KR102086565B1 - (E)-N'-ARYLIDENE-2-(3-OXO-2,3-DIHYDRO-4H-BENZO[b][1,4]OXAZIN-4-YL) ACETOHYDRAZIDES AND AN ANTICANCER COMPOSITION COMPRISING THE SAME AS AN ACTIVE INGREDIENT - Google Patents

Abstract

The present invention relates to (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide and It relates to an anticancer composition comprising this as an active ingredient. Specifically, (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) according to the present invention Acetohydrazide activates procaspase-3 and promotes its conversion to caspase-3, and thus can be used as a proliferative inhibitor against various cancer cells. The compounds according to the invention are expected to be developed as active ingredients of potent anticancer agents.

Description

(E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide and its active ingredient Anticancer composition comprising as {(E) -N'-ARYLIDENE-2- (3-OXO-2,3-DIHYDRO-4H-BENZO [b] [1,4] OXAZIN-4-YL) ACETOHYDRAZIDES AND AN ANTICANCER COMPOSITION COMPRISING THE SAME AS AN ACTIVE INGREDIENT}

The present invention relates to (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide and It relates to an anticancer composition comprising this as an active ingredient. More specifically, the present invention relates to (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [3] as an activator of procaspase-3. b] [1,4] oxazin-4-yl) acetohydrazide and an anticancer composition comprising the same as an active ingredient.

Cancer is one of two leading causes of death worldwide. The main cause of cancer development and metastasis is the dysregulation of apoptosis. Compounds such as p53 protein and XIAP or Bcl-2 inhibitors act directly on proteins during apoptosis, leading to cell death and leading to cancer cell death. Caspase is a family of cysteine proteolytic enzymes important for maintaining homeostasis through the regulation of cell death. Many studies have demonstrated that the direct activation of procaspase-3 by small molecules may have advantages over compounds that induce apoptosis, because procaspase-3 has colon cancer, lung cancer, melanoma, and liver cancer. This is due to overexpression in various tumors, including breast cancer, lymphoma and neuroblastoma.

Despite the important role of caspase in cancer pathology, caspase specific activators have been rarely developed to date. PAC-1 is the first procaspase activating compound reported to show antitumor activity in in vivo (Formula 1).

[Formula 1]

Studies on the structure and activity of PAC-1 and other related compounds have revealed that the acylhydrazone moiety plays a very important role in activity, which is a functional group with acylhydrazone and zinc. Results were obtained from complex formation between.

Prior literature

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The present inventors have made diligent efforts to develop novel compounds that enable the conversion or activation of procaspase-3 to caspase-3, resulting in the synthesis of quinazoline-based acetohydrazide compounds. The present invention was completed by experimentally confirming their caspase-3 inducing activity and proliferation inhibitory activity against cancer cells.

Accordingly, the present invention provides (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [as an activator of procaspase-3. 1,4] oxazin-4-yl) acetohydrazide or a pharmaceutically acceptable salt thereof.

The invention also relates to (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide Or it is an object to provide an anticancer composition comprising a pharmaceutically acceptable salt thereof as an active ingredient.

According to the first embodiment,

The present invention

(E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] as an activator of procaspase-3 Oxazin-4-yl) acetohydrazide or a pharmaceutically acceptable salt thereof.

As used herein, the term “pharmaceutically acceptable salts” retains the biological efficacy and properties of such compounds and is conventional acid addition salts or base addition salts formed from suitable non-toxic organic or inorganic acids, or non-toxic organic or inorganic bases. Means. Examples of acid addition salts include acid addition salts derived from inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, sulfamic acid, phosphoric acid and nitric acid, p-toluenesulfonic acid, salicylic acid, methanesulfonic acid, oxalic acid, succinic acid, citric acid, malic acid, lactic acid, Acid addition salts derived from organic acids such as fumaric acid and the like. Examples of base addition salts include base addition salts derived from ammonium, potassium, sodium, and quaternary ammonium hydroxides such as tetramethylammonium hydroxide. In order to obtain improved physical and chemical stability, hygroscopicity, flowability and solubility of the compounds, the chemical modification of the pharmaceutical compounds (ie drugs) with salts is a technique well known to pharmacy chemists, as described in H. . Ansel et. al., Pharmaceutical Dosage Forms and Drug Delivery Systems (6th Ed. 1995) at pp. 196 and 1456-1457, which is incorporated herein by reference.

As used herein, the term “pharmaceutically acceptable” means, for example, that a pharmaceutically acceptable carrier, excipient, etc. may be pharmacologically acceptable and substantially non-toxic to the patient to which the particular compound is administered. do.

In the compound according to the present invention or a pharmaceutically acceptable salt thereof, the compound or a pharmaceutically acceptable salt thereof is characterized in that the compound represented by the following formula (1) or (2) or a pharmaceutically acceptable salt thereof:

[Formula 1]

[Formula 2]

(In the formula 1 or 2,

R is hydrogen, halogen, OH, C1-6 alkyl or C1-6 alkoxy)

In the compounds according to the invention or pharmaceutically acceptable salts thereof, the halogen is characterized in that fluorine (F), chlorine (Cl), bromine (Br) or iodine (I).

In a compound according to the invention or a pharmaceutically acceptable salt thereof, the compound is characterized in that any one of the following compounds:

(E) -N '-(2-hydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydra Jide;

(E) -N '-(2,3-dihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl Acetohydrazide;

(E) -N '-(2,4-dihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl Acetohydrazide;

(E) -N '-(2,5-dihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl Acetohydrazide);

(E) -N '-(2-hydroxy-4-methoxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazine-4 -Yl) acetohydrazide;

(E) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) -N '-(3-oxoindolin-2-yl Den) acetohydrazide;

(E) -N '-(5-chloro-3-oxoindolin-2-ylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] jade Photo-4-yl) acetohydrazide.

According to the second embodiment,

The present invention is to provide an anticancer composition comprising the compound or a pharmaceutically acceptable salt thereof as an active ingredient.

As used herein, the term "cancer" refers to the aggressive nature of cells dividing and growing, ignoring normal growth limits, the invasive nature of infiltrating surrounding tissue, and spreading to other parts of the body. It is a generic term for diseases caused by cells having metastatic properties.

In the anticancer agent composition according to the present invention, the cancer is breast cancer, lung cancer, stomach cancer, liver cancer, hematologic cancer, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, skin or eye melanoma, uterine sarcoma, ovarian cancer, rectal cancer, anal cancer, colon Cancer, fallopian tube cancer, endometrial cancer, cervical cancer, small intestine cancer, endocrine cancer, thyroid cancer, parathyroid cancer, kidney cancer, soft tissue tumor, urethral cancer, prostate cancer, bronchial cancer, or bone marrow cancer.

In the anticancer agent composition according to the present invention, the compound or a pharmaceutically acceptable salt thereof is characterized by inducing conversion to caspase-3 through activation of procaspase-3.

In the anticancer agent composition according to the present invention, the anticancer agent composition comprises (i) a pharmaceutically effective amount of a compound represented by Formula 1 or 2 or a pharmaceutically acceptable salt thereof; And (ii) characterized in that it is provided in the form of a pharmaceutical composition comprising a pharmaceutically acceptable carrier.

Carriers that may be included in the anticancer composition according to the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone , Cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like.

The anticancer composition according to the present invention may further comprise a lubricant, a humectant, a sweetener, a flavoring agent, an emulsifier, a suspending agent or a preservative. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

Suitable dosages of the anticancer agent compositions according to the invention can be determined in various ways depending on factors such as the method of formulation, mode of administration, age, weight, sex, morbidity, food, time of administration, route of administration, rate of excretion and response to the patient. It may be prescribed.

The dosage of the anticancer agent composition according to the present invention may preferably be 0.001-1000 mg / kg body weight per day.

The anticancer composition according to the present invention may be administered orally or parenterally, and when administered parenterally, it may be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, or the like.

The concentration of the active ingredient included in the anticancer agent composition according to the present invention is determined in consideration of the purpose of treatment, the condition of the patient, the period of need, the severity of the disease and the like, and is not limited to a specific range of concentration.

The anticancer agent composition according to the present invention is in unit dose form by formulating with a pharmaceutically acceptable carrier and / or excipient according to a method which can be easily carried out by one of ordinary skill in the art. It can be made or prepared by incorporating into a multi-dose container. In this case, the formulation may be in the form of a solution, suspension or emulsion in an oil or aqueous medium, or may be in the form of extracts, powders, granules, tablets or capsules, and may further include a dispersant or stabilizer.

(E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide according to the present invention Activation of procaspase-3 may promote the conversion to caspase-3, and exhibits antiproliferative activity against various cancer cells. Therefore, it is expected that the compounds according to the invention can be developed as active ingredients of potent anticancer agents.

1 is (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazine- in accordance with an embodiment of the present invention; It shows the route to synthesize 4-yl) acetohydrazide.
2 is (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazine- in accordance with an embodiment of the present invention; Cytotoxic effect of 4-day) acetohydrazide on SW620, PC3 and NCI-H23.
3 is (E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazine- according to an embodiment of the present invention. 4-day) shows the caspase activity of acetohydrazide.

Hereinafter, various examples are presented to help understand the invention. The following examples are merely provided to more easily understand the invention, but the protection scope of the invention is not limited to the following examples.

Experimental Materials and Methods

chemical substance

Thin layer chromatography was performed on Whatman® 250 μm Silica Gel GF Uniplates and visualized with UV light at 254 nm to confirm reaction progress and preliminary evaluation of compound homogeneity. Melting points were measured using Gallenkamp Melting Point Apparatus (LabMerchant, London, United Kingdom) and were not calibrated. Purification using chromatography was done via open flash silica gel column chromatography using Merck silica gel 60 (240-400 mesh). Nuclear magnetic resonance spectra (1H NMR) were measured on a Bruker 500 MHz spectrometer using tetramethylsilane as internal standard and dimethylsulfoxide-d6 (DMSO-d6) as solvent unless otherwise specified. Chemical shifts are reported in parts per million (ppm) downfield from tetramethylsilane, an internal standard. Electron ionization (EI), electrospray ionization (ESI), and high resolution mass spectra were obtained from PE Biosystems API 200 (Perkin Elmer, Palo Alto, CA, USA) and Mariner® (Azco Biotech, Inc. Oceanside, USA). CA, USA) was measured using a mass spectrometer. Elemental (C, H, N) analysis was performed using a Perkin Elmer model 2400 element analyzer. Reagents and solvents are designated Aldrich or Fluka Chemical Corp. unless otherwise specified. (Milwaukee, WI, USA) or Merck was used.

<Example>

(E) -N'-arylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5a-n , 6a-d) was carried out according to the method described in FIG. The specific method is as follows.

1.Synthesis of Compounds 5 and 6

To a solution of isobutyl methyl ketone (0.6 mL) containing 2-aminophenol ( 1 ) (1.0 mmol) was added NaHCO ₃ (200 mg, 2.39 mmol) and water (0.6 mL) and the resulting mixture was cooled in a water bath. . Chloroacetyl chloride (130 mg, 1.15 mmol) was added dropwise while stirring and the cooling mixture was left to room temperature, then refluxed for 4 hours and cooled to afford 1,4-benzooxazine-3 (4H) -one ( 2 ) Was obtained in a yield of 93%.

To acetone (10 mL) solution containing 1,4-benzooxazine-3 (4H) -one ( 2 ) (0.224 g, 1.5 mmol) was added K ₂ CO ₃ (345 mg, 2.5 mmol). The resulting mixture was stirred for 30 min at 60 ° C., then KI (166 mg, 1 mmol) was added. After an additional 15 minutes, 0.50 mL of ethyl chloroacetate (4.5 mmol) was slowly added dropwise to the reaction mixture. The reaction mixture was stirred again at 60 ° C. for 3 hours. At the end of the reaction, the solvent was evaporated. The residue was taken up in 10 mL of water and extracted three times with dichloromethane. The combined organic layer was then evaporated to afford ethyl 2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) ( 3 ) as an intermediate.

Solution of ethinol (10 mL) containing ethyl 2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) ( 3 ) (1.0 mmol) To the slowly added 3.0 ml of hydrazine monohydrate (3.0 mmol). The mixture was stirred at 0 ° C. until the starting material was consumed completely. The white precipitate formed was filtered off and washed with cold-ethanol (3 times). The off-white solid product (4, 2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide) was collected and dried in vacuo. And used in the next step without further purification.

The acetohydrazide ( 4 ) (0.7 mmol) was dissolved in ethanol (20 mL) and 2 drops of concentrated acetic acid followed by benzaldehyde or isatin derivative (1.0 mmol). The mixture was refluxed until the reaction was complete. The precipitate formed was filtered off and washed three times with ethanol. The off-white solid was collected, dried in vacuo and then re-crystallized in ethanol or column chromatography (MeOH: DCM) to give the compounds according to the invention ( 5a-n, 6a-d ).

(E) -N'-benzylidene-2- (4-oxoquinazolin-3 (4H) -yl) acetohydrazide (5a)

White solid; Yield: 60% mp: 179.7-181.5 ° C. Rf = 0.53 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3506 (NH); 3236 (N = C-H aromatic); 3060, 2990 (CH, aren); 2830 (CH 2); 1719, 1680, 1531 (C = O, C = N); 1608 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.90, 11.84 (˜26%, 74%) (s, 1H, CONH); 8.39 (s, 1 H, H 2); 8.24, 8.07 (-22%, 78%) (s, 1H, N = CH); 8.17 (d, J = 7.5 Hz, 1H, H 5); 7.87 (t, J = 7.5 Hz, 1H, H 7); 7.76-7.71 (m, 3H, H2 ', H6', H8); 7.58 (t, J = 7.5 Hz, 1H, H 6); 7.48-7.45 (m, 3H, H3 ', H4', H5 '); 5.24, 4.80 (~ 23%, 77%) (s, 2H, NCH 2 CO). 13 C NMR (125 MHz, DMSO-d6, ppm): δ 168.3 (CONH), 160.3 (C = O), 148.6 (C8 = CN = C2), 148.1 (C2), 144.3 (N = CH), 134.5 (C7 ), 133.9 (C1 '), 130.1 (C4'), 128.9 (C2 ', C6'), 127.2 (C6), 127.1 (C8), 126.9 (C3 ', C5'), 126.1 (C5), 121.5 (C5 = CC = O), 47.0 (NCH2CO). MS (ESI) m / z 307.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 14 N 4 O 2 (306.1117): C, 66.66; H, 4.61; N, 18.29. Found: C, 66.63; H, 4. 64; N, 18.32.

(E) -N'-Benzylidene-2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5a)

White solid; Yield: 43%. mp: 177.3-178.5 oC. Rf = 0.68 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3442 (NH); 3203 (N = C-H aromatic); 3091, 2949 (CH, arene); 2854 (CH2); 1701, 1668, 1504 (C = O, C = N); 1618 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.75, 11.73 (˜25%, 75%) (s, 1H, CONH); 8.24, 8.06 (-25%, 75%) (s, 1H, N = CH); 7.75, 7.71 (˜75%, 25%) (dd, J = 8.0 Hz, 2.0 Hz, 2H, H 2 ′, H 6 ′); 7.47-7.43 (m, 3H, H3 ', H4', H5 '); 7.06-7.00 (m, 4H, H5, H6, H7, H8); 5.09, 4.68 (˜75%, 25%) (s, 2H, NCH 2 CO); 4.73, 4.72 (-25%, 75%) (s, 2H, OCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.4, 165.0 (CONH); 165.1, 163.8 (C3 = 0); 147.7, 144.5 (N = CH); 145.1 (C8-C-0); 134.5 (C1 ′); 130.6, 130.5 (C4 ′); 129.6, 129.5 (C5 = C-N); 129.3 (C3 ', C5'), 127.6, 127.5 (C2 ', C6'), 124.2, 124.0 (C7), 123.2 (C6), 117.0 (C5), 116.0, 115.9 (C8), 67.5 (C1), 43.4 , 42.8 (NCH2CO). MS (ESI) m / z 309.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 3 (309.1113): C, 66.01; H, 4.89; N, 13.58. Found: C, 66.05; H, 4.92; N, 13.61.

(E) -N '-(2-Chlorobenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5b)

White solid; Yield: 32%. mp: 178.7-180.5 o C. Rf = 0.69 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3564 (NH); 3269 (N = C-H aromatic); 3115, 2991 (CH, arene); 2829 (CH 2); 1687, 1504 (C = O, C = N); 1608 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.98, 11.91 (˜25%, 75%) (s, 1H, CONH); 8.63, 8.44 (-25%, 75%) (s, 1H, N = CH); 8.09, 7.95 (˜75%, 25%) (dd, J1 = 7.5 Hz, J2 = 2.0 Hz, 1H, H6 '); 7.56 (dd, J1 = 9.0 Hz, J2 = 1.0 Hz, 1H, H3 '); 7.48-7.42 (m, 2H, H4 ', H5'); 7.03-7.07 (m, 4H, H5, H6, H7, H8); 5.11, 4.69 (-75%, 25%) (s, 2H, NCH-2CO); 4.73, 4.72 (-25%, 75%) (s, 2H, OCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.5, 165.0 (CONH); 165.1, 163.9 (C3 = O), 145.0 (C8 = C-O), 140.6 (N = CH), 133.6, 133.1 (C1 '); 132.0, 131.9 (C2 ′); 130.4 (C4 ′); 129.5, 129.4 (C5 = C-N); 128.1, 128.0 (C3 '), 127.5, 127.3 (C5', C6 '); 124.1, 123.9 (C7), 123.2 (C6), 117.0 (C5), 115.9 (C8), 67.4 (C1); 43.4, 42.8 (NCH2CO). MS (ESI) m / z 343.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 14 ClN 3 O 3 (343.0724): C, 59.40; H, 4.10; N, 12.22. Found: C, 59.37; H, 4. 12; N, 12.25.

(E) -N '-(3-Chlorobenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5c)

White solid; Yield: 48%. mp: 179.7-181.5 o C. Rf = 0.69 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3456 (NH); 3184 (N = C-H aromatic); 3066, 2949 (CH, arene); 2850 (CH2); 1699, 1668, 1504 (C = O, C = N); 1595 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.89, 11.84 (˜25%, 75%) (s, 1H, CONH); 8.22, 8.04 (-25%, 75%) (s, 1H, N = CH); 7.86, 7.77 (˜75%, 25%) (s, 1 H, H 2 ′); 7.71-7.68 (m, 1H, H 6 '); 7.50-7.48 (m, 2H, H4 ', H5'); 7.03-7.02 (m, 4H, H5, H6, H7, H8); 5.11, 4.69 (˜75%, 25%) (s, 2H, OCH 2 CO); 4.73 (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.6, 165.1 (CONH); 166.5, 165.0 (C3 = 0); 145.0 (C8 = C-O); 145.9, 142.9 (N = CH); 136.8, 136.7 (C1 ′); 134.1, 134.0 (C3 ′); 131.2, 131.1 (C4 ′); 130.2, 130.1 (C5 ′); 129.5, 129, 4 (C5 = C-N); 126.9, 126.4 (C2 ′), 126.5, 126.1 (C6 ′), 124.1, 123.96 (C7); 123.1 (C6), 117.0, 116.9 (C5); 115.97, 115.90 (C8); 67.5 (OCH 2 CO); 43.3, 42.9 (NCH2CO). MS (ESI) m / z 343.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 14 ClN 3 O 3 (343.0724): C, 59.40; H, 4.10; N, 12.22. Found: C, 59.43; H, 4.08; N, 12.18.

(E) -N '-(4-Chlorobenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5d)

White solid; Yield: 61%. mp: 177.8-179.3 o C. Rf = 0.69 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3427 (NH); 3107, 2949 (CH, arene); 2846 (CH2); 1699, 1666, 1506 (C = O, C = N); 1554 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.82, 11.80 (˜25%, 75%) (s, 1H, CONH); 8.24, 8.06 (-25%, 75%) (s, 1H, N = CH); 7.80, 7.75 (˜75%, 25%) (d, J = 8.5 Hz, 2H, H 2 ′, H 6 ′); 7.61, 7.52 (-25%, 75%) (d, J = 8.5 Hz, 2H, H3 ', H5'); 7.05-7.01 (m, 4H, H5, H6, H7, H8); 5.09, 4.68 (˜75%, 25%) (s, 2H, OCH 2 CO); 4.73, 4.72 (˜75%, 25%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.4, 165.1 (CONH); 165.1, 163.9 (C3 = 0); 145.0 (C8 = C-O); 146.2, 143.2 (N = CH), 135.0, 134.9 (C4 ′); 133.5, 133.4 (C1 ′); 130.5, 129.4 (C5 = C-N); 129.6, 129.5 (C2 ′, C6 ′); 129.4, 129.3 (C3 ′); 129.2, 129.1 (C5 ′); 124.1, 123.9 (C7); 123.1 (C6); 117.0,116.9 (C5); 115.9, 115.8 (C8); 67.5 (OCH 2 CO); 43.7, 42.8 (NCH2CO). MS (ESI) m / z 343.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 14 ClN 3 O 3 (343.0724): C, 59.40; H, 4.10; N, 12.22. Found: C, 59.42; H, 4.13; N, 12.26.

(E) -N '-(4-Florobenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5e)

White solid; Yield: 45%. mp: 180.1-181.6 o C. Rf = 0.67 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3471 (NH); 3190 (N = C-H aromatic); 3080, 2949 (CH, arene); 2850 (CH2); 1699, 1668, 1506 (C = O, C = N); 1606 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.76, 11.74 (˜25%, 75%) (s, 1H, CONH); 8.25, 8.06 (-25%, 75%) (s, 1H, N = CH); 7.84, 7.79 (˜75%, 25%) (dd, J1 = 9.0 Hz, J2 = 5.0 Hz, 2H, H2 ', H6'); 7.30 (t, J = 9.0 Hz, 2H, H3 ', H5'); 7.06-7.01 (m, 4H, H5, H6, H7, H8); 5.09, 4.68 (˜75%, 25%) (s, 2H, OCH 2 CO); 4.73, 4.72 (˜25%, 75%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.3 (CONH); 165.1, 165.0 (C 3 ═O); 162.6 (C4 ′); 145.1 (C8 = C-O); 146.5, 143.3 (N = CH); 133.1, 133.0 (C5 = C-N); 129.8, 129.7 (C1 ′); 129.6, 129.5 (C2 ′, C6 ′); 124.1, 123.9 (C7); 123.1 (C6); 117.0, 116.9 (C5); 116.4, 116.2 (C3 ′, C5 ′); 115.95, 115.90 (C8); 67.4 (OCH 2 CO); 43.3, 42.8 (NCH2CO). MS (ESI) m / z 327.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 14 FN 3 O 3 (327.1019): C, 62.38; H, 4.31; N, 12.84. Found: C, 62.41; H, 4. 34; N, 12.87.

(E) -N '-(4-Bromobenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5f)

White solid; Yield: 33%. mp: 181.0-182.0 o C. Rf = 0.65 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3448 (NH); 3346 (N = C-H aromatic); 3082, 2949 (CH, arene); 2852 (CH2); 1701, 1666, 1504 (C = O, C = N); 1612 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.82, 11.80 (˜25%, 75%) (s, 1H, CONH); 8.22, 8.03 (-25%, 75%) (s, 1H, N = CH); 7.75, 7.73 (˜25%, 75%) (d, J = 8.5 Hz, 2H, H 2 ′, H 6 ′); 7.67 (d, J = 7.5 Hz, 2H, H3 ', H5'); 7.05-7.03 (m, 4H, H5, H6, H7, H8); 5.09, 4.68 (˜75%, 25%) (s, 2H, OCH 2 CO); 4.73, 4.72 (˜25%, 75%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.4 (CONH); 165.1, 165.0 (C 3 ═O); 145.1 (C8 = C-O); 146.4, 143.3 (N = CH); 133.7 (C1 ′); 132.5, 132.2 (C3 ′, C5 ′); 130.7 (C5 = C-N); 129.5, 129.4, 129.3 (C2 ', C6'); 124.1, 123.9 (C7); 123.8, 123.7 (C4 ′); 123.1 (C6); 117.0, 116.9 (C5); 116.0, 115.9 (C8); 67.4 (OCH 2 CO); 43.3, 42.8 (NCH2CO). MS (ESI) m / z 389.8 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 14 BrN 3 O 3 (for Br 81 isotope, 389.0198): C, 52.60; H, 3.63; N, 10.82. Found: C, 52.71; H, 3.72; N, 10.76.

(E) -N '-(4-Methoxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5 g)

White solid; Yield: 39%. mp: 185.1-186.3 o C. Rf = 0.68 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3427 (NH); 3186 (N = C-H aromatic); 3078, 2951 (CH, arene); 2839 (CH 2); 1701, 1666, 1504 (C = O, C = N); 1608 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.60 (s, 1H, CONH); 8.18, 8.00 (-25%, 75%) (s, 1H, N = CH); 7.70, 7.66 (˜75%, 25%) (d, J = 8.5 Hz, 2H, H 2 ′, H 6 ′); 7.05-7.01 (m, 6H, H5, H6, H7, H8, H3 ', H5'); 5.06, 4.66 (˜75%, 25%) (s, 2H, OCH 2 CO); 4.72 (s, 2H, NCH 2 CO); 3.81 (s, 3 H, 4′-OCH 3). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.0 (CONH); 165.0, 164.9 (C 3 ═O); 161.3, 161.2 (C4 ′); 147.5, 145.1 (C8 = C-O); 144.4 (N = CH); 129.6, 129.5 (C5 = C-N); 129.0, 129.0 (C2 ', C6'); 127.0 (C1 ′); 124.1, 123.9 (C7); 123.1 (C6); 117.0, 116.9 (C5); 115.9, 115.8 (C8); 114.8 (C3 ′, C5 ′); 67.4 (OCH 2 CO); 55.7 (4′-OCH 3); 43.3, 42.8 (NCH2CO). MS (ESI) m / z 339.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 18 H 17 N 3 O 4 (339.1219): C, 63.71; H, 5.05; N, 12.38. Found: C, 63.74; H, 5.02; N, 12.41.

(E) -N '-(2-Hydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5h)

White solid; Yield: 36%. mp: 175.2-177.0 o C. Rf = 0.56 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3487 (NH); 3431 (OH); 3078, 2949 (CH, arene); 2854 (CH2); 1669, 1666, 1506 (C = O, C = N); 1608 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.95, 11.66 (˜43%, 57%) (s, 1H, CONH); 10.95, 10.06 (˜43%, 57%) (s, 1H, 2′-OH); 8.46, 8.36 (-43%, 57%) (s, 1H, N = CH); 7.78, 7.57 (~ 57%, 43%) (dd, J1 = 8.0 Hz, J2 = 1.5 Hz, 1H, H6 '); 7.31-7.24 (m, 1 H, H 4 '); 7.08-7.03 (m, 4H, H5, H6, H7, H8); 6.93-6.86 (m, 2H, H3 ', H5'); 5.06, 4.69 (˜57%, 43%) (s, 2H, OCH 2 CO); 4.73, 4.72 (˜43%, 57%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 168.0 (CONH), 165.0 (C 3 ═O); 157.7, 156.8 (C2 ′); 147.7 (N = CH); 145.0 (C8 = C-O); 131.9, 131.7 (C4 ′); 129.5, 129.4 (C5 = C-N); 126.8 (C6 ′); 124.2, 123.9 (C7); 123.2 (C6); 120.6 (C5 ′); 119.9, 119.8 (C1 ′); 117.0, 116.8 (C3 ′); 116.9, 116.6 (C5); 116.0, 115.9 (C8); 67.4 (OCH 2 CO); 43.2, 42.7 (NCH2CO). MS (ESI) m / z 325.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 4 (325.1063): C, 62.76; H, 4.65; N, 12.92. Found: C, 62.79; H, 4.61; N, 12.95.

(E) -N '-(4-Hydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5i)

White solid; Yield: 65%. mp: 176.7-178.5 oC. Rf = 0.58 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3454 (NH); 3414 (OH); 2987 (CH, arene); 2831 (CH2); 1685, 1658, 1504 (C = O, C = N); 1606 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.53 (s, 1H, CONH); 9.93, 9.92 (-25%, 75%) (s, 1H, 4'-OH); 8.13, 7.95 (-25%, 75%) (s, 1H, N = CH); 7.57, 7.53 (˜75%, 25%) (d, J = 8.5 Hz, 2H, H 2 ′, H 6 ′); 7.05-6.99 (m, 4H, H5, H6, H7, H8); 6.84 (d, J = 8.5 Hz, 2H, H3 ', H5'); 5.04, 4.65 (˜75%, 25%) (s, 2H, OCH 2 CO); 4.72, 4.71 (-25%, 75%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 167.9, 164.9 (CONH); 165.0, 163.3 (C3 = 0); 159.9, 159.8 (C4 ′); 145.0 (C8 = C-O); 144.8 (N = CH); 129.6, 129.5 (C5 = C-N); 129.3, 129.2 (C2 ', C6'); 125.5 (C1 ′); 123.9 (C7); 123.1 (C6); 117.0, 116.9 (C5); 116.1 (C3 ', C5'); 115.9, 115.8 (C8); 67.4 (OCH2CO); 43.3, 42.7 (NCH2CO). MS (ESI) m / z 325.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 4 (325.1063): C, 62.76; H, 4.65; N, 12.92. Found: C, 62.77; H, 4.63; N, 12.90.

(E) -N '-(2,3-Dihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5j)

White solid; Yield: 42%. mp: 172.0-173.4 o C. Rf = 0.51 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3496 (NH); 3182 (N = C-H aromatic); 3076, 2941 (CH, arene); 2850 (CH2); 1695, 1660, 1502 (C = O, C = N); 1610 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.96, 11.66 (˜44%, 56%) (s, 1H, CONH); 10.71, 9.57 (˜56%, 44%) (s, 1H, 2′-OH); 9.25, 9.17 (˜56%, 44%) (s, 1H, 3′-OH); 8.42, 8.36 (-44%, 56%) (s, 1H, N = CH); 7.22, 6.99 (d, J = 7.0 Hz, 1H, H6 '); 7.22-6.98 (m, 4H, H5, H6, H7, H8); 6.87-6.84 (m, 1 H, H 4 '); 6.84-6.68 (m, 1 H, H5 '); 5.05, 4.70 (˜56%, 44%) (s, 2H, OCH 2 CO); 4.74, 4.72 (~ 44%, 56%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 167.9 (CONH); 165.0 (C3 = 0), 163.6 (C2 '), 148.6 (C3'); 146.1, 146.0 (C8 = C-O); 142.6 (N = CH); 129.6, 129.5 (C5 = C-N); 123.9, 123.1 (C7); 124.2 (C6 '), 123.1 (C6); 121.1 (C5 ′); 120.2, 119.6 (C1 '); 119.7, 119.2 (C4 '); 117.9, 117.2 (C5); 116.0; 115.9 (C8); 67.4 (OCH 2 CO); 43.3, 42.7 (NCH2CO). MS (ESI) m / z 341.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 5 (341.1012): C, 59.82; H, 4. 43; N, 12.31. Found: C, 59.79; H, 4. 45; N, 12.34.

(E) -N '-(2,4-Dihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5k)

White solid; Yield: 47%. mp: 171.2-173.6 o C. Rf = 0.52 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3425 (NH); 3184 (OH); 3115 (N = C-H aromatic); 2953 (CH, arene); 2845 (CH2); 1687, 1660, 1502 (C = O, C = N); 1631 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.76, 11.47 (˜49%, 51%) (s, 1H, CONH); 11.12, 9.99 (˜49%, 51%) (s, 1H, 2′-OH); 9.96, 9.82 (˜49%, 51%) (s, 1H, 4′-OH); 8.32, 8.23 (˜49%, 51%) (s, 1H, N═CH); 7.56, 7.34 (˜49%, 51%) (d, J = 8.5 Hz, 1H, H 6 ′); 7.06-7.00 (m, 4H, H5, H6, H7, H8); 6.36-6.33 (m, 2H, H3 ', H5'); 5.01, 4.67 (˜49%, 51%) (s, 2H, OCH 2 CO); 4.73, 4.71 (˜51%, 49%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 167.5 (CONH); 165.01 (C 3 ═O); 163.2 (C2 '); 161.2, 161.0 (C4 ′); 145.1, 145.0 (C8 = C-O); 143.0 (N = CH); 131.5 (C6 '); 129.5, 129.4 (C5 = C-N); 124.1, 123.9 (C7); 123.2 (C6); 117.0, 116.9 (C5); 115.97, 115.91 (C8); 110.8 (C1 '); 108.3, 108.2 (C5 '); 103.1, 102.8 (C3 '); 67.4 (OCH 2 CO); 43.2, 42.7 (NCH2CO). MS (ESI) m / z 341.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 5 (341.1012): C, 59.82; H, 4. 43; N, 12.31. Found: C, 59.85; H, 4. 46; N, 12.29.

(E) -N '-(2,5-Dihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5l)

White solid; Yield: 63%. mp: 172.4-173.8 oC. Rf = 0.45 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3606 (OH); 3415 (NH); 3194 (N = C-H aromatic); 3011, 2949 (CH, arene); 2856 (CH2); 1687, 1653, 1502 (C = O, C = N); 1585 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.83, 11.62 (˜43%, 57%) (s, 1H, CONH); 10.08, 9.37 (˜43%, 57%) (s, 1H, 2′-OH); 8.97, 8.90 (˜43%, 57%) (s, 1H, 5′-OH); 8.39, 8.30 (-43%, 57%) (s, 1H, N = CH); 7.17-6.99 (m, 5H, H6 ', H5, H6, H7, H8); 6.76-6.70 (m, 2H, H3 ', H4'); 5.06, 4.68 (~ 57%, 43%) (s, 2H, OCH-2CO); 4.73, 4.72 (˜43%, 57%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 167.9 (CONH); 165.1, 165.0 (C 3 ═O); 150.5, 150.4 (C2 '); 150.0, 149.9 (C5 ′); 145.1, 145.0 (C8 = C-O); 142.0 (N = CH); 129.5, 129.4 (C5 = C-N); 124.1, 123.9 (C7); 123.2 (C6); 120.8 (C3 '); 119.5, 119.3 (C4 ′); 117.5 (C6 ′); 117.0, 116.9 (C5); 116.0, 115.9 (C8); 113.7, 111.6 (C1 '); 67.4 (OCH 2 CO); 43.3, 42.6 (NCH2CO). MS (ESI) m / z 341.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 5 (341.1012): C, 59.82; H, 4. 43; N, 12.31. Found: C, 59.80; H, 4. 44; N, 12.30.

(E) -N '-(2,3,4-Trihydroxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5m )

White solid; Yield: 74%. mp: 171.8-172.6 o C. Rf = 0.45 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3417 (NH); 3101, 2981 (CH, arene); 2845 (CH2); 1670, 1626, 1504 (C = O, C = N); 1607 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.80, 11.51 (˜58%, 42%) (s, 1H, CONH); 11.10, 9.57 (˜58%, 42%) (s, 1H, 2′-OH); 9.48, 9.38 (˜58%, 42%) (s, 1H, 4′-OH); 8.56, 8.49 (˜42%, 58%) (s, 1H, 3′-OH); 8.27, 8.21 (˜58%, 42%) (s, 1H, N═CH); 7.09-7.00 (m, 4H, H5, H6, H7, H8); 6.81, 6.40 (-42%, 58%) (d, J = 8.5 Hz, 1H, H6 '); 6.39 (dd, J1 = 8.5 Hz, J2 = 2.0 Hz, 1H, H5 '); 5.02, 4.68 (˜42%, 58%) (s, 2H, OCH 2 CO); 4.73, 4.72 (˜58%, 42%) (s, 2H, NCH 2 CO). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 167.4 (CONH); 165.0 (C 3 ═O); 149.9, 149.2 (C3 '); 148.8, 147.8 (C2 '); 145.0 (C8 = C-O); 144.3 (N = CH); 133.2, 133.1 (C4 '); 129.5, 129.4 (C5 = C-N); 124.1, 123.9 (C7); 123.1, 121.5 (C6); 118.5 (C6 '); 117.0, 116.9 (C5); 115.9, 115.3 (C8); 112.7, 111.1 (C1 '); 108.2, 108.1 (C5 '); 67.4 (OCH 2 CO); 43.1, 42.6 (NCH2CO). MS (ESI) m / z 357.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 17 H 15 N 3 O 6 (357.0961): C, 57.14; H, 4. 23; N, 11.76. Found: C, 57.11; H, 4. 25; N, 11.79.

(E) -N '-(2-Hydroxy-4-methoxybenzylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) acetohydrazide (5n )

White solid; Yield: 83%. mp: 176.3-177.9 oC. Rf = 0.59 (DCM: MeOH = 14: 1). IR (KBr, cm-1): 3458 (NH); 3186 (N = C-H aromatic); 3080, 2945 (CH, arene); 2843 (CH2); 1699, 1664, 1504 (C = O, C = N); 1570 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.84, 11.54 (˜46%, 54%) (s, 1H, CONH); 11.28, 10.17 (˜46%, 54%) (s, 1H, 2′-OH); 8.36, 8.26 (˜46%, 54%) (s, 1H, N═CH); 7.66, 7.46 (˜46%, 54%) (d, J = 8.5 Hz, 1H, H 6 ′); 7.08-7.01 (m, 4H, H5, H6, H7, H8); 6.54-6.51 (m, 2H, H3 ', H5'); 5.03, 4.68 (˜46%, 54%) (s, 2H, OCH 2 CO); 4.73, 4.72 (-54%, 46%) (s, 2H, NCH 2 CO); 3.77, 3.76 (~ 54%, 46%) (s, 4'-OCH3). 13 C-NMR (125 MHz, DMSO-d 6, ppm): δ 167.6 (CONH), 165.0 (C 3 ═O); 163.4, 159.7 (C4 '); 162.6, 158.4 (C2 ′); 145.1 (C8 = C-O); 142.5 (N = CH); 131.3 (C6 '); 129.5, 129.4 (C5 = C-N); 124.1, 123.9 (C7); 123.2, 123.1 (C6); 117.0, 116.9 (C5); 116.0, 115.9 (C8), 112.1 (C1 '); 106.9, 106.8 (C5 '); 101.6, 101.3 (C3 '); 67.4 (OCH 2 CO); 55.7, 55.6 (OCH 3); 43.2, 42.7 (NCH2CO). MS (ESI) m / z 355.9 [M + H] &lt; + &gt;. Anal. Calcd. For C 18 H 17 N 3 O 5 (355.1168): C, 60.84; H, 4. 82; N, 11.83. Found: C, 60.80; H, 4.79; N, 11.86.

(E) -2- (3-Oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) -N '-(3-oxoindolin-2-ylidene) acetohydrazide (6a )

Light yellow solid; Yield: 57%. mp: 179.7-181.5 o C. Rf = 0.67 (DCM: MeOH = 9: 1). IR (KBr, cm-1): 3460 (NH); 3234 (N = C-H aromatic); 2985 (CH, arene); 2951 (CH2); 1703, 1668, 1504 (C = O, C = N); 1622 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 13.22, 12.69 (~ 25%, 75%) ((s, 1H, CONH); 11.30 (s, 1H, NH-isatine); 7.60 (s, 1H, H4 '); 7.40 (t, J = 7.5 Hz, 1H, H6'); 7.14- 7.02 (m, 5H, H5 ', H5, H6, H7, H8); 6.97 (d, J = 7.5 Hz, 1H, H3 '); 5.23, 4.92 (~ 75%, 25%) (s, 2H, OCH2CO); 4.89, 4.75 (~ 25%, 75%) (s, 2H, NCH2CO) .13C-NMR (125 MHz , DMSO-d6, ppm): δ 165.2 (CONH); 162.9 (C3 = O); 145.1 (C8 = CO); 143.0 (C7'-C-NH); 132.3 (NC-C5, C6 ', C2') ; 124.1 (C7), 123.2 (C5 '); 123.1 (C6); 121.3 (C4'); 120.1 (C5); 117.0 (O = C3'-C-C4 '); 116.0 (C8); 111.6 (C7' 67.4 (OCH 2 CO); 42.5 (NCH 2 CO) .MS (ESI) m / z 350.9 [M + H] + .Anal.Calcd.For C 18 H 14 N 4 O 4 (350.3282): C, 61.71; H, 4.03; N, 15.99. : C, 61.74; H, 4.06; N, 15.96.

(E) -N '-(5-Chloro-3-oxoindolin-2-ylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl acetohydrazide (6b)

Light yellow solid; Yield: 39%. mp: 180.2-181.9 oC. Rf = 0.70 (DCM: MeOH = 9: 1). IR (KBr, cm-1): 3458 (NH); 3280 (N = C-H aromatic); 3051 (CH, arene); 2854 (CH2); 1747, 1726, 1504 (C = O, C = N); 1602 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.80 (s, 1H, CONH); 10.97 (s, 1H, NH-isatine); 8.38 (s, 1 H, H 6 '); 7.46 (d, J = 8.5 Hz, 1H, H 3 ′); 7.10-7.02 (m, 4H, H5, H6, H7, H8); 6.93 (d, J = 8.5 Hz, 1H, H 4 ′); 5.13 (s, 2H, OCH 2 CO); 4.75 (s, 2 H, NCH 2 CO). 13 C NMR (125 MHz, DMSO-d 6, ppm): δ 165.2 (CONH); 164.7 (C 3 ═O); 145.0 (C8 = C-O); 143.0 (C7'-C-NH); 132.5 (C2 ', N-C-C5, C6'); 129.3 (C5 ′); 126.3 (C4 '); 124.1 (C7); 123.1 (C5, C6); 117.0 116.7 (C7 ′); 115.9 (C8); 112.4 (C3'-C-C4 '); 67.4 (O-CH 2 -CO); 43.3 (NCH2CO). MS (ESI) m / z 384.8 [M + H] &lt; + &gt;. Anal. Calcd. For C 18 H 13 ClN 4 O 4 (384.7732): C, 56.19; H, 3.41; N, 14.56. Found: C, 56.15; H, 3. 44; N, 14.58.

(E) -N '-(5-Methyl-3-oxoindolin-2-ylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl acetohydrazide (6c)

Light yellow solid; Yield: 44%. mp: 178.6-179.3 o C. Rf = 0.68 (DCM: MeOH = 9: 1). IR (KBr, cm-1): 3244 (N = C-H aromatic); 3149, 2983 (CH, arene); 2949 (CH2); 1705, 1668, 1504 (C = O, C = N); 1622 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 11.51 (s, 1H, CONH), 10.73 (s, 1H, NH-isatine); 8.06 (s, 1 H, H 4 '); 7.22 (d, J = 8.0 Hz, 1H, H 6 ′); 7.09-7.04 (m, 4H, H5, H6, H7, H8); 6.82 (d, J = 8.0 Hz, 1H, H 3 ′); 5.13 (s, 2H, OCH 2 CO); 4.76 (s, 2H, NCH 2 CO); 2.31 (s, 3 H, 5′-CH 3). 13 C NMR (125 MHz, DMSO-d 6, ppm): δ 165.2 (CONH); 164.9 (C 3 ═O); 145.0 (C8 = C-O); 142.0 (C7'-C-NH); 133.5 (C2 ', N-C-C5); 131.2 (C6 '); 129.4 (C5 ′); 127.2 (C4 '); 124.1 (C7); 123.1 (C5); 117.0 (C6); 116.7 (C7 '); 115.9, 115.7 (C8); 110.8 (C3'-C-C4 '); 67.4 (O-CH 2 -CO); 43.3 (NCH 2 CO); 20.9 (5′-CH 3). MS (ESI) m / z 365.0 [M + H] &lt; + &gt;. Anal. Calcd. For C 19 H 16 N 4 O 4 (364.3547): C, 62.63; H, 4. 43; N, 15.38. Found: C, 62.60; H, 4. 45; N, 15.41.

(E) -N '-(5-Methoxy-3-oxoindolin-2-ylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl acetohydrazide (6d)

Light yellow solid; Yield: 53%. mp: 180.1-181.7 o C. Rf = 0.72 (DCM: MeOH = 9: 1). IR (KBr, cm-1): 3277 (N = C-H aromatic); 3136, 3051 (CH, arene); 2858 (CH2); 1660, 1533 (C = O, C = N); 1606 (C = C). 1 H-NMR (500 MHz, DMSO-d 6, ppm): δ 12.71 (s, 1H, CONH), 11.12 (s, 1H, NH-isatine); 6.85-7.21 (m, 8H, H5, H6, H7, H8, H3 ', H4', H5 '); 5.25 (s, 2H, OCH 2 CO); 4.75 (s, 2H, NCH 2 CO); 3.77 (s, 3 H, 5′-OCH 3). 13 C NMR (125 MHz, DMSO-d6, ppm): δ 165.2 (CONH), 163.1 (C3 = O), 155.9 (C6 '), 145.1 (C8 = CO), 136.7 (C1', C2 ', NC-C5 ), 123,2 (C5, C4 '); 120.8 (C7), 118.5 (C7'-C-NH), 117.0 (C6), 116.0 (C8), 112.5 (C5 '), 106.4 (C7'); 67.4 (OCH 2 CO); 56.1 (5′-OCH 3). MS (ESI) m / z 381.0 [M + H] &lt; + &gt;. Anal. Calcd. For C 18 H 13 N 4 O 5 (380.3541): C, 60.00; H, 4. 24; N, 14.73. Found: C, 60.12; H, 4.13; N, 14.62.

Experimental Example

1. Cytotoxicity Assay

Toxicity of the synthesized compounds was evaluated using SW620 (colon cancer), PC3 (prostate cancer) and NCI-H23 (lung cancer). The cell lines were purchased from a cancer cell bank at the Korea Research Institute of Bioscience and Biotechnology (KRIBB). Media, serum and other reagents used in cell culture were GIBCO Co. Ltd. (Grand Island, New York, USA). The cells were incubated in DMEM. The cells were then suspended at a concentration of 3 × 10 ⁴ cells / mL after trypsinization. On day 0, 180 μL of cell suspension was seeded into each well of a 96-well plate. The plates were then incubated at 37 ° C. for 24 hours in an incubator containing 5% CO 2. The compound was first dissolved and diluted in dimethyl sulfoxide (DMSO) and used. 20 μL of each compound sample prepared in the above example was added to each well of a 96-well plate seeded with cell suspension and incubated for 24 hours at different concentrations. The plates were further incubated for 48 hours. Cytotoxicity of the compound was determined by a method [18-21] with slight modifications to the contents described in known literature [17]. The value measured for each compound is shown as the average value of 3 independent measurement results (SD≤10%). Cell growth percentages (CGP) of SW620 cancer cells in the presence of 30 μg / mL of each compound were measured, and the results are shown in Table 1 and FIG. 2 using 5-fluorouracil as a positive control.

Cpd code
R Cell growth percentage (CGP,%) ¹ / Cell lines ² SW620 PC3 NCI-H23 5a H 75.48 ± 0.13 68.43 ± 6.06 70.70 ± 3.91 5b 2-Cl 60.85 ± 4.93 71.46 ± 5.87 68.77 ± 4.64 5c 3-Cl 70.88 ± 1.81 77.74 ± 7.06 63.49 ± 4.12 5d 4-Cl 75.98 ± 3.42 77.65 ± 5.54 79.99 ± 1.42 5e 4-F 61.92 ± 3.89 64.31 ± 5.69 75.62 ± 4.89 5f 4-Br 78.96 ± 7.29 78.21 ± 4.27 76.61 ± 2.87 5 g 4-OCH ₃ 63.63 ± 3.61 70.09 ± 5.10 66.19 ± 3.23 5h 2-OH -15.32 ± 6.77 -12.65 ± 5.47 -16.00 ± 1.75 5i 4-OH 66.23 ± 4.61 77.74 ± 4.24 75.75 ± 3.19 5j 2,3- (OH) ₂ -8.64 ± 0.79 -17.20 ± 7.28 -10.69 ± 5.87 5k 2,4- (OH) ₂ 13.51 ± 6.91 5.50 ± 2.90 13.30 ± 2.83 5l 2,5- (OH) ₂ -13.66 ± 5.43 -20.73 ± 4.45 -17.11 ± 1.13 5 m 2,3,4- (OH) ₃ 70.59 ± 2.38 77.29 ± 4.04 75.67 ± 3.73 5n 2-OH-4-OCH ₃ 11.35 ± 6.07 9.64 ± 6.78 13.93 ± 3.28 6a H 25.29 ± 2.92 16.52 ± 2.86 21.78 ± 4.93 6b 5-Cl 23.79 ± 6.51 19.25 ± 1.11 14.88 ± 2.71 6c 5-CH ₃ 73.48 ± 2.59 77.85 ± 3.93 75.23 ± 4.63 6d 7-Cl 76.36 ± 3.93 75.94 ± 3.50 73.94 ± 4.88 5-FU ³ 17.35 ± 9.01 25.45 ± 1.21 23.22 ± 2.35

As can be seen from Table 1, 5h, 5j, 5k, 5l, 5n, 6a and 6b of the compounds according to the present invention synthesized in the above examples are cancer cell lines SW620 (colon cancer), PC3 (prostate cancer) and NCI -H23 (lung cancer) was found to show good cytotoxicity (Figure 2.)

2. Caspase-3 activation assay

U937 cells (5 × 10 ⁵ cells / well) were plated in 6-well plates and allowed to stabilize overnight. Cells were treated with PAC-1 or compounds 5k and 5j (50 μM in 0.1% dimethyl sulfoxide) according to the invention. After 24 hours in incubation, cells were harvested and washed twice with PBS. The cells were lysed with 50 μL of quench cell lysis buffer at 4 ° C. for 10 minutes. Cell lysates (200 μg / 100 μL / well) were mixed using Ac-DEVD-pNA (200 μM). OD was measured every 30 minutes at 405 nm for 12 hours.

As a result, it was confirmed that the compounds (5k and 5j) according to the present invention showed significantly superior caspase activity compared to PAC-1 (FIG. 3).

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that the specific technology is merely a preferred embodiment, which is not intended to limit the scope of the present invention. Therefore, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

Claims (7)

A compound represented by the following 2 or a pharmaceutically acceptable salt thereof:
[Formula 2]

(In the above formula 2,
R is hydrogen, halogen, OH, C1-6 alkyl or C1-6 alkoxy)
The method of claim 1,
The halogen is a compound or a pharmaceutically acceptable salt thereof, characterized in that fluorine (F), chlorine (Cl), bromine (Br), iodine (I).
The method of claim 1,
The compound or a pharmaceutically acceptable salt thereof, characterized in that the compound is any one of the following compounds:
(E) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] oxazin-4-yl) -N '-(3-oxoindolin-2-yl Den) acetohydrazide; And
(E) -N '-(5-chloro-3-oxoindolin-2-ylidene) -2- (3-oxo-2,3-dihydro-4H-benzo [b] [1,4] jade Photo-4-yl) acetohydrazide.
An anticancer agent composition comprising the compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof as an active ingredient.
The method of claim 4, wherein
The cancer may be breast cancer, lung cancer, stomach cancer, liver cancer, blood cancer, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, skin or eye melanoma, uterine sarcoma, ovarian cancer, rectal cancer, anal cancer, colon cancer, fallopian tube cancer, endometrial cancer, Cervical cancer, small intestine cancer, endocrine cancer, thyroid cancer, parathyroid cancer, kidney cancer, soft tissue tumor, urethral cancer, prostate cancer, bronchial cancer, or bone marrow cancer.
The method of claim 4, wherein
The compound or a pharmaceutically acceptable salt thereof is characterized in that the induction of the conversion to caspase-3 through the activation of procaspase-3, anticancer agent composition.
The method of claim 4, wherein
The anticancer agent composition may comprise (i) a pharmaceutically effective amount of a compound represented by Formula 2 or a pharmaceutically acceptable salt thereof; And (ii) in the form of a pharmaceutical composition comprising a pharmaceutically acceptable carrier.

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