KR20220051816A - Thiobenzimidazole derivatives or its pharmaceutically acceptable salts and use thereof - Google Patents

Abstract

The present invention relates to a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, and a composition for preventing or treating cancer, comprising the derivative as an active ingredient. The thiobenzimidazole derivative of the present invention exhibits cell toxicity by blocking the cell cycle of a cancer cell and inducing apoptosis when administered to an individual, as the derivative inhibits tubulin polymerization by being activated in the cancer cell. Therefore, the thiobenzimidazole derivative can be used for prevention or treatment of cancer, desirably for prevention or treatment of triple-negative breast cancer.

Description

Thiobenzimidazole derivatives or its pharmaceutically acceptable salts and use thereof

The present invention relates to a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, and uses thereof.

Triple-negative breast cancer (TNBC; ER-, PR-, HER2-) patients account for 10% to 15% of all breast cancer patients, and hormone receptors (ER (estrogen receptor), PR (progesterone receptor)) and Because it lacks the HER2 protein, it does not benefit from hormone therapy or HER2-targeted therapies. Currently, standard treatment for triple-negative breast cancer is completely dependent on general cytotoxic anticancer drugs (Taxene-based or Anthracycline-based). Due to the lack of established targeted therapeutics, other subtypes do not have as diverse therapeutic strategies as for breast cancer. What is more serious is that, after surgery or chemotherapy, recurrence occurs within 2-3 years in most patients, and metastasis to other organs such as lung, liver, brain, and bone is easily induced, which affects the survival rate of patients.

The 5-year overall survival rate of patients diagnosed with stage III (stage-III) is 55% or less, and the 5-year survival rate of patients who have already progressed to cancer metastasis (advanced-stage) is 30% or less. very low Most of these patients are very serious diseases that ultimately lead to death all within a few years.

Microtubules are a major component of the cytoskeleton and are composed of tubulin heteropolymers composed of α and β subunits. Microtubules perform various cellular functions, such as intracellular transport, maintenance of polarity, intracellular signal transduction, and cell migration and proliferation. During mitosis, a spindle is formed, chromosomes are arranged in the center of the cell, and then the process of bipolar separation is performed. If the spindle does not function properly, cell division is inhibited and apoptosis occurs, and thus it is attracting attention as a target for anticancer drugs.

Drugs targeting microtubules are largely divided into two groups: drugs that stabilize microtubules and drugs that destabilize microtubules. First, microtubule stabilizers include taxane, paclitaxel (Taxol), decetaxel, etc., which prevent depolymerization of microtubules and enhance polymerization. Most microtubule stable substances bind to the taxane-binding site or the overlapping site of β-tubulin. Second, microtubule destabilizers include colchicine and vinca alkaloid, which bind to the colchicine binding site or the vinca binding site. It is the same that drugs that target these microtubules themselves are effective at lower concentrations than drugs that affect microtubule polymers, and consequently inhibit cell mitosis. Therefore, there is a need to develop a potential inhibitor of tubulin polymerization as an anticancer agent.

On the other hand, flubendazole and albendazole are tubulin polymerization inhibitors that are marketed as anthelmintic drugs, but can confirm apoptosis by cell cycle arrest, and have a killing effect on slow-growing cancer cells has been identified and is attracting attention as a therapeutic agent for various cancers. However, due to its low solubility in water, it is difficult to absorb into the body, and side effects such as an increase in liver enzyme levels when consumed in large amounts appear, so the development of derivatives with a new structure is required.

Accordingly, the present inventors confirmed that a novel thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof as a tubulin polymerization inhibitor induces apoptosis by cell cycle arrest of cancer cells. and completed the present invention.

Indian Journal of Chemistry, Vol. 52B, April 2013, pp 535-545.

An object of the present invention is to provide a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof.

An object of the present invention is to provide a method for preparing the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof.

Another object of the present invention is to provide a pharmaceutical composition for preventing or treating cancer comprising the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof as an active ingredient.

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems not mentioned will be clearly understood by those skilled in the art from the following description.

In order to solve the above problems, the present invention provides a thiobenzimidazole derivative represented by the following [Formula 1] or a racemate, isomer, solvate or pharmaceutically acceptable salt thereof:

[Formula 1]

In Formula 1,

R ₁ is -SR ₂ or -SSR ₂ ,

R ₂ may be any one selected from the group consisting of a substituted or unsubstituted cyclic alkyl group, a heterocycloalkyl group, an aryl group, a heteroaryl group, and an alkylaryl group, preferably a substituted or unsubstituted C ₃ to C It may be a ₂₀ aryl group, a substituted or unsubstituted C ₃ to C ₂₀ heteroaryl group, or a substituted or unsubstituted C ₃ to C ₂₀ alkylaryl group.

As an embodiment of the present invention, in [Formula 1], a substituted aryl group, a heteroaryl group, or an alkylaryl group is a substituted or unsubstituted alkyl group, a cyclic alkyl group, a heterocycloalkyl group, an aryl group, a heteroaryl group, may be substituted with any one or more selected from the group consisting of a halogen group, a hydroxyl group, an alkoxy group, and an amino group, preferably a substituted or unsubstituted C ₃ to C ₁₂ heterocycloalkyl group, a substituted or unsubstituted It may be substituted with a C ₃ to C ₁₂ heteroaryl group, or a halogen group.

As another embodiment of the present invention, the thiobenzimidazole derivative represented by the [Formula 1] may be any one selected from the group consisting of the following compounds:

[Formula 1-1]

[Formula 1-2]

[Formula 1-3]

[Formula 1-4]

[Formula 1-5]

[Formula 1-6]

[Formula 1-7]

[Formula 1-8]

[Formula 1-9]

[Formula 1-10]

[Formula 1-11]

[Formula 1-12]

[Formula 1-13]

[Formula 1-14]

[Formula 1-15]

[Formula 1-16]

[Formula 1-17]

[Formula 1-18]

[Formula 1-19]

[Formula 1-20]

[Formula 1-21]

및

and

[Formula 1-22]

.

.

In another embodiment of the present invention, the thiobenzimidazole derivative may inhibit tubulin polymerization.

In another embodiment of the present invention, the pharmaceutically acceptable salts of the thiobenzimidazole derivatives are hydrochloride, bromate, sulfate, phosphate, nitrate, citrate, acetate, lactate, tartrate, maleate, gluconate, and succinic acid. Salt, formate, trifluoroacetate, oxalate, fumarate, glutarate, adipate, methanesulfonate, benzenesulfonate, paratoluenesulfonate, camphorsulfonate, sodium salt, potassium salt, lithium salt , calcium salt, and may be any one or more selected from the group consisting of a magnesium salt, preferably a hydrochloride salt.

In addition, the present invention provides a method for preparing a hydrochloride salt of the thiobenzimidazole derivative comprising the following steps.

(1) preparing a suspension of the thiobenzimidazole derivative represented by the following [Formula 1];

[Formula 1]

(2) injecting hydrogen chloride (HCl) into the suspension and evaporating under reduced pressure;

(3) adding isopropyl alcohol to the product of step (2), heating and cooling; and

(4) adding isopropyl ether to the product of step (3), stirring, and then filtering.

In addition, the present invention provides a composition for preventing or treating cancer comprising the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a method for preventing or treating cancer, comprising administering the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof to a subject.

In addition, the present invention provides the use of the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the prevention or treatment of cancer.

The present invention also provides a method for diagnosing cancer, comprising administering the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof to a subject.

In addition, the present invention provides the use of the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for diagnosing cancer.

In one embodiment of the present invention, the pharmaceutical composition may be to induce apoptosis by stopping the cell cycle of cancer cells (cell cycle arrest).

In another embodiment of the present invention, the cancer is skin cancer, breast cancer, uterine cancer, esophageal cancer, stomach cancer, brain tumor, colon cancer, rectal cancer, colorectal cancer, lung cancer, ovarian cancer, cervical cancer, endometrial cancer, vulvar cancer, kidney cancer, blood It may be any one or more selected from the group consisting of cancer, pancreatic cancer, prostate cancer, testicular cancer, laryngeal cancer, head and neck cancer, thyroid cancer, liver cancer, bladder cancer, osteosarcoma, lymphoma, leukemia, thymus cancer, urethral cancer, and bronchial cancer, preferably It may be breast cancer, more preferably triple negative breast cancer.

The present invention relates to a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, and a composition for preventing or treating cancer comprising the derivative as an active ingredient, wherein the thiobenzimidazole derivative of the present invention is activated in cancer cells. It inhibits tubulin polymerization and exhibits cytotoxicity by blocking the cancer cell cycle and inducing apoptosis when administered to an individual. there is.

1 is a triple-negative breast cancer (TNBC) cell line after treatment with the thiobenzimidazole derivatives of Formulas 1-1, 1-2, 1-3 and 1-5 in MDA-MB-231 cell line; The survival rate results are shown.
2 is a HER2 positive breast cancer (HER2+ BC) cell line, JIMT-1, after treatment with the thiobenzimidazole derivatives of Formulas 1-1, 1-2, 1-3, and 1-5 cell viability results is shown.
3 is a triple negative breast cancer (TNBC) cell line MDA-MB-231 of Formulas 1-6, 1-7, 1-8, 1-10, 1-11, 1-12, 1-13 and 1-14 The results of cell viability after treatment with thiobenzimidazole derivatives are shown.
4 is thiobenzimidazole derivatives of Formulas 1-6, 1-7, 1-8, 1-10, 1-11, 1-12, 1-13 and 1-14 in JIMT-1, a HER2-positive breast cancer cell line. Shows the results of cell viability after treatment.
5 is a triple negative breast cancer (TNBC) cell line, MDA-MB-231, thiobenzimi of Formulas 1-15, 1-16, 1-17, 1-19, 1-20, 1-21, and 1-22 The results of cell viability after treatment with dazole derivatives are shown.
6 is a HER2-positive breast cancer cell line JIMT-1 treated with the thiobenzimidazole derivatives of Formulas 1-15, 1-16, 1-17, 1-19, 1-20, 1-21, and 1-22. The results of cell viability afterward are shown.
7 shows the cell viability results after treatment with the thiobenzimidazole derivatives of Formula 1-3 in HER2-positive breast cancer cell lines SKBR3, BT474, and JIMT-1.
8 shows the results of confirming the induction of apoptosis (Sub-G1 accumulation) and G2/M phase cell cycle stasis of cancer cells by the thiobenzimidazole derivative of Formula 1-3.
9 shows the results of confirming the early and late apoptosis of cancer cells by the thiobenzimidazole derivative of Formula 1-3.
10 shows the cell viability results after treatment with the thiobenzimidazole derivative of Formula 1-3 and its hydrochloride (Compound 1-23) in HER2-positive breast cancer cell lines BT474 and JIMT-1.
11 shows the cell viability results after treatment with the hydrochloride salt of a thiobenzimidazole derivative of Formula 1-3 (Compound 1-23) in triple negative breast cancer (TNBC) cell lines MDA-MB-231, BT549 and 4T1.
12 is a Western blot after treatment with the hydrochloride salt of a thiobenzimidazole derivative of Formula 1-3 (Compound 1-23) in a triple-negative breast cancer (TNBC) cell line, MDA-MB-231, and a HER2-positive breast cancer cell line, JIMT-1. (Western blot) shows the results.
13 is a hematologic cancer cell line HL-60, colon cancer cell line HCT116, non-small cell lung cancer cell lines H1299 and A549, ovarian cancer cell line SKOV3, prostate cancer cell line Du145, and liver cancer cell line HepG2 of Formula 1-3 The cell viability results are shown after treatment with the hydrochloride salt of a thiobenzimidazole derivative (Compound 1-23).

The present inventors completed the present invention by confirming the anticancer activity of a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, as a result of intensive research.

More specifically, it was confirmed that the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof is activated in cancer cells to inhibit tubulin polymerization to block the cancer cell cycle and induce apoptosis, thereby exhibiting cytotoxicity.

From the above results, the present invention can provide a thiobenzimidazole derivative represented by the following [Formula 1] or a pharmaceutically acceptable salt thereof.

[Formula 1]

In Formula 1,

R ₁ is -SR ₂ or -SSR ₂ ,

R ₂ is a substituted or unsubstituted C ₃ to C ₂₀ aryl group, a substituted or unsubstituted C ₃ to C ₂₀ heteroaryl group, and a substituted or unsubstituted C ₃ to C ₂₀ alkylaryl group from the group consisting of It may be any one selected.

In the present invention, the term “substitution” refers to a reaction in which an atom or group included in the molecule of a compound is replaced with another atom or group.

In the present invention, the term “chain” refers to a molecule having a chain structure, and the chain structure is a chemical structure in which carbon atoms are connected in a chain shape, and there are straight chain ones and branched ones.

In the present invention, the term “cyclic” refers to a structure in which both ends of the chain in the backbone of the organic compound are connected to form a ring.

In the present invention, the term “chain or cyclic alkyl group” means a monovalent linear or branched or cyclic saturated hydrocarbon residue having 1 to 20 carbon atoms and consisting only of carbon and hydrogen atoms. Examples of such alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, 2-butyl, 3-butyl, pentyl, n-hexyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like. not limited

In the present invention, the term "heterocycloalkyl group" usually refers to a saturated or unsaturated (but not aromatic) cyclohydrocarbon, which may be optionally unsubstituted, monosubstituted or polysubstituted, and in its structure At least one is selected from a heteroatom of N, O or S.

In the present invention, the term "aryl group" refers to an unsaturated aromatic ring compound having 3 to 12 carbon atoms having a single ring (eg, phenyl) or a plurality of condensed rings (eg, naphthyl). Examples of such aryl groups include, but are not limited to, phenyl, naphthyl, and the like.

In the present invention, the term “heteroaryl group” refers to a single ring or a plurality of condensed rings having at least one heteroatom of N, O or S among atoms constituting the ring. Examples of the heteroaryl group include, but are not limited to, a pyridyl group, a pyrimidinyl group, a pyrazinyl group, an oxazolyl group, a furyl group, and the like.

In the present invention, the term “alkoxy group” refers to an alkyl group (-O-R) bonded to oxygen. Examples of such an alkoxy group include, but are not limited to, a methoxy group, an ethoxy group, a propoxy group, a butoxy group, and the like.

In the present invention, the “halogen group” may be fluorine (F), chloride (Cl), bromine (Br), or iodine (I).

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As a preferred embodiment of the present invention, the compound represented by Formula 1 is preferably at least one selected from the group consisting of the following compounds:

[Formula 1-1]

[Formula 1-2]

[Formula 1-3]

[Formula 1-4]

[Formula 1-5]

[Formula 1-6]

[Formula 1-7]

[Formula 1-8]

[Formula 1-9]

[Formula 1-10]

[Formula 1-11]

[Formula 1-12]

[Formula 1-13]

[Formula 1-14]

[Formula 1-15]

[Formula 1-16]

[Formula 1-17]

[Formula 1-18]

[Formula 1-19]

[Formula 1-20]

[Formula 1-21]

및

and

[Formula 1-22]

.

.

The thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof of the present invention is activated specifically for cancer cells to inhibit tubulin polymerization and induce cell death. It can be used as a pharmaceutical composition for preventing or treating cancer comprising an acceptable salt as an active ingredient.

As used herein, the term “pharmaceutically acceptable salt” refers to a formulation of a compound that does not cause serious irritation to an organism to which the compound is administered and does not impair the biological activity and properties of the compound. The pharmaceutically acceptable salt may be prepared by reacting the compound of the present invention with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid, sulfonic acid such as methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, tartaric acid, formic acid, citric acid, acetic acid, It can be obtained by reaction with an organic carbonic acid such as trichloroacetic acid, trifluoroacetic acid, capric acid, isobutanoic acid, malonic acid, succinic acid, phthalic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid, maleic acid, salicylic acid, and the like. Further, by reacting the compound of the present invention with a base, an ammonium salt, an alkali metal salt such as sodium or potassium salt, an alkaline earth metal salt such as a calcium or magnesium salt, dicyclohexylamine, N-methyl-D-glucamine, It can also be obtained by forming salts of organic bases such as tris (hydroxymethyl) methylamine and salts of amino acids such as arginine and lysine.

In addition, the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof may include all salts, hydrates and solvates that can be prepared by conventional methods as well as pharmaceutically acceptable salts.

In addition, the present invention may provide a method for preventing, treating, and/or diagnosing cancer, comprising administering the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof to a subject.

As used herein, the term "prevention" refers to any action that inhibits or delays the occurrence, spread, or recurrence of cancer by administration of the composition of the present invention, and "treatment" refers to the symptoms of the disease by administration of the composition of the present invention. means any action that is improved or is changed for the better.

In the present invention, the term "pharmaceutical composition" means one prepared for the purpose of preventing or treating a disease, and each may be formulated in various forms according to a conventional method and used. For example, it may be formulated in oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, and syrups, and may be formulated in the form of external preparations, suppositories, and sterile injection solutions.

In the present invention, "included as an active ingredient" means that the ingredient is included in an amount necessary or sufficient to realize a desired biological effect. The amount may be determined taking into account other non-toxic factors, and may vary depending on various factors, such as, for example, the disease or condition being treated, the form of the composition being administered, the size of the subject, or the severity of the disease or condition. A person of ordinary skill in the art to which the present invention pertains can empirically determine the effective amount of an individual composition without undue experimentation.

In addition, the pharmaceutical composition of the present invention may include one or more pharmaceutically acceptable carriers in addition to the active ingredients described above according to each formulation.

The pharmaceutically acceptable carrier may be saline, sterile water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol, and a mixture of one or more of these components, if necessary, an antioxidant, a buffer, a bacteriostatic agent It may further include other conventional additives, such as. In addition, diluents, dispersants, surfactants, binders and lubricants may be additionally added to form an injectable formulation such as an aqueous solution, suspension, emulsion, etc., pills, capsules, granules or tablets. Furthermore, it may be preferably formulated according to each disease or component using an appropriate method in the art or a method disclosed in Remington's Pharmaceutical Science (Mack Publishing Company, Easton PA).

The composition of the present invention may be administered orally or parenterally in a pharmaceutically effective amount according to a desired method, and the term “pharmaceutically effective amount” of the present invention means a disease with a reasonable benefit/risk ratio applicable to medical treatment. It means an amount sufficient to treat the drug and does not cause side effects, and the effective dose level is the patient's health status, severity, drug activity, drug sensitivity, administration method, administration time, administration route and excretion rate, treatment The duration, formulation, or concomitant use may be determined by factors including drugs and other factors well known in the medical field.

Accordingly, cancer can be prevented, treated, and/or diagnosed by administering the pharmaceutical composition of the present invention to an individual, and the cancer is skin cancer, breast cancer, uterine cancer, esophageal cancer, stomach cancer, brain tumor, colon cancer, rectal cancer, colorectal cancer, Lung cancer, ovarian cancer, cervical cancer, endometrial cancer, vulvar cancer, kidney cancer, blood cancer, pancreatic cancer, prostate cancer, testicular cancer, laryngeal cancer, head and neck cancer, thyroid cancer, liver cancer, bladder cancer, osteosarcoma, lymphoma, blood cancer, thymus cancer, urethral cancer It may be cancer, or bronchial cancer, etc., but is not limited thereto, and preferably has a higher acidity than normal cells, and may be a cancer of which cytotoxicity is inhibited by a tubulin polymerization inhibitor, as a non-limiting example of which is breast cancer, preferably Examples include triple-negative breast cancer.

As used herein, the term “individual” is not limited as long as it is a mammal such as livestock or human in need of prevention, treatment, and/or diagnosis of cancer, but preferably a human.

The pharmaceutical composition of the present invention may be formulated in various forms for administration to an individual, and a representative formulation for parenteral administration is an injection formulation, preferably an isotonic aqueous solution or suspension. Formulations for injection may be prepared according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. For example, each component may be dissolved in saline or buffer to be formulated for injection. In addition, formulations for oral administration include, for example, ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups and wafers. , sucrose, mannitol, sorbitol, cellulose and/or glycine) and glidants (eg silica, talc, stearic acid and its magnesium or calcium salts and/or polyethylene glycol). The tablet may contain a binder such as magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and/or polyvinylpyrrolidine, and optionally starch, agar, alginic acid or its It may further contain disintegrating agents such as sodium salts, absorbents, coloring agents, flavoring agents and/or sweetening agents. The formulation may be prepared by conventional mixing, granulating or coating methods.

In addition, the pharmaceutical composition of the present invention may further contain adjuvants such as preservatives, wettable powders, emulsification accelerators, salts or buffers for osmotic pressure control, and other therapeutically useful substances, and may be formulated according to a conventional method. .

The pharmaceutical composition according to the present invention may be administered through several routes including oral, transdermal, subcutaneous, intravenous or intramuscular, and the dosage of the active ingredient may vary depending on the route of administration, age, sex, weight, and severity of the patient. It may be appropriately selected according to several factors. In addition, the composition of the present invention may be administered in parallel with a known compound capable of enhancing the desired effect.

The pharmaceutical composition according to the present invention may be administered orally or parenterally, such as intravenously, subcutaneously, intranasally or intraperitoneally, to humans and animals. Oral administration also includes sublingual application. Parenteral administration includes injection methods such as subcutaneous injection, intramuscular injection and intravenous injection and drip method.

In the pharmaceutical composition of the present invention, the total effective amount of the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof according to the present invention may be administered to a patient in a single dose, and multiple doses ) can be administered by a fractionated treatment protocol that is administered for a long period of time. The pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the severity of the disease, but may be administered repeatedly several times a day at an effective dose of 100 μg to 3,000 mg when administered once for adults. . However, the concentration of the thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof depends not only on the route of administration and the number of treatments, but also on various factors such as the patient's age, weight, health status, sex, severity of disease, diet and excretion rate. Taking into account the effective dosage for the patient can be determined.

In addition, the pharmaceutical composition according to the present invention is not particularly limited in its formulation, administration route and administration method as long as the effect of the present invention is exhibited, and the pharmaceutical composition of the present invention is the thiobenzimidazole derivative or its active ingredient. It may further include a known anticancer agent in addition to the pharmaceutically acceptable salt, and may be used in combination with other known treatments for the treatment of these diseases.

Terms used in the examples are used for the purpose of description only, and should not be construed as limiting. The singular expression includes the plural expression unless the context clearly dictates otherwise. In this specification, terms such as "comprise" or "have" are intended to designate that a feature, number, step, operation, component, part, or a combination thereof described in the specification exists, but one or more other features It is to be understood that this does not preclude the possibility of the presence or addition of numbers, steps, operations, components, parts, or combinations thereof.

Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which the embodiment belongs. Terms such as those defined in commonly used dictionaries should be interpreted as having a meaning consistent with the meaning in the context of the related art, and should not be interpreted in an ideal or excessively formal meaning unless explicitly defined in the present application. does not

Hereinafter, embodiments will be described in detail with reference to the accompanying drawings. However, since various changes may be made to the embodiments, the scope of the patent application is not limited or limited by these embodiments. It should be understood that all modifications, equivalents and substitutes for the embodiments are included in the scope of the rights.

In addition, in the description with reference to the accompanying drawings, the same components are given the same reference numerals regardless of the reference numerals, and the overlapping description thereof will be omitted. In describing the embodiment, if it is determined that a detailed description of a related known technology may unnecessarily obscure the gist of the embodiment, the detailed description thereof will be omitted.

[Example]

Example 1. Preparation of thiobenzimidazole derivatives

All chemical reagents were commercially available. ¹ H NMR spectra were recorded on a Bruker Avance III 400 MHz and Bruker Fourier 300 MHz and TMS was used as an internal standard.

LCMS were taken on a quadrupole Mass Spectrometer of an Agilent LC/MSD 1200 series (column: ODS 2000 (50 × 4.6 mm, 5 μm)) operating in ES (+) or (-) ionization mode; T = 30 °C, flow rate = 1.5 mL/min; Detected wavelength: 214 nm.

1.1 Synthesis of Formula 1-1

The synthesis process of the thiobenzimidazole derivative of Formula 1-1 is shown in Scheme 1 below.

[Scheme 1]

1.1.1. Synthesis of 5-(4-fluorophenylthio)-2-nitrobenzeneamine

4-Fluorothiol (1.1 g, 8.69 mmol) was dissolved in 10 mL, and K ₂ CO ₃ (1.24 g, 9.01 mmol) was added thereto. Then, 2-Nitro-5-Chloroaniline (1.5 g, 8.69 mmol) was dissolved in 3 mL and added dropwise. Then, the mixture was stirred at 90 °C for 3 hours. After confirming the completion of the reaction by TLC, it was cooled to room temperature. 50 mL of purified water was poured, and the mixture was extracted with 100 mL of ethyl acetate. This was dried over MgSO ₄ anhydrous, filtered and distilled under reduced pressure to obtain a yellow solid, which was recrystallized with ethyl acetate/n-hexane to obtain a yellow solid 5-(4-fluorophenylthio)-2-nitrobenzeneamine (1.87 g). (yield 82%).

R _f : 0.37 (EtOAc/n-Hexane, 1:5)

¹ H NMR (400 MHz. CDCl ₃ yield: 82 %, δ, ppm): 6.05 (brs, 2H, -NH ₂ ), 6.33~8.02 (m, 7H, ArH)

1.1.2. Synthesis of 4-(4-fluorophenylthio)benzene-1,2-diamine

5-(4-fluorophenylthio)-2-nitrobenzeneamine (1.0 g, 0.378 mmol) was dissolved in 20 mL of acetic acid, and then Zn (1.23 g, 18.9 mmol) was slowly added under an ice-bath. After stirring at room temperature for 10 hours, the completion of the reaction was confirmed by TLC, followed by filtration. The filtrate was distilled under reduced pressure, and the pH was adjusted to 8 with 5M-NaOH solution, followed by extraction with 100 mL of ethyl acetate. After drying with MgSO ₄ anhydrous, filtration and distillation under reduced pressure to obtain a dark brown liquid, which was then columned to obtain 4-(4-fluorophenylthio)benzene-1,2-diamine (0.84 g) as a brown liquid (yield 95.4 %) .

R _f : 0.06 (EtOAc/n-Hexane, 1:4)

¹ H NMR (400 MHz. CDCl ₃ , δ, ppm): 3.41 (brs, 2H, -NH ₂ ), 3.50 (brs, 2H, -NH ₂ ), 6.68-7.28 (m, 7H, ArH)

1.1.3. Synthesis of methyl 5-(4-fluorophenylthio)-1H-benzo[d]imidazol-2-ylcarbamate

4-(4-fluorophenylthio)benzene-1,2-diamine (0.7 g, 2.98 mmol) and 1,3-Bis(methoxycarbonyl) -S-methylisothiourea (1.6 g, 7.77 mmol) in 10 mL of 5 %-AcOH in Ethanol After dissolving in , it was heated and refluxed for 4 hours. After completion of the reaction by TLC, the resulting precipitate was filtered and cooled to room temperature. During filtration, it was washed thoroughly with MeOH and dried under vacuum at 50 ° C. to obtain Methyl 5-(4-fluorophenyl-thio)-1H-benzo[d]imidazol-2-ylcarbamate (Formula 1-1, 1.02 g) as a white solid ( yield 78%).

Melting Point: : 232-233 ℃

¹ H NMR (400 MHz. CDCl ₃ , δ, ppm): 3.85(s, 3H, -OCH ₃ ), 6.95-7.69(m, 7H, ArH), 11.74(brs, 2H, (-NH) ₂ )

1.2 Synthesis of Formula 1-2

The synthesis process of the thiobenzimidazole derivative of Formula 1-2 is shown in Scheme 2 below.

[Scheme 2]

1.2.1. Synthesis of tert-butyl 4-(4-((ethoxycarbonothioyl)thio)phenyl)piperazine-1-carboxylate

Concentrate of H ₂ SO ₄ (1.96 g, 20.0 mmol) in a solution of tert-butyl 4-(4-aminophenyl)piperazine-1-carboxylate (2.77 g, 10.2 mmol) in THF/H ₂ O (40 mL/40 mL) was added. Then, NaNO ₂ (2.10 g, 30 mmol) dissolved in water (3.0 mL) was slowly added at -5 °C to 0 °C and stirred for 2 h. Potassium O-ethyl carbonodithioate (9.60 g, 60 mmol) was added and the mixture was stirred at room temperature for 2 h. After the reaction was completed, the mixture was poured into water (100 mL) and extracted with EA (100 mL * 3). The organic layer was dried over Na ₂ SO ₄ , concentrated and purified by reverse column (ACN/H ₂ O) as an orange solid in 2.00 g of tert-butyl 4-(4-((ethoxycarbonothioyl)thio)phenyl)piperazine-1- A carboxylate was obtained (yield 52%).

1.2.2. Synthesis of tert-butyl 4-(4-((3-amino-4-nitrophenyl)thio)phenyl)piperazine-1-carboxylate

5 to a solution of tert-butyl 4-(4-((ethoxycarbonothioyl)thio)phenyl)piperazine-1-carboxylate (1.15 g, 3.00 mmol) in THF/MeOH/H ₂ O (30 mL/50 mL/10 mL) -chloro-2-nitroaniline (520 mg, 3.00 mmol) and NaOH (480 mg, 12 mmol). The mixture was stirred at 65 °C for 16 h. After completion of the reaction, the mixture was adjusted to pH = 7-8 with AcOH, concentrated and purified by reverse column (ACN/H ₂ O) to obtain 600 mg of tert-butyl 4-(4-((3- amino-4-nitrophenyl)thio)phenyl)piperazine-1-carboxylate was obtained (yield 46%).

1.2.3. Synthesis of tert-butyl 4-(4-((2-((methoxycarbonyl)amino)-1H-benzo[d]imidazol-6-yl)thio)phenyl)piperazine-1-carboxylate

1,3-bis(methoxycarbonyl) in a solution of tert-butyl 4-(4-((3-amino-4-nitrophenyl)thio)phenyl)piperazine-1-carboxylate (500 mg, 1.16 mmol) in AcOH (30 mL) )-2-methyl-2-thiopseudoeura (478 mg, 2.32 mmol) and Zn powder (603 mg, 9.28 mmol) were added, and the mixture was stirred at 75 °C for 20 h. After the reaction was completed, the solvent was removed. The mixture was dissolved in MeOH (50 mL), filtered, the filtrate was concentrated and purified by reverse column (ACN/H ₂ O) to 300 mg of tert-butyl 4-(4-((2-((methoxycarbonyl) amino)-1H-benzo[d]imidazol-6-yl)thio)phenyl)piperazine-1-carboxylate was obtained as a yellow solid (yield 53%).

1.2.4. Synthesis of methyl (6-((4-(piperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

tert-butyl 4-(4-((2-((methoxycarbonyl)amino)-1H-benzo[d]imidazol-6-yl)thio)phenyl)piperazine-1-carboxylate (300 mg) in dioxane (6 mL) , 0.62 mmol) was added HCl/dioxane (6 mL, 6 mol/L), and the mixture was stirred at room temperature for 2 h. After the reaction was completed, the solvent was removed and Et ₃ N was added. The mixture was concentrated and purified by prep-HPLC to give 100 mg of methyl (6-((4-(piperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate (Formula 1- 2) was obtained as an off-white solid (yield 42%).

¹ H-NMR (400 MHz, DMSO-d ₆ ): δ 7.33 (d, 1H, J = 8.0 Hz), 7.29 (s, 1H), 7.21 (d, 1H, J = 8.8 HZ), 7.03-7.00 ( m, 1H), 6.90 (d, 2H, J = 8.8 Hz), 3.74 (s, 3H), 3.06-3.03 (m, 4H), 2.82-2.79 (m, 4H)

1.3 Synthesis of Formula 1-3

The synthesis process of the thiobenzimidazole derivative of Formula 1-3 is shown in Scheme 3 below.

[Scheme 3]

1.3.1. Synthesis of (5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitrophenyl)carbamate

To a solution of tert-butyl (5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate (9.0 g, 21.2 mmol, 1.0 equiv) in tolenium (50 mL), 1-methylpiperazine (4.2 g, 42.4 mmol, 2.0 equiv.) ) and Pd ₂ (dba) ₃ (1.9 g, 2.12 mmol, 0.1 eq) were sequentially added. To this were added Xantphos (2.0 g, 4.24 mmol, 0.2 eq) and t-BuONa (4.1 g, 42.4 mmol, 2.0 eq), and the mixture was stirred at 100° C. for 16 h. After confirming that the reaction was complete by LCMS, the reaction product was extracted with DCM, and the organic layer was concentrated and purified by TLC to form a yellow solid tert-butyl (5-((4-(4-methylpiperazin-1-yl)phenyl)thio )-2-nitrophenyl)carbamate (4 g) was synthesized (yield 43%).

1.3.2. Synthesis of 5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitroaniline

2 (5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitrophenyl)carbamate (3 g, 6.75 mmol, 1.0 equiv) in HCl/dioxane (30 mL) at room temperature After stirring for an hour, the reaction was confirmed to be complete by LCMS. NaHCO ₃ was added to the mixture, the reaction was extracted with DCM, the organic layer was concentrated and purified by TLC, and a yellow solid 5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitroaniline (2.3 g ) was obtained (yield 95%).

1.3.3. Preparation of 4-((4-(4-methylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine

Fe in EtOH (50 mL) and H ₂ O (10 mL) solution of 5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitroaniline (2.3 g, 6.68 mmol, 1.0 eq) (1.8 g, 33.4 mmol, 5.0 eq) and NH ₄ Cl (3.7 g, 66.8 mmol, 5.0 eq) were added. After the mixture was stirred at 80 °C for 2 hours, the reaction was confirmed by LCMS, and the reaction product was extracted with DCM, the organic layer was concentrated and purified by TLC as a brown solid 4-((4-(4-methylpiperazin-1) -yl)phenyl)thio)benzene-1,2-diamine (1.9 g) was obtained (yield 90%).

¹ H NMR (400 MHz, DMSO-d ₆ ): δ 7.03 (d, J=8.8 Hz, 2H), 6.84 (d, J=8.8 Hz, 2H), 6.56 (s, 1H), 6.46 (s, 2H) ), 4.57-4.66 (m,4H), 3.06-3.08 (m, 4H), 2.40-2.42 (m, 4H), 2.19 (s, 1H)

1.3.4. Preparation of (5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

4-((4-(4-methylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine (800 mg, 2.54 mmol, 1.0 equiv) in CH ₃ COOH (20 mL) solution in 1,3- Bis(methoxycarbonyl)-S-methylisothiourea (575 mg, 2.79 mmol, 1.1 equiv) was added, and the mixture was stirred at 80 °C for 2 hours, and LCMS confirmed that the reaction was complete. The reaction mixture was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and an aliquot was purified by Prep-TLC to form methyl (5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl) as a white solid. Carbamate (632 mg) was obtained (yield 63%).

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 7.28-7.32 (m,2H), 7.20 (d, J=8.8 Hz, 2H), 6.99-7.01 (m, 1H), 6.91 (d, J=8.8 Hz, 2H), 3.71 (s, 3H), 3.11-3.14 (m, 4H), 2.41-2.43 (m, 4H), 2.20 (s, 3H)

1.4 Synthesis of Formula 1-4

The synthesis process of the thiobenzimidazole derivative of Formula 1-4 is shown in Scheme 4 below.

[Scheme 4]

1.4.1. Synthesis of methyl (5-thiocyanato-1H-benzo[d]imidazol-2-yl) carbamate

In a solution of 2-nitro-4-thiocyanatoaniline (1.95 g, 10.0 mmol) in AcOH (50 mL) Zn powder (5.00 g, 70.0 mmol) and 1,3-bis(methoxycarbonyl)-2-methyl-2-thiopseudoeura ( 2.70 g, 13.0 mmol). The mixture was stirred at 90 °C for 20 min. After the reaction was completed, the mixture was filtered. The organic layer was concentrated in vacuo. The residue was purified by reverse column (CH ₃ CN/H ₂ O) to obtain 300 mg of methyl (5-thiocyanato-1H-benzo[d]imidazol-2-yl) carbamate as a yellow solid.

1.4.2. Synthesis of methyl (5-(phenyldisulfanyl)-1H-benzo[d]imidazol-2-yl) carbamate

In a solution of methyl (5-thiocyanato-1H-benzo[d]imidazol-2-yl) carbamate (300 mg, 1.20 mmol) in EtOH (120 mL) PhSNa (319 mg, 2.40 mmol) and KOH (101 mg, 1.80) mmol) was added. The mixture was stirred at room temperature for 16 h. After the reaction was completed, EtOH was removed. The residue was purified by prep-HPLC to give 90 mg of formula 1-4 as an off-white solid (TFA salt, yield 22%).

¹ H-NMR (400 MHz, DMSO-d ₆ ): δ 7.64 (s, 1H), 7.55-7.53 (m, 2H), 7.46-7.44 (d, 1H), 7.42-7.74 (m, 2H), 7.34 -7.30 (m, 2H), 3.79 (s, 3H)

1.5 Synthesis of Formula 1-5

The synthesis process of the thiobenzimidazole derivative of Formula 1-5 is shown in Scheme 5 below.

[Scheme 5]

1.5.1. Synthesis of 5-(benzylthio)-2-nitroaniline

To a solution of phenylmethanethiol (2.40 g, 20.0 mmol) in DMSO (150 mL) was added 5-chloro-2-nitroaniline (3.40 g, 20.0 mmol) and Cs ₂ CO ₃ (13.0 g, 40.0 mmol), and the mixture was Stirred at 80 °C for 16 hours. After the reaction was completed, the mixture was poured into water (700 mL), extracted with EA (400 mL * 3), dried over Na ₂ SO ₄ , concentrated and purified by reverse column (ACN/H ₂ O) to 6.00 as a red oil. g of 5-(benzylthio)-2-nitroaniline was obtained.

1.5.2. Synthesis of 4-(benzylthio)benzene-1,2-diamine

To a solution of 5-(benzylthio)-2-nitroaniline (2.00 g, 7.70 mmol) in AcOH (50 mL) was added Zn (4.00 g, 61.0 mmol). The mixture was stirred at 90 °C for 3 h. After the reaction was completed, the mixture was filtered; The filtrate was concentrated to obtain 2.20 g of 4-(benzylthio)benzene-1,2-diamine as a brown oil.

1.5.3. Preparation of methyl (5-(benzylthio)-1H-benzo[d]imidazol-2-yl) carbamate

1,3-bis(methoxycarbonyl)-2-methyl-2-thiopseudoeura (1.50 g, 7.20 mmol) in a solution of 4-(benzylthio)benzene-1,2-diamine (2.00 g, 8.70 mmol) in AcOH (40 mL) ) was added. The mixture was stirred at 85 °C for 16 h. After the reaction was completed, AcOH was removed and extracted with EA (50 mL * 3). The organic layer was dried over Na ₂ SO ₄ and concentrated. The crude was purified by prep-HPLC to obtain 75 mg of methyl (5-(benzylthio)-1H-benzo[d]imidazol-2-yl)carbamate (Formula 1-5) as a yellow solid (yield 11%).

¹ H-NMR (400 MHz, DMSO-d ₆ ): δ11.61 (s, 1H), 7.38 (s, 1H), 7.32-7.30 (m, 1H), 7.26-7.25 (m, 1H), 7.23 7.19 (m, 4H), 7.10-7.07 (m, 1H), 4.14 (s, 2H), 3.75 (s, 3H)

1.6 Synthesis of Formula 1-6

The synthesis process of the thiobenzimidazole derivative of Formula 1-6 is shown in Scheme 6 below.

[Scheme 6]

1.6.1. Synthesis of methyl 3-((4-(piperidin-1-yl)phenyl)thio)propanoate

Compound 1-(4-bromophenyl)piperidine (5.0 g, 20.83 mmol, 1.0 eq) in dry dioxane (50 ml) solution in methyl 3-mercaptopropanoate (16.13 mL, 145.81 mmol, 7.0 eq), Xantphos (2.4 g, 4.17) mmol, 0.2 eq) was added. After the addition of N,N-Diisopropylethylamine (DIEA, 10.9 mL, 62.49 mmol, 3.0 eq) and Pd ₂ (dba) ₃ (1.9 g, 2.08 mmol, 0.1 eq) was completed to this mixture, the reactant was heated to 110 under nitrogen gas. The mixture was stirred at °C for 16 hours. After the reaction was complete, the mixture was cooled to room temperature. The reaction was filtered and the filtrate was concentrated. The residue was purified by silica gel column chromatography (HE:EtOAC=40:1) to obtain the compound methyl 3-((4-(piperidin-1-yl)phenyl)thio)propanoate as a yellow oil (5.9 g, yield) 100%).

1.6.2. Synthesis of sodium 4-(piperidin-1-yl)benzenethiolate

To a solution of the compound methyl 3-((4-(piperidin-1-yl)phenyl)thio)propanoate (3.6 g, 12.9 mmol, 1.0 equiv) in dry THF (60 mL) at room temperature 20% NaOEt (6.59 g, 19.355 mmol) , 1.5 eq) was added. The reaction was heated to room temperature for 1 hour. After the reaction was completed, the mixture was concentrated to obtain the compound sodium 4-(piperidin-1-yl)benzenethiolate as a brown compound (3.45 g, crude).

1.6.3. Synthesis of 2-Nitro-5-((4-(piperidin-1-yl)phenyl)thio)aniline

Compound sodium 4-(piperidin-1-yl)benzenethiolate (3.45 g, 16.05 mmol, 1.0 equiv), 4-fluoro-2-di(tert-butoxycarbonyl) nitroaniline (5.98 g, 16.05 mmol, 1.0 equiv) and K ₂ CO A solution of ₃ (6.65 g, 48.15 mmol, 3.0 equiv) of anhydrous 1-Methyl-2-pyrrolidinone (NMP, 60 mL) was stirred at 130 °C under nitrogen charge for 16 hours. After the reaction was completed, the residue was purified by silica gel column chromatography (HE:EtOAC=20:1) to form a yellow solid 2-nitro-5-((4-(piperidin-1-yl)phenyl)thio)aniline (630 mg) was obtained.

1.6.4. Synthesis of 4-((4-(Piperidin-1-yl)phenyl)thio)benzene-1,2-diamine

2-nitro-5-((4-(piperidin-1-yl)phenyl)thio)aniline (1-4, 450 mg, 1.37 mmol, 1.0 eq) and Pd/C (110 mg) in MeOH (30 mL) To the solution was added MeOH/NH ₃ (10 mL), and stirred in a hydrogen (H ₂ ) stream under atmospheric pressure at room temperature for 18 hours. The reaction product was filtered and the filtrate was concentrated to obtain the compound 4-((4-(piperidin-1-yl)phenyl)thio)benzene-1,2-diamine as a purple solid (370 mg) (yield 90.5%).

¹ H NMR (400 MHz, CDCl ₃ ): δ ppm 7.21-7.25 (m, 2H), 6.85 (d, J = 6.8 Hz, 2H), 6.69-6.74 (m, 2H), 6.62 (d, J = 8.0) Hz, 1H), 3.34-3.43 (m, 4H), 3.14-3.16 (m, 4H), 1.55-1.69 (m, 6H)

1.6.5. Synthesis of (5-((4-(Piperidin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

Compound 4-((4-(piperidin-1-yl)phenyl)thio)benzene-1,2-diamine (100 mg, 0.334 mmol, 1.0 equiv) in acetic acid (5 mL) solution with 1,3-Bis( methoxycarbonyl)-S-methylisothiourea (76 mg, 0.367 mmol, 1.1 equiv) was added, and the mixture was stirred and reacted at 80 °C for 2 hours. The reaction was extracted with DCM and washed with saturated NaHCO ₃ . The organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and the resulting mixture was purified by Prep-TLC to produce a white solid product methyl (5-((4-(piperidin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl) It was obtained as carbamate (Formula 1-6, 20 mg) (yield 15.7%).

¹ H NMR (400 MHz, DMSO-d6): δ 7.33 (d, J = 8.4 Hz, 1H), 7.28 (d, J = 1.2 Hz, 1H), 7.20 (d, J = 8.8 Hz, 2H), 7.01 -7.03 (m, 1H), 6.91 (d, J = 8.8 Hz, 2H), 6.08 (brs, 2H), 3.74 (s, 3H), 3.11-3.19 (m, 4H), 1.53-1.59 (m, 6H) )

1.7 Synthesis of Formula 1-7

The synthesis process of the thiobenzimidazole derivative of Formula 1-7 is shown in Scheme 7 below.

[Scheme 7]

1.7.1. Synthesis of methyl 3-((4-morpholinophenyl)thio)propanoate

In a solution of 4-(4-bromophenyl)morpholine (7.0 g, 30.9 mmol, 1.0 eq) in dry dioxane (50 mL), methyl 3-mercaptopropanoate (18.6 g, 154.9 mmol, 5.0 eq), Xantphos (3.6 g, 6.2 mmol) , 0.2 eq) was added. DIEA (12.2 g, 92.9 mmol, 3.0 eq) and Pd ₂ (dba) ₃ (1.4 g, 1.6 mmol, 0.05 eq) were sequentially added thereto, and the reaction was stirred at 110° C. for 16 hours. After the reaction was complete, the mixture was cooled to room temperature. The reaction was filtered and the filtrate was concentrated. The residue was separated and purified by silica gel column chromatography (HE:EtOAC=8:1) to obtain methyl 3-((4-morpholinophenyl)thio)propanoate (7.5 g) as a yellow oil (yield 91.2%).

1.7.2. Synthesis of sodium 4-morpholinobenzenethiolate

To a solution of methyl 3-((4-morpholinophenyl)thio)propanoate (7.4 g, 26.3 mmol, 1.0 equiv) in dry THF (100 mL) at room temperature was added 20% NaOEt (10.7 g, 31.6 mmol, 1.5 equiv). The reaction was reacted at room temperature for 1 hour. After the reaction was completed, the mixture was concentrated to obtain a brown oily liquid compound sodium 4-morpholinobenzenethiolate (5.2 g) (yield 88.9%).

1.7.3. Synthesis of 4-morpholino-2-nitroaniline

5-fluoro-2-nitroaniline (4.5 g, 28.7 mmol, 1.2 equiv) and K ₂ CO ₃ in anhydrous 1-Methyl-2-pyrrolidinone (60 mL) solution of 4-morpholinobenzenethiolate (5.2 g, 23.9 mmol, 1.0 equiv) (9.9 g, 71.8 mmol, 3.0 equiv) was added under nitrogen charge, and then stirred at 130 °C for 16 hours. After the reaction was completed, the residue was purified by column chromatography on silica gel (HE:EtOAC=20:1) to obtain the compound 4-morpholino-2-nitroaniline as a yellow solid (1.2 g) (yield 15.2%).

1.7.4. Synthesis of 4-morpholinobenzene-1,2-diamine

Pd/C (300 mg) was added to a solution of 4-morpholino-2-nitroaniline (1.2 g, 3.6 mmol, 1.0 equiv) in MeOH (20 mL), and the mixture was stirred at room temperature in the presence of hydrogen gas (H ₂ ) for 18 hours. . The reaction product was filtered, and the filtrate was used to obtain a purple solid 4-morpholinobenzene-1,2-diamine (640 mg) (yield 60.1%).

¹ H NMR (400 MHz, DMSO-d6): δ7.85 (d, J = 9.2 Hz, 1H), 7.39-7.44 (m, 3H), 7.06 (d, J = 8.8 Hz, 2H), 6.52 (d , J = 1.6 Hz, 1H), 6.24-6.27 (m, 1H), 3.69 - 3.76 (m, 4H), 3.19-3.23 (m, 4H)

1.7.5. Synthesis of methyl (5-((4-morpholinophenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

1,3-Bis(methoxycarbonyl)-S-methylisothiourea (75 mg, 0.365 mmol, 1.1 eq) was added to a solution of 4-morpholinobenzene-1,2-diamine (100 mg, 0.332 mmol, 1.0 eq) in acetic acid (5 mL). After addition, the mixture was stirred at 80 °C for 2 hours, and the reaction was confirmed to be complete by LCMS. The reaction was extracted with DCM, washed with saturated bicarbonate water, and the organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and the material was purified by Prep-TLC to give the product as a white solid (Formula 1-7, 30 mg) (yield 23.6%).

¹ H NMR (400 MHz, DMSO-d6): δ ppm 7.34 (d, J = 8.4 Hz, 1H), 7.30 (s, 1H), 7.23 (d, J = 8.8 Hz, 2H), 7.02-7.05 (m, 1H), 6.93 (d, J = 8.8 Hz, 2H), 3.74 (s, 3H), 3.71-3.73 (m, 4H), 3.10-3.12 (m, 4H)

1.8 Synthesis of Formula 1-8

The synthesis process of the thiobenzimidazole derivative of Formula 1-8 is shown in Scheme 8 below.

[Scheme 8]

1.8.1. Synthesis of 1-(4-bromophenyl)-4-ethylpiperazine

1-(4-bromophenyl)piperazine (10.0 g, 41.83 mmol, 1.0 eq) in anhydrous acetonitrile (50 mL) solution in ethyl iodide (4.0 mL, 49.78 mmol, 1.2 eq) and K ₂ CO ₃ (11.47 g, 82.96 mmol) , 2.0 eq) was added. After the addition was complete, the reaction was stirred under N ₂ at 70 °C for 2 h. The reaction was filtered and the filtrate was concentrated. The residue was purified by silica gel (HE:EtOAC=5:1) column chromatography to obtain compound 1-(4-bromophenyl)-4-ethylpiperazine as a white solid (9.84 g) (yield 84.2 %).

1.8.2. Synthesis of methyl 3-((4-(4-ethylpiperazin-1-yl)phenyl)thio)propanoate

In a dry dioxane (50 mL) solution of 1-(4-bromophenyl)-4-ethylpiperazine (9.4 g, 34.9 mmol, 1.0 eq), methyl 3-mercaptopropanoate (27.1 mL, 244.4 mmol, 7.0 eq), Xantphos (4.04 g) , 6.98 mmol, 0.2 eq) was added. After the addition of DIEA (18.3 mL, 104.7 mmol, 3.0 eq) and Pd ₂ (dba) ₃ (3.2 g, 3.5 mmol, 0.1 eq) was completed, the reaction was stirred at 110° C. under a N ₂ stream for 16 hours. The reaction was filtered and the concentrate was concentrated. The residue was purified by silica gel column chromatography (Hexane:EtOAC=2:1) to give the compound methyl 3-((4-(4-ethylpiperazin-1-yl)phenyl)thio)propanoate (9.5 g) as a yellow oil. obtained (yield 88.29%).

1.8.3. Synthesis of sodium 4-(4-ethylpiperazin-1-yl)benzenethiolate

To a solution of 3-((4-(4-ethylpiperazin-1-yl)phenyl)thio)propanoate (2.81 g, 9.1 mmol, 1.0 equiv) in dry THF (60 mL), 20% sodium ethoxide (4.64 g, 13.65 mmol, 1.5 eq) was added and then the reaction was heated to room temperature for 20 min. After the reaction was completed, the mixture was concentrated to obtain a brown solid compound sodium 4-(4-ethylpiperazin-1-yl)benzenethiolate (2.8 g).

1.8.4. Synthesis of 5-((4-(4-ethylpiperazin-1-yl)phenyl)thio)-2-nitroaniline

Sodium 4-(4-ethylpiperazin-1-yl)benzenethiolate (2.8 g, 11.5 mmol, 1.0 equiv) was dissolved in dry N-methylpyrrolidinedimethyl (60 mL), tert-butyl (tert-butoxycarbonyl) (5 -chloro-2-nitrophenyl)carbamate (4.29 g, 11.5 mmol, 1.0 equiv) and K ₂ CO ₃ (4.77 g, 34.5 mmol, 3.0 equiv) were sequentially added thereto. The solution was stirred at 130 °C under a N ₂ stream for 16 hours. . After the reaction was completed, the residue was purified by silica gel column chromatography (DCM:MeOH=20:1) to form an orange solid 5-((4-(4-ethylpiperazin-1-yl)phenyl)thio)-2- nitroaniline (520 mg) was obtained.

1.8.5. Synthesis of 4-((4-(4-ethylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine

5-((4-(4-ethylpiperazin-1-yl)phenyl)thio)-2-nitroaniline (520 mg, 1.45 mmol, 1.0 equiv) in MeOH (15 mL) and MeOH/NH ₃ (5 mL) solution Pd/C (150 mg) was added, and the solution was stirred in the presence of hydrogen (H ₂ ) at room temperature for 18 hours. The reaction product was filtered and the filtrate was concentrated to obtain a purple solid compound 4-((4-(4-ethylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine (470 mg) (yield 99.5%).

¹ H NMR (400 MHz, CDCl ₃ ): δ 7.22 (d, J = 8.4 Hz, 2H), 6.82 (d, J = 8.8 Hz, 2H), 6.73-6.78 (m, 2H), 6.64 (d, J ) = 7.6 Hz, 1H), 3.26-3.46 (m, 8H), 2.78-2.92 (m, 4H), 1.51-1.64 (m, 2H), 1.29-1.30 (m, 3H)

1.8.6. Synthesis of methyl (5-((4-(4-ethylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

4-((4-(4-ethylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine (100 mg, 0.305 mmol, 1.0 equiv) was added to a solution of acetic acid (5 mL). The mixture of 1,3-Bis(methoxycarbonyl)-S-methylisothiourea (69 mg, 0.335 mmol, 1.1 equiv) was stirred at 80 °C for 2 hours, and the reaction was confirmed by LCMS. The reaction was extracted with DCM and washed with saturated NaHCO ₃ . The filtrate was concentrated and the product was purified by Prep-TLC to give the product as a white solid methyl (5-((4-(4-ethylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2 -yl) carbamate (Formula 1-8, 44 mg) was obtained (yield 32%).

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 7.33 (d, J = 8.0 Hz, 1H), 7.29 (s, 1H), 7.21 (d, J = 8.8 Hz, 2H), 7.00-7.03 (m, 1H), 6.92 (d, J = 8.8 Hz, 2H), 3.74 (s, 3H), 3.13-3.15 (m, 4H), 2.46-2.48 (m, 4H), 2.32-2.37 (m, 2H), 1.01 -1.04 (m, 3H)

1.9 Synthesis of Formulas 1-9

The synthesis process of the thiobenzimidazole derivative of Formula 1-9 is shown in Scheme 9 below.

[Scheme 9]

1.9.1. Synthesis of 1-(3-bromophenyl)-4-methylpiperazine

1-methylpiperazine (19.2 g, 192.4 mmol, 5.0 equiv), BINAP (4.8 g, 7.7 mmol, 0.2 eq), DBU in toluene (100 mL) solution of 1,3-dibromobenzene (9.0 g, 38.5 mmol, 1.0 equiv) (17.5 g, 115.5 mmol, 3.0 eq) and Pd ₂ (dba) ₃ (2.2 g, 1.9 mmol, 0.05 eq) were added respectively, and the reaction mixture was heated to 60 ° C. and then freshly ground sodium tert-butoxide (12.9 g , 192.4 mmol, 3.0 eq) were added in one portion. After the reaction was stirred at 60 °C under N ₂ for 16 h, the reaction was concentrated. The residue was separated and purified by silica gel column chromatography (HE:EtOAC=60:1) to give 1-(3-bromophenyl)-4-methylpiperazine (5.5 g) as a yellow oil (yield 56.3%).

1.9.2. Synthesis of methyl 3-((3-(4-methylpiperazin-1-yl)phenyl)thio)propanoate

To a solution of 1-(3-bromophenyl)-4-methylpiperazine (5.5 g, 21.65 mmol, 1.0 equiv) in anhydrous dioxane (50 mL) was added methyl 3-mercaptopropanoate (18.2 mg, 155.55 mmol, 7.0 equiv). Xantphos (2.5 g, 4.33 mmol, 0.2 equiv), DIEA (8.4 g, 64.95 mmol, 3.0 equiv) and Pd ₂ (dba) ₃ (2.0 g, 2.165 mmol, 0.1 eq) were sequentially added to this, after completion of the reaction. was stirred under N ₂ at 110 °C for 24 hours. The reaction was filtered and the filtrate was concentrated. The residue was purified by silica gel column chromatography (DCM:MeOH=20:1) to give methyl 3-((3-(4-methylpiperazin-1-yl)phenyl)thio)propanoate as an orange oil (4.5 g). (yield 70.7%).

1.9.3. Synthesis of sodium 3-(4-methylpiperazin-1-yl)benzenethiolate

To a solution of methyl 3-((3-(4-methylpiperazin-1-yl)phenyl)thio)propanoate (2.3 g, 7.82 mmol, 1.0 equiv) in dry THF (60 mL), 20% NaOEt (3.8 g, 11.73 mmol, 1.5 eq) was added. The reaction at room temperature was heated for 10 minutes. After the reaction was completed, the mixture was concentrated to obtain a brown solid (2.05 g) of sodium 3-(4-methylpiperazin-1-yl)benzenethiolate.

1.9.4. Synthesis of 5-((3-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitroaniline

Sodium 3-(4-methylpiperazin-1-yl)benzenethiolate (2.05 g, 8.91 mmol, 1.0 equiv) in dry N-methylpyrrolidinedimethyl (60 mL) solution in tert-butyl (tert-butoxycarbonyl)(5-chloro- 2-nitrophenyl)carbamate (3.3 g, 8.91 mmol, 1.0 equiv) and K ₂ CO ₃ (3.7 g, 26.73 mmol, 3.0 equiv) were sequentially added and stirred under N ₂ at 130 °C for 16 h. After the reaction was completed, the residue was purified by silica gel column chromatography (DCM:MeOH=20:1) to obtain an orange color of 5-((3-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitroaniline. A solid compound (3.1 g) was obtained.

1.9.5. Synthesis of 4-((3-(4-methylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine

5-((3-(4-methylpiperazin-1-yl)phenyl)thio)-2-nitroaniline (3.1 g, 9.0 mmol, 1.0 equiv) and Pd/C (1.0 g) in MeOH solution of (60 mL) and MeOH/NH ₃ (20 mL) were added and stirred in the presence of H ₂ for 18 h. The reaction was filtered, and the filtrate was concentrated to obtain 4-((3-(4-methylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine (600 mg) as a gray solid.

¹ H NMR (400 MHz, CDCl ₃ ): δ 7.07-7.12 (m, 1H), 6.84-6.89 (m, 2H), 6.78-6.80 (m, 1H), 6.66-6.70 (m, 2H), 6.60- 6.63 (m, 1H), 3.49 (s, 2H), 3.36 (s, 2H), 3.14-3.17 (m, 4H), 2.52-2.55 (m, 4H), 2.34 (d, J= 5.6 Hz, 3H)

1.9.6. Synthesis of methyl (5-((3-(4-methylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

4-((3-(4-Methylpiperazin-1-yl)phenyl)thio)benzene-1,2-diamine (100 mg, 0.318 mmol, 1.0 equiv) in CH ₃ COOH (10 mL) solution in 1,3- Bis(methoxycarbonyl)-S-methylisothiourea (72 mg, 0.35 mmol, 1.1 eq) was added. After the reaction mixture was stirred at 80 °C for 2 hours, it was confirmed by LCMS that the reaction was complete. The reaction was extracted with DCM and sat. NaHCO ₃ The organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and the mixture was purified by Prep-TLC to form methyl (5-((3-(4-methylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate as a white solid. (Formula 1-9, 20 mg) was obtained (yield 15.87%).

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 7.49 (s, 1H), 7.43 (d, J=8.0 Hz, 1H), 7.16-7.18 (m, 1H), 7.08-7.12 (m, 1H), 6.77 (d, J= 6 Hz, 2H), 6.47 (d, J=7.6 Hz, 1H), 3.76 (s, 3H), 3.04-3.07 (m, 4H), 2.38-2.40 (m, 4H), 2.19 (s, 3H)

1.10 Synthesis of Formula 1-10

The synthesis process of the thiobenzimidazole derivative of Formula 1-10 is shown in Scheme 10 below.

[Scheme 10]

1.10.1. Synthesis of tert-butyl (tert-butoxycarbonyl)(5-chloro-2-nitrophenyl)carbamate

In a solution of 5-chloro-2-nitroaniline (20 g, 116.2 mmol, 1.0 equiv) in THF (300 mL), di-tert-butyl decarbonate (50 g, 232.4 mmol, 2.0 equiv) and DMAP (14 g, 116.2 mmol, 1.0 equivalent) was added. The reaction mixture was stirred at 70 °C for 1 h. It was confirmed by LCMS that the reaction was complete. The reaction was concentrated and purified by silica gel column chromatography (eluting with HE:EA=10:1) to give tert-butyl (tert-butoxycarbonyl)(5-chloro-2-nitrophenyl)carbamate (36 g) as a yellow solid. (yield 85%).

1.10.2. Synthesis of tert-butyl (5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate

4-bromobenzenethiol (21 g, 112.9 mmol, 1.5 eq) in DMF (100 mL) solution of tert-butyl (tert-butoxycarbonyl)(5-chloro-2-nitrophenyl)carbamate (28 g, 75.2 mmol, 1.0 eq) K ₂ CO ₃ (20.7 g, 150.4 mmol, 2eq) was added sequentially, and the reaction mixture was stirred at 100° C. for 16 h. LCMS confirmed that the reaction was complete. The reaction was extracted with DCM and washed with brine. The organic layer was dried and concentrated to obtain tert-butyl(5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate (10.2 g, yield 35%) as a yellow solid.

1.10.3. Synthesis of tert-butyl (2-nitro-5-((4-(thiophen-3-yl)phenyl)thio)phenyl)carbamate

A solution of tert-butyl (5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate (4.3 g, 10.1 mmol, 1.0 equiv) in DMF (20 mL) and H ₂ O (4 mL) solution of thiophen-3- yl boronic acid (2.6 g, 20.2 mmol, 2.0 eq) was added. To this mixture, Pd(PPh ₃ ) ₄ (1.2 g, 1.01 mmol, 0.1 eq) and K ₂ CO ₃ (4.2 g, 30.3 mmol, 3.0 eq) were sequentially added and stirred at 100° C. for 16 hours. After confirming that the reaction was complete by LCMS, the reaction was extracted with DCM and washed with brine. The organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated, and the residue was purified by silica gel column chromatography (eluting with HE:EA=3:1) as a yellow solid tert-butyl (2-nitro-5-((4-(thiophen-3-yl) )phenyl)thio)phenyl)carbamate (2.7 g) was obtained (yield 62%).

1.10.4. Synthesis of 2-nitro-5-((4-(thiophen-3-yl)phenyl)thio)aniline

A solution of tert-butyl (2-nitro-5-((4-(thiophen-3-yl)phenyl)thio)phenyl)carbamate (2.6 g, 6.07 mmol, 1.0 equiv) in HCl/dioxane (20 mL) was brought to room temperature. was stirred for 2 hours. After confirming that the reaction was complete by LCMS, the reaction mixture was poured into 100 mL of an ice solution of NaHCO ₃ . The mixture was extracted with DCM and washed with brine. The organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and purified by silica gel column chromatography eluting with DCM:MeOH=10:1 to give a yellow solid 2-nitro-5-((4-(thiophen-3-yl)phenyl)thio)aniline (1.5 g) was obtained (yield 78%).

1.10.5. Synthesis of 4-((4-(thiophen-3-yl)phenyl)thio)benzene-1,2-diamine

2-nitro-5-((4-(thiophen-3-yl)phenyl)thio)aniline (1.3 g, 3.96 mmol, 1.0 equiv) in EtOH (20 mL) and H ₂ O (5 mL) solution of Fe ( 1.1 g, 19.8 mmol, 5.0 eq) and NH ₄ Cl (1.1 g, 19.8 mmol, 5.0 eq) were added and the mixture was stirred at 80° C. for 2 h. After completion of the reaction, monitored by LCMS, the mixture was extracted with DCM and washed with brine. The organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and purified by silica gel column chromatography eluting with DCM:MeOH=10:1 to form a brown solid 4-((4-(thiophen-3-yl)phenyl)thio)benzene-1,2-diamine ( 900 mg) (yield 76%).

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 7.77-7.78(m,1H), 7.57-7.76(m,3H), 7.47-7.49 (m,1H), 7.03-7.06 (m,2H), 6.68 (s, 1H), 6.56-6.61 (m, 2H), 4.67-4.84 (m, 4H)

1.10.6. Synthesis of methyl (5-((4-(thiophen-3-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

In a CH ₃ COOH (5 mL) solution of 4-((4-(thiophen-3-yl)phenyl)thio)benzene-1,2-diamine (100 mg, 0.335 mmol, 1.0 equiv) 1,3-Bis( methoxycarbonyl)-S-methylisothiourea (76 mg, 0.369 mmol, 1.1 equiv) was added. The reaction mixture was stirred at 80 °C for 2 h. After confirming that the reaction was complete by LCMS, the product of the reaction was extracted with DCM and washed with a saturated solution. The organic layer was washed with saturated NaHCO ₃ , dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated, and this material was purified by Prep-TLC to form a white solid methyl (5-((4-(thiophen-3-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate ( Formula 1-10, 50 mg) (yield 39.1%).

¹ H NMR (400 MHz, DMSO-d ₆ ): δ ppm 7.82-7.84 (m, 1H), 7.61-7.66 (m, 3H), 7.45-7.54 (m, 3H), 7.10-7.23 (m, 3H), 3.76 (s, 3H)

1.11 Synthesis of Formula 1-11

The synthesis process of the thiobenzimidazole derivative of Formula 1-11 is shown in Scheme 11 below.

[Scheme 11]

1.11.1. Synthesis of methyl 3-(furan-2-ylthio)propanoate

2-Bromofuran (6.0 g, 40.8 mmol, 1.0 equiv) in anhydrous dioxane (50 mL) solution in methyl 3-mercaptopropanoate (31.6 mL, 285.6 mmol, 7.0 equiv), Xantphos (4.787 g, 8.16 mmol, 0.2 eq)) , DIEA (21.4 mL, 122.4 mmol, 3.0 eq) and Pd ₂ (dba) ₃ (3.623 g, 4.08 mmol, 0.1 eq) were added sequentially. After the addition was complete, the reaction was stirred under N ₂ at 110 °C for 16 h. The reaction was filtered and the filtrate was concentrated. The obtained product was purified by column chromatography on silica gel (HE:EtOAC=40:1) to obtain yellow oily liquid methyl 3-(furan-2-ylthio)propanoate (6.34 g) (yield 83.4 %).

1.11.2. Synthesis of sodium furan-2-thiolate

To a solution of methyl 3-(furan-2-ylthio)propanoate (6.34 g, 34.08 mmol, 1.0 equiv) in dry THF (60 mL) was added 20% NaOEt (14.06 g, 41.32 mmol, 1.2 equiv) at room temperature. After the reaction was completed by heating the reaction mixture for 1 hour, the mixture was concentrated to obtain sodium furan-2-thiolate (4.61 g) as a brown solid.

1.11.3. Synthesis of 5-(furan-2-ylthio)-2-nitroaniline

of sodium furan-2-thiolate (4.61 g, 37.86 mmol, 1.1 equiv), 5-fluoro-2-nitroaniline (5.39 g, 34.4 mmol, 1.0 equiv) and K ₂ CO ₃ (14.26 g, 103.2 mmol, 3.0 equiv) Anhydrous N-methylpyrrolidinedimethyl (60 mL) solution was stirred under N ₂ at 130 °C for 16 hours. After the reaction was completed, the residue was purified by silica gel column chromatography (HE: EtOAC = 20: 1) to obtain 5-(furan-2-ylthio)-2-nitroaniline (860 mg) as a yellow solid.

1.11.4. Synthesis of 4-(furan-2-ylthio)benzene-1,2-diamine

5-(furan-2-ylthio)-2-nitroaniline (860 mg, 3.624 mmol, 1.0 eq) and Pd/C (360 mg) in a mixed solvent of MeOH (36 mL) and MeOH/NH ₃ (12 mL) and stirred in the presence of H ₂ at room temperature for 18 h. The reaction mixture was filtered and the filtrate was concentrated to obtain 4-(furan-2-ylthio)benzene-1,2-diamine (750 mg) as a purple solid (yield 100%).

¹ H NMR (400 MHz, CDCl ₃ ): δ ppm 7.49-7.50 (m, 1H), 6.69-6.74 (m, 2H), 6.59-6.62 (m, 2H), 6.39-6.41 (m, 1H), 3.37 (brs, 4H)

1.11.5. Synthesis of methyl (5-(furan-2-ylthio)-1H-benzo[d]imidazol-2-yl)carbamate

In a solution of 4-(furan-2-ylthio)benzene-1,2-diamine (100 mg, 0.485 mmol, 1.0 equiv) in CH ₃ COOH (5 mL), 1,3-Bis(methoxycarbonyl)-S-methylisothiourea (100 mg, 0.485 mmol, 1.1 equiv) and the mixture was stirred at 80 °C for 2 h. The reaction was monitored by LCMS to confirm that the reaction was complete, then extracted with DCM and washed with saturated NaHCO ₃ solution. The organic layer was dried over anhydrous Na ₂ SO ₄ and filtered. The filtrate was concentrated and the reaction residue was purified by Prep-TLC to form a white solid methyl (5-(furan-2-ylthio)-1H-benzo[d]imidazol-2-yl)carbamate (Formula 1-11, 20 mg) was obtained (yield 14.2%).

¹ H NMR (400 MHz, DMSO-d ₆ ): δ ppm 7.88-7.89 (m, 1H), 7.40 (d, J = 8.4 Hz, 1H), 7.30 (d, J = 8.4 Hz, 1H), 7.07- 7.10 (m, 1H), 6.92-6.93 (m, 1H), 6.61-6.63 (m, 1H), 3.78 (s, 3H)

1.12 Synthesis of Formula 1-12

Using 3,4-difluorobenzenethiol as a starting material, in the same manner as in Formula 1-1 of Example 1.1, methyl (5-((3,4-difluorophenyl)thio)-1H-benzo[d]imidazol-2-yl) Carbamate was synthesized (Formula 1-12, yield 69.2%).

¹ H NMR (CDCl ₃ , 400 MHz) 3.76 (s, 3H), 6.93 (m, 1H), 7.17 (m, 2H), 7.19 (d, J=2.32 Hz, 1H), 7.23 (q, 1H), 7.38 (d, J=10.64 Hz), 7.55 (s, 1H), 11.53 (m, 1H), 12.02 (m, 1H)

1.13 Synthesis of Formula 1-13

Using 2,4-difluorobenzenethiol as a starting material, in the same manner as in Formula 1-1 of Example 1.1, methyl (5-((2,4-difluorophenyl)thio)-1H-benzo[d]imidazol-2-yl) Carbamate was synthesized (Formula 1-13, yield 75.4%).

¹ H NMR (DMSO-d6, 400 MHz) 3.75(s, 3H), 7.12(m, 3H), 7.33(m, 1H), 7.38(m, 1H), 7.47(s, 1H), 11.73(m, 2H)

1.14 Synthesis of Formula 1-14

Using 5-chloropyridine-2-thiol as a starting material, methyl (5-((5-chloropyridin-2-yl)thio)-1H-benzo[d]imidazol- 2-yl) carbamate was synthesized (Formula 1-14, yield 75.2%).

¹ H NMR (DMSO-d6, 400 MHz) 3.77 (s, 3H), 6.75 (d, J=8.68 Hz, 1H), 7.31 (d, J=1.68 Hz), 7.53 (d, J=8.2 Hz, 1H) ), 7.64(s, 1H), 7.69(d, J=2.6 Hz), 8.44(s, 1H), 11.96(m, 2H)

1.15 Synthesis of Formula 1-15

The synthesis process of the thiobenzimidazole derivative of Formula 1-15 is shown in Scheme 15 below.

[Scheme 15]

1.15.1. Synthesis of 5-((4-Bromophenyl)thio)-2-nitroaniline

DMF (3 mL) solution was added. The reaction mixture was stirred at 90 °C for 3 hours, cooled to room temperature, poured into cold water (500 mL), and the precipitate was filtered and dried to give a yellow solid product 5-((4-Bromophenyl)thio)-2-nitroaniline (2.8 g). obtained (yield 99%).

1.15.2. Synthesis of tert-Butyl (5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate

To a solution of 5-((4-Bromophenyl)thio)-2-nitroaniline (2.8 g, 8.7 mmol) in THF (30 mL), di-tert-butyl dicarbonate (5.14 g, 23.5 mmol) and DMAP (0.146 g) were added. The reaction was heated to reflux for 1 hour. After confirming the completion of the reaction by TLC, the reaction product was cooled to room temperature and evaporated under reduced pressure to remove the solvent. The product was dissolved in methanol (30 mL), potassium carbonate (5.2 g) was added, stirred at room temperature for 6 hours, and then treated after reaction tert-Butyl (5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate ( 1.48 g) was obtained (yield 40%).

1.15.3. Synthesis of tert-butyl (2-nitro-5-((4-(thiophen-2-yl)phenyl)thio)phenyl)carbamate

tert-Butyl (5-((4-bromophenyl)thio)-2-nitrophenyl)carbamate (0.7 g, 1.6 mmol) in DMF (5 mL) solution with thiophen-2-ylboronic acid (0.42 g, 3.28 mmol, 2.0 eq ), Tetrakis(triphenylphosphine)palladium (0.18 g, 0.16 mmol, 0.1 eq), potassium carbonate (0.66 g, 4.80 mmol, 3 eq) were added, stirred at 100 ° C. for 15 hours, cooled to room temperature, and then added with methylene chloride solution. After extraction, it was washed once with Brine solution, dried over anhydrous magnesium sulfate, evaporated under reduced pressure, and separated and purified by silica-Gel column chromatography using ethyl acetate and hexane (7:3, v/v) eluent to obtain a pale yellow solid product tert-butyl (2-nitro-5-((4-(thiophen-2-yl)phenyl)thio)phenyl)carbamate (0.48 g) was obtained (yield 70%).

1.15.4. Synthesis of 2-nitro-5-((4-(thiophen-2-yl)phenyl)thio)aniline

tert-butyl (2-nitro-5-((4-(thiophen-2-yl)phenyl)thio)phenyl)carbamate (0.48 g, 1.1 mmol) in HCl/dioxane (6 mL) for 2 h at room temperature After reacting for a while, it was poured into a cold saturated aqueous sodium bicarbonate solution (30 mL), extracted with an excess of methylene chloride solution, dried over anhydrous sodium sulfate, filtered, the solvent was removed by evaporation under reduced pressure, and purified by silica gel column chromatography 2- nitro-5-((4-(thiophen-2-yl)phenyl)thio)aniline (413 mg) was obtained (yield 78%).

1.15.5. Synthesis of 4-((4-(thiophen-2-yl)phenyl)thio)benzene-1,2-diamine

To a solution of 2-nitro-5-((4-(thiophen-2-yl)phenyl)thio)aniline (413 mg, 1.2 mmol) in ethanol (15 mL), tin(II) chloride dihydrate (1.74 g, 7.7 mmol, 6.4 eq) was added, heated to reflux for 3 hours, cooled to room temperature, and water (50 mL) was added. After extraction with an excess of ethyl acetate solvent, the solvent was evaporated under reduced pressure to obtain a solid reaction mixture. This was separated and purified by column chromatography using silica gel using a solvent of ethyl acetate and hexane (1:3, v/v) to 4-((4-(thiophen-2-yl)phenyl)thio)benzene-1 ,2-diamine (0.20 g) was obtained (yield 57%).

1.15.6. Synthesis of methyl (5-((4-(thiophen-2-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

4-((4-(thiophen-2-yl)phenyl)thio)benzene-1,2-diamine (0.1 g, 0.33 mmol) and 1,3-Bis(methoxycarbonyl)-S-methylisothiourea (0.55 g, 2.6 mmol) , 8 eq) was dissolved in 3 ml of 5%-AcOH in Ethanol and reacted at 80 °C for 2 hours. The completion of the reaction was confirmed by TLC, extraction was performed with an excess of methylene chloride, neutralized with saturated sodium bicarbonate water, dried over anhydrous sodium sulfate, and separated by silica gel column chromatography to obtain a light brown solid, Methyl (5-((4-( thiophen-2-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate (Formula 1-15, 90 mg) was obtained (yield 72%).

¹ H NMR (DMSO-d ₆ , 400 MHz) 3.76 (s, 3H), 7.12 (m, 3H), 7.23 (d, J=8.16 Hz, 1H), 7.47 (m, 2H), 7.51-7.58 (m) , 4H), 11.77 (m, 2H)

1.16 Synthesis of Formula 1-16

The synthesis process of the thiobenzimidazole derivative of Formula 1-16 is shown in Scheme 16 below.

[Scheme 16]

1.16.1. Preparation of 5-((4-(1H-imidazole-1-yl)phenylthio)-2-nitroaniline

1H-imidazole (0.23 g, 3.3 mmol) was added to a DMSO (5 mL) solution of 5-((4-fluorophenyl)thio)-2-nitroaniline (0.85 g, 3.2 mmol), and after stirring for 5 minutes, potassium t-butoxide ( 0.36 g, 3.2 mmol) was slowly added, and then reacted at 90° C. for 2 hours. The reaction mixture was extracted with ethyl acetate, washed with water, and the solvent was removed by evaporation under reduced pressure to obtain a mixed product. This was separated and purified by column chromatography using silica gel using a solvent of ethyl acetate and hexane (3:1, v/v) to 5-((4-(1H-imidazole-1-yl)phenylthio)-2-nitroaniline ( 0.32 g) (yield 32%).

1.16.2. Preparation of 5-((4-(1H-imidazole-1-yl)phenylthio)-1,2-diamine

To a solution of 5-((4-(1H-imidazole-1-yl)phenylthio)-2-nitroaniline (0.309 g, 1.0 mmol) in ethanol (10 mL) was added tin chloride two hydrate (6 eq) and heated for 4 hours. , refluxed, cooled to room temperature, added water (50 mL), extracted with an excess of ethyl acetate solvent, and then the solvent was removed by evaporation under reduced pressure to obtain a solid reaction mixture. /v) separation and purification by column chromatography using silica gel using a solvent to obtain 5-((4-(1H-imidazole-1-yl)phenylthio)-1,2-diamine (0.30 g) (yield 98%) ).

1.16.3. Synthesis of methyl (5-((4-(1H-imidazol-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate.

5-((4-(1H-imidazole-1-yl)phenylthio)-1,2-diamine (0.28 g, 1.0 mmol) in a solution of 5% acetic acid in ethanol (3.0 mL) with 1,3-Bis( After adding methoxycarbonyl)-2-methyl-2-thiopseudourea (0.52 g, 2.5 mmol), it was heated and refluxed for 5 hours, The reaction mixture was filtered, washed thoroughly with methanol, and methyl (5-((4-(1H-imidazol) -1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate (Formula 1-16, 0.26 g) was obtained (yield 70%).

¹ H NMR (DMSO-d ₆ , 400 MHz) 3.76(s, 3H), 7.18(s, 1H), 7.23(m, 3H), 7.48(m, 1H), 7.58(m, 3H), 7.68(s) , 1H), 8.20 (s, 1H), 11.46 (m, 1H), 11.94 (m, 1H)

1.17 Synthesis of Formula 1-17

Using 4,6-dimethylpyrimidine-2-thiol as a starting material, methyl(5-((4,6-dimethylpyrimidin-2-yl)thio)-1H-benzo[ d]imidazol-2-yl)carbamate was synthesized (Formula 1-17, yield 75.7%).

¹ H NMR (DMSO-d6, 400 MHz) 2.25(s, 6H), 3.77(s, 3H), 6.93(s, 1H), 7.26(d, J=8.24 Hz, 1H), 7.45(d, J= 8.20 Hz, 1H), 7.61 (s, 1H), 11.74 (m, 2H)

1.18 Synthesis of Formula 1-18

Synthesis of methyl(5-(pyridin-2-ylthio)-1H-benzo[d]imidazol-2-yl)carbamate in the same manner as in Formula 1-1 of Example 1.1 using pyridine-2-thiol as a starting material was (Formula 1-18, yield 72.5%).

¹ H NMR (DMSO-d6, 400 MHz) 3.77 (s, 3H), 6.73 (d, J=8.08, 1H), 7.08 (m, 1H), 7.28 (d, J=6.72 Hz, 1H), 7.55 ( m, 2H), 7.64 (s, 1H), 8.37 (d, J=3.36 Hz, 1H), 11.89 (m, 2H).

1.19 Synthesis of Formula 1-19

The synthesis process of the thiobenzimidazole derivative of Formula 1-19 is shown in Scheme 19 below.

[Scheme 19]

1.19.1. Synthesis of 5-((4-(1H-1,2,4-triazol-1-yl)phenyl)thio)-2-nitroaniline

1H-1,2,4-triazole (2.2 g, 32 mmol) was added to a DMSO (30 mL) solution of 5-((4-fluorophenyl)thio)-2-nitroaniline (2.0 g, 29 mmol) and stirred for 5 minutes Then potassium t-butoxide (3.6 g, 33 mmol) was slowly added thereto, and then reacted at 90° C. for 4 hours. The reaction mixture was extracted with ethyl acetate, washed with water, and the solvent was removed by evaporation under reduced pressure to obtain a mixed product. This was separated and purified by column chromatography using silica gel using a solvent of ethyl acetate and hexane (3:1, v/v) to 5-((4-(1H-1,2,4-triazol-1-yl) phenyl)thio)-2-nitroaniline (0.94 g) was obtained (yield 47%).

1.19.2. Synthesis of 4-((4-(1H-1,2,4-triazol-1-yl)phenyl)thio)benzene-1,2-diamine

SnCl ₂ 2H ₂ O in ethanol (100 mL) solution of 5-((4-(1H-1,2,4-triazol-1-yl)phenyl)thio)-2-nitroaniline (0.94 g, 3.0 mmol) (6 eq) was added, heated to reflux for 5 hours, cooled to room temperature, and water (300 mL) was added. After extraction with an excess of ethyl acetate solvent, the solvent was evaporated under reduced pressure to obtain a solid reaction mixture. This was separated and purified by column chromatography using silica gel using a solvent of ethyl acetate and hexane (3:1, v/v) to 4-((4-(1H-1,2,4-triazol-1-yl) )phenyl)thio)benzene-1,2-diamine (0.85 g) was obtained (yield 99%).

1.19.3. Synthesis of methyl (5-((4-(1H-1,2,4-triazol-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate

4-((4-(1H-1,2,4-triazol-1-yl)phenyl)thio)benzene-1,2-diamine (0.85 g, 3.0 mmol) in 5 % acetic acid in ethanol (10 mL) After adding 1,3-Bis(methoxycarbonyl)-2-methyl-2-thiopseudourea (1.5 g, 7.7 mmol) to the solution, it was heated and refluxed for 5 hours. The reaction mixture was cooled to room temperature and then filtered. At this time, wash thoroughly with methanol and methyl (5-((4-(1H-1,2,4-triazol-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate (Formula 1- 19, 0.81 g) (yield 74%).

¹ H NMR (DMSO-d ₆ , 400 MHz) 3.76 (s, 3H), 7.25 (m, 1H), 7.28 (d, J=8.6 Hz, 2H), 7.47 (m, 1H), 7.57 (s, 1H) ), 7.78 (d, J=8.6 Hz, 2H), 8.21 (s, 1H), 9.23 (s, 1H), 11.80 (m, 2H)

1.20 Synthesis of Formula 1-20

methyl(5-((4-(4-methyl-1H-imidazol-1-yl)phenyl)thio) in the same manner as in Formula 1-19 of Example 1.19 using 4-methyl-1H-imidazole as a starting material -1H-benzo[d]imidazol-2-yl)carbamate was synthesized (Formula 1-20, yield 78.1%).

¹ H NMR (DMSO-d6, 400 MHz) 2.14 (s, 3H), 3.76 (s, 3H), 7.23 (d, J=8.4 Hz, 3H), 7.37 (s, 1H), 7.47 (m, 1H) , 7.53 (d, J=8.8 Hz, 3H), 8.07 (s, 1H), 11.47 (m, 1H), 12.01 (M, 1H)

1.21 Synthesis of Formula 1-21

Using 3,5-dimethyl-1H-1,2,4-triazole as a starting material, methyl (5-((4-(3,5-dimethyl-1H- 1,2,4-triazol-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate was synthesized (Formula 1-21, yield 71.9%).

¹ H NMR (DMSO-d6, 400 MHz) 2.24(s, 3H), 2.37(s, 3H), 3.76(s, 3H), 7.20(d, J=8.8 Hz, 2H), 7.28(m, 1H) , 7.43 (d, J=8.8 Hz, 2H), 7.15 (m, 1H), 7.60 (s, 1H), 11.41 (m, 1H), 12.37 (m, 1H)

1.22 Synthesis of Formula 1-22

Using 3-methyl-1H-1,2,4-triazole as a starting material, methyl (5-((4-(3-methyl-1H-1,2, 4-triazol-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate was synthesized (Formula 1-22, yield 70.4%).

¹ H NMR (DMSO-d6, 400 MHz) 2.37(s, 3H), 3.76(s, 3H), 7.26(m, 3H), 7.50(m, 1H), 7.56(s, 1H), 7.73(d, J=8.4 Hz, 2H), 9.08 (s, 1H), 11.46 (m, 1H), 12.04 (m, 1H)

Example 2. Measurement of anticancer activity of thiobenzimidazole derivatives

2.1 Confirmation of cell viability of thiobenzimidazole derivatives

Human triple-negative breast cancer (TNBC) cell line MDA-MB-231 (Cell seeding numbers: 1(M231) x 10 ⁴ cells / wells (confluency ≥ 25%) and HER2 positive breast cancer , HER2+ BC) cell line JIMT-1 (Cell seeding numbers: 1.2(JIMT) x 10 ⁴ cells / wells (confluency ≥ 25 %)) was used for the experiment.

The cell lines were prepared in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS), streptomycin-penicillin (100 U/mL) and Fungizone (0.625 μg/mL), respectively, in 5% CO ₂ , 37 °C environment. cultured.

Formulas 1-1, 1-2, 1-3, 1-5, 1-6, 1-7, 1-8, 1-10, 1-11 in the human breast cancer cell lines MDA-MB-231 and JIMT-1 , 1-12, 1-13, 1-14, 1-15, 1-16, 1-17, 1-19, 1-20, 1-21, and 1-22 of the thiobenzimidazole derivatives 0 , 0.5, 1, 5 μM after treatment for 72 hours at various concentrations, the cell viability was measured by the MTS assay technique. MTS assay was performed by attaching cells to a 96-well plate for 24 hours, then treating the thiobenzimidazole derivative for 72 hours, and MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-) After color development with carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) for 4 hours, absorbance was measured at 490 nm using a Spectramax Plus384 microplate analyzer.

The measurement results are shown in FIGS. 1 to 6 . 1, 3, and 5 confirm cell viability for the MDA-MB-231 cell line, and FIGS. 2, 4, and 6 confirm cell viability for the JIMT-1 cell line.

As a result, in the human breast cancer cell lines MDA-MB-231 and JIMT-1, Chemical Formulas 1-1, 1-2, 1-3, 1-5, 1-6, 1-7, 1-8, 1-10, Thiobenziimidazole derivatives of 1-11, 1-12, 1-13, 1-14, 1-15, 1-16, 1-17, 1-19, 1-20, 1-21, and 1-22 It was confirmed that most of them inhibited cell viability in a concentration-dependent manner.

Additionally, HER2-positive breast cancer cell lines sensitive to trastuzumab, SKBR3 and BT474, and TRAstuzumab-resistant HER2-positive breast cancer cell line, JIMT-1, were treated with the thiobenzimidazole derivative of Formula 1-3 in a concentration-dependent manner for 72 hours. It was confirmed that all three cell lines showed a survival inhibitory effect (FIG. 7).

2.2 Confirmation of cell cycle arrest and apoptosis by thiobenzimidazole derivatives

The degree of apoptosis of cancer cells induced by the thiobenzimidazole derivative of Formula 1-3 was measured through DNA content analysis using flow cytometry. For SKBR3, BT474 and JIMT-1 cell lines, control (DMSO), a derivative of Formula 1-3 at concentrations of 0.1, 0.25, and 0.5 μM for 72 hours, the cells were harvested, and 0.5% for 24 hours It was fixed with 95% ethanol containing Tween-20 and stained with propidium iodide (PI, 50 μg/mL) and RNase (50 μg/mL) for 30 minutes. Then, the degree of apoptosis of cancer cells was analyzed using a flow cytometer.

In general, the cell cycle is divided into G1 (cell growth phase)-S (cell replication phase)-G2/M (cell division phase) depending on the amount of DNA in the cell, and when apoptosis is induced, DNA fragmentation phenomenon (DNA fragmentation), the DNA content in each cell is significantly lower than that in the G1 phase. The result of such apoptosis is shown on the cell cycle as a Sub G1 region, and the ratio of Sub G1 is expressed as a numerical value and is shown in FIG. 8 .

As a result, it was confirmed that the derivative of Formula 1-3 significantly induced apoptosis (Sub-G1 population) in SKBR3, BT474, and JIMT-1 breast cancer cell lines, and stagnated the G2/M phase cell cycle. The apoptosis experiment was independently performed three times, and the significance was verified by the unpaired Students t-test (*p<0.01; DMSO control vs NCT-58 or NCT-407).

In addition, in order to confirm the apoptosis-inducing effect of the derivative of Formula 1-3, control (DMSO), the derivative of Formula 1-3 was treated at concentrations of 0.1, 0.25, and 0.5 μM for 72 hours, followed by Annexin V/ The degree of apoptosis was confirmed by PI staining. As a result, it was confirmed that early and late apoptosis was effectively induced in a concentration-dependent manner by treatment with the derivative of Formula 1-3 (FIG. 9).

Example 3. Preparation of hydrochloride salt of thiobenzimidazole derivative

The synthesis process of Compound 1-23, which is the hydrochloride salt of the thiobenzimidazole derivative of Formula 1-3, is shown in Scheme 23 below.

[Scheme 23]

In a suspension of methyl (5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate (370 mg, 0.93 mmol) in methanol (75 mL) Hydrogen chloride (HCl) gas was injected at an interval of 2 minutes until pH was 1 or less. After the reaction product was evaporated under reduced pressure to obtain an oily product, isopropyl alcohol (Isopropyl alcohol, 37 mL) was added thereto, dissolved by heating, and then cooled to room temperature to form a solid suspension. Then, isopropyl ether (Isopropyl ether, 35 mL) was added to the solid suspension in portions, stirred for 3 hours, and then filtered. At this time, wash with IPA and IPE mixed solvent and dry with hot air at 50 ℃ for 1 hour, 40 Methyl(5-((4-(4-methylpiperazin-1-yl)phenyl)thio)-1H-benzo[d]imidazol-2-yl)carbamate hydrochloric acid salt (Formula 1 -23, off-white solid, 350 mg) was obtained (yield 87%).

¹ H NMR (D ₂ O, 400 MHz) δ 2.93(s, 3H), 3.08-3.14 & 3.19-3.25 (2m, 4H), 3.61 (d, J = 12.5 Hz, 2H), 3.64 (d, J = 12.5 Hz, 2H), 3.89 (s, 3H), 7.04-7.44 (m, 7H)

Example 4. Measurement of anticancer activity of hydrochloride salt of thiobenzimidazole derivatives

4.1 Confirmation of cell viability of hydrochloride salt of thiobenzimidazole derivatives

In BT474, a HER2-positive breast cancer cell line sensitive to trastuzumab, and JIMT-1, a HER2-positive breast cancer cell line resistant to trastuzumab, the thiobenzimidazole derivative of Formula 1-3 and its hydrochloride (Compound 1-23) were mixed at the same concentration ( 0, 0.1, 0.5, 1, 5, 10 μM) for 72 hours, cell viability and IC ₅₀ were measured by MTS assay. As a result, in the BT474 cell line, the IC ₅₀ of the derivative of Formula 1-3 was 2.184 μM and the IC ₅₀ of the hydrochloride (Compound 1-23) was 0.347 μM, which was 6 times lower, and in the JIMT-1 cell line, the The IC ₅₀ of the derivative of Formula 1-3 was 0.448 μM, and the IC ₅₀ of the hydrochloride salt (Compound 1-23) was 0.228 μM, which was almost half of the value ( FIG. 10 ).

In addition, in the triple-negative breast cancer cell lines MDA-MB-231, BT549, and 4T1, the hydrochloride salt (Compound 1-23) of the thiobenzimidazole derivative of Formula 1-3 prepared above was added at different concentrations (0, 0.1, 0.5, 1). , 5, 10 μM) for 72 hours, and then the cell viability and IC ₅₀ were measured by MTS assay. As a result, the three cell lines exhibited low IC ₅₀ of 0.523 μM, 0.559 μM, and 0.267 μM, respectively ( FIG. 11 ).

The above results suggest that hydrochloride (Compound 1-23) has improved solubility than the derivative of Formula 1-3, and thus has more excellent cell viability inhibition ability.

4.2 Confirmation of cell cycle arrest of hydrochloride salts of thiobenzimidazole derivatives

In the HER2-positive breast cancer cell line JIMT-1 and the triple-negative breast cancer cell line MDA-MB-231, the hydrochloride salt of the thiobenzimidazole derivative of Formula 1-3 (Compound 1-23) was added by concentration (0, 0.1, 0.25, 0.5 μM). After each treatment for 72 hours, western blot was performed.

As a result, the hydrochloride salt of the present invention decreases β-tubulin, a major target protein of benzimidazole-based anthelmintics, and increases p -Histone H3 (S10), a G2/M cell cycle marker, according to inhibition of tubulin synthesis. was confirmed (FIG. 12).

4.3 Confirmation of anticancer activity of hydrochloride salt of thiobenzimidazole derivatives in other cancer cell lines

In addition to the breast cancer cell line, the hematologic cancer cell line HL-60, the colon cancer cell line HCT116, the non-small cell lung cancer cell lines H1299 and A549, the ovarian cancer cell line SKOV3, the prostate cancer cell line Du145, and the liver cancer cell line HepG2 After treatment with the hydrochloride salt of a thiobenzimidazole derivative (Compound 1-23) at each concentration (0, 0.1, 0.25, 0.5 μM) for 72 hours, cell viability and IC50 were measured. As a result, excellent cells in other cancer cell lines as well It was confirmed that there is an effect of inhibiting the survival rate (FIG. 13).

As described above, although the embodiments have been described with reference to the limited drawings, those skilled in the art may apply various technical modifications and variations based on the above. For example, the described techniques are performed in an order different from the described method, and/or the described components of the system, structure, apparatus, circuit, etc. are combined or combined in a different form than the described method, or other components Or substituted or substituted by equivalents may achieve an appropriate result.

Therefore, other implementations, other embodiments, and equivalents to the claims are also within the scope of the following claims.

Claims (12)

A thiobenzimidazole derivative represented by the following [Formula 1] or a pharmaceutically acceptable salt thereof:
[Formula 1]

In Formula 1,
R ₁ is -SR ₂ or -SSR ₂ ,
R ₂ is a substituted or unsubstituted C ₃ to C ₂₀ aryl group, a substituted or unsubstituted C ₃ to C ₂₀ heteroaryl group, and a substituted or unsubstituted C ₃ to C ₂₀ alkylaryl group from the group consisting of whichever one is chosen.
The method of claim 1,
In the above [Formula 1], a substituted aryl group, a heteroaryl group, or an alkylaryl group is a substituted or unsubstituted C ₃ to C ₁₂ heterocycloalkyl group, a substituted or unsubstituted C ₃ to C ₁₂ heteroaryl group, and a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, characterized in that it is substituted with any one or more selected from the group consisting of a halogen group.
The method of claim 1,
The thiobenzimidazole derivative represented by the [Formula 1] is a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, characterized in that any one selected from the group consisting of the following compounds:
[Formula 1-1]

[Formula 1-2]

[Formula 1-3]

[Formula 1-4]

[Formula 1-5]

[Formula 1-6]

[Formula 1-7]

[Formula 1-8]

[Formula 1-9]

[Formula 1-10]

[Formula 1-11]

[Formula 1-12]

[Formula 1-13]

[Formula 1-14]

[Formula 1-15]

[Formula 1-16]

[Formula 1-17]

[Formula 1-18]

[Formula 1-19]

[Formula 1-20]

[Formula 1-21]

and
[Formula 1-22]

.
The method of claim 1,
The thiobenzimidazole derivative represented by the [Formula 1] is a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, characterized in that any one selected from the group consisting of the following compounds:
[Formula 1-1]

[Formula 1-2]

[Formula 1-3]

[Formula 1-5]

[Formula 1-6]

[Formula 1-7]

[Formula 1-8]

[Formula 1-10]

[Formula 1-11]

[Formula 1-12]

[Formula 1-13]

[Formula 1-14]

[Formula 1-15]

[Formula 1-16]

[Formula 1-17]

[Formula 1-19]

[Formula 1-20]

[Formula 1-21]

and
[Formula 1-22]

.
According to claim 1,
The thiobenzimidazole derivative is a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, characterized in that it inhibits tubulin polymerization.
The method of claim 1,
The pharmaceutically acceptable salts of the thiobenzimidazole derivatives include hydrochloride, bromate, sulfate, phosphate, nitrate, citrate, acetate, lactate, tartrate, maleate, gluconate, succinate, formate, and trifluoroacetic acid. salt, oxalate, fumarate, glutarate, adipate, methanesulfonate, benzenesulfonate, paratoluenesulfonate, camphorsulfonate, sodium salt, potassium salt, lithium salt, calcium salt, and magnesium salt A thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, characterized in that at least one selected from the group.
The method of claim 1,
The pharmaceutically acceptable salt of the thiobenzimidazole derivative is a thiobenzimidazole derivative or a pharmaceutically acceptable salt thereof, characterized in that the hydrochloride salt.
(1) preparing a suspension of the thiobenzimidazole derivative represented by the following [Formula 1];
[Formula 1]

(2) injecting hydrogen chloride (HCl) into the suspension and evaporating under reduced pressure;
(3) adding isopropyl alcohol to the product of step (2), heating and cooling; and
(4) adding isopropyl ether to the product of step (3), stirring, and then filtering;
As a method for producing a hydrochloride salt of thiobenzimidazole, comprising:
In Formula 1,
R ₁ is -SR ₂ or -SSR ₂ ,
R ₂ is a substituted or unsubstituted C ₃ to C ₂₀ aryl group, a substituted or unsubstituted C ₃ to C ₂₀ heteroaryl group, and a substituted or unsubstituted C ₃ to C ₂₀ alkylaryl group from the group consisting of Any one selected, the manufacturing method.
A pharmaceutical composition for preventing or treating cancer, comprising the thiobenzimidazole derivative according to any one of claims 1 to 7, or a pharmaceutically acceptable salt thereof, as an active ingredient.
10. The method of claim 9,
The pharmaceutical composition is a pharmaceutical composition for preventing or treating cancer, characterized in that it induces apoptosis by stopping the cancer cell cycle (cell cycle arrest).
10. The method of claim 9,
The cancer is skin cancer, breast cancer, uterine cancer, esophageal cancer, stomach cancer, brain tumor, colon cancer, rectal cancer, colorectal cancer, lung cancer, ovarian cancer, cervical cancer, endometrial cancer, vulvar cancer, kidney cancer, blood cancer, pancreatic cancer, prostate cancer, testicular cancer , laryngeal cancer, head and neck cancer, thyroid cancer, liver cancer, bladder cancer, osteosarcoma, lymphoma, blood cancer, thymus cancer, urethral cancer, and a pharmaceutical composition for cancer prevention or treatment, characterized in that any one or more selected from the group consisting of bronchial cancer.
10. The method of claim 9,
The cancer is a pharmaceutical composition for preventing or treating cancer, characterized in that triple-negative breast cancer.

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