WO2005054250A1 - Tricyclacetalactonin, Preparation and Use there of - Google Patents

Abstract

The invention relates to tricyclacetalactonin, preparation and use there of. Said tricyclacetalactonin is prepared from Streptomyces albogriseolus of A2-2002. As being proved by the experiments, said compounds can be used as cell cycle inhibitor, apoptosis inducer or anti­tumor agent.

Description

 Tricyclic acetal lactone and its preparation method and use

Technical field:

 The present invention relates to a tricyclic acetalactin compound, and a method for preparing a tricyclic acetalactin compound by using Streptomyces spp., And the present invention also relates to the preparation of a cell cycle inhibitor and apoptosis induction of this type of compound Agent or antineoplastic agent. Background technique:

Compounds with cell cycle inhibition, apoptosis induction, or anti-tumor activity in microbial fermentation products or plant materials have been documented in the literature [Nagata Hiyuki et al., New regulatory substance U ⁷ 'Mouth: ^ Xi, Zi o manufacturing method, cell cycle encumber Ji anti-neoplasm Ji, Japanese Patent Laid-open public information (A), Laid-open 9-59275, announcement date of the 1997 (1997) March 4; Nagata Hiroyuki and other new regulations substance 7 seven BU ⁷ evening U, Cell cycle obstruction J: Anti-ulcer ulcer, Japanese Patent, Japanese Patent Publication (A), Japanese Patent Application Laid-Open No. 9-87269, Announcement date Heisei 9 (1997) March 31] and literature [Cui Chengbin et al., Carbazole Bio Alkali cell cycle inhibitors, apoptosis inducers and their preparations, Chinese patent CN1309964A; Cui Chengbin et al., Carbazole alkaloid anticancer drugs and their preparation, Chinese patent CN1357327A] have been reported. However, the chemical structures of the compounds reported in these documents are different parent structures such as carbazoles. Summary of the invention:

 The present invention aims to provide a compound with a novel chemical skeleton structure having anti-tumor activity such as cell cycle inhibition, apoptosis induction, and direct killing of cancer cells.

 The inventors have found a new chemical skeleton structure of tricyclic acetalactin through tenacious efforts

(tricyclacetalactonin) new compounds, as shown in formula I:

式 I中， 为氨基或羟基； R₂为羟基、 氨基或氢。

In formula I, is amino or hydroxyl; R ₂ is hydroxyl, amino or hydrogen.

 Its structural characteristics are: The molecular skeleton structure is a tricyclic skeleton structure composed of a five-membered carbocyclic ring and two six-membered oxo rings, wherein the five-membered carbocyclic ring is the basic carbon skeleton of the structure, An acetal ether type six-membered oxo ring formed by condensation of an aldehyde group on the five-membered carbocyclic ring with an enol hydroxyl group on a side chain carbon adjacent to the aldehyde group, and another six-membered oxo The ring is an acetal lactone six-membered oxo ring formed by the condensation of a hemiacetal hydroxyl group produced by condensation with an enol hydroxyl group and a hydroxyl group on a carboxyl group connected to a five-membered carbon ring. Different substituents are attached at different positions of the above-mentioned tricyclic skeleton structure,

 In the present invention, the method of sulforhodamine B (SRB) and flow cytometry combined with a method for detecting cell morphological characteristics under a microscope are used to test compounds of formula I on mouse breast cancer tsFT210 cells and human chronic myeloid leukemia K562. Cell and human colorectal cancer HCT-15 cells have the effects of cell proliferation inhibition, cell cycle inhibition, apoptosis induction, and direct killing of the cells. Experiments have shown that the compounds of formula I can exert their anti-tumor effect on tumor cells by inhibiting cell cycle turnover, inducing cancer cell apoptosis or direct killing, etc., by showing biological activities that inhibit tumor cell proliferation.

 The compounds of formula I of the present invention are therefore useful as cell cycle inhibitors, apoptosis inducers, or tumor cell killers.

 The compound of formula I is compatible with various pharmaceutically acceptable carriers, excipients or excipients, and can be made into antitumor drugs for use in the treatment of tumors.

 The compounds of formula I can also be used as life-molecular research as low-molecular biological probes that inhibit the cell cycle or induce apoptosis. When the compound of formula I is used as a cell cycle inhibitor or apoptosis inducer in life science research, it can be dissolved in methanol, water or aqueous methanol, and it can also be used in an aqueous solution of dimethylsulfoxide. The compound of formula I of the present invention can be obtained by fermenting and culturing a microorganism capable of producing tricyclic lactones, obtaining a fermented product containing tricyclic lactones, and then separating and purifying the fermented products.

 The separation and purification includes conventional methods for separation and purification of natural products, which are well known to those skilled in the art, such as liquid-liquid extraction, column chromatography, thin-layer chromatography and recrystallization.

 The microorganism capable of producing tricyclic acetal lactone includes Streptomyces genus-producing bacteria and the like, such as Streptomyces albogriseolus

In an experimental example of the present invention, the used protobacter was isolated from a marine mud sample in Qingdao offshore and identified as a strain A2-2002 of Streptomyces albogriseolus by taxonomic research. This strain was deposited in the General Microbiology Center of the China Microbial Strain Collection Management Committee (Deposit No .: CGMCC 1005) on September 18, 2003. The Streptomyces albogriseolus A2-2002 CGMCC 1005 strain has the following microbiological characteristics: 1 Culture characteristics and morphological characteristics on various media

 Cultivation characteristics: Synthetic No. 1 agar, Sucrose Chard's agar, Glycerol aspartin agar, Kirschner's synthesis

No. 1 agar, inorganic salt starch agar, oat flour agar, potato infusion agar and other 7 media were cultured at 28 ° C for 7-12 days, and the mycelial color and pigment production were observed. The characteristics are shown in Table 1. Table 1 Culture characteristics of A2-2002 strains on 7 media

 S S dagger m

 Aerial mycelium basal mycelium soluble pigment ~~ Gao's synthetic No. 1 Qiongyue white light gray

 Sucrose agar white light gray

 Glycerol Aspartate

 Kirschner Synthetic No. 1 Qiongyue White White White White White

 Color color color color color

 Inorganic salt starch agar

 Oatmeal Agar

 Potato agar agar morphological characteristics No. 1 agar and sucrose agar agar apricot brown

 After cultivating the yellow-brown kernel tooth brown-yellow brown at 28 ° C for 7-10 days, take the insert and observe the brownish yellowish yellow color of the mycelium with an optical microscope and an electron microscope. As a result, the intracellular mycelium of the strain was observed. No transverse septum, no break; aerial mycelium grows abundantly; spore filaments are tight and open spiral; spores are ovoid, with rough surface with warts (photographs of optical microscope and electron microscope are omitted). Faint

 2 Chemical taxonomic characteristics

 Analysis of the chemical composition of the color cell wall According to the thin layer chromatography (TLC) method of Hasegawa, the whole cell hydrolysate DAP (diaminopimelate) amino acid and glycoform analysis were performed. The results showed that the A2- 2002 strain whole cell hydrolysate Contains LL-DAP (Diaminopimelic acid), glycine; no characteristic sugar (glycoform C). The chemical composition of the cell wall is type I.

3 physiological and biochemical characteristics

 Physiological and biochemical identification of the strains was performed according to the content of "Bergey, s Manual of Systematic Bacteriology" Vol. IV. The physiological and biochemical characteristics of the A2-2002 strain are shown in Table 2 Table 2

 Feature result feature result

D-glucose + gelatin liquefaction +

 Milk coagulation

 L-Arabinose + 'Milk Softened +

D-xylose-starch hydrolysis + D-fructose-nitrate reduction + Growth on sucrose + cellulose―

Rhamnose-H ₂ S production-Inositol.-Tyrosinase production-Lactose + melanoid production-

 D-mannitol +

Raffinose ― Based on the results of the above test, based on the principle of morphological characteristics combined with cell wall chemical components, the basal mycelium of the A2-2002 strain has no transverse septum and does not break; the spore filaments are spiral, and the spores have a rough surface. Wart, cell wall chemical component type I, belongs to the genus Streptomyces (_Stre _P tom _y . Based on the culture characteristics and physiological and biochemical characteristics of the principle of breeding, the aerial mycelium of the A2-2002 strain is light gray, grayish white, endophytes wire yellow, brown tones, physiological and biochemical features of the integrated results are very similar to Streptomyces albogriseolus, so A2-2002 strain named Streptomyces albogriseolus {Streptomyces albogriseolus) _a particular note is made by fermenting microorganism The method for taking the compound of the formula I of the present invention can use any other Streptomyces microorganism capable of producing tricyclic acetal lactone compounds, and any Streptomyces microorganism capable of producing tricyclic acetal lactone compounds can be used as a product Bacillus bacteria are used to prepare compounds of formula I. Brief description of the drawings:

FIG. 1A is a single crystal X-ray diffraction crystal structure of Compound 1. FIG. FIG. 1B is a flow cytometry histogram of mouse breast cancer tsFT210 cells after being treated with different concentrations of Compound I for 17 hours. The upper left figure is the blank control group, and the remaining five are different concentrations of Compound I. (For the concentration, see the upper right of each figure) Treatment group, the curve part in the figure is the measured data, and the black filled part is the calculated value. FIG. 2A is a flow cytometry histogram of human chronic myeloid leukemia K562 cells treated with different concentrations of Compound I for 24 hours. The upper left figure is the blank control group, and the remaining five are different concentrations of Compound I (see the upper right of each figure) for the treatment group. The curve in the figure is the measured data, and the black filled part is the calculated value. FIG. 2B is a flow cytometry histogram of human colorectal cancer HCT-15 cells after being treated with different concentrations of Compound I for 24 hours. The upper left figure is the blank control group, and the remaining five are different concentrations of Compound I (see the upper right of each figure) for the treatment group. The curve in the figure is the measured data, and the black filled part is the calculated value. Figure 3A is a flow cytometry measured after 17 hours of treatment of mouse breast cancer tsFT210 cells with different concentrations of compound II Cell histogram. The upper left figure is the blank control group, and the remaining five are compounds with different concentrations (see the upper right of each figure) for the treatment group. The curve in the figure is the measured data, and the black filled part is the calculated value. FIG. 3B is a flow cytometry histogram of human chronic myelogenous leukemia K562 cells after being treated with different concentrations of Compound II for 24 hours. The upper left figure is the blank control group, and the remaining five are compounds with different concentrations (see the upper right of each figure) for the treatment group. The curve in the figure is the measured data, and the black filled part is the calculated value. FIG. 4 is a flow cytometry histogram of human colorectal cancer HCT-15 cells after being treated with different concentrations of Compound II for 24 hours. The upper left figure is the blank control group, and the remaining five are compounds with different concentrations (see the upper right of each figure) for the treatment group. The curve in the figure is the measured data, and the black filled part is the calculated value. detailed description:

 The chemical structure of compound I referred to in the following examples is: a compound of formula I, wherein Ri is amino; is a hydroxyl group; the chemical structure of compound II is: a compound of formula I, wherein is amino; is hydrogen:

式 I

Formula I

 In the formula, the Arabic numeral is the standard position of the carbon atom in the chemical structure. Example 1 Fermentation Production and Isolation of Compound I

1 Fermentation production

 Fermentation culture of vegetative bacteria according to the conventional method of cultivating microorganisms, take an appropriate amount of Streptomyces albogriseolus A2-2002 strain, inoculate it on Gaussian synthetic agar solid slant culture medium, and culture in a 28 ° C incubator for 4 days .

Take an appropriate amount of Streptomyces albogriseolus) A2-2002 strain cultured on a slant for 4 days. Planting a culture medium containing 100 ml of seed [Media composition (g / l): glucose 20.0, K ₂ HP0 ₄ 0.5, MgS0 ₄ 0.5, beef extract 3.0, corn slurry 3.0, yeast extract 10.0, starch 10.0, CaC0 ₃ 2.0 , PH 7.0] in an Erlenmeyer flask, cultured at 28 ° C, 120 rpm for 48 hours in a shaker to obtain a seed culture solution of Streptomyces spp. Take an appropriate amount of the seed culture solution and inoculate 100 cultured production culture liquids containing 100 ml of production culture medium at a 5% inoculation amount [medium composition (g / l): glucose 20.0, K ₂ HP0 ₄ 0.5, MgS0 ₄ 0.5, beef extract 3.0, Corn pulp 3.0, yeast extract 10.0, starch 10.0, CaC0 ₃ 2.0, pH 7.0] in a conical flask, loaded at 28 ° C, 120 rpm on a shaker for 7 days of production fermentation to obtain substituted tricyclic acetal About 10 liters of the fermentation culture of Streptomyces albogriseolus A2-2002 strain of the lactone target compound. 2 Preparation of crude extract of bacterial cells containing compound I

 The fermented culture of Streptomyces albogriseolus K2-20Q2 strain (about 10 liters) was suction filtered, and the filtrate was discarded to obtain 830 g of bacteria. The mycelium was immersed in 1.5 liters of 80% acetone aqueous solution and stirred overnight at room temperature for extraction, centrifuged at 4000 rpm for 15 minutes, the supernatant was taken, and concentrated under reduced pressure to contain no acetone. The ester was extracted three times, and the combined ethyl acetate extracts were concentrated under reduced pressure to obtain a crude mycelium extract (2.5 g) containing compound I.

3 Isolation and purification of compound I

! A mixture of Streptomyces albogriseolus Streptomyces albogriseolus hi- i I is ^a compound of the fermentation culture mycelium crude extract (2.5 g), washed with 10 ml of chloroform - methanol (9: 1) mixed solvent was added to dissolve 10 200-300 mesh silica gel G (product of Qingdao Ocean Chemical Group Co., Ltd.), mixed with 30 grams of silica gel G produced by Qingdao Ocean Chemical Group Co., Ltd. on a glass decompression column with petroleum ether-chloroform- The methanol mixture was used as the elution solvent system. The column was subjected to reduced pressure column chromatography. The polarity of the elution solvent was increased by increasing the amount of chloroform in petroleum ether or methanol in chloroform. Each fraction was 50 ml. After combined with thin layer chromatography, 14 components were obtained, namely Fr-1 (20 mg, petroleum ether eluate) and Fr-2 (65 mg, petroleum ether-chloroform 1: 1 eluate). ), Fr-3 (120 mg, petroleum ether-chloroform 1: 1 eluate), Fr-4 (260 mg, chloroform and chloroform-methanol 99: 1 eluate), Fr-5 (70 mg, chloroform- Methanol 99: 1 eluate), Fr-6 (80 mg, chloroform-methanol 99: 1-98: 2 eluate), Fr-7 (90 mg, Chloroform-methanol 98: 2 eluate), Fr-8 (150 mg, chloroform-methanol 97: 3 eluate), Fr-9 (124 mg, chloroform-methanol 97: 3-96: 4 eluate) , Fr-10 (190 mg, chloroform-methanol 96: 4 eluate), Fr-11 (108 mg, chloroform-methanol 95: 5 eluate), Fr-12 (252 mg, chloroform-methanol 94: 6 — 90:10 eluate), Fr-13 (250 mg, chloroform-methanol 90: 10-60: 40 eluate), Fr-14 (93 mg, chloroform-methanol 50: 50—100: 0 eluent Thing). Then, a screening model of temperature-sensitive mouse breast cancer tsFT210 cells using flow cytometry combined with morphological detection was used to determine cell cycle inhibition, apoptosis induction and cell necrosis. The activity is an anti-cancer index, and the activity of each component is tested to determine that Fr-11 has the relevant biological activity, thereby obtaining the chromatographic component Fr-11 (108 mg) containing Compound I. In the process of recovering the solvent under reduced pressure, the component Fr-11 gave the pale yellow crude crystals of the compound I, which was purified by repeated recrystallization in methanol to obtain 49 mg of pure colorless transparent granular crystals of the compound I.

Compound I colorless transparent square crystal, mp 236 · 9- 237.9 ° C, [σ] ¾ ⁷ -377.9 ° (c 1.0, MeOH), molecular formula C _{1 ()} H ₉ N0 ₅ , TOF-MS m / z: 224 [M + H] ⁺ , 246 [M + Na] ⁺ ; Positive HR-TOF-MS m / z, found 224.0585 [M + H] ⁺ , calculated 224.0559 (C _IO H ₁₀ N0 ₅ [M + H] ⁺ ); Negative HR-TOF-MS m / z-. Found 222.0379 [M-H]-, Calculated 222.0402 (job [M-H] —). UV nm (log ε) in MeOH: 219 (3.99), terminal absorption. IR v _mx cm ^-1 (KBr): 3437 (NH), 3314, 3184 (OH), 3095, 3002 (olefin protons), 2947 (methine protons), 1746 (lactone carbonyl), 1690 (amide I absorption, — C0N <), 1654, 1602 (amide II absorption, N¾), 1406 (C = C), 1324, 1181 (= C-0), 1136, 1115, 1097 (C_0), 1004, 953 (m), 937 (m ), 917 (vs), 847 (m), 791 (m), 775 (m), 697 (m), 675 (m), 642 (m), 576 (s), 530 (m), 419 (m ). See Table 3 for ¾ and ¹³ C NMR data. Table 3 600 MHz ¾ and 150 MHz ¹³ C NMR data of compound I in deuterated chloroform ^a) Position number ¾-¾ C0SY ^b) HMBC ^0>

 1 163. 00 s 3

 2 144. 27 s 3, 4, 9, N-Ha

3 6.82 d (5.8) 4 106. 35 d 4

4 3.39 td (ca.5.8, 3, 8, 9 ^d> 49. 30 d 3, 6, 7, 9

 ιτΝ

 5 85., 03 s 4, 6, 7,8

6 6.43 d (5.9) 7 143. 37 d 7, 8

7 6.28 dd (5.9, 3.3) 6, 8 133., 18 d 6, 8

8 2.95 m 4, 7, 9 41.. 30 d 3, 4, 6, 7

9 6.16 dd (ca.1.8, 4 ^d) , 8 97., 42 d 4, 6 ^e) ,

 irN

 10 172. .84 s 4, 9

N-Ha 8.62 br s N-Hb

 N-Hb 8.30 br s N-Ha

5-OH 9.06 br s a) The signal assignment of this table is based on the analysis results of DEPT, PFG ¾-¾ C0SY, PFG HMQC and PFG HMBC. The multiplicity of the carbon signal is determined using the DEPT method and is represented by s (single peak), d (doublet), t (triplet), and q (quadruplex). b) The numbers and codes in this column represent the ¾ cores in the PFG ¾-¾ COSY spectrum that give coupling-related signals in the corresponding row. c) The numbers and codes in this column represent the ¹ H cores in the PFG HMBC (™ = 8 Hz) that give the remote heteronuclear correlation (HMBC) signal with the carbon signal in the corresponding row. d) From the PFG ¾-¾ COSY spectrum, a weak but significant remote correlation signal separated by four bonds is detected between 4-H and 9-H. e) From the PFG HMBC spectrum (™ = 8 Hz), a remote heteronuclear correlation signal separated by four bonds between C-9 and 6-H was detected.

Example 2 Fermentation Production and Isolation of Compound II

1 Fermentation production

 Take an appropriate amount of Streptomyces albogriseolus A2-2002 strain, and follow the same methods and steps as described in Example 1 under the same conditions for production fermentation to obtain the white light containing the substituted tricyclic acetal lactones. The fermentation culture of Streptomyces albogriseolus A2-2002 strain was about 10 liters.

2 Preparation of crude extract of fermentation broth containing compound II

 The fermented culture of Streptomyces albogriseolus A2-2002 strain (about 10 liters) was suction-filtered, and the bacterial cells were directly washed with a suitable amount of distilled water on a suction filter funnel and filtered twice. . The fermentation filtrate (10 liters) was concentrated to 1 liter under reduced pressure, and extracted three times with equal volumes of ethyl acetate. The ethyl acetate extracts were combined and concentrated under reduced pressure to obtain a crude extract of the fermentation filtrate containing compound II (6 g). . 3 Isolation and purification of compound II

 As with the process of isolating and purifying Compound I, each step of the following experiments to isolate and purify Compound II uses a screening model of flow cytometry combined with morphological detection of thermosensitive mouse breast cancer tsFT210 cells. Cycle inhibition, apoptosis induction and cell necrosis activity are anti-cancer indicators. On the basis of detecting the activity of each component and determining the active component, the active component is selected and further separation operation is performed.

Take the crude extract of the filtrate of the fermentation culture of Streptomyces albogriseolus A2-2002 strain containing compound II (6 g), dissolve it in 5 ml of chloroform, and add 5 g of 200-300 mesh silica gel G (Qingdao Ocean Chemical Group Company) Product) Mix the sample and load it onto a glass decompression column packed with 15 g of silica gel G for thin layer chromatography produced by Qingdao Ocean Chemical Group Co. After changing the polarity of the elution solvent from petroleum ether to chloroform, the gradient was gradually increased by increasing the amount of methanol in chloroform. Each fraction was received in 20 ml. After the detection by thin layer chromatography, the combined results were 5 Components, namely A-1 (200 mg, petroleum ether Elution and chloroform eluate), A-2 (2.5 g, chloroform, chloroform-methanol 98: 2-95: 5 eluate), A-3 (580 mg, chloroform-methanol 95: 5-92: 8 Eluate), A-4 (740 mg, chloroform-methanol 92: 8-90: 10 eluate), A-5 (910 mg, chloroform-methanol 80: 20-50: 50 and methanol eluate) .

 Take the active component A-2 (2.5 g), dissolve it with an appropriate amount of methanol, load it on a Sephadex LH-20 column, elute with a single methanol as the eluent, and accept it in the order of elution, and divide it into five components B-1 (50 Mg), B-2 (2 g), B-3 (130 mg), B-4 (83 mg), B-5 (127 mg). Among them, B-2 is an active component.

 Take B-2 (2g), dissolve in 2ml of chloroform, and add 4g 200-300 mesh silica gel G (product of Qingdao Ocean Chemical Group Co., Ltd.), and fill it with 10g of thin layer produced by Qingdao Ocean Chemical Group Co., Ltd. Silica gel G was used on a glass decompression column, and chloroform-methanol solvent system was used as the eluent for decompression column chromatography. The polarity of the elution solvent was gradually increased by increasing the amount of methanol in chloroform. 10 ml, combined with thin layer chromatography, and divided into 4 components, namely C-1 (54 mg, chloroform eluate), C-2 (70 mg, chloroform eluate), C -3 (1.4 g, chloroform and chloroform-methanol 95: 5-90: 10 eluate), C-4 (192 mg, chloroform-methanol 80:20 and methanol eluate). Among them, C-3 is an active component.

 The component C-3 (1.g) was wet-loaded onto a reversed-phase silica gel (RP-18 6g) column (Φ2.2 cm X7.5 cm) using a water-methanol-chloroform solvent gradient to elute through a thin layer. Test and combine to obtain 5 components, D-1 (25 mg, water eluate), D-2 (49 mg, 5% methanol water eluate), D-3 (1.1 g, 5% -30% Methanol water eluate), D-4 (72 mg, 50% methanol and methanol eluate), D-5 (34 mg, chloroform eluate). Among them, D-3 is the active component.

 Take component D-3 (1.1 g), depressurize on a silica gel G (5 g) column (Φ2.5 cm X 7 cm) on a wet method, and elute with a gradient of petroleum ether-chloroform-methanol solvent system. Combine the relevant fractions to obtain 10 components, E-1 (27 mg, petroleum ether and petroleum ether-chloroform 80: 20-70: 30 eluate eluate), E-2 (700 mg, petroleum ether -Chloroform 70:30, chloroform and chloroform-methanol 99: 1-98: 2 eluate), E- 3 (46 mg, chloroform-methanol 97: 3 eluate), E-4 (50 mg, chloroform- Methanol 96: 4-95: 5 eluate), E-5 (46 mg, chloroform-methanol 95: 5-94: 6 eluate), E-6 (31 mg, chloroform-methanol 93: 7 eluate) ), E-7 (23 mg, chloroform-methanol 92: 8 → 90: 10 eluate), E-8 (30 mg, chloroform-methanol 90: 10 eluate), E-9 (51 mg, Chloroform-methanol 90: 10-60: 40 eluate), E-10 (12 mg, methanol eluate). Among them, E-2 is the active component.

 Take component E-2 (700 mg), wet-pressurize a silica gel G (10 g) column (Φ2.5 cm X 7 cm), and use a chloroform-methanol-water (10: 3: 3) mixed solvent with a constant gradient Elution, receiving the fractions in the elution order, and combining them after thin layer chromatography, to obtain 7 components, F-1 (300 mg), F-2 (33 mg), F-3 (24 mg) , F-4 (15 mg), F-5 (17 mg), F-6 (8 mg), F-7 (59 mg). Among them, F-1 is an active component.

Dissolve the active ingredient F-1 (300 mg) containing compound II with an appropriate amount of methanol, and soak it in methanol. On a Sephadex LH-20 column, elution chromatography with methanol was used to intercept the eluted fractions containing the target compound and concentrated under reduced pressure to obtain 190 mg of a pure product of the oily compound Π. Compound II light brown oil, [^] ³ ^ -531.7 ° (c0.5, CHC1 ₃ ), molecular formula CHANOA, T0F-MS m / z: 208 [M + H] ⁺ , 225 [M + H ₂ 0] ⁺ , 230 [M + Na] ⁺ ; Positive HR-TOF-MS m / z, found 208.0585 [M + H] ⁺ , calculated 208.0559 (C ₁₀ H ₁₀ N0 ₄ [M + H] ⁺ ); Negative HR -TOF-MS m / z: Found 207.0379 [MH] ", Calculated 207.0402 (C _1Q H ₈ N0 ₄ [MH]") UV _max nm (log s) in MeOH: 216 (3.95), terminal absorption. IR v _m cm ¹ (KBr): 3428 (NH), 3085 (olefin protons), 2987, 2930 (raethine protons), 1754 (lactone carbonyl), 1687 (amide I absorption,-C0N 〈), 1649, 1595 (amide II absorption, NH ₂ ), 1406 (OC), 1369, 1320 (two C-0), 1199, 1122, 1096, 1066 (C-0), 991 (m), 964 (s), 911 (m), 840 (m), 805 (), 771 (m), 695 (m), 638 (w), 596 (m), 554 (w), 474 (vw;). See Table 4 for ¾ and ¹³ C NMR data. 600 MHz ¾ and 150 MHz ¹³ C NMR data for compound Π in deuterated pyridine position label J in Hz) ¾-¾ C0SY ^b> 5 _C draw C ^c >

 1 163. 23 s 3

 2 144. 04 s 3, 4, 9, N-Ha

3 6.59 d (5.8) 4 107. 43 d 4

 4 3.02 m 3, 5, 8, 9 * 42. 11 d 3, 6, 7, 8, 9

5 3.42 m 4, 6 52. 29 d 4, 6, 7, 8

6 6.27 AB type 5 136. .69 d 7, 8

7 6.27 AB type 8 135., 22 d 5, 6

8 2.66 m "4, 7, 9 38., 93 d 3, 4, 5, 6, 7

9 6.06 dd (ca 1.8, 4 ^d) , 8 96.. 29 d 4

 10 168. .78 s 4, 5, 6, 9

N-Ha 8.59 br s + N-Hb

 N-Hb 8.29 br s N-Ha

谱中与相应行中的碳信号给出远程杂核相关 （HMBC) 信号的¹ H核。 d) 从 PFG ¾-¾ COSY谱中在 4-H和 9-H之间检测到虽然弱但比较显著的相隔四根键的远程 相关信号。 a) The signal assignment of this table is based on the analysis results of DEPT, PFG ¾-¾ C0SY, PFG HMQC and PFG HMBC. The multiplicity of the carbon signal is determined using the DEPT method and is represented by s (single peak), d (doublet), t (triplet), and q (quadruplex). b) The numbers and codes in this column represent the ¾ cores in the PFG ¾-¾ COSY spectrum that give coupling-related signals in the corresponding row. c) The numbers and codes in this column represent the PFG HMBC

The ¹ H nucleus of the remote heteronuclear correlation (HMBC) signal is given in the spectrum by the carbon signal in the corresponding row. d) from In the PFG ¾-¾ COSY spectrum, although weak, but significant remote correlation signals separated by four bonds are detected between 4-H and 9-H.

Example 3 Testing of cancer cell proliferation inhibition, cell cycle inhibition, apoptosis induction and cell killing activity 1 Experimental samples and experimental methods

 Formulation of the test sample solution The test samples were pure compounds I and compound I isolated and purified in Examples 1 and 2. Accurately weigh an appropriate amount of sample and use methanol to prepare a solution of the required concentration for testing activity.

 Cell lines and cell subcultures The mammalian cancer cell lines such as mouse breast cancer tsFT210 cells, human chronic myeloid leukemia K562 cells, and human colorectal cancer HCT-15 cells were used for the test. All cells were subcultured with RPMI-1640 medium containing 10% FBS at 32 ° C (tsFT210 cells) or 37 ° C (K562 cells and HCT-15 cells) in a 5% CO2 incubator. to cultivate. Cell proliferation inhibitory activity test method (SRB method)

 This method uses the SRB (sulforhodaraine B) method to test and evaluate the inhibitory activity of the tested sample on the proliferation of cancer cells. The SRB method is a new colorimetric method recently developed and used for screening and evaluation of anticancer drugs. Lissamine Rhodamine B or SRB is a bright pink aminoxanthene dye with two sulfonic acid groups in the molecule. Under slightly acidic conditions, SRB can quantitatively bind to basic amino acid residues in intracellular proteins fixed with trichloroacetic acid, thereby providing a very sensitive quantitative indicator for determining the protein content of cells. The intracellular protein content has a linear relationship with the density of living cells. Therefore, the SRB method can be used to evaluate the inhibitory activity of anticancer drugs on the proliferation of cancer cells.

For the activity test, take tsFT210, K562 or HCT-15 cells in logarithmic growth phase and prepare fresh cell suspension with RPMI-1640 medium at a density of 2 X 105 ⁵ cells per ml. 200 μl per well Inoculate in 96-well plates, add 2 microliters of different concentration of sample solution to each well, and culture for 17 hours at 32 ° C (tsFT210 cells) or 24 hours at 37 ° C (K562 cells and HCT-15 cells). Take the cells cultured under the action of the drug, first observe the morphological changes caused by drug treatment under an optical microscope, and determine whether there are morphological characteristics of cell cycle inhibition, apoptosis or cell necrosis, and then at 4 ° (, 3000 rpm / Centrifuge under the conditions of 3 minutes for 3 minutes and aspirate the supernatant. Add 50 microliters of 20% trichloroacetic acid to each cell, fix at 4 ° C for 1 hour, rinse with water 5 times and air dry. Add 0.4 to each well. 50 microliters of acetic acid solution of% SRB and left at room temperature for 30 minutes. Wash 4 'times with 1% acetic acid water to remove unbound free SRB dye. Add 150 microliters of Tris buffer (10 mmol / L) to each well. , pH 10. 5) The protein-binding dye was dissolved and the optical density (0D) of each well at 520 nm was measured using a SPECTRA MAX Plus microplate reader manufactured by MD. Each concentration of the sample was in the same 96-well plate. Set three wells, and set another three wells as blank control. Take the average 0D value of the three wells as IR% = (0D s self-load-0D-like „ _¾ ) / 0D _s ;;" X100% formula to calculate cell proliferation at each concentration T N2003 / 001039 Inhibition rate (IR%), and then the Bliss method was used to determine the half inhibitory concentration (IC ₅ ) from the inhibition rate (IR%) at various concentrations. The same test and calculation were performed three times to obtain IC ₅ . Mean and standard deviation. Flow cytometry test method for cell cycle inhibition and apoptosis-inducing activity

Take tsFT210, K562 or HCT-15 cells in logarithmic growth phase, prepare fresh cell suspension with a density of ² × 10 ⁵ cells per ml using fresh RPMI-1640 medium, and inoculate 0.5 ml per well into a 24-well plate. Add 5 μl of sample solution of different concentration to each well, and culture for 17 hours at 32 ° C (tsFT210 cells) or 24 hours at 37 ° C (K562 cells and HCT-15 cells). Take the cells cultured under the action of the drug, and first observe the morphological changes caused by the drug treatment under an optical microscope. Determine whether there are morphological characteristics of cell cycle inhibition, apoptosis or cell necrosis, and take pictures if necessary. The cells were then transferred from a 24-well plate to a 1.5 ml Eppendorf centrifuge tube, 3000 rpm at 4 ° C for 3 minutes, and the supernatant was aspirated and washed with 0.5 ml phosphate buffer solution (PBS) and shaken once, under the same conditions Collect the cells by centrifugation, add 150 μl of propidium iodide (PI) aqueous solution (containing 5 mg of PI, 100 mg of sodium citrate and 200 mg of NP-40 in 100 ml of water), stain at 30 ° C for 30 minutes, then add 150 microliters of PBS was diluted and analyzed by flow cytometry to determine the DNA content distribution in the cells. The distribution of cells in each phase of the cell cycle was analyzed and calculated using computer software WinCycle made by Kurt.

2 experimental results

 Cell proliferation inhibitory activity of compound I and compound II: tsFT210 cells were treated with different concentrations of compounds I and II for 17 hours, and K562 and HCT-15 cells were each treated for 24 hours by SRB method. The results are shown in Table 5 and Table 6. Table 5 SRB method test results of compound I cell proliferation inhibitory activity (mean ± standard deviation, n = 3) Concentration Inhibition rate%

 tsFT210 cells K562 cells HCT-15 cells

0.39 5.2 ± 6. 1 4. 6 ± 2.9 9.8 ± 11 · 4

 0.78 7.1 ± 4. 9 13. 8 ± 1.7 '6.2 ± 2. 6

 1.56 14.1 ± 6. 7 23. 1 ± 2.7 16.2 ± 2. 6

 3.13 23.8 ± 10. 6 39. 3 ± 1.5 21.5 ± 17.4

 6.25 35.0 ± 8. 5 52. 0 ± 1.6 33 · 8 ± 15 · 0

 12.50 41.8 ± 6 6 57. 1 ± 3.6 35.3 ± 18.8

 25.0 49.2 ± 1. 2 59.1 ± 4.1 38.4 ± 13.6

 50.0 57.6 ± 7. 2 59. 8 ± 0.8 47.0 ± 1.4

 100.0 63.6 ± 5. 9 60. 2 ± 1.3 59.6 ± 6.5

IC50 20.4 ± 5. 9 5. 6 ± 0.4 55.1 ± 13. 2 Table 6 SRB method test results of compound II cell proliferation inhibitory activity (mean soil standard deviation, n = 3) Concentration inhibition rate%

 (P g / ml)

 tsFT210 cells K562 cells HCT-15 cells

0.39 10.4 ± 11.6 8.3 ± 2.9 6.6 ± 3.8

 0.78 6.8 ± 5.8 14.1 ± 2.8 7.0 ± 5.3

 1.56 12.2 ± 7.8 13.4 ± 3.6 6.2 ± 2.8

 3.13-15 ± 12.8 20.1 ± 2.2 6.3 ± 3.8

 6.25-4.2 ± 9.0 23.4 ± 7.5 13.6 ± 1.8

 12.50-4.4 ± 2.7 33.7 ± 5.3 22.1 ± 4.8

 25.0 7.2 ± 7.5 49.7 ± 1.8 33.9 ± 6, 4

 50.0 13.4 ± 2.5 56.2 ± 2.3 45.1 ± 1.5

 100.0 24.8 ± 5.2 59.4 ± 0.7 47.6 ± 1.2

 IC50> 100 29.6 ± 6.0> 100 Compound I activity by flow cytometry analysis Test results: tsFT210 cells were treated with compound I at different concentrations for 17 hours, K562 and HCT-15 cells were each 24 hours later, and analyzed by flow cytometry The test results of the cell cycle inhibition and apoptosis induction activity of the compound I on cancer cells are shown in Table 7, Table 8 and Table 9. Test results of tsFT210 cells treated with Compound I for 17 hours (mean ± SD), concentration

 su ^ G0 / Gl% G0 / G1% S% G2 / M%

(P g / ml)

 Cotroll 0.6 ± 0 · 3 31.7 ± 4.2 47.5 ± 0.8 20.8 ± 3.3

0.39 0.8 ± 0.6 31.8 ± 4.9 46.4 ± 1.1 21.8 ± 4.2

0.78 0.9 ± 0.5 31.3 ± 3.2 45.9 ± 3.2 22.7 ± 6.4

1.56 1.0 ± 0.6 22.1 ± 4.4 47.5 ± 5.0 30.4 ± 1.3

3.13 1.5 ± 1.0 3.1 ± 4.1 53.4 ± 26.5 43.5 ± 30.5

6.25 2.8 ± 1.9 1 · 1 ± 0 · 5 41.7 soil 1.9 57.2 ± 2.4

12.50 4.3 ± 2.1 15.8 ± 3.1 44.5 ± 19.4 39.6 ± 16 · 4

25.0 6 · 8 ± 8.8 17.0 ± 2 · 9 46.9 ± 8.1 36.2 ± 10.0

50.0 6.5 ± 8.9 17.5 ± 1.1 46.5 ± 11.1 36.0 ± 12.1

100.0 0.9 ± 0.5 · 14.1 ± 2.9 56.3 ± 2.7 29.7 ± 5.5 Note: The data in this table are the results of flow cytometry analysis of tsFT210 cells after propidium iodide staining. Table 8 Test results of K562 cells treated with Compound I for 24 hours (mean soil standard deviation, n = 3)

 sut ^ G0 / Gl% G0 / G1% S% G2 / M%

(g / ml)

 Cotroll 2.832.9 36.9 ± 3.0 48 · 2 ± 1.8 14.9 ± 1.4

0.39 2.2 ± 1.8 36.8 ± 3.8 47.1 ± 3.2 16.1 ± 1.3

0.78 2.7 ± 2.4 35.2 ± 3.8 47.9 ± 5.7 16.9 ± 1.8

1.56 2.5 ± 1.8 24.5 ± 10.6 55.4 ± 3.8 20.1 ± 6.9

3.13 4 · 7 ± 1.4 7.43 ± 11.6 70.1 ± 1.3 22.4 ± 12.0

6.25 16.7 ± 6.9 23.7 ± 0.9 59.0 ± 2.0 17.3 ± 2.9

12.50 10.6 ± 5.0 23.9 ± 2.0 57.4 ± 2.3 18.7 ± 1.2

25.0 6.0 ± 3.4 27.4 ± 2.0 63.1 ± 2.3 9.57 ± 1.1

50.0 3.6 ± 1.6 27.2 ± 4.6 68.3 ± 5.1 4.53 ± 1.7

100.0 1 · 8 ± 1 · 2 15.3 ± 11.6 70.9 ± 10.0 13 · 8 ± 8.1 Note: The data in this table are obtained by flow cytometry analysis after staining K562 cells with propidium iodide staining.

Table 9 Test results of HCT-15 cells treated with Compound I for 24 hours (mean soil standard deviation, n = 3)

 OTb-G0 / Gl% G0 / G1% S% G2 / M%

 Og / ml)

 Cotroll 3.6 ± 1.9 38.5 ± 2.6 39.2 ± 4.0 22.3 ± 2.1

0.39 5.4 ± 4. 6 34.8 ± 4.4 39.5 ± 5.5 25.7 ± 1.1

0.78 5.8 ± 2.6 24.3 ± 6.1 43.6 ± 1.2 32.1 ± 5.7

1.56 9.8 ± 5.3 4.40 ± 3.6 32.1 ± 5.2 63.5 ± 1.6

3.13 13.9 ± 12.5 6.70 ± 10.8 41.3 ± 24.4 52.0 ± 14.0

6.25 30.2 ± 6.6 12.8 ± 5.2 56.8 ± 12.4 30.4 ± 7.3

12.50 22.6 soil 6.2 12.2 ± 4.6 64.6 ± 2.6 23.2 ± 7.1

25.0 20.3 ± 7.5 28.9 ± 2 · 7 24.6 ± 7.9 46.5 ± 5.7

50.0 14.8 ± 6.1 16.5 ± 9.8 50.5 ± 3.4 32.9 ± 7.9

100.0 35.7 ± 16.1 29.6 ± 17.5 30.7 ± 20.0 39.7 ± 6.1 Note: The data in this table are obtained by flow cytometry analysis of HCT-15 cells stained with propidium iodide.

Flow cytometry analysis of compound Π activity test results: tsFT210 cells were treated with different concentrations of compound II for 17 hours, K562 and HCT-15 cells were each 24 hours later, and compound I was measured by flow cytometry on cancer cells The results of cell cycle inhibition and apoptosis-inducing activity tests are shown in Tables 10, 11 and 12. Table 10 Test results of tsFT210 cells treated with Compound II for 17 hours (mean ± SD, n-3)

 wb-G0 / Gl% G0 / G1% S% G2 / M%

(g / ml)

 Cotroll 0.3 ± 0.2 30.1 ± 1.0 47.7 ± 1 · 6 21.9 ± 1 · 7

0.39 2.6 ± 1.6 31.3 ± 1.3 37.1 ± 0.4 21.6 ± 1.0

0.78 3.8 ± 0.9 35.3 ± 4.3 46.4 ± 1.3 18.3 ± 3.1

1.56 3.9 ± 1.2 35.6 ± 4.9 46.5 ± 2.1 18.0 ± 2.9

3.13 5.1 ± 0.8 37.9 ± 7.2 46.1 ± 2.8 16.0 ± 4.8

6.25 4.8 ± 1.6 36.0 ± 4.2 47.1 ± 1.8 16.9 ± 2.5

12.50 5.3 ± 2.4 35.8 ± 2.9 48.1 ± 0.8 16.0 ± 2.2

25.0 4.4 ± 1.8 26.8 ± 3.4 52.5 ± 1.8 20.5 ± 2.9

50.0 9.8 ± 3.2 10.0 ± 5.2 54.7 ± 1.6 35.3 ± 5.3

100.0 10.0 ± 2.9 18.9 ± 0.7 51.8 ± 0.4 29.2 ± 0.5 Note: The data in this table are obtained by flow cytometry analysis of tsFT210 cells stained with propidium iodide.

Table 11 Test results of K562 cells treated with compound Π for 24 hours (mean ± SD), concentration

 5i / Z G0 / Gl% G0 / G1% S% G2 / M% (u g / ml)

 Cotroll 2.6 ± 1.0 45.8 ± 7.4 44.2 ± 10.0 10.0 ± 2.5

0.39 2.1 ± 1.8 43.9 ± 4.3 46.0 ± 7.2 10.0 ± 2.9

0.78 2.1 ± 1.9 44.1 ± 4.7 45.5 ± 12.0 10.4 ± 3.3

1.56 2.0 ± 1.9 42.5 ± 3.5 46.5 ± 12.2 11.0 ± 4.7

3.13 2.3 ± 2.1 42.9 ± 4.4 46.2 ± 7.6 10.8 ± 3.3

6.25 1.7 ± 1.3 37.0 ± 1.0 47.6 ± 7.5 15.4 ± 7.4

12.50 2.0 ± 1.7 17.1 ± 15.4 54.2 ± 3.1 28.7 ± 17.6

25.0 5.2 ± 1, 2 2.8 ± 3.0 51.2 ± 13.7 46.1 ± 16.3

50.0 7.1 ± 1.6 11.5 ± 0.9 57.0 ± 2 · 6 32.9 ± 2.8

100.0 3.37 ± 0.7 11.3 ± 3.2 63.2 ± 17.9 25.5 ± 14.9 Note: The data in this table are the results of flow cytometry analysis after K562 cells were stained with propidium iodide.

Table 12 Test results of HCT-15 cells treated with Compound II for 24 hours (mean soil standard deviation, n = 3)

 s "b ~ G0 / Gl% G0 / G1% S% G2 / M%

(g / ml)

Cotroll 5.2 ± 2.0 43.8 ± 2.1 40.6 ± 1.5 15.5 ± 0.9 0.39 5.1 ± 1.6 40.7 ± 2 · 7 44.3 ± 1.4 14.9 ± 1.4

0.78 5.1 ± 0.7 41.0 ± 2. 6 44. 8 ± 0. 8 14. 3 ± 1.9

1.56 4.3 ± 0.3 39.1 ± 2 · 0 46. 4 ± 1. 1 14. 5 ± 1.9

3.13 4.2 ± 0.4 39. 4 ± 0. 6 46. 6 ± 1. 4 14. 1 ± 0.8

6.25 5.2 ± 0.7 34.2 ± 1. 8 45. 4 ± 1. 0 20. 4 ± 1.0

12.50 9.1 ± 3.2 16. 6 ± 0. 6 39. 5 ± 1. 4 43. 9 ± 1.9

25.0 22.4 ± 0.8 24.3 ± 9. 4 55. 3 ± 9. 5 20. 4 ± 3.2

50.0 9.8 ± 3.4 28. 7 ± 1. 0 26. 6 ± 2. 8 44. 6 ± 3.6

100.0 19.9 ± 2.8 45. 5 ± 5. 9 16. 6 ± 7. 5 38. 0 ± 5.4 Note: The data in this table are obtained by flow cytometry analysis after HCT-15 cells were stained with propidium iodide. .

Morphological test results of compound I and compound II on cancer cell killing activity:

 It was observed under an optical inverted microscope that when tsFT210 cells were treated with compound I and compound II respectively, the cell morphology gradually changed with increasing sample concentration. At low concentrations (compound I below 0.78 ig / ml, compound II below 12.5 gM, as in the blank control group, the full field is mostly small G0 / G1 cells. As the concentration of the sample increases, When I was 1.56 Pg / m and compound II was 25 g / ffll, the G2 / M phase cells in the visual field with full body and larger volume gradually increased, showing a typical G2 / M phase inhibition, and the compound I was 3.13- 6.25 ng / m Compound II peaked at 50 wg / ml. In addition, a part of snow petal-like or membrane-wrapped apoptotic cells were observed in tsFT210 cells treated with Compound II high concentration (100 ^ g / inl). .

 Similarly, when treated with different concentrations of Compound I and Compound II, the morphological changes of K562 cells were similar to the above tsFT210 cells, starting from Compound I at 1.56 g / ml and Compound II at 6.25 Pg / ml. G2 in the field of vision Cells in the / M phase gradually increased, showing typical G2 / M phase inhibition, and peaked at the G2 / M phase when Compound I was 3.13 PgA and Compound II was 25 g / fnl. At the same time, with the increase of drug concentration, from the time when Compound I was 3.13 g / ml and Compound II was 25 PgM, some apoptotic cells were observed from the visual field.

When HCT-15 cells were treated with different concentrations of Compound I and Compound II, the morphological changes were similar to those of tsFT210 and K562 cells, starting from 0.78 g / m of Compound I and 6.25 gM of Compound II. G2 / M in the field of vision Cells gradually increased, showing typical G2 / M phase inhibition, and peaked at G2 / M phase when Compound I was 1.56-3.13 g / m and Compound II was 12.5 Pg / ml. However, HCT-15 cells seem to be more sensitive to Compound I and Compound II, and more susceptible to apoptosis induced by Compound I and II. From Compound I at 1.56 gA and Compound II at 12.5 g / ml, they can be observed from the visual field. Some of the most apoptotic cells, and when the concentration of Compound I is 6.25 ng / ml and Compound II is 25 g / ml or more, Very significant apoptotic cells were observed.

 In addition, when the concentrations of Compound I and Compound II exceeded 100 g / l, both tsFT210 and K562 cells and HCT-15 cells had the morphological characteristics of typical necrotic cells with swollen body and dark opacity, indicating high concentration Compounds I and II have a direct killing effect on these cancer cells.

 The above morphological observations are in full agreement with the results of flow cytometry analysis (see Figure 10-Figure 15 and Table

7-Table 12). In short, both compound I and compound II can significantly inhibit the proliferation of tsFT210, K562, HCT-15 and other cancer cells, and the half-inhibition concentration (IC ₅ ) of compound I inhibiting the proliferation of these cancer cells is 20.4 ± 5.9 g / ml (tsFT210 Cells), 5, 6 ± 0.4 / g / ml (K562 cells), 55.1 ± 13.2 Pg / ml (HCT-15 cells); IC _{5 of} compound II. IC ₅₀ > 100 jug / ml (tsFT210 cells, IR% of inhibition rate at 100 g / ml is 24.8 ± 5.2%), 29.6 ± 6.0 ^ g / ml (K562 cells), IC ₅ . > 100 Pg / ml (HCT-15 cells, inhibition rate of 11¾% at 100 β g / ral is 47.6 ± 1.2%).

 The MIC values of Compound I on cell cycle inhibition of tsFT210, K562 and HCT-15 cells were 1.56 g / ml (tsFT210 cells), 0.78 g / l (K562 cells) and 0.39 Pg / ml (HCT-15 cells). The MIC values of apoptosis induction were 6.25 PgMl (tsFT210 cells), 3.13 g / l (K562 cells), and 0.39 g / ml (HCT-15 cells). The MIC values of Compound II on cell cycle inhibition of tsFT210, K562 and HCT-15 cells were 25 g / ml (tsFT210 cells), 6.25 / g / ml (K562 cells) and 6.25 gl (HCT-15 cells), showing the cells The MIC values of apoptosis induction were 25 g / ml (tsFT210 cells), 50 g / ml (1 (562 cells), and 6.25 g / ml (HCT-15 cells).

3 Conclusion

 Compounds I and II have anti-tumor effects such as direct killing (necrotic cell killing effect), cell cycle inhibition, and apoptosis induction on mammalian-derived cancer cells including humans.

Claims

Claim 1. A compound of formula I, wherein is amino or hydroxyl; is hydroxyl, amino or hydrogen.

 Formula I  2. The compound of formula I according to claim 1, wherein is an amino group; is a hydroxyl group or hydrogen. 3. The method for preparing a compound of formula I according to claim 1, characterized in that the tricyclic acetalactin-producing bacteria of the genus Streptomyces are fermented and cultured, a fermented product containing the tricyclic lactones compound is obtained, and The compound of formula I was isolated and purified from the fermentation. The method for preparing a compound of formula I according to claim 3, wherein the tricyclic acetalactin producing strain of the genus Streptomyces is Streptomyces albogriseohis A2-2002 CGMCC 1005. 5. Use of a compound of formula I according to claim 1 as a cell cycle inhibitor, an apoptosis inducer and a tumor cell killer. 6. The use of a compound of formula I according to claim 1 for inhibiting tumor cell proliferation. 7. Use of a compound of formula I according to claim 1 for the preparation of an antitumor drug. 8. Streptomyces white light gray „myces albogriseolus) A2-2002 CGMCC 1005. 9. Use of a strain according to claim 8 for the production of a compound of formula I according to claim 1.