KR20110115048A - New arylsulfonylimidazolone derivatives and an anti-cancer pharmaceutical composition comprising the same - Google Patents

Abstract

(식에서, R은 CH₃ 또는 CH(CH₃)₂이고, R'는 CH₂CH(CH₃)₂ 또는 CH₂Ph이다.)The present invention relates to an arylsulfonylimidazolone derivative compound having an enhanced water solubility and anticancer target represented by Formula I or a pharmaceutically acceptable salt thereof, a method for preparing the same, and an anticancer composition containing the same:
Formula I

(Wherein R is CH ₃ or CH (CH ₃ ) ₂ and R 'is CH ₂ CH (CH ₃ ) ₂ or CH ₂ Ph.)

Description

New arylsulfonylimidazolone derivatives and an anticancer agent composition containing the same {New arylsulfonylimidazolone derivatives and an anti-cancer pharmaceutical composition comprising the same}

The present invention relates to an arylsulfonylimidazolone derivative compound or a pharmaceutically acceptable salt thereof having increased water solubility and cancer cell targeting, a method for preparing the same, and an anticancer agent composition containing the same.

Sulfonylurea compounds exhibit various physiological activities such as hypoglycemic action, herbicides and antifungal agents. In particular, diarylsulfonyl urea (diarylsulfonylurea (DSU) has been reported to exhibit anticancer activity. The compound also expressed significant activity against solid cancer models that were not well treated by existing anticancer agents, and then modified the structure of the compound to further enhance sulofenur (Sulofenur, LY186641, J. Med. Chem., 1990, 33, 2393). Although these diarylsulfonyl urea derivative compounds showed excellent anticancer effects in transplantation of colorectal cancer xenografts, the development of anticancer drugs was stopped due to side effects such as methemoglobinemia and hemolytic anemia. Was found to be due to the ρ-chloroaniline derivative, a metabolite of the DSU material.

U.S. Patent No. 5,929,103 discloses a new arylsulfonylimidazolone-based compound that has superior anticancer activity than the sulfofenur and does not produce aniline-based metabolites that cause the aforementioned side effects. These arylsulfonylimidazolone compounds are antimitotic agents that express toxicity by inhibiting tubulin polymerization and resting the cell cycle in the G2 / M phase. According to the present invention, it shows excellent cytotoxicity against various cancer cells (HCT116, A-549, NCI-H460), and some derivatives have not been shown to express the multi-resistance of the paclitaxel or vinca alkaloid anticancer drugs currently used in the clinic. (See Patent Application No. 10-2008-0013100, Reference Experiment Example). Applicants conducted a preclinical study of the compounds (e.g., the following Formula II compounds or JSH2282) showing particularly good anticancer effects, and as a result, a large portion of the administered drug is deposited in the intestine of the experimental animal, causing toxicity. It was discovered that the development as a new drug was stopped. In addition, the solubility in water was very low at 23 μg / mL, making it impossible to use as an intravenous injector, and the intestinal toxicity was intensified due to poor dissolution and absorption of intestinal drugs.

In this regard, the present applicant has conducted a study to solve the problem of poor solubility of water of the arylsulfonylimidazolone-based compound and its intestinal toxicity, and thus a novel arylsulfonylimidazolone-based compound having a markedly increased water solubility. Was developed and filed as a patent application No. 10-2008-0013100.

On the other hand, anti-cancer substances used for cancer treatment show cytotoxicity not only to cancer cells but also to normal cells, thereby showing side effects. Therefore, studies have been made to minimize side effects by increasing the selectivity of cancer cells for anticancer drugs, and further maximize the anticancer activity.

U.S. Patent 5,929,103, Patent Application 10-2008-0013100

J. Med. Chem., 1990, 33, 2393

The present invention shows a very excellent effect on anticancer activity and resistance, but by solving the problems of the conventional arylsulfonyl imidazolone-based compound that its use is limited due to side effects such as enterotoxicity due to poor water solubility The present invention aims to provide a new arylsulfonylimidazolone-based compound which can increase intracellular solubility and can be administered intravenously, has no risk of enterotoxicity, and minimizes side effects by maximizing targeting to cancer cells.

The present invention relates to an arylsulfonylimidazolone compound of Formula I or a pharmaceutically acceptable salt thereof, a method for preparing the same, and an anticancer composition containing the same as an active ingredient. :

Formula I

Wherein R is CH ₃ or CH (CH ₃ ) ₂ and R 'is CH ₂ CH (CH ₃ ) ₂ or CH ₂ Ph. Ph here refers to phenol.

Especially preferred are compounds of formula I, wherein R is CH (CH ₃ ) ₂ , R 'is CH ₂ CH (CH ₃ ) ₂ , and compounds of formula I, wherein R is CH ₃ and R' is CH ₂ Ph.

The compound of formula I of the present invention is prepared according to Reaction Scheme 1 below, specifically, using a PABA (p-amino benzyl alcohol) as a linker to the compound of formula II as a peptide group D-Val-Leu-Lys- or D It is prepared by combining -Ala-Phe-Lys-.

[Reactivity 1]

Thus prepared formula I compound of the present invention is efficiently transformed into a formula II compound, a parent drug that exhibits anticancer activity by plasmin, a protease that specifically expresses cancer cells upon administration to the cancer cells. Targetability is increased.

The formula I compound of the present invention has a marked increase in water solubility of 450 to 580 times compared to the formula II compound of the parent drug, and exhibits high water solubility in water of 10 mg / ml or more, and thus can be easily formulated as an injection. This effectively improves the poor solubility in water, which is a problem of the conventional arylsulfonylimidazolone compounds, facilitates formulation into intravenous injections, and can effectively avoid the problems of enterotoxicity due to oral administration. have.

The preparation in the form of prodrugs as described above is a useful method for overcoming physicochemical (eg, solubility, lipophilic) or biological (eg, bioavailability) limitations in vivo. In the design of prodrugs, the desired prodrugs can be synthesized, have good stability at room temperature or in vitro (such as good half-life), and can be converted into parent drugs in a timely and fully in vivo manner. Shall be allowed. In addition, prodrugs are usually produced only by enzymatic reactions in vivo to satisfy all of the above conditions, and in particular, the compound of formula I of the present invention is plasmin, which is one of the proteases that specifically expresses cancer cells. Selectively disintegrates. Plasmin is present in the body, most of plasminogen inactive-type chair Imogen form, α ₂ present in vivo-anti-plasmin and α ₂ - is rapidly inhibited by macroglobulin, the u-PA that are overexpressed in cancer cells Is an enzyme that is activated locally. Since the compound of formula I of the present invention is more degraded by plasmin in cancer cells than in blood, it is present in the form of a relatively inactive prodrug in the blood, and is then regenerated into a formula II compound, which is the parent drug in cancer cells. It exhibits strong anticancer activity in cancer cells, and as a result, the compounds of the present invention significantly increase the selectivity for cancer cells and reduce side effects.

Compounds of the present invention are lung cancer, colon cancer, rectal cancer, colon cancer, breast cancer, cervical cancer, endometrial cancer, fallopian tube carcinoma, ovarian cancer, vaginal carcinoma, vulvar carcinoma, liver cancer, gastric cancer, esophageal cancer, small intestine cancer, pancreatic cancer, gallbladder cancer, kidney cancer , Bladder cancer, urethral cancer, penile cancer, prostate cancer, testicular cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, non-small cell lung cancer, bone cancer, skin cancer, head or neck cancer, skin or eye melanoma, Hodgkin's disease, endocrine May be used for the treatment of cancers such as adenocarcinoma, chronic or acute leukemia, lymphocytic lymphoma, central nervous system tumor, spinal cord tumor, brain stem glioma and pituitary adenoma, preferably lung cancer, colorectal cancer, rectal cancer, uterine cancer, ovarian cancer, or It can be used for the treatment of leukemia.

In addition, the compounds of the present invention can also be used for the treatment of all cancer diseases caused by cancer cells resistant to existing anticancer agents.

In addition, the compounds of the present invention are particularly effective in the treatment of cancer diseases caused by cancer cells expressing plasmin.

As used herein, "pharmaceutically acceptable salts" means hydrochloride, hydrobromide, hydroiodide, hydrogen fluoride, sulfate, sulfonate, citrate, camphorate, maleate, acetate, lactate, nicotinate, Nitrate, Succinate, Phosphate, Malonate, Dried salt, Salicylate, Phenyl acetate, Stearate, Palmitate, Pyridine, Ammonium, Piperazine, Diethylamine, Nicotinamide, Formate, Fumarate, Urea, Sodium Potassium, Calcium, Magnesium, Zinc, Lithium, Cinnamic Acid, Methylamino, Methanesulfonate, Pycholate, P-Toluenesulfonate, Naphthalenesulfonate, Tartarate, Triethylamino, Dimethylamino, and Tri (hydroxymethyl Salts commonly used in the pharmaceutical arts, such as aminomethane, with hydrochloride particularly preferred.

Those skilled in the art will appreciate that the compounds of the present invention contain one or more asymmetric carbon atoms and therefore include enantiomers or diastereomers. Diastereomeric mixtures can be separated into individual diastereomers on the basis of their physicochemical differences by known methods such as chromatography, recrystallization or fractional crystallization, and enantiomers can be used to convert enantiomeric mixtures into suitable optically active compounds. Can be converted into diastereomeric mixtures and the diastereomers are separated and separated in conventional manner by converting each diastereomer to the corresponding pure enantiomer. Diastereomers, enantiomers and mixtures thereof are considered part of the present invention, wherein in the formula I, the position where phenyl is attached to the imidazolidinone is in the S configuration, and the amino acids are all L-amino acids.

The composition of the present invention may be an aqueous solution or the powder itself, or may be a pharmaceutical composition of a suitable formulation prepared using known additives such as excipients, binders or lubricants as necessary.

In one embodiment of the invention, in the preparation of injectables, the powder comprising the compound of formula I as an active ingredient is readily dissolved in an aqueous isotonic solution prepared using distilled water or sodium chloride and sugars (e.g., glucose, mannitol, inositol, etc.). do. After dissolution, the injection containing the active ingredient may be administered intravenously, intramuscularly, subcutaneously, or directly to a lesion or tumor cut such as a tumor at an organ with an effective drug concentration in vivo for the disease to be treated.

As another example of the present invention, when preparing oral preparations, it may be prepared in tablets, capsules, granules, granules, binders, liquids and the like. In the preparation of these formulations, it may further comprise inert ingredients, including pharmaceutically acceptable carriers. As used herein, a "pharmaceutically acceptable carrier" is a term that refers to a composition, specifically an ingredient other than the active substance of the pharmaceutical composition. Examples of pharmaceutically acceptable carriers include binders, disintegrants, diluents, fillers, glidants, excipients, lubricants, dispersants, stabilizers, colorants, absorption enhancers, solubilizers or emulsifiers and salts.

In addition, sugars, preservatives, stabilizers, antistatic agents, and the like can be added to improve the properties of the powder obtained (packing capacity in storage containers or vials, specific amounts, etc.).

The powder may also be formulated in preparations other than injections or oral preparations according to conventional methods. Examples of such agents are preparations, transdermal and implants administered to the mucous membranes of the nose, mouth, sublingual, rectum, vagina or uterus. Each of the above formulations may be molded into various controlled release or targeted therapeutic agents, and the compositions of the present invention may be used as raw materials for such formulations.

As described above, the formula I compound of the present invention solves the poor solubility problem of water of the conventional arylsulfonylimidazolone-based compounds, which allows for intravenous administration, and therefore, rapid systemic circulation immediately after intravenous administration. It is relatively safe for the living body because it enables drug delivery, does not express enterotoxicity, and in particular, increases targetability to cancer cells, thereby minimizing side effects. Thus, various formulations such as injections, oral preparations, preparations administered to the oral cavity (e.g., troches, oral preparations), sublingual agents, eye drops, syrups, external preparations administered to the skin, nasal preparations, preparations administered through the lungs, Formulations applied to rectal suppositories or mucous membranes are very useful in pharmaceutical or veterinary compositions and as anticancer agents for use in humans or non-human mammals (eg, monkeys, cattle, dogs, etc.).

Hereinafter, experimental examples and production examples for confirming the effect of the compound of the present invention will be described. However, the scope of the present invention is not limited thereto.

Therefore, the novel arylsulfonylimidazolone compounds of the present invention exhibit excellent water solubility and can be prepared as injections, thereby avoiding the fear of enterotoxicity, which is a problem of conventional systemic anticancer agents, and inducing cancer specific expression enzymes in vivo. It is selectively regenerated by phosphorus plasmin to increase the target for cancer cells, thereby minimizing side effects and exerting an excellent anticancer effect.

1 and 2 show the conversion rate of the Examples 1 and 2 of the present invention to the parent drug in human plasma, respectively.
3 and 4 show the conversion rate to the parent drug in the human plasmin of Example compounds 1 and 2 of the present invention, respectively.

Synthetic example : Reactant Alloc Tripeptide PABA Synthesis of

Reactants Alloc- tripeptide-PABA used in the preparation of the derivative compounds of the present invention is a publication such as de Groot Synthesis and biological evaluation of novel prodrugs of anthracyclines for selective activation by the tumor-associated protease plasmin. J. Med . Chem . 42 , 5277-5283 (1999) and Synthesis and biological evaluation of 2'-carbamate-linked and 2'-carbonate-linked prodrugs of paclitaxel: Selective activation by the tumor-associated protease plasmin. J. Med . Chem . 43 , 3093-3102 (2000), according to the method described in the following reaction scheme 2 was used.

[Reactivity 2]

Synthetic example  One: Alloc -D- Val - Leu - Lys ( Alloc ) - PABA  Synthesis of 14a

D- ( N - tert - Butoxycarbonylvalyl ) -L-leucine methyl  Ester (7a)

Reagents and conditions; a) HOBt / H ₂ O, DCC, TEA, CH ₂ Cl ₂ / DMF

Under nitrogen gas, D- ( N- Boc) -valine (5a) (5.0 g, 23 mmol), leucine methyl ester hydrochloride (6a) (4.2 g, 23 mmol), 1-hydroxybenzotriazole (HOBt) (3.2 g, 23 mmol) was dissolved in dry DMF (35 mL) and stirred at 0 ° C. for 30 min. Triethylamine (3.2 mL, 23 mmol) was slowly added dropwise to this mixed solution. Dicyclohexylcarbodiimide (5.3 g, 25 mmol) was dissolved in this mixed solution in 55 mL of CH ₂ Cl ₂ , and then slowly added dropwise, followed by stirring at 0 ° C. for 4 hours and at room temperature for 24 hours. After completion of the reaction, dicyclohexylurea was filtered through a filter paper, and the filtrate was concentrated under reduced pressure. Then diluted with EtOAc (50 mL), extracted three times with 10% citric acid (30 mL), saturated NaHCO ₃ (30 mL), brine (30 mL), and the organic layer was dehydrated with anhydrous and concentrated under reduced pressure to give a white solid 7a (7.53). g, 95.0%).

mp: 89.0-90.0 ° C

FT-IR (cm ⁻¹ ,) 3317, 2957, 1647, 1539, 1172;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 0.94 (m, 12H), 1.45 (s, 9H), 1.54-1.67 (m, 3H), 2.19 (m, 1H), 3.73 (s, 3H) , 3.97 (m, 1H), 4.62 (m, 1H), 5.04 (d, J = 8.4 Hz, 1H), 6.45 (d, J = 8.0 Hz, 1H).

D- ( N - tert - Butoxycarbonylvalyl ) -L-leucine (8a)

Reagents and conditions; b) 1N-NaOH, acetone / DMF

Under nitrogen gas, D- ( N - tert -butoxycarbonylvalyl) -L-leucine methylester (7a) (7 g, 20 mmol) is dissolved in acetone / DMF (120 mL + 20 mL) and 1 M NaOH (10 mL) is added dropwise. After stirring at room temperature for 6 hours. After completion of the reaction, the solvent was concentrated under reduced pressure, dissolved in distilled water (100 mL), and then washed three times with EtOAc (50 mL). Citric acid was added dropwise to the distilled water layer and acidified to pH 2-3, followed by extraction with EtOAc (100 mL) dropwise. After layer separation, the organic layer was washed three times with distilled water (50 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to obtain a white solid 8a (5.83 g, 87.0%).

mp: 143.0-144.0 ° C.,

FT-IR (cm ⁻¹ ,) 3342, 2946, 1652, 1523, 1170;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 0.95 (m, 12H), 1.44 (s, 9H), 1.61-1.71 (m, 3H), 2.14 (m, 1H), 4.09 (m, 1H) , 4.62 (m, 1H), 5.37 (d, J = 8.4 Hz, 1H), 6.95 (d, J = 8.0 Hz, 1H).

D- ( N - tert - Butoxycarbonylvalyl ) -L- Leucinel -L- ( N ^ω - tert - Butoxycarbonyl ) Lysine methyl ester (10a)

Reagents and conditions; c) HOSu, NMM, DCC, TEA, CH ₂ Cl ₂ / DMF

Under nitrogen gas, D- ( N - tert -butoxycarbonylvalyl) -L-leucine (8a) (3.3 g, 10 mmol), N ^ω -BOC-lysine methyl ester hydrochloride (9) (3.0 g, 10 mmol) , N -hydroxypyrrolidine-2,5-dione (HOSu) (1.15 g, 10 mmol) was dissolved in dry DMF (35 mL) and stirred at 0 ° C. for 30 minutes. Triethylamine (1.4 mL, 10 mmol) and N -methylmorpholine (NMM) (1.1 mL, 10 mmol) were slowly added dropwise to this mixed solution. Dicyclohexylcarbodiimide (2.3 g, 11 mmol) was dissolved in this mixed solution in CH ₂ Cl ₂ (55 mL), and then slowly added dropwise, followed by stirring at 0 ° C. for 4 hours and at room temperature for 24 hours. After completion of the reaction, dicyclohexylurea was filtered through a filter paper, and the filtrate was concentrated under reduced pressure. Then diluted with EtOAc (50 mL), extracted three times with 10% citric acid (30 mL), saturated NaHCO ₃ (30 mL), brine (30 mL), and then the organic layer was dehydrated with anhydrous and concentrated under reduced pressure to give a white solid 10a (3.07). g, 53.7%).

mp: 147.0-148.0 ° C.,

FT-IR (cm ⁻¹ ,) 3270, 2957, 1646, 1521, 1167;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 0.91 (m, 12H), 1.25-2.18 (m, 28H), 3.10 (m, 2H), 3.69 (s, 3H), 3.87 (m, 1H) , 4.45-4.51 (m, 2H), 4.85-5.11 (m, 1H), 5.36-5.70 (m, 1H), 6.61-6.77 (m, 2H).

D- (N- tert - Butoxycarbonylvalyl ) -L- Leucinel -L- (N- tert - Butoxycarbonyl Lysine (11a)

Reagents and conditions: d) 1N-NaOH, acetone / DMF

Under nitrogen gas, D- (N - tert - butoxycarbonyl balril) -L- ryusi the carbonyl -L- (N ^ω - - tert-butoxycarbonyl) lysine methyl ester (10a) (3g, 5mmol) acetone / It was dissolved in DMF (60mL + 10mL), 1M NaOH (5mL) was added dropwise, and stirred at room temperature for 6 hours. After completion of the reaction, the solvent was concentrated under reduced pressure, dissolved in distilled water (50 mL), and then washed three times with EtOAc (30 mL). Citric acid was added dropwise to the distilled water layer and acidified to pH 2-3, followed by extraction with EtOAc (50mL) dropwise. After layer separation, the organic layer was washed three times with distilled water (30 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to obtain a white solid 11a (1.97 g, 67.3%).

mp: 63.0-64.0 ° C.,

FT-IR (cm ⁻¹ ,) 3341, 2940, 1652, 1522, 1166;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.81-0.88 (m, 12H), 1.17-1.68 (m, 28H), 2.65 (m, 1H), 3.35 (d, J = 5.6 Hz, 2H), 3.76 (m, 1H), 4.08 (m, 1H), 4.33 (m, 1H), 6.70 (d, J = 8.0 Hz, 1H), 6.79 (m, 1H), 8.03 (m, 2H), 12.30 (s , 1H).

H-D- Val - Leu - Lys - OH  (12a)

Reagents and conditions; e) TFA, CH ₂ Cl ₂

Under nitrogen gas, D- (N- tert - butoxycarbonyl balril) -L- ryusi carbonyl -L- (N ^ω - tert - butoxycarbonyl) lysine (11a) (1g, ₂ mmol) was dissolved in CH ₂ Cl ₂ (50 mL). TFA (10 mL) was slowly added dropwise to this mixed solution, followed by stirring at room temperature for about 6 hours. After completion of the reaction, the solvent was completely dried under reduced pressure and then crystallized using ether to obtain a white solid 12a (0.79 g, 75.2%).

mp: 85.0-86.0 ° C.,

FT-IR (cm ⁻¹ ,) 2957, 1652, 1187, 1136;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.84-0.94 (m, 12H), 1.33-1.70 (m, 9H), 2.07 (m, 1H), 2.75 (m, 2H), 3.63 (m, 1H ), 4.09 (m, 1H), 4.41 (m, 1H), 8.35 (d, J = 7.2 Hz, 1H), 8.60 (d, J = 8.0 Hz, 1H).

Alloc - ONSu

Reagents and Conditions: TEA, Acetonitrile

Under nitrogen gas, 1-hydroxysuccinimide (11.5 g, 100 mmol) and triethylamine (14 mL, 100 mmol) were dissolved in acetonitrile (100 mL) and stirred at 0 ° C. for 30 minutes. In this mixed solution, Alloc-Cl (11 mL, 100 mmol) was dissolved in acetonitrile (100 mL), and slowly added dropwise thereto, followed by stirring for 1 hour. After completion of the reaction, triethylammonium chloride was filtered and the filtrate was concentrated under reduced pressure. The crude product was dissolved in EtOAc (100 mL), washed three times with distilled water (50 mL) and 5% KHSO ₄ (50 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to give a clear liquid (18.1 g, 91.0%). Got it.

mp: 24.0-25.0 ° C.,

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 2.78 (s, 4H), 4.69-4.85 (m, 2H), 5.32 (dd, J = 9.2 Hz, 18.4 Hz, 1H), 5.46 (t, J = 18.0 Hz, 1H), 5.93 (m, 1H).

Alloc -D- Val - Leu - Lys ( Alloc ) - OH  (13a)

Reagents and conditions; f) Alloc-ONSu, TEA, acetonitrile / H ₂ O

Under nitrogen gas, D-Val-Leu-Lys-OH (12a) (3 g, 5.1 mmol) was dissolved in H ₂ O / acetonitrile (50 mL / 50 mL), and triethylamine (2.2 mL, 15.4 mmol) was added dropwise. The mixture was adjusted to pH 9.0-9.5 and stirred at room temperature for 30 minutes. In this mixed solution, Alloc-ONSu (2.23 g, 11.2 mmol) was dissolved in acetonitrile (20 mL), and then slowly added dropwise thereto. After stirring for 6 hours at room temperature, 0.5M HCl was added dropwise, acidified to pH 3.0, extracted three times with CH ₂ Cl ₂ (100 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to give a white solid 13a (1.6 g, 60.0%).

mp: 135.0-136.0 ° C.

FT-IR (cm ⁻¹ ,) 3320, 2957, 1652, 1538, 1246, 1165;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 0.89-0.97 (m, 12H), 1.26-2.09 (m, 10H), 3.14 (m, 2H), 3.95 (m, 1H), 4.45-4.62 ( m, 6H), 5.19-5.32 (m, 4H), 5.91 (m, 2H).

Alloc -D- Val - Leu - Lys ( Alloc ) - PABA  (14a)

Reagents and conditions; 4-aminobenzyl alcohol (PABA), NMM, i-BuOCOCl

Under a nitrogen gas, Alloc-D-Val-Leu-Lys (Alloc) -OH (13a) (450 mg, 0.85 mmol) was dissolved in anhydrous THF (30 mL) and stirred at -20 ° C for 30 minutes. Isobutylchloroformate (122 μL, 0.94 mmol) and N -methyl morpholine (104 μL, 0.94 mmol) were added dropwise to the mixed solution, followed by stirring at −20 ° C. for 3 hours. 4-aminobenzyl alcohol (132 mg, 1.07 mmol) and N -methyl morpholine (118 μL, 1.07 mmol) were dissolved in anhydrous THF (5 mL) and slowly added dropwise to the mixed solution. After dropping, the temperature was gradually raised and stirred at room temperature for 24 hours. After completion of the reaction, the mixture was concentrated under reduced pressure, diluted with CH ₂ Cl ₂ (50 mL), and washed three times with saturated NaHCO ₃ (30 mL), 0.5 N KHSO ₄ (30 mL), and brine (30 mL). The organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure. The crude product thus obtained was separated in a Si-gel column with a mixed solvent of n-HX: EtOAc = 1: 1 → n-HX: EtOAc: MeOH = 10: 10: 1 to obtain a white solid 14a (350 mg, 66.3%). .

mp: 153.0-154.0 ° C.,

FT-IR (cm ⁻¹ ,) 3320, 2957, 1652, 1538, 1366, 1165;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 0.93-1.08 (m, 12H), 1.46-2.15 (m, 10H), 3.09 (m, 2H), 3.70 (m, 1H), 4.07 (m, 1H), 4.37-4.62 (m, 6H), 5.06 (m, 2H), 5.22-5.33 (m, 5H), 5.60 (m, 1H), 5.91 (m, 1H), 7.32 (d, J = 7.2 Hz , 2H), 7.70 (d, J = 7.2 Hz, 2H).

Synthetic example  2: Alloc -D- Ala - Phe - Lys ( Alloc ) - PABA  (14b) synthesis

D- ( N - tert - Butoxycarbonylalanyl ) -L-phenylalanine methyl  Ester (7b)

Reagents and conditions; a) HOBt / H ₂ O, DCC, TEA, CH ₂ Cl ₂ / DMF

Under nitrogen gas, D- (N-Boc) -alanine (5b) (4.35 g, 23 mmol), phenylalanine methyl ester hydrochloride (6b) (4.95 g, 23 mmol), 1-hydroxybenzotriazole (HOBt) (3.2 g, 23 mmol) was dissolved in dry DMF (35 ml) and stirred at 0 ° C. for 30 minutes. Triethylamine (3.2 mL, 23 mmol) was slowly added dropwise to this mixed solution. Dicyclohexylcarbodiimide (5.3 g, 25 mmol) was dissolved in this mixed solution in CH ₂ Cl ₂ (55 mL), and then slowly added dropwise, followed by stirring at 0 ° C. for 4 hours and at room temperature for 24 hours. After completion of the reaction, dicyclohexylurea was filtered through a filter paper, and the filtrate was concentrated under reduced pressure. Then diluted with EtOAc (50 mL), extracted three times with 10% citric acid (30 mL), saturated NaHCO ₃ (30 mL), brine (30 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to give a white solid 7b (6.45). g, 80.1%).

mp: 92.0-93.0 ° C.,

FT-IR (cm ⁻¹ ,) 3338, 3302, 2982, 2934, 1660, 1519;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 1.29 (d, J = 7.2 Hz, 2H), 1.43 (s, 9H), 3.09-3.15 (m, 2H), 3.72 (s, 3H), 4.16 (m, 1H), 4.86 (m, 1H), 5.04 (m, 1H), 6.60 (m, 1H), 7.09-7.28 (m, 5H).

D- ( N - tert - Butoxycarbonylalanyl ) -L-phenylalanine (8b)

Reagents and conditions: b) 1N-NaOH, acetone / DMF

Under nitrogen gas, dissolve D- ( N - tert -butoxycarbonylalanyl) -L-phenylalanine methyl ester (7b) (6 g, 17 mmol) in acetone / DMF (120 mL + 20 mL) and dissolve 1 M NaOH (10 mL). After dropping, the mixture was stirred at room temperature for 6 hours. After completion of the reaction, the solvent was concentrated under reduced pressure, dissolved in distilled water (100 mL), and then washed three times with EtOAc (50 mL). Citric acid was added dropwise to the distilled water layer and acidified to pH 2-3 and then EtOAc (100 mL) was added dropwise to extract. After layer separation, the organic layer was washed three times with distilled water (50 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to obtain a white solid 8b (5.12 g, 88.7%).

mp: 55.0-56.0 ° C.,

FT-IR (cm ⁻¹ ,) 3313, 3246, 2958, 2873, 1700, 1635, 1388, 1167;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 01.27 (d, J = 7.2 Hz, 2H), 1.42 (s, 9H), 2.86-3.21 (m, 2H), 4.28 (m, 1H), 4.86 (m, 1H), 5.26 (d, J = 5.6 Hz 1H), 6.74 (m, 1H), 7.15-7.28 (m, 5H).

D- ( N - tert - Butoxycarbonylalanyl ) -L- Phenylalanyl -L- ( N - tert ^ω - Butoxycarbonyl ) Lysine methyl ester  (10b)

Reagents and conditions: c) HOSu, NMM, DCC, TEA, CH ₂ Cl ₂ / DMF

Under nitrogen gas, D- ( N - tert -butoxycarbonylalanyl) -L-phenylalanine (8b) (3.4 g, 10 mmol), N ^ω -BOC-lysinemethylester hydrochloride (9) (3.0 g, 10 mmol ), N -hydroxypyrrolidone-2,5-dione (HOSu) (1.15 g, 10 mmol) was dissolved in dry DMF (35 ml) and stirred at 0 ° C. for 30 minutes. Triethylamine (1.4 mL, 10 mmol) and N -methylmorpholine (NMM) (1.1 mL, 10 mmol) were slowly added dropwise to this mixed solution. Dicyclohexylcarbodiimide (2.3 g, 11 mmol) was dissolved in this mixed solution in CH ₂ Cl ₂ (55 mL), and then slowly added dropwise, followed by stirring at 0 ° C. for 4 hours and at room temperature for 24 hours. After completion of the reaction, dicyclohexylurea was filtered through a filter paper, and the filtrate was concentrated under reduced pressure. Then diluted with EtOAc (50 mL), extracted three times with 10% citric acid (30 mL), saturated NaHCO ₃ (30 mL), brine (30 mL), and then the organic layer was dehydrated with anhydrous and concentrated under reduced pressure to give a white solid 10b (2.79). g, 47.7%).

mp: 87.0-88.0 ° C.,

FT-IR (cm ⁻¹ ,) 3257, 3072, 2935, 1653, 1558, 1507;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.89 (d, J = 7.2 Hz, 3H), 1.23-1.69 (m, 24H), 2.71 (m, 1H), 2.88 (m, 2H), 3.06 ( m, 1H), 3.62 (s, 3H), 3.90 (m, 1H), 4.20 (m, 1H), 4.54 (m, 1H), 6.82 (m, 1H), 6.87 (d, J = 7.2 Hz, 1H ), 7.17-7.26 (m, 5H), 8.08 (d, J = 7.6 Hz, 1H), 8.29 (d, J = 6.8 Hz, 1H).

D- ( N - tert - Butoxacarbonylalanyl ) -L- Phenylalanyl -L- ( N ^ω - tert - Butoxycarbonyl Lysine (11b)

Reagents and conditions; d) 1N-NaOH, acetone / DMF

Under nitrogen gas, D- ( N - tert -butoxycarbonylalanyl) -L-phenylalanyl-L- ( N ^ω - tert -butoxycarbonyl) lysinemethyl ester (10b) (2.5 g, 4.3 mmol ) Was dissolved in acetone / DMF (60 mL + 10 mL), 1 M NaOH (5 mL) was added dropwise, and the mixture was stirred at room temperature for 6 hours. After completion of the reaction, the solvent was concentrated under reduced pressure, dissolved in distilled water (50 mL), and then washed three times with EtOAc (30 mL). Citric acid was added dropwise to the distilled water layer and acidified to pH 2-3, followed by extraction with EtOAc (50mL) dropwise. After separation of the layers, the organic layer was washed three times with distilled water (30 mL), and the organic layer was dehydrated with anhydrous manganese and concentrated under reduced pressure to obtain a white solid 11b (2.30 g, 94.2%).

mp: 121.0-122.0 ° C.,

FT-IR (cm ⁻¹ ,) 3299, 2969, 2933, 1729, 1647, 1538, 1165;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.89 (d, J = 6.8 Hz, 3H), 1.27-1.71 (m, 24H), 2.72 (m, 1H), 2.88 (m, 2H), 3.08 ( m, 1H), 3.90 (m, 1H), 4.14 (m, 1H), 4.55 (m, 1H), 6.82-6.89 (m, 2H), 7.17-7.28 (m, 5H), 8.08 (d, J = 7.6 Hz, 1H), 8.29 (d, J = 6.8 Hz, 1H), 12.28 (br, 1H).

H-D- Ala - Phe - Lys - OH  (12b)

Reagents and conditions; e) TFA, CH ₂ Cl ₂

Under nitrogen gas, D- ( N - tert -butoxacarbonylalanyl) -L-phenylalanyl-L- ( N ^ω -tert -butoxycarbonyl) lysine (11b) (2 g, 3.5 mmol) was dissolved in CH ₂ Cl ₂ (50 mL). TFA (10 mL) was slowly added dropwise to this mixed solution, followed by stirring at room temperature for about 6 hours. After completion of the reaction, the solvent was completely dried under reduced pressure and then crystallized using ether to give a white solid 12b (1.44g, 68.6%).

mp: 142.0-143.0 ° C.,

FT-IR (cm ⁻¹ ,) 3257, 3075, 2940, 1653, 1508, 1183, 1134;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.99 (d, J = 6.8 Hz, 1H), 1.33-1.75 (m, 6H), 2.65-2.77 (m, 3H), 3.09 (dd, J = 3.6 Hz, 13.6 Hz, 1H), 3.74 (m, 1H), 4.22 (q, J = 8.0 Hz, 13.2 Hz, 1H), 4.72 (m, 1H), 7.16-7.29 (m, 5H).

Alloc -D- Ala - Phe - Lys ( Alloc ) - OH  (13b)

Reagents and conditions; f) Alloc-ONSu, TEA, H ₂ O / acetonitrile

Under nitrogen gas, D-Ala-Phe-Lys-OH (12b) (3 g, 4.9 mmol) was dissolved in H ₂ O / acetonitrile (50 mL / 50 mL), and triethylamine (1.9 mL, 13.3 mmol) was added dropwise. The mixture was adjusted to pH 9.0-9.5 and stirred at room temperature for 30 minutes. In this mixed solution, Alloc-ONSu (1.97 g, 9.9 mmol) was dissolved in acetonitrile (20 mL), and then slowly added dropwise thereto. After stirring for 6 hours at room temperature, 0.5M HCl was added dropwise, acidified to pH 3.0, extracted three times with CH ₂ Cl ₂ (100 mL), and the organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure to give a white solid 13b (1.95 g, 74.0%).

mp: 139.0-140.0 ° C.,

FT-IR (cm ⁻¹ ,) 3271, 3081, 2941, 1637, 1540, 1237;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.92 (d, J = 7.2 Hz, 3H), 1.34-1.71 (m, 6H), 2.73 (m, 1H), 2.96 (m, 2H), 3.07 ( m, 1H), 3.97 (m, 1H), 4.14 (m, 1H), 4.45 (m, 4H), 4.56 (m, 1H), 5.14-5.28 (m, 4H), 5.88 (m, 2H), 7.16 -7.28 (m, 7H), 8.04 (d, J = 8.8 Hz, 1H), 8.16 (d, J = 7.6 Hz, 1H), 12.50 (s, 1H).

Alloc -D- Ala - Phe - Lys ( Alloc ) - PABA  (14b)

Reagents and conditions; 4-aminobenzyl alcohol (PABA), NMM, i-BuOCOCl

Under a nitrogen gas, Alloc-D-Ala-Phe-Lys (Alloc) -OH (13a) (455 mg, 0.85 mmol) was dissolved in anhydrous THF (30 mL) and stirred at -20 ° C for 30 minutes. Isobutylchloroformate (122 μL, 0.94 mmol) in this mixed solution And N -methylmorpholine (104 μL, 0.94 mmol) were added dropwise and stirred at −20 ° C. for 3 hours. 4-aminobenzyl alcohol (132 mg, 1.07 mmol) and N -methyl morpholine (118 μL, 1.07 mmol) were dissolved in anhydrous THF (5 mL) and slowly added dropwise to the mixed solution. After dropping, the temperature was gradually raised and stirred at room temperature for 24 hours. After completion of the reaction, the mixture was concentrated under reduced pressure, diluted with CH ₂ Cl ₂ (50 mL), and washed three times with saturated NaHCO ₃ (30 mL), 0.5 N KHSO ₄ (30 mL), and brine (30 mL). The organic layer was dehydrated with anhydrous forget-me-not and concentrated under reduced pressure. The crude product thus obtained was separated in a silica gel column with a mixed solvent of n-HX: EtOAc = 1: 1 → n-HX: EtOAc: MeOH = 10: 10: 1 to obtain a white solid 14b (245 mg, 44.9%).

mp: 159.0-160.0 ° C.,

FT-IR (cm ⁻¹ ,) 3271, 1635, 1522, 1255; ¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.96 (d, J = 6.8 Hz, 3H), 1.14-1.85 (m, 7H), 2.77-3.06 (m, 2H), 2.95 (d, J = 6.0 Hz, 2H), 4.00 (m, 1H), 4.41-4.53 (m, 8H), 5.07-5.25 (m, 4H), 5.84 (m, 2H), 7.21 (m, 7H), 7.37 (d, J = 7.2 Hz, 1H), 7.54 (d, J = 8.0 Hz, 2H), 8.10 (d, J = 7.2 Hz, 1H), 8.15 (d, J = 8.0 Hz, 1H), 9.77 (s, 1H).

Synthetic example  3: (S) -1- (1- (4-aminobenzolyl) Indolin -5- Ilsulfonyl )-4- Phenylimidazoline 2-one (formula II  compound)

Formula II compound used as a parent drug and starting material of the derivative compounds of the present invention is Jung, SH et al. Synthesis and evaluation of cytotoxicity of novel ary sufonylimidaxolidinones containing sulfonylurea pharmacophore. Arch . Pharm . Res . Prepared according to the preparation disclosed in 1996, 19 (6), 570-580. Hereinafter referred to as Formula II compound or JSH2282.

Example  1: D- Val - Leu - Lys - PABA - JSH2282  (·2 HCl ) 16a

Alloc -D- Val - Leu - Lys ( Alloc ) - PABA - JSH2282  (15a)

Under nitrogen gas, the formula II compound (100 mg, 0.22 mmol) was dissolved in anhydrous THF (30 mL), and 20% phosgene (114 μL, 0.22 mmol) in toluene was added dropwise. The mixture solution was refluxed for 2 hours, concentrated under reduced pressure, and then dissolved in toluene (30 mL), followed by Alloc-D-Val-Leu-Lys (Alloc) -PABA (14a) (134 mg,) prepared in Synthesis Example 1. 0.22 mmol) was added dropwise and refluxed for 3 hours. After completion of the reaction was concentrated under reduced pressure. The crude product thus obtained was separated in a silica gel column with a mixed solvent of n-HX: EtOAc = 1: 1 → n-HX: EtOAc: MeOH = 10: 10: 1 to obtain a white solid 15a (106 mg, 43.8%).

mp: 144.0-145.0 ° C.,

FT-IR (cm ⁻¹ ,) 3318, 2957, 1652, 1521, 1217, 1156, 1077;

¹ H-NMR (400 MHz, CDCl ₃ -d ₁ ) δ 0.93-1.08 (m, 12H), 1.42-2.17 (m, 10H), 3.06 (m, 2H), 3.15 (t, J = 8.4 Hz, 2H) , 3.31 (m, 1H), 3.66 (q, J = 7.6 Hz, 9.6 Hz, 1H), 3.80 (m, 1H), 4.14 (m, 2H), 4.28 (t, J = 8.8 Hz, 1H), 4.40 -4.61 (m, 4H), 4.74 (t, J = 8.0 Hz, 1H), 5.01-5.31 (m, 7H), 5.54-5.89 (m, 3H), 7.21-7.37 (m, 6H), 7.50-7.67 (m, 7H), 7.75 (d, J = 7.6 Hz, 1H), 7.87 (s, 1H), 8.61 (s, 1H).

D- Val - Leu - Lys - PABA - JSH2282  (·2 HCl ) (16a)

Reagents and conditions; Bu ₃ SnH, Pd (PPh ₃ ) ₄ , 0.5M HCl in methanol

Under nitrogen gas, Alloc-D-Val-Leu-Lys (Alloc) -PABA-JSH2282 (15a) (100 mg, 0.09 mmol) was dissolved in anhydrous THF and stirred at room temperature. Acetic acid (26 μL, 0.45 mmol), tributyl tin hydride (73 μL, 0.27 mmol), and Pd (PPh ₃ ) ₄ (10 mg) were added dropwise to this mixed solution, followed by stirring at room temperature for 1 hour, followed by tributyl tin hydride. (73μL, 0.27mmol) was further added dropwise and stirred again for 2 hours. To this mixed solution, 1.25M HCl (2 mL) in methanol was slowly added dropwise, stirred for 3 hours, and concentrated under reduced pressure. The crude product was crystallized with ether, and the obtained material was again dissolved in distilled water, followed by centrifugation (3000 rpm, 10 minutes), and the supernatant was lyophilized to obtain a pale yellow solid 16a (43 mg, 47.0%).

mp: 196.0-197.0 ° C.,

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.84-0.95 (m, 12H), 1.22-2.07 (m, 10H), 2.76 (m, 2H), 3.15 (t, J = 8.4 Hz, 2H), 3.49 (q, J = 7.6 Hz, 9.6 Hz, 1H), 3.67 (m, 1H), 4.12 (t, J = 8.4 Hz, 2H), 4.29 (t, J = 8.8 Hz, 1H), 4.38 (m, 2H), 4.79 (t, J = 8.0 Hz, 1H), 5.12 (s, 2H), 7.22 (d, J = 6.8 Hz, 2H), 7.31-7.38 (m, 5H), 7.58-7.67 (m, 6H ), 7.74-7.78 (m, 3H), 8.19 (s, 1H), 8.35 (d, J = 8.0 Hz, 1H), 8.79 (d, J = 7.6 Hz, 1H), 10.04 (s, 1H), 10.13 (s, 1H); FT-IR (cm ⁻¹ ,) 3313, 2957, 1652, 1521, 1386, 1218, 1157, 1073.

Example  2: D- Ala - phe - Lys - PABA - JSH2282 (·2 HCl ) Preparation of 16b

Alloc -D- Ala - Phe - Lys ( Alloc ) - PABA - JSH2282  (15b)

Reagents and Conditions: 1) phosgene, THF, reflux; 2) toluene, reflux

Under nitrogen gas, the formula II compound (100 mg, 0.22 mmol) was dissolved in anhydrous THF (30 mL), and 20% phosgene (114 μL, 0.22 mmol) in toluene was added dropwise. The mixed solution was refluxed for 2 hours, concentrated under reduced pressure, and then dissolved in toluene (30 mL), followed by Alloc-D-Ala-Phe-Lys (Alloc) -PABA (14b) (138 mg, prepared in Synthesis Example 2). 0.22 mmol) was added dropwise and refluxed for 3 hours. After completion of the reaction was concentrated under reduced pressure. The crude product thus obtained was separated in a silica gel column with a mixed solvent of n-HX: EtOAc = 1: 1 → n-HX: EtOAc: MeOH = 10: 10: 1 to obtain a white solid 15b (156 mg, 64.1%).

mp: 147.0-148 ° C.,

FT-IR (cm ⁻¹ ,) 3273, 1636, 1557, 1357, 1233, 1155;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.96 (d, J = 6.8 Hz, 3H), 1.22-1.85 (m, 6H), 2.77 (m, 1H), 2.95 (m, 2H), 3.14 ( m, 3H), 3.49 (q, J = 6.0 Hz, 10.0 Hz, 1H), 4.00 (m, 1H), 4.12 (t, J = 8.0 Hz, 2H), 4.27 (t, J = 7.2 Hz, 1H) , 4.37-4.43 (m, 5H), 4.50 (m, 1H), 4.80 (t, J = 6.8 Hz, 1H), 5.11-5.25 (m, 6H), 5.84 (m, 2H), 7.20-7.24 (m , 7H), 7.31-7.39 (m, 6H), 7.58-7.65 (m, 6H), 7.74-7.78 (m, 3H), 8.20 (m, 3H), 9.88 (s, 1H), 10.05 (s 1H) .

D- Ala - phe - Lys - PABA - JSH2282 (·2 HCl ) (16b)

Reagents and conditions; Bu ₃ SnH, Pd (PPh ₃ ) ₄ , 0.5M HCl in methanol

Under nitrogen gas, Alloc-D-Ala-Phe-Lys (Alloc) -PABA-JSH2282 (15b) (100 mg, 0.09 mmol) was dissolved in anhydrous THF and stirred at room temperature. Acetic acid (26 μL, 0.45 mmol), tributyl tin hydride (73 μL, 0.27 mmol) and Pd (PPh ₃ ) ₄ (10 mg) were added dropwise to this mixed solution, followed by stirring at room temperature for 1 hour, followed by tributyl tin hydride. (73 μL, 0.27 mmol) was further added dropwise, followed by further stirring for 2 hours. To this mixed solution, 1.25M HCl (2 mL) in methanol was slowly added dropwise, stirred for 3 hours, and concentrated under reduced pressure. The crude product was crystallized with ether, and the obtained material was again dissolved in distilled water, followed by centrifugation (3000 rpm, 10 minutes), and the supernatant was lyophilized to obtain a pale yellow solid 16b (40 mg, 43.7%).

mp: 192.0-193.0 ° C.,

FT-IR (cm ⁻¹ ,) 3277, 1636, 1522, 1375, 1247;

¹ H-NMR (400 MHz, DMSO- d ₆ ) δ 0.99 (d, J = 6.8 Hz, 3H), 1.22-1.80 (m, 6H), 2.74 (m, 3H), 3.14 (m, 3H), 3.47 ( q, J = 6.8 Hz, 9.2 Hz, 1H), 3.75 (m, 1H), 4.12 (t, J = 8.4 Hz, 2H), 4.26 (t, J = 9.2 Hz, 1H), 4.40 (m, 1H) , 4.77 (m, 2H), 5.12 (s, 2H), 7.15-7.39 (m, 13H), 7.57-7.98 (m, 9H), 8.18 (s, 1H), 8.50 (d, J = 8.0 Hz, 1H ), 8.64 (d, J = 8.8 Hz, 1H), 10.04 (s, 1H), 10.23 (s, 1H).

Experimental Example  1: solubility test

The solubility in distilled water was measured in comparison with the derivative compound of formula II of the present invention. Solubility was measured by Casini et al. (Casini, A .; Scozzafava, A .; Mincione, F .; Menabuoni, L .; Ilies, MA; Supuran, CT Carbonic Anhydrase Inhibitors: Water-Soluble 4-Sulfamoylphenylthioureas as Topical Intraocular Pressure-Lowering Agents with Long- Lasting Effects. J. Med. Chem. 2000, 43, 4884-92).

Dissolve 1 mg of each correctly weighed substance in MeOH (10 mL) for HPLC to make a standard sample, then UV maximum absorbance (UV absorption maxium, λ _max) for this standard sample. in UV spectrophotometer Mini 1240, Shimadzu, Japan) was determined. After making a saturated aqueous solution for each material using a small amount of distilled water (250 μL), the insolubles were removed by passing through a 0.45 μm syringe filter (Whatman, USA). The saturated aqueous solution thus prepared was scanned at the maximum absorption wavelength of each material determined using a standard sample to determine its absorbance. The solubility of each substance in water was determined by the following equation. The results are as shown in Table 1.

C '= (A' × C) ÷ A

From here,

C = concentration of standard solution (mg / mL),

A = absorbance of standard solution,

A '= absorbance of saturated solution,

C '= concentration of the aqueous solution (mg / mL).

compound Solubility (mg / mL) ^a Example 1 Compound 13.94 Example 2 Compound 10.79   Control compound 0.024

^a Solubility was measured at 25 ° C. in distilled water according to Casini et al.

As can be seen from Table 1, Example Compound 1 (16a) and Example Compound 2 (16b) of the present invention have a sharp increase in solubility of about 450, 580 times when compared to the control compound Formula II compound, the parent drug. Effect. Derivative compounds of the present invention incorporating PABA (paraaminobenzyl alcohol) linkers can be formulated as injections with a solubility in water of at least 10 mg / ml.

Experimental Example  2: In vitro  Confirmation of cytotoxicity against human cancer cells- On plasmin  Target identification

In particular, human breast cancer cell of MDA-MB-231 (high levels of plasmin) expressing high plasmin and human breast cancer cell expressing low plasmin of human breast cancer cells of derivative compounds of the present invention line, low levels of plasmin) was confirmed for its cytotoxicity. The cell lines were distributed and used by the Institute of Biotechnology, and their cytotoxicity values (IC ₅₀ , μM) were measured by MTT assay (Ming, Z .; Lanrong, B .; Wei, W .; Chao, W. Michele, BF; Jingfang, J .; Shiqi, P. Synthesis and cytoxic activities of β-carboline amino acid ester conjugates.Bioorg.Med.Chem. 2006, 14, 6998-7010; Brij, BS; Lei, Z .; Meirong.H .; Eileen, K .; Meena, K .; Ozgur, O .; Arkadiy, B .; Premila, R. Boc-lysinated-betulonic acid: A potent, anti-prostate cancer agent.Bioorg.Med.Chem 2006, 14, 6349-6358). The culture solution was a bag of DMEM medium containing L-glutamine in sterile injectable distilled water, 100 mL of bovine serum (FBS) inactivated by heat treatment in a 60 ° C water bath for 30 minutes, 3.7 g of NaHCO ₃ , and penicillin (10000 unit / mL). After dissolving, streptomycin (10 mg / mL) was added, the pH was adjusted to 0.1 L with 0.1 N hydrochloric acid, and prepared by bacterial filtration. Cells were maintained in subcultures every 3 days, using trypsin (2.5 g / L) and EDTA (0.38 g / L) in a solution of phosphate buffered saline to separate the cells from the adherent surface. It was.

The cells to be used for the experiment were separated from the attachment surface with trypsin-EDTA solution and seeded at 5 × 10 ³ cells per well of 96 well plates and incubated for 24 hours in a 5% CO ₂ incubator (37 ° C.). The sample was dissolved in dimethyl sulfoxide (DMSO) and diluted in stages to the concentration of the final DMSO to 0.2% or less by diluting with experimental medium or tertiary distilled water to the concentration required for the experiment. 2 μL of each diluted sample was added to each well of a 96 well plate and incubated for 72 hours in a 37 ° C., 5% CO ₂ incubator. At the end of the incubation, 20 μL MTT solution per well (thiazolyl blue tetrazolium bromide: 5 mg / mL in PBS, Sigma M2128) was added to incubate for another 4 hours, and then all the media was removed.

To each well, 100 μL of lysis buffer (10% SDS in 0.01 N HCl) was added again to completely lyse the formazan produced by overnight cell lysis, followed by microplates. The optical density (OD) value of each well was measured using a reader (microplate reader, 570 nm, Tecan A-5082, Salzburg, Austria). Using this value, the IC ₅₀ for cancer cells of each substance (50% inhibitory dose) was determined by Excel program. The results are as shown in Table 2.

compound
Aprotinin ^b
IC ₅₀ (μM) ^a MDA-MB-231 MCF-7 Example 1 Compound
- 0.283 5.683 + 6.632 6.216 Example 2 Compound
- 0.242 6.327 + 4.909 8.376 Control compound 0.265 1.268

^a All experiments were performed at least three times with minor modifications to the MTT assay as described above.

^b cells Incubated with or without 400 KIU of aprotinin (Sigma Aldrich) per ml.

As can be seen from Table 2, the derivative compounds of the present invention are the parent drug (IC ₅₀ ) in the absence of aprotinin in MDA-MB-231 cells expressing a lot of plasmin (IC ₅₀ = 0.265 μM). On the other hand, when aprotinin is included, the cytotoxicity values (IC ₅₀ ) are 6.632 and 4.909 μM, respectively, compared to the parent compound (IC _50). = 0.265 μΜ) activity was reduced by about 19-25 fold. This is due to the plasmin inhibitory action of aprotinin. In addition, plasmin-free MCF-7 cells exhibited cytotoxicity values (IC ₅₀ ) of 5.638 and 6.327 μM without aprotinin, respectively, reducing activity by about 5-6 fold compared to the parent drug. Including tinin, the activity was reduced by about 5-7 times compared to the control compound. That is, since MCF-7 cells are less expressed in plasmin, regardless of the presence of aprotinin, the derivative compounds of the present invention are not regenerated into the parent compound, which is considered to have low activity. Through the above experimental results, the derivative compounds of the present invention can be selectively exhibited by the plasmin of cancer cells, and in the plasmin-expressing cancer cells can be as strong cytotoxic as the parent compound.

Experimental Example  3: human plasma and human Plasmin  Within Mother drug  ( II Regeneration experiment

For example compounds of the present invention synthesized using the compound of formula II as the parent drug, conversion (regeneration) to parent drug (II) in human plasma and plasmin was confirmed by kinetic experiments.

end. Human plasma

Human plasma used in the experiment was supplied from Chungnam National University Hospital, and the regeneration test conditions in human plasma are as follows.

450 μL of cold human plasma (0 ° C., ice bath) is mixed with 50 μL of 1 mM stock solution of each compound (0.1 M Tris / HCl buffer (pH = 7.3) prepared in advance, and then the plasma suspension is incubated without delay (37 ° C.). ) And incubated at predetermined times (0 h, 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 8 h, and 24 h) and incubated and incubated to the compound of formula II in each plasma. The content of the compound was calculated by HPLC, and the content of the compound of formula II at each time period relative to the content when 1 mg of each compound was completely regenerated into the parent drug (II) was calculated as a ratio, and the results are shown in Table 3 and below. 1 and 2 are shown.

compound Time (min) Average peak area Content (μM) % ratio Example 1 Compound
(Maximum content of compound II: 11.28 μM)


15 1455 0.26 2.28 30 2062 0.36 3.23 60 4144 0.73 6.48 120 9785 1.72 15.31 240 15260 2.69 23.87 480 17664 3.12 27.64 1440 31997 5.65 50.06 Example 2 Compound (Maximum Content of Compound II: 11.22 μM)

15 2369 0.42 3.73 30 4047 0.71 6.37 60 25233 4.45 39.71 120 13777 2.43 21.68 240 16227 2.86 25.53 480 20857 3.68 32.82 1440 25188 4.45 39.64 II Compound-Standard
Average peak area content 113325 20 μM

As can be seen in Figures 1 and 2, the derivative compounds of the present invention showed a conversion rate to the II compound which is about 50.1, 39.6%, respectively, at 24 hours. This suggests that the compounds of the present invention are converted to the parent drug by cleaving peptide bonds by various enzymes in human plasma, but it can be confirmed that the conversion rate is not so high. In particular, if the conversion rate by human plasmin is higher than the conversion rate in human plasma, the derivative compounds of the present invention will be converted into the parent drug in cancer cells expressing plasmin more than in plasma, so it is an excellent prodrug that can target cancer cells. Will act as Therefore, the conversion rate of the compound of the present invention to the II compound by human plasmin was subsequently confirmed.

I. human Plasmin

Human plasmin used in the experiment was purchased from Sigma (Plasmin from human plasma, EC 3. 4. 21. 7., ≥ 2.0 units / mg protein).

The flask and 5 μL in 1 mM stock solution (0.1 M Tris / HCl buffer (pH = 7.3)) of each compound prepared in 983 μL (0 ^o C, ice bath) in cold 0.1 M Tris / HCl buffer (pH = 7.3) After mixing 12 μL in a Min solution (1 mg / L, 0.1 M Tris / HCl buffer (pH = 7.3)), the plasmin suspension is transferred to an incubator (37 ° C.) without delay and incubated for a defined time (5 minutes, 15 minutes, Sampling (50 μL) at 30 min, 45 min, 1 h, 2 h, 4 h, 8 h, 24 h), add 450 μL of ice-cold human plasma, add 250 μL of acetone, stir vigorously and store at -80 ° C It was. 50 μL of the pretreated sample (750 μL) and aqueous zinc sulfate solution were mixed and vortexed for 10 minutes, followed by centrifugation (4000 g, 15 min) to separate the supernatant. The separated supernatant was dried by evaporation with nitrogen and diluted with 125 μL of distilled water. After the concentration was made to μM the content of the compound of formula II was measured using HPLC. The results are shown in Table 4 and FIGS. 3 and 4.

compound Time (min) Average peak area Content (μM) % ratio Example 1 Compound
(Max Content of Compound II: 0.90 μM)


5 1531 0.27 29.94 15 2118 0.37 41.42 30 3904 0.69 76.34 45 3125 0.55 61.11 60 2763 0.49 54.03 120 3262 0.58 63.79 240 2836 0.50 55.46 480 3783 0.67 73.97 1440 4326 0.76 84.59 Example 2 Compound (Maximum Content of Compound II: 0.90 μM)

5 2135 0.38 41.99 15 2498 0.44 49.13 30 4290 0.76 84.38 45 3387 0.60 66.62 60 2898 0.51 57.00 120 3236 0.57 63.65 240 3243 0.57 63.79 480 4133 0.73 81.29 1440 4257 0.75 83.73 II Compound-Standard
Average peak area content 11333 2 μM

As can be seen in the table and Figures 3 and 4, the derivative compounds of the present invention after 24 hours of mixing with human plasmin, the conversion to the formula II compound, the parent drug, was found to be very high as 84.6, 83.7%, T _1/2 values were 7.3 minutes, from 3.7 minutes (WinNonlin, also calculated in version 2.1). Through this, the derivative compounds of the present invention can be confirmed that the human plasmin is very quickly converted to the formula II compound of the parent drug. When comprehensively examined with the result of regeneration of human plasma, the derivative compounds of the present invention can be found to be more selectively cleaved by human plasmin and quickly converted to the formula II compound as a parent drug. Therefore, it can be confirmed that the target for cancer cells is increased.

Experimental Example  4: In vivo  Confirmation of anticancer activity

In contrast, the derivative compound of the present invention was confirmed in vivo anticancer activity compared to the formula II compound as a parent drug.

As a tumor model, human colon adenocarcinoma (SW620) cells were used, and animals used for experimental animals were 7-week-old female Balb / c nu / nu mice supplied by Orient Bio Co., Ltd. (Seongnam, Korea). . Animals were used for testing with a 1-week acclimation period and were raised aseptically at 22 ° C.

SW620 tumor cells cultured in vitro were subcutaneously transplanted into mice at 2 × 10 ⁶ cells per horse and raised until tumors grew to a certain size. When the tumor grows to 50-100 mm ³ , the groups are separated so that the group average tumor volume is constant, and the parent formula (II) compound as a reference drug is orally administered, the derivative of Example 1 compound (16a) and Example 2 Compound (16b) was administered intravenously, respectively. Formula II compound was administered orally once a day for a total of 5 mg at a dose of 70 mg / kg, and Example compounds were dissolved in sterile saline solution once a day for 3 consecutive days at a dose of 35 mg / kg. Administered.

Tumor size and animal weight were measured twice a week and tumor size was calculated by the following formula. The test was terminated at the first (300 mm ³ ), secondary (700 mm ³ ), and tertiary (1500 mm ³ ) growth based on the start date of drug administration. After 10 days and 15 days after the first administration of the drug, the tumor growth inhibition rate was calculated and shown in Table 5 below.

Tumor size (mm ³ ) = [length × (width) ² ] ÷ 2

compound Tumor growth inhibition rate 10 days after initiation Example 1 Compound 82% Example 2 Compound 89% Control compound 48%

As can be seen from Table 5, the derivative compounds of the present invention 10 days after the start of the tumor growth inhibition rate of about 82%, 89%, compared with 48% tumor growth inhibition rate of the orally administered control compound is very good Inhibition rate was shown. Through this, the derivative compound of the present invention can be administered via intravenous injection, it can be seen that more selectively targeted to cancer cells than the control compound, and can exhibit increased anti-cancer activity.

Claims (6)

Arylsulfonylimidazolone derivatives of Formula I:
Formula I

In the formula,
R is CH ₃ or CH (CH ₃ ) ₂ ,
R 'is CH ₂ CH (CH ₃ ) ₂ or CH ₂ Ph. The arylsulfonylimidazolone derivative according to claim 1, wherein R is CH (CH ₃ ) ₂ and R 'is CH ₂ CH (CH ₃ ) ₂ . The arylsulfonylimidazolone derivative according to claim 1, wherein R is CH ₃ and R 'is CH ₂ Ph. Lung cancer, colorectal cancer, rectal cancer, colon cancer, breast cancer, cervical cancer, endometrial cancer, fallopian tube carcinoma, ovarian cancer, vaginal carcinoma, vulvar carcinoma, liver cancer, gastric cancer, esophageal cancer, small intestine, comprising the derivative of formula I according to claim 1 Cancer, pancreatic cancer, gallbladder cancer, kidney cancer, bladder cancer, urethral cancer, penile cancer, prostate cancer, testicular cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, non-small cell lung cancer, bone cancer, skin cancer, head or neck cancer, skin or eyeball A pharmaceutical composition for the treatment of a disease selected from the group consisting of melanoma, Hodgkin's disease, endocrine adenocarcinoma, chronic or acute leukemia, lymphocyte lymphoma, central nervous system tumor, spinal cord tumor, brain stem glioma and pituitary adenoma. The pharmaceutical composition of claim 4, wherein the disease is selected from the group consisting of lung cancer, colorectal cancer, rectal cancer, uterine cancer, ovarian cancer and leukemia. The pharmaceutical composition of claim 4, wherein the cancer is caused by cancer cells resistant to an existing anticancer agent.

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