KR100318499B1 - Cis-Fumagillin, a Novel Angiogenesis Inhibitor and Anti-angiogenic Composition Containing Same - Google Patents

Abstract

The present invention provides a novel compound cis-fumagillin or its esterified product thereof, which inhibits angiogenesis, the new strain Penicillium janczewskii F2641, the strain which produces the same . The present invention relates to a method for producing cis-fumaziline from and purifying cis-fumaziline from the production culture, and an angiogenesis-inhibiting composition containing cis-fumaziline or its esterified product. Since the novel compound cis-fumarzyline of the present invention shows excellent selective inhibitory activity against methionine aminopeptidase type 2 (MetAP2) involved in neovascularization, the pharmaceutical composition comprising the same may be used for cancer, rheumatoid arthritis, chronic inflammation and diabetes mellitus. It can be usefully used for the prevention and treatment of sexual retinopathy or hemangioma.

Description

Novel Compound Cis-Fumaziline Inhibits Angiogenesis and Angiogenesis Inhibiting Composition Containing The Same {Cis-Fumagillin, a Novel Angiogenesis Inhibitor and Anti-angiogenic Composition Containing Same}

The present invention is a novel compound cis- fumaziline or esterified product thereof that inhibits neovascular formation, the new strain Penicillium janczewskii F2641, which produces the same, by culturing this strain to produce cis-fumarzyline and this culture And a neovascularization inhibiting composition containing cis-fumaziline or its esterified product.

The reason why the cancer is fatally recognized is that the solid cancer is threatened by the hidden size of the metastatic cancer rather than the primary mass of the cancer that can be diagnosed medically. Cancers that have metastasized along the blood vessels can proliferate into new organs or tissues and cause new cancers at any time, even if the primary cancer has been surgically removed. Therefore, it is considered that the treatment probability of cancer is well over 500,000, and the cancer to which surgical surgery or other chemotherapy can be applied is extremely limited depending on the type, treatment time, or the condition of the patient.

On the other hand, the growth of cancer tissue requires oxygen and nutrient supply, and angiogenesis is involved to make such a supply passage. In addition, since cancer cells are transferred to other tissues through this neovascularization, it is recognized as a new cancer therapy to control the neovascularization process essential for cancer growth and metastasis.

In 1998, a study by Dr. Folkman of the Harvard Medical School, which was developed by Entermed Corporation, showed the results of a large-scale study of animal tests of endostatin and angiostatin, which are neovascularization inhibitors. There is a bar. As a result of animal experiments in rats, endostatin and angiostatin showed antitumor growth ability, which raised expectations for the development of new anticancer drugs. These drugs are expected to be tested in humans at the end of 1999, and the potential for new anticancer therapies of neovascularization inhibitors has been proven, but difficulties in mass production methods are increasingly being revealed.

In addition, angiogenesis inhibitors have been found from low molecular weight secondary metabolites produced by microorganisms in addition to endostatin and angiostatin. Fumaziline known to date is limited to the trans form synthesized from the genus Aspergillus , the structural analogue TNP-470 (AGM-1470), which is a parent nucleus, also shows potent angiogenesis inhibitory effects and enters the clinical stage 3 In the process.

Around 1997, the protein p67 to which the inhibitors specifically bind (covalently) was found by using the TNP-470 or a synthetic angiogenesis inhibitor, ovalicin and HUVEC cells involved in angiogenesis. It has been confirmed that the protein is methionine aminopeptidase type 2 (MetAP2) (Griffith, EC et al., 'Methionine aminopeptidase (type 2) is the common target for angiogenesis inhibitors AGM-1470 and ovalicin', Chem. Biology, 4 , pp. 461-471 (1997); and Sin, N. et al., 'The anti-angiogenic agent fumagillin covalently binds and inhibits the methionine aminopeptidase, MetAP-2', Proc. Natl. Acad. , vol. 94 , pp. 6099-6103 (1997). This methionine aminopeptidase type 2 acts as an enzyme as a metalloproteinase that requires Co ⁺⁺ ions and as a phosphorylation inhibitor protein of eukaryotic initiation factor 2 (eif-2). It has two functions. Therefore, substances that bind to methionine aminopeptidase type 2 and inhibit its enzymatic function but do not affect eif phosphorylation inhibition function may be an angiogenesis inhibitor. This discovery led to the discovery of a mechanism of action to inhibit angiogenesis, which led to an active study of novel angiogenesis inhibitors having methionine aminopeptidase type-2 as a functioning point. In particular, in November 1998, methionine aminopeptidase type As the crystal structure of -2 was revealed, it was possible to design new drugs at the molecular level, thereby increasing the feasibility of cancer therapeutics having angiogenesis inhibitory function (Liu, S. et al., 'Structure of human methionine aminopeptidase'). -2 complexed with fumagillin ', Science, 282 : 1324-1327 (1998).

In addition, platelet factor-4 (PF-4) or its synthetic peptides, ursolic acid, herbimycin A, cartilage-induced neovascularization inhibitors have been discovered or synthesized. Inhibitors (cartilage-derived inhibitors, CDI) or atanomaloid compounds AP-F and the like.

As such, the discovery of a substance that inhibits neovascularization is a clue to the cancer growth mechanism, and may be applied to early diagnosis, prevention, or treatment of cancer, as well as rheumatoid arthritis and chronic diseases related to neovascularization. It can be widely used in the treatment of diseases such as inflammation, diabetic retinopathy, hemangioma, etc., and academia and industry continue to research and develop new materials having this function.

Accordingly, the present inventors have found that a new strain of the genus Penisillium binds to the above-described methionine aminopeptidase type 2 and effectively inhibits its enzyme function while hardly affecting the eif phosphorylation inhibitory function. The present invention has been found to produce cis-fumaziline which inhibits effectively.

It is an object of the present invention to provide novel compounds that inhibit angiogenesis.

Another object of the present invention is to provide a novel microorganism producing the novel compound.

Another object of the present invention is to provide a method for producing the new compound from the new microorganism.

Still another object of the present invention is to provide a method for purifying the novel compound produced from the culture medium of the novel microorganism.

It is another object of the present invention to provide angiogenesis inhibiting composition containing the novel compound.

Figure 1 shows the hydrogen nuclear magnetic resonance spectrum of the cis-fumaziline methyl ester,

Figure 2 shows the carbon nuclear magnetic resonance spectrum of the cis-fumazilin methyl ester.

In accordance with the above object, the present invention provides a novel compound cis-fumaziline or a pharmaceutically acceptable ester thereof, which exhibits angiogenesis inhibitory activity:

Formula 1

The compound cis-fumaziline is represented by the molecular formula C ₂₆ H ₃₄ O ₇ , cis-fumazyline or its esterified product has a methionine aminopeptidase-2 involved in the neovascularization to the point of action and covalently binds to its enzymatic activity Inhibits and acts as angiogenesis inhibitors.

Examples of the esterified product of cis-fumagillin include cis-fumagillin alkyl esters, cis-fumagillin C _1-4 alkylesters are preferred, and cis-fumagilin methyl esters are most preferred.

In the present invention, the angiogenesis inhibitory activity is measured from the inhibitory activity of methionine aminopeptidase-2. For example, the IC ₅₀ of the inhibitory activity of the cis-fumaziline methyl ester of methionine aminopeptidase-2 is about 3.0 ng / ml. It shows very good neovascularization inhibitory activity.

In addition, the cis-fumazyline-based compound of the present invention is preferably separated and purified from the culture medium in the form of a stable esterified product, for example, cis-fumazyline methyl ester, and then the compound is effectively used together with a pharmaceutically acceptable carrier. It can be mixed as a component to prepare an angiogenesis inhibiting composition. The composition may further include conventionally used excipients, disintegrants, sweeteners, lubricants, flavoring agents and the like, and may be tablets, capsules, powders, granules, suspensions, emulsifiers, syrups, solutions or by conventional methods or It may be formulated in unit dosage forms or multiple dosage forms of pharmaceutical preparations, such as preparations for parenteral administration.

Angiogenesis-inhibiting composition of the present invention containing the cis-fujimaline-based compound of the present invention as an active ingredient can be parenterally or orally administered as desired. The amount of 1 to 50 mg, preferably 10 to 15 mg, more preferably 12 to 13 mg per sugar can be administered in one to several doses. Dosage levels for a particular patient may vary depending on the patient's weight, age, sex, health condition, diet, time of administration, method of administration, rate of excretion, severity of the disease, and the like.

Also provided in the present invention is a new strain Penicillium janczewskii F2641 which produces cis- fumazilin. Known trans-fumazilins are produced from the genus Aspergillus, whereas cis-fumazilins of the present invention are produced from penicillium xanthoskis species and cannot be synthesized chemically from trans-fumazilins, so It is clearly distinguished. The taxonomic characteristics of the strain Penicillium xantzwiskey F2641 of the present invention cultured on MEA (malt extract agar; malt extract 20 g, peptone 1.0 g, glucose 20 g, agar 20 g and distilled water 1,000 ml) plate medium are shown in the following table. 1, Penicillium Zantz Whiskey F2641 has been deposited with Accession No. KCTC 0636BP dated June 14, 1999 to the Institute of Biotechnology Gene Bank (KCTC).

Morphological structure Microscope observation Conidiophores Formed from parasitic mycelium Penicilli Typically strongly branched Bottles Thin and smooth walls, usually 50-250 μm in length Metulae About 8-15 x 2-3 μm and have a variety of strongly branched forms Horseshoe (phialides) Complex form mainly forming 6-12 clusters, somewhat branched and having a length of 5-9 μm Conidia Spiny spherical, 3-4 μm in diameter, yellowish brown Taxonomic location Typical Divaricatum and Furcatum Genus

To cultivate penicillium xant whiskey F2641 to produce cis-fumaziline or its esterified product, it is necessary to use a peniche in a spawning medium such as YM medium (yeast extract 0.3, malt extract 0.3, dextrose 1 and peptone 0.5). After inoculating the bacteriophage of Silium Zanzi whiskey F2641 and spawning for 3 to 5 days at 24 to 26 ° C., the seed culture is produced, for example, maltose 0.5-1.2, soluble starch 1.0-2.0, cottonseed powder 0.5 Incubate in medium consisting of -1.5, corn immersion solids 0.3-1.5, NZ-amine 0.5 and allofosite 0.2-3.0 to carry out this culture for 4-6 days at 24-26 ° C. In this culture, fermentation tanks of appropriate size are selected to maintain the air supply and the stirring speed at the optimum culture conditions.

The present invention also provides a method for isolating and purifying cis-fumazyline-based compounds from the culture solution of penicillium xanthoskis F2641. The method of the present invention specifically includes the following steps. First, the culture solution of Penicillium xantzwiskey F2641 was filtered under reduced pressure with a filter and divided into cells and culture supernatant, and each was extracted with acetone and ethyl acetate. The cell extract was concentrated under reduced pressure to make an aqueous solution, which was then extracted with ethyl acetate, concentrated under reduced pressure with ethyl acetate extract of the culture supernatant, and dried. The solvent extract was separated by silica gel column chromatography using a methanol / chloroform mixture of 3:97 to 10:90 (v / v) as an eluent, and then the active substance was separated from 50:50 to 100: 0 (v / v). The methanol / distilled water mixture of v) is separated by column chromatography using eluent, for example ODS column chromatography, to obtain a partially purified active substance. Subsequently, the partially purified active material is separated by liquid chromatography (HPLC) using a 60:40 to 80:20 (v / v) methyl cyanide / distilled water mixture as an eluent to separate and purify pure cis-fumarzyline. . Preferably, in order to increase the resolution of the active substance, the carboxyl group at the terminal of the active substance partially purified with diazomethane is methylated, divided into several fractions by thin layer chromatography, and the fraction showing the best activity is finally selected and purified by HPLC. Can be. According to the method of the present invention, pure cis-fumaziline can be obtained in a yield of 0.86 mg per liter of fermentation broth of penicillium xanthosky F2641.

Hereinafter, the specific configuration and operation of the present invention will be described with reference to Examples, but the scope of the present invention is not limited only to the following Examples.

Example 1 Selection of Production Strains

10 ml of sterile liquid (6 yeast extract, 0.01MgSO ₄ -7H ₂ O and 0.02 twin 80) was added to 1 g of soil collected from Daejeon and Gangwon-do, Korea, and then ground for 30 seconds using a blender. Shake incubation for 15 minutes at. This was sequentially diluted with sterile distilled water, plated in a separation medium, and incubated at 27 ° C. for 4 weeks. In this case, Starch-Nitrate Agar medium (Kuster, E and ST Williams, 'Selection of media for isolation of Streptomyces', Nature, 202 : 928-929 (1964)) and HV Agar medium (see Hayakawa, M and H. Nonomura, 'Humic acid-vitamine agar, a new medium for the seletive isolation of soil Actinomycetes'.J. Ferment.Technol., 65 (5) : 501-509 (1987)]. The composition thereof is as follows; Starch-Nitrate Agar medium (pH 7.0)-starch 10g, KNO ₃ 2g, NaCl 2g, K ₂ HPO ₄ 2g, casamino acid 0.3g, MgSO ₄ · 7H ₂ O 0.01g, CaCO ₃ 0.02g, cyclo 50 mg heximide, 50 mg nystatin, 20 g agar and 1 liter of distilled water; And HV agar medium (pH 7.2)-1 g of humic acid, 0.05 g of Na ₂ HPO ₄ , 1.7 g of KCl, 0.05 g of MgSO ₄ · 7H ₂ O, 0.01 g of FeSO ₄ · 7H ₂ O, 0.02 g of CaCO ₃ , Thiamine 0.5mg, Riboflavin 0.5mg, Niacin 0.5mg, Pyridoxal-HCl 0.5mg, Inositol 0.5mg, Ca-pantothenate 0.5mg, P -benzoic acid 0.5mg, Biotin 0.25mg, Cyclohex 50 mg of mead, 50 mg of nystatin, 20 g of agar and 1 liter of distilled water. After incubation, a single colony was isolated to isolate a strain named F2641 from the group of microorganisms showing MetAP2 inhibitory activity, and John Eye. John I. Pitt's method (John I. Pitt, 'A Laboratory Guide to Common Penicillium Species', ISBN 0-643-03949-X (1985), Published by Commonwealth Scientific and Industrial Research Organization (CSIRO) , Division of Food Research (located in PO Box 52, North Ryde, NSW 2113, Australia). To identify isolated strains, CYA medium (1.0 g K ₂ HPO ₄ , 10 ml Czapek concentrate (NaNO ₃ 30 g, KCl 5 g, MgSO ₄ 7H ₂ O 5 g, FeSO ₄ 7H ₂ O 0.1 g and 100 ml of distilled water), 5.0 g of yeast extract, 30 g of sucrose, 15 g of agar and 1,000 ml of distilled water); MEA medium (20 g malt extract, 1.0 g peptone, 20 g glucose, 20 g agar and 1,000 ml distilled water); G25N medium (0.75 g K ₂ HPO ₄ , 7.5 ml Xzapec concentrate, 3.7 g yeast extract, 250 g glycerol, 12 g agar and 750 ml distilled water); And inoculated in CzA medium (10 ml of Xzapec concentrate, 30 g of sucrose and 15 g of agar) and observed for 7 to 14 days of incubation at 25 to 29 ° C., respectively, and the colonies exhibited the characteristics shown in Table 2 below.

Flatbed Type CYA MEA G25N CzA Diameter (mm) 23-45 18-30 12-15 20-25 Texture Genital, wrinkled, downy Smooth or fluffy surface Have down Have down Mycelium color White White White White Conidia color Greenish gray Usually tan or greenish gray Usually green or light green Pale green Exudate Transparency Sometimes transparent - Transparency Conigenation degree usually usually usually slightly Badge back Dark orange yellow to brown Dark yellowish brown Usually yellow Pale yellowish brown Czapek yeast extract agar (CYA), malt extract agar (MEA), 25glycerol nitrate agar (G25N), and Czapek agar (CzA)

In addition, the strain was observed under a microscope to confirm microbiological characteristics as shown in Table 1, and from these results, the F2641 strain was found to be the penicillium zantzwhiskey species.

The present inventors deposited Penicillium Zantz Whiskey F2641 with the Accession No. KCTC 0636BP dated June 14, 1999 to the Gene Technology Bank.

Example 2 Isolation and Purification of Angiogenesis Inhibitors Produced by Penicillium Zantz Whiskey F2641

(Step 1) Cultivation of Strains

F2641 strains grown in PDA medium (potato dextrose agar; potato blend 20, dextrose 2 and agar 1.5) were inoculated in YM medium (yeast extract 0.3, malt extract 0.3, dextrose 1 and peptone 0.5) and incubated at 25 ° C. After 4 days of spawning, the spawning medium was produced at a concentration of 3 (v / v) (maltose 0.5-1.2, water-soluble starch 1.0-2.0, cottonseed powder 0.5-1.5, corn steep solid 0.3-1.5, NZ). Inoculated in 0.5 amine and allophosite 0.2-3.0) and incubated at 25 ° C. At this time, 1.5 liters of medium in a 5 L flask was incubated for 5 days, and cultured for 5 days in a fermenter of 14 L using 8 liters of the same composition. At this time, aeration rate of 4 L per minute was maintained, and the stirring speed was maintained at a speed of 400 rpm with a flat blade impeller.

(Step 2) Isolation and Purification of Angiogenesis Inhibitors

Penicillium Zantz Whiskey obtained in (Step 1) Eight liters of the culture medium of F2641 was prepared by Whatman No. Filtration under reduced pressure with one filter, the cells and the culture supernatant were separated and extracted with 8 L of acetone and ethyl acetate. The cell extract was concentrated under reduced pressure to make an aqueous solution, which was then extracted with ethyl acetate and concentrated under reduced pressure with ethyl acetate extract of the culture supernatant and dried. The solvent extracted active material was subjected to silica gel column chromatography (column: Kieselgel 60 (manufactured by Merck, 230-400 mesh)) using a methanol / chloroform mixture having a methanol concentration of 3 to 10 as an eluent. The eluted fraction was separated as an active material. The separated active substance was separated by column chromatography (column: Lichroprep RP-18 (registered trademark, manufactured by Merck, 40-63 μm)) using a methanol / distilled water mixture having a methanol concentration of 50 to 100 as an eluent. To obtain a partially purified active material at a methanol concentration of 75 to 80. Subsequently, the terminal carboxyl group of this active material was methylated with diazomethane, followed by silica thin layer chromatography (mobile phase: 5MeOH / CHCl).₃) Divided into several fractions, and then divided into 20MeOH / CHCl.₃Was dissolved in the solution to investigate MetAP2 inhibitory activity (see the method described in Example 4 below), and the fraction showing the best activity was selected. (Ultracarb) 10 ODS (250 x 21.2 mm) was carried out to isolate the pure compound that inhibits neovascularization, and structural analysis confirmed that it is cis-fumagilin methyl ester. At this time, the yield of the finally obtained pure cis-fumarzyline methyl ester was 0.86 mg per 1 liter of culture.

Example 3: Structure analysis of angiogenesis inhibitors

In order to determine the structure of the cis-fumazyline methyl ester separated and purified in Example 2, molecular weight and nuclear magnetic resonance (NMR) analysis was performed.

(1) molecular weight analysis

As a result of high resolution mass spectrometry using a mass spectrometer (VG70-SEQ mass spectrometry), it was found that the molecular weight of the separated and purified cis-fumarzyline methyl ester was 472 and the molecular formula was C ₂₇ H ₃₆ O ₇ .

(2) nuclear magnetic resonance (NMR) analysis

5 mg of cis-fumaziline methyl ester was dissolved in CD ₃ OD, placed in a 5 mm NMR tube, and subjected to NMR analysis with a Bruker AMX 500. ¹ H-NMR was 300 MHz and ¹³ C-NMR was 75 MHz. Measured. The results are shown in FIGS. 1 and 2, from which it was found that the cis-fumazilin methyl ester has the same structure as in Chemical Formula 2.

Example 4: Angiogenesis Inhibitory Activity Assay

Cis-fumarzyline methyl ester and conventional trans-fumarzyline isolated in Example 2 were used to measure human MetAP2, yeast MetAP1 and A172 glioma cell inhibitory activity (WT Lowther et al. , Proc. Natl. Acad. Sci. Vol. 95 , pp12153-12157 (October, 1998); and SA Cohen and DP Michaud, Analytical Biochemistry, 211 , pp 279-289 (1993); and YH Chang, Ph.D., University of St. Louis, USA (Location: 1402 S. Grand Blvd., St. Louis MO 63104, USA).

First, in human MetAP2 and yeast MetAP1, enzymes (MetAP1 or MetAP2) are used as substrates of norleucine-Ala-Ala-Glu-Glu at various concentrations of fumarziline (cis-fumarizin methylester or trans-fuma). Gilin) for 2 minutes preincubation (preincubation) at 30 ℃. Subsequently, 15 µl of a mixture of pH 7.5 consisting of 0.05 µg of enzyme, 4 mM of substrate, 50 mM of Hepes, 50 mM of KCl, and ₁ mM of CoCl ₂ was incubated at 30 ° C. for 10 minutes with various concentrations of fumigillin, and then EDTA was added thereto. The reaction was stopped. After the reaction was completed, the inhibitory activity against the enzyme was measured using an AccQ-Fluor reagent kit (manufactured by Waters). The degree of removal of N-terminal norleucine was determined by the amount of remaining peptide measured using micellar electrokinetic chromatography on Hewlett Packard HP ^3D capillary electrophoresis system.

Subsequently, in A172 glycoma cells, the cell line was placed in a collagen-coated microtiter plate, incubated for 24 hours, and treated with various concentrations of fumigillin to measure the growth inhibition effect of the glycoma cells. . The experimental results are shown in Table 3 below.

Experimental compound IC ₅₀ measurements Human MetAP2 (ng / ml) Yeast MetAP1 (μg / ml) A172 glycoma cells (μg / ml) Trans-Fumaziline 4.5 50 One Cis-fumazilin methyl ester 3.0 100 0.5

From Table 3 above, cis-fumaziline methyl ester has a higher MetAP2 inhibitory effect than the trans-fumazilin, while MetAP1 inhibitory effect is significantly lower, which is superior to the specificity of the inhibitory activity, and also inhibits the growth of tumor cells (glioma cells). It can be seen that high.

Example 5: Acute toxicity test of angiogenesis inhibitors

The acute toxicity test was performed on 5 weeks old ddy mice (Korean Association of Experimental Animals) weighing about 25 g for cis-fumigillin methyl ester. First, a test substance (cis-fumarzyline methyl ester) was mixed with sterile distilled water and prepared immediately before administration. Sterile distilled water was used as a control. The test substance was forcibly administered to the test system fasted for about 3 hours by using oral administration for sonde. On the day of administration, changes in general condition, symptoms of poisoning and the presence of dead animals were observed every hour from 1 hour to 6 hours after administration and once daily from the next day of administration to 14 days. After the end of the observation period, heat dissipation under CO ₂ anesthesia was performed to observe the visual abnormality of internal organs in detail. As a result, even when the compound was administered up to 100 mg once and continuously observed for 14 days, it was confirmed that there was no great toxicity.

Formulation example

The following formulation example is an example for formulation of the anticancer agent which contains the cis-fumarzyline methyl ester which concerns on this invention as an active ingredient. The anticancer agent according to the present invention is not limited by the following formulation example.

Formulation Example 1 Preparation of Tablet for Oral Administration

Oral administration tablets were prepared according to a conventional method after the following composition was prepared using cis-fumarzyline methyl ester as an effective drug.

ingredient content

Cis-fumazillin methyl ester 100 mg

10 mg of hard silicic anhydride

Microcrystalline Cellulose 190 mg

Magnesium stearate 5 mg

Sodium starch glycolate 60 mg

Calcium monohydrogen phosphate 135 mg

Formulation Example 2: Preparation of Injection

Injectables were prepared according to a conventional method after the following composition was prepared using cis-fumarzyline methyl ester as an effective drug.

ingredient content

Cis-fumazillin methyl ester 100 mg

Mannitol 180 mg

_{Na 2 HPO 4 · 12H 2 O} 26 mg

Distilled water 2,974 mg

The cis-fumarzyline-based compound of the present invention is excellent in selective inhibitory activity against methionine aminopeptidase type 2 (MetAP2), and thus excellent in inhibitory activity on neovascularization, and thus the pharmaceutical composition comprising the same is anticancer and rheumatoid. It can be usefully used to treat diseases such as arthritis, chronic inflammation, diabetic retinopathy and hemangioma.

Claims (8)

Cis-fumaziline or a pharmaceutically acceptable ester thereof thereof Formula 1 The method of claim 1, Cis-fumaziline methyl ester of formula (II) Formula 2 Penicillium janczewskii F2641 strain (KCTC 0636BP), which produces the cis-fumaziline of claim 1. Inoculating penicillium xantzwiskey F2641 in the spawning medium and spawning for 3 to 5 days at 24 to 26 ℃, the seed culture is inoculated into the production medium and incubated for 4 to 6 days at 24 to 26 ℃ A process for producing the cis-fumaziline of claim 1 from penicillium xantzwiskey F2641. (a) extracting a culture solution of penicillium xanthosky F2641 with an organic solvent, concentrating under reduced pressure and drying to obtain a crude extract; (c) separating the crude extract by silica gel column chromatography using a methanol / chloroform mixture as eluent to obtain an active fraction: (d) separating the active fraction obtained in step (c) by column chromatography using a methanol / distilled water mixture as an eluent to obtain an active fraction; (e) esterifying the active fraction obtained in step (d) and then obtaining the active fraction by thin layer chromatography; And (f) separating the active fraction obtained in step (e) by liquid chromatography using methane cyanide / distilled water mixture as a developing solvent, A method for isolating and purifying the cis-fumazinic esterified product of claim 1 from a culture solution of penicillium xantzwiskey F2641. The method of claim 5, In step (a), the culture solution is filtered, divided into cells and the culture supernatant, and then the cells are continuously extracted with acetone and ethyl acetate and the culture supernatant is extracted with ethyl acetate. Angiogenesis-inhibiting composition containing, as an active ingredient, cis-fumaziline or a pharmaceutically acceptable ester thereof and a pharmaceutically acceptable carrier: Formula 1 The method of claim 7, wherein A composition, which is used for the prevention or treatment of cancer, rheumatoid arthritis, chronic inflammation, diabetic retinopathy or hemangioma.