Itraconazole analogs and uses thereof

Abstract

Itraconazole, a widely used antifungal drug, has been found to possess potent antiangiogenic and anti-hedgehog activities and has shown promising antitumor activity in several human clinical studies. However, its widespread use in cancer treatment has been limited by its potent inhibition of the drug-metabolizing enzyme CYP3A4, which can cause drug-drug interactions. In an effort to eliminate itraconazole's CYP3A4 inhibition while retaining its antiangiogenic activity, we synthesized a series of itraconazole derivatives. The newly synthesized itraconazole analogs were evaluated for their cytotoxicity and CYP3A4 inhibitory activity against human umbilical vein endothelial cells (HUVECs).

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This patent application claims the benefit under 35 U.S.C. §119(e) of U.S. Provisional Patent Application No. 62 / 742,046, filed on October 5, 2018. This application also claims the benefit of application No. 16 / 152,008, filed October 4, 2018; application No. 16 / 152,008 is a continuation-in-part of application No. 15 / 162,524, filed May 23, 2016, now abandoned; application No. 15 / 162,524 is a continuation-in-part of application No. 14 / 343,040, filed April 10, 2014, now issued as U.S. Patent No. 9,346,791; application No. 14 / 343,040 is a 35 U.S.C. §371 national phase application of International Application No. PCT / US2012 / 054306, filed September 7, 2012; International Application No. PCT / US2012 / 054306 claims the benefit of U.S. Provisional Patent Application No. 61 / 531,819 under 35 U.S.C. §119(e). The disclosure of each of the foregoing applications is considered part of the disclosure of the present application and is incorporated by reference in its entirety into the disclosure of the present application herein.

[0003] Government Support Statement

[0004] This disclosure was made with government support under Grant No. 1R01CA184103-01A1 awarded by the National Institutes of Health and the Flight Attendant Medical Research Institute (FAMRI). The government has certain rights in this disclosure.

[0005] field

[0006] The present disclosure relates generally to derivatives of itraconazole, and more particularly to itraconazole analogs and compositions as drugs for treating disease.

[0007] background

[0008] Itraconazole is well known for its use as a clinical agent for treating a broad spectrum of fungal infections. However, it has been shown to also possess potent in vitro and in vivo anti-angiogenic activity, and additionally inhibits both Hedgehog (Hh) signaling and the growth of murine medulloblastoma (MB) allografts with dysregulated Hh activity. These observations have led to an expansion of itraconazole's potential therapeutic applications and have even triggered the evaluation of the compound in four ongoing cancer clinical trials.

[0009] Itraconazole has been found to possess potent anti-angiogenic and anti-hedgehog activities, showing promising antitumor activity in several human clinical studies. However, its widespread use in the treatment of cancer has been limited by its potent inhibition of human hepatic cytochrome P450 3A4 (CYP3A4), a drug-metabolizing enzyme that can cause drug-drug interactions. A major limitation of itraconazole as a novel anticancer drug is its potent inhibitory activity against CYP3A4. CYP3A4 is a major xenobiotic-metabolizing enzyme and contributes to the metabolism of approximately 50% of prescription drugs and most anticancer drugs. CYP3A4 inhibition can lead to reduced metabolism of other drugs that may cause undesirable side effects, thus preventing its combination with other drugs in cancer therapy. Many anticancer drugs, particularly those that inhibit angiogenesis, are most effective when used in combination with other drugs. Therefore, there is a need to develop novel itraconazole analogs with reduced or no CYP3A4 inhibition while retaining their anti-angiogenic activity.

[0010] Overview

[0011] The present disclosure is based on the seminal discovery of a series of itraconazole analog compounds that are potent anti-angiogenic agents with reduced CYP3A4 inhibition.

[0012] Provided herein are compounds of structural formula (I), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0013]

[0014] wherein U is selected from the group consisting of hydrogen, alkyl, arylalkyl, alkoxyalkyl, arylalkoxy, alkynylalkyl, alkylalkynylalkyl, alkenylalkyl, alkylalkenylalkyl, cycloalkyl, cyanoalkyl, cycloalkylalkyl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroarylalkyl, heterocycloalkylalkyl, haloalkyl, halogen, amino, amido, nitro, and cyano, any of which may be optionally substituted;

[0015] R 1 、R 2 、R 3 and R 4 each independently selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0016] A is CR 5 or N;

[0017] B is CR 6 or N;

[0018] E is CR 7 or N;

[0019] L is CR 8 or N;

[0020] Q is O or CH2;

[0021] R 5 、R 6 、R 7 and R 8 each independently selected from the group consisting of hydrogen, alkoxy, alkyl, alkynyl, amino, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0022] T is OR 9 or hydrogen;

[0023] R 9 is hydrogen or optionally substituted alkyl;

[0024] G is (CH2) z , or G and R 9 Together with the atom(s) to which they are attached, may optionally be linked together to form a monocyclic heterocycle, including but not limited to dioxolane;

[0025] z is an integer between 0 and 2;

[0026] m is an integer between 0 and 5;

[0027] n and q are each independently an integer between 0 and 2;

[0028] p is an integer between 0 and 4;

[0029] D is selected from the group consisting of:

[0030]

[0031] in, Is a single bond or a double bond;

[0032] R 10 selected from the group consisting of hydrogen, alkyl, arylalkyl, alkoxyalkyl, arylalkoxy, alkynylalkyl, alkylalkynylalkyl, alkenylalkyl, alkylalkenylalkyl, cycloalkyl, cyanoalkyl, cycloalkylalkyl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroarylalkyl, heterocycloalkylalkyl, and alkylsulfonyl, any of which may be optionally substituted;

[0033] I is (CH2) r or NH;

[0034] J is (CH2) s or NH;

[0035] K is (CH2) t or NH;

[0036] r, s and t are each independently an integer between 0 and 4.

[0037] In certain embodiments, U is not

[0038] Also provided herein are compounds of structural formula (II), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0039]

[0040] R 1 and R 2 each independently selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0041] n is an integer between 0 and 5;

[0042] R 3 Selected from the group consisting of hydrogen, alkyl, arylalkyl, alkoxyalkyl, arylalkoxy, alkynylalkyl, alkylalkynylalkyl, alkenylalkyl, alkylalkenylalkyl, cycloalkyl, cyanoalkyl, cycloalkylalkyl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroarylalkyl, heterocycloalkylalkyl and alkylsulfonyl, any of which may be optionally substituted.

[0043] Also provided herein are compounds of structural formula (III), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0044]

[0045] R 1 selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0046] R 2 、R 3 and R 4 each independently selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0047] n and p are each independently an integer between 0 and 2;

[0048] m is an integer between 0 and 4.

[0049] Also provided herein are compounds of structural formula (IV), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0050]

[0051] Wherein U is selected from the group consisting of: and optionally substituted phenyl;

[0052] R 1 、R 2 、R 3 and R 4 each independently selected from the group consisting of hydrogen, trifluoromethyl, alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0053] each of A, B, D, E, M, N, X and Y is independently CH or N;

[0054] m is an integer between 0 and 5;

[0055] n and p are each independently an integer between 0 and 2;

[0056] q is 0, 1, 2, or 3.

[0057] Also disclosed herein is a method for treating a disease in a subject, the method comprising administering an effective amount of a compound according to formula (I), formula (II), formula (III) or formula (IV). In some embodiments, the disease is cancer. In some embodiments, the cancer is selected from the group consisting of: central nervous system (CNS) cancer, lung cancer, breast cancer, colorectal cancer, prostate cancer, gastric cancer, liver cancer, cervical cancer, esophageal cancer, bladder cancer, non-Hodgkin's lymphoma, leukemia, pancreatic cancer, kidney cancer, endometrial cancer, head and neck cancer, lip cancer, oral cancer, thyroid cancer, brain cancer, ovarian cancer, kidney cancer, melanoma, gallbladder cancer, laryngeal cancer, multiple myeloma, nasopharyngeal cancer, Hodgkin's lymphoma, testicular cancer and Kaposi's sarcoma.

[0058] In some embodiments, the disease can be dependent on angiogenesis. In some embodiments, angiogenesis-dependent disease can be selected from the group consisting of: macular degeneration, diabetic retinopathy, hemangioma, colon polyps, precancerous skin lesions, uveitis, ocular melanoma, corneal neovascularization, primary pterygium, HSV stromal keratitis, HSV-1-induced corneal lymphangiogenesis, retinopathy of prematurity, retinal vein occlusion, corneal transplant rejection, neovascular glaucoma and iris redness (rubeosis). In some embodiments, the method further comprises administering a chemotherapeutic agent. The compound can be administered before, simultaneously with, or after the administration of the chemotherapeutic agent.

[0059] Also disclosed herein is a pharmaceutical preparation comprising a compound of formula (I), formula (II), formula (III) or formula (IV) and a pharmaceutically acceptable carrier. In some embodiments, the compound can be formulated into a delayed release product, a slow release product, an extended release product or a controlled release product. In some embodiments, the compound can be provided in a dosage form selected from an injectable dosage form, an infusible dosage form, an inhalable dosage form, an edible dosage form, an oral dosage form, a topical dosage form and a combination thereof. In some embodiments, the dosage form includes an enteric coating. BRIEF DESCRIPTION OF THE DRAWINGS

[0061] Figure 1 The structures of itraconazole and ketoconazole are shown;

[0062] Figure 2 The CYP3A4 enzyme activity in the presence of different concentrations of itraconazole, compound 59, compound 51, compound 45, and compound 48 is shown;

[0063] Figure 3 A- Figure 3Figure B shows that compound 48 inhibits HUVEC tube formation. HUVECs were seeded on Matrigel and treated with 5 μM itraconazole, 1 μM compound 48, 5 μM compound 48, or DMSO for 20 h. (A) The cells were then stained with calcein-AM, and the vascular network was imaged using fluorescence microscopy. (B) The total tube length from the fluorescence images was quantified using imageJ software and plotted using GraphPad Prism. Data, mean ± SE of three independent experiments. (*p < 0.01);

[0064] Figure 4 It is shown that itraconazole and compound 48 induce NPC phenotype at 0.2 μM. HUVECs were treated with 0.2 μM itraconazole, 0.2 μM compound 48, or DMSO for 24 h. Intracellular cholesterol was visualized using filipin staining, and fluorescence images were captured under a confocal microscope.

[0065] Figure 5 Compound 48 was shown to dose-dependently activate AMPK and inhibit mTOR in HUVECs. HUVECs were treated with 0.01 μM, 0.03 μM, 0.1 μM, 0.3 μM, 1 μM of compound 48 or DMSO for 24 h. Cell lysates were subjected to Western blotting.

[0066] Figure 6 Shown are the dose-response curves for CYP3A4 enzyme inhibition by itraconazole and compound 4, compound 8, compound 14, compound 15, and compound 101;

[0067] Figure 7A Inhibition of HUVEC tube formation is shown when cells were stained with Calcein-AM and the vascular network was imaged using fluorescence microscopy. HUVECs were seeded on Matrigel-coated plates and treated with DMSO or 3 μM of compound for 24 h;

[0068] Figure 7B Inhibition of HUVEC tube formation is shown when total tube length from fluorescent images was quantified using ImageJ software and plotted using GraphPad Prism. HUVECs were seeded on Matrigel-coated plates and treated with DMSO or 3 μM of compound for 24 h. Data represent the mean ± SD of three independent experiments.

[0069] Figure 8Induction of the NPC phenotype is shown. HUVECs were treated with 0.2 μM itraconazole, compound 8, compound 14, compound 15, compound 24, or DMSO for 24 h. Intracellular cholesterol was stained with filipin, and fluorescence images were captured using an LSM710 confocal microscope with a 25× objective.

[0070] Figure 9 A shows the predicted binding modes of itraconazole and compound 14 to NPC1 (PDB code: 5I31) using AutoDock Vina software;

[0071] Figure 9 B shows the predicted interaction between compound 14 and SSD;

[0072] Figure 10A Inhibition of VEGFR2 and AMPK / mTOR in HUVECs was shown, wherein HUVECs were treated with 0.05 μM, 0.25 μM, 0.5 μM, 1 μM or 2 μM compound 14 or DMSO for 24 hours. VEGFR2, p-ACC, total ACC, p-S6K, total S6K and β-actin were analyzed by Western blot; and

[0073] Figure 10B Inhibition of VEGFR2 and AMPK / mTOR in HUVECs is shown, wherein HUVECs were treated with DMSO, 2 μM compound 14, or itraconazole, wherein the cells were stained with VEGFR2, GM130 antibodies, and DAPI. Images were captured using an LSM 700 confocal microscope.

[0074] Details

[0075] Angiogenesis, the formation of new blood vessels, plays a key role in the onset and progression of cancer and many other human diseases. Inhibition of angiogenesis has become an important strategy in the fight against cancer, as highlighted by the clinical introduction of numerous angiogenesis inhibitors. In an effort to repurpose existing drugs as novel angiogenesis inhibitors, we previously discovered that the antifungal drug itraconazole possesses potent antiangiogenic activity. The molecular target underlying itraconazole's antifungal activity is lanosterol 14α-demethylase (14-DM). However, itraconazole only exhibits weak inhibition of human 14-DM, which excludes 14-DM as a relevant target for itraconazole's antiangiogenic activity. Instead, we have identified voltage-dependent anion channel (VDAC) 1 and Niemann-Pick type C (NPC) 1 as direct targets of itraconazole. We have shown that binding of itraconazole to NPC1 leads to inhibition of cholesterol transport from endolysosomes, which in turn induces the NPC1 phenotype. Itraconazole binds to VDAC1, blocking mitochondrial ATP biosynthesis, increasing the cytosolic AMP / ATP ratio and activating AMPK. The inhibition of cholesterol transport and activation of AMPK lead to synergistic inhibition of mTOR signaling. Itraconazole's unique mechanism of action distinguishes itraconazole from rapamycin (a direct inhibitor of mTOR) and most other triazole antifungal drugs, such as ketoconazole, which lack angiogenic activity. This new mechanistic insight, along with other preclinical results, has led to the entry of itraconazole into multiple Phase II clinical trials for the treatment of prostate cancer, non-small cell lung cancer, basal cell carcinoma, and other cancers.

[0076] A major limitation of itraconazole as a novel anticancer drug is that it is a potent inhibitor of human hepatic cytochrome P450 3A4 (CYP3A4). CYP3A4 is the major xenobiotic-metabolizing enzyme and contributes to the metabolism of approximately 50% of prescription medications and most anticancer drugs. Inhibition of CYP3A4 can lead to reduced metabolism of other drugs that may cause undesirable side effects, thus preventing the combination of itraconazole with those drugs in cancer therapy. Many anticancer drugs, particularly those that inhibit angiogenesis, are most effective when used in combination with other drugs. Therefore, there is a need to develop novel itraconazole analogs with reduced or no CYP3A4 inhibition while retaining their antiangiogenic activity.

[0077] In previous work, we identified cis-stereochemical itraconazole stereoisomers with a dioxalane moiety (cis-2R,4S) that exhibited enhanced antiangiogenic activity and significantly reduced hepatotoxicity. See U.S. Patent No. 9,346,791. We also found that a sec-butyl side chain, or a side chain of similar length, is required for antiangiogenic activity. The triazole moiety of itraconazole is a key pharmacophore required for its binding to the heme group of the antifungal target 14-DM and the heme group of CYP3A4. However, little is known about the importance of the triazole moiety in itraconazole's antiangiogenic activity.

[0078] In an effort to identify novel itraconazole analogs with reduced or no CYP3A4 inhibition while retaining their antiangiogenic potency, we systematically modified 1,2,4-triazole (R 1 ) moiety. Here, we report the SAR (structure-activity relationship) studies of novel triazole analogs of itraconazole for their antiangiogenic activity and CYP3A4 inhibition, as well as the successful identification of a class of itraconazole analogs that possess potent antiangiogenic activity but significantly reduced CYP3A4 inhibitory activity compared to itraconazole.

[0079] Provided herein are compounds of structural formula (I), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0080]

[0081] wherein U is selected from the group consisting of hydrogen, alkyl, arylalkyl, alkoxyalkyl, arylalkoxy, alkynylalkyl, alkylalkynylalkyl, alkenylalkyl, alkylalkenylalkyl, cycloalkyl, cyanoalkyl, cycloalkylalkyl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroarylalkyl, heterocycloalkylalkyl, haloalkyl, halogen, amino, amido, nitro, and cyano, any of which may be optionally substituted;

[0082] R 1 、R 2 、R 3 and R 4 each independently selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0083] A is CR 5 or N;

[0084] B is CR 6or N;

[0085] E is CR 7 or N;

[0086] L is CR 8 or N;

[0087] Q is O or CH2;

[0088] R 5 、R 6 、R 7 and R 8 each independently selected from the group consisting of hydrogen, alkoxy, alkyl, alkynyl, amino, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0089] T is OR 9 or hydrogen;

[0090] R 9 is hydrogen or optionally substituted alkyl;

[0091] G is (CH2) z , or G and R 9 Together with the one or more atoms to which they are attached, may optionally be linked together to form a monocyclic heterocycle, including but not limited to dioxolane;

[0092] z is an integer between 0 and 2;

[0093] m is an integer between 0 and 5;

[0094] n and q are each independently an integer between 0 and 2;

[0095] p is an integer between 0 and 4;

[0096] D is selected from the group consisting of:

[0097]

[0098] in, Is a single bond or a double bond;

[0099] R 10 selected from the group consisting of hydrogen, alkyl, arylalkyl, alkoxyalkyl, arylalkoxy, alkynylalkyl, alkylalkynylalkyl, alkenylalkyl, alkylalkenylalkyl, cycloalkyl, cyanoalkyl, cycloalkylalkyl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroarylalkyl, heterocycloalkylalkyl, and alkylsulfonyl, any of which may be optionally substituted;

[0100] I is (CH2) r or NH;

[0101] J is (CH2) s or NH;

[0102] K is (CH2) t or NH;

[0103] r, s and t are each independently an integer between 0 and 4.

[0104] In certain embodiments, U is not

[0105] Also provided herein are compounds of structural formula (II), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0106]

[0107] R 1 and R 2 each independently selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0108] n is an integer between 0 and 5;

[0109] R 3 Selected from the group consisting of hydrogen, alkyl, arylalkyl, alkoxyalkyl, arylalkoxy, alkynylalkyl, alkylalkynylalkyl, alkenylalkyl, alkylalkenylalkyl, cycloalkyl, cyanoalkyl, cycloalkylalkyl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroarylalkyl, heterocycloalkylalkyl and alkylsulfonyl, any of which may be optionally substituted.

[0110] Also provided herein are compounds of structural formula (III), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0111]

[0112] R 1 selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0113] R 2 、R 3 and R 4each independently selected from the group consisting of alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0114] n and p are each independently an integer between 0 and 2;

[0115] m is an integer between 0 and 4.

[0116] Also provided herein are compounds of structural formula (IV), or optically pure stereoisomers or pharmaceutically acceptable salts, hydrates, solvates or prodrugs thereof,

[0117]

[0118] Wherein U is selected from the group consisting of: and optionally substituted phenyl;

[0119] R 1 、R 2 、R 3 and R 4 each independently selected from the group consisting of hydrogen, trifluoromethyl, alkoxy, alkyl, alkynyl, amino, amido, halogen, hydroxy, haloalkyl, perhaloalkyl, perhaloalkoxy, nitro, and cyano, any of which may be optionally substituted;

[0120] each of A, B, D, E, M, N, X and Y is independently CH or N;

[0121] m is an integer between 0 and 5;

[0122] n and p are each independently an integer between 0 and 2;

[0123] q is 0, 1, 2, or 3.

[0124] As used herein, the following terms have the indicated meanings.

[0125] When a range of values is disclosed and the notation "from n1 ... to n2" or "between n1 ... and n2" is used, where n1 and n2 are numbers, then unless otherwise specified, the notation is intended to include the numbers themselves and the range between them. The range can be between and include the end values being integers or continuous. By way of example, the range "from 2 to 6 carbons" is intended to include two, three, four, five, and six carbons, since carbon appears in integer units. In contrast, by way of example, the range "from 1 μM to 3 μM (micromolar)" is intended to include 1 μM, 3 μM, and everything in between up to any number of significant figures (e.g., 1.255 μM, 2.1 μM, 2.9999 μM, etc.). When n is set to 0 in the context of "0 carbon atoms," it is intended to indicate a bond or null.

[0126] As used herein, the term "about" is intended to qualify the numerical value it modifies, expressing such value as a variation within a margin of error. When no specific margin of error is recited, such as the standard deviation of the mean value given in a graph or data table, the term "about" should be understood to mean a range that will encompass the recited value and that will be included by rounding up or down to that number, taking into account significant figures.

[0127] As used herein, alone or in combination, the term "acyl" refers to a carbonyl group attached to an alkenyl, alkyl, aryl, cycloalkyl, heteroaryl, heterocycle, or any other moiety, wherein the atom attached to the carbonyl group is carbon. An "acetyl" group refers to a -C(O)CH3 group. An "alkylcarbonyl" or "alkanoyl" group refers to an alkyl group attached to the parent molecular moiety through a carbonyl group. Examples of such groups include methylcarbonyl and ethylcarbonyl. Examples of acyl groups include formyl, alkanoyl, and aroyl.

[0128] As used herein, alone or in combination, the term "alkenyl" refers to a straight or branched hydrocarbon group having one or more double bonds and containing from 2 to 20 carbon atoms. In certain embodiments, the alkenyl will contain from 2 to 6 carbon atoms. The term "alkenylene" refers to a carbon-carbon double bond system attached at two or more positions, such as vinylene [(-CH=CH-), (-C::C-)]. Examples of suitable alkenyl groups include vinyl, propenyl, 2-methylpropenyl, 1,4-butadienyl and similar groups. Unless otherwise specified, the term "alkenyl" may include "alkenylene" groups.

[0129] As used herein, the term "alkoxy" refers to an alkyl ether group, wherein the term alkyl is as defined below. Examples of suitable alkyl ether groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy, and the like.

[0130] As used herein alone or in combination, the term "alkyl" refers to a straight or branched chain alkyl group containing from 1 to 20 carbon atoms. In certain embodiments, the alkyl group will comprise from 1 to 10 carbon atoms. In other embodiments, the alkyl group will comprise from 1 to 6 carbon atoms. The alkyl group can be optionally substituted as defined herein. The example of an alkyl group includes methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, hexyl, octyl, nonyl and similar groups. As used herein alone or in combination, the term "alkylene" refers to a saturated aliphatic group derived from a straight or branched chain saturated hydrocarbon attached at two or more positions, such as methylene (-CH2-). Unless otherwise specified, the term "alkyl" can include an "alkylene" group.

[0131] The term "alkylamino," as used herein, alone or in combination, refers to an alkyl group attached to the parent molecular moiety through an amino group. Suitable alkylamino groups can be monoalkylated or dialkylated to form groups such as, for example, N-methylamino, N-ethylamino, N,N-dimethylamino, N,N-ethylmethylamino and the like.

[0132]

[0046] The term "alkylidene," as used herein, alone or in combination, refers to an alkenyl group wherein one carbon atom of the carbon-carbon double bond belongs to the moiety to which the alkenyl group is attached.

[0133] As used herein, "alkylthio" refers to an alkylthioether (RS-) group, wherein the term alkyl is as defined above, and wherein the sulfur may be mono- or di-oxidized. Examples of suitable alkylthioether groups include methylthio, ethylthio, n-propylthio, isopropylthio, n-butylthio, isobutylthio, sec-butylthio, tert-butylthio, methylsulfonyl, ethylsulfinyl, and the like.

[0134] As used herein alone or in combination, the term "alkynyl" refers to a straight or branched hydrocarbon group having one or more triple bonds and containing from 2 to 20 carbon atoms. In certain embodiments, the alkynyl comprises from 2 to 6 carbon atoms. In other embodiments, the alkynyl comprises from 2 to 4 carbon atoms. The term "alkynylene" refers to a carbon-carbon triple bond attached at two positions, such as ethynylene (-C:::C-, -C≡C-). The example of an alkynyl group includes ethynyl, propynyl, hydroxypropynyl, butyn-1-yl, butyn-2-yl, pentyn-1-yl, 3-methylbutyn-1-yl, hexyn-2-yl and similar groups. Unless otherwise specified, the term "alkynyl" may include an "alkynylene" group.

[0135] As used herein, alone or in combination, the terms "amido" and "carbamoyl" refer to an amino group as described below attached to the parent molecular moiety through a carbonyl group, or vice versa. The term "Camido," as used herein, alone or in combination, refers to a C(=O)NR2 group with R as defined herein. The term "Namido," as used herein, alone or in combination, refers to a RC(=O)NH group with R as defined herein. The term "acylamino," as used herein, alone or in combination, includes an acyl group attached to the parent moiety through an amino group. An example of an "acylamino" group is acetylamino (CH3C(O)NH-).

[0136] As used herein, the term "amino" refers to -NRR', wherein R and R' are independently selected from the group consisting of hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may itself be optionally substituted. In addition, R and R' may combine to form heterocycloalkyl, any of which may be optionally substituted.

[0137] As used herein, alone or in combination, the term "aryl" means a carbocyclic aromatic system containing one, two or three rings, wherein such polycyclic ring systems are fused together. The term "aryl" includes aromatic groups such as phenyl, naphthyl, anthracenyl and phenanthrenyl.

[0138]

[0066] The term "arylalkenyl" or "aralkenyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkenyl group.

[0139]

[0066] The term "arylalkoxy" or "aralkoxy," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkoxy group.

[0140]

[0066] The term "arylalkyl" or "aralkyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkyl group.

[0141]

[0066] The term "arylalkynyl" or "aralkynyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkynyl group.

[0142]

[0014] The term "arylalkanoyl" or "aralkanoyl" or "aroyl," as used herein, alone or in combination, refers to an acyl radical derived from an aryl-substituted alkanecarboxylic acid, such as benzoyl, naphthoyl, phenylacetyl, 3-phenylpropionyl (hydrocinnamoyl), 4-phenylbutyryl, (2-naphthyl)acetyl, 4-chlorohydrocinnamoyl, and the like.

[0143] The term aryloxy, as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an oxy group.

[0144]

[0026] The terms "benzo" and "benz," as used herein, alone or in combination, refer to a divalent radical C6H4= derived from benzene. Examples include benzothiophene and benzimidazole.

[0145] The term "carbamate," as used herein, alone or in combination, refers to an ester of a carbamic acid (-NHCOO-), which can be attached to the parent molecular moiety from either the nitrogen terminus or the acid terminus, and which can be optionally substituted as defined herein.

[0146] The term "Ocarbamyl," as used herein, alone or in combination, refers to an OC(O)NRR' group with R and R' as defined herein.

[0147] The term "N-carbamyl," as used herein, alone or in combination, refers to a ROC(O)NR' group with R and R' as defined herein.

[0148] As used herein, the term "carbonyl," when alone includes formyl [-C(O)H], and when in combination is a -C(O)- group.

[0149] As used herein, the term "carboxyl" or "carboxy" refers to -C(O)OH or the corresponding "carboxylate" anion, such as in a carboxylate salt. An "Ocarboxyl" group refers to a RC(O)O- group, where R is as defined herein. A "Ccarboxyl" group refers to a -C(O)OR group, where R is as defined herein.

[0150]

[00146] The term "cyano," as used herein, alone or in combination, refers to -CN.

[0151] As used herein alone or in combination, the term "cycloalkyl" or alternatively "carbocycle" refers to a saturated or partially saturated monocyclic, bicyclic or tricyclic alkyl group, wherein each ring portion comprises from 3 to 12 carbon atom ring members, and it can optionally be an optionally substituted benzo-fused ring system as defined herein. In certain embodiments, the cycloalkyl will comprise from 5 to 7 carbon atoms. Examples of such cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, tetrahydronaphthyl, indanyl, octahydronaphthyl, 2,3-dihydro-1H-indenyl, adamantyl and similar groups. As used herein, "bicyclic" and "tricyclic" are intended to include fused ring systems such as decahydronaphthalene, octahydronaphthalene and polycyclic (multi-center) saturated or partially unsaturated types. The latter type of isomer is typically exemplified by bicyclo [1,1,1] pentane, camphor, adamantane and bicyclo [3,2,1] octane.

[0152]

[00146] The term "ester," as used herein, alone or in combination, refers to a carboxyl group bridging two moieties linked at carbon atoms.

[0153]

[00146] The term "ether," as used herein, alone or in combination, refers to an oxy group bridging two moieties linked at carbon atoms.

[0154]

[0046] The term "halo" or "halogen," as used herein, alone or in combination, refers to fluoro, chloro, bromo, or iodo.

[0155]

[0066] The term "haloalkoxy," as used herein, alone or in combination, refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom.

[0156] As used herein alone or in combination, the term "haloalkyl" refers to an alkyl group with the implication as defined above, wherein one or more hydrogens are replaced by halogen. Specifically include monohaloalkyl groups, dihaloalkyl groups and polyhaloalkyl groups. For an example, a monohaloalkyl group can have an iodine atom, a bromine atom, a chlorine atom or a fluorine atom in the group. Dihaloalkyl groups and polyhaloalkyl groups can have a combination of two or more identical halogen atoms or different halo groups. The example of a haloalkyl group includes fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl and dichloropropyl. "haloalkylene" refers to a haloalkyl group attached at two or more positions. Example includes fluoromethylene (-CFH-), difluoromethylene (-CFH-), chloromethylene (-CHCl-) and similar groups.

[0157] As used herein alone or in combination, term " assorted alkyl " refers to a stable straight chain or branched or cyclic hydrocarbon group or its combination comprising from 1 to 3 degrees of unsaturation, by the carbon atom of the number stated and from one to three heteroatoms selected from the group consisting of O, N and S, and wherein nitrogen-atoms and sulphur atoms can be optionally oxidized, and nitrogen heteroatoms can be optionally quaternized. One or more heteroatoms (heteroatom(s)) O, N and S can be located at any internal position of assorted alkyl group. Up to two heteroatoms can be continuous, such as, for example, -CH2-NH-OCH3.

[0158] As used herein, alone or in combination, the term "heteroaryl" refers to a 3- to 7-membered unsaturated heteromonocyclic ring, or a fused monocyclic, bicyclic, or tricyclic ring system comprising at least one atom selected from the group consisting of O, S, and N, wherein at least one of the fused rings is aromatic. In certain embodiments, the heteroaryl group will contain from 5 to 7 carbon atoms. The term also includes fused polycyclic groups wherein a heterocyclic ring is fused to an aryl ring, wherein a heteroaryl ring is fused to another heteroaryl ring, wherein a heteroaryl ring is fused to a heterocycloalkyl ring, or wherein a heteroaryl ring is fused to a cycloalkyl ring. Examples of heteroaryl groups include pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridinyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl, pyranyl, furanyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolinyl, isoquinolinyl, quinoxalinyl, quinazolinyl, indazolyl, benzotriazolyl, , benzodioxolyl, benzopyranyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzofuranyl, benzothienyl, chromonyl, coumarinyl, tetrahydroquinolinyl, tetrazolypyridazinyl, tetrahydroisoquinolinyl, thienopyridinyl, furopyridinyl, pyrrolopyridinyl and the like. Exemplary tricyclic heterocyclic groups include carbazolyl, benzindolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl and the like.

[0159] As used herein, alone or in combination, the terms "heterocycloalkyl" and, interchangeably, "heterocycle" each refer to a saturated, partially unsaturated, or fully unsaturated monocyclic, bicyclic, or tricyclic heterocyclic group comprising at least one heteroatom as a ring member, wherein each of the heteroatoms may be independently selected from the group consisting of nitrogen, oxygen, and sulfur. In certain embodiments, the heterocycloalkyl will comprise from 1 to 4 heteroatoms as ring members. In further embodiments, the heterocycloalkyl will comprise from 1 to 2 heteroatoms as ring members. In certain embodiments, the heterocycloalkyl will comprise from 3 to 8 ring members in each ring. In further embodiments, the heterocycloalkyl will comprise from 3 to 7 ring members in each ring. In yet further embodiments, the heterocycloalkyl will comprise from 5 to 6 ring members in each ring. "Heterocycloalkyl" and "heterocycle" are intended to include sulfones, sulfoxides, N-oxides of tertiary nitrogen ring members, and carbocyclic-fused and benzo-fused ring systems; in addition, both terms also include systems in which the heterocycle is fused to an aryl group as defined herein or to another heterocyclic group. Examples of heterocyclic groups include aziridinyl, azetidinyl, 1,3-benzodioxolyl, dihydroisoindolyl, dihydroisoquinolinyl, dihydrocinnolinyl, dihydrobenzodioxinyl, dihydro[1,3]oxazolo[4,5-b]pyridinyl, benzothiazolyl, dihydroindolyl, dihydropyridinyl, 1,3-dioxanyl, 1,4-dioxanyl, 1,3-dioxolanyl, isoindolinyl, morpholinyl, piperazinyl, pyrrolidinyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl, and the like. Unless specifically prohibited, heterocyclic groups may be optionally substituted.

[0160] The term "hydrazino," as used herein, alone or in combination, refers to two amino groups linked by a single bond, ie, -NN-.

[0161]

[0046] The term "hydroxy," as used herein, alone or in combination, refers to -OH.

[0162] The term "hydroxyalkyl," as used herein, alone or in combination, refers to a hydroxy group attached to the parent molecular moiety through an alkyl group.

[0163] The term "imino," as used herein, alone or in combination, refers to =N-.

[0164] The term "iminohydroxy," as used herein, alone or in combination, refers to =N(OH) and =NO-.

[0165] The phrase "in the backbone" refers to the longest continuous or adjacent chain of carbon atoms starting at the point of attachment of a group to the compound of any of the formulae disclosed herein.

[0166] The term "isocyanato" refers to an -NCO group.

[0167] The term "isothiocyanato" refers to a -NCS group.

[0168] The phrase "linear chain of atoms" refers to the longest straight chain of atoms independently selected from carbon, nitrogen, oxygen and sulfur.

[0169] As used herein, alone or in combination, the term "lower," unless otherwise specifically defined, means containing from 1 to 6 and including 6 carbon atoms.

[0170] The term "lower aryl," as used herein, alone or in combination, means phenyl or naphthyl, which may be optionally substituted as specified.

[0171] As used herein, the term "lower heteroaryl," alone or in combination, means: 1) a monocyclic heteroaryl comprising five or six ring members, of which between one and four may be heteroatoms selected from the group consisting of O, S, and N; or 2) a bicyclic heteroaryl wherein each of the fused rings comprises five or six ring members, between which one and four heteroatoms selected from the group consisting of O, S, and N.

[0172] As used herein, alone or in combination, the term "lower cycloalkyl" means a monocyclic cycloalkyl group having between three and six ring members. Lower cycloalkyl groups may be unsaturated. Examples of lower cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0173] As used herein, the term "lower heterocycloalkyl" means a monocyclic heterocycloalkyl having between three and six ring members, of which between one and four ring members may be heteroatoms selected from the group consisting of O, S, and N. Examples of lower heterocycloalkyl groups include pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, and morpholinyl. Lower heterocycloalkyl groups may be unsaturated.

[0174] As used herein, alone or in combination, the term "lower amino" refers to -NRR', wherein R and R' are independently selected from the group consisting of hydrogen, lower alkyl, and lower heteroalkyl, any of which may be optionally substituted. In addition, R and R' of the lower amino group may be combined to form a five-membered or six-membered heterocycloalkyl, any of which may be optionally substituted.

[0175] The term "mercaptyl," as used herein, alone or in combination, refers to an RS- group, where R is as defined herein.

[0176]

[0014] The term "nitro," as used herein, alone or in combination, refers to -NO2.

[0177]

[00146] The term "oxy" or "oxa," as used herein, alone or in combination, refers to -O-.

[0178] The term "oxo," as used herein, alone or in combination, refers to =0.

[0179] The term "perhaloalkoxy" refers to an alkoxy group in which all of the hydrogen atoms are replaced by halogen atoms.

[0180]

[0046] The term "perhaloalkyl," as used herein, alone or in combination, refers to an alkyl group in which all of the hydrogen atoms are replaced by halogen atoms.

[0181] As used herein, the terms "sulfonate," "sulfonic acid," and "sulfonic acid group," alone or in combination, refer to the -SO3H group and its anion as the sulfonic acid is used in salt formation.

[0182] The term "sulfanyl," as used herein, alone or in combination, refers to -S-.

[0183]

[00146] The term "sulfinyl," as used herein, alone or in combination, refers to -S(O)-.

[0184] The term "sulfonyl," as used herein, alone or in combination, refers to -S(O)2-.

[0185] The term "N-sulfonylamino" refers to an RS(=0)2NR' group with R and R' as defined herein.

[0186] The term "S-sulfonylamino" refers to a S(=O)2NRR' group with R and R' as defined herein.

[0187] The terms "thia" and "thio," as used herein, alone or in combination, refer to an -S- group or an ether wherein the oxygen is replaced by sulfur. The oxygenated derivatives of the thio group, i.e., sulfinyl and sulfonyl, are included in the definitions of thia and thio.

[0188] The term "thiol," as used herein, alone or in combination, refers to a -SH group.

[0189] As used herein, the term "thiocarbonyl" when alone includes thioformyl -C(S)H, and when in combination is a -C(S)- group.

[0190] The term "N-thiocarbamyl" refers to a ROC(S)NR'- group with R and R' as defined herein.

[0191] The term "O-thiocarbamyl" refers to a -OC(S)NRR' group with R and R' as defined herein.

[0192] The term "thiocyanato" refers to a -CNS group.

[0193] The term "trihalomethanesulfonylamino" refers to a X3CS(O)2NR- group wherein X is a halogen and R is as defined herein.

[0194] The term "trihalomethanesulfonyl" refers to a X3CS(O)2- group wherein X is a halogen.

[0195] The term "trihalomethoxy" refers to a X3CO- group where X is a halogen.

[0196] As used herein, the term "trisubstituted silyl," alone or in combination, refers to an organosilicon group substituted at its three free valencies with groups as listed herein under the definition of substituted amino. Examples include trimethylsilyl, tert-butyldimethylsilyl, triphenylsilyl, and the like.

[0197] Any definition herein may be used in combination with any other definition to describe a composite structural group. By convention, the tail element of any such definition is the element attached to the parent moiety. For example, the composite group alkylamido would represent an alkyl group attached to the parent molecule via an amido group, and the term alkoxyalkyl would represent an alkoxy group attached to the parent molecule via an alkyl group.

[0198] When a group is defined as "empty", it means that the group does not exist.

[0199] The term "optionally substituted" means that the preceding group may be substituted or unsubstituted. When substituted, the substituents of the "optionally substituted" group may include, but are not limited to, one or more substituents independently selected from the following groups or specifically specified groups, alone or in combination: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloalkoxy, oxo , lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyl ester, lower formamido, cyano group, hydrogen, halogen, hydroxyl, amino, lower alkylamino, arylamino, acylamino, nitro, thiol, lower alkylthio, lower haloalkylthio, lower perhaloalkylthio, arylthio, sulfonate, sulfonic acid, trisubstituted silyl, N3, SH, SCH3, C(O)CH3, CO2CH3, CO2H, pyridyl, thiophene, furyl, lower carbamate and lower urea. Two substituents can be linked together to form a fused five-, six- or seven-membered carbocyclic or heterocyclic ring consisting of zero to three heteroatoms, for example, to form methylenedioxy or ethylenedioxy. Optionally substituted groups can be unsubstituted (e.g., -CH2CH3), fully substituted (e.g., -CF2CF3), monosubstituted (e.g., -CH2CH2F)) or substituted at any level between fully substituted and monosubstituted (e.g., -CH2CF3). Where substituents are recited without qualification as to substitution, both substituted and unsubstituted forms are encompassed. Where a substituent is qualified as "substituted," substituted forms are explicitly contemplated. Furthermore, different sets of optional substituents for a particular moiety may be defined as desired; in these cases, the optional substitutions will be as defined, typically immediately following the phrase "optionally substituted with."

[0200] Unless otherwise defined, otherwise term R or term R ', when occurring individually and not having number designation, refer to the part selected from the group consisting of: hydrogen, alkyl, cycloalkyl, heteroalkyl, aryl, heteroaryl and heterocycloalkyl, any of which can be optionally substituted. Such R groups and R ' groups should be understood as optionally substituted as defined herein. Regardless of whether R groups have number designation, each R group (including R, R ' and Rn, wherein n=(1,2,3, ... n)), each substituent and each term should be understood as being independent of one another in terms of selecting from group. If any variable, substituent or term (for example, aryl, heterocycle, R etc.) occurs more than once in a formula or general structure, then its definition is independent of the definition when other occur at each time when it occurs. Those skilled in the art will also recognize that some groups can be attached to the parent molecule, or can occupy the position in the element chain from either end as written. Thus, by way of example only, an asymmetric group such as -C(O)N(R)- may be attached to the parent moiety at either the carbon or the nitrogen.

[0201] Asymmetric centers are present in the compounds disclosed herein. These centers are designated by the symbols "R" or "S," depending on the configuration of the substituents around the chiral carbon atom. It should be understood that the present disclosure encompasses all stereochemical isomeric forms, including diastereomeric, enantiomeric, and epimeric forms, as well as d-isomers and l-isomers and mixtures thereof. Individual stereoisomers of a compound can be synthetically prepared from commercially available starting materials containing a chiral center, or by preparing a mixture of enantiomeric products, followed by separation such as conversion to a mixture of diastereoisomers, followed by separation or recrystallization, chromatography techniques, direct separation of enantiomers on a chiral chromatographic column, or any other suitable method known in the art. Specific stereochemical starting compounds are commercially available or can be prepared and resolved by techniques known in the art. In addition, the compounds disclosed herein can exist as geometric isomers. The present disclosure includes all cis (cis) isomers, trans (trans) isomers, cis (syn) isomers, trans (anti) isomers, cis (entgegen) (E) isomers and trans (zusammen) (Z) isomers and appropriate mixtures thereof. In addition, the compounds can exist as tautomers; all tautomeric isomers are provided by the present disclosure. In addition, the compounds disclosed herein can exist in unsolvated form as well as in solvated form with pharmaceutically acceptable solvents such as water, ethanol and similar solvents. Generally, the solvated form is considered equivalent to the unsolvated form.

[0202] The term "bond" refers to a covalent bond between two atoms or moieties when the atoms connected by the bond are considered to be part of a larger substructure. Unless otherwise specified, a bond can be a single bond, a double bond, or a triple bond. A dashed line between two atoms in a diagram of a molecule indicates that an additional bond may or may not be present at that position.

[0203] The term "optically pure stereoisomer" refers to the absolute difference between stereoisomers, such as enantiomeric or diastereomeric excess or the mole fraction of each enantiomer or diastereomer.

[0204] As used herein, the term "disease" is intended to be generally synonymous and used interchangeably with the terms "disorder" and "condition" (as in medical condition), as all reflect an abnormal condition of the human or animal body, or a part of the human or animal body, that impairs normal function, is usually manifested by distinctive signs and symptoms, and causes the human or animal to have reduced survival time or reduced quality of life.

[0205] The term "combination therapy" means the administration of two or more therapeutic agents to treat the therapeutic conditions or disorders described in this disclosure. Such administration encompasses the co-administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule having a fixed ratio of active ingredients or in multiple separate capsules for each active ingredient. In addition, such administration also encompasses the use of each type of therapeutic agent in a sequential manner. In either case, the treatment regimen will provide the beneficial effects of the drug combination in treating the conditions or disorders described herein.

[0206] As used herein, the term "inhibition" (and, by extension, "inhibitor") encompasses all forms of functional protein (e.g., enzyme, kinase, receptor, channel, etc.) inhibition, including neutral antagonism, inverse agonism, competitive inhibition, and non-competitive inhibition (such as allosteric inhibition). Inhibition can be expressed as IC50 as defined below.

[0207] "Inhibitor" is used herein to refer to a compound that exhibits an IC50 activity of no more than about 100 μM and more typically no more than about 50 μM relative to its target, as measured in an assay generally described herein below. "IC50" is the concentration of an inhibitor that reduces the activity of an enzyme to half the maximum level. It has been found that certain representative compounds of the present disclosure exhibit inhibition of the VEGFR2 or Hedgehog (Hh) pathway. In certain embodiments, the compound will exhibit an IC50 of no more than about 10 μM relative to the VEGFR2 or Hh pathway; in further embodiments, the compound will exhibit an IC50 of no more than about 5 μM relative to the VEGFR2 or Hh pathway; in yet further embodiments, the compound will exhibit an IC50 of no more than about 1 μM relative to the VEGFR2 or Hh pathway, as measured in a VEGFR2 or Hh pathway assay described herein. In yet further embodiments, the compound will exhibit an IC50 of no more than about 200 nM relative to the VEGFR2 or Hh pathway.

[0208] The term "therapeutically effective" is intended to qualify the amount of active ingredient used to treat a disease or disorder. This amount will achieve the goal of reducing or eliminating the disease or disorder.

[0209] The term "therapeutically acceptable" refers to those compounds (or salts, prodrugs, tautomers, zwitterionic forms, etc.) that are suitable for use in contact with the tissues of patients without excessive toxicity, irritation, and allergic response, commensurate with a reasonable benefit / risk ratio, and effective for their intended use.

[0210] As used herein, reference to "treatment" of a patient is intended to include prevention. The term "patient" refers to all mammals, including humans. Examples of patients include humans, cattle, dogs, cats, goats, sheep, pigs, and rabbits. Preferably, the patient is human.

[0211] The term "prodrug" refers to a compound that is made more active in vivo. Certain compounds disclosed herein may also exist as prodrugs, as described in Hydrolysis in Drug and Prodrug Metabolism: Chemistry, Biochemistry, and Enzymology (Testa, Bernard and Mayer, Joachim M. Wiley-VHCA, Zurich, Switzerland 2003). The prodrug of a compound described herein is a structurally modified form of the compound that readily undergoes chemical changes under physiological conditions to provide the compound. In addition, a prodrug can be converted into a compound by chemical or biochemical methods in an ex vivo environment. For example, a prodrug can be slowly converted into a compound when placed in a transdermal patch reservoir together with a suitable enzyme or chemical reagent. Prodrugs are generally useful because, in some cases, they can be more easily administered than a compound or parent drug. For example, they can be bioavailable by oral administration, while the parent drug is not bioavailable by oral administration. Prodrugs can also have improved solubility in pharmaceutical compositions compared to the parent drug. A variety of prodrug derivatives are known in the art, such as those that rely on hydrolytic cleavage or oxidative activation of the prodrug. Examples of prodrugs, but not limited thereto, would be compounds that are administered as esters ("prodrugs") but are then metabolically hydrolyzed to the carboxylic acid (active entity). Additional examples include peptidyl derivatives of the compounds.

[0212] The compounds disclosed herein may exist as therapeutically acceptable salts. The present disclosure includes the compounds listed above in the form of salts (including acid addition salts). Suitable salts include salts formed with organic and inorganic acids. Such acid addition salts will generally be pharmaceutically acceptable. However, salts that are not pharmaceutically acceptable salts may have utility in the preparation and purification of the compound in question. Base addition salts may also be formed and are pharmaceutically acceptable. For a more complete discussion of the preparation and selection of salts, reference is made to Pharmaceutical Salts: Properties, Selection, and Use (Stahl, P. Heinrich. Wiley-VCHA, Zurich, Switzerland, 2002).

[0213] As used herein, term " acceptable salt for the treatment of " represents salt or zwitterion form of compound disclosed herein, which are soluble or dispersible in water or oil, and are acceptable for the treatment as defined herein. Salt can be prepared during the final separation and purification of compound, or prepared individually by reacting the appropriate compound in the form of free alkali with suitable acid. Representative acid addition salts include acetate, adipate, alginate, L- ascorbate, aspartate, benzoate, benzenesulfonate (benzenesulfonate) (benzenesulfonate (besylate)), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, enanthate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate (hydroxyethylsulfonate), lactate, maleate, malonic acid The compounds disclosed herein include quaternary ammonium salts of methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; sulfates of dimethyl, diethyl, dibutyl, and diamyl; chlorides, bromides, and iodides of decyl, lauryl, myristyl, and steryl; and bromides of benzyl and phenethyl. Examples of acids that can be used to form therapeutically acceptable addition salts include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid and phosphoric acid, and organic acids such as oxalic acid, maleic acid, succinic acid and citric acid. Salts can also be formed by coordination of the compound with an alkali metal or alkaline earth metal ion. Thus, the present disclosure contemplates sodium, potassium, magnesium and calcium salts of the compounds disclosed herein and similar salts.

[0214] Abbreviations. HUVEC, human umbilical vein endothelial cells; NPC1, Niemann-Pick disease type C1; VDAC1, voltage-dependent anion channel 1; 14-DM, lanosterol 14-alpha demethylase; mTORC, mammalian target of rapamycin complex; SAR, structure–activity relationship; IC50, half-maximal inhibitory concentration; DMSO, dimethyl sulfoxide; DMF, dimethylformamide; EC50, half-maximal effective concentration; AMPK, 5′AMP-activated protein kinase; ATP, adenosine triphosphate; AMP, adenosine monophosphate; CYP3A4, cytochrome P450 3A4; THF, tetrahydrofuran; DCM, dichloromethane; TfOH, trifluoromethanesulfonic acid; NaH, sodium hydride; ACC1, acetyl CoA carboxylase 1; S6K, p70S6 kinase.

[0215] Base addition salts can be prepared by reacting a carboxyl group with a suitable alkali such as hydroxide, carbonate or bicarbonate of a metal cation or with ammonia or an organic primary amine, secondary amine or tertiary amine during the final separation and purification of the compound. The positively charged ion of the acceptable salt for treatment includes lithium, sodium, potassium, calcium, magnesium and aluminum, and nontoxic quaternary ammonium cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, N, N-dimethylaniline, N-methylpiperidine, N-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N, N-dibenzylphenethylamine, 1-diphenylhydroxymethylamine and N, N'-dibenzylethylenediamine. Other representative organic amines that can be used for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidines and piperazine.

[0216] While it is possible that the compounds of the subject disclosure are administered as raw chemicals, it is also possible to present them as pharmaceutical preparations. Thus, provided herein are pharmaceutical preparations comprising one or more of certain compounds disclosed herein, or one or more pharmaceutically acceptable salts, esters, prodrugs, amides or solvates thereof, together with one or more pharmaceutically acceptable carriers thereof and optionally one or more other therapeutic ingredients. The carrier(s) must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not harmful to the recipient thereof. Appropriate formulations depend on the chosen route of administration. Any of the well-known techniques, carriers and excipients may be used where appropriate and as understood in the art; for example, in Remington's Pharmaceutical Sciences. The pharmaceutical compositions disclosed herein can be manufactured in any manner known in the art, for example, by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, embedding or compression processes.

[0217] The formulations include those suitable for oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous, intraarticular and intramedullary), intraperitoneal, transmucosal, transdermal, rectal and topical (including cutaneous, buccal, sublingual and intraocular) administration, although the most suitable route may depend on, for example, the condition and disorder of the recipient. The formulations can be conveniently presented in unit dosage form and can be prepared by any method well known in the pharmaceutical art. Typically, these methods include the step of associating a compound of the present disclosure or a pharmaceutically acceptable salt, ester, amide, prodrug or solvate thereof ("active ingredient") with a carrier constituting one or more auxiliary ingredients. Typically, the formulations are prepared by uniformly and closely associating the active ingredient with a liquid carrier or a finely divided solid carrier or both, and then, if necessary, shaping the product into the desired formulation.

[0218] Formulations of the compounds disclosed herein suitable for oral administration can be presented as discrete units such as capsules, cachets, or tablets, each containing a predetermined amount of the active ingredient; a powder or granules; a solution or suspension in an aqueous or non-aqueous liquid; or an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient can also be presented as a bolus, electuary, or paste.

[0219] Pharmaceutical products that can be used orally include tablets, push-fit capsules made of gelatin, and soft sealed capsules made of gelatin and plasticizers such as glycerol or sorbitol. Tablets can be prepared by compression or molding together with one or more auxiliary ingredients. Compressed tablets can be prepared by compressing an active ingredient in a free-flowing form such as a powder or granule, optionally mixed with a binder, an inert diluent or lubricant, a surfactant or a dispersant, in a suitable machine. Molded tablets can be prepared by molding a mixture of powdered compounds moistened with an inert liquid diluent in a suitable machine. Tablets can be optionally coated or scored and can be formulated to provide a slow release or controlled release of the active ingredient therein. All preparations for oral administration should be in a dosage suitable for such administration. Push-fit capsules can contain an active ingredient mixed with a filler such as lactose, a binder such as starch, and / or a lubricant such as talc or magnesium stearate and optionally a stabilizer. In soft capsules, active compound can be dissolved or suspended in suitable liquid, such as fatty oil, liquid paraffin or liquid polyethylene glycol. In addition, stabilizing agent can be added. Dragee core is provided with suitable coating. For this purpose, concentrated sugar solution can be used, which can optionally contain gum arabic, talcum powder, polyvinyl pyrrolidone, carbomer gel, polyethylene glycol and / or titanium dioxide, lacquer solution (lacquer solution) and suitable organic solvent or solvent mixture. Dye or pigment can be added to tablet or dragee coating, for identification or in order to characterize the different combinations of active compound dosage.

[0220] Compound can be formulated for parenteral administration by injection, for example, by push injection or continuous infusion. The preparation for injection can be presented in unit dosage form, for example, in ampoule or in multidose container together with the preservative added. Composition can take the form of suspension, solution or emulsion such as in oily vehicle or aqueous vehicle, and can include formulation (formulatory agent), such as suspending agent, stabilizer and / or dispersant. Preparation can be presented in unit dose container or multidose container such as sealed ampoule and vial, and can be stored in powder form or in freeze drying (lyophilization) condition, only need to add sterile liquid carrier such as saline or sterile pyrogen-free water before about to use. Instant injection solution and suspension can be prepared by sterile powder, granules and tablet of the kind described previously.

[0221] Preparations for parenteral administration include aqueous and non-aqueous (oily) sterile injection solutions of the active compound, which may include antioxidants, buffers, antibacterials, and solutes that render the preparation is isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions, which may include suspending agents and thickening agents. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may include substances that increase the viscosity of the suspension, such as sodium carboxymethylcellulose, sorbitol, or dextran. Optionally, the suspension may also include suitable stabilizers or agents that increase the solubility of the compound to allow the preparation of highly concentrated solutions.

[0222] In addition to the preparations described previously, the compound can also be formulated as a reservoir product. Such a long-acting formulation can be administered by implantation (e.g., subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the compound can be formulated with a suitable polymeric material or hydrophobic material (e.g., as an emulsion in an acceptable oil) or an ion exchange resin, or be formulated as a slightly soluble derivative, e.g., as a slightly soluble salt.

[0223] For buccal or sublingual administration, the composition may take the form of tablets, lozenges, pastilles or gels formulated in conventional manner. Such compositions may comprise the active ingredient in a flavored basis such as sucrose and acacia or tragacanth.

[0224] The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, eg, containing conventional suppository bases such as cocoa butter, polyethylene glycol, or other glycerides.

[0225] Certain compounds disclosed herein can be administered topically, i.e., by non-systemic administration. This includes applying compounds disclosed herein externally to the epidermis or buccal cavity, as well as instilling such compounds into the ears, eyes, and nose so that the compound does not appreciably enter the bloodstream. In contrast, systemic administration refers to oral, intravenous, intraperitoneal, and intramuscular administration.

[0226] Preparations suitable for topical application include liquid products or semi-liquid products suitable for penetrating into the inflammation site through the skin, such as gels, liniments, lotions, creams, ointments or pastes, and drops suitable for application to eyes, ears or noses. The active ingredient for topical application can account for, for example, from 0.001% w / w to 10% w / w (by weight) of the preparation. In certain embodiments, the active ingredient can account for up to 10% w / w. In other embodiments, it can account for less than 5% w / w. In certain embodiments, the active ingredient can account for from 2% w / w to 5% w / w. In other embodiments, it can account for from 0.1% w / w to 1% w / w of the preparation.

[0227] In addition to the active ingredient, the topical eye preparations, ear preparations and nasal preparations of the present disclosure may include excipients. Excipients commonly used in such preparations include, but are not limited to, tonicity agents, preservatives, chelating agents, buffers and surfactants. Other excipients include solubilizers, stabilizers, comfort enhancers, polymers, emollients, pH regulators and / or lubricants. Any of a variety of excipients can be used for the preparations of the present disclosure, including water, mixtures of water and water-miscible solvents such as C1-C7 alkanols, vegetable oils or mineral oils containing from 0.5% to 5% non-toxic water-soluble polymers, natural products such as alginates, pectins, tragacanth gum, karaya gum, guar gum, xanthan gum, carrageenan, agar and gum arabic, starch derivatives such as starch acetate and hydroxypropyl starch, and other synthetic products such as polyvinyl alcohol, polyvinyl pyrrolidone, polyvinyl methyl ether, polyethylene oxide, preferably cross-linked polyacrylic acid and mixtures of those products. The concentration of the excipient is generally from 1 to 100,000 times the concentration of the active ingredient. In preferred embodiments, the excipients to be included in the formulation are generally selected based on their inertness toward the active ingredient components of the formulation.

[0228] With respect to ophthalmic, otic, and nasal preparations, suitable tonicity modifiers include, but are not limited to, mannitol, sodium chloride, glycerol, sorbitol, and the like. Suitable buffers include, but are not limited to, phosphates, borates, acetates, and the like. Suitable surfactants include, but are not limited to, ionic and nonionic surfactants (although nonionic surfactants are preferred), RLM 100, POE 20, cetearyl ether such as CS20 and poloxamer such as F68.

[0229] The formulations described herein may contain one or more preservatives. Examples of such preservatives include parabens, sodium perborate, sodium chlorite, alcohols such as chlorobutanol, benzyl alcohol or phenylethyl alcohol, guanidine derivatives such as polyhexamethylene biguanide, polyquaternium-1, amino alcohols such as AMP-95 or sorbic acid. In certain embodiments, the formulations may be self-preserving, such that no preservative is required.

[0230] For ophthalmic administration, ear administration or nasal administration, the formulation can be a solution, suspension or gel. In a preferred aspect, the formulation is used for topical application to the eye, nose or ear in the form of drops in an aqueous solution. The term "aqueous" generally refers to an aqueous formulation, wherein the formulation is by weight>50%, more preferably>75% and particularly>90% water. These drops can be delivered from single-dose ampoules, which can preferably be sterile and therefore render the antibacterial component of the formulation unnecessary. Alternatively, the drops can be delivered from multi-dose bottles, which can preferably contain a device for extracting any preservative from the formulation as the formulation is delivered, such devices being known in the art.

[0231] For ocular disorders, the components of the present disclosure may be delivered to the eye as a concentrated gel or similar vehicle, or as a dissolvable insert placed under the eyelid.

[0232] The preparations suitable for topical application to the eye of the present disclosure are preferably isotonic or slightly hypotonic to counteract any hypertonicity of the tears caused by evaporation and / or disease. This may require a tonicity agent to bring the osmolality of the preparation to a level at or near 210-320 milliosmoles / kilogram (mOsm / kg). The preparations of the present disclosure typically have an osmolality in the range of 220mOsm / kg-320mOsm / kg, and preferably have an osmolality in the range of 235mOsm / kg-300mOsm / kg. Eye preparations will typically be formulated as sterile aqueous solutions.

[0233] In certain ophthalmic embodiments, the compositions of the present disclosure are formulated with one or more tear substitutes. A variety of tear substitutes are known in the art and include, but are not limited to: monomeric polyols such as glycerol, propylene glycol, and ethylene glycol; polymeric polyols such as polyethylene glycol; cellulose esters such as hydroxypropyl methylcellulose, sodium carboxymethylcellulose, and hydroxypropyl cellulose; dextran such as dextran 70; vinyl polymers such as polyvinyl alcohol; and carbomers such as carbomer 934P, carbomer 941, carbomer 940, and carbomer 974P. Certain formulations of the present disclosure can be used with contact lenses or other ophthalmic products.

[0234] Preferred formulations are prepared using a buffer system that maintains the formulation at a pH of about 4.5 to a pH of about 8. The most preferred formulation pH is from 6 to 8.

[0235] In specific embodiments, the preparation of the present disclosure is used once a day. Yet, preparation can also be formulated for using with any frequency of administration, comprise once a week, every 5 days once, every 3 days once, every 2 days once, twice a day, three times a day, four times a day, five times a day, six times a day, eight times a day, per hour or any higher frequency.Depend on treatment regimen, such administration frequency also keeps continuing different persistent periods.The persistent period of specific treatment regimen can change from disposable administration to the scheme of continuing several months or several years.Preparation is used with different dosages, but typical dosage is one to two drops of using at each time, or a considerable amount of gel or other preparations.Those of ordinary skill in the art will be familiar with determining the treatment regimen for specific indications.

[0236] Gels for topical or transdermal administration can generally comprise a mixture of a volatile solvent, a non-volatile solvent, and water. In certain embodiments, the volatile solvent component of the buffered solvent system can include lower (C1-C6) alkyl alcohols, lower alkyl glycols, and lower glycol polymers. In another embodiment, the volatile solvent is ethanol. The volatile solvent component is considered to act as a penetration enhancer and also has a cooling effect on the skin as it evaporates. The non-volatile solvent portion of the buffered solvent system is selected from lower alkylene glycols and lower glycol polymers. In certain embodiments, propylene glycol is used. The non-volatile solvent slows down the evaporation of the volatile solvent and reduces the vapor pressure of the buffered solvent system. Like the volatile solvent, the amount of this non-volatile solvent component is determined by the pharmaceutical compound or drug used. When too little non-volatile solvent is in the system, the pharmaceutical compound may crystallize due to the evaporation of the volatile solvent, and excessive non-volatile solvent may lead to a lack of bioavailability due to poor release of the drug from the solvent mixture. The buffer component of the buffered solvent system can be selected from any buffer commonly used in the art; in certain embodiments, water is used. A common ratio of ingredients is about 20% non-volatile solvent, about 40% volatile solvent, and about 40% water. There are several optional ingredients that can be added to the topical composition. These include, but are not limited to, chelating agents and gelling agents. Suitable gelling agents can include, but are not limited to, semisynthetic cellulose derivatives (such as hydroxypropyl methylcellulose), synthetic polymers, galactomannan polymers (such as guar gum and its derivatives), and cosmetic agents.

[0237] Lotions include those suitable for application to the skin or eyes. Eyewashes may comprise a sterile aqueous solution optionally containing a bactericide and may be prepared by methods similar to those used to prepare drops. Lotions or liniments for application to the skin may also contain an agent to accelerate drying and cool the skin, such as alcohol or acetone, and / or a moisturizer such as glycerol or an oil such as castor oil or peanut oil.

[0238] Creams, ointments or pastes are semisolid preparations of active ingredients for external application. They can be prepared by mixing the active ingredient in a finely divided or powdered form, either alone or in a solution or suspension in an aqueous or non-aqueous fluid, with an oily or non-oily matrix using a suitable machine. The matrix may include hydrocarbons such as hard paraffin, soft paraffin or liquid paraffin, glycerol, beeswax, metallic soaps; mucus; oils of natural origin such as almond oil, corn oil, peanut oil, castor oil or olive oil; lanolin or its derivatives or fatty acids such as stearic acid or oleic acid and alcohols such as propylene glycol or macrogranular gel. The formulation may contain any suitable surfactant, such as anionic, cationic or nonionic surfactants, such as sorbitan esters or polyoxyethylene derivatives thereof. It may also contain suspending agents such as natural gums, cellulose derivatives or inorganic materials such as siliceous silica and other ingredients such as lanolin.

[0239] Drops can include sterile aqueous or oily solutions or suspensions, and can be prepared by dissolving the active ingredient in a suitable aqueous solution of a bactericide and / or fungicide and / or any other suitable preservative, and in certain embodiments, include a surfactant. The resulting solution can then be clarified by filtration, transferred to a suitable container, which is then sealed and sterilized by autoclaving or by keeping at 98°C-100°C for half an hour. Alternatively, the solution can be sterilized by filtration and transferred to the container by aseptic technique. Examples of bactericides and fungicides suitable for inclusion in drops are phenylmercuric nitrate or phenylmercuric acetate (0.002%), benzalkonium chloride (0.01%), and chlorhexidine acetate (0.01%). Suitable solvents for the preparation of oily solutions include glycerol, diluent alcohol, and propylene glycol.

[0240] Formulations for topical administration in the mouth, e.g., buccal or sublingual, include lozenges comprising the active ingredient in a flavored basis such as sucrose and acacia or tragacanth, and pastilles comprising the active ingredient in a basis such as gelatin and glycerin or sucrose and acacia.

[0241] For administration by inhalation, the compound can be conveniently delivered from an insufflator, a nebulizer pressurized pack, or other convenient device for delivering an aerosol spray. The pressurized pack can contain a suitable propellant, such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas. In the case of a pressurized aerosol, the dosage unit can be determined by providing a valve to deliver a metered amount. Alternatively, for administration by inhalation or insufflation, the compound according to the present disclosure can take the form of a dry powder composition, for example a powder mixture of the compound and a suitable powder base such as lactose or starch. The powder composition can be presented in unit dosage form, for example, in capsules, cartridges, gelatin, or blister packs, and the powder can be administered from the blister pack by means of an insufflator or insufflator.

[0242] Preferred unit dosage formulations are those containing an effective dose, as hereinbelow recited, or an appropriate fraction thereof, of the active ingredient.

[0243] It should be understood that in addition to the ingredients particularly mentioned above, the formulations described above may include other agents conventional in the art having regard to the type of formulation in question, for example those suitable for oral administration may include flavoring agents.

[0244] The compound can be taken orally or via injection at a dosage of 0.1 mg / kg to 500 mg / kg per day. The dosage range for adults is generally from 5 mg / day to 2 g / day. Tablets or other presentations provided in discrete units can conveniently contain an amount of one or more compounds that are effective at such dosages or multiples of such dosages, for example, units comprising 5 mg to 500 mg, typically about 10 mg to 200 mg.

[0245] The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration.

[0246] The compound can be administered in a variety of modes, such as orally, topically or by injection. The precise amount of compound administered to the patient will be the responsibility of the attending physician. The specific dosage level for any particular patient will depend on a variety of factors, including the activity of the specific compound employed, age, body weight, general health, sex, diet, time of administration, route of administration, excretion rate, drug combination, the exact disorder being treated, and the severity of the indication or condition being treated. In addition, the route of administration can vary depending on the condition and its severity.

[0247] In some cases, it may be appropriate to administer at least one of the compounds described herein (or a pharmaceutically acceptable salt, ester, or prodrug thereof) in combination with another therapeutic agent. By way of example only, if one of the side effects experienced by a patient after receiving one of the compounds described herein is hypertension, then it may be appropriate to administer an antihypertensive agent in combination with the initial therapeutic agent. Alternatively, by way of example only, the therapeutic effectiveness of one of the compounds described herein may be enhanced by administering an adjuvant (i.e., the adjuvant itself may have only minimal therapeutic benefit, but when combined with another therapeutic agent, the overall therapeutic benefit to the patient is enhanced). Alternatively, by way of example only, the benefit experienced by a patient may be increased by administering one of the compounds described herein together with another therapeutic agent (which also includes a treatment regimen) that also has therapeutic benefit. By way of example only, in the treatment of diabetes involving the administration of one of the compounds described herein, an increased therapeutic benefit may be obtained by also providing the patient with another diabetes therapeutic agent. In any case, regardless of the disease, disorder, or condition being treated, the overall benefit experienced by the patient may be simply the sum of the two therapeutic agents, or the patient may experience a synergistic benefit.

[0248] In any case, the multiple therapeutic agents (at least one of which is a compound disclosed herein) can be administered in any order or even simultaneously. If simultaneously, the multiple therapeutic agents can be provided in a single, unified form, or in multiple forms (by way of example only, as a single pill or as two separate pills). One of the therapeutic agents can be given in multiple doses, or both can be given as multiple doses. If not simultaneously, the time between multiple doses can be any duration ranging from a few minutes to four weeks.

[0249] The term "cancer" refers to a group of diseases characterized by abnormal and uncontrolled cell proliferation that begins in one site (primary site) with the potential to invade and spread to other sites (secondary sites, metastases), which distinguishes cancer (malignant tumors) from benign tumors. Almost all organs can be affected, resulting in more than 100 types of cancer that can affect humans. Cancer can be caused by many causes, including genetic predisposition, viral infection, exposure to ionizing radiation, exposure to environmental pollutants, smoking and / or drinking, obesity, poor diet, lack of physical activity, or any combination thereof.

[0250] Exemplary cancers described by the National Cancer Institute include: Acute Lymphoblastic Leukemia in Adults; Acute Lymphoblastic Leukemia in Children; Acute Myeloid Leukemia in Adults; Adrenocortical Carcinoma; Adrenocortical Carcinoma in Children; AIDS-Related Lymphoma; AIDS-Related Malignancies; Anal Cancer; Cerebellar Astrocytoma in Children; Cerebral Astrocytoma in Children; Extrahepatic Bile Duct Carcinoma; Bladder Cancer; Bladder Cancer in Children; Bone Cancer, Osteosarcoma / Malignant Fibrous Histiocytoma; Brain Stem Glioma in Children; Cerebellar Astrocytoma in Children; Cerebral Astrocytoma / Malignant Glioma in Children; Ependymoma Brain Tumor in Children; Medulloblastoma Brain Tumor in Children; Brain Tumor, Supratentorial Primitive Neuroectodermal Tumors, Childhood; Visual Pathway and Hypothalamic Glioma Brain Tumors in Children; Childhood (Other) Brain Tumors; Breast Cancer and Pregnancy Pregnancy); Childhood Breast Cancer; Male Breast Cancer; Childhood Bronchial Adenoma / Carcinoid Tumor; Childhood Carcinoid Tumor; Gastrointestinal Carcinoid Tumor; Adrenocortical Carcinoma; Islet Cell Carcinoma; Carcinoma of Unknown Primary; Primary Central Nervous System Lymphoma; Childhood Cerebellar Astrocytoma; Childhood Cerebral Astrocytoma / Glioma; Cervical Cancer; Childhood Cancer; Chronic Lymphocytic Leukemia; Chronic Myeloid Leukemia; Chronic Myeloproliferative Disorders; Clear Cell Sarcoma of the Tenosynovium; Colon Cancer; Childhood Colorectal Cancer; Cutaneous T-Cell Lymphoma; Endometrial Cancer Ependymoma in children; Ovarian epithelial carcinoma; Esophageal cancer; Esophageal cancer in children; Ewing family of tumors; Extracranial germ cell tumors in children; Extragonadal germ cell tumors; Extrahepatic bile duct cancer; Intraocular melanoma; Retinoblastoma; Gallbladder cancer; Gastric (stomach) cancer; Gastric (stomach) cancer in children; Gastrointestinal carcinoid tumors; Extracranial germ cell tumors in children; Extragonadal germ cell tumors; Ovarian germ cell tumors; Gestational trophoblastic tumors; Brainstem gliomas in children; Visual pathway and hypothalamic gliomas in children Visual Pathway and Hypothalamic; Hairy Cell Leukemia; Head and Neck Cancer; Hepatocellular Carcinoma (Primary) (Adult); Hepatocellular Carcinoma (Primary) (Childhood); Hodgkin's Lymphoma (Adult); Hodgkin's Lymphoma (Childhood); Hodgkin's Lymphoma (During Pregnancy); Hypopharyngeal Cancer; Hypothalamic and Visual Pathway Glioma (Childhood);Intraocular melanoma; Islet cell carcinoma (endocrine pancreas); Kaposi sarcoma; Kidney cancer; Laryngeal cancer; Laryngeal cancer (childhood); Leukemia, Acute Lymphoblastic (Adult); Leukemia, Acute Lymphoblastic (Childhood); Acute Myeloid Leukemia (Adult); Acute Myeloid Leukemia (Childhood); Chronic Lymphocytic Leukemia; Chronic Myeloid Leukemia; Hairy cell leukemia; Lip and oral cavity cancer; Liver cancer (primary) in adults; Liver cancer (primary) in children; Non-small cell lung cancer; Small cell lung cancer; Lymphoblastic Leukemia (Adult); Lymphoblastic Leukemia (Childhood) Acute; chronic lymphocytic leukemia; AIDS-related lymphoma; central nervous system (primary) lymphoma; cutaneous T-cell lymphoma; adult Hodgkin's lymphoma; childhood Hodgkin's lymphoma; Hodgkin's lymphoma during pregnancy Pregnancy; Non-Hodgkin's Lymphoma (Adult); Non-Hodgkin's Lymphoma (Child); Non-Hodgkin's Lymphoma (Child); Pregnancy; Primary Central Nervous System Lymphoma; Waldenstrom's Macroglobulinemia; Male Breast Cancer; Malignant Mesothelioma (Adult); Malignant Mesothelioma (Child); Malignant Thymoma; Medulloblastoma (Child); Melanoma; Intraocular Melanoma; Merkel Cell Carcinoma; Malignant Mesothelioma; Occult Primary Metastatic Squamous Neck Cancer; Multiple Endocrine Neoplasia Syndrome (Child); Multiple Myeloma / Plasma Cell Neoplasm; Mycosis Fungoides; Myelodysplasia Syndrome; Chronic Myeloid leukemia; childhood acute myeloid leukemia; multiple myeloma; chronic myeloproliferative disorder; nasal and paranasal sinus cancer; nasopharyngeal cancer; childhood nasopharyngeal cancer; neuroblastoma; adult non-Hodgkin's lymphoma; childhood non-Hodgkin's lymphoma; pregnancy non-Hodgkin's lymphoma; non-small cell lung cancer; childhood oral cancer; oral cavity and lip cancer; oropharyngeal cancer; osteosarcoma / malignant fibrous histiocytoma of the bone; childhood ovarian cancer; ovarian epithelial cancer; ovarian germ cell tumor; ovarian low malignant potential tumor; pancreatic cancer; childhood pancreatic cancer; islet cell pancreatic cancer; paranasal sinus and nasal cavity cancer; parathyroid cancer; penile cancer; pheochromocytoma; childhood pineal and supratentorial primitive neuroectodermal tumors; pituitary tumors; plasma cell neoplasms / multiple myeloma; pleuropulmonary blastoma; pregnancy complicated by breast cancer; pregnancy complicated by Hodgkin's lymphoma; pregnancy complicated by non-Hodgkin's lymphoma;Primary central nervous system lymphoma; Primary liver cancer in adults; Primary liver cancer in children; Prostate cancer; Rectal cancer; Renal cell (kidney) cancer; Renal cell carcinoma in children; Transitional cell carcinoma of the renal pelvis and ureter; Retinoblastoma; Rhabdomyosarcoma in children; Salivary gland cancer; Salivary gland cancer in children; Ewing family of tumors; Kaposi's sarcoma; Osteosarcoma / malignant fibrous histiocytoma sarcoma of the skeleton; Rhabdomyosarcoma in children; Soft tissue sarcoma in adults; Soft tissue sarcoma in children; Sézary syndrome; Skin cancer; Skin cancer in children; Skin cancer (melanoma); Merkel cell skin cancer; Small cell lung cancer; Small bowel cancer; Adults Soft tissue sarcoma; childhood soft tissue sarcoma; occult primary metastatic squamous neck cancer; stomach (gastric) cancer; childhood stomach (gastric) cancer; childhood supratentorial primitive neuroectodermal tumor; cutaneous T-cell lymphoma; testicular cancer; childhood thymoma; malignant thymoma; thyroid cancer; childhood thyroid cancer; transitional cell carcinoma of the renal pelvis and ureter; gestational trophoblastic tumor; childhood cancer of unknown primary site; rare childhood cancers; transitional cell carcinoma of the ureter and renal pelvis; urethral cancer; uterine sarcoma; vaginal cancer; pediatric visual pathway and hypothalamic gliomas; vulvar cancer; Waldenstrom's macroglobulinemia; and Wilms' tumor.

[0251] In certain aspects, the cancer includes lung cancer, breast cancer, colorectal cancer, prostate cancer, stomach cancer, liver cancer, cervical cancer, esophageal cancer, bladder cancer, non-Hodgkin lymphoma, leukemia, pancreatic cancer, kidney cancer, endometrial cancer, head and neck cancer, lip cancer, oral cancer, thyroid cancer, brain cancer, ovarian cancer, melanoma, gallbladder cancer, laryngeal cancer, multiple myeloma, nasopharyngeal cancer, Hodgkin lymphoma, testicular cancer, and Kaposi's sarcoma.

[0252] In other aspects, the method further comprises administering a chemotherapeutic agent. The compounds of the present disclosure can be administered in combination with one or more additional therapeutic agents. The phrases "combination therapy," "in combination with," and the like refer to the simultaneous use of more than one drug or treatment to increase response. The compounds of the present disclosure can be used, for example, in combination with other drugs or treatments for the treatment of cancer. In various aspects, the compounds are administered before, simultaneously with, or after the administration of the chemotherapeutic agent.

[0253] The term "anti-cancer therapy" refers to any therapy or treatment that can be used to treat cancer. Anti-cancer therapies include, but are not limited to, surgery, radiation therapy, chemotherapy, immunotherapy, and targeted therapy.

[0254] Examples of chemotherapeutic or anticancer agents include, but are not limited to, dactinomycin, azacitidine, azathioprine, bleomycin, bortezomib, carboplatin, capecitabine, cisplatin, chlorambucil, cyclophosphamide, cytarabine, daunorubicin, docetaxel, doxifluridine, doxorubicin, epirubicin, epothilone, etoposide, fluorouracil, gemcitabine, hydroxyurea, idarubicin, imatinib, irinotecan, nitrogen mustard, mercaptopurine, methotrexate, mitoxantrone, oxaliplatin, paclitaxel, pemetrexed, teniposide, thioguanine, topotecan, valrubicin, vinblastine, vincristine, vindesine, vinorelbine, panitumumab, erbitux (cetuximab), matuzumab, IMC-IIF 8, TheraCIM hR3, denosumab, Avastin (bevacizumab), Humira (adalimumab), Herceptin (trastuzumab), Remicade (infliximab), Rituximab, Synagis (palivizumab), Mylotarg (gemtuzumab ozogamicin), Raptiva (efalizumab), Tysabri (natalizumab), Zenapax (daclizumab), NeutroSpec (technetium (99mTc) fanolesomab), tocilizumab, ProstaScint (indium-Ill-labeled card (Rosumab pendetide), Bexxar (tositumomab), Zevalin (ibritumomab tiuxetan conjugated to yttrium-90 (IDEC-Y2B8)), Xolair (omalizumab), MabThera (rituximab), ReoPro (abciximab), MabCampath (alemtuzumab), Simulect (basiliximab), LeukoScan (thiosomab), CEA-Scan (acitumomab), Verluma (nofetumomab), Panorex (edrecolomab), alemtuzumab, CDP 870, natalizumab, Gilotrif (afatinib), Lynparza (olaparib), Perjeta (pertuzumab), Otdivo (nivolumab), Bosulif (bosutinib), Cabometyx (cabozantinib), Ogivri (trastuzumab-DKST), Sutent (sunitinib malate), Adcetris (brentuximab), Alecensa (alecin), Calquence (acalabrutinib), Yescarta (ciloleucel), Verzenio (pomacicillin),Keytruda (pembrolizumab), Aliqopa (copanlisib), Nerlynx (neratinib), Imfinzi (durvalumab), Darzalex (daratumumab), Tecentriq (atezolizumab), and Tarceva (erlotinib). Examples of immunotherapeutic agents include, but are not limited to, interleukins (Il-2, Il-7, Il-12), cytokines (interferon, G-CSF, imiquimod), chemokines (CCL3, CCl26, CXCL7), and immunomodulatory imide drugs (thalidomide and its analogs).

[0255] In the treatment of, the dosage of agent is optionally in the range of from about 0.0001mg / kg to about 100mg / kg, about 0.01mg / kg to about 5mg / kg, about 0.15mg / kg to about 3mg / kg, 0.5mg / kg to about 2mg / kg and about 1mg / kg to about 2mg / kg of the body weight of experimenter.In other embodiments, dosage is in the range of from about 100mg / kg to about 5g / kg, about 500mg / kg to about 2mg / kg and about 750mg / kg to about 1.5g / kg of the body weight of experimenter.For example, depending on the type and the order of severity of the disease, the agent of about 1 μ g / kg to 15mg / kg (for example, 0.1mg / kg-20mg / kg) is for being applied to the candidate dosage of patient, no matter whether it is for example by one or more times being used separately or by continuous infusion.Depending on the factors mentioned above, typical daily dose is in the range of from about 1 μ g / kg to 100mg / kg or more. For repeated administration over several days or longer, depending on the situation, continued treatment occurs until desired disease symptoms are suppressed. However, other dosage regimens may also be useful. The unit dose may be, for example, in the range of about 5 mg to 500 mg, such as 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, and 300 mg. The progress of therapy is monitored by conventional techniques and assays.

[0256] In some embodiments, the agent is administered to a human patient in an effective amount (or dosage) of less than about 1 μg / kg, for example, about 0.35 μg / kg to about 0.75 μg / kg or about 0.40 μg / kg to about 0.60 μg / kg. In some embodiments, the dosage of the agent is about 0.35 μg / kg, or about 0.40 μg / kg, or about 0.45 μg / kg, or about 0.50 μg / kg, or about 0.55 μg / kg, or about 0.60 μg / kg, or about 0.65 μg / kg, or about 0.70 μg / kg, or about 0.75 μg / kg, or about 0.80 μg / kg, or about 0.85 μg / kg, or about 0.90 μg / kg, or about 0.95 μg / kg, or about 1 μg / kg. In various embodiments, the absolute dose of an agent is about 2 μg / subject to about 45 μg / subject, or about 5 μg / subject to about 40 μg / subject, or about 10 μg / subject to about 30 μg / subject, or about 15 μg / subject to about 25 μg / subject. In some embodiments, the absolute dose of an agent is about 20 μg, or about 30 μg, or about 40 μg.

[0257] In various embodiments, the dosage of an agent can be determined by the weight of the human patient. For example, for a pediatric human patient of about 0 kg to about 5 kg (e.g., about 0 kg, or about 1 kg, or about 2 kg, or about 3 kg, or about 4 kg, or about 5 kg), the absolute dosage of the agent is about 2 μg; or for a pediatric human patient of about 6 kg to about 8 kg (e.g., about 6 kg, or about 7 kg, or about 8 kg), about 3 μg; or for a pediatric human patient of about 9 kg to about 13 kg (e.g., 9 kg, or about 10 kg, or about 11 kg, or about 12 kg, or about 13 kg), about 5 μg; or for a pediatric human patient of about 14 kg to about 20 kg (e.g., about 14 kg, or about 16 kg, or about 18 kg, or about 20 kg), about 8 μg; or for a pediatric human patient of about 21 kg to about 30 kg (e.g., about 21 kg, or about 23 kg, or about 25 kg, or about 27 kg, or about 30 kg). or about 12 μg for a pediatric human patient of about 31 kg to about 33 kg (e.g., about 31 kg, or about 32 kg, or about 33 kg); or about 20 μg for an adult human patient of about 34 kg to about 50 kg (e.g., about 34 kg, or about 36 kg, or about 38 kg, or about 40 kg, or about 42 kg, or about 44 kg, or about 46 kg, or about 48 kg, or about 50 kg); or about 30 μg for an adult human patient of about 51 kg to about 75 kg (e.g., about 51 kg, or about 55 kg, or about 60 kg, or about 65 kg, or about 70 kg, or about 75 kg); or about 45 μg for an adult human patient greater than about 114 kg (e.g., about 114 kg, or about 120 kg, or about 130 kg, or about 140 kg, or about 150 kg).

[0258] In addition to those compounds presented in the Examples, the following compounds further illustrate the scope of the present disclosure:

[0259] Table 1. Summary of compounds synthesized and tested in this disclosure.

[0260]

[0261]

[0262]

[0263]

[0264]

[0265]

[0266]

[0267]

[0268]

[0269]

[0270]

[0271]

[0272]

[0273]

[0274]

[0275]

[0276]

[0277]

[0278]

[0279]

[0280]

[0281]

[0282]

[0283]

[0284]

[0285] Table 2. HUVEC antiproliferative activity and CYP3A4 enzyme inhibition of itraconazole analogs.

[0286]

[0287]

[0288]

[0289]

[0290] a IC of HUVECs 50 use[ 3H]-thymidine incorporation assay was performed for evaluation. b CYP3A4 enzyme inhibition use CYP3A4 Green Screening Assay was used for evaluation. Values are % enzyme inhibition at 1 μM of the indicated drug / % enzyme inhibition at 1 μM of itraconazole. Values represent the mean of at least two separate experiments performed in triplicate.

[0291] SAR studies of new itraconazole analogs were performed using HUVEC proliferation and CYP3A4 enzymatic assays.

[0292] The antiangiogenic activity of novel itraconazole analogs was determined using a HUVEC proliferation assay [ 3 [H] Thymidine incorporation was used as a readout and CYP3A4 inhibition was initially determined using a cell-free fluorescence-based assay at a final concentration of 1 μM (Table 2).

[0293] To evaluate the importance of the triazole moiety for the antiangiogenic activity and CYP3A4 inhibition of itraconazole, we removed the 1,2,4-triazole (compound 62) and replaced it with an aliphatic ethyl (compound 59), bromomethyl (compound 57), cyclopentyl (compound 61) group, or phenyl (compound 60) group. As expected, in the initial screening, compound 57 and compounds 59-62, which do not have a nitrogen atom in the R1 position, did not exhibit CYP3A4 inhibition at 1 μM. Further dose-response assays showed that compound 59 completely lost CYP3A4 inhibition ( Figure 2 ). In contrast, analogs from the same group suffered only a 2-3 fold reduction in antiproliferative activity against HUVEC, indicating a differential dependence of these two activities on the presence of the triazole moiety.

[0294] Next, tertiary amines were introduced at the R1 position using dimethylamine, morpholine, piperidinyl, or N-methylpiperidinyl groups (Compounds 63–66). Compared to the parent itraconazole, analogs 63–66 possessed the additional characteristic of improved solubility. With the exception of Compound 64, analogs containing aliphatic amines at the R1 position inhibited CYP3A4 at 1 μM. Structurally, Compounds 63, 65, and 66 possess electron-donating nitrogen or oxygen atoms at the position corresponding to the N4 of the 1,2,4-triazole in itraconazole. The results suggest that aliphatic nitrogen and oxygen atoms are also capable of interacting with the heme group in CYP3A4. Compared to itraconazole, Compounds 63–66 exhibited a 2- to 9-fold decrease in antiproliferative activity in HUVEC. Overall, the results confirm that itraconazole's CYP3A4 inhibition is highly dependent on the 1,2,4-triazole, particularly the basic N4 atom of the triazole. They also showed that replacement of the triazole moiety with alkylamines or aliphatic amines was not sufficient to reduce CYP3A4 inhibition without compromising antiangiogenic activity.

[0295] Given the importance of the triazole moiety in the anti-angiogenic activity of itraconazole, we decided to make less drastic structural changes to the triazole by increasing the steric hindrance around the nitrogen atom at the 4 position, which might reduce its access to the heme iron in CYP3A4. As a start, we introduced methyl or trifluoromethyl substitutions into 1,2,4-triazole. The analog compound 50 with 3-methyl-1H-1,2,4-triazole substitution and compound 51 with dimethyl substitution showed reduced potency in terms of CYP3A4 inhibition. The trifluoromethyl substituted compound 47 showed no CYP3A4 inhibition at 1 μM and weak inhibition at higher concentrations ( Figure 2). The IC50 values of compound 50, compound 47, and compound 51 for inhibiting HUVEC proliferation were 0.27 μM, 0.22 μM, and 0.27 μM, respectively, which are slightly higher than the IC50 value of itraconazole (0.17 μM). Compound 33 contains an imidazole moiety instead of 1,2,4-triazole and exhibits a higher efficacy against HUVEC proliferation than itraconazole. We synthesized a series of analogs containing imidazole groups with various substitutions (compounds 35, 43, 45, and 46). Similar to compound 51, compound 39 with a 2-methyl-4-trifluoromethyl-1H-imidazolyl moiety has both steric hindrance and electron-withdrawing effects, and in addition, the N3 nitrogen no longer reacts with heme, so CYP3A4 inhibition is eliminated at 1 μM. However, the disubstituted compounds (compounds 39-compound 41) also suffered a 2-3-fold reduction in anti-angiogenic activity. Interestingly, among the monosubstituted imidazolyl compounds, the 2-isopropyl-1H-imidazolyl analog, compound 45, exhibited the most potent HUVEC inhibitory activity, with an IC50 of 0.084 μM. Compounds with substituents smaller or larger than the isopropyl group were less potent than compound 45. CYP3A4 inhibition, on the other hand, showed a different trend—the larger the substituent on the imidazole, the less CYP3A4 inhibition.

[0296] In contrast to triazole and imidazole, which are basic aromatic rings, tetrazoles are relatively acidic due to the addition of another nitrogen to the 5-membered ring. Because itraconazole's inhibitory activity against CYP3A4 requires coordination of the basic nitrogen to the heme iron, we reasoned that tetrazoles should have reduced CYP3A4 inhibition compared to imidazole or triazole compounds. We synthesized the 1-tetrazolyl analog compound 48; it showed improved antiproliferative activity in HUVECs, with an IC50 of 0.073 μM, and significantly reduced CYP3A4 inhibition, with an IC50 greater than 20 μM ( Figure 2 In contrast, the structurally related 2-tetrazolyl analog, compound 49, was less potent in HUVEC and exhibited greater CYP3A4 inhibition than compound 48. Addition of a 5-methyl or 5-phenyl substituent to the tetrazole group did not result in further improvement of analog compound 48, making compound 48 the best of all itraconazole analogs. Therefore, we selected compound 48 for further biological evaluation.

[0297] Inhibition of tube formation. To further evaluate the anti-angiogenic activity of compound 48, we employed an in vitro tube formation assay. In the tube formation assay, HUVECs were seeded on Matrigel and after 20 hours, the cells migrated and elongated to form a tube-like network, which is reminiscent of neovascularization. Figure 3As shown in , treatment of HUVECs with 5 μM itraconazole inhibited 45% of HUVEC tube formation, as judged by total tube length. At the same concentration, compound 48 inhibited 61% of HUVEC tube formation, further demonstrating that compound 48 is a more potent anti-angiogenesis inhibitor.

[0298] Niemann-Pick C phenotype and mTOR inhibition. We have previously shown that the mechanism underlying the antiangiogenic activity of itraconazole is mediated in part by inhibition of endolysosomal cholesterol transport and mTOR inhibition. Therefore, we determined whether compound 48 shares the same mechanism as itraconazole. Intracellular cholesterol was detected using the cholesterol-binding fluorescent dye filipin. Figure 4 As shown in , similar to itraconazole, compound 48 induced an NPC phenotype at a concentration of 0.2 μM, as judged by the accumulation of cholesterol in late endosomes / lysosomes. Analog compound 48 also increased the phosphorylation of AMPK at Thr172 in a dose-dependent manner ( Figure 5 ). Phosphorylation of the AMPK substrate acetyl-CoA carboxylase 1 (ACC1) was also increased as expected. AMPK activation is known to lead to mTOR inhibition. Treatment of HUVECs with compound 48 indeed resulted in a decrease in the phosphorylation of mTOR and its downstream effector p70 S6 kinase (S6K). Taken together, these results suggest that compound 48 inhibits HUVEC proliferation and angiogenesis by the same target and underlying mechanism as itraconazole.

[0299] CYP3A4 inhibition limits the use of itraconazole as an anticancer agent due to metabolic interactions with other anticancer drugs. To overcome this limitation, we have synthesized a series of azole and nonazole derivatives of itraconazole and evaluated their antiangiogenic activity and CYP3A4 inhibition. Based on our SAR studies, we conclude that itraconazole inhibition of CYP3A4 is highly dependent on the coordination of the 1,2,4-triazole to the heme iron and that modification of the triazole can reduce or eliminate CYP3A4 inhibition. Nonazole analogs successfully abolished CYP3A4 inhibition by disrupting heme binding, but unfortunately, the compounds were significantly less potent than itraconazole against HUVEC. Substituted azoles with sterically hindered and electron-withdrawing groups also abolished CYP3A4 inhibition, as seen in analogs 51 and 39. The 1H-tetrazolyl analog 48 showed an EC50 above 20 μM against CYP3A4. More importantly, it also exhibits more potent anti-angiogenic activity than itraconazole. The anti-angiogenic potency of compound 48 was further confirmed by HUVEC tube formation. Like itraconazole, compound 48 acts by the same mechanism as itraconazole through NPC1 phenotype induction and mTOR inhibition. In summary, our results strongly suggest that the tetrazole-containing analog compound 48 is a promising new lead for the development of next-generation itraconazole analogs with improved anti-angiogenic potency and minimal CYP3A4 inhibition, which can be used in combination with most other known anticancer drugs for the treatment of a wide variety of cancers.

[0300]

[0301] Scheme 1. Synthetic procedures of Compounds 57-62.

[0302] The synthetic route for non-azole itraconazole analogs (Compounds 57–62) is outlined in Scheme 1. The synthesis begins with commercially available 1,3-dichlorobenzene and a series of different acid chlorides. Intermediates 3(af), 5(af), and 6(af) can vary depending on the product structure. The intermediate 2,4-dichlorobenzaldehyde (Intermediate 3a) is commercially available. The other intermediates, Intermediates 3b–3f, were efficiently prepared in satisfactory yields (70%–90%) by acylation of 1,3-dichlorobenzene with a series of acid chlorides (Intermediates 2b–2f) under Friedel-Craft acylation conditions. Our previous results indicate that the 2S,4R-cis stereochemistry on the 1,3-dioxolane ring is more potent in antiangiogenic activity than alternative stereochemical configurations. Therefore, we construct cis-1,3-dioxolane (intermediates 5a-intermediates 5f) via acid-assisted ketalization of 2,4-dichloroacetophenone (intermediates 3a-intermediates 3f) with optically pure glycerol tosylate in the presence of methanesulfonic acid (TfOH) in toluene, producing cis-diastereomers and trans-diastereomers in good yields in a 3:1 ratio. Those two diastereomers are easily separated by column chromatography. The obtained phenol fragment 7a is synthesized, followed by a previously reported synthetic route. See U.S. Patent No. 9,346,791. Phenol fragment intermediate 7a is subjected to o-toluenesulfonate in the presence of NaH to provide the final product (compound 57-compound 62). Analogs with tertiary amine groups (compound 63-compound 66) are prepared by nucleophilic substitution reactions of bromo-substituted compound 57 and different secondary amines (Scheme 2).

[0303]

[0304] Scheme 2. Synthetic procedures of Compound 63-Compound 66.

[0305]

[0306] Scheme 3. Synthetic procedures of Compound 1-Compound 23, Compound 33-Compound 53, and Compound 67-Compound 90.

[0307] The synthetic routes of itraconazole analogs containing substituted azoles (Compound 1-Compound 23, Compound 33-Compound 53 and Compound 67-Compound 90) are outlined in Scheme 3. Depending on the structure of the product, intermediate 9, intermediate 10 and intermediate 11 may be different. The itraconazole analogs were synthesized using a similar route to that outlined in Scheme 1. Intermediate (9) was prepared by N-alkylation of 2-bromo-1-(2,4-dichlorophenyl)ethanone (Intermediate 8) with a series of azoles in DCM at room temperature. Ketalization of the acetone intermediate 9 with glycerol tosylate (Intermediate 4) gave intermediates as cis diastereomers, intermediate 10, in low to moderate yields (10%-65%). Finally, the o-toluenesulfonate intermediate 108 was treated with fragment intermediate 7b to give the desired product. The syntheses of Compound 24-Compound 32, Compound 54-Compound 56, and Compound 91-Compound 98 can be found in US Pat. No. 9,346,791.

[0308] Despite its potential as a novel anti-angiogenic drug, it has three major limitations. First, inhibition of cytochrome P450 3A4 (CYP3A4) prevents itraconazole from being used in combination therapy with most other anticancer drugs. Second, itraconazole's high lipophilicity (logP = 5.3) leads to its accumulation in adipose tissue, with concentrations in skin and adipose tissue 19-fold and 17-fold greater than those in plasma, respectively. Third, itraconazole is relatively insoluble in water and has limited oral bioavailability. Although clinically used suspension formulations can improve its bioavailability, itraconazole has highly variable absorption, and plasma concentrations are often difficult to predict between individuals. The high lipophilicity and low solubility of itraconazole, although tolerable for the treatment of fungal infections, pose major problems in the treatment of cancer.

[0309]

[0310] Scheme 4. Structures of itraconazole, IT-C (Compound 99) and Compound 24.

[0311] In an attempt to improve the efficacy and reduce the toxicity of itraconazole, a systematic structure-activity relationship (SAR) study was conducted on both ends of itraconazole. Compound IT-C (compound 99), which has cis-(2S,4R,2'S) stereochemistry in the dioxolane ring, in which the isobutyl side chain has a 2'S configuration, showed the strongest antiangiogenic activity among the eight stereoisomers of itraconazole and significantly reduced hepatotoxicity. Compound 24, which replaces 1,2,4-triazole with a tetrazole in the head position, has increased activity and significantly reduced CYP3A4 inhibition, while methyl substitution in the same position results in a loss of antiangiogenic activity. Furthermore, a sec-butyl tail (or similar alkyl group) is required for angiogenesis inhibition.

[0312] Although modifications at the "head" and "tail" portions of itraconazole effectively reduce CYP3A4 inhibition and hepatotoxicity, hydrophobicity and the accompanying low water solubility remain to be addressed. The hydrophobicity of itraconazole can be largely attributed to the two phenyl groups (W1-piperazin-1-yl-W2, Scheme 4) in the "core" region of the molecule. The phenyl-piperazin-1-yl-phenyl core portion has a symmetrical and rigid configuration. Symmetrical and rigid compounds are reported to have high crystal packing energies and, therefore, low solubility in water and in organic solvents. In an effort to further improve the properties of itraconazole analog compounds, we have used pyridyl to replace the phenyl group or added fluorine substitutions to the phenyl ring. The synthesis and anti-angiogenic activity characteristics of the novel analogs are discussed below.

[0313]

[0314]

[0315] Reagents and conditions: (a) azoles, K2CO3, CAN, rt, 16h, 40%-70%; (b) TfOH, toluene, rt, 60h, 55%; (c) Pd2(dba)3, BINAP, NaOtBu, toluene, 80℃, 16h, 80%; (d) 10% Pd / C, N2H4.H2O, EtOH, reflux, 3.5h, quantitative; (e) phenyl chloroformate, pyridine Pyridine, rt, 16h, 82%; (f) i. NH2NH2.H2O, 1,4-dioxane, reflux, 3h; ii. Formamidine acetate, 1-propanol, reflux, 3h, 66%; (g) 2-bromobutane, K2CO3, DMSO, 80℃, 16h, 82%; (h) 48% aqueous HBr, 110℃, 87%; (i) NaH, DMF, 80℃, 16h, 60-75%.

[0316] Scheme 5. Synthetic procedures for additional compounds of the disclosure.

[0317]

[0318] Scheme 6. Synthetic procedures of Compound 6, Compound 7, Compound 102 and Compound 103.

[0319] The synthesis of additional novel itraconazole analogs was accomplished using modified synthetic routes shown in Schemes 5 and 6. Nucleophilic displacement of the chlorine in intermediate 12 with 1,2,4-triazole, imidazole, or tetrazole, followed by ketal ring closure with enantiomerically pure glycerol tosylate intermediate 4 under strongly acidic conditions, provided the 1,3-dioxolane intermediate as a 3:1 mixture of cis and trans forms. The cis-2S,4R diastereoisomer, intermediate 14, was separated by normal phase column chromatography. Another portion of the analogs was synthesized by two different routes, determined by the commercial availability of the starting materials.

[0320] For analogs where the W1 group is pyridyl, Buchwald-Hartwig cross coupling of bromide intermediate 15 with intermediate 16 is used to generate nitrobenzene intermediate 17. The nitro group is then reduced to the amine using 10% Pd / C in ethanol and hydrazine monohydrate. The triazolone ring is constructed in three steps by sequentially reacting the aniline intermediate with phenyl chloroformate, hydrazine monohydrate, and formamidine acetate. N-alkylation of triazolone intermediate 20 with 2-bromobutane under basic conditions provides intermediate 21. The methyl protecting group in intermediate 21 is demasked using 48% aqueous hydrobromic acid (HBr) at 110°C. The resulting phenol intermediate 22 was coupled with the tosylate intermediate 14 to give the desired Compound 1, Compound 2, Compound 4, Compound 5, Compound 8, Compound 14, Compound 15, Compound 100, Compound 101, Compound 104, Compound 105 and Compound 106.

[0321] For analogs where the W1 ring is phenyl and W2 is pyridyl, nitro intermediate 25 was constructed via a CN cross-coupling reaction of 2-bromo-5-nitropyridine (intermediate 24a) or 5-bromo-2-nitropyridine (intermediate 24b) with piperazine intermediate 23, using Pd2(dba)3 as a catalyst, 2,2′-bis(diphenylphosphino)-1,1′-binaphthyl (BINAP) as a ligand, and sodium tert-butoxide as a base. The resulting nitro-containing intermediate intermediate 25 was subjected to the same six-step transformation as intermediate 22 to produce the final compounds 6, 7, 102, and 103.

[0322]

[0323] Reagents and conditions: (a) 1 H-tetrazole, K2CO3, CAN, rt, 16h; (b) TfOH, toluene, rt, 60h; (c) Pd2(dba)3, BINAP, NaOtBu, toluene, 80℃, 16h; (d) 10% Pd / C, NH2NH2, EtOH, reflux, 3.5h; (e) phenyl chloroformate, pyridine, rt, 16h; (f) 1.NH2NH2.H2O, 1,4-dioxane, reflux, 3h; 2. formamidine acetate, 1-propanol, reflux, 3h; (g) K2CO3, 18-crown-6, DMSO, room temperature; (h) 48% aqueous HBr, 110℃; (i) NaH, DMF, 80℃, 16h.

[0324] Scheme 7. Synthetic procedures of Compound 130 and Compound 131.

[0325] Scheme 7 shows the synthesis of pure diastereomers such as compounds 130 and 131 on the isobutyl side chain. The synthesis of tetrazole intermediate 14c follows the procedure described in the previous section. The stereocenter on the isobutyl side chain can be introduced through intermediates 26a and 26b.

[0326] Table 3. Inhibition of HUVEC proliferation, aqueous solubility, and logP of disclosed itraconazole analog compounds.

[0327]

[0328]

[0329] a IC of HUVECs 50 use[ 3 [H]-thymidine incorporation was assessed. Values represent the mean ± SD of three independent experiments performed in triplicate. b logP was predicted using ALOGPS2.1 software. c Thermodynamic solubility in 0.001 N HCl was measured using HPLC.

[0330] use[ 3[H]-thymidine incorporation assay measures the inhibitory effect of novel analogs on HUVEC proliferation. ALOGPS2.1 software was used to calculate logP values, which serve as an indicator of lipophilicity. Aqueous solubility is a key physicochemical property associated with oral absorption. We determined the solubility of each analog in 0.001N HCl (pH = 3), which represents acidic human gastric fluid. The IC50, logP, and solubility of all analogs against HUVEC proliferation are shown in Table 3.

[0331] Itraconazole is slightly soluble in 0.001N HCl (10.1ng / mL). In order to destroy the symmetry of the phenyl-piperazine-1-yl-phenyl core (W1-piperazine-1-yl-W2), we use pyridyl to replace one of the phenyl rings in the W1 or W2 position, or use 3 '-fluorophenyl to replace W2. Both pyridine and fluorobenzene are reasonable benzene isosteres and are not expected to cause significant steric perturbations in the W1-W2 portion of the molecule. Initially, compounds 100-106 with 1,2,4-triazole in the R1 position were synthesized and characterized. Pyridyl substitution at the W1 (compound 100 and compound 101) or W2 (compound 102 and compound 103) position and 3 '-fluoro-phenyl at the W2 (compound 104-compound 106) position all resulted in a 5-84-fold increase in solubility. The combination of pyridin-2-yl and 3'-fluorophenyl (compound 106) resulted in the best solubility (847.7 ng / mL) among the analogs in this series. Pyridine (=N), benzene (=CH), and fluorine-substituted benzene (=CF) are similar in van der Waals radius, but very different in lipophilicity. The lipophilicity of the three groups follows the following order: pyridinyl < benzyl < fluorobenzyl. As expected, the analogs containing pyridine (compounds 100-compound 103) showed a decrease in logP, while the fluorophenyl analog (compound 104) had an increase in logP. In the HUVEC proliferation assay, compounds 103, 104, and 106 showed an activity similar to that of itraconazole, while other modifications resulted in a decrease in efficacy.

[0332] Next, six analogs with imidazole in the R1 position were synthesized and characterized. With the exception of compound 2 (pyridin-3-yl) and compound 7 (pyridin-2'-yl), the other four analogs (compound 1, compound 4, compound 5, and compound 6) resulted in increased activity for inhibiting HUVEC proliferation. In general, the change from triazole to imidazole resulted in a significant increase in solubility in the range of 6 μg / mL to 12 μg / mL, or 600 to 1200 times greater than itraconazole. However, imidazole compounds are more lipophilic (higher logP values) than their 1,2,4-triazole counterparts, which is not conducive to drug distribution. In addition, 1,2,4-triazole-containing compounds and imidazole-containing compounds still showed inhibition of CYP3A4, although weaker than itraconazole. For example, the IC50s for CYP3A4 inhibition of compound 101 and compound 4 were 3.4 μM and 2.6 μM, respectively ( Figure 6 ).

[0333] Our previous SAR studies have shown that a 1H-tetrazol-1-yl group in the R1 position significantly reduced CYP3A4 inhibition while increasing anti-angiogenic potency. Therefore, three new analogs were synthesized that have a pyridin-2-yl, a 3'-fluorophenyl group, or a combination in the core region and a tetrazole group in the R1 position. As expected, the resulting tetrazole-containing analogs, Compound 8, Compound 14, and Compound 15, had reduced CYP3A4 inhibition, with an IC50 value of 95 μM for Compound 14, an IC50 value of over 100 μM for Compound 8, and an IC50 value of 37 μM for Compound 15. Figure 6 To our delight, the calculated logP values of the three new analogs were further reduced, and the logP of compound 14 was reduced to 4.36, falling within the range of orally active drugs according to Lipinski's rule of five. Among the three tetrazole-containing analogs, compound 14 also had the highest HUVEC inhibitory potency (IC50 of 77 nM) and solubility. Compared to itraconazole, the solubility of compound 14 increased by more than 90 times.

[0334] Inhibition of HUVEC tube formation. Tetrazole compounds 8, 14, and 15 were selected for endothelial cell tube formation assays to further evaluate their anti-angiogenic potential, with compound 24 serving as a positive control. In this assay, HUVECs assembled into three-dimensional networks and formed tubular structures in Matrigel-coated wells, which recapitulate many key aspects of in vivo neovascularization. As shown in Figure 7, compound 14 inhibited HUVEC tube formation by 70% at 3 μM, as judged by total tube length. The ranking ability to inhibit tube formation was compound 14 > compound 15 > compound 8, which is consistent with their antiproliferative activity in HUVEC.

[0335] Inhibition of NPC1. NPC1 plays an important role in cholesterol export from endolysosomes. Previously, it was reported that itraconazole and its structurally related drug posaconazole directly bind to NPC1 and induce an NPC phenotype in endothelial cells. Using filipin staining, we observed that compound 14 caused a large accumulation of cholesterol in perinuclear structures in the same manner as itraconazole and other tetrazole-containing analogs, compounds 24, 8, and 15 ( Figure 8 ). These results indicate that pyridyl analogs and fluorophenyl analogs retain NPC1 inhibitory activity. To evaluate the binding of compound 14 to NPC1 protein, we used AutoDock Vina software to dock compound 14 and itraconazole into the pocket within the sterol sensing domain (SSD). Figure 9 As shown in A, itraconazole and compound 14 are predicted to bind to NPC1 in a similar manner. Figure 9 B), the linear pyridyl-piperazin-1-yl-phenyl core of compound 14 is positioned within a hydrophobic channel created by the transmembrane helix. The isobutyl tail faces the open entrance of this channel. The tetrazole moiety points toward the closed end of the pocket and interacts with the hydroxyl group of Tyr1225 via hydrogen bonding. These results provide a plausible explanation for how benzene bioisosteric replacements can alter the physicochemical properties of itraconazole without compromising NPC1 inhibition.

[0336] mTOR and VEGFR2 inhibition. We then determined the effect of compound 14 on AMPK / mTOR activity and VEGFR2 glycosylation. Similar to itraconazole, compound 14 activated AMPK in a dose-dependent manner as judged by phosphorylation of its substrate acetyl-CoA carboxylase (ACC). Figure 10A). AMPK activation and NPC1 inhibition were shown to result in synergistic mTOR inhibition by itraconazole. In fact, compound 14 effectively inhibited the phosphorylation of the mTOR substrate p70S6 kinase (S6K). At higher doses of 1 μM and 2 μM, S6K phosphorylation was completely abolished. We have previously reported that itraconazole inhibits VEGFR2 glycosylation and surface expression. We observed two VEGFR2 bands by Western blotting, representing differentially glycosylated forms of the receptor, with the higher molecular weight band being more dominant in untreated HUVECs. Treatment with compound 14 caused a shift in mobility toward the lower molecular weight, hypoglycosylated band. Following treatment with 0.5 μM or higher concentrations of compound 14, the high molecular weight species of VEGFR2 disappeared ( Figure 10A We next used immunofluorescence to determine whether compound 14 and other analogs affected the cellular localization of VEGFR2 ( Figure 10B ). In cells treated with compound 14 and itraconazole, VEGFR2 accumulated in the perinuclear region and co-localized with the Golgi marker GM130. In contrast, in untreated cells, VEGFR2 was evenly distributed in small puncta throughout the cytoplasm. As a key component of lipid rafts, cholesterol is critical for intracellular transport and cell signaling. In our previous studies, we demonstrated that hypoglycosylation of VEGFR2 was rescued by supplementing cellular cholesterol. Therefore, it is possible that the relocation and inhibition of VEGFR2 by itraconazole and its analogs can be mediated by inhibiting NPC1. In short, these results indicate that compound 14 inhibits endothelial cell growth and angiogenesis by simultaneously inhibiting NPC1 and VDAC1, leading to the activation of AMPK and the inhibition of mTOR and VEGFR2 signaling.

[0337] Antiangiogenic therapy has been clinically validated for the treatment of a variety of diseases, including cancer, autoimmune disorders, retinopathy, obesity, macular degeneration, and others. Itraconazole, a newly identified angiogenesis inhibitor, holds great potential and is being investigated in multiple clinical trials. However, its widespread use as an antiangiogenic agent in general, and in combination with other cancer treatment drugs in particular, has been limited by its inhibition of CYP450 and unfavorable physicochemical properties. To improve its solubility and reduce its lipophilicity, we replaced the phenyl group in the core region of itraconazole with a pyridyl or fluorophenyl group. Among the newly synthesized analogs, compound 14, which possesses a 2-pyridyl group at W1 and a 1H-tetrazol-1-yl group at the R1 position, exhibited improved antiangiogenic activity, solubility, and hydrophilicity, with negligible effects on CYP3A4. The antiangiogenic activity of compound 14 was further validated using a tube formation assay. Furthermore, compound 14 exhibits all hallmarks of itraconazole activity in endothelial cells, including activation of AMPK and inhibition of mTOR, induction of cholesterol accumulation in endolysosomes and binding to NPC1, and inhibition of VEGFR2 glycosylation, suggesting that the structural changes required to improve its pharmacological properties do not alter its mechanism of action. This work paves the way for compound 14 to undergo further preclinical studies as a novel antiangiogenic and anticancer drug candidate. Tetrazolium-containing compounds have also been reported to exhibit promising antifungal activity. It will be interesting to determine whether compound 14 also possesses antifungal activity.

[0338] General experimental conditions. Reactions were carried out in oven-dried glassware. Unless otherwise noted, all reagents were purchased from commercial sources and used without further purification. Unless otherwise stated, all reactions were carried out under an argon atmosphere, monitored by Merck pre-coated silica gel 60F-254 plates and visualized using 254nm UV light. Column chromatography was performed on a normal phase silica gel flash column (RediSepRf). NMR data were collected on a Bruker Avance III (500MHz 1H, 125MHz 13C) machine at the Department of Pharmacology and Molecular Sciences, the Johns Hopkins University School of Medicine. 1 H NMR spectroscopy and 13C NMR spectra were obtained in deuterated chloroform (CDCl3) with tetramethylsilane (TMS, δ = 0.00 for 1H) as an internal reference. Chemical shifts are reported in ppm (δ). Data are presented in the following form: chemical shift (multiplicity, coupling constant, and integration). Low-resolution ESI-MS and HPLC purity were recorded on an Agilent 6120 quadrupole LC / MS. The reported purity values were obtained using a Pursuit XRs diphenyl column (150×4.5 mm) and a diode array detector (DAD). A flow rate of 1.0 ml / min was used with a mobile phase of acetonitrile in H2O with 0.1% modifier (formic acid, v / v).

[0339] The general experimental procedure of compound 57, compound 59-62, compound 33-35, compound 38-41, compound 43, compound 45, compound 46 and compound 48-53. Under argon atmosphere, sodium hydride (NaH, 60% dispersion in mineral oil, (1.5 equivalents)) is added to the solution of toluenesulfonic acid ester 103 or 108 (1 equivalent) in dry DMF. Afterwards, reaction mixture is stirred at 50 DEG C for 1 hour. At the same temperature, the solution of 105a (1.2 equivalents) in DMF is slowly added. After interpolation, the temperature is increased to 90 DEG C and stirring is continued for another 3 hours. Reaction mixture is quenched by saturated sodium chloride, and the resulting mixture is extracted twice with dichloromethane. By organic fraction through Na2SO4 drying, filter and concentrate under vacuum, to produce crude product, this crude product is purified by column chromatography, and the product of expectation is provided with medium to good yield.

[0340] 1-(2,4-dichlorophenyl)-2-(5-methyl-1H-tetrazol-1-yl) ethyl-1-one (intermediate 13c). To a solution of 1.44g tetrazole and 3.1g anhydrous KCO in 150mL acetonitrile (ACN) was added dropwise 4.2g 2-chloro-1-(2,4-dichlorophenyl) ketene (intermediate 12). Before adding water, the reaction mixture was stirred at room temperature for 16h. The mixture was extracted three times with ethyl acetate (EtOAc), washed with salt water, and dried over MgSO. The solution was concentrated under reduced pressure and purified by column chromatography (1.9g, 40% yield). 1H NMR (500MHz, CDCl3, δH): 7.71 (d, J = 8.5Hz, 1H), 7.48 (d, J = 2.0Hz, 1H), 7.37 (m, 1H), 5.80 (s, 2H), 2.46 (s, 3H).13C NMR (125MHz, CDCl3, δC): 190.1, 153.2, 140.0, 133.2, 132.6, 131.8, 131, 131.0, 130.4, 128.1, 55.3, 8.7. ESI-MS[M+H]+257.2.

[0341] ((2S,4S)-2-((1H-tetrazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolane-4-yl)methyl 4-methylbenzenesulfonate (Intermediate 14c). Intermediate 12c (1 g) and Intermediate 13 (1.1 g) were dissolved in 15 mL of dry toluene under an ice bath. Trifluoromethanesulfonic acid (TfOH, 1.53 mL) was added and the ice bath was removed. The reaction was stirred at room temperature for 60 h. The progress of the reaction was monitored by TLC. The reaction was quenched by the addition of saturated aqueous NaHCO3, followed by extraction with ethyl acetate, washed with brine and dried over Na2SO4. The solvent was removed under reduced pressure, and the crude product was purified by column chromatography to provide the cis-dioxolane product (Intermediate 14c) (0.98 g, 55% yield) as a light yellow oil. 1H NMR (500MHz, CDCl3, δH): 8.41 (s, 1H), 7.79 (d, J = 8.5Hz, 1H), 7.46 (d, J = 2Hz, 1H), 7.43-7.38 (m, 3H), 7.21 (dd, J = 8.5, 2.0Hz, 1H), 5 .30-5.17(m,2H),4.27-4.25(m,1H),3.92-3.88(m,1H),3.86-3.84(m,1H),3.76-3.74(m,1H),3.61-3.57(m,1H),2.48(s,3H).13C NMR (125MHz, CDCl3, δC): 154.4, 151.9, 121.0, 118.9, 117.3, 114.6, 112.8, 112.5, 55.6, 51.0, 49.6. ESI-MS[M+H]+485.3.

[0342] 1-(2-Fluoro-4-nitrophenyl)-4-(4-methoxyphenyl)piperazine (Intermediate 17c). In a dry flask, 1-(2-fluoro-4-nitrophenyl)piperazine (Intermediate 16b, 550 mg) was mixed with 1-bromo-4-methoxybenzene (Intermediate 15a, 500 mg), tris(dibenzylideneacetone)dipalladium (Pd2(dba)3, 120 mg), sodium tert-butoxide (NaOt-Bu, 380 mg), and (±)-2,2′-bis(diphenylphosphino)-1,1′-binaphthyl (BINAP, 245 mg) in 20 mL of dry toluene. The mixture was stirred at 80°C under argon overnight. After cooling to room temperature, water was added and the organic phase was separated. The aqueous phase was extracted three times with dichloromethane (DCM). The combined organic layers were dried over Na 2 SO 4 , which was purified by column chromatography to give Intermediate 17c (2.5 g, yield 75%) as a light yellow solid. 1H NMR (500MHz, CDCl3, δH): 8.01 (dd, J=8.8, 2.4Hz, 1H), 7.93 (dd, J=12.8, 2.4Hz, 1H), 6.9 9-6.97(m,3H),6.89-6.87(m,2H),3.79(s,3H),3.47(bs,4H),3.26(d,J=4.8Hz,4H).13C NMR (125MHz, CDCl3, δC): 154.4, 151.9, 121.0, 118.9, 117.3, 114.6, 112.8, 112.5, 55.6, 51.0, 49.6. ESI-MS[M+H]+332.2.

[0343] 1-(sec-butyl)-4-(3-fluoro-4-(4-(4-methoxyphenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (intermediate 21c). To a suspension of 0.55 g of triazolone intermediate 20c in 20 mL of dimethyl sulfoxide was added 0.4 g of KCO. The resulting mixture was stirred at room temperature for 6 h. An aliquot of 0.25 mL of 2-bromobutane was added dropwise at room temperature. The temperature was then increased to 80 ° C and stirred overnight. After cooling, the reaction mixture was diluted with water and extracted with EtOAc. The combined organic layers were dried over NaSO, filtered and concentrated under reduced vacuum to produce a crude product, which was purified by column chromatography to obtain intermediate 21c (0.55 g, 87% yield). 1H NMR (500MHz, CDCl3, δH): 7.64 (s, 1H), 7.37 (dd, J=13.0, 2.2Hz, 1H), 7.26-7.24 (m, 1H), 7.04 (t, J=8.8Hz, 1H), 6.99-6.97 (m, 2H), 6.88-6.84 ( m,2H),4.31-4.26(m,1H),3.77(s,3H),3.26(bs,8H),1.86-1.81(m,1H),1.73-1.71(m,1H),1.38(d,J=6.8Hz,3H),0.89(t,J=7.4Hz,3H).13C NMR(125MHz, CDCl3, δC):154.6,154.2,151.6,139.4,133.3,128.4,119.4,1 18.7,118.0,114.5,111.0,110.8,55.6,52.8,51.0,50.5,28.4,19.3,10.8. ESI-MS[M+H]+:425.5.

[0344] 1-(sec-Butyl)-4-(3-fluoro-4-(4-(4-hydroxyphenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Intermediate 22c). 0.5 g of triazolone intermediate 21c was added to aqueous HBr (48%, 10 mL) in a flask. The reaction was heated to 120°C and refluxed overnight. After cooling to room temperature, the solution was neutralized with cold saturated Na2CO3 and extracted three times with DCM. The combined organic layers were dried over Na2SO4, filtered and concentrated to give a crude product, which was purified by column chromatography to obtain intermediate 22c (0.4 g, 83% yield). 1H NMR (500MHz, CDCl3, δH): 7.57 (s, 1H), 7.24 (dd, J = 10.2, 2.0Hz, 1H), 7.15 (dd, J = 6.8, 1.6Hz, 1H), 6.92 (t, J = 7.0Hz, 1H), 6.79 (d, J = 6.8Hz, 2H), 6. 67(d,J=7.2Hz,2H),4.24-4.20(m,1H),3.17-3.16(m,8H),1.82-1.77(m, 1H),1.66-1.63(m,1H),1.32(d,J=5.6Hz,3H),0.82(t,J=5.8Hz,3H).13C NMR (125MHz, CDCl3, δC): 156.3, 154.3, 150.8, 144.9, 139.7, 133.6, 127.8, 119.4, 118.9, 116.0, 111.4, 53.1, 51.2, 50.5, 28.4, 19.3, 10.8. ESI-MS[M+H]+:411.5.

[0345] 4-(4-(4-(4-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 1). 1H NMR (500MHz, CDCl3, δH): 8.08 (s, 1H), 7.64 (s, 1H), 7.59 (d, J = 8.5Hz, 1H), 7.49 (d, J = 2.0Hz, 1H), 7.40 (d, J = 2.0Hz, 1H), 7.39 (dd, J = 13.0 ,2.5Hz,1H),7.30(dd,J=8.5,2.0Hz,1H),7.25-7.23(m,2H),7.15-7.12(m,2H),7.08-7.04(m,1H),7.01(d,J=8.5Hz,1H),6.82(d,J=9.0 Hz,1H),4.61(q,J=14.5Hz,2H),4.39-4.36(m,1H),4.32-4.27(m,1H),3.93(t,J=8.5Hz,1H),3.82-3.79(m,1H),3.75-3.73(m,1H),3.59 -3.56(m,1H),3.35-3.33(m,4H),3.30-3.29(m,4H),1.90-1.84(m,1H),1.76-1.70(m,1H),1.41(d,J=7.0Hz,3H),0.91(t,J=7.0Hz,3H). 13C NMR (125 MHz, CDCl3, δC): 156.4, 152.4, 151.8, 146.1, 140.4, 136.6, 134.3, 133.8, 133.0, 131.5, 129.6, 128.2, 127.8, 119.4, 118.9, 115.7, 111.2, 109.8, 107.2, 74.8, 70.7, 68.7, 67.4, 52.2, 50.4, 48.9, 28.4, 19.3, 10.8. HRMS (ESI) calculated for C36H38Cl2FN7O4: 722.2425; found 722.2429. HPLC purity: 95.8%, tR=5.6min.

[0346] 4-(4-(4-(6-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)piperazin-1-yl)phenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 2). 1H NMR (500MHz, CDCl3, δH): 8.22 (s, 1H), 7.57 (s, 1H), 7.40 (m, 1H), 7.39-7.37 (m, 3H), 7.32 (dd, J = 9.5, 3Hz, 1H), 7.18 (dd, J = 8 .5,2Hz,1H),7.14(s,1H),7.01(s,1H),6.99(d,J=9.0Hz,2H),6.74(d,J=3.0Hz,1H),6.52(d,J=9.0Hz,1H),4.58(q,J=14.5H z,2H),4.35-4.32(m,1H),4.26-4.21(m,1H),4.05-4.01(m,1H),3.94-3.91(m,1H),3.59-3.56(m,1H),3.51-3.46(m,1H),3 .35-3.32(m,4H),3.06(t,J=5.0Hz,4H),1.87-1.78(m,1H),1.72-1.63(m,1H),1.36(d,J=7.0Hz,3H),0.86(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 156.3, 152.6, 151.3, 146.4, 136.0, 138.9, 134.2, 133.8, 133.2, 131.6, 130.1, 129.5, 127.3, 123.4, 117.0, 111.2, 107.4, 74.8, 67.2, 64.9, 53.8, 52.8, 50.6, 49.3, 41.0, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C35H38Cl2N8O4: 705.2471; found: 705.2463. HPLC purity: 96.3%, tR = 3.6 min.

[0347] 4-(4-(4-(5-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)piperazin-1-yl)phenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 4). 1H NMR (500MHz, CDCl3, δH): 7.90-7.87(m,2H),7.62(s,1H),7.61(d,J=8.0Hz,1H),7.49(d,J=2.5,1H),7.44(m,3H),7.30( dd,J=8.0,2Hz,1H),7.19-7.17(m,1H),7.11(s,1H),7.04(d,J=9.0Hz,2H),6.76(d,J=8.5Hz,1H),4.57(q,J=14.5Hz,2H ),4.38-4.35(m,1H),4.32-4.28(m,1H),3.93-3.90(m,1H),3.78-3.76(m,1H),3.74-3.71(m,1H),3.64-3.62(m,4H),3. 51-3.48(m,1H),3.35-3.34(m,4H),1.91-1.84(m,1H),1.75-1.70(m,1H),1.41(d,J=6.5Hz,3H),0.91(t,J=6.5Hz,3H). C NMR (125 MHz, CDCl, δC): 156.4, 152.1, 150.6, 138.7, 136.3, 134.2, 134.0, 133.0, 132.0, 131.6, 129.5, 127.4, 126.0, 123.6, 116.8, 107.7, 98.1, 74.8, 68.5, 52.7, 48.9, 46.3, 29.7, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C35H38Cl2N8O4: 705.2471; found: 705.2473. HPLC purity: 95.4%, tR = 3.7 min.

[0348] 4-(4-(4-(5-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 5). 1H NMR (500MHz, CDCl3, δH): 8.02 (d, J = 2.5Hz, 1H), 7.75 (s, 1H), 7.66-7.65 (m, 2H), 7.49 (d, J = 2.0Hz, 1H), 7.40 (t, J = 2.0Hz ,1H),7.37(t,J=2.0Hz,1H),7.30-7.27(m,1H),7.26-7.24(m,1H),7.15(dd,J=9.0,2.5Hz,1H),7.07-7.01(m,2H),6.71 (t,J=8.5Hz,1H),4.55(q,J=14.5Hz,2H),4.38-4.27(m,2H),3.92-3.88(m,1H),3.77-3.72(m,2H),3.63-3.60(m,4H),3 .45-3.41(m,1H),3.25-3.21(m,4H),1.91-1.82(m,1H),1.77-1.68(m,1H),1.40(d,J=7.5Hz,3H),0.91(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 156.6, 155.2, 151.7, 148.0, 141.3, 136.1, 134.6, 133.3, 133.0, 131.5, 129.5, 127.4, 125.7, 119.5, 118.0, 110.8, 108.3, 74.8, 67.3, 52.9, 50.3, 47.0, 46.5, 28.4, 19.3, 10.8. HRMS (ESI) calcd for C35H37Cl2FN8O4: 723.2377; found: 723.2371. HPLC purity: 95.8%, tR = 4.3 min.

[0349] 4-(6-(4-(4-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)pyridin-3-yl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 6). 1H NMR (500MHz, CDCl3, δH): 8.26 (d, J = 3.0Hz, 1H), 7.75 (dd, J = 9.0Hz, 3.0Hz, 1H), 7.63 (s, 1H), 7.60-7.57 (m, 2 H),7.48(d,J=2.0Hz,1H),7.28(dd,J=8.0,2.0Hz,1H),7.04-7.00(m,2H),6.95-6.93(m,2H),6.80-6.76(m,3 H),4.52(q,J=14.5Hz,2H),4.38-4.27(m,2H),3.91-3.86(m,1H),3.77-3.72(m,6H),3.38-3.34(m,1H),3.2 0-3.18(t,J=4.5Hz,4H),1.92-1.83(m,1H),1.77-1.69(m,1H),1.41(d,J=7.0Hz,3H),0.91(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 158.4, 152.7, 152.1, 146.0, 142.4, 136.0, 134.4, 133.8, 132.9, 131.5, 129.6, 127.3, 121.3, 118.6, 115.3, 107.9, 107.1, 74.9, 67.7, 52.9, 50.5, 45.4, 28.5, 19.3, 10.8. HRMS (ESI) calculated for C35H38Cl2N8O4: 705.2471; found: 705.2468. HPLC purity: 96.1%, tR = 4.6 min.

[0350] 4-(5-(4-(4-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)pyridin-2-yl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 7). 1H NMR (500MHz, CDCl3, δH): 8.34 (s, 1H), 8.22 (d, J = 9.0Hz, 1H), 8.22 (d, J = 3.0Hz, 1H), 7.77 (s, 1H), 7.60 (d, J = 8.5Hz, 1 H),7.76(d,J=2.0Hz,1H),7.41(dd,J=9.0,3.0Hz,1H),7.28(dd,J=8.5,2.0Hz,1H),7.05-7.01(m,2H),6.96(d,J=9.0 Hz,1H),6.81(d,J=9.0Hz,1H),4.54(q,J=14.5Hz,2H),4.38-4.28(m,2H),3.91-3.88(m,1H),3.78-3.72(m,2H),3.39 -3.36(m,4H),3.27-3.25(m,5H),1.90-1.85(m,1H),1.75-1.71(m,1H),1.41(d,J=7.0Hz,3H),0.91(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 152.8, 145.8, 140.2, 136.1, 134.3, 133.0, 132.5, 131.5, 129.6, 127.3, 125.6, 118.6, 115.3, 113.7, 107.8, 74.9, 67.7, 52.6, 50.5, 49.0, 28.4, 19.3, 10.8. HRMS (ESI) calcd for C35H38Cl2N8O4: 705.2471; found: 705.2463. HPLC purity: 95.6%, tR = 5.5 min.

[0351] 4-(4-(4-(4-(((2S,4R)-2-((1H-tetrazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 8). 1H NMR (500MHz, CDCl3, δH): 8.46 (s, 1H), 7.64 (s, 1H), 7.56 (d, J = 8.0Hz, 1H), 7.48 (s, 1H), 7.39 (d, J = 3.0Hz, 1 H),7.27-7.23(m,3H),7.05(t,J=8.5Hz,1H),6.94(d,J=8.5Hz,2H),6.80(d,J=8.5Hz,2H),5.35(q,J=14.0 Hz,2H),4.40-4.38(m,1H),4.31-4.27(m,1H),3.96(t,J=8.0Hz,1H),3.89-3.83(m,2H),3.53(t,J=8.0Hz, 1H), 3.41-3.38 (m, 8H), 1.90-1.83 (m, 1H), 1.76-1.70 (m, 1H), 1.41 (d, J = 7.0Hz, 3H), 0.91 (t, J = 7.0Hz, 3H). 13C NMR (125 MHz, CDCl3, δC): 156.9, 154.9, 153.1, 152.2, 146.7, 140.0, 136.9, 134.3, 134.0, 132.1, 130.2, 129.0, 128.9, 127.9, 120.1, 119.1, 118.7, 116.1, 111.8, 111.6, 108.2, 75.9, 69.0, 68.7, 57.8, 54.1, 51.9, 51.8, 29.8, 20.7, 12.5. HRMS (ESI) calculated for C34H36Cl2FN9O4: 724.2330; found 724.2328. HPLC purity: 96.7%, tR=12.7 min.

[0352] 4-(4-(4-(5-(((2S,4R)-2-((1H-tetrazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)piperazin-1-yl)phenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 14). 1H NMR (500MHz, CDCl3, δH): 8.47 (s, 1H), 7.88 (d, J = 2.5Hz, 1H), 7.61 (s, 1H), 7.56 (d, J = 8.5Hz, 1H), 7.49 (d, J = 1.5Hz, 1H),7.43(d,J=9.0Hz,2H),7.26-7.24(m,1H),7.15(dd,J=9.0,3.0Hz,1H),7.04(d,J=9.0Hz,2H),6.70(d,J=9.0Hz, 1H),5.35(q,J=14.0Hz,2H),4.41-4.36(m,1H),4.32-4.28(m,1H),3.96(t,J=8.0Hz,1H),3.88-3.82(m,2H),3.62- 3.57(m,5H),3.35-3.33(m,4H),1.92-1.83(m,1H),1.77-1.68(m,1H),1.41(d,J=7.0Hz,3H),0.91(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 155.7, 152.5, 151.1, 148.5, 136.9, 135.3, 134.2, 134.0, 132.2, 130.0, 127.9, 126.7, 126.5, 124.3, 117.5, 109.0, 75.8, 69.8, 68.5, 57.8, 53.9, 50.2, 47.5, 29.9, 20.8, 12.4. HRMS (ESI) calcd for C33H36Cl2N10O4: 707.2376; found: 707.2379. HPLC purity: 98.9%, tR = 8.2 min.

[0353] 4-(4-(4-(5-(((2S,4R)-2-((1H-tetrazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 15). 1H NMR (500MHz, CDCl3, δH): 8.47 (s, 1H), 7.88 (d, J = 3.0Hz, 1H), 7.64 (s, 1H), 7.56 (d, J = 8.5Hz, 1H), 7.49 (d, J = 2.0Hz, 1H ),7.39(dd,J=13.0,2.5Hz,1H),7.25-7.24(m,2H),7.14(dd,J=9.0,3.0Hz,1H),7.04(t,J=9.0Hz,1H),6.69(d,J=9.0 Hz,1H),5.35(q,J=14.5Hz,2H),4.39-4.37(m,1H),4.30-4.28(m,1H),3.98-3.94(m,1H),3.88-3.82(m,1H),3.65-3. 56(m,5H),3.23(t,J=5.0Hz,3H),1.89-1.83(m,1H),1.76-1.69(m,1H),1.40(d,J=6.5Hz,3H),0.91(t,J=6.5Hz,3H). 13C NMR (125MHz, CDCl3, δC):156.0,155.0,154.1,152.4,151.3,147.7,139.2,136.1,134.5,133.3,133.1,131.3,129.4,12 8.2,127.1,125.6,119.3,117.8,111.0,110.8,108.2,107.4,75.0,69.0,67.6,56.9,53.2,50.7,46.8,29.0,19.9,11.5. HRMS (ESI) calculated for C33H35Cl2FN10O4: 725.2282; found: 725.2281. HPLC purity: 99.0%, tR = 10.8 min.

[0354] 4-(4-(4-(4-(((2S,4R)-2-((1H-imidazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1-sec-butyl-1H-1,2,4-triazol-5(4H)-one (Compound 33). 1H NMR (500MHz, CDCl3, δH): 7.61 (s, 1H), 7.58 (d, J = 8.0Hz, 1H), 7.53 (bs, 1H), 7.46 (d, J = 2.5Hz, 1H), 7.42 (d, J = 9.5Hz, 2H), 7.26 (dd, J=8.2,2.2Hz,2H),7.03(d,J=9Hz,2H),7.00-6.98(m,1H),6.93(d,J=9.0Hz,2H),6.78(d,J=9.5Hz,2H),4.51(d,J=15.0Hz,1H),4. 41(d,J=15.0Hz,1H),4.41(d,J=15.0Hz,1H),4.34–4.28(m,2H),3.87(dd,J=8.5,6.5Hz,1H),3.74–3.72(m,2H),3.36(t,J=5.0Hz, 4H),3.32–3.31(m,1H),3.23(t,J=5.0Hz,4H),1.89–1.83(m,1H),1.74–1.71(m,1H),1.39(d,J=7.0Hz,3H),0.90(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 152.7, 152.1, 150.6, 146.0, 136.0, 134.6, 134.0, 133.0, 131.4, 129.5, 127.3, 125.9, 123.6, 118.5, 116.7, 115.3, 108.0, 74.8, 67.7, 67.6, 52.7, 50.6, 49.3, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C36H39Cl2N7O4: 704.2519; found: 704.2525. HPLC purity: 95.3%, tR = 8.3 min.

[0355] 4-(4-(4-(((2S,4R)-2-((1H-pyrazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1-sec-butyl-1H-1,2,4-triazol-5(4H)-one (Compound 34). 1H NMR (500MHz, CDCl3, δH): 7.62 (s, 1H), 7.58 (d, J = 8.5Hz, 1H), 7.49 (t, J = 2.2Hz, 2H), 7.45 (d, J = 2.0Hz, 1H), 7.43 (d, J = 9.0Hz, 2H), 7.22 (dd,J=9.5,2.0Hz,1H),7.03(d,J=9.5Hz,2H),6.93(d,J=8.5Hz,2H),6.77(d,J=9.0Hz,2H),6.22(t,J=2.0Hz,1H),4.79(d,J=14.5Hz,1 H),4.68(d,J=15.0Hz,1H),4.36-4.33(m,1H),4.31-4.27(m,1H),3.87(dd,J=8.5,6.5Hz,1H),3.80(dd,J=8.5,4.5Hz,1H),3.77(dd,J =8.5,5.0Hz,1H),3.37-3.31(m,5H),3.24(bs,4H),1.90–1.84(m,1H),1.74–1.70(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.2Hz,3H). C NMR (125 MHz, CDCl, δC): 152.1, 139.4, 135.6, 135.0, 134.0, 133.2, 133.3, 131.2, 129.7, 127.1, 123.6, 118.5, 116.7, 115.3, 108.5, 105.8, 74.6, 68.0, 67.6, 56.1, 52.7, 50.7, 49.3, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C36H39Cl2N7O4: 704.2519; found: 704.2542. HPLC purity: 95.9%, tR = 11.6 min.

[0356] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2-methyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 35). 1H NMR (500MHz, CDCl3, δH): 7.63 (d, J = 8.5 Hz, 1H), 7.61 (s, 1H), 7.47 (d, J = 2.0 Hz, 1H), 7.42 (d, J = 9.0 Hz, 2H), 7.29 (dd, J = 8 .5,2.0Hz,1H),7.03(d,J=9.0Hz,2H),6.94-6.90(m,3H),6.77(d,J=9.5Hz,2H),4.41(d,J=15.0Hz,1H),4.34(d,J=15.0H z,1H),4.32–4.26(m,2H),3.85(dd,J=8.5,6.5Hz,1H),3.76-3.73(m,1H),3.66(dd,J=9.5,5.0Hz,1H),3.36(t,J=4.5Hz, 4H), 3.24-3.22 (m, 5H), 2.49 (s, 3H), 1.89–1.84 (m, 1H), 1.74–1.69 (m, 1H), 1.39 (d, J = 6.5Hz, 3H), 0.90 (t, J = 7.5Hz, 3H). C NMR (125 MHz, CDCl, δC): 152.7, 152.1, 150.6, 146.0, 136.0, 134.0, 133.1, 131.6, 129.6, 127.3, 125.9, 123.6, 118.5, 116.7, 115.3, 108.6, 74.8, 67.6, 67.4, 52.7, 50.6, 49.3, 43.2, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C37H41Cl2N7O4: 718.2675; found: 718.2645. HPLC purity: 95.0%, tR = 5.3 min.

[0357] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((4-methyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 38). 1H NMR (500MHz, CDCl3, δH): 7.61 (s, 1H), 7.58 (d, J = 8.5Hz, 1H), 7.49 (bs, 1H), 7.46 (d, J = 2.0Hz, 1H), 7.42 (d, J = 9.0Hz, 2H), 7.27-7.25 (m, 1H ),7.03(d,J=9.0Hz,2H),6.94(dd,J=9.0,2.0Hz,2H),6.78(d,J=9.0Hz,2H),6.69(bs,1H),4.43(d,J=15.0Hz,1H),4.34(d,J=15.0Hz,1H), 4.33–4.27(m,2H),3.86(dd,J=8.5,6.5Hz,1H),3.79(dd,J=8.5,4.5Hz,1H),3.71(dd,J=9.5,5.0Hz,1H),3.36(t,J=4.5Hz,4H),3.29(dd, J=9.5,6.5Hz,1H),3.23(t,J=5.0Hz,4H),2.18(s,3H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.7, 152.1, 150.6, 146.0, 136.0, 134.6, 134.0, 133.0, 131.4, 129.6, 127.3, 125.9, 123.6, 118.5, 116.7, 115.2, 108.0, 74.8, 67.7, 67.5, 52.7, 51.4, 50.7, 49.3, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C37H41Cl2N7O4: 718.2675; found: 718.2697. HPLC purity: 95.7%, tR = 5.3 min.

[0358] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2-methyl-4-(trifluoromethyl)-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 39). 1H NMR (500MHz, CDCl3, δH): 7.64 (d, J = 8.5Hz, 2H), 7.61 (s, 1H), 7.48 (d, J = 1.5Hz, 1H), 7.42 (d, J = 9Hz, 2H), 7 .31-7.29(m,2H),7.03(d,J=9Hz,2H),6.93(d,J=9Hz,2H),6.75(d,J=9Hz,2H),4.44-4.41(m,1H),4.37-4 .26(m,3H),3.85(t,J=8.5Hz,1H),3.78-3.76(m,1H),3.65(q,J=4.5Hz,1H),3.32-3.38(m,4H),3.40-3.2 0(m,4H),2.48(s,3H),1.90-1.84(m,1H),1.75-1.69(m,1H),1.40(d,J=6.5Hz,3H),0.91(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.7, 152.1, 150.6, 146.0, 136.1, 134.5, 133.0, 132.0, 129.6, 127.2, 125.9, 123.6, 121.9, 118.6, 116.5, 107.2, 74.7, 67.8, 52.7, 50.8, 49.3, 28.5, 19.3, 14.1, 10.8. HRMS (ESI) calcd for C38H40Cl2F3N7O4: 786.2549; found: 786.2562. HPLC purity: 95.7%, tR = 11.9 min.

[0359] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2,4-dimethyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 40). 1H NMR (500MHz, CDCl3, δH): 7.65 (d, J = 8.5Hz, 1H), 7.61 (s, 1H), 7.43-7.40 (m, 3H), 7.24 (s, 1H), 7.03 (d, J = 8.5Hz, 2H),6.89(d,J=8.5Hz,2H),6.73(s,1H),6.66(d,J=8.5Hz,2H),4.35-4.24(m,3H),4.18-4.15(m,1H),3.94(t,J =7.5Hz,1H),3.89-3.86(m,1H),3.82-3.79(m,1H),3.75(t,J=7.5Hz,1H),3.33-3.37(m,4H),3.20-3.22(m,4H) ,2.50(s,3H),2.20(s,3H),1.90-1.82(m,1H),1.75-1.70(m,1H),1.40(d,J=6.5Hz,3H),0.91(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.6, 152.0, 150.2, 146.9, 136.2, 143.0, 134.1, 133.8, 131.2, 129.6, 127.2, 125.5, 122.5, 118.2, 117.5, 115.2, 108.2, 75.3, 67.9, 67.4, 52.5, 50.7, 49.4, 28.7, 15.6, 14.2, 10.8. HRMS (ESI) calculated for C38H43Cl2N7O4: 732.2832; found: 732.2803. HPLC purity: 94.8%, tR = 5.0 min.

[0360] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2,5-dimethyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 41). 1H NMR (500MHz, CDCl3, δH): 7.73 (d, J = 8.5 Hz, 1H), 7.61 (s, 1H), 7.45-7.41 (m, 3H), 7.24 (s, 1H), 7.03 (d, J = 8.5 Hz,2H),6.89(d,J=8.5Hz,2H),6.67-6.65(m,3H),4.33-4.27(m,2H),4.24-4.21(m,1H),4.02-3.98(m,1H),3 .85-3.84(d,J=4.5Hz,1H),3.81-3.76(m,2H),3.72(t,J=1.5Hz,1H),3.37-3.34(m,4H),3.24-3.20(m,4H),2 .51(s,3H),2.30(s,3H),1.90-1.85(m,1H),1.75-1.70(m,1H),1.41(d,J=6.5Hz,3H),0.93(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.7, 152.0, 150.2, 146.9, 136.2, 142.7, 134.1, 133.8, 131.2, 128.0, 127.2, 125.5, 122.0, 118.2, 117.5, 115.2, 108.2, 75.3, 67.9, 67.4, 52.5, 50.7, 49.7, 28.7, 14.0, 10.8. HRMS (ESI) calculated for C38H43Cl2N7O4: 732.2832; found: 732.2837. HPLC purity: 95.0%, tR = 5.2 min.

[0361] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2-ethyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 43). 7.65(d,J=8.5Hz,1H),7.62(s,1H),7.48(d,J=2Hz,1H),7.43(d,J=9.0Hz,2H),7.30(dd,J=8.5,2.0Hz,1H),7.04(d,J=9Hz,2H),6.95-6 .92(m,4H),6.77(d,J=9.0Hz,2H),4.51(d,J=15.0Hz,1H),4.44(d,J=15.0Hz,1H),4.38(d,J=15.0Hz,1H),4.35–4.27(m,2H),3.86(dd,J =8.5,6.5Hz,1H),3.76(dd,J=8.5,4.5Hz,1H),3.65(dd,J=9.5,5.2Hz,1H),3.37(t,J=5.0Hz,4H),3.24(t,J=5.0Hz,4H),3.15(t,J=8.5H z,1H),2.8(q,J=7.5Hz,2H),1.90–1.84(m,1H),1.75–1.70(m,1H),1.41(d,J=6.5Hz,3H),1.34(3t,J=7.5Hz,3H),0.91(t,J=7.5Hz,3H). 13C NMR (125 MHz, CDCl3, δC): 153.2, 152.6, 151.3, 151.1, 146.4, 136.5, 135.7, 133.8, 132.1, 130.2, 128.0, 127.2, 126.6, 124.3, 122.3, 119.2, 117.4, 116.0, 109.5, 75.9, 68.7, 68.6, 53.9, 51.9, 50.8, 50.5, 29.9, 21.5, 20.8, 13.4, 12.4. HRMS (ESI) calculated for C38H43Cl2N7O4: 732.2832; found 732.2855. HPLC purity: 98.2%, tR=5.6 min.

[0362] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2-isopropyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 45). 1H NMR (500MHz, CDCl3, δH): 7.63(d,J=8.5Hz,1H),7.61(s,1H),7.48(d,J=2.5Hz,1H),7.42(d,J=9.0Hz,2H),7.29(dd,J=8.5,2.0Hz,1H),7.03(d,J=9Hz ,2H),6.97(bs,1H),6.94(s,1H),6.93-6.92(m,2H),6.75(d,J=9.0Hz,2H) ,4.51(d,J=15.0Hz,1H),4.44(d,J=15.0Hz,1H),4.38(d,J=15.0Hz,1H),4. 35–4.27(m,2H),3.85(dd,J=8.5,6.5Hz,1H),3.76(dd,J=8.5,4.5Hz,1H), 3.66(dd,J=9.5,5.2Hz,1H),3.36(t,J=5.2Hz,4H),3.26(t,J=7.0Hz,1H),3 .23(t,J=5.0Hz,4H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=6.5H z, 3H), 1.34 (d, J = 7.0Hz, 3H), 1.31 (d, J = 6.5Hz, 3H), 0.90 (t, J = 7.5Hz, 3H). 13C NMR (125 MHz, CDCl3, δC): 154.4, 152.6, 152.0, 150.6, 146.0, 136.0, 133.9, 133.1, 131.5, 129.6, 127.4, 125.9, 123.6, 121.3, 118.5, 116.7, 115.2, 108.3, 74.8, 67.6, 67.6, 52.7, 50.6, 49.3, 28.5, 25.5, 22.0, 21.6, 19.3, 10.8. HRMS (ESI) calculated for C39H45Cl2N7O4: 746.2988; found 746.2996. HPLC purity: 95.8%, tR=6.0 min.

[0363] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((2-phenyl-1H-imidazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 46). 1H NMR (500MHz, CDCl3, δH): 7.61 (s, 2H), 7.60 (d, J = 3.5Hz, 1H), 7.51 (d, J = 8.5Hz, 1H), 7.43-7.40 (m, 5H), 7.30 (d, J = 2.5Hz, 1H), 7. 25(d,J=1.5Hz,1H),7.22(dd,J=8.5,2.5Hz,1H),7.13(s,1H),7.03(d,J=9.5Hz,2H),6.94(d,J=9.5Hz,2H),6.79(d,J=9.5Hz,2H ),4.56(s,2H),4.38–4.27(m,2H),3.89-3.87(m,2H),3.80(dd,J=9.5,5.0Hz,1H),3.50(dd,J=9.5,6.5Hz,1H),3.37-3.34(m,5H ),3.23(t,J=5.5Hz,4H),3.20-3.19(m,1H),1.88–1.83(m,1H),1.74–1.70(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.6, 152.0, 150.6, 146.0, 133.9, 131.4, 129.6, 129.5, 128.5, 127.1, 125.9, 123.6, 123.0, 118.9, 116.0, 115.3, 108.4, 74.7, 67.9, 67.3, 52.7, 50.6, 49.3, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C₄₂H₄₃Cl₂NₐO₄: 780.2832; found 780.2819. HPLC purity: 95.6%, tR = 6.5 min.

[0364] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((3,5-dimethyl-1H-1,2,4-triazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 47). 1H NMR (500MHz, CDCl3, δH): 7.67 (d, J = 8.5 Hz, 1H), 7.61 (s, 1H), 7.47 (d, J = 2.0 Hz, 1H), 7.43 (d, J = 9.0 Hz, 2H), 7.29 (dd, J = 8.5, 2.0 Hz, 1H), 7. 03(d,J=9.5Hz,2H),6.97(d,J=9.5Hz,2H),6.77(dd,J=7.0,2.5Hz,2H),4.64(d,J=15.0Hz,1H),4.54(d,J=15.0Hz,1H),4.36(dt,J=6.5,5 .0,2.0Hz,1H),4.29(qt,J=8.5,6.5,4.0Hz,1H),3.84(t,J=5.0Hz,1H),3.67(dd,J=5.0,6.5Hz,1H),3.39(t,J=5.0Hz,4H),3.29-3.38(m, 1H),3.24(t,J=5.0Hz,4H),2.51(s,3H),2.30(s,3H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.5Hz,3H). 13C NMR (125 MHz, CDCl3, δC): 159.0, 154.5, 152.1, 150.5, 136.0, 134.7, 134.0, 133.1, 131.4, 129.8, 127.3, 127.1, 126.0, 123.6, 123.6, 118.7, 116.8, 116.2, 115.2, 108.6, 74.6, 67.6, 67.3, 52.7, 51.9, 50.8, 49.2, 28.5, 19.3, 13.7, 10.8. HRMS (ESI) calculated for C37H42Cl2N8O4: 733.2784; found 733.2764. HPLC purity: 95.4%, tR=7.9 min.

[0365] 4-(4-(4-(((2S,4R)-2-((1H-tetrazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1-sec-butyl-1H-1,2,4-triazol-5(4H)-one (Compound 48). 1H NMR (500MHz, CDCl3, δH): 8.46 (s, 1H), 7.61 (s, 1H), 7.55 (d, J = 8.5Hz, 1H), 7.48 (d, J = 2.0Hz, 1H), 7.43 (d, J = 9Hz, 2H), 7.24(dd,J=8.5,2.0Hz,1H),7.03(d,J=9.0Hz,2H),6.81(d,J=9.0Hz,2H),5.36(d,J=14.0Hz,1H),5.27(d,J=14.0Hz, 1H),4.38(t,J=5.0Hz,1H),4.31-4.27(m,1H),3.95(dd,J=8.5,6.5Hz,1H),3.88–3.83(m,2H),3.53(dd,J=9.5,6.5Hz ,1H),3.38(bs,4H),3.26(bs,4H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=7.0Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 162.5, 152.8, 152.7, 152.0, 136.3, 133.9, 133.3, 131.5, 130.1, 129.6, 127.2, 123.6, 116.8, 115.4, 107.4, 74.8, 67.9, 67.6, 56.6, 52.7, 36.5, 31.0, 28.5, 19.2, 10.8. HRMS (ESI) calcd for C₃₄H₃ₐCl₂N₆O₄: 706.2424; found: 706.2425. HPLC purity: 95.9%, tR = 10.8 min.

[0366] 4-(4-(4-(((2S,4R)-2-((2H-tetrazol-2-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1-sec-butyl-1H-1,2,4-triazol-5(4H)-one (Compound 49). 1H NMR (500MHz, CDCl3, δH): 8.79 (s, 1H), 7.99 (s, 1H), 7.61 (s, 1H), 7.58 (d, J = 8.5Hz, 1H), 7.48 (d, J = 2.0Hz, 1H), 7.4 2-7.40(m,2H),7.28-7.26(m,1H),7.01(d,J=9.0Hz,2H),6.79(d,J=9.0Hz,2H),5.03(d,J=4.0Hz,2H),4.34(t,J=9 .0Hz,1H),4.29-4.25(m,1H),3.91(dd,J=8.5,7.0Hz,1H),3.77–3.74(m,2H),3.56(dd,J=10.0,5.5Hz,1H),3.35( bs,4H),3.23(d,J=5.0Hz,4H),1.88–1.82(m,1H),1.73–1.67(m,1H),1.37(d,J=7.0Hz,3H),0.88(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 162.9, 151.9, 151.7, 151.5, 136.3, 133.9, 133.9, 131.5, 131.0, 129.6, 126.9, 123.4, 116.6, 115.4, 107.4, 74.8, 67.9, 67.6, 56.6, 52.7, 36.5, 31.2, 28.5, 19.2, 10.7. HRMS (ESI) calcd for C₃₄H₃ₐCl₂N₆O₄: 706.2424; found: 706.2425. HPLC purity: 94.6%, tR = 10.4 min.

[0367] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((3-methyl-1H-1,2,4-triazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 50). 1H NMR (500MHz, CDCl3, δH): 8.08 (bs, 1H), 7.62 (s, 1H), 7.60 (s, 1H), 7.46 (d, J = 2.0Hz, 1H), 7.43 (d, J = 9.0Hz, 2H), 7.26 (dd, J = 8.0, 2.0Hz, 1H), 7.03(d,J=9.5Hz,2H),6.98-6.97(m,2H),6.80(d,J=9.0Hz,2H),4.75(d,J=15.0Hz,1H),4.65(d,J=15.0Hz,1H),4.36(dt,J=6.5,5.0,2.5Hz, 1H), 4.29 (dd, J=8.5, 6.5Hz, 1H), 3.90 (dd, J=8.5, 6.5Hz, 1H), 3.83 (dd, J=8.5, 4.5Hz, 1H), 3.77 (dd, J=9.5, 5.0Hz, 1H), 3.43 (dd, J=9.5, 6.5 Hz,1H),3.39(m,4H),3.25(t,J=5.0Hz,4H),2.35(s,3H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.1, 145.3, 136.0, 134.3, 134.0, 133.2, 131.5, 129.7, 127.3, 123.7, 123.6, 118.7, 116.8, 115.3, 107.7, 74.7, 67.7, 67.5, 53.3, 52.7, 50.9, 49.2, 28.5, 19.3, 13.9, 10.8. HRMS (ESI) calcd for C36H40Cl2N8O4: 719.2628; found: 719.2614. HPLC purity: 97.4%, tR = 9.2 min.

[0368] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((3-(trifluoromethyl)-1H-1,2,4-triazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 51). 1H NMR (500MHz, CDCl3, δH): 8.32 (s, 1H), 7.62 (bs, 1H), 7.60 (s, 1H), 7.49 (d, J = 2.0Hz, 1H), 7.43 (d, J = 8.5Hz, 2H), 7.29 (dd, J = 8.5, 2.0Hz ,1H),7.03(d,J=9.0Hz,2H),6.94-6.91(m,2H),6.79(d,J=8.0Hz,2H),4.87(d,J=15.0Hz,1H),4.80(d,J=15.0Hz,1H),4.37(t,J=5.0H z,1H),4.29(dd,J=8.5,6.5,2.0Hz,1H),3.93(dd,J=8.5,7.0Hz,1H),3.82(dd,J=8.5,5.0Hz,1H),3.80(dd,J=10.0,4.5Hz,1H),3.49( dd,J=9.5,6.0Hz,1H),3.37(bs,4H),3.25(bs,4H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=7.0Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl₃, δC): 162.6, 152.1, 146.6, 136.3, 134.0, 133.6, 133.2, 131.5, 129.7, 127.4, 123.6, 116.7, 115.2, 107.2, 74.7, 67.4, 67.4, 54.1, 52.7, 49.2, 36.5, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C₃₆H₃ₐ₃N₈O₄: 773.2345; found: 773.2357. HPLC purity: 95.5%, tR = 12.2 min.

[0369] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((5-methyl-1H-tetrazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 52). 1H NMR (500MHz, CDCl3, δH): 7.61 (s, 1H), 7.59 (d, J = 8.5Hz, 1H), 7.48 (d, J = 2.0Hz, 1H), 7.43 (d, J = 9Hz, 2H), 7.26 (dd, J = 8.0 ,2.5Hz,1H),7.03(d,J=9.0Hz,2H),6.80(d,J=8.5Hz,2H),5.27(d,J=14.5Hz,1H),5.15(d,J=14.5Hz,1H),4.39(dt,J=6. 5,5.0,1.5Hz,1H),4.31-4.27(m,1H),3.95-3.87(m,2H),3.80(dd,J=9.5,4.7Hz,1H),3.76(dd,J=9.0,7.0Hz,1H),3.38 (bs,4H),3.27(bs,4H),2.47(s,3H),1.89–1.83(m,1H),1.74–1.69(m,1H),1.39(d,J=7.0Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 171.2, 162.9, 152.1, 133.9, 133.5, 131.5, 129.6, 127.2, 123.6, 117.7, 115.4, 107.5, 74.7, 67.9, 67.5, 60.4, 56.4, 52.7, 28.5, 21.1, 19.3, 14.2, 10.8. HRMS (ESI) calcd for C35H39Cl2N9O4: 720.2580; found: 720.2571. HPLC purity: 94.8%, tR = 11.3 min.

[0370] 1-sec-butyl-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((5-phenyl-1H-tetrazol-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 53). 1H NMR (500MHz, CDCl3, δH): 8.15-8.13 (m, 2H), 8.02 (bs, 1H), 7.63-7.59 (m, 2H), 7.50 (d, J = 2.5Hz, 1H), 7.48-7.46 (m, 2H), 7.44-7.41 (m, 2H),7.26(dd,J=8.5,2.0Hz,1H),7.03(d,J=9.0Hz,2H),6.67(d,J=9.0Hz,2H),5.35(d,J=14.5Hz,1H),5.24(d,J=14.5Hz,1H),4.41-4 .34(m,1H),4.29(dd,J=9.0,6.5Hz,1H),3.94(dd,J=9.0,6.5Hz,1H),3.88(dd,J=8.5,4.5Hz,1H),3.81(dd,J=9.5,4.5Hz,1H),3.44(d d,J=9.5,6.5Hz,1H),3.35(bs,4H),3.20(bs,4H),1.89–1.83(m,1H),1.75–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 196.5, 133.9, 133.4, 131.5, 129.7, 128.9, 127.0, 123.6, 118.4, 115.3, 107.5, 57.4, 56.7, 52.7, 32.2, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C₄₀H₄₁₂N₆O₄: 782.2737; found: 782.2749. HPLC purity: 95.7%, tR = 13.7 min.

[0371] 4-(4-(4-(4-(((2S,4R)-2-(bromomethyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1-sec-butyl-1H-1,2,4-triazol-5(4H)-one (Compound 57). 1H NMR (500MHz, CDCl3, δH): 7.65 (d, J=8.5Hz, 1H), 7.61 (s, 1H), 7.43-7.42 (m, 3H), 7.27 (dd, J=8.5, 2.0Hz, 1H), 7.03 (d, J= 9.0Hz,2H),6.95(bs,2H),6.90(d,J=9.0Hz,2H),4.46-4.42(m,1H),4.31-4.27(m,1H),4.22(dd,J=9.5,5.0Hz,1H),4.1 4(dd,J=8.5,5.5Hz,1H),4.09(dd,J=8.5,6.5Hz,1H),4.02(dd,J=8.5,7.0Hz,1H),3.96(d,J=11.5,1H),3.86(d,J=11.5 Hz,1H),3.37(bs,4H),3.25(bs,4H),1.89–1.84(m,1H),1.75–1.69(m,1H),1.39(d,J=7.0Hz,3H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.1, 135.7, 134.7, 133.9, 133.0, 131.3, 120.0, 127.0, 123.6, 118.5, 116.7, 115.5, 107.8, 74.8, 68.6, 68.4, 52.7, 50.7, 49.3, 35.4, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C33H36BrCl2N5O4: 716.1406; found: 716.1422. HPLC purity: 95.7%, tR = 13.3 min.

[0372] 1-sec-butyl-4-(4-(4-(4-(((2R,4R)-2-(2,4-dichlorophenyl)-2-ethyl-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 59). NMR (500MHz, CDCl3, δH): 7.61 (s, 1H), 7.57 (d, J = 8.0Hz, 1H), 7.43 (d, J = 9.0Hz, 2H), 7.40 (d, J = 2.0Hz, 1H), 7.23 (dd, J = 8. 5,2.0Hz,1H),7.03(d,J=9.0Hz,2H),6.95(bs,2H),6.89(d,J=9.0Hz,2H),4.33-4.27(m,2H),4.12(dd,J=9.5,5.0Hz,1H) ,3.98(dd,J=8.5,4.5Hz,1H),3.96(dd,J=9.5,6.5Hz,1H),3.86(dd,J=8.5,7.0Hz,1H),3.37(bs,4H),3.24(bs,4H),2.18 –2.09(m,2H),1.89–1.83(m,1H),1.75–1.69(m,1H),1.39(d,J=7.0Hz,3H),0.90(t,J=7.5Hz,3H),0.89(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 152.1, 137.4, 134.5, 133.9, 132.8, 131.2, 129.8, 126.8, 123.6, 118.5, 116.7, 115.2, 111.2, 73.7, 69.3, 67.1, 52.7, 50.7, 49.3, 30.7, 28.5, 19.3, 10.8, 7.7. HRMS (ESI) calcd for C34H39Cl2N5O4: 652.2457; found: 652.2426. HPLC purity: 95.0%, tR = 13.4 min.

[0373] 4-(4-(4-(4-(((2R,4R)-2-benzyl-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1-sec-butyl-1H-1,2,4-triazol-5(4H)-one (Compound 60). 1H NMR (500MHz, CDCl3, δH): 7.62 (s, 1H), 7.46-7.42 (m, 4H), 7.43-7.42 (m, 3H), 7.22 (bs, 5H), 7.16 (dd, J=8. 5,2.0Hz,1H),7.03(d,J=9.0Hz,2H),6.93(bs,2H),6.73(d,J=9.0Hz,2H),4.32-4.26(m,2H),3.79-3.77(m ,2H),3.68(dd,J=9.5,5.0Hz,1H),3.45(d,J=14.0,1H),3.37(bs,4H),3.34(d,J=14.0Hz,1H),3.32-3.28 (m,1H),3.25(bs,4H),1.89–1.84(m,1H),1.75–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.90(t,J=7.2Hz,3H). C NMR (125 MHz, CDCl, δC): 152.1, 137.7, 135.2, 134.6, 133.9, 132.9, 131.3, 131.1, 129.5, 127.7, 123.6, 116.7, 115.3, 110.1, 74.0, 68.4, 67.2, 52.7, 49.2, 43.7, 36.5, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C39H41Cl2N5O4: 714.2614; found: 714.2590. HPLC purity: 95.1%, tR = 14.8 min.

[0374] 1-sec-butyl-4-(4-(4-(4-(((2R,4R)-2-(cyclopentylmethyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 61). 1HNMR (500MHz, CDCl3, δH): 7.62 (s, 1H), 7.58 (d, J = 8.0Hz, 1H), 7.43 (d, J = 9.0Hz, 2H), 7.39 (d, J = 2.5Hz, 1H), 7.22 (dd, J =8.5,2.0Hz,1H),7.03(d,J=9.0Hz,2H),6.95(bs,2H),6.89(d,J=9.0Hz,2H),4.31-4.27(m,2H),4.12(dd,J=9.2,4.5Hz, 1H),3.99-3.95(m,2H),3.84(dd,J=8.0,7.0Hz,1H),3.37(bs,4H),3.24(bs,4H),2.24–2.16(m,2H),1.90–1.81(m,2H),1 .75–1.69(m,3H),1.57–1.53(m,2H),1.44–1.41(m,2H),1.39(d,J=7.0Hz,3H),1.14–1.06(m,2H),0.90(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.1, 137.8, 134.5, 133.9, 133.0, 131.2, 129.8, 126.6, 123.6, 118.5, 116.7, 115.5, 111.1, 73.6, 69.1, 66.8, 52.7, 50.7, 49.3, 43.6, 35.4, 33.6, 33.5, 28.5, 25.1, 25.0, 19.3, 10.8. HRMS (ESI) calcd for C38H45Cl2N5O4: 706.2927; found: 706.2915. HPLC purity: 95.4%, tR = 15.2 min.

[0375] 1-sec-butyl-4-(4-(4-(4-(((4R)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (mixture of trans and cis) (Compound 62). Trans compound: 1H NMR (500MHz, CDCl3, δH): 7.62 (s, 1H), 7.57 (d, J = 8.5Hz, 1H), 7.43 (d, J = 8.5Hz, 2H), 7.40 (t, J = 2.0Hz, 1H), 7.28 (t, J=2.0Hz,1H),7.27(bs,1H),7.03(d,J=9.0Hz,2H),6.95(bs,1H),6.92-6.88(m,2H),6.27(s,1H),4.66-4.60(m,1H ),4.34(dd,J=8.2,6.7Hz,1H),4.31-4.27(m,1H),4.18-4.15(m,1H),4.14-4.07(m,1H),4.03(dd,J=8.5,6.5Hz,1H ),3.37(bs,4H),3.25(bs,4H),1.90–1.84(m,1H),1.75–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.91(t,J=7.2Hz,3H). 13C NMR (125MHz, CDCl3, δC):152.1,135.8,135.7,134.4,134.3,133.9,129.6,128.8,127.4,127.2 ,123.6,118.5,116.7,115.5,100.7,76.5,74.7,68.7,68.6,67.9,52.7,50.7,28.5,19.3,10.8.Cis compound: 1H NMR (500 MHz, CDCl3, δH): 7.62 (s, 1H), 7.57 (d, J = 8.5 Hz, 1H), 7.43 (d, J = 8.5 Hz, 2H), 7.40 (t, J = 2.0 Hz, 1H), 7.29 (d, J = 2.0 Hz, 1H), 7.27 (bs, 1H), 7.03 (d, J = 9.0 Hz, 2H), 6.95 (bs, 1H), 6.92-6.88 (m, 2H), 6.16 (s, 1H), 4.66-4.60 (m, 1H ),4.31-4.27(m,1H),4.21(dd,J=8.2,6.7Hz,1H),4.18-4.15(m,1H),4.14-4.07(m,1H),4.03(dd,J=8.5,6.5Hz,1H ),3.37(bs,4H),3.25(bs,4H),1.90–1.84(m,1H),1.75–1.69(m,1H),1.39(d,J=6.5Hz,3H),0.91(t,J=7.2Hz,3H). C NMR (125 MHz, CDCl₃, δC): 152.1, 135.8, 135.7, 134.4, 134.3, 133.9, 129.6, 128.8, 127.4, 127.2, 123.6, 118.5, 116.7, 115.5, 100.7, 76.5, 74.7, 68.7, 68.6, 67.9, 52.7, 50.7, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C₃₂H₃₅Cl₂N₅O₄: 624.2144; found: 624.2114. HPLC purity: 95.9%, tR = 13.7 min.

[0376] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-(morpholinomethyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 63). 1HNMR (500MHz, CDCl3, δH): 7.63 (s, 1H), 7.61 (d, J = 9.0Hz, 1H), 7.45 (d, J = 8.5Hz, 2H), 7.41 (d, J = 1.5Hz, 1H), 7.25 ( dd,J=8,1.5Hz,1H),7.06(d,J=9.0Hz,2H),6.98(d,J=9.0Hz,2H),6.92(d,J=8.5Hz,2H),4.36-4.41(m,1H),4.28-3. 34(m,1H),4.14-4.18(m,1H),4.05-4.10(m,2H),3.89(t,J=7.0Hz,1H),3.57(t,J=4Hz,4H),3.36-3.40(m,4H),3.2 6(t,J=5Hz,4H),2.59(t,J=4Hz,4H),1.95-1.85(m,1H),1.79-1.70(m,1H),1.42(d,J=7Hz,3H),0.93(t,J=7Hz,3H). C NMR (125 MHz, CDCl, δC): 152.6, 151.1, 146.5, 135.3, 133.6, 131.6, 130.6, 127.2, 126.6, 124.3, 119.3, 117.5, 116.2, 75.2, 68.4, 56.0, 53.9, 51.9, 50.5, 31.1, 29.9, 20.8, 12.4. HRMS (ESI) calcd for C37H44Cl2N6O5: 723.2828; found: 723.2826. HPLC purity: 96.2%, tR = 5.0 min.

[0377] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((dimethylamino)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 64). 1H NMR (500 MHz, CDCl3, δH): 8.41 (s, 1H), 7.74-7-75 (m, 2H), 7.59 (d, J = 9 Hz, 2H), 7.19 (d, J = 9 Hz, 2H), 7.06 (d, J = 9 Hz, 2H), 7.00 (d, J = 9 Hz, 2H), 4.53-4.48 (m, 1H), 4.23-4 .18(m,3H),4.10-4.03(m,4H),3.43-3.38(m,4H),3.27-3.24(m,4H),2.59(s,6H), 1.84-1.78(m,1H),1.76-1.70(m,1H),1.38(d,J=6.5Hz,3H),0.88(t,J=6.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.8, 150.2, 146.5, 135.5, 134.2, 130.2, 127.0, 126.6, 124.3, 119.3, 117.5, 116.2, 77.2, 72.2, 68.4, 60.7, 50.4, 47.6, 29.9, 20.8, 12.4. HRMS (ESI) calculated for C36H40Cl2N8O4: 681.2723; found: 681.2714. HPLC purity: 97.1%, tR = 4.7 min.

[0378] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-((4-methylpiperazin-1-yl)methyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 65). 1H NMR (500MHz, CDCl3, δH): 8.03(s,1H),7.62(s,1H),7.58(d,J=8.5Hz,1H),7.43(d,J=7.5Hz,2H),7.38(s,1H),7.23(d,J =8.5Hz,1H),7.04(d,J=8Hz,2H),6.96(d,J=8Hz,2H),6.90(d,J=8Hz,2H),4.36-4.34(m,1H),4.32-4.28(m,1H),4.15-4 .12(m,1H),4.06-4.02(m,2H),3.87(t,J=7.5Hz,1H),3.60-3.58(m,4H),3.37-3.36(m,4H),3.25-3.22(m,4H),3.02(s, 2H),2.70-2.67(m,4H),2.33(s,3H),1.89-1.84(m,1H),1.75-1.69(m,1H),1.41(d,J=6.5Hz,3H),0.91(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 153.5, 152.6, 151.1, 146.5, 137.4, 135.2, 133.7, 131.6, 130.6, 127.2, 126.6, 124.3, 119.3, 117.5, 116.2, 75.2, 70.1, 68.2, 63.6, 55.8, 54.3, 53.9, 51.9, 50.5, 46.7, 29.9, 20.8, 12.4. HRMS (ESI) calculated for C38H47Cl2N7O4: 736.7291; found: 736.7252. HPLC purity: 96.3%, tR = 7.5 min.

[0379] 1-(sec-butyl)-4-(4-(4-(4-(((2S,4R)-2-(2,4-dichlorophenyl)-2-(piperazin-1-ylmethyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)phenyl)-1H-1,2,4-triazol-5(4H)-one (Compound 66). 1H NMR (500MHz, CDCl3, δH): 7.62 (s, 1H), 7.60 (d, J = 8.5Hz, 2H), 7.44 (d, J = 9Hz, 2H), 7.39 (d, J = 1.5Hz, 1H), 7.24 (dd, J = 8.5, 2. 0Hz,1H),7.04(d,J=8.5Hz,2H),6.96(d,J=9Hz,2H),6.90(d,J=9Hz,2H),4.39-4.34(m,1H),4.32-4.27(m,1H),4.16-4.13(m ,1H),4.08-4.03(m,2H),3.89(t,J=7.5Hz,1H),3.56(t,J=4.5Hz,4H),3.39-3.35(m,4H),3.26-3.23(m,4H),2.99(d,J=3Hz, 3H), 2.57 (t, J = 4.5Hz, 4H), 2.19 (s, 1H), 1.90-1.85 (m, 1H), 1.76-1.70 (m, 1H), 1.41 (d, J = 6.5Hz, 3H), 0.92 (t, J = 7.5Hz, 3H). C NMR (125 MHz, CDCl, δC): 153.5, 151.1, 146.5, 137.4, 135.2, 133.6, 131.6, 130.6, 127.2, 126.6, 124.3, 119.3, 117.5, 116.2, 75.2, 70.0, 68.4, 68.2, 64.4, 56.0, 53.9, 51.9, 50.5, 29.9, 20.8, 12.4. HRMS (ESI) calculated for C37H45Cl2N7O4: 722.7049; found: 722.7043. HPLC purity: 95.1%, tR = 5.1 min.

[0380] 4-(4-(4-(6-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)piperazin-1-yl)phenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 100). 1H NMR (500MHz, CDCl3, δH): 8.12 (s, 1H), 7.76 (s, 1H), 7.74 (d, J = 2.0Hz, 1H), 7.51 (s, 1H), 7.45 (d, J = 8.5Hz, 1H) 7.35-7 .29(m,4H),7.12(dd,J=8.0,2.0Hz,1H),6.97(d,J=8.5Hz,2H),6.67(d,J=9.0Hz,1H),4.71(q,J=14.5Hz,2H),4.29-4 .25(m,1H),4.20-4.15(m,1H),4.13-4.09(m,1H),4.04-4.00(m,1H),3.81(t,J=7.0Hz,1H),3.65-3.61(m,1H),3.31 -3.29(m,4H),3.20-3.15(m,4H),1.78-1.71(m,1H),1.65-1.56(m,1H),1.29(d,J=7.0Hz,3H),0.79(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 158.3, 152.0, 151.3, 136.0, 134.9, 134.2, 133.8, 133.2, 131.4, 130.4, 129.7, 127.2, 123.6, 117.2, 111.3, 107.6, 74.8, 67.2, 64.9, 53.8, 52.8, 50.6, 49.3, 41.0, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C₃₄H₃ₐCl₂N₆O₄: 706.2424; found: 706.2422. HPLC purity: 98.2%, tR = 10.7 min.

[0381] 4-(4-(4-(5-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)piperazin-1-yl)phenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 101). 1H NMR (500MHz, CDCl3, δH): 8.09 (s, 1H), 7.78 (s, 1H), 7.76 (d, J = 3.0Hz, 1H), 7.51 (s, 1H), 7.47 (d, J = 8.5Hz, 1H), 7.36 (d, J = 2.0H z,1H),7.32(d,J=9.0Hz,2H),7.16(dd,J=8.5,2Hz,1H),7.06(dd,J=8.5,2Hz,1H),6.92(d,J=9.0Hz,2H),6.61(d,J=9.0Hz,1H ),4.71(q,J=14.5Hz,2H),4.26-4.22(m,1H),4.19-4.15(m,1H),3.82-3.79(m,1H),3.71-3.65(m,2H),3.51(t,J=5.5Hz,4H), 3.45-3.42(m,1H),3.23(t,J=5.5Hz,4H),1.78-1.71(m,1H),1.63-1.57(m,1H),1.28(d,J=6.5Hz,3H),0.79(t,J=6.5Hz,3H). C NMR (125 MHz, CDCl, δC): 152.0, 151.4, 150.5, 147.9, 136.1, 134.0, 133.1, 131.5, 129.6, 127.3, 126.0, 123.6, 116.7, 107.7, 74.6, 68.5, 67.2, 53.6, 52.7, 48.9, 46.3, 41.0, 28.4, 19.28, 10.8. HRMS (ESI) calcd for C₃₄H₃ₐCl₂N₆O₄: 706.2424; found: 706.2415. HPLC purity: 95.4%, tR = 6.3 min.

[0382] 4-(6-(4-(4-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)pyridin-3-yl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 102). 1H NMR (500MHz, CDCl3, δH): 8.27-8.26 (m, 2H), 7.91 (s, 1H), 7.75 (dd, J = 8.5Hz, 4Hz, 1H), 8.60-8.5 7(m,2H),7.48(d,J=4.0Hz,1H),7.28(m,1H),6.98-6.96(m,2H),6.82-6.76(m,3H),4.85(q,J=1 4.5Hz,2H),4.39-4.26(m,2H),3.94-3.90(m,1H),3.82-3.75(m,7H),3.50-3.46(m,1H),3.21-3 .18(m,4H),1.90-1.81(m,1H),1.77-1.68(m,1H),1.41(d,J=7.0Hz,3H),0.91(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 158.3, 152.1, 142.3, 136.1, 134.1, 133.8, 133.1, 133.0, 131.4, 129.6, 127.3, 121.3, 118.7, 115.3, 107.6, 107.1, 74.7, 67.6, 53.7, 52.8, 50.6, 45.3, 41.0, 28.5, 19.3, 10.8. HRMS (ESI) calcd for C₃₄H₃ₐCl₂N₆O₄: 706.2424; found: 706.2417. HPLC purity: 95.6%, tR = 10.5 min.

[0383] 4-(5-(4-(4-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)pyridin-2-yl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 103). 1H NMR (500MHz, CDCl3, δH): 8.35 (s, 1H), 8.27-8.20 (m, 2H), 8.10 (d, J=4.0Hz, 1H), 7.80-7.95 (brs, 1H), 7.60 (dd, J= 12.0Hz,1H),7.48(d,J=4.0Hz,1H),7.42(dd,J=8.0,2.2Hz,1H),7.28(m,1),6.99(d,J=8.0Hz,2H),6.83(d,J=8.0H z,2H),4.84(q,J=14.5Hz,2H),4.40-4.25(m,2H),3.94-3.90(m,1H),3.84-3.78(m,2H),3.50-3.46(m,1H),3.41-3 .35(m,4H),3.28-3.26(m,4H),1.91-1.84(m,1H),1.77-1.67(m,1H),1.41(d,J=7.0Hz,3H),0.90(t,J=7.0Hz,3H). C NMR (125 MHz, CDCl, δC): 151.1, 145.7, 140.2, 136.1, 134.0, 133.1, 132.5, 131.5, 129.6, 127.3, 125.7, 118.7, 115.3, 113.7, 107.6, 74.7, 67.6, 53.6, 52.5, 50.7, 48.9, 28.4, 19.3, 10.8. HRMS (ESI) calcd for C₃₄H₃ₐCl₂N₆O₄: 706.2424; found: 706.2420. HPLC purity: 98.2%, tR = 10.6 min.

[0384] 4-(4-(4-(4-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)phenyl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 104). 1H NMR (500MHz, CDCl3, δH): 8.12 (s, 1H), 7.79 (s, 1H), 7.67 (d, J = 8.5Hz, 1H), 7.53 (s, 1H), 7.47 (d, J = 8.5Hz, 1H), 7.36(d,J=2.0Hz,1H),7.29-7.26(m,1H),7.20-7.13(m,2H),6.95(t,J=9.0Hz,2H),6.71(d,J=9.0Hz,2H),4.73 (q,J=14.5Hz,2H),4.28-4.23(m,1H),4.19-4.15(m,1H),3.80(t,J=7.0Hz,1H),3.72-3.66(m,2H),3.40-3.36 (m,1H),3.26-3.14(m,8H),1.77-1.69(m,1H),1.64-1.56(m,1H),1.28(d,J=7.5Hz,3H),0.78(t,J=7.5Hz,3H). C NMR (125 MHz, CDCl, δC): 156.4, 154.4, 151.7, 136.1, 134.0, 133.3, 133.2, 131.5, 130.1, 129.6, 128.1, 127.3, 119.5, 117.9, 115.4, 111.0, 110.8, 107.7, 74.7, 73.7, 67.7, 67.4, 53.7, 52.9, 41.0, 28.4, 19.3, 10.8. HRMS (ESI) calcd for C35H37Cl2FN8O4: 723.2377; found: 723.2365. HPLC purity: 99.0%, tR = 10.3 min.

[0385] 4-(4-(4-(6-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-3-yl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 105). 1H NMR (500MHz, CDCl3, δH): 8.13 (s, 1H), 7.76-7.73 (m, 2H), 7.61 (d, J = 0.5Hz, 1H), 7.49 (d, J = 8.0Hz, 1H) 7.34-7 .31(m,3H),7.18-7.15(m,1H),7.13(dd,J=8.5Hz,2Hz,1H),6.95(t,J=9.0Hz,1H),6.60(d,J=9.0Hz,1H),4.7 1(q,J=14.5Hz,2H),4.29-4.26(m,1H),4.18-4.09(m,2H),4.03-3.99(m,1H),3.83-3.79(m,1H),3.64-3.61( m,1H),3.32-3.14(m,8H),1.77-1.70(m,1H),1.63-1.57(m,1H),1.27(d,J=7.0Hz,3H),0.78(t,J=7.0Hz,3H). 13C NMR (125MHz, CDCl3, δC): 156.3, 154.3, 151.5, 145.4, 136.1, 135.8, 134.1, 133.3, 133.0, 132.3, 131.3, 130.0, 129.6, 129.5, 128.5, 127. 9,127.1,119.5,117.8,111.1,110.9,110.7,107.5,74.77,73.57,69 .3,67.1,66.7,64.7,61.6,53.8,52.7,50.2,40.7,28.3,19.2,10.7. HRMS (ESI) calculated for C34H36Cl2FN9O4: 724.2330; found: 724.2328. HPLC purity: 95.3%, tR = 11.2 min.

[0386] 4-(4-(4-(5-(((2S,4R)-2-((1H-1,2,4-triazol-1-yl)methyl)-2-(2,4-dichlorophenyl)-1,3-dioxolan-4-yl)methoxy)pyridin-2-yl)piperazin-1-yl)-3-fluorophenyl)-2-(sec-butyl)-2,4-dihydro-3H-1,2,4-triazol-3-one (Compound 106). 1H NMR (500MHz, CDCl3, δH): 8.10 (s, 1H), 7.78 (s, 1H), 7.77 (d, J = 3.0Hz, 1H), 7.55 (s, 1H), 7.47 (d, J = 8.0Hz, 1H), 7.36 (d, J = 2.0H z,1H),7.29(dd,J=8.0,2.5Hz,1H),7.16-7.13(m,2H),7.06(dd,J=9.0,3.0Hz,1H),6.93(t,J=9.0Hz,1H),6.60(d,J=9.0Hz,1 H),4.72(q,J=14.5Hz,2H),4.26-4.22(m,1H),4.19-4.14(m,1H),3.83-3.79(m,1H),3.71-3.65(m,2H),3.52(t,J=5.5Hz,4H) ,3.45-3.42(m,1H),3.11(t,J=5.5Hz,4H),1.79-1.70(m,1H),1.64-1.56(m,1H),1.28(d,J=7.0Hz,3H),0.78(t,J=7.0Hz,3H). 13C NMR (125 MHz, CDCl3, δC): 156.4, 154.4, 151.6, 151.4, 147.9, 139.3, 136.1, 133.9, 133.3, 133.1, 131.5, 129.6, 127.3, 119.5, 117.9, 111.0, 110.8, 108.5, 107.7, 74.6, 68.5, 67.2, 53.6, 52.8, 50.3, 46.5, 41.0, 28.4, 19.3, 10.8. HRMS (ESI) calculated for C34H36Cl2FN9O4: 724.2330; found 724.2326. HPLC purity: 96.0%, tR=7.2 min.

[0387] Water solubility test. In a 1.5mL microcentrifuge tube, 2 μl of 10mM compound stock solution in DMSO was added to 1mL 0.001N HCl aqueous buffer to obtain a saturated solution. The resulting mixture was shaken at room temperature for 24h at 600rpm. The tube was then centrifuged for 5min at 13,000rpm to remove undissolved compounds. 100 μl of supernatant was injected into an Agilent 6120LC / MS for analysis. Mass spectral data were collected in SIM / Scan mode. The ions of compound ExactMass+1 and ExactMass+23 were monitored for peak area integration using selective ion monitoring (SIM). Solubility was calculated based on the concentration standard curve of each compound.

[0388] HUVEC culture and proliferation assay. HUVEC (Lonza) were grown in EGM-2 bullet kit medium (Lonza) and used at passage 8 or lower. H thymidine incorporation assay was performed as previously described. Briefly, cells were seeded at a concentration of 2000 cells / well in 96-well plates and allowed to settle overnight. Drugs were added to each well in triplicate. After 24 hours, cells were treated with 1 μCi of [ 3 [H] Thymidine treatment was continued for 6 h. Cells were then harvested and transferred to a filter mat. Scintillation was counted and IC50 was determined using Prism software (version 6.0).

[0389] CYP3A4 enzyme assay. CYP3A4 enzyme activity was tested using the Vivid™ CYP3A4 Green Screening Kit (ThermoFisher, #P2857) using the manufacturer's protocol. Briefly, CYP3A4 baculosomes and drug were incubated at 37°C for 10 min. substrate, and fluorescence (excitation / emission (ex / em): 485 / 520): every 1 min for 60 min.

[0390] Tube formation assay. 24-well plates were coated with 250 μl of Matrigel (BD, #CB-40234C) / well. 70,000 HUVECs were added to a plate with 500 μl of culture medium containing different drugs. After 24 h, 2 μM calcein AM was added and incubated for 15 min. After replacing the culture medium with new medium, the tube network was photographed using a fluorescence microscope. Total tube length was calculated using ImageJ.

[0391] Filipin staining. 2000 HUVECs were plated on a chamber slide with 1 ml of culture medium and allowed to stand overnight. Cells were treated with 0.1 μM of drug or DMSO for 14 hours. Cells were fixed with 4% paraformaldehyde for 15 min. After washing, cells were incubated in the dark with 50 μg / ml of 500 μl of filipin solution for 1 hour. The cells were then washed twice with PBS, fixed, and covered with a coverslip. Images were taken using a confocal microscope at 360 nm / 460 nm.

[0392] Western blotting. HUVEC cells were treated with different concentrations of compound 48 for 24 hours. The cells were then lysed using RIPA buffer, and protein concentrations were measured and normalized. After electrophoresis and transfer to a 0.45 μm nitrocellulose membrane, the membrane was bolted with 5% BSA for 1 hour and then incubated at 4°C overnight. Secondary antibodies were applied to each membrane for 1 hour. After adding chemiluminescent substrate, the blots were imaged using a Syngene PXi imaging system. The following antibodies were used in the assay: AMPKα (1:1,000, Cell Signaling, #2532s), phospho-AMPKα (1:1,000, Cell Signaling, #2535s), ACC (1:1,000, Cell Signaling, #3662s), phospho-ACC (1:1,000, Cell Signaling, #3661s), mTOR (1:1,000, Cell Signaling, #2972S), phospho-mTOR (1:1,000, Cell Signaling, #9234s), p70S6 kinase (1:1,000, Senta Cruz, #sc-8418), phospho-p70S6 kinase (1:1,000, Cell Signaling, #3662s), and anti-rabbit IgG (1:10,000, GE Lifesciences, #NA934V).

[0393] Immunofluorescence assay. HUVEC (5000) were seeded in 4-well glass slides (Nunc TM Lab-Tek TMII) and cultured overnight. The cells were treated with 2 μM drug or DMSO for 24 hours. The culture medium was then aspirated and washed three times with PBS. The cells were fixed with 4% paraformaldehyde for 15 minutes, washed three times with PBS, permeabilized with 0.1% Triton X and washed again. 2% BSA blocking buffer was applied to the cells for 1 hour. Primary antibodies for VEGFR2 (1:50, Cell Signaling, #2479) and GM130 (1:1000, BD, #610823) were incubated at 4°C overnight. After washing three times with 0.5% BSA in PBS, secondary antibody solution (1:1000 goat anti-rabbit Alexa Fluor 488, Invitrogen, #A11008 and 1:1000 goat anti-mouse Alexa Fluor 594, Invitrogen, A11005) was applied in the dark at room temperature for 1 hour. After washing 3 times, the slides were incubated in 0.1 μg / mL DAPI solution for 5 min and rinsed 2 times with PBS. Coverslips were fixed with Immunount (Fisher) and sealed with nail polish. Confocal images were taken using a Zeiss LSM700 confocal microscope with a 20× lens.

[0394] Molecular docking. The binding modes of itraconazole and 17 to NPC1 (PDB: 5I31) were simulated using AutoDock Vina3. The PDB structures of itraconazole and 17 were generated using Babel, and pdbqt files for both the protein and ligand were created using the AutoDock tool. The grid box configuration was set as follows: center_x = 28.5, center_y = 51, center_z = 44.5; size_x = 24, size_y = 24, size_z = 22. PyMOL software was used to visualize the ligand-protein interaction.

[0395] Although the present disclosure has been described with reference to the above examples, it will be understood that modifications and variations are encompassed within the spirit and scope of the present disclosure. Accordingly, the present disclosure is limited only by the following claims.

Claims

1. A compound selected from: or a pharmaceutically acceptable salt thereof.

2. A pharmaceutical preparation comprising the compound according to claim 1 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

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