CN114366739A - Application of lercanidipine in preparation of medicine for treating and/or preventing colorectal cancer - Google Patents

Abstract

The invention discloses an application of lercanidipine in preparation of a drug for treating and/or preventing colorectal cancer, and belongs to the technical field of biological medicines. The lercanidipine can effectively inhibit proliferation, migration invasion and apoptosis of colorectal cancer cells. Experiments show that lercanidipine with the effective concentration of 2-18 mu M can effectively inhibit proliferation, migration invasion and apoptosis of colorectal cancer cells. The invention provides a novel drug for treating and/or preventing colorectal cancer, and provides a novel therapeutic drug and strategy for treating and/or preventing colorectal cancer.

Description

Application of lercanidipine in the preparation of medicaments for the treatment and/or prevention of colorectal cancer

technical field

The invention belongs to the technical field of biomedicine, and particularly relates to the application of lercanidipine in the preparation of medicaments for treating and/or preventing colorectal cancer.

Background technique

Colorectal cancer is one of the most common malignant tumors. According to the 2020 Global Cancer Statistics Report, its incidence and mortality ranks third and second in the world, respectively, and the overall survival rate is very poor. Because colorectal cancer is located in the colorectum, its early development is very secretive, so patients are often in the middle and late stages when diagnosed, and because of the high metastatic and invasive nature of colorectal cancer, the treatment of advanced colorectal cancer is more difficult; At the same time, due to the resistance of colorectal cancer cells to chemotherapy drugs, the difficulty of colorectal cancer treatment is exacerbated. Drug resistance has become an important joint in overcoming colorectal cancer. Reversing drug resistance and finding new anticancer drugs have become two major problems in the treatment of colorectal cancer. It is relatively clear in nature. If such drugs with potent anti-cancer effects can be found, it will greatly reduce the cost of anti-cancer drug research and development, shorten the research and development cycle, increase the choice of clinical colorectal cancer treatment, and facilitate the rapid application of drugs and enable patients. benefit. Under this circumstance, finding more stable, effective and safe anti-colorectal cancer drugs has become a top priority.

Lercanidipine (Lercanidipine) is a lipophilic third-generation dihydropyridine-calcium channel blocker (DHP-CCB), which has good and long-lasting efficacy in the treatment of hypertension, whether used alone or in combination It is a first-line antihypertensive drug for the treatment of essential hypertension; at the same time, lercanidipine is well tolerated and has a lower incidence of side effects, especially in renal Low toxicity. The research of lercanidipine in anti-hypertension, anti-inflammatory and other aspects has been very in-depth, but so far, there is no report on the therapeutic effect of lercanidipine on colorectal cancer, and its molecular mechanism is unclear.

SUMMARY OF THE INVENTION

The primary purpose of the present invention is to overcome the shortcomings and deficiencies of the prior art, and to provide the application of lercanidipine in the preparation of medicaments for treating and/or preventing colorectal cancer.

Above-mentioned purpose of the present invention is achieved through the following technical solutions:

Application of lercanidipine in the preparation of medicaments for treating and/or preventing colorectal cancer.

The structural formula of described lercanidipine is shown in formula (I):

The effective concentration of the lercanidipine is preferably 2-18 μM.

The drug for treating and/or preventing colorectal cancer is preferably a drug that can inhibit the growth, proliferation and/or migration and invasion of colorectal cancer cells.

The colorectal cancer is preferably at least one of colorectal adenocarcinoma and colon cancer.

A medicine used to treat and/or prevent colorectal cancer, including lercanidipine.

The medicament for treating and/or preventing colorectal cancer further comprises pharmaceutically acceptable excipients.

The pharmaceutically acceptable adjuvant is preferably at least one of slow release agent, excipient, filler, binder, wetting agent, disintegrant, absorption enhancer, adsorption carrier, surfactant and lubricant. kind.

The administration form of the medicament for treating and/or preventing colorectal cancer preferably includes but is not limited to oral administration.

Compared with the prior art, the present invention has the following advantages and effects:

The lercanidipine of the present invention can effectively inhibit the proliferation, migration, invasion and apoptosis of colorectal cancer cells. The experiment found that lercanidipine with an effective concentration of 2-18 μM could effectively inhibit the proliferation, migration, invasion and apoptosis of colorectal cancer cells. The present invention provides a new drug for treating and/or preventing colorectal cancer, and provides new therapeutic drugs and strategies for treating and/or preventing colorectal cancer.

Description of drawings

Figure 1 is a structural diagram of lercanidipine.

Figure 2 is a graph showing the effect of lercanidipine treatment time and treatment concentration on the proliferation ability of DLD-1 cells, HT-29 cells and HCT-116 cells.

Figure 3 is a graph showing the effect of different concentrations of lercanidipine on the migration ability of DLD-1 cells.

Figure 4 is a graph showing the effect of different concentrations of lercanidipine on the migration ability of HT-29 cells.

Figure 5 is a graph showing the effect of different concentrations of lercanidipine on the migration ability of HCT-116 cells.

Figure 6 is a graph showing the crystal violet staining results of the effect of different concentrations of lercanidipine on the migration ability of DLD-1 cells, HT-29 cells and HCT-116 cells.

Figure 7 is a graph showing the crystal violet staining results of the effect of different concentrations of lercanidipine on the invasive ability of DLD-1 cells, HT-29 cells and HCT-116 cells.

Figure 8 is a flow cytometry result of the apoptosis of DLD-1 cells, HT-29 cells and HCT-116 cells by different concentrations of lercanidipine.

Detailed ways

The present invention will be described in further detail below with reference to the embodiments and the accompanying drawings, but the embodiments of the present invention are not limited thereto.

Example 1

(1) Experimental materials

Human colorectal adenocarcinoma epithelial cells (DLD-1), human colon cancer cells (HT-29) and human colon cancer cells (HCT-116) were purchased from ATCC; Lercanidipine was purchased from Selleck Chemicals (Huston, TX, USA), the structural formula is shown in Figure 1; CCK-8 kit was purchased from Japan Tongjin Institute of Chemistry; 1640 basal medium, fetal bovine serum, penicillin-streptomycin, PBS buffer (pH7.4) were purchased from Gibco, USA; DMSO was purchased from sigma, USA, Lercanidipine was dissolved in DMSO, and stored at a storage concentration of 10 mM in -80 °C refrigerator; Annexin V-FITC/PI fluorescence double staining apoptosis detection kit was purchased from KGI Biotech, Nanjing, China Technology Development Co., Ltd.; Transwell chambers and Matrigel were purchased from Corning, United States.

(2) CCK-8 experiment

DLD-1 cells, HT-29 cells and HCT-116 cells were seeded in 96-well plates at 50 μL/1000 cells per well. After the cells adhered, 10 mM lercanidipine was diluted with 1640 complete medium to contain Lercanidipine medium; wherein, the concentrations of lercanidipine in the medium containing lercanidipine are 4 μM, 8 μM, 12 μM, 16 μM, 20 μM, 24 μM, 28 μM, 32 μM and 36 μM, respectively. 50 μL of the above medium containing lercanidipine, while the medium without lercanidipine was used as a control, that is, the concentrations of lercanidipine were 0, 2 μM, 4 μM, 6 μM, 8 μM, 10 μM, 12 μM, 14 μM, 16 μM and 18 μM. cell treatment group. On day 0, day 1, day 2, day 3, day 4, and day 5, add 10 μL of CCK-8 reagent to each well according to the instructions of the CCK-8 kit, and store at 37°C, 5% CO. ₂ Incubate for 2 hours in an incubator and detect with an enzyme-linked immunosorbent assay at a wavelength of 450 nm.

The results are shown in Figure 2. With the increase of lercanidipine concentration and treatment time, the killing effect (inhibition of cell proliferation) of lercanidipine on colorectal cancer cells (DLD-1, HT-29, HCT-116) was significantly increased (lecanidipine). Compared with the control group, the candipine-treated group had fewer viable cells and significantly decreased OD450 readings), indicating that the killing effect of lercanidipine on DLD-1 cells, HT-29 cells and HCT-116 cells was dose- and time-dependent ( The longer the time, the higher the concentration of lercanidipine, and the stronger the killing effect on colorectal cancer cells).

(3) Cell scratch test

Take well-growing cells (DLD-1 cells, HT-29 cells and HCT-116 cells), digest and count to adjust the cell density, and inoculate appropriate amount of DLD-1 cells, HT-29 cells and HCT-116 cells in 6-well plates respectively. (2 mL complete medium per well), 3 duplicate wells were set in each group, and control wells were set, and cultured overnight in a 37° C., 5% CO ₂ constant temperature incubator. Use a 10 μL pipette tip to streak the holes to create gaps between cells, aspirate the complete medium, wash twice with PBS buffer (pH 7.4), and try to wash the gaps and detached cells as much as possible, and add 2 mL containing 2% FBS and 1640 complete medium containing corresponding concentrations of lercanidipine (0 μM, 8 μM, 16 μM). Mark the vertical line on the back of the 6-well plate to confirm that the position of each photo is consistent. Every 24 hours (ie, day 0, day 1, and day 2), the cells at the line in each group are recorded by microscopy. The growth and gap size were determined according to the gap distance to determine the inhibitory effect of different concentrations of lercanidipine on the migration of colorectal cancer cells.

(4) Cell migration and invasion assay

Appropriate numbers of DLD-1 cells, HT-29 cells, and HCT-116 cells were resuspended in serum-free 1640 basal medium containing various concentrations of lercanidipine (0 μM, 8 μM, 16 μM) and added to the corresponding Transwell upper chambers , add 1640 complete medium containing 20% FBS to the lower chamber. Then, the culture plate was incubated in a 37°C incubator for 24 hours and detected by crystal violet staining. For invasion experiments, Matrigel (Corning, USA) was diluted 1:25 with FBS-free 1640 basal medium and spread evenly on the surface of the upper ventricular membrane of the Transwell chamber before seeding cells. After the Matrigel solidified, cells were seeded as follows. The steps are the same as in the cell migration experiments.

(5) Apoptosis experiment

Cells (DLD-1 cells, HT-29 cells and HCT-116 cells) were cultured in lercanidipine 1640 complete medium with concentrations of 0μM, 8μM, and 16μM for 48 hours, and apoptosis was double-stained with Annexin V-FITC/PI fluorescence. According to the instructions of the detection kit, gently collect the cells with trypsin without EDTA, and resuspend the cells with an appropriate amount of binding buffer to obtain a cell suspension. In the dark, add 5 μL each of Annexin V-FITC and propidium iodide (PI) to the cell suspension, mix well, and then react at room temperature for 5-15 minutes to ensure sufficient dye binding. Within 1 hour, the number of apoptotic cells was determined by flow cytometry and the FACSCalibur system (BD Biosciences).

The result is shown in Figure 3-5. As can be seen from Figure 3-5, the scratch width of DLD-1 cells, HT-29 cells and HCT-116 cells treated with 8 μM and 16 μM lercanidipine was significantly wider than that of cells treated with 0 μM lercanidipine , indicating that lercanidipine can significantly attenuate the migration ability of cancer cells (DLD-1 cells, HT-29 cells and HCT-116 cells). It can be seen from Figure 6 that in the cell migration experiment, as the treatment concentration of lercanidipine increased, the number of DLD-1 cells, HT-29 cells and HCT-116 cells migrated from the upper surface to the lower surface of the Transwell chamber membrane Significantly reduced, indicating that lercanidipine can significantly attenuate the migration ability of cancer cells (DLD-1 cells, HT-29 cells and HCT-116 cells); it can be seen from Figure 7 that in the cell invasion experiment, with the increase of lercanidipine The number of DLD-1 cells, HT-29 cells and HCT-116 cells invading from the upper surface to the lower surface of the transwell membrane decreased significantly with the increase of the treatment concentration, indicating that lercanidipine can significantly attenuate the invasive ability of colorectal cancer cells; It can be seen from Figure 8 that in the apoptosis experiment, with the increase of the treatment concentration of lercanidipine, DLD-1 cells, HT-29 cells and HCT-116 cells undergo a significant increase in apoptosis after drug treatment. The proportion of apoptotic cells in the total counted cells increased, indicating that lercanidipine could significantly induce apoptosis in cancer cells (DLD-1 cells, HT-29 cells and HCT-116 cells).

The above-mentioned embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited by the above-mentioned embodiments, and any other changes, modifications, substitutions, combinations, The simplification should be equivalent replacement manners, which are all included in the protection scope of the present invention.

Claims (9)

1. Application of lercanidipine in preparation of medicine for treating and/or preventing colorectal cancer.

2. Use according to claim 1, wherein lercanidipine has the formula (i):

3. the use according to claim 1, wherein the effective concentration of lercanidipine is 2 to 18 μ M.

4. The use according to claim 1, wherein the medicament for the treatment and/or prevention of colorectal cancer is a medicament capable of inhibiting the growth, proliferation and/or migratory invasion of colorectal cancer cells.

5. The use of claim 1, wherein the colorectal cancer is at least one of colorectal adenocarcinoma and colon carcinoma.

6. A medicament for the treatment and/or prevention of colorectal cancer, comprising lercanidipine.

7. The medicament for treating and/or preventing colorectal cancer according to claim 6, wherein the medicament for treating and/or preventing colorectal cancer further comprises pharmaceutically acceptable auxiliary materials.

8. The drug for treating and/or preventing colorectal cancer according to claim 7, wherein the pharmaceutically acceptable excipient is at least one of a sustained release agent, an excipient, a filler, a binder, a wetting agent, a disintegrant, an absorption enhancer, an adsorption carrier, a surfactant, and a lubricant.

9. The medicament for the treatment and/or prevention of colorectal cancer according to claim 6,

the administration form of the medicament for treating and/or preventing colorectal cancer includes, but is not limited to, oral administration.

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