CN109400632B

CN109400632B - A kind of bis-fluoroquinolone-based oxadiazole urea derivatives containing N-methyl enoxacin and its preparation method and application

Info

Publication number: CN109400632B
Application number: CN201811425995.0A
Authority: CN
Inventors: 周敬; 臧锋磊; 李阳杰; 胡国强
Original assignee: Zhengzhou University of Industrial Technology
Current assignee: Zhengzhou University of Industrial Technology
Priority date: 2018-11-27
Filing date: 2018-11-27
Publication date: 2021-07-06
Anticipated expiration: 2038-11-27
Also published as: CN109400632A

Abstract

本发明公开了一种含N‑甲基依诺沙星的双‑氟喹诺酮基噁二唑脲类衍生物及其制备方法和应用，其化学结构通式如下式I所示：

通式I中，R为‑CH₂‑CH₃、环丙基或‑CH₂‑CH₂F；L为‑Cl、‑F、1‑哌嗪基、取代的哌嗪‑1‑基或含氮稠杂环；X为‑CH、‑N、‑CF或‑COCH₃；或R与X共同构成噁嗪环或噻嗪环。本发明的双‑氟喹诺酮噁二唑脲类衍生物，实现了双‑氟喹诺酮骨架及噁二唑杂环和功能基脲类的有机拼合，进而实现不同药效团的迁越与叠加，增加了氟喹诺酮的抗肿瘤活性和选择性，降低对正常细胞的毒副作用，可以作为抗肿瘤活性物质开发全新结构的抗肿瘤药物。The invention discloses a bis-fluoroquinolone-based oxadiazole urea derivative containing N-methyl enoxacin and a preparation method and application thereof. The general chemical structure of the derivative is shown in the following formula I:

In general formula I, R is -CH ₂ -CH ₃ , cyclopropyl or -CH ₂ -CH ₂ F; L is -Cl, -F, 1-piperazinyl, substituted piperazine-1-yl or containing nitrogen-fused heterocycle; X is -CH, -N, -CF or -COCH ₃ ; or R and X together form an oxazine ring or a thiazine ring. The bis-fluoroquinolone oxadiazole urea derivatives of the present invention realize the organic combination of the bis-fluoroquinolone skeleton, the oxadiazole heterocycle and the functional group urea, thereby realizing the transition and superposition of different pharmacophores, increasing the The anti-tumor activity and selectivity of fluoroquinolones can reduce the toxic and side effects on normal cells, and can be used as anti-tumor active substances to develop anti-tumor drugs with new structures.

Description

Bis-fluoroquinolone oxadiazole urea derivative containing N-methylenoxacin and preparation method and application thereof

Technical Field

The invention belongs to the technical field of new drug discovery and innovative drug synthesis, and particularly relates to a bis-fluoroquinolone oxadiazole urea derivative containing N-methylenoxacin, a preparation method of the derivative, and application of the derivative in antitumor drugs.

Background

The research and development of new drugs originate from the discovery of leads, and the structural optimization of the leads is a key link for promoting the development of the leads to patent drugs. Based on a rational drug design strategy of a structure or a mechanism, a novel small molecule primer which has the functions of treating and regulating serious diseases such as malignant tumor is created by utilizing the dominant skeleton or pharmacophore fragment of the existing drug, and the strategy is the most economical and effective strategy for developing new drugs. Based on the above, on one hand, the Fluoroquinolone (FQs) is considered as a class of antibacterial drugs widely used in clinic, the antibacterial dominant pharmacophore skeleton of the Fluoroquinolone (FQs) is quinoline (naphthyridine) -4-ketone-3-carboxylic acid, and as the action target of the Fluoroquinolone (FQs), Topoisomerase (TOPO) is also an important target enzyme of the antitumor drugs, the antibacterial activity of the fluoroquinolone can be converted into the antitumor activity through a structural modification strategy, and then the antitumor fluoroquinolone guide with a new structure is discovered. Meanwhile, the structure-activity relationship research finds that fluoroquinolone C-3 carboxyl is not a pharmacophore necessary for anti-tumor activity although necessary for anti-bacterial activity, and the heterocyclic ring or the fused heterocyclic ring is used as an isostere of the C-3 carboxyl to obviously improve the anti-tumor activity, so that a new thought is provided for converting an anti-bacterial fluoroquinolone medicament into an anti-tumor FQ molecule. However, the problem is that selecting what structure type of carboxyl isostere and how to link with the fluoroquinolone skeleton is beneficial to the discovery of targeting small molecule leads, and the discovery of further innovating and driving targeting anti-tumor fluoroquinolone drugs is still a problem to be solved at present. On the other hand, targeted antitumor drug molecules constructed based on Protein Tyrosine Kinase (PTK) targets have made great progress, and numerous small molecule Protein Tyrosine Kinase Inhibitors (PTKIs) enter clinical targeted therapy of tumors, thereby stimulating discovery and research and development of targeted therapeutic drugs. Meanwhile, the structures of the marketed targeting PTKIs molecules are analyzed, and the structures can be divided into four structural types (shown as follows) of arylaminopyrimidines such as imatinib (a), arylaminoquinazolines such as gefitinib (B), diarylureas such as regorafenib (C), and α, β -unsaturated ketones such as sunitinib (D):

if the fluoroquinolone can be combined with the targeting PTKIs molecules, the structural characteristics of the antitumor fluoroquinolone are reserved, and the urea structural characteristics of the targeting PTKIs molecules are reflected, so that a new thought is provided for the development of the targeting tumor treatment medicine.

Disclosure of Invention

The invention provides a bis-fluoroquinolone oxadiazole urea derivative containing N-methylenoxacin, aiming at the technical problem of combining fluoroquinolone with targeted PTKIs molecules, and the prepared derivative has the effects of synergism and toxicity reduction and can be developed as an anti-tumor medicament with a brand new structure.

In order to achieve the above object, the technical scheme adopted by the invention is as follows: the bis-fluoroquinolone oxadiazole urea derivative containing N-methyl enoxacin has the following structural general formula I:

in the general formula I, R is-CH₂-CH₃Cyclopropyl or-CH₂-CH₂F；

L is-Cl, -F, 1-piperazinyl, substituted piperazin-1-yl, or a nitrogen-containing fused heterocycle;

x is-CH, -N, -CF or-COCH₃；

Or R and X together form an oxazine ring or a thiazine ring.

More preferably, the N-methylenoxacin-containing bis-fluoroquinolone oxadiazole urea derivative is a compound having a structure shown in the following formulas I-1 to I-18:

the preparation method of the bis-fluoroquinolone oxadiazole urea derivative containing N-methyl enoxacin comprises the following specific preparation processes:

1) n-methyl enoxacin (12) is used as a raw material, condensed with semicarbazide hydrochloride in polyphosphoric acid (PPA), and post-treated to prepare an N-methyl enoxacin C-3 oxadiazolamide intermediate II, wherein the synthetic preparation route is as follows:

2) carrying out condensation reaction on fluoroquinolone carboxylic acid (marked as FQ-COOH) shown in formulas 1-18 and N ', N-carbonyl diimidazole (marked as CDI) in N, N-Dimethylformamide (DMF) respectively to prepare corresponding fluoroquinolone carboxylic acid imidazole amide (shown in formulas 1 ' -18 '); the obtained fluoroquinolone carboxylic acid imidazole amide (1 'to 18') is respectively reacted with hydroxylamine hydrochloride in pyridine (By) to conveniently prepare corresponding fluoroquinolone hydroximic acid (1 'to 18');

wherein the molar ratio of the fluoroquinolone carboxylic acid shown in the formulas 1-18 to the N, N' -carbonyldiimidazole is 1: 1.0-2.0;

the molar ratio of the fluoroquinolone carboxylic acid imidazole amide shown in the formula 1 'to 18' to hydroxylamine hydrochloride is 1: 1.0-5.0.

3) Fluoroquinolone hydroxamic acid (1 ' -18 ') is rearranged into fluoroquinolone isocyanate under the auxiliary catalysis of N ', N-carbonyl diimidazole through Lossen, directly undergoes condensation reaction with an N-methyl enoxacin C-3 oxadiazolamide intermediate II without separation, and is subjected to post-treatment to prepare the bis-fluoroquinolone-based oxadiazomide derivatives containing N-methyl enoxacin shown in formulas I-1 to I-18, wherein the synthetic route is as follows:

wherein the molar ratio of the fluoroquinolone hydroximic acid shown in the formula 1 ' to 18 ' to the N, N ' -carbonyldiimidazole is 1: 1.0-2.0, and the molar ratio of the fluoroquinolone hydroximic acid shown in the formula 1 ' to 18 ' to the N-methylenoxacin C-3 oxadiazolidine intermediate II is 1:1.

Wherein the fluoroquinolone carboxylic acid (FQ-COOH) comprises: the ofloxacin injection comprises ofloxacin (1), levofloxacin (2), ofloxacin (3), levofloxacin (4), norfloxacin (5), pefloxacin (6), N-acetylnorfloxacin (7), ciprofloxacin (8), N-methylcyclociprofloxacin (9), N-acetylciprofloxacin (10), enoxacin (11), N-methylenoxacin (12), N-acetylenoxacin (13), fleroxacin (14), N-methyllomefloxacin (15), N-methylgatifloxacin (16), N-methylmoxifloxacin (17) and rufloxacin (18), and the structures are shown as follows.

The corresponding fluoroquinolone carboxylic acid imidazolamides (1 'to 18') are: ofloxacin imidazole amide (1 '), levofloxacin imidazole amide (2'), ofloxacin imidazole amide (3 '), levofloxacin imidazole amide (4'), norfluorocarboxylic acid imidazole amide (5 '), pefloxacin imidazole amide (6'), N-acetylnorfloxacin imidazole amide (7 '), cyclopropanecarboxylic acid imidazole amide (8'), N-methylcyclofloxacin imidazole amide (9 '), N-acetylciprofloxacin imidazole amide (10'), enoxacarboxylic acid imidazole amide (11 '), N-methylenoxacin imidazole amide (12'), N-acetylenoxacin imidazole amide (13 '), fleroxacin imidazole amide (14'), N-methyllomefloxacin imidazole amide (15 '), N-methylglucfloxacin imidazole amide (16'), N-methylmoxifloxacin imidazole amide (17 ') and rufloxacin imidazole amide (18'), the structure is as follows:

the corresponding fluoroquinolone hydroxamic acid (1 "-18"): ofloxacin hydroxamic acid (1 '), levofloxacin hydroxamic acid (2'), ofloxacin hydroxamic acid (3 '), levofloxacin hydroxamic acid (4'), norfloxacin hydroxamic acid (5 '), pefloxacin hydroxamic acid (6'), N-acetylnorfloxacin hydroxamic acid (7 '), ciprofloxacin hydroxamic acid (8'), N-methylcyclociprofloxacin hydroxamic acid (9 '), N-acetylciprofloxacin hydroxamic acid (10'), enoxacarboxylic acid (11 '), N-methylenoxacillin hydroxamic acid (12'), N-acetylenoxacin hydroxamic acid (13 '), fleroxacin hydroxamic acid (14'), N-methyllomefloxacin hydroxamic acid (15 '), N-methylgatifloxacin hydroxamic acid (16'), N-methylmoxifloxacin hydroxamic acid (17 '), and norfloxacin hydroxamic acid (18'), the structure is as follows:

the application of the bis-fluoroquinolone oxadiazole urea derivative containing N-methyl enoxacin in preparing antitumor drugs.

Specifically, the anti-tumor drug is a drug for treating lung cancer, liver cancer, stomach cancer, pancreatic cancer, skin melanoma, leukemia or gefitinib-resistant cancer.

The invention is based on an effective bioisostere oxadiazole heterocycle of a pharmacophore urea structural fragment of a targeted tyrosine kinase inhibitor and a fluoroquinolone C-3 carboxyl, designs and synthesizes a 'bis-fluoroquinolone urea derivative' by utilizing a pharmacophore mosaic drug molecule design principle and taking oxadiazole urea as a connecting chain to connect a bis-fluoroquinolone framework, realizes the migration and complementation of superior pharmacophore structures among drugs with different action mechanisms, further innovates a new structure of a drug molecule, achieves the effects of synergy and toxicity reduction, and can be developed as an anti-tumor drug with a brand new structure.

Detailed Description

the preparation operation steps of the N-methylenoxacin C-3 oxadiazolidine II intermediate are as follows: 10.0 g (30.0mmol) of N-methylenoxacin (12) and 2.6g (35.0mmol) of semicarbazide are uniformly mixed and then added into 150g of PPA, and the mixture is stirred and reacted for 12 hours at 140 ℃ in an oil bath. The reaction mixture was cooled to 80 deg.C, 350mL of water was slowly added dropwise, refluxed for 2 hours, cooled to 0 deg.C, adjusted to pH 10.0 with concentrated ammonia water, and left overnight. The resulting solid was collected by filtration, neutralized with water and dried. Recrystallizing the crude product with ethanol-DMF to obtain 7.6g of an analytically pure intermediate II, wherein m.p.226-228 ℃;

the general method for preparing fluoroquinolone carboxylic acid imidazole amide (1 'to 18') is as follows: dissolving fluoroquinolone carboxylic acid FQ-COOH (0.10mol) in anhydrous N, N-diformylamide (500mL), adding 16.2-32.4 g (0.10-0.20 mol) of CDI, stirring and reacting for 10.0-24.0 hours in a water bath at 80-90 ℃, after the reaction is finished, evaporating the solvent under reduced pressure, adding ethyl acetate (500mL), fully stirring and dispersing the solid, filtering and collecting the solid, washing with ethyl acetate, and drying to obtain a fluoroquinolone carboxylic acid imidazole amide crude product, wherein the fluoroquinolone carboxylic acid imidazole amide crude product is directly used for the next reaction without purification.

The fluoroquinolone hydroxamic acid (1 "-18") is prepared by the general method as follows: suspending the crude fluoroquinolone carboxylic acid imidazole amide (0.10mol) in 500mL of pyridine, adding 7.0-35 g (0.1-0.50 mol) of hydroxylamine hydrochloride, stirring in a water bath at 60-75 ℃ for 8.0-24.0 hours for reaction, cooling to room temperature, filtering to collect a solid, washing the obtained solid with pyridine, drying in vacuum at 60-70 ℃, dispersing in a saturated sodium bicarbonate solution (500mL) again, stirring in a water bath at 50-65 ℃ for 3-5 hours, filtering to collect the solid, washing with deionized water to pH 7.0, drying to obtain a crude product, recrystallizing the obtained crude product with absolute ethyl alcohol (or an absolute ethyl alcohol-DMF mixed solvent), and obtaining analytically pure crystalline fluoroquinolone hydroxamic acid (1 '-18').

The general preparation method of the target compound bis-fluoroquinolone oxadiazole urea derivative containing N-methyl enoxacin comprises the following steps: 1.0g of each fluoroquinolone hydroxamic acid (1 '-18') is suspended in an appropriate amount of acetonitrile, CDI (the molar weight of CDI is 1.0-2.0 times that of fluoroquinolone hydroxamic acid) is added, the mixture is stirred and dissolved at normal temperature, an N-methylenoxacin C-3 oxadiazolamine II intermediate (the molar weight of CDI is 1.0 time that of fluoroquinolone hydroxamic acid) is added, the mixture is stirred in a water bath at 55-60 ℃ for 10-24 hours, the mixture is cooled to room temperature, the generated solid is collected through filtration, and the target compound is obtained through recrystallization by using an appropriate solvent.

The preparation of the object compounds of the formulae I-1 to I-18 according to the invention is illustrated in detail below by means of specific examples.

In the following examples, unless otherwise specified, the volume ratio of DMF to ethanol in a DMF-ethanol mixed solvent was 1: 5.

Example 1

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 6-fluoro-7- (4-methylpiperazin-1-yl) -8,1- (1, 3-oxopropyl) -quinolin-4 (1H) -one-3-yl ] -urea (I-1) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: after 1.0g (2.7mmol) of ofloxacin hydroxamic acid (1') was suspended in 25mL of acetonitrile, 0.79g (4.9mmol) of Carbonyldiimidazole (CDI) was added, and the mixture was stirred at room temperature until the material was dissolved. Then, 1.01g (2.7mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 15 hours in a water bath at the temperature of 55-60 ℃. Left overnight, filtered and the resulting solid collected and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-1), wherein the yield is 54 percent, and the m.p. is 215-217 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.57(brs,1H,NH),9.48(s,1H,NH),9.25,8.97(2s,2H,2×2′-H),8.76,8.25(2d,2H,2×5′-H), 5.12～4.74(m,5H,OCH₂CHN and NCH₂) 3.62 to 3.45(m,8H,2 XPiperazine-H), 2.63 to 2.38(m,14H,2 XPiperazine-H and 2 XPNCH)₃),1.64～1.58(m,6H,2×CH₃)；MS(m/z):732[M+H]⁺Calculating the value: 731.77。

example 2

(S) -1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 6-fluoro-7- (4-methylpiperazin-1-yl) -8,1- (1, 3-oxopropyl) -quinolin-4 (1H) -one-3-yl ] -urea (I-1) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.7mmol) of levofloxacin hydroxamic acid (2') is suspended in 25mL of acetonitrile, 0.70g (4.3mmol) of Carbonyldiimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 1.01g (2.7mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 10 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using ethanol to obtain a light yellow crystal (I-2) with the yield of 46 percent and m.p.212-214 ℃.¹H NMR(400MHz,DMSO-d₆)δ:11.57(brs,1H,NH), 9.46(s,1H,NH),9.27,9.06(2s,2H,2×2′-H),8.77,8.23(2d,2H,2×5′-H),5.16～4.73(m,5H, OCH₂CHN and NCH₂) 3.62 to 3.47(m,8H,2 XPiperazine-H), 2.65 to 2.37(m,14H,2 XPiperazine-H and 2 XPNCH)₃),1.66～1.58(m,6H,2×CH₃)；MS(m/z):732[M+H]⁺Calculating the value: 731.77.

example 3

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3-

[6, 7-difluoro-8, 1- (1, 3-oxopropyl) -quinolin-4 (1H) -one-3-yl ] -urea (I-3) having the chemical structure:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: oxofluorocarboxylic acid hydroxamic acid (3 ″)1.0g (3.4mmol) was suspended in 25mL of acetonitrile and Carbonyldiimidazole (CDI)0 was added.82g (5.1mmol) were stirred at room temperature until the material dissolved. Then, 1.27g (3.4mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 22 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-3), wherein the yield is 62%, and the m.p.220-222 ℃.¹H NMR(400MHz,DMSO-d₆)δ:11.56 (brs,1H,NH),9.46(s,1H,NH),9.23,8.98(2s,2H,2×2′-H),8.67,8.18(2d,2H,2×5′-H), 4.97～4.68(m,5H,OCH₂CHN and NCH₂) 3.62 to 3.43(m,4H, piperazine-H), 2.65 to 2.36(m,7H, piperazine-H and NCH)₃),1.64～1.57(m,6H,2×CH₃)；MS(m/z):652[M+H]⁺Calculating the value: 651.61.

example 4

(S) -1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [6, 7-difluoro-1, 8- (1, 3-oxopropyl) -quinolin-4 (1H) -one-3-yl ] -urea (I-4) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (3.4mmol) of levofloxacin hydroxamic acid (4') was suspended in 25mL of acetonitrile, 0.82g (5.1mmol) of Carbonyldiimidazole (CDI) was added, and the mixture was stirred at room temperature until the material was dissolved. Then, 1.27g (3.4mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 18 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-4), wherein the yield is 52 percent, and the m.p. is 216-218 ℃.¹H NMR(400MHz,DMSO-d₆)δ:11.55 (brs,1H,NH),9.41(s,1H,NH),9.24,9.08(2s,2H,2×2′-H),8.64,8.15(2d,2H,2×5′-H), 4.98～4.67(m,5H,OCH₂CHN and NCH₂) 3.63 to 3.42(m,4H, piperazine-H), 2.64 to 2.37(m,7H, piperazine-H and NCH)₃),1.62～1.58(m,6H,2×CH₃)；MS(m/z):652[M+H]⁺Calculating the value: 651.61.

example 5

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6-fluoro-7-chloro-quinolin-4 (1H) -one-3-yl ] -urea (I-5) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (3.5mmol) of norfluorocarboxylic acid hydroxamic acid (5') was suspended in 25mL of acetonitrile, 0.98g (6.0mmol) of Carbonyldiimidazole (CDI) was added thereto, and the mixture was stirred at room temperature until the materials were dissolved. Then, 1.31g (3.5mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 24 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-5), wherein the yield is 57 percent, and the m.p.224-226 ℃.¹H NMR(400MHz,DMSO-d₆)δ:11.54 (brs,1H,NH),9.45(s,1H,NH),9.18,8.94(2s,2H,2×2′-H),8.67～8.13(m,2H,2×5′-H),7.54(d, 1H,8′-H),4.84～4.42(m,4H,2×NCH₂) 3.62 to 3.42(m,4H, piperazine-H), 2.63 to 2.37(m,7H, piperazine-H and NCH)₃),1.62～1.40(m,6H,2×CH₃)；MS(m/z):640[M+H]⁺(³⁵Cl), calculated: 640.05.

example 6

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-6) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.9mmol) of pefloxacin hydroxamic acid (6') was suspended in 25mL of acetonitrile, 0.61g (3.7mmol) of Carbonyldiimidazole (CDI) was added, and the mixture was stirred at room temperature until the material was dissolved. Then adding N-methyl enoxacin C-3 oxadiazolidine II intermediate1.08g (2.9mmol) of the compound is stirred for 20 hours in a water bath at 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-6) with the yield of 55 percent and m.p. 226-228 ℃.¹H NMR(400MHz,DMSO-d₆)δ:11.58 (brs,1H,NH),9.46(s,1H,NH),9.25,9.14(2s,2H,2×2′-H),8.72～8.24(m,2H,2×5′-H),7.56(d, 1H,8′-H),4.86～4.46(m,4H,2×NCH₂) 3.65 to 3.46(m,8H,2 XPiperazine-H), 2.66 to 2.37(m,14H,2 XPiperazine-H and 2 XPNCH)₃),1.68～1.45(m,6H,2×CH₃)；MS(m/z):704[M+H]⁺Calculating the value: 703.76.

example 7

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6-fluoro-7- (4-acetylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-7) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.7mmol) of N-acetyl norfloxacin hydroxamic acid (7') was suspended in 25mL of acetonitrile, 0.75g (5.4mmol) of Carbonyldiimidazole (CDI) was added, and the mixture was stirred at room temperature until the material was dissolved. Then, 1.01g (2.7mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 20 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-7) with the yield of 62 percent and m.p.225-227 ℃.¹H NMR(400MHz,DMSO-d₆) Delta.11.62 (brs,1H, NH),9.53(s,1H, NH),9.32,9.18(2s,2H, 2X 2 ' -H),8.74 to 7.66(m,3H, 2X 5 ' -H and 8 ' -H), 4.94 to 4.47(m,4H, 2X NCH)₂) 3.63 to 3.45(m,8H,2 XPiperazine-H), 2.57 to 2.36(m,14H,2 XPiperazine-H, NCH)₃and Ac),1.68～1.47(m,6H,2×CH₃)；MS(m/z):732[M+H]⁺Calculating the value: 731.77.

example 8

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-cyclopropyl-6-fluoro-7-chloro-quinolin-4 (1H) -one-3-yl ] -urea (I-8) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (3.4mmol) of cyclopropyl carboxylic acid hydroxamic acid (8') was suspended in 25mL of acetonitrile, 0.97g (6.0mmol) of Carbonyldiimidazole (CDI) was added, and the mixture was stirred at room temperature until the materials were dissolved. Then, 1.27g (3.4mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 24 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-8), wherein the yield is 75%, and the m.p.232-234 ℃.¹H NMR(400MHz,DMSO-d₆)δ:11.58 (brs,1H,NH),9.46(s,1H,NH),9.27,9.06(2s,2H,2×2′-H),8.72～7.56(m,3H,5′-H and 8′-H), 4.87～4.57(m,3H,NCH₂and CH), 3.62-3.43 (m,4H, piperazine-H), 2.57-2.36 (m,7H, piperazine-H and CH)₃),1.64～0.93(m,7H,CH₃and CH₂CH₂)；MS(m/z):652[M+H]⁺(³⁵Cl), calculated: 652.07.

example 9

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-cyclopropyl-6-fluoro-7- (4-methylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-9) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.8mmol) of N-methyl ciprofloxacin hydroxamic acid (9') is suspended in 25mL of acetonitrile, 0.81g (5.0mmol) of Carbonyl Diimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then adding 1.04g (2.8mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate, stirring 1 in water bath at 55-60 DEG CFor 6 hours. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-9), wherein the yield is 58 percent, and the m.p.221-223 ℃.¹H NMR(400MHz,DMSO-d₆) Delta.11.62 (brs,1H, NH),9.48(s,1H, NH),9.27,9.15(2s,2H, 2X 2 ' -H), 8.78-7.66 (m,3H, 2X 5 ' -H and 8 ' -H), 4.94-4.61 (m,3H, NCH)₂and CH),3.65 to 3.44(m,8H,2 XPiperazine-H), 2.63 to 2.38(m,14H,2 XPiperazine-H and 2 XPNCH)₃),1.68～1.16(m,7H,CH₃and CH₂CH₂)；MS(m/z):716[M+H]⁺Calculating the value: 715.77.

Example 10

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-cyclopropyl-6-fluoro-7- (4-acetylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-10) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.6mmol) of N-acetyl ciprofloxacin hydroxamic acid (10') is suspended in 25mL of acetonitrile, 0.75g (4.6mmol) of Carbonyl Diimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 0.97g (2.6mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate is added, and the mixture is stirred for 22 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal (I-10), wherein the yield is 65 percent, and the m.p.223-225 ℃.¹H NMR(400MHz,DMSO-d₆) Delta.11.65 (brs,1H, NH),9.51(s,1H, NH),9.34,9.18(2s,2H, 2X 2 ' -H),8.84 to 7.74(m,3H, 2X 5 ' -H and 8 ' -H), 4.95 to 4.63(m,3H, NCH)₂and CH), 3.65-3.46 (m,8H,2 XPiperazine-H), 2.67-2.37 (m,14H,2 XPiperazine-H, NCH)₃and Ac),1.68～1.25(m,7H,CH₃and CH₂CH₂)；MS(m/z):744[M+H]⁺Calculating the value: 743.78.

Example 11

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6-fluoro-7-chloro- [1,8] naphthyridin-4 (1H) -one-3-yl ] -urea (I-11) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (3.5mmol) of enoxohydroxamic acid (11') was suspended in 25mL of acetonitrile, 1.13g (7.0mmol) of Carbonyldiimidazole (CDI) was added, and the mixture was stirred at room temperature until the material was dissolved. Then, 1.31g (3.5mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 24 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal target product (I-11), wherein the yield is 68 percent, and the m.p.231-233 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.58(brs,1H,NH),9.46(s,1H,NH),9.25,9.17(2s,2H,2×2′-H),8.86～8.37(m,2H,2×5′-H), 4.88～4.82(m,4H,2×NCH₂) 3.64 to 3.43(m,4H, piperazine-H), 2.62 to 2.36(m,7H, piperazine-H and NCH)₃), 1.67～1.60(m,6H,2×CH₃)；MS(m/z):641[M+H]⁺(³⁵Cl), calculated: 641.07.

example 12

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -urea (I-12) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.9mmol) of N-methylenoxacin hydroximic acid (12') is suspended in 25mL of acetonitrile, 0.65g (4.0mmol) of Carbonyldiimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then adding 1.08g (2.9mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate, and stirring 1 in water bath at 55-60 DEG CFor 8 hours. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a golden yellow crystal target (I-12), wherein the yield is 53%, and the m.p. is 218-220 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.65(brs,1H,NH),9.54(s,1H,NH),9.35,9.26(2s,2H,2×2′-H),8.87,8.52(2d,2H,2×5′-H),, 4.96～4.87(m,4H,2×NCH₂) 3.63 to 3.46(m,8H,2 XPiperazine-H), 2.66 to 2.37(m,14H,2 XPiperazine-H and 2 XPNCH)₃),1.72～1.68(m,6H,2×CH₃)；MS(m/z):705[M+H]⁺Calculating the value: 704.75.

example 13

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6-fluoro-7- (4-acetylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -urea (I-13) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.7mmol) of N-acetyl enoxacin hydroximic acid (13') is suspended in 25mL of acetonitrile, 0.84g (5.2mmol) of Carbonyl Diimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 1.01g (2.7mmol) of the intermediate of N-methylenoxacin C-3 oxadiazolidine II is added, and the mixture is stirred for 20 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a golden yellow crystal target (I-13) with the yield of 62 percent and m.p. 226-228 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.67(brs,1H,NH),9.55(s,1H,NH),9.36,9.28(2s,2H,2×2′-H),9.14,8.78(2d,2H,2×5′-H), 5.04～4.87(m,4H,2×NCH₂) 3.65 to 3.46(m,8H,2 XPiperazine-H), 2.67 to 2.38(m,14H,2 XPiperazine-H, NCH)₃and Ac),1.76～1.68(m,6H,2×CH₃)；MS(m/z):733[M+H]⁺Calculating the value: 732.76.

example 14

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [1- (2-fluoroethyl) -6, 8-difluoro-7- (4-methylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-14) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.6mmol) of fleroxacin hydroxamic acid (14') is suspended in 25mL of acetonitrile, 0.62g (3.8mmol) of Carbonyl Diimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 0.97g (2.6mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate is added, and the mixture is stirred for 16 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a golden yellow crystal target product (I-14), wherein the yield is 62%, and m.p.227-229 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.63(brs,1H,NH),9.52(s,1H,NH),9.34～8.57(m,4H,2×2′-H and 2×5′-H),4.94～4.72(m,6H, FCH₂CH₂and NCH₂) 3.65 to 3.46(m,8H,2 XPiperazine-H), 2.67 to 2.38(m,14H,2 XPiperazine-H and 2 XPNCH)₃),1.48(t,3H,CH₃)；MS(m/z):740[M+H]⁺Calculating the value: 739.74.

example 15

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-ethyl-6, 8-difluoro-7- (3, 4-dimethylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-15) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.6mmol) of N-methyl lomefloxacin hydroxamic acid (15') is suspended in 25mL of acetonitrile, 0.55g (3.4mmol) of Carbonyl Diimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 0.97g (2.6mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate is added, and the mixture is stirred for 16 hours in a water bath at the temperature of 55-60 ℃. Standing overnight, and collecting the solidBody, acetonitrile washes. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a golden yellow crystal target (I-15) with the yield of 46 percent and m.p.217-219 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.58(brs,1H,NH),9.46(s,1H,NH),9.22,9.05(2s,2H,2×2′-H),8.74～8.13(m,2H,2×5′-H), 4.88～4.47(m,4H,2×NCH₂) 3.63 to 3.45(m,8H,2 XPiperazine-H), 2.64 to 2.37(m,13H,2 XPiperazine-H and 2 XPNCH)₃),1.66～1.47(m,9H,3×CH₃)；MS(m/z):736[M+H]⁺Calculating the value: 735.78.

example 16

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-cyclopropyl-6-fluoro-8-methoxy-7- (3, 4-dimethylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-16) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.5mmol) of N-methyl gatifloxacin hydroxamic acid (16') is suspended in 25mL of acetonitrile, 0.75g (4.6mmol) of Carbonyldiimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 0.93g (2.5mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate is added, and the mixture is stirred for 15 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a golden yellow crystal target (I-16) with the yield of 56 percent and the m.p. of 218-220 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.62(brs,1H,NH),9.46(s,1H,NH),9.27,9.08(2s,2H,2×2′-H),8.76,7.76(2d,2H,2×5′-H), 4.87～4.62(m,3H,NCH₂and CH),3.88(s,3H,OCH₃) 3.63 to 3.45(m,8H,2 XPiperazine-H), 2.58 to 2.37(m,13H,2 XPiperazine-H and 2 XPNCH)₃),1.68～1.15(m,10H,2×CH₃and CH₂CH₂)；MS (m/z):760[M+H]⁺Calculating the value: 759.82.

example 17

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 1-cyclopropyl-6-fluoro-8-methoxy-7- (1-methyl-octahydropyrrolo [3,4-b ] pyridin-6-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-17) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.3mmol) of N-methylmoxifloxacin hydroximic acid (17') is suspended in 25mL of acetonitrile, 0.65g (4.0mmol) of Carbonyldiimidazole (CDI) is added, and the mixture is stirred at normal temperature until the materials are dissolved. Then, 0.86g (2.3mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate is added, and the mixture is stirred for 15 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal target product (I-17), wherein the yield is 46%, and the m.p.223-225 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.62(brs,1H,NH),9.54(s,1H,NH),9.23,9.15(2s,2H,2×2′-H),8.68～8.25(m,2H,2×5′-H), 4.88～4.58(m,3H,NCH₂and CH),3.92(s,3H,OCH₃) 3.66 to 3.32(m,8H, piperazine-H and pyrrolidine ring-H), 2.62 to 2.28(m,13H, piperazine-H and piperidine ring-H and 2 XNCH)₃),2.17～1.13(m,12H,CH₃Piperidine ring-H and CH₂CH₂)；MS(m/z):786[M+H]⁺Calculating the value: 785.86.

example 18

1- {2- [ 1-ethyl-6-fluoro-7- (4-methylpiperazin-1-yl) - [1,8] naphthyridin-4 (1H) -one-3-yl ] -1,3, 4-oxadiazol-5-yl } -3- [ 6-fluoro-8, 1-thienyloxy-7- (4-methylpiperazin-1-yl) -quinolin-4 (1H) -one-3-yl ] -urea (I-18) having the chemical formula:

the preparation method of bis-fluoroquinolone oxadiazole urea in this example is as follows: 1.0g (2.6mmol) of rufloxacin hydroxamic acid (18') is suspended in 25mL of acetonitrile, 0.72g (4.4mmol) of Carbonyldiimidazole (CDI) is added, and the mixture is stirred at normal temperature to obtain a productAnd (4) dissolving the materials. Then, 0.97g (2.6mmol) of N-methylenoxacin C-3 oxadiazolidine II intermediate is added, and the mixture is stirred for 18 hours in a water bath at the temperature of 55-60 ℃. After standing overnight, the resulting solid was collected by filtration and washed with acetonitrile. Recrystallizing the crude product by using a DMF-ethanol mixed solvent to obtain a light yellow crystal target product (I-18), wherein the yield is 57 percent, and the m.p.225-227 ℃.¹H NMR(400MHz,DMSO-d₆)δ: 11.58(brs,1H,NH),9.45(s,1H,NH),9.18,9.06(2s,2H,2×2′-H),8.72,8.16(2d,2H,2×5′-H), 4.84(q,2H,NCH₂),4.32～3.36(m,12H,SCH₂CH₂) and 2 XPiperazine-H), 2.58 to 2.37(m,14H,2 XPiperazine-H and 2 XPCH)₃),1.65(t,3H,CH₃)；MS(m/z):734[M+H]⁺Calculating the value: 733.81.

test examples

In vitro antitumor Activity assay of bis-fluoroquinolone oxadiazolyl Urea derivatives prepared in examples 1-18

1. Test sample

Using 18 new bis-fluoroquinolone oxadiazole urea derivatives provided in examples 1-18 and the classical antitumor topoisomerase inhibitor 10-Hydroxycamptothecin (HC), the urea tyrosine kinase inhibitor Regorafenib (RRF), Cabozantinib (CZT), and the parent compound N-Methylenoxacin (MEX) as test samples, 22 samples in total, wherein HC, RRF, CZT, and MEX are positive control test groups, and examples 1-18 are sample test groups;

3- (4, 5-dimethyl-2-thiazole) -2, 5-diphenyl tetrazolium bromide (MTT), HC, RRF, CZT and MEX are all products of Sigma company; the RPMI-1640 culture solution is a product of GIBCO company; other used reagents are all domestic analytical pure reagents.

The experimental cancer cell strains are respectively a human non-small cell lung cancer cell strain A549, a human liver cancer cell strain SMCC-7721, a human gastric cancer cell strain HGC27, a human pancreatic cancer cell strain Capan-1, a human skin melanoma cell strain A375 and a human leukemia cell strain HL60 which are purchased from a Shanghai cell bank of Chinese academy of sciences; gefitinib-resistant cell strain K562G was purchased from Tianjin hematopathy institute; normal cells were VerO cells, which were obtained from Shanghai city, park, Inc.

2. Measurement method

The determination method comprises the following specific steps:

(1) firstly, the 22 samples were dissolved in dimethyl sulfoxide (DMSO) to prepare 1.0X 10^-4mol·L^-1The stock solution was diluted to have a concentration gradient of 5 (specifically, concentration set to 50, 10, 5, 1.0, 0.1. mu. mol. L) by 10-fold dilution method using 10% by mass of calf serum in RPMI-1640 culture solution^-1) The working fluid of (1);

the first set of experiments: inoculating 5000 cells per well of cancer cell strains HL60, Capan-1 and K562G in logarithmic growth phase to 96-well plate, culturing overnight, adding working solution with 5 concentration gradients, discarding culture medium after 48 hr, adding 1 g.L per well^–1100 mu L of 3- (4, 5-dimethyl-2-thiazole) -2, 5-diphenyl tetrazolium bromide (MTT) solution, then continuously culturing for 4 hours, removing supernatant, adding 150 mu L of DMSO into each hole, slightly shaking for 30 minutes, and then measuring the absorbance (OD) value at the wavelength of 570nm by using a microplate reader;

the second set of experiments: taking cancer cell strains A549, SMCC-7721, HGC27, VERO and A375 in logarithmic growth phase, inoculating 7000 cells per well into a 96-well plate, respectively adding working solution with 5 concentration gradients, and adding 5 g.L per well after 48 hours^–110 mu L of MTT solution is continuously cultured for 4 hours, then 100 mu L of Sodium Dodecyl Sulfate (SDS) solution with the mass percentage concentration of 10 percent is added for culture overnight, and then an OD value is measured at the wavelength of 570nm by using an enzyme-labeling instrument;

(2) the inhibition rate of the test samples with different concentrations on the experimental cancer cells is calculated according to the following formula:

the cancer cell inhibition rate ═ 1 [ (experimental OD value)/control OD value ] × 100%,

then, linear regression is carried out on the cancer cell inhibition rate corresponding to each concentration by the pair value of each concentration of the test sample to obtain a dose-effect equation, and the half Inhibition Concentration (IC) of each test sample to the experimental cancer cell is calculated from the obtained dose-effect equation₅₀) (ii) a Each data was measured in parallel five times and averaged, the results are shown in Table 1.

As can be seen from table 1, the bis-fluoroquinolone oxadiazolyl urea N-methylenoxacin derivatives provided in examples 1 to 18 all have significant proliferation inhibitory activity against 7 experimental cancer cell lines, and particularly show higher activity against human non-small cell lung cancer cell line a549, human pancreatic cancer cell line Capan-1 and human skin melanoma cell line a375, which is significantly stronger than the activity of the parent compound N-methylenoxacin, and at the same time stronger than the activity of the control topoisomerase inhibitor Hydroxycamptothecin (HC), and the activity of most compounds is also stronger than the activity of the control tyrosine kinase inhibitors regorafenib (RRF) and Cabozantinib (CZT). More significantly, the compounds provided in examples 1-18 also showed very strong sensitivity to gefitinib resistant cell line K562G, while showing low toxicity to normal cell VERO cells, with the property of being druggable. Therefore, according to the general approach of drug development, the conventional antitumor in vitro screening is carried out, and then the targeted research is carried out, so that the compound has strong antitumor and anti-drug resistance activity and lower toxicity, and can be used for preparing antitumor drugs by salifying with acid acceptable for human bodies or mixing with medicinal carriers.

TABLE 1 antitumor Activity (IC) of the test samples₅₀)

Claims

1. a bis-fluoroquinolone-based oxadiazole urea derivative containing N-methyl enoxacin is characterized in that: be specifically the compound of the structure shown in the following formula I-1 to I-18:

2. the preparation method of the bis-fluoroquinolone-based oxadiazole urea derivatives containing N-methyl enoxacin according to claim 1, is characterized in that, concrete preparation step comprises:

1) N-methyl enoxacin and semicarbazide hydrochloride are condensed in polyphosphoric acid, and after post-processing, N-methyl enoxacin C-3 oxadiazolamine intermediate II is obtained, and its structural formula is as follows :

2) The fluoroquinolone carboxylic acid series shown in structural formulas 1 to 18 are respectively subjected to condensation reaction with N, N'-carbonyldiimidazole to generate corresponding fluoroquinolone carboxylic acid imidazole amide series, whose structural formula is such as 1' to 18'; the aforementioned preparation The obtained fluoroquinolone carboxylic acid imidazole amide series are respectively reacted with hydroxylamine hydrochloride to obtain the corresponding fluoroquinolone hydroxamic acid series, whose structural formula is shown in 1″～18″:

Formulas 1 to 18:

Formulas 1′～18′:

Formula 1″～18″

3) The fluoroquinolone hydroxamic acid series prepared in step 2) are respectively rearranged into corresponding fluoroquinolone isocyanate series by Lossen under the assisted catalysis of N,N'-carbonyldiimidazole, and are directly prepared with step 1) without treatment. The N-methyl enoxacin C-3 oxadiazolamine intermediate II undergoes a condensation reaction, and after post-treatment, compounds with structures shown in formulas I-1 to I-18 can be prepared.

3. The preparation method of N-methylenoxacin-containing bis-fluoroquinolone-based oxadiazole urea derivatives according to claim 2, characterized in that: in step 2), the formulas 1-18 The shown molar ratio of fluoroquinolone carboxylic acid to N,N'-carbonyldiimidazole is 1:1.0-2.0.

4. The preparation method of N-methylenoxacin-containing bis-fluoroquinolone-based oxadiazole urea derivatives according to claim 2, characterized in that: in step 2), the formula 1'～ The molar ratio of the fluoroquinolone carboxylic acid imidazole amide represented by 18' to the hydroxylamine hydrochloride is 1:1.0-5.0.

5. The preparation method of the N-methyl enoxacin-containing bis-fluoroquinolone-based oxadiazole urea derivatives according to claim 2, characterized in that: in step 3), the formula 1"～ The molar ratio of the fluoroquinolone hydroxamic acid represented by 18" to N,N'-carbonyldiimidazole is 1:1.0 to 2.0, and the fluoroquinolone hydroxamic acid represented by the formula 1" to 18" and the N-methyl quinolone hydroxamic acid are The molar ratio of norfloxacin C-3 oxadiazolamine intermediate II was 1:1.

6. The application of the N-methylenoxacin-containing bis-fluoroquinolone-based oxadiazole urea derivatives of claim 1 in the preparation of antitumor drugs.

7. The application of the N-methylenoxacin-containing bis-fluoroquinolone-based oxadiazole urea derivatives according to claim 6 in the preparation of an antitumor drug, wherein the antitumor drug is Drugs to treat lung, liver, stomach, pancreatic, skin melanoma, leukemia, or gefitinib-resistant cancers.