Use of flavonoid compound and medicine for preventing and / or treating familial intestinal polyps

Abstract

The present invention belongs to the field of biotechnology, and specifically relates to a use of a flavonoid compound and a drug for preventing and / or treating familial intestinal polyps. The present invention first discovered through experiments that the compound shown in Formula I can correct the obstacle of β-catenin accumulation in the nucleus caused by APC gene mutation, promote the transfer of β-catenin to the cell membrane, promote the restoration of normal morphology of dysplastic and deformed intestinal villi, and reduce the number of intestinal polyps caused by APC mutation, thereby achieving the effect of preventing and treating familial intestinal polyps. This provides more options for the drug treatment of familial intestinal polyps and has good application prospects.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and specifically relates to the use of a flavonoid compound and a medicine for preventing and / or treating familial intestinal polyps. Background Art

[0002] Familial Adenomatous Polyposis (FAP) is an inherited disorder that is usually caused by mutations in the APC gene. Polyps are small bumps or masses in the intestine that are usually benign, but they can become malignant in people with the disease. In addition, people with FAP may also develop polyps in other organs, most commonly the duodenum. Polyps in the duodenum may also develop into cancer. Polyps that develop in the colon are usually monitored and removed through regular colonoscopies that continue throughout life, usually starting in adolescence. To reduce the risk of cancer, colectomy, in which the entire colon is surgically removed, is a common treatment for FAP.

[0003] β-catenin is a multifunctional protein that plays a key role in intracellular signal transduction and gene transcription regulation. It is an important structural protein that participates in the formation of intercellular connections and can interact with a variety of proteins to regulate cell growth, differentiation, migration and apoptosis. Under normal physiological conditions, the level of β-catenin in cells is strictly regulated. However, when certain abnormalities occur, such as gene mutations or abnormal signal transduction, the level of β-catenin in cells will change, leading to uncontrolled cell growth and ultimately the occurrence of tumors. Therefore, the study of β-catenin is of great significance for understanding cell biology and tumorigenesis. At present, there is no relevant literature report on how to regulate β-catenin to achieve the treatment of FAP.

[0004] Some drugs, such as nonsteroidal anti-inflammatory drugs (NSAIDs) or selective cyclooxygenase-2 inhibitors (such as Celecoxib), may be helpful in the treatment of FAP. These drugs can slow the growth of polyps, but they are not a radical treatment. Therefore, there is an urgent need to develop new FAP treatment drugs in this field.

[0005] Small molecule compound YS434, molecular formula:

[0006]

[0007] And, the small molecule compound YS392, the molecular formula is:

[0008]

[0009] According to reports (CN201810380559.X Flavonoid compounds for preventing and treating colorectal cancer, CN202210977121.6 A new use of flavonoid compounds for treating ulcerative colitis), this type of compound has the effect of preventing and treating colorectal cancer and ulcerative colitis. However, there is no literature report on the therapeutic effect of this type of small molecule compound on familial intestinal polyps. Summary of the invention

[0010] In view of the problems in the prior art, the present invention provides a use of a flavonoid compound and a medicine for preventing and / or treating familial intestinal polyps.

[0011] Use of the compound represented by formula I, or its derivative, or its pharmaceutically acceptable salt, or its crystal form, or its stereoisomer, or its optical isomer in the preparation of a drug for preventing and / or treating familial intestinal polyps, characterized in that the structural formula of the compound represented by formula I is as follows:

[0012]

[0013] Wherein, R is selected from C1-C 10 of an alkoxy or amine group.

[0014] Preferably, R is selected from a C2 alkoxy group or an amine group.

[0015] Preferably, the compound represented by formula I is:

[0016]

[0017] Preferably, the drug is used to improve the accumulation of β-catenin in the cell nucleus and promote the transfer of β-catenin to the cell membrane;

[0018] And / or, the drug is used to promote the restoration of normal morphology of abnormally developed and / or deformed intestinal villi.

[0019] Preferably, the amino acid sequence of β-catenin is as shown in SEQ ID NO.1.

[0020] Preferably, the drug is in the form of pills, tablets, solutions, inhalants or sprays.

[0021] The present invention also provides a drug for preventing and / or treating familial intestinal polyps, which is prepared by using the compound represented by formula I, or its derivative, or its pharmaceutically acceptable salt, or its crystal form, or its stereoisomer, or its optical isomer as an active ingredient, and adding pharmaceutically acceptable excipients or auxiliary ingredients;

[0022] The structural formula of the compound shown in Formula I is as follows:

[0023]

[0024] Wherein, R is selected from C1-C 10 of an alkoxy or amine group.

[0025] Preferably, R is selected from a C2 alkoxy group or an amine group.

[0026] Preferably, the compound represented by formula I is:

[0027]

[0028] Preferably, the drug is used to improve the accumulation of β-catenin in the cell nucleus and promote the transfer of β-catenin to the cell membrane;

[0029] And / or, the drug is used to promote the restoration of normal morphology of abnormally developed and / or deformed intestinal villi.

[0030] Preferably, the drug is in the form of pills, tablets, solutions, inhalants or sprays.

[0031] In the present invention, the amino acid sequence of β-catenin (SEQ ID NO.1) is as follows:

[0032] MATQADLMELDMAMEPDRKAAVSHWQQQSYLDSGIHSGATTTAPSLSGKGNPEEEDVDTSQVLYEWEQGFSQSFTQEQVADIDGQYAMTRAQRVRAAMFPETLDEGMQIPSTQFDAAHPTNVQRLAEPSQMLKHAV VNLINYQDDAELATRAIPELTKLLNDEDQVVVNKAAVMVHQLSKKEASRHAIMRSPQMVSAIVRTMQNTNDVETARCTAGTLHNLSHHREGLAIFKSGGIPALVKMLGSPVDSVLFYAITTLHNLLLHQEGAKMA VRLAGGLQKMVALLNKTNVKFLAITTDCLQILAYGNQESKLIILASGGPQALVNIMRTYTYEKLLWTTSRVLKVLSVCSSNKPAIVEAGGMQALGLHLTDPSQRLVQNCLWTLRNLSDAATKQEGMEGLLGTLVQL LGSDDINVVTCAAGILSNLTCNNYKNKMMVCQVGGIEALVRTVLRAGDREDITEPAICALRHLTSRHQEAEMAQNAVRLHYGLPVVVKLLHPPSHWPLIKATVGLIRNLALCPANHAPLREQGAIPRLVQLLVRAH QDTQRRTSMGGTQQQFVEGVRMEEIVEGCTGALHILARDVHNRIVIRGLNTIPLFVQLLYSPIENIQRVAAGVLCELAQDKEAAEAAIEAEGATAPLTELLHSRNEGVATYAAAVLFRMS EDKPQDYKKRLSVELTSSLFRTEPMAWNETADLGLDIGAQGEPLGYRQDDPSYRSFHSGGYGQDALGMDPMMEHEMGGHHPGADYPVDGLPDLGHAQDLMDGLPPGDSNQLAWFDTDL.

[0033] The present invention provides a new use for the flavonoid compound shown in formula I. Experiments have found that the compound can correct the obstacle of β-catenin accumulation in the cell nucleus caused by APC gene mutation, promote the transfer of β-catenin to the cell membrane, promote the restoration of normal morphology of dysplastic and deformed intestinal villi, and reduce the number of intestinal polyps caused by APC mutation, thereby achieving the effect of preventing and treating familial intestinal polyps. The present invention provides a new option for the development of drugs for familial intestinal polyps and has a good application prospect.

[0034] Obviously, according to the above contents of the present invention, in accordance with common technical knowledge and customary means in the art, without departing from the above basic technical ideas of the present invention, other various forms of modification, replacement or change may be made.

[0035] The above contents of the present invention are further described in detail below through specific implementation methods in the form of embodiments. However, this should not be understood as the scope of the above subject matter of the present invention being limited to the following examples. All technologies realized based on the above contents of the present invention belong to the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 The figures are the experimental results of YS434 preventing β-catenin from entering the cell nucleus, wherein A is a representative detection result of Western-blot of cytoplasmic protein and nuclear protein extracted after 48 hours of YS434 treatment; B is the statistical analysis result of the detection results of Western-blot experiment (*: P<0.05, **: P<0.001).

[0037] Figure 2 The figures are the experimental results of YS434 promoting the transfer of β-catenin to the cell membrane, wherein A is a representative detection result of Western-blot experiment in which cell membrane proteins were extracted after 48 hours of YS434 treatment; B is the statistical analysis result of the detection results of the Western-blot experiment (*: P<0.05, ***: P<0.0001).

[0038] Figure 3 The figures show the experimental results of YS434 promoting the transfer of β-catenin to the cell membrane, wherein A is a landmark experimental result of immunofluorescence staining using anti-E-cadherin and anti-β-catenin antibodies after YS434 treatment for 48 hours; B is the result of co-localization analysis of antibody-labeled E-cadherin and β-catenin (shown as the Pearson correlation coefficient of localization, **: P<0.001).

[0039] Figure 4 APC for YS434 min / + The experimental results of the decrease in the number of intestinal polyps in mice, where A is the statistical result of the number of polyp nodules in the small intestine of the control group (give corn oil) mice and the treatment group (give YS434) mice one month after administration, and B is the statistical result of the number of polyp nodules in the colorectal area of ​​the control group and the treatment group mice one month after administration (*: P<0.05).

[0040] Figure 5 APC for YS434 min / +The experimental results of the mouse intestinal villus morphology returning from abnormality to normal, where A is the HE staining result of the paraffin section of the intestinal tissue of 5-month-old wild-type mice (i.e., healthy mice without APC gene mutation); B is the HE staining result of the paraffin section of the intestinal tissue of 4-month-old APC min / + HE staining results of paraffin sections of intestinal tissue of mice fed corn oil for 1 month; C is APC at 4 months old min / + HE staining results of paraffin sections of intestinal tissue of mice administered YS434 for 1 month. DETAILED DESCRIPTION

[0041] In the following examples and experimental examples, reagents and raw materials not specifically described are all commercially available products.

[0042] Example 1 Therapeutic effect of small molecule compound YS434 on familial intestinal polyps

[0043] It should be noted that normal human intestinal epithelial cells are difficult to culture in vitro, and their performance in in vitro culture, especially in 2D culture, is very different from that of in vivo intestinal epithelial cells, and they are usually not used as a model for epithelial cell function research. Caco-2 cells are morphologically similar to normal human small intestinal epithelial cells, have the same cell polarity and tight junctions, and have similar molecular characteristics to normal intestinal epithelial cells. As a general model, they are widely used in the study of the physiological functions of intestinal epithelial cells. Human primary colon cancer cells are an ideal model for this application and related research on cell differentiation induction because of their differentiation potential.

[0044] 1. YS434 prevents β-catenin from entering the cell nucleus in cell experiments

[0045] 1. Experimental methods

[0046] Human colon cell line Caco-2 was cultured in DMEM + 10% fetal bovine serum medium. Different concentrations of YS434 were added to the medium. After treating the cells for 48 hours, the cells were collected and the nuclear and cytoplasmic proteins were extracted using the Pierce NE-PER nuclear and cytoplasmic protein extraction kit produced by Thermo-Fisher, respectively, for Western-blot detection.

[0047] 2. Experimental results

[0048] The results are as follows Figure 1 As shown in the figure, as the concentration of YS434 increased, the β-catenin content in the cell nucleus decreased, indicating that YS434 could significantly inhibit the entry of β-catenin into the cell nucleus.

[0049] 2. YS434 promotes the transfer of β-catenin to the cell membrane in cell experiments

[0050] 1. Experimental methods

[0051] Method 1: Primary human colon cancer cells were cultured in DMEM + 10% fetal bovine serum medium. Different concentrations of YS434 were added to the medium. After treating the cells for 48 hours, the cells were collected and the cell membrane proteins were extracted using the Mem-PER Plus membrane protein extraction kit from Thermo-Fisher for Western-blot detection.

[0052] Method 2: Primary human colon cancer cells were seeded in a culture dish with a matrix gel base and cultured in DMEM + 10% fetal bovine serum medium. YS434 (2 μg / ml) was added to the culture medium. After 48 hours of treatment, β-catenin and cell membrane marker E-cadherin were detected by immunofluorescence staining.

[0053] 2. Experimental results

[0054] The experimental results of method 1 are as follows Figure 2 As shown in the figure, after YS434 treatment, there was a dose-dependent increase in β-catenin on the cell membrane (the internal reference membrane protein ATPase was positive, and the nuclear protein lamin and cytoplasmic protein GAPDH were negative, indicating that the test sample was a membrane protein), indicating that YS434 treatment can specifically promote the localization of β-catenin on the cell membrane. The experimental results of method 2 are shown in the figure. Figure 3 As shown in the figure, the β-catenin staining in the cell nucleus indicated by DAPI staining decreased, indicating that YS434 treatment inhibited the entry of β-catenin into the cell nucleus. At the same time, β-catenin increased near the cell membrane indicated by E-cadherin staining, and statistical analysis confirmed that the co-localization coefficient of β-catenin and membrane protein E-cadherin increased, indicating that YS434 treatment caused β-catenin to aggregate at the cell membrane.

[0055] 3. YS434 leads to APC min / + The number of intestinal polyps in mice was significantly reduced

[0056] 1. Experimental methods

[0057] Take 4-month-old APC min / + Mice (purchased from Jicui Pharmaceuticals) were given YS434 (dissolved in corn oil) by gavage at 20 mg / kg body weight every 5 days, while the control group was only given corn oil by gavage for 1 month. After 1 month, the intestines of mice in the control group and YS434 treatment group were collected to count the number of nodules.

[0058] 2. Experimental results

[0059] The results are as follows Figure 4As shown, after YS434 treatment, the number of intestinal polyp nodules was significantly lower than that of the control group, indicating that YS434 effectively inhibited and reversed the occurrence of intestinal polyps caused by APC mutation.

[0060] 4. YS434 leads to APC min / + The morphology of mouse intestinal villi returned from abnormality to normal

[0061] 1. Experimental methods

[0062] Take 4-month-old APC min / + Mice were given YS434 (dissolved in corn oil) by gavage at 20 mg / kg body weight every 5 days, while the control group was only given corn oil by gavage for 1 month. After 1 month, the intestinal tissues of the control group and YS434-treated group were obtained, paraffin sections were prepared, and H&E staining was performed for observation.

[0063] 2. Experimental results

[0064] like Figure 5 As shown, APC min / + The villi of the small intestine of mice were swollen, twisted and deformed, and YS434 treatment restored the morphology of the villi to normal.

[0065] From the above experimental data, it can be seen that the compound provided by the present invention can correct the obstacle of β-catenin accumulation in the cell nucleus caused by APC gene mutation, promote the transfer of β-catenin to the cell membrane, promote the restoration of normal morphology of dysplastic and deformed intestinal villi, and reduce the number of intestinal polyps caused by APC mutation, thereby achieving the effect of preventing and treating familial intestinal polyps. This provides more options for the drug treatment of familial intestinal polyps and has a good application prospect.

Claims

1. Use of a compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating familial intestinal polyps, characterized in that: The structural formula of the compound is as follows: 。 2. The use according to claim 1, characterized in that: The drug is in the form of pills, tablets, solutions, inhalers or sprays.

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