, 1H), 4.53 - 4.03 (m, 4H), 3.32 (s, 1H), 3.18 - 3.08 (m, 1H), 3.06 (s, 1H), 1.94 - 1.65 (m, 4H), 1.65 - 1.52 (m, 1H), 1.47 (s, 6H), 1.36 - 0.90 (m, 5H). MS: (ES) m / zC 30H 34F 3N 4O 3[M+H] + Calculated value 555.3, experimental value 555.3. [Example]

[11] [:] [, N , ] [-(] [1] [-(] [Cyclopropylmethyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [1] [-(] [Pyrimidine] [-] [2] [-] [base] [)] [Cyclopropyl] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0170] Step a: LiOH (240 mg, 6.0 mmol) was added to a solution of 3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-3-(2-ethoxy-2-t-oxyethyl)acrylidine-1-carboxylic acid tributyl ester (600 mg, 1.2 mmol) in a 4:1 THF / H₂O (5 mL) mixture. The reaction mixture was stirred overnight at room temperature. The solvent was removed under vacuum and the residue was treated with 1 N HCl. The mixture was then diluted with EtOAc and washed with H₂O. The organic layer was concentrated and the crude material was purified by silicone column chromatography to give 2-(1-(tributyloxycarbonyl)-3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acrylidine-3-yl)acetic acid. MS: (ES) m / zC 20H 22F 2N 3O 6[M+H] + Calculated value 438.1, experimental value 438.2.

[0171] Step b: Add 1-(pyrimidin-2-yl)cyclopropyl-1-amine dihydrochloride (110 mg, 0.52 mmol), DIPEA (0.36 mL, 2.08 mmol), and HATU (220 mg, 0.58 mmol) to a solution of 2-(1-(tributoxycarbonyl)-3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acrylic acid (230 mg, 0.52 mmol) in DCM (2 mL). Stir the reaction mixture at room temperature for 1 hour. The solvent was removed under vacuum and the residue was purified by silicone column chromatography to give tributyl 3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-3-(2-sideoxy-2-((1-(pyrimidin-2-yl)cyclopropyl)amino)ethyl)acetidine-1-carboxylic acid. MS: (ES) m / z C 27H 29F 2N 6O 5[M+H] + Calculated value 555.2, experimental value 555.2.

[0172] Step c: Stir a mixture of 3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-3-(2-sideoxy-2-((1-(pyrimidin-2-yl)cyclopropyl)amino)ethyl)acidine-1-carboxylic acid tributyl ester (288 mg, 0.52 mmol) in 1:1 DCM / TFA (2 mL) for 1 hour at room temperature. Remove the solvent under vacuum to give 5-(2,4-difluorophenyl)-N-(3-(2-sideoxy-2-((1-(pyrimidin-2-yl)cyclopropyl)amino)ethyl)acidine-3-yl)isoazol-3-methamido)amine.

[0173] Step d: Add NaBH(OAc)3 (42 mg, 0.2 mmol) to a mixture of 5-(2,4-difluorophenyl)-N-(3-(2-sideoxy-2-((1-(pyrimidin-2-yl)cyclopropyl)amino)ethyl)acetidin-3-yl)isoazol-3-methamide (57 mg, 0.1 mmol), triethylamine (0.05 mL, 0.4 mmol), and cyclopropaneformaldehyde (15 mg, 0.2 mmol) in 4:1 DCM / MeOH (2 mL). The contents were stirred at room temperature for 1 hour, then filtered and purified by preparative HPLC to obtain N-(1-(cyclopropylmethyl)-3-(2-sideoxy-2-((1-(pyrimidin-2-yl)cyclopropyl)amino)ethyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.60 (dd, J= 5.1, 5.1 Hz, 2H), 8.05 (dd, J= 6.4, 6.4 Hz, 1H), 7.31 - 7.17 (m, 3H), 7.12 (dd, J= 11.9, 3.5 Hz, 1H), 4.69 (d, J= 10.6 Hz, 2H), 4.56 (d, J= 12.4 Hz, 2H), 3.25 (d, J= 7.3 Hz, 1H), 3.20 - 3.09 (m, 3H), 1.66 (q, J= 4.4 Hz, 2H), 1.31 (dd, J= 3.9, 3.9 Hz, 2H), 1.04 (s, 1H), 0.70 (s, 2H), 0.41 (s, 2H). MS: (ES) m / z C 26H 27F 2N 6O 3[M+H] + Calculated value 509.2, experimental value 509.2. [Example]

[12] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-((] [3] [-] [Methyloxetine] [-] [3] [-] [base] [)] [methyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [Pyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine]

[0174] Step a: Add NaBH(OAc)3 (551 mg, 2.0 mmol) to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acetidin-3-yl)acetate (600 mg, 1.29 mmol), triethylamine (0.55 mL, 4.0 mmol), and 3-methyloxetane-3-carboxaldehyde (260 mg, 2.0 mmol) in a 4:1 DCM / MeOH (10 mL). After stirring at room temperature for 1 hour, quench the reaction mixture with saturated NaHCO3 (aqueous solution) and extract the aqueous layer with DCM. Combine the organic layers, dry over MgSO4, filter, and concentrate under vacuum. The crude residue was purified by silicone column chromatography to give ethyl acetate 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-((3-methyloxacyclobut-3-yl)methyl)acetidine-3-yl). MS: (ES) m / z C 22H 26F 2N 3O 5[M+H] + Calculated value 450.2, experimental value 450.2.

[0175] Step b: LiOH (224 mg, 5.34 mmol) was added to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-((3-methyloxetane-3-yl)methyl)acetidine-3-yl)acetic acid (400 mg, 0.89 mmol) in a 4:1 THF / H₂O (5 mL). The reaction mixture was stirred overnight at room temperature. The volatiles were removed under vacuum and the residue was treated with 1 N HCl. The solid was filtered, washed with H₂O, and collected to give 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-((3-methyloxetane-3-yl)methyl)acetidine-3-yl)acetic acid. MS: (ES) m / zC 20H 22F 2N 3O 5[M+H] + Calculated value 422.1, experimental value 422.1.

[0176] Step c: To a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-((3-methyloxacyclobut-3-yl)methyl)acetidin-3-yl)acetic acid (45 mg, 0.10 mmol) in DCM (2 mL), 2-(pyridin-2-yl)prop-2-amine dihydrochloride (23 mg, 0.10 mmol), DIPEA (0.07 mL, 0.40 mmol), and HATU (46 mg, 0.12 mmol) were added. The reaction mixture was stirred for 1 hour, followed by concentration under vacuum. The crude residue was purified by preparative HPLC to obtain 5-(2,4-difluorophenyl)-N-(1-((3-methyloxacyclobut-3-yl)methyl)-3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.59 (s, 1H), 8.27 (s, 1H), 8.06 (ddd, J= 8.5,8.5, 6.2 Hz, 1H), 7.89 (s, 1H), 7.70 (s, 1H), 7.32 - 7.17 (m, 2H), 7.12 (s, 1H), 4.76 - 4.47 (m, 6H), 4.36 (d, J= 6.3 Hz, 2H), 3.68 (d, J= 20.0 Hz, 2H), 3.22 (d, J= 20.0 Hz, 2H), 1.70 (s, 6H), 1.39 (s, 3H). MS: (ES) m / zC 28H 32F 2N 5O 4[M+H] + Calculated value 540.2, experimental value 540.3. [Example]

[13] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [2] [,] [6] [-] [Dimethylpyridine] [-] [4] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine]

[0177] To a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (60 mg, 0.13 mmol) in DMF (2 mL), 2-(2,6-dimethylpyridin-4-yl)propyl-2-amine (22 mg, 0.13 mmol), DIPEA (0.06 mL, 0.33 mmol), and HATU (61 mg, 0.16 mmol) were added. The contents were stirred for 1 hour and then concentrated under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(2,6-dimethylpyridin-4-yl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0178] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.05 ̶ 8.00 (m, 1H), 7.63 (s, 2H), 7.30 ̶ 7.11 (m, 3H), 4.61 - 4.38 (m, 4H), 3.24 ̶ 3.18 (m, 2H), 2.68 (s, 6H), 2.01 - 1.96 (m, 2H), 1.75 ̶ 1.48 (m, 13H), 1.22 (s, 3H). MS: (ES) m / z C 32H 40F 2N 5O 4[M+H] + Calculated value 596.3, experimental value 596.5.

[0179] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.08 ± 8.00 (m, 1H), 7.63 - 7.61 (m, 2H), 7.30 ± 7.12 (m, 3H), 4.58 - 4.37 (m, 4H), 3.30 ± 3.19 (m, 2H), 2.67 (s, 6H), 1.82 - 1.73 (m, 13H), 1.48 ± 1.38 (m, 2H), 1.21 (s, 3H). MS: (ES) m / z C 32H 40F 2N 5O 4[M+H] + Calculated value 596.3, experimental value 596.5. [Example]

[14] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-] [Isobutyl] [-] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Methylpyridine] [-] [4] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine]

[0180] Step a: Add NaBH3CN (52 mg, 0.8 mmol) to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acetidin-3-yl)acetate (300 mg, 0.8 mmol), DIPEA (0.29 mL, 1.6 mmol), acetic acid (0.19 mL, 3.2 mmol), and isobutyraldehyde (118 mg, 1.6 mmol) in 2:1 DCM / MeOH (9 mL). Stir the contents at room temperature for 1 hour, quench the reaction mixture with saturated NaHCO3 (aqueous solution), and extract with DCM. Dry the combined organic layers with MgSO4, filter, and concentrate under vacuum. The crude residue was purified by silicone column chromatography to give ethyl acetate 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-isobutylacrylidine-3-yl). MS: (ES) m / z C 21H 26F 2N 3O4 [M+H] + Calculated value 422.2, experimental value 422.2.

[0181] Step b: LiOH (49 mg, 1.2 mmol) was added to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-isobutylacrylidine-3-yl)acetate (164 mg, 0.4 mmol) in 1:1:1 THF / H₂O / MeOH (9 mL). The mixture was stirred at room temperature for 2 hours, concentrated under vacuum, and then treated with 1 N HCl (10 mL) and concentrated under vacuum to give 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-isobutylacrylidine-3-yl)acetic acid. MS: (ES) m / z C₁₉H₂₂F₂N₃O₄[M+H]⁺ Calculated value 394.2, experimental value 394.2.

[0182] Step c: Add 2-(2-methylpyridin-4-yl)propyl-2-amine (21 mg, 0.14 mmol), DIPEA (0.06 mL, 0.35 mmol), and HATU (64 mg, 0.17 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isobutyl-3-methylamino)-1-isobutylacryl-3-yl)acryl-3-yl)isobutyl-3-methylamino ... 1H NMR (400 MHz, DMSO- d 6 ) δ 9.56 (d, J= 4.6 Hz, 1H), 8.72 - 8.69 (m, 1H), 8.48 - 8.44 (m, 1H), 8.15 - 8.07 (m, 1H), 7.65 - 7.53 (m, 2H), 7.47 - 7.37 (m, 1H), 7.35 - 7.28 (m, 1H), 7.27 - 7.23 (m, 1H), 4.56 - 4.39 (m, 2H), 4.35 - 4.22 (m, 2H), 3.15 - 2.99 (m, 4H), 2.51 (s, 3H), 1.89 - 1.78 (m, 1H), 1.51 (s, 6H), 0.89 - 0.86 (m, 6H). MS: (ES) m / zC 28H 34F 2N 5O 3[M+H] + Calculated value 526.3, experimental value 526.3. [Example]

[15] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Chlorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-] [Isobutylacetidine] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0183] 2-(3-chlorophenyl)propane-2-amine (17 mg, 0.10 mmol), DIPEA (0.04 mL, 0.25 mmol), and HATU (46 mg, 0.12 mmol) were added to a solution of 2-(3-(5-(2,4-difluorophenyl)propane-2-amino)-1-isobutylacrylidine-3-yl)acetic acid (40 mg, 0.10 mmol) in DMF (2 mL). The mixture was stirred for 1 hour, filtered, and then purified by preparative HPLC to obtain N-(3-(2-((2-(3-chlorophenyl)propane-2-yl)amino)-2-sideoxyethyl)-1-isobutylacrylidine-3-yl)-5-(2,4-difluorophenyl)isobutyl-3-amino)acetic acid. MS: (ES) m / zC 28H 32ClF 2N 4O 3[M+H] + Calculated value 545.2, experimental value 545.2. [Example]

[16] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [Pyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)-] [1] [-(] [2] [-] [Oxygen snail] [[] [3] [.] [5] []] [the ninth of the ten Heavenly Stems] [-] [7] [-] [base] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine]

[0184] Step a: Add NaBH3CN (7 mg, 0.1 mmol) to a mixture of 5-(2,4-difluorophenyl)-N-(3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine (70 mg, 0.1 mmol), DIPEA (0.04 mL, 0.2 mmol), acetic acid (0.03 mL, 0.4 mmol), and 2-oxospiro[3.5]non-7-one (30 mg, 0.2 mmol) in 2:1 DCM / MeOH (3 mL). Stir the contents at room temperature for 1 hour, then quench the reaction mixture with saturated NaHCO3 (aqueous solution) and extract with DCM. Dry the combined organic layers with MgSO4, filter, and concentrate under vacuum. The crude material was purified by silicone column chromatography to obtain 5-(2,4-difluorophenyl)-N-(3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)-1-(2-oxospi[3.5]non-7-yl)acetidin-3-yl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.39 (dd, J= 4.9, 1.8 Hz, 1H), 8.07 - 8.01 (m, 1H), 7.69 - 7.64 (m, 1H), 7.41 (d, J= 8.1 Hz, 1H), 7.30 - 7.14 (m, 3H), 7.07 (d, J= 3.5 Hz, 1H), 4.42 (s, 2H), 4.34 (s, 2H), 3.77 - 3.66 (m, 4H), 3.05 (s, 2H), 2.39 - 2.30 (m, 1H), 2.15 (d, J= 13.5 Hz, 2H), 1.79 - 1.75 (m, 2H), 1.60 (s, 6H), 1.50 - 1.43 (m, 2H), 1.05 - 0.95 (m, 2H). MS: (ES) m / zC 31H 36F 2N 5O 4[M+H] + Calculated value 580.3, experimental value 580.2. [Example]

[17] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Methylpyridine] [-] [4] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0185] NaBH(OAc)3 (43 mg, 0.20 mmol) was added to a mixture of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(2-methylpyridin-4-yl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methamide (65 mg, 0.10 mmol), Et3N (0.05 mL, 0.4 mmol), and 4-hydroxy-4-methylcyclohexane-1-one (26 mg, 0.20 mmol) in a 4:1 DCM / MeOH mixture (2 mL). The contents were stirred at room temperature for 1 hour, followed by concentration under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-((2-(2-methylpyridin-4-yl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0186] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.50 (d, J = 6.3 Hz, 1H), 8.05 (ddd, J = 8.5, 8.5, 6.0 Hz, 1H), 7.84 (s, 1H), 7.77 (d, J = 5.6 Hz, 1H), 7.24 (dddd, J = 22.5, 8.7, 8.7, 2.6 Hz, 2H), 7.11 (d, J = 4.0 Hz, 1H), 4.60 - 4.35 (m, 4H), 3.30 - 3.10 (m, 3H), 2.75 (s, 3H), 1.98 (s, 2H), 1.74 - 1.61 (m, 8H), 1.56 - 1.36 (m, 4H), 1.21 (s, 3H). MS: (ES) m / zC 31H 38F 2N 5O 4[M+H] + Calculated value 582.3, experimental value 582.3.

[0187] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.50 (d, J = 6.2 Hz, 1H), 8.06 (ddd, J = 8.6, 8.6, 6.2 Hz, 1H), 7.83 (d, J = 8.0 Hz, 1H), 7.77 (s, 1H), 7.32 - 7.17 (m, 2H), 7.13 (s, 1H), 4.57 - 4.35 (m, 4H), 3.38 - 3.10 (m, 3H), 2.72 (d, J = 4.5 Hz, 3H), 1.85 - 1.70 (m, 4H), 1.68 - 1.50 (m, 8H), 1.48 - 1.36 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 31H 37F 2N 5O 4[M+H] + Calculated value 582.3, experimental value 582.3. [Example]

[18] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-((] [2] [-(] [4] [-] [Methylpyrimidine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0188] NaBH(OAc)₃ (60 mg, 0.28 mmol) was added to a mixture of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(4-methylpyrimidin-2-yl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methamide (70 mg, 0.14 mmol), Et₃N (0.05 mL, 0.4 mmol), and 4-hydroxy-4-methylcyclohexane-1-one (36 mg, 0.28 mmol) in a 4:1 DCM / MeOH mixture (2 mL). The contents were stirred at room temperature for 1 hour, followed by concentration under vacuum. The residue was purified by preparative HPLC to obtain 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-((2-(4-methylpyrimidin-2-yl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.49 (dd, J= 11.3, 5.2 Hz, 1H), 8.05 (s, 1H), 7.30 - 7.06 (m, 4H), 4.59 - 4.38 (m, 4H), 3.14 - 3.06 (m, 3H), 2.55 - 2.42 (m, 3H), 1.98 (s, 2H), 1.75 - 1.56 (m, 2H), 1.63 (s, 3H), 1.62 (s, 3H), 1.56 - 1.30 (m, 4H), 1.20 (d, J= 8.0 Hz, 3H). MS: (ES) m / zC 30H 37F 2N 6O 4[M+H] + Calculated value 583.3, experimental value 583.3. [Example]

[19] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Methoxypyridine] [-] [4] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0189] To a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (50 mg, 0.10 mmol) in DCM (1 mL), 2-(2-methoxypyridin-4-yl)propyl-2-amine (17 mg, 0.10 mmol), DIPEA (0.052 mL, 0.3 mmol), and HATU (40 mg, 0.11 mmol) were added. The contents were stirred for 1 hour, followed by concentration under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-((2-(2-methoxypyridin-4-yl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0190] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.08 (d, J = 8.4 Hz, 1H), 7.95 (dd, J = 17.0, 6.0 Hz, 1H), 7.25 (dt, J = 17.1, 9.1 Hz, 2H), 7.16 - 7.09 (m, 1H), 6.96 (dd, J = 19.8, 5.9 Hz, 1H), 6.80 (d, J = 18.6 Hz, 1H), 4.58 - 4.44 (m, 3H), 4.39 (d, J = 11.3 Hz, 1H), 3.86 (s, 3H), 3.31 - 3.20 (m, 1H), 3.16 (d, J = 10.7 Hz, 2H), 1.97 (s, 2H), 1.57 (s, 6H), 1.70 - 1.24 (m, 6H), 1.20 (d, J = 1.6 Hz, 3H). MS: (ES) m / zC 31H 38F 2N 5O 5[M+H] + Calculated value 598.3, experimental value 598.3.

[0191] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.12 - 7.97 (m, 2H), 7.31 - 7.07 (m, 4H), 7.00 (d, J = 19.9 Hz, 1H), 4.57 - 4.42 (m, 3H), 4.37 (d, J = 11.7 Hz, 1H), 3.97 (s, 3H), 3.30 - 3.10 (m, 3H), 1.80 - 1.68 (m, 4H), 1.65 - 1.50 (m, 2H), 1.59 (s, 6H), 1.50 - 1.36 (m, 2H), 1.20 (d, J = 3.6 Hz, 3H). MS: (ES) m / z calculated C 31H 38F 2N 5O 5[M+H] + calculated value 598.3, experimental value 598.3. [Example]

[20] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [4] [,] [6] [-] [Dimethylpyrimidine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0192] Step a: At room temperature, a solution of 3.0 M methylmagnesium bromide (10 mL, 30 mmol) was added to a solution of 4,6-dimethylpyrimidin-2-carboxynitrile (1.34 g, 10 mmol) in diethyl ether (35 mL), followed by the addition of titanium isopropoxide (IV) (3.04 mL, 10 mmol). The reaction mixture was stirred overnight at 40 °C, followed by quenching with H₂O, and then with 10% NaOH (aqueous solution). The contents were diluted with DCM and passed through a diatomaceous earth stopper. The filtrate was concentrated and the crude material was purified by silica gel column chromatography to give 2-(4,6-dimethylpyrimidin-2-yl)propyl-2-amine.

[0193] Step b: Add 2-(4,6-dimethylpyrimidin-2-yl)propyl-2-amine (21 mg, 0.12 mmol), DIPEA (0.04 mL, 0.24 mmol), and HATU (52 mg, 0.14 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (60 mg, 0.12 mmol) in DCM (2 mL). Stir the contents for 1 hour and then concentrate under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(4,6-dimethylpyrimidin-2-yl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methamide were obtained.

[0194] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.07 - 7.98 (m, 1H), 7.23 (ddd, J = 18.7, 9.3, 9.3 Hz, 2H), 7.13 - 7.06 (m, 1H), 7.04 (s, 1H), 5.49 (s, 1H), 4.58 - 4.32 (m, 4H), 3.50 - 3.20 (m, 1H), 3.11 (d, J = 9.8 Hz, 2H), 2.40 (s, 6H), 1.99 (s, 2H), 1.75 - 1.32 (m, 6H), 1.62 (s, 6H), 1.20 (s, 3H). MS: (ES) m / zC 31H 39F 2N 6O 4[M+H] + Calculated value 597.3, experimental value 597.3.

[0195] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.07 - 7.98 (m, 1H), 7.23 (ddd, J= 18.7, 9.3, 9.3 Hz, 2H), 7.14 - 7.06 (m, 1H), 7.04 (s, 1H), 4.58 - 4.38 (m, 4H), 3.30 - 3.10 (m, 3H), 2.40 (s, 6H), 1.82 - 1.70 (m, 4H), 1.62 (s, 6H), 1.62 - 1.50 (m, 2H), 1.48 - 1.36 (m, 2H), 1.21 (s, 3H). MS: (ES) m / zC 31H 39F 2N 6O 4[M+H] + Calculated value 597.3, experimental value 597.3. [Example] [twenty one] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [6] [-(] [Trifluoromethyl] [)] [Pyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0196] To a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (60 mg, 0.12 mmol) in DMF (1 mL), 2-(6-(trifluoromethyl)pyridin-2-yl)propyl-2-amine (25 mg, 0.12 mmol), DIPEA (0.052 mL, 0.3 mmol), and HATU (52 mg, 0.14 mmol) were added. The contents were stirred for 1 hour and then concentrated under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-(6-(trifluoromethyl)pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0197] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.06 (d, J = 8.1 Hz, 1H), 7.87 (ddd, J = 15.4, 7.9, 7.9 Hz, 1H), 7.68 - 7.54 (m, 2H), 7.31 - 7.17 (m, 2H), 7.14 - 7.06 (m, 1H), 4.56 - 4.30 (m, 4H), 3.40 - 3.20 (m, 1H), 3.15 (d, J = 13.0 Hz, 2H), 1.96 (s, 2H), 1.63 (s, 6H), 1.74 - 1.30 (m, 6H), 1.19 (s, 3H). MS: (ES) m / zC 31H 35F 5N 5O 4[M+H] + Calculated value 636.3, experimental value 636.3.

[0198] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.06 (d, J = 8.5 Hz, 1H), 7.87 (ddd, J = 8.4, 8.4, 8.4 Hz, 1H), 7.69 - 7.52 (m, 2H), 7.24 (ddd, J = 17.8, 9.3, 9.3 Hz, 2H), 7.10 (dd, J = 10.7, 3.5 Hz, 1H), 4.56 - 4.30 (m, 4H), 3.40 - 3.10 (m, 3H), 1.85 - 1.70 (m, 4H), 1.62 (s, 6H), 1.62 - 1.35 (s, 4H), 1.19 (s, 3H). MS: (ES) m / zC 31H 35F 5N 5O 4[M+H] + Calculated value 636.3, experimental value 636.3. [Example] [twenty two] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [3] [-(] [Trifluoromethoxy] [)] [Phenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0199] Add 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (45 mg, 0.093 mmol) to a mixture of 2-(3-(trifluoromethoxy)phenyl)prop-2-amine (65 mg, 0.30 mmol), HATU (57 mg, 0.15 mmol), and DIPEA (0.061 mL, 0.35 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (45 mg, 0.093 mmol) in DMF (1.5 mL). The contents were stirred for 1 hour, quenched with water, and purified by preparative HPLC to obtain two isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-(3-(trifluoromethoxy)phenyl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine.

[0200] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.58 (s, ¹H), 8.49 (s, ¹H), 8.02 ̶ 8.10 (m, ¹H), 7.15 ̶ 7.34 (m, 5H), 7.03 ̶ 7.14 (m, 2H), 4.54 (d, J = 11.6 Hz, ¹H), 4.66 (s, 2H), 4.37 (d, J = 11.6 Hz, 1H), 3.22 ̶ 3.40 (m, 1H), 3.15 (d, J = 12.4 Hz, 2H), 1.96 (bs, 2H), 1.30 ̶ 1.56 (m, 12H), 1.20 (s, 3H). MS: (ES) m / zC 32H 36F 5N 4O 5[M+H] + Calculated value 651.3, experimental value 651.5.

[0201] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.52 (s, 1H), 8.49 (s, 1H), 8.02 ̶ 8.11 (m, 1H), 7.15 ̶ 7.40 (m, 5H), 7.03 ̶ 7.14 (m, 2H), 4.42 ̶ 4.54 (m, 3H), 4.36 (d, J= 12.4 Hz, 1H), 3.10 ̶ 3.35 (m, 3H), 1.70 ̶ 1.90 (m, 4H), 1.50 ̶ 1.62 (m, 8H), 1.34 ̶ 1.48 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 32H 36F 5N 4O 5[M+H] + Calculated value 651.3, experimental value 651.5. [Example] [twenty three] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [,] [5] [-] [Dichlorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0202] Add 2-(3,5-dichlorophenyl)prop-2-amine (25 mg, 0.12 mmol), HATU (30 mg, 0.079 mmol), and Et 3N (0.040 mL, 0.28 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (30 mg, 0.062 mmol) in DMF (1.5 mL). The mixture was stirred for 0.5 hours, quenched with water, and purified by preparative HPLC to obtain two isomers of N-(3-(2-((2-(3,5-dichlorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.61 (s, 1H), 8.51 (s, 1H), 8.00 ̶ 8.10 (m, 1H), 7.15 ̶ 7.32 (m, 5H), 7.06 ̶ 7.13 (m, 1H), 4.54 (d, J= 12.4 Hz, 1H), 4.47 (s, 2H), 4.39 (d, J= 12.0 Hz, 1H), 3.22 ̶ 3.40 (m, 1H), 3.15 (d, J= 12.4 Hz, 2H), 1.97 (bs, 2H), 1.30 ̶ 1.75 (m, 12H), 1.20 (s, 3H). MS: (ES) m / zC 31H 35Cl 2F 2N 4O 4[M+H] + Calculated value 635.2, experimental value 635.5. [Example] [twenty four] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Ethylpyridine] [-] [4] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0203] Step a: Under N2, add a solution of 3.0 M methylmagnesium bromide (7.56 mL, 22.7 mmol) and titanium isopropoxide (IV) (8.82 mL, 29.8 mmol) to a flask containing 50 mL of diethyl ether (1.00 g, 7.56 mmol). Stir the mixture overnight at 40 °C, cool to 0 °C, quench with water, and dilute with a 10% NaOH (aqueous solution). Filter the contents through diatomaceous earth and wash with DCM. Dry the organic layer of the filtrate with NaSO4, filter, concentrate under vacuum, and purify by silica gel column chromatography to give 2-(2-ethylpyridin-4-yl)propyl-2-amine. MS: (ES) m / z C 10H 17N 2[M+H]+ Calculated value 165.1, experimental value 165.0.

[0204] Step b: Add 2-(2-ethylpyridin-4-yl)prop-2-amine (25 mg, 0.15 mmol), HATU (45 mg, 0.12 mmol), and Et 3N (0.052 mL, 0.37 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (45 mg, 0.093 mmol) in DMF (1.5 mL). The contents were stirred for 1 hour, quenched with water, and purified by preparative HPLC to obtain two isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(2-(2-ethylpyridin-4-yl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methylamine.

[0205] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.85 (s, ¹H), 8.83 (s, ¹H), 8.52 (d, J = 6.0 Hz, ¹H), 8.02 ̶ 8.10 (m, ¹H), 7.86 (s, ¹H), 7.80 (s, ¹H), 7.18 ̶ 7.31 (m, 2H), 7.12 (s, ¹H), 4.36 ̶ 4.63 (m, 4H), 3.26 ̶ 3.40 (m, ¹H), 3.16 ̶ 3.26 (m, 2H), 2.97 ̶ 3.08 (m, 2H), 1.93 ̶ 2.02 (m, 2H), 1.66 ± 1.74 (m, 2H), 1.64 (s, 6H), 1.40 ± 1.56 (m, 4H), 1.35 (t, J = 7.6 Hz, 3H), 1.21 (s, 3H). MS: (ES) m / zC 32H 40F 2N 5O 4[M+H] + Calculated value 596.3, experimental value 596.4.

[0206] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.85 (s, ¹H), 8.81 (s, ¹H), 8.52 (d, J = 6.0 Hz, ¹H), 8.02 ̶ 8.10 (m, ¹H), 7.74 ̶ 7.87 (m, 2H), 7.18 ̶ 7.31 (m, 2H), 7.12 (s, ¹H), 4.34 ̶ 4.60 (m, 4H), 3.16 ̶ 3.26 (m, ¹H), 3.18 (s, 2H), 2.98 ̶ 3.04 (m, 2H), 1.72 ̶ 1.84 (m, 4H), 1.64 (s, 6H). 1.37 ̶ 1.63 (m, 4H), 1.34 (t, J= 7.4 Hz, 3H), 1.20 (s, 3H). MS: (ES) m / zC 32H 40F 2N 5O 4[M+H] + Calculated value 596.3, experimental value 596.4. [Example]

[25] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-] [Isobutyl] [-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [Pyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0207] Add NaBH(OAc)3 (50 mg, 0.23 mmol) to a mixture of 5-(2,4-difluorophenyl)-N-(3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methamide (35 mg, 0.071 mmol), isobutyraldehyde (30 mg, 0.042 mmol), and Et3N (0.050 mL, 0.35 mmol) in DCM (2 mL). The contents were stirred for 1 hour, quenched with water, concentrated, and purified by preparative HPLC to obtain 5-(2,4-difluorophenyl)-N-(1-isobutyl-3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.64 (ddd, J= 7.6, 2.0, 1.2 Hz, 1H), 8.43 (dd, J= 10.4, 10.4 Hz, 1H), 7.97 ̶ 8.10 (m, 2H), 7.83 (ddd, J= 8.8, 8.8, 1.6 Hz, 1H), 7.16 ̶ 7.29 (m, 2H), 7.09 (d, J= 4.8 Hz, 1H), 4.25 ̶ 4.75 (m, 4H), 3.00 ̶ 3.40 (m, 4H), 1.85 ̶ 2.00 (m, 1H), 1.72 (s, 6H), 0.97 (d, J = 9.2 Hz, 6H). MS: (ES) m / z C 27H 32F 2N 5O 3[M+H] + Calculated value 512.2, experimental value 512.4. [Example]

[26] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [1, H , ] [-] [pyrrolo] [[] [2] [,] [3] [-] [, b , ] []] [Pyridine] [-] [4] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0208] Step a: Under N2 conditions, a solution of 3.0 M methylmagnesium bromide (7.0 mL, 21 mmol) was added to a flask containing 1 H-pyrrolo[2,3-b]pyridin-4-carboxylonitrile (1.00 g, 7.0 mmol) in ether (50 mL) and THF (50 mL), followed by the addition of titanium isopropoxide (IV) (2.07 mL, 7.0 mmol). The mixture was stirred overnight at 40 °C, cooled to 0 °C, quenched with water, and diluted with a 10% NaOH (aqueous solution). The contents were filtered through diatomaceous earth and washed with DCM. The organic layer of the filtrate was dried over NaSO4, filtered, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(1 H-pyrrolo[2,3-b]pyridin-4-yl)propyl-2-amine. MS: (ES) m / zC 10H 14N 3[M+H] + Calculated value 176.1, experimental value 176.1.

[0209] Step b: Add 2-(1H-pyrrolo[2,3-b]pyridin-4-yl)prop-2-amine (25 mg, 0.14 mmol), HATU (30 mg, 0.079 mmol), and Et 3N (0.040 mL, 0.28 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acryl-3-yl)acetic acid (35 mg, 0.072 mmol) in DMF (1.5 mL). The contents were stirred for 0.5 hours, quenched with water, and purified by preparative HPLC to obtain N-(3-(2-((2-(1H-pyrrolo[2,3-b]pyridin-4-yl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 9.03 (s, 1H), 8.94 (s, 1H), 8.27 (d, J= 5.2 Hz, 1H), 8.08 (d, J= 6.8 Hz, 1H), 7.41 (d, J= 6.0 Hz, 1H), 7.15 ̶ 7.32 (m, 4H), 7.07 (s, 1H), 6.81 (d, J= 14.8 Hz, 1H), 4.22 ̶ 4.49 (m, 4H), 3.12 ̶ 3.30 (m, 3H), 1.89 (s, 2H), 1.76 (s, 6H), 1.25 ̶ 1.71 (m, 6H), 1.17 (s, 3H). MS: (ES) m / z C 32H 37F 2N 6O 4[M+H] + Calculated value 607.3, experimental value 607.5. [Example]

[27] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-((] [2] [-(] [4] [-] [Methylpyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0210] To a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (0.075 g, 0.17 mmol) in DCM (1 mL), 2-(4-methylpyridin-2-yl)propyl-2-amine (0.082 g, 0.55 mmol), DIPEA (0.09 mL, 0.50 mmol), and HATU (0.130 g, 0.33 mmol) were added. The mixture was stirred overnight at room temperature and then concentrated under vacuum. By preparative HPLC purification of the residue, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-((2-(4-methylpyridin-2-yl)propyl-2-yl)amino)-2-sideoxyethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0211] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.40 (s, ¹H), 8.05 (m, ¹H), 7.75 (s, ¹H), 7.59 (d, J = 6.0 Hz, ¹H), 7.34 - 7.16 (m, 2H), 7.11 (s, 1H), 4.58 - 4.34 (m, 3H), 3.21 - 3.11 (m, 3H), 2.55 (s, 3H), 2.02 - 1.92 (m, 2H), 1.82 - 1.72 (m, 1H), 1.68 (s, 6H), 1.58 - 1.45 (m, 5H), 1.44 - 1.32 (m, 1H). 1.21 (s, 3H). MS: (ES) m / z C 31H 38F 2N 5O 4[M+H] + Calculated value 582.3, experimental value 582.5.

[0212] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.40 (s, 1H), 8.05 (m, 1H), 7.76 (s, 1H), 7.61 (d, J = 6.0 Hz, 1H), 7.30 - 7.18 (m, 2H), 7.13 - 7.08 (m, 1H), 4.58 - 4.31 (m, 4H), 3.22 - 3.07 (m, 3H), 2.55 (s, 3H), 1.85 - 1.70 (m, 3H), 1.68 (s, 6H), 1.63 - 1.49 (m, 2H), 1.47 - 1.36 (m, 3H), 1.20 (s, 3H). MS: (ES) m / zC 31H 38F 2N 5O 4[M+H] + Calculated value 582.3, experimental value 582.5. [Example]

[28] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [4] [,] [6] [-] [Dimethylpyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0213] To a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (0.075 g, 0.15 mmol) in DMF (1 mL), 2-(4,6-dimethylpyridin-2-yl)propyl-2-amine (0.033 g, 0.20 mmol), HATU (0.129 g, 0.34 mmol), and DIPEA (0.09 mL, 0.50 mmol) were added. The reaction mixture was stirred at room temperature for 5 hours, followed by concentration under vacuum. The crude material was purified by preparative HPLC to obtain 5-(2,4-difluorophenyl)-N-(3-(2-((2-(4,6-dimethylpyridin-2-yl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methylamine. 1H NMR (400 MHz, CD 3OD) δ 8.08 - 8.00 (m, 1H), 7.69 (s, 1H), 7.57 (s, 1H), 7.34 - 7.16 (m, 2H), 7.10 (s, 1H), 4.65 - 4.35 (m, 4H), 3.40 - 3.15 (m, 2H), 2.70 - 2.60 (m, 1H), 2.57 (s, 6H), 2.05 - 1.90 (m, 2H), 1.75 - 1.70 (m, 1H), 1.70 (s, 6H), 1.57 - 1.33 (m, 5H), 1.21 (s, 3H). MS: (ES) m / zC 32H 40F 2N 5O 4[M+H] + Calculated value 596.3, experimental value 596.5. [Example]

[29] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [Pyrimidine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] [] []

[0214] To a solution of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (0.06 g, 0.13 mmol) in DMF (1 mL), 2-(pyrimidin-2-yl)propyl-2-amine dihydrochloride (0.06 g, 0.29 mmol), DIPEA (0.07 mL, 0.40 mmol), and HATU (0.10 g, 0.26 mmol) were added. The reaction mixture was stirred overnight at room temperature, followed by concentration under vacuum. The aqueous layer was extracted with EtOAc, and the organic layers were combined, dried over sodium sulfate, filtered, and concentrated. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-(pyrimidin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0215] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.68 (dd, J = 9.2, 4.9 Hz, 2H), 8.15 - 7.96 (m, 1H), 7.32 - 7.27 (m, 2H), 7.27 - 7.16 (m, 1H), 7.11 (dd, J = 8.1, 3.5 Hz, 1H), 4.58 - 4.37 (m, 4H), 3.27 - 3.20 (m, 1H), 3.09 (d, J = 16.3 Hz, 2H), 1.97 (bs, 2H), 1.75 - 1.67 (m, 2H), 1.64 (s, 3H), 1.63 (s, 3H) 1.55 - 1.45 (m, 2H), 1.43 - 1.32 (m, 2H), 1.21 (s, 3H). MS: (ES) m / zC 29H 35F 2N 6O 4[M+H] + Calculated value 569.3, experimental value 569.4.

[0216] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.67 (dd, J = 7.5, 4.9 Hz, 2H), 8.05 (m, 1H), 7.33 - 7.26 (m, 2H), 7.26 - 7.16 (m, 1H), 7.11 (dd, J = 10.7, 3.4 Hz, 1H), 4.56 - 4.36 (m, 4H), 3.19 - 3.05 (m, 3H), 1.87 - 1.70 (m, 4H), 1.63 (s, 3H), 1.62 (s, 3H), 1.61 - 1.51 (m, 2H), 1.48 - 1.35 (m, 2H), 1.21 (s, 3H). MS: (ES) m / zC 29H 35F 2N 6O 4[M+H] + Calculated value 569.3, experimental value 569.4. [Example]

[30] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [2] [,] [2] [-] [Dimethylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [Pyridine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0217] Step a: Add NaBH(OAc)3 (840 mg, 4.0 mmol) to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)acetidin-3-yl)acetate (800 mg, 2.0 mmol), Et 3N (0.55 mL, 4.0 mmol), and 2,2-dimethylcyclohexanone (505 mg, 4.0 mmol) in a 4:1 DCM / MeOH (20 mL). Stir the reaction mixture at room temperature for 1 hour, then quench with saturated NaHCO3 (aqueous solution) and extract with DCM. Dry the combined organic layers in MgSO4, filter, and concentrate under vacuum. The crude residue was purified by silicone column chromatography to give ethyl acetate 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(2,2-dimethylcyclohexyl)acetidin-3-yl). MS: (ES) m / z C 25H 32F 2N 3O 4[M+H] + Calculated value 476.2, experimental value 476.5.

[0218] Step b: Add NaOH (300 mg, 7.5 mmol) to a mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(2,2-dimethylcyclohexyl)acetidin-3-yl)acetic acid (600 mg, 1.2 mmol) in 1:1 THF / H₂O (20 mL). Stir the mixture overnight at room temperature, then treat with 1 N HCl and MeCN. Concentrate the contents under vacuum and coarsely grind the residue with acetone, then filter through diatomaceous earth. Concentrate the filtrate to produce 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(2,2-dimethylcyclohexyl)acetidin-3-yl)acetic acid. MS: (ES) m / zC 23H 28F 2N 3O 4[M+H] + Calculated value 448.2, experimental value 448.4.

[0219] Step c: Add 2-(pyridin-2-yl)propane-2-amine dihydrochloride (43 mg, 0.21 mmol), DIPEA (0.14 mL, 0.82 mmol), and HATU (82 mg, 0.22 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(2,2-dimethylcyclohexyl)acridin-3-yl)acetic acid (100 mg, 0.21 mmol) in DCM (2 mL). Stir the contents for 1 hour and then concentrate under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(2,2-dimethylcyclohexyl)-3-(2-sideoxy-2-((2-(pyridin-2-yl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0220] Isomer 1: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.81 - 9.50 (m, 2H), 8.51 - 8.37 (m, 2H), 8.17 - 8.06 (m, 1H), 7.69 - 7.48 (m, 1H), 7.41 - 7.10 (m, 4H), 4.57 - 4.22 (m, 3H), 3.36 - 3.25 (m, 1H), 3.17 - 3.01 (m, 2H), 2.26 - 2.14 (m, 1H), 1.76 - 1.10 (m, 14H), 0.90 - 0.79 (m, 6H). MS: (ES) m / zC 31H 38F 2N 5O 3[M+H] + Calculated value 566.3, experimental value 566.3.

[0221] Isomer 2: ¹H NMR (400 MHz, DMSO-d⁶) δ 9.55 (s, ¹H), 9.40 (s, ¹H), 8.50 - 8.37 (m, 2H), 8.18 - 8.03 (m, ¹H), 7.63 - 7.44 (m, 2H), 7.42 - 7.10 (m, 3H), 4.44 - 4.25 (m, ¹H), 3.18 (s, 2H), 1.79 - 1.63 (m, 2H), 1.79 - 1.63 (m, 3H), 1.59 - 1.46 (m, 6H), 1.24 (s, 7H), 1.06 - 1.00 (m, 3H), 0.94 - 0.88 (m, 3H). MS: (ES) m / zC 31H 38F 2N 5O 4[M+H] + Calculated value 566.3, experimental value 566.3. [Example]

[31] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [1] [-(] [Pyrimidine] [-] [2] [-] [base] [)] [Cyclopropyl] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine]

[0222] Add 1-(pyrimidin-2-yl)cyclopropylamine dihydrochloride (42 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.105 mL, 0.60 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acridin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. The crude material was purified by preparative HPLC to obtain 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((1-(pyrimidin-2-yl)cyclopropyl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine. 1H NMR (400 MHz, DMSO- d 6 ) δ 9.57 (s, 1H), 8.89 (s, 1H), 8.57 - 8.50 (m, 2H), 8.13 - 8.00 (m, 1H), 7.65 - 7.54 (m, 1H), 7.39 - 7.28 (m, 1H), 7.25 - 7.16 (m, 2H), 4.48 - 4.35 (m, 4H), 3.01 - 2.94 (m, 2H), 1.79 - 1.38 (m, 9H), 1.33 - 1.12 (m, 4H), 1.08 (s, 3H). MS: (ES) m / zC 29H 33F 2N 6O 4[M+H] + Calculated value 567.3, experimental value 567.5. [Example]

[32] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Methylphenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine]

[0223] Add 2-(o-tolyl)propyl-2-amine (34 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. The crude material was purified by preparative HPLC to obtain 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-(o-tolyl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine. ¹H NMR (400 MHz, CD 3OD) δ 8.14 - 7.99 (m, 1H), 7.34 - 6.88 (m, 7H), 4.58 - 4.30 (m, 4H), 3.25 - 3.03 (m, 3H), 2.26 - 2.14 (m, 3H), 1.90 - 1.68 (m, 4H), 1.68 - 1.48 (m, 8H), 1.48 - 1.31 (m, 2H), 1.20 (s, 3H). MS: (ES) m / z C 32H 39F 2N 4O 4[M+H] + Calculated value 581.3, experimental value 581.3. [Example]

[33] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Chlorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0224] Add 2-(2-chlorophenyl)propane-2-amine (36 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the crude material, two isolated isomers of N-(3-(2-((2-(2-chlorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methamide were obtained.

[0225] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.14 - 8.02 (m, 1H), 7.56 - 7.48 (m, 1H), 7.33 - 7.18 (m, 3H), 7.18 - 7.04 (m, 3H), 4.56 - 4.29 (m, 4H), 3.27 - 3.08 (m, 3H), 2.00 - 1.88 (m, 2H), 1.77 - 1.64 (m, 7H), 1.59 - 1.26 (m, 5H), 1.22 - 1.16 (m, 3H). MS: (ES) m / zC 31H 36ClF 2N 4O 4[M+H] + Calculated value 601.2, experimental value 601.3.

[0226] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.13 - 8.02 (m, ¹H), 7.54 - 7.48 (m, ¹H), 7.32 - 7.18 (m, 3H), 7.17 - 7.04 (m, 3H), 4.58 - 4.25 (m, 4H), 3.24 - 3.04 (m, 3H), 1.83 - 1.66 (m, ¹⁰H), 1.63 - 1.33 (m, 4H), 1.23 - 1.16 (m, 3H). MS: (ES) m / z C 3¹H 3⁶ClF ₂N ₄O ₄[M+H]⁺ Calculated value 601.2, experimental value 601.3. [Example]

[34] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Fluorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0227] Add 2-(2-fluorophenyl)propane-2-amine (34 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(2-(2-fluorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0228] Isomer 1: ¹H NMR (400 MHz, CD₃OD) δ 8.13 - 8.03 (m, ¹H), 7.40 - 7.01 (m, 6H), 6.89 - 6.73 (m, ¹H), 4.59 - 4.29 (m, 4H), 3.27 - 3.07 (m, 3H), 2.02 - 1.89 (m, 2H), 1.74 - 1.55 (m, 8H), 1.55 - 1.27 (m, 4H), 1.20 (s, 3H). MS: (ES) m / z C₃¹H₃₆F₃N₄O₄[M+H]⁺ Calculated value 585.3, experimental value 585.3.

[0229] Isomer 2: ¹H NMR (400 MHz, CD₃OD) δ 8.13 - 8.04 (m, ¹H), 7.38 - 7.01 (m, 6H), 6.85 - 6.75 (m, ¹H), 4.54 - 4.30 (m, 4H), 3.15 - 3.05 (m, 3H), 1.88 - 1.32 (m, ¹⁴H), 1.20 (s, 3H). MS: (ES) m / z C₃¹H₃₆F₃N₄O₄[M+H]⁺ Calculated value 585.3, experimental value 585.3. [Example]

[35] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)-] [3] [-(] [2] [-] [Side group] [-] [2] [-((] [2] [-(] [m-Toluyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)] [Ethyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0230] Add 2-(m-Tolyl)propyl-2-amine (34 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(1-(4-hydroxy-4-methylcyclohexyl)-3-(2-sideoxy-2-((2-(m-tolyl)propyl-2-yl)amino)ethyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0231] Isomer 1: 1H NMR (400 MHz, CD 3OD) δ 8.11 - 8.01 (m, 1H), 7.32 - 7.18 (m, 2H), 7.16 - 7.01 (m, 4H), 6.98 - 6.90 (m, 1H), 4.61 - 4.27 (m, 4H), 3.39 - 3.18 (m, 1H), 3.16 - 3.07 (m, 2H), 2.25 - 2.16 (m, 3H), 2.02 - 1.88 (m, 2H), 1.74 - 1.27 (m, 12H), 1.24 - 1.16 (m, 3H). MS: (ES) m / zC 32H 39F 2N 4O 4[M+H] + Calculated value 581.3, experimental value 581.3.

[0232] Isomer 2: ¹H NMR (400 MHz, CD₃OD) δ 8.14 - 7.99 (m, 1H), 7.34 - 6.88 (m, 7H), 4.58 - 4.30 (m, 4H), 3.25 - 3.03 (m, 3H), 2.26 - 2.14 (m, 3H), 1.90 - 1.68 (m, 4H), 1.68 - 1.48 (m, 8H), 1.48 - 1.31 (m, 2H), 1.20 (s, 3H). MS: (ES) m / z C₃₂H₃₹F₂N₄O₄[M+H]⁺ Calculated value 581.3, experimental value 581.3. [Example]

[36] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Chlorine] [-] [5] [-] [Fluorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0233] Step a: 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) solution was added dropwise to a solution of 3-chloro-5-fluorobenzonitrile (1.00 g, 6.37 mmol) in ether (100 mL), followed by titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The resulting mixture was refluxed overnight under N2, then cooled to 0°C and quenched with water, followed by 10% NaOH (aqueous solution). The contents were filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na2SO4, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(3-chloro-5-fluorophenyl)prop-2-amine. MS: (ES) m / z C9H12ClFN [M+H]+ Calculated value 188.1, experimental value 188.1.

[0234] Step b: Add 2-(3-chloro-5-fluorophenyl)propane-2-amine (38 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the crude residue, two isolated isomers of N-(3-(2-((2-(3-chloro-5-fluorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)isoazol-3-methamide were obtained.

[0235] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.11 - 7.98 (m, 1H), 7.30 - 7.07 (m, 4H), 7.04 - 6.93 (m, 2H), 4.58 - 4.33 (m, 4H), 3.40 - 3.22 (m, 1H), 3.14 (s, 2H), 2.04 - 1.91 (m, 2H), 1.78 - 1.46 (m, 10H), 1.46 - 1.29 (m, 2H), 1.21 (s, 3H). MS: (ES) m / z C 31H 35ClF 3N 4O 4[M+H] + Calculated value 619.2, experimental value 619.2.

[0236] Isomer 2: 1H NMR (400 MHz, CD 3OD) δ 8.10 - 7.99 (m, 1H), 7.30 - 7.18 (m, 2H), 7.15 - 7.07 (m, 2H), 7.02 - 6.94 (m, 2H), 4.57 - 4.32 (m, 4H), 3.15 - 3.10 (m, 3H), 1.86 - 1.69 (m, 4H), 1.59 - 1.55 (m, 8H), 1.47 - 1.35 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 31H 35ClF 3N 4O 4[M+H] + Calculated value 619.2, experimental value 619.2. [Example]

[37] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Fluoro-based] [-] [5] [-] [Methoxyphenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0237] Step a: 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) solution was added dropwise to a solution of 3-fluoro-5-methoxybenzonitrile (1.00 g, 6.99 mmol) in ether (100 mL), followed by the addition of titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The resulting mixture was refluxed overnight under N₂, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The contents were filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na₂SO₄, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(3-fluoro-5-methoxyphenyl)prop-2-amine. MS: (ES) m / z C₁₀H₁₅FNO₃ [M+H]⁺ Calculated value 184.1, experimental value 184.1.

[0238] Step b: Add 2-(3-fluoro-5-methoxyphenyl)prop-2-amine (38 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methoxyphenyl)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(3-fluoro-5-methoxyphenyl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methylamine were obtained.

[0239] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.12 - 8.00 (m, 1H), 7.31 - 7.17 (m, 2H), 7.13 - 7.06 (m, 1H), 6.65 (s, 1H), 6.62 - 6.57 (m, 1H), 6.49 - 6.44 (m, 1H), 4.55 - 4.33 (m, 4H), 3.72 - 3.64 (m, 3H), 3.26 - 3.05 (m, 3H), 1.88 - 1.69 (m, 4H), 1.66 - 1.48 (m, 8H), 1.48 - 1.34 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 32H 38F 3N 4O 5[M+H] + Calculated value 615.3, experimental value 615.3.

[0240] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.12 - 8.00 (m, 1H), 7.33 - 7.17 (m, 2H), 7.14 - 7.06 (m, 1H), 6.71 - 6.56 (m, 2H), 6.52 - 6.43 (m, 1H), 4.58 - 4.33 (m, 4H), 3.72 - 3.64 (m, 3H), 3.39 - 3.20 (m, 1H), 3.17 - 3.08 (m, 2H), 2.04 - 1.89 (m, 2H), 1.74 - 1.28 (m, 12H), 1.20 (s, 3H). MS: (ES) m / zC 32H 38F 3N 4O 5[M+H] + Calculated value 615.3, experimental value 615.3. [Example]

[38] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Fluoro-based] [-] [5] [-] [Methoxyphenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0241] Step a: 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) solution was added dropwise to a solution of 2-fluoro-5-methoxybenzonitrile (1.00 g, 6.99 mmol) in diethyl ether (100 mL), followed by the addition of titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The resulting mixture was refluxed overnight under N₂, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The mixture was filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na₂SO₄, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(2-fluoro-5-methoxyphenyl)prop-2-amine. MS: (ES) m / z C₁₀H₁₅FNO₃ [M+H]⁺ Calculated value 184.1, experimental value 184.1.

[0242] Step b: Add 2-(2-fluoro-5-methoxyphenyl)propane-2-amine (38 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methoxyphenyl)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). The reaction mixture was stirred at room temperature for 1 hour, followed by quenching with 3 drops of H₂O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(2-(2-fluoro-5-methoxyphenyl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methamide were obtained.

[0243] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.12 - 8.02 (m, 1H), 7.32 - 7.18 (m, 2H), 7.14 - 7.07 (m, 1H), 6.88 - 6.81 (m, 1H), 6.80 - 6.67 (m, 2H), 4.57 - 4.32 (m, 4H), 3.71 (s, 3H), 3.28 - 3.02 (m, 3H), 2.03 - 1.90 (m, 2H), 1.75 - 1.27 (m, 12H), 1.20 (s, 3H). MS: (ES) m / zC 32H 38F 3N 4O 5[M+H] + Calculated value 615.3, experimental value 615.3.

[0244] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.12 - 8.03 (m, 1H), 7.32 - 7.18 (m, 2H), 7.14 - 7.07 (m, 1H), 6.87 - 6.81 (m, 1H), 6.74 - 6.67 (m, 2H), 4.58 - 4.29 (m, 4H), 3.70 (s, 3H), 3.15 - 3.05 (m, 3H), 1.85 - 1.70 (m, 4H), 1.67 - 1.33 (m, 10H), 1.20 (s, 3H). MS: (ES) m / zC 32H 38F 3N 4O 5[M+H] + Calculated value 615.3, experimental value 615.3. [Example]

[39] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [2] [-] [Fluoro-based] [-] [5] [-] [Methylphenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)] [Isotoxorubicin] [-] [3] [-] [Methionine] []

[0245] Step a: 3.0 M methylmagnesium bromide (7.30 mL, 21.9 mmol) solution was added dropwise to a solution of 2-fluoro-5-methylbenzonitrile (1.00 g, 7.87 mmol) in diethyl ether (100 mL), followed by the addition of titanium isopropoxide (IV) (2.16 mL, 7.30 mmol). The resulting mixture was refluxed overnight under N₂, then cooled to 0 °C and quenched with water, followed by 10% NaOH (aqueous solution). The mixture was filtered through a diatomaceous earth stopper and washed with DCM. The organic layer of the filtrate was collected, dried over Na₂SO₄, concentrated under vacuum, and purified by silica gel column chromatography to give 2-(2-fluoro-5-methylphenyl)prop-2-amine. MS: (ES) m / z C₁₀H₁₅FN [M+H]⁺ Calculated value 168.1, experimental value 168.1.

[0246] Step b: Add 2-(2-fluoro-5-methylphenyl)prop-2-amine (38 mg, 0.20 mmol), HATU (77 mg, 0.20 mmol), and DIPEA (0.070 mL, 0.40 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)isoazol-3-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acrylin-3-yl)acetic acid (50 mg, 0.10 mmol) in DMF (2 mL). Stir the contents at room temperature for 1 hour, then quench with 3 drops of H2O. By preparative HPLC purification of the crude material, two isolated isomers of 5-(2,4-difluorophenyl)-N-(3-(2-((2-(2-(2-fluoro-5-methylphenyl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)isoazol-3-methamide were obtained.

[0247] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.13 - 8.01 (m, 1H), 7.35 - 7.17 (m, 2H), 7.17 - 7.06 (m, 2H), 7.05 - 6.91 (m, 1H), 6.78 - 6.58 (m, 1H), 4.57 - 4.30 (m, 4H), 3.39 - 3.15 (m, 1H), 3.15 - 2.96 (m, 2H), 2.28 - 2.18 (m, 3H), 2.02 - 1.88 (m, 2H), 1.75 - 1.26 (m, 12H), 1.20 (s, 3H). MS: (ES) m / zC 32H 38F 3N 4O 4[M+H] + Calculated value 599.3, experimental value 599.3.

[0248] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.14 - 8.00 (m, 1H), 7.32 - 7.18 (m, 2H), 7.16 - 7.08 (m, 2H), 7.01 - 6.92 (m, 1H), 6.70 - 6.60 (m, 1H), 4.53 - 4.31 (m, 4H), 3.15 - 3.05 (m, 3H), 2.23 (s, 3H), 1.85 - 1.69 (m, 4H), 1.67 - 1.33 (m, 10H), 1.20 (s, 3H). MS: (ES) m / zC 32H 38F 3N 4O 4[M+H] + Calculated value 599.3, experimental value 599.3. [Example]

[40] [:] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)-] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [4] [,] [6] [-] [Dimethylpyrimidine] [-] [2] [-] [base] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [1] [,] [3] [,] [4] [-] [Thiadiazole] [-] [2] [-] [Methionine] []

[0249] Step a: HATU (3.75 g, 9.86 mmol) was added to a solution of 3-amino-3-(2-ethoxy-2-thiadiazole-1-carboxylic acid tributyl ester (2.31 g, 8.96 mmol) and 5-(2,4-difluorophenyl)-1,3,4-thiadiazole-2-carboxylic acid lithium (2.22 g, 8.96 mmol) in 20 mL of DMF. The mixture was stirred at room temperature for 1 hour, followed by quenching with H₂O. The precipitated solid was filtered, washed with water, and dried in a vacuum oven to give 3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazole-2-methamino)-3-(2-ethoxy-2-thiadiazole-2-carboxylic acid tributyl ester). MS: (ES) m / zC 21H 25F 2N 4O 5S [M+H] + Calculated value 483.1, experimental value 483.2.

[0250] Step b: The mixture of 3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazole-2-methamido)-3-(2-ethoxy-2-sideoxyethyl)acetidine-1-carboxylic acid tributyl ester (2.36 g, 4.89 mmol) in dimethyl ether (20 mL) containing 4 M HCl was stirred for 1 hour. The contents were concentrated under vacuum to give ethyl acetate 2-(3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazole-2-methamido)acetidine-3-yl). MS: (ES) m / z C 16H 17F 2N 4O 3S [M+H] + Calculated value 383.1, experimental value 383.1.

[0251] Step c: A mixture of ethyl 2-(3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazol-2-methamido)acetidin-3-yl)acetate (2.0 g, 5.23 mmol), pyridine (0.85 mL, 10.46 mmol), and 4-hydroxy-4-methylcyclohexane-1-one (1.0 g, 7.85 mmol) in a 4:1 DCM / MeOH mixture was stirred for 1 hour at room temperature. NaBH(OAc)3 (2.22 g, 10.46 mmol) was added to the contents. The reaction mixture was stirred for another 1 hour at room temperature, followed by quenching with saturated NaHCO3 (aqueous solution) and extraction with DCM. The combined organic layers were dried over MgSO4, filtered, and concentrated under vacuum. The crude residue was purified by silicone column chromatography to give two isolated isomers of ethyl acetate: 2-(3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazol-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl). MS: (ES) m / z C 23H 29F 2N 4O 4S [M+H] + Calculated value 495.2, experimental value 495.2.

[0252] Step d: LiOH (48 mg, 1.2 mmol) was added to a solution of ethyl 2-(3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazole-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (472 mg, 0.95 mmol) in a 4:1 THF / H₂O (2 mL) solution. The mixture was stirred overnight at room temperature, then concentrated to dryness, acidified to pH 3 to 4, and extracted with CHCl₃ / IPA (2:1) solution. The organic layers were combined, dried, filtered, and concentrated to give 2-(3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazole-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid. MS: (ES) m / zC 21H 25F 2N 4O 4S [M+H] + Calculated value 466.2, experimental value 466.2.

[0253] Step e: Add 2-(4,6-dimethylpyrimidin-2-yl)prop-2-amine (17 mg, 0.10 mmol), DIPEA (0.04 mL, 0.24 mmol), and HATU (42 mg, 0.11 mmol) to a mixture of 2-(3-(5-(2,4-difluorophenyl)-1,3,4-thiadiazol-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (47 mg, 0.10 mmol) in DMF (2 mL). The contents were stirred for 1 hour, and then purified by preparative HPLC to obtain the desired product 5-(2,4-difluorophenyl)-N-(3-(2-((2-(4,6-dimethylpyrimidin-2-yl)propyl-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-1,3,4-thiadiazole-2-methylamine.

[0254] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.44 (ddd, J = 8.0, 8.0, 8.0 Hz, 1H), 7.37 - 7.20 (m, 2H), 7.05 (s, 1H), 4.64 - 4.40 (m, 4H), 3.43 - 3.22 (m, 1H), 3.13 (d, J = 9.3 Hz, 2H), 2.40 (s, 6H), 1.99 (s, 2H), 1.75 - 1.35 (m, 6H), 1.62 (s, 6H), 1.21 (s, 3H). MS: (ES) m / zC 30H 38F 2N 7O 3S [M+H] + Calculated value 614.3, experimental value 614.0.

[0255] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.50 - 8.40 (m, 1H), 7.35 - 7.22 (m, 2H), 7.05 (s, 1H), 4.62 - 4.40 (m, 4H), 3.39 - 3.22 (m, 3H), 2.39 (s, 6H), 1.84 - 1.71 (m, 4H), 1.61 (s, 6H), 1.62 - 1.35 (m, 4H), 1.21 (s, 3H). MS: (ES) m / z C 30H 37F 2N 7O 3S [M+H] + Calculated value 614.3, experimental value 614.0. [Example]

[41] [:] [, N , ] [-(] [3] [-(] [2] [-((] [2] [-(] [3] [-] [Chlorophenyl] [)] [C] [-] [2] [-] [base] [)] [Amine] [)-] [2] [-] [Side-oxyethyl] [)-] [1] [-(] [4] [-] [Hydroxy] [-] [4] [-] [Methylcyclohexyl] [)] [Acidin] [-] [3] [-] [base] [)-] [5] [-(] [2] [,] [4] [-] [Difluorophenyl] [)] [Thiazole] [-] [2] [-] [Methionine] []

[0256] Step a: Pd(OAc)₂ (48 mg, 0.21 mmol), Xantphos (98 mg, 0.21 mmol), and NMM (0.52 mL, 4.7 mmol) were added to a solution containing (2,4-difluorophenyl)boronic acid (370 mg, 2.3 mmol) and ethyl 5-bromothiazol-2-carboxylate (500 mg, 2.1 mmol) in a 2:1 mixture of toluene / H₂O (4.2 mL / 2.1 mL). After two hours, the mixture was extracted with EtOAc. The organic layers were combined, dried over sodium sulfate, filtered, and concentrated. The crude material was purified by silicone column chromatography to give ethyl 5-(2,4-difluorophenyl)thiazol-2-carboxylate. MS: (ES) m / zC 12H 10F 2NO 2S [M+H] + Calculated value 270.0, experimental value 270.1.

[0257] Step b: Add a solution of 1 M LiOH (5.4 mL) to a solution of ethyl 5-(2,4-difluorophenyl)thiazol-2-carboxylate (488 mg, 1.8 mmol) in 5.4 mL THF. Stir the reaction mixture at room temperature for 16 hours, then quench with 1 N HCl. Filter the contents and collect the solid, drying it to give 5-(2,4-difluorophenyl)thiazol-2-carboxylic acid. MS: (ES) m / z C 10H 6F 2NO 2S [M+H] + Calculated values ​​242.0, 241.9.

[0258] Step c: DIPEA (0.59 mL, 3.4 mmol) was added to a solution containing 5-(2,4-difluorophenyl)thiazol-2-carboxylic acid (450 mg, 1.9 mmol) and 3-amino-3-(2-ethoxy-2-sideoxyethyl)acetidine-1-carboxylic acid tributyl ester (440 mg, 1.7 mmol) in 6.8 mL of DMF, followed by the addition of HATU (780 mg, 2.1 mmol). The contents were stirred at room temperature for 16 h, then concentrated under vacuum and purified by silicone column chromatography to obtain 3-(5-(2,4-difluorophenyl)thiazol-2-methamino)-3-(2-ethoxy-2-sideoxyethyl)acetidine-1-carboxylic acid tributyl ester. MS: (ES) m / zC 22H 26F 2N 3O 5S [M+H] + Calculated value 482.2, experimental value 482.2.

[0259] Step d: A solution of 4.0 M HCl in 3.6 mL of dimethyl methacrylate (3.6 mL, 14.2 mmol) was added to a solution of 3-(5-(2,4-difluorophenyl)thiazol-2-methamido)-3-(2-ethoxy-2-toxyethyl)acetidine-1-carboxylic acid tributyl ester (820 mg, 1.7 mmol) in 3.6 mL of dimethyl methacrylate. The contents were stirred at room temperature for 16 hours, followed by concentration to give ethyl acetate of 2-(3-(5-(2,4-difluorophenyl)thiazol-2-methamido)acetidine-3-yl). MS: (ES) m / z C 17H 18F 2N 3O 3S [M+H] + Calculated value 382.1, experimental value 382.2.

[0260] Step e: Pyridine (0.15 mL, 1.4 mmol) was added to a solution of ethyl 2-(3-(5-(2,4-difluorophenyl)thiazolyl-2-methamido)acetidin-3-yl)acetate (360 mg, 0.94 mmol) and 4-hydroxy-4-methylcyclohexane-1-one (165 mg, 1.3 mmol) in 3.4 mL DCM. After stirring at room temperature for 10 min, NaBH(OAc)3 (360 mg, 1.7 mmol) was added and the contents were stirred for another 16 h. The reaction mixture was quenched with H2O and extracted with EtOAc. The combined organic layers were dried over sodium sulfate, filtered, and concentrated. The crude material was purified by silicone column chromatography to give ethyl acetate 2-(3-(5-(2,4-difluorophenyl)thiazolyl-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl). MS: (ES) m / z C 24H 30F 2N 3O 4S [M+H]+ Calculated value 494.2, experimental value 494.4.

[0261] Step f: Add a solution of 1 M LiOH (2 mL, 2.0 mmol) to a solution of ethyl 2-(3-(5-(2,4-difluorophenyl)thiazol-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (339 mg, 0.69 mmol) in 2 mL THF. Stir the contents at room temperature for 16 hours, then quench with 1 N HCl and concentrate to dryness to give 2-(3-(5-(2,4-difluorophenyl)thiazol-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid. MS: (ES) m / z C 22H 26F 2N 3O 4S [M+H] + Calculated value 466.2, experimental value 466.3.

[0262] Step g: Add DIPEA (0.19 mL, 1.1 mmol) to a solution of 2-(3-(5-(2,4-difluorophenyl)thiazolyl-2-methamido)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)acetic acid (160 mg, 0.35 mmol) and 2-(3-chlorophenyl)prop-2-amine (70 mg, 0.41 mmol) in 2 mL of DMF, followed by the addition of HATU (245 mg, 0.65 mmol). Stir the contents at room temperature for 4 hours, then concentrate under vacuum. By preparative HPLC purification of the crude material, two isolated isomers of N-(3-(2-((2-(3-chlorophenyl)prop-2-yl)amino)-2-sideoxyethyl)-1-(4-hydroxy-4-methylcyclohexyl)acetidin-3-yl)-5-(2,4-difluorophenyl)thiazolyl-2-methamide were obtained.

[0263] Isomer 1: ¹H NMR (400 MHz, CD 3OD) δ 8.31 (m, 1H), 7.88 (ddd, J = 8.1, 8.1, 8.1 Hz, 1H), 7.38 - 7.05 (m, 6H), 4.68 - 4.29 (m, 4H), 3.13 (m, 2H), 2.05 - 1.90 (m, 2H), 1.78 - 1.62 (m, 2H), 1.59 (s, 3H), 1.57 (s, 3H), 1.55 - 1.44 (m, 3H), 1.44 - 1.28 (m, 2H), 1.20 (s, 3H). MS: (ES) m / zC 31H 36ClF 2N 4O 3S [M+H] + Calculated value 617.2, experimental value 617.4.

[0264] Isomer 2: ¹H NMR (400 MHz, CD 3OD) δ 8.49 (d, J = 10.6 Hz, ¹H), 8.32 (d, J = 3.4 Hz, ¹H), 7.92 - 7.84 (m, ¹H), 7.30 - 7.27 (m, ¹H), 7.25 - 7.17 (m, 2H), 7.17 - 7.10 (m, 4H), 4.59 - 4.43 (m, 4H), 4.37 (d, J = 11.8 Hz, ¹H), 3.17 - 2.94 (m, 3H), 1.85 - 1.70 (m, 3H), 1.64 - 1.58 (m, ¹H), 1.57 (s, 3H). 1.56 (s, 3H), 1.55 - 1.49 (m, 1H), 1.47 - 1.35 (m, 2H), 1.20 (s, 3H). MS: (ES) m / z C 31H 36ClF 2N 4O 3S [M+H] + Calculated value 617.2, experimental value 617.4. [Biological Examples] [1]

[0265] CXCR7 is fused co-framed with the small enzyme donor fragment ProLink and co-expressed in CHO cells that stably express the fusion protein of a larger N-terminal deletion mutant of β-repressor protein and β-galactosidase (called the enzyme receptor or EA). Activation of CXCR7 stimulates the binding of β-repressor protein to the ProLink-tagged CXCR7 and promotes complementarity between the two enzyme fragments, leading to the formation of active β-galactosidase. The enzyme activity of β-galactosidase is measured using a fluorescent acceptor.

[0266] CHO-CXCR7 cells (0.22 × 10⁶ / mL) were cultured in Ham's F-12 medium with 10% fetal bovine serum (FBS), and the transgenic gene was maintained using hygromycin B (200 μg / mL) and G418 (250 μg / mL). The day before analysis, cells were isolated from the culture dish using 0.25% trypsin-EDTA (Corning, catalog number 25-053-CI) and plated into 96-well plates (2.2 × 10⁵ cells / mL, 100 μL / well), and incubated overnight at 37°C with 5% CO₂. On the day of analysis, the cell culture medium was removed, and 100 μL of analysis buffer (PBS or FBS) was added to each well. 1 µL of the compound serially diluted in DMSO was added to each well. Subsequently, 5 µL of human SDF-1a (Pepro Tech, catalog number 300-28A, at the EC50 concentration predetermined on the same day) was added and mixed to induce CXCR7-mediated recruitment of β-repressor protein. The culture dish was incubated at 37°C for 1.5 hours. The analysis buffer was removed, 100 µL of acceptor solution was added, and the reaction was carried out at 37°C for 30 minutes. The acceptor solution was prepared by mixing 100 mL of phosphate buffer (1 M, Sigma, catalog number P3619-1GA), 100 mL of 10% Triton X 100 (Sigma, catalog number T8787.), 5 mL of MgCl2 (1 M, Sigma, catalog number M1028.), and 1.5 mL of β-mercaptoethanol (Gibco, catalog number 21985-023.). Add 0.4 mM of 3-carboxyum-umbelliferone β-D-piperanopyranoside (ThermoFisher Scientific, catalog number F2905) just before use. The reaction was then stopped by adding 50 µL of stop solution (2.1% w / v Na₂CO₃) to each well. Fluorescence intensity was measured using a FlexStation 3 Molecular Devices microplate reader with the following settings: excitation: 360 nm, emission: 465 nm, manual gain: 55. IC₅₀ values ​​were calculated using a GraphPad Prism with 3-parameter nonlinear regression. [verify]

[0267] The compounds initially identified as being of interest using any of the aforementioned screening methods can be further tested to validate their apparent activity in vivo. Preferably, such studies are conducted using suitable animal models. The basic form of such methods involves administering a lead compound identified during the initial screening to an animal serving as a disease model in humans, and subsequently determining whether the disease (e.g., cancer, myocardial infarction, wound healing, inflammatory diseases, or other diseases associated with CXCR7) is actually modulated and / or whether the disease or condition is improved. Animal models used in validation studies are generally any type of mammal. Specific examples of suitable animals include, but are not limited to, primates, mice, rats, and zebrafish.

[0268] The compounds in the table below were prepared as described above. Without providing specific synthetic details, minor variations were made to the above method by substituting the chosen reagents to prepare the compounds. The activities are provided as follows: IC50 < 5 nM (++++); 5 nM up to 100 nM (+++); 100 nM up to 1000 nM (++); and 1000 nM up to 20,000 nM (+). [, , ] [] [Specific Instance] [(] [surface] [1] [)] [] []

[0269] Those skilled in the art will recognize from the provided descriptions, drawings, and examples that various embodiments of the invention can be modified and altered without departing from the scope of the invention as defined by the following claims and their equivalents.

[0270] All patents, patent applications, publications, and reprints mentioned herein are incorporated herein by reference in their entirety. Any conflict between any references cited herein and the teachings of this specification shall be interpreted in accordance with this specification. Similarly, any conflict between the definitions of words or phrases generally accepted in this art and the definitions of words or phrases provided in this specification shall be interpreted in accordance with this specification.

Claims

1. A compound or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer thereof, having the formula (I): , wherein HAr is a five-membered heteroaryl ring; Ar1 ​​is selected from the group consisting of: phenyl, pyridyl, pyrimidinyl and pyridine; Ar2 is an aryl or heteroaryl, each of which is independently a monocyclic or fused bicyclic ring; subscript m is 0, 1 or 2; subscript n is 0, 1, 2 or 3; subscript p is 0, 1, 2 or 3; subscript q is 0, 1, 2, 3 or 4; each R1 is independently selected from the group consisting of: halogen, CN, C1-4 alkyl, C1-4 haloalkyl, -NRaRb, -ORa, -CO2Ra and -C(O)NRaRb; Each of the R2 groups is independently selected from members of the following groups: halogen, CN, C1-4 alkyl, C1-4 haloalkyl, -NRaRb, -ORa, -CO2Ra and -C(O)NRaRb; Each of the R3 groups is selected from members of the following groups: C1-4 alkyl, C1-4 haloalkyl, C1-4 hydroxyalkyl, -CO2Ra, -X-CO2Ra, -C(O)NRaRb and -XC(O)NRaRb; Each of the R4a and R5a groups is independently selected from members of the following groups: H, C1-6 alkyl, C1-6 haloalkyl, C1-6 hydroxyalkyl, -X-ORa, -CO2Ra, -X-CO2Ra, -X-NRaRb, -C(O)NRaRb and -XC(O)NRaRb; Each of R4 and R5 is independently selected from a member of the group consisting of: H, C1-6 alkyl, C1-6 haloalkyl, C1-6 hydroxyalkyl, -X-ORa, -CO2Ra, -X-CO2Ra, -X-NRaRb, -C(O)NRaRb and -XC(O)NRaRb; or R4 and R5 are combined to form a three- to five-membered ring having 0 or 1 heteroatom ring vertices selected from O, S or N, wherein the three- to five-membered ring is unsubstituted or substituted by 1 to 4 substituents independently selected from the group consisting of: halogen, CN, C1-4 alkyl, C1-4 haloalkyl, C1-4 alkoxy and C1-4 haloalkoxy;Each R6 group is independently selected from members of the following groups: halogen, CN, -X-CN, C1-4 alkyl, C1-4 haloalkyl, C3-6 cycloalkyl, C1-4 hydroxyalkyl, -ORa, -CO2Ra, -X-CO2Ra, -NRaRb, -X-NRaRb, -C(O)NRaRb, and -XC(O)NRaRb. R7 is a member selected from the group consisting of: C1-8 alkyl, C3-8 hydroxyalkyl, C1-4 alkoxy-C2-4 alkyl, -C(O)NH-C1-8 alkyl, -C(O)-C1-8 alkyl, -S(O)2-C1-8 alkyl, C3-8 cycloalkyl, -X-C3-8 cycloalkyl, C6-9 spirocycloalkyl, -X-C6-9 spirocycloalkyl, 4- to 7-membered heterocycloalkyl, -X-4- to 7-membered heterocycloalkyl, 7- to 11-membered spiroheterocycloalkyl and -X-7- to 11-membered spiroheterocycloalkyl, wherein each R7 is substituted with zero to four independent substituents selected from the group consisting of: hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy, ethoxy and cyclopropyl; Each Ra and Rb group is independently selected from the group consisting of: H, C1-4 alkyl, C1-4 haloalkyl, C3-6 cycloalkyl, and C3-6 cycloalkyl-C1-4 alkyl; each X group consists of a C1-4 alkyl-linked group, wherein any of the methylene moiety of X is unsubstituted or substituted with one or two methyl groups.

2. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the HAr group is selected from the group consisting of: isothiazolium, isothiazole, imidazole, pyrazole, thiazole, 1,2,4-isodiazole, 1,3,4-isodiazole, 1,2,4-thiadiazole, 1,3,4-thiadiazole, 1,2,3-triazole and 1,2,4-triazole.

3. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein HAr is selected from the group consisting of isothiazoles and thiadiazoles.

4. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein Ar1 is phenyl.

5. The compound of claim 4 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the subscript q is 1, 2 or 3; and each R1 is independently selected from a member of the group consisting of: halogen, CN, C1-4 alkyl and C1-4 haloalkyl.

6. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the Ar2 group is selected from the group consisting of: pyridinyl, pyrimidinyl, pyridine, phenyl, indolyl, thiazolyl, pyrazolyl, indazole, and pyrrolopyridinyl.

7. The compound of claim 6 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein Ar2 is selected from the group consisting of pyrimidinyl, pyridinyl and phenyl groups.

8. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enriched or mirror-image isomer enriched form or rotational isomer, wherein the Ar2 group is selected from the group consisting of: 2-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl, 2-thiazolyl, 4-pyrazolyl, phenyl and indolyl; and each R6 group is independently selected from the group consisting of: halogen, CN, C1-4 alkyl, C1-4 haloalkyl, C3-6 cycloalkyl and C1-4 alkoxy.

9. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein -Ar2-(R6)p is selected from the group consisting of:

10. The compound of claim 1 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein R7 is selected from the group consisting of: .

11. The compound of any one of claims 1 to 8 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the subscript m is 0.

12. The compound of any one of claims 1 to 8 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the subscript n is 0.

13. The compound of any one of claims 1 to 8 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the subscript p is 0, 1 or 2.

14. The compound of any one of claims 1 to 8 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the subscript q is 1 or 2.

15. The compound of claim 1 or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope enriched or mirror isomer enriched or rotational isomer thereof, wherein the compound has formula (Ia): .

16. The compound of claim 15 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein HAr is selected from the group consisting of isothiazoles and thiadiazoles.

17. The compound of claim 15 or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope enriched or mirror isomer enriched or rotational isomer thereof, wherein the compound has the formula (Ia1): .

18. The compound of claim 17 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein the compound has the formula (Ia2): .

19. The compound of claim 18 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein HAr is selected from the group consisting of isothiazoles and thiadiazoles.

20. The compound of claim 1 or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope enriched or mirror isomer enriched form or rotational isomer thereof, wherein the compound has formula (Ib), formula (Ic) or formula (Id): .

21. The compound of claim 20 or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope enriched or mirror isomer enriched form or rotational isomer thereof, wherein the compound has the formula (Ib1), (Ic1) or (Id1): .

22. The compound of claim 21 or its pharmaceutically acceptable salt, hydrate, N-oxide, isotope enrichment or mirror isomer enrichment or rotational isomer, wherein HAr is selected from the group consisting of isothiazoles and thiadiazoles.

23. A compound of any one of claims 1 to 8 or 15 to 22, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotopically enriched or mirror-isomer enriched form or rotational isomer thereof, wherein R7 is a member selected from the group consisting of: C1-8 alkyl, C3-8 hydroxyalkyl, C1-4 alkoxy-C2-4 alkyl, C3-8 cycloalkyl, C6-9 spirocycloalkyl, 4- to 7-membered heterocycloalkyl and 7- to 11-membered spiroheterocycloalkyl, wherein each R7 is substituted with zero to four independent substituents selected from the group consisting of: hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy, ethoxy and cyclopropyl.

24. A compound of any one of claims 1 to 8 or 15 to 22, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotopically enriched or mirror-isomer enriched form or rotational isomer thereof, wherein R7 is a member selected from the group consisting of: -X-C3-8 cycloalkyl, -X-C6-9 spirocycloalkyl, -X-4 to 7-membered heterocycloalkyl and -X-7 to 11-membered spiroheterocycloalkyl, wherein each R7 is substituted by zero to four independent substituents selected from the group consisting of: hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy, ethoxy and cyclopropyl.

25. A compound of any one of claims 1 to 8 or 15 to 22, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer thereof, wherein R7 is selected from the group consisting of cyclohexyl, cyclopentyl, piperidinyl, tetrahydropiperanyl and tetrahydrofuranyl, each of which is substituted by zero or two independent substituents selected from the group consisting of hydroxyl, methyl, ethyl, hydroxymethyl, fluoro, chloro, methoxy and ethoxy.

26. The compound of claim 1 or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer thereof, wherein the compound is selected from the following: .

27. A pharmaceutical composition comprising a compound as claimed in any one of claims 1 to 26, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotopically enriched or mirror-isomer-enriched form or rotational isomer thereof, and a pharmaceutically acceptable excipient.

28. The pharmaceutical composition of claim 27, wherein the compound is selected from the following:

29. Use of a compound of any one of claims 1 to 26, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer thereof, or a pharmaceutical composition of claim 27 or 28, for the preparation of a medicament for the treatment of a group of diseases or conditions selected from the group consisting of: cancer, inflammation, and neurological or progenitor cell / stem cell diseases.

30. Use of a compound of any one of claims 1 to 26, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotopically enriched or mirror-isomer-enriched form or rotational isomer, or a pharmaceutical composition of claim 27 or 28, for the preparation of a medicament for inhibiting the binding of chemokines I-TAC or SDF-1 to the CXCR7 receptor, wherein the inhibition comprises contacting a compound of any one of claims 1 to 26, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotopically enriched or mirror-isomer-enriched form or rotational isomer, or a pharmaceutical composition of claim 27 or 28, with a cell expressing the CXCR7 receptor for a duration sufficient to inhibit the binding of the chemokines to the CXCR7 receptor.

31. As claimed in claim 30, wherein the compound is selected from the following:

32. Use of a compound of any one of claims 1 to 26, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotopically enriched or mirror-isomer-enriched form or rotational isomer thereof, or a pharmaceutical composition of claim 27 or 28, for the preparation of a medicament for imaging a tumor, organ, or tissue, the imaging comprising: (a) to deliver to an individual requiring such imaging a radiolabeled or detectable form of any of the compounds described in claims 1 to 26, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer, or a pharmaceutical composition as described in claims 27 or 28; and (b) to detect the compound, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer, or the pharmaceutical composition, to determine where the compound, or a pharmaceutically acceptable salt, hydrate, N-oxide, isotope-enriched or mirror-isomer-enriched form or rotational isomer, or the pharmaceutical composition is concentrated in the individual.

33. As claimed in claim 32, wherein the compound is radiolabeled.

34. A method for detecting elevated CXCR7 content in a sample, the method comprising: (a) contacting a sample suspected of having elevated CXCR7 content with a radiolabeled or detectable form of a compound as claimed in any of claims 1 to 26; (b) determining the content of a CXCR7-bound compound present in the sample to determine the content of CXCR7 present in the sample; and (c) comparing the content determined in step (b) with a control sample to determine whether elevated CXCR7 content is present in the sample.

35. The method of claim 34, wherein the compound is radiolabeled.