Application of 10-gingerol in the preparation of drugs that enhance the antitumor activity of paclitaxel

Abstract

This invention provides the application of 10-gingerol in the preparation of drugs that enhance the antitumor activity of paclitaxel, and an antitumor composition. The inventors innovatively discovered, through in vitro and in vivo experiments, that 10-gingerol can effectively enhance the ability of paclitaxel to inhibit the proliferation of triple-negative breast cancer cells and promote paclitaxel-induced apoptosis in triple-negative breast cancer cells, thereby effectively inhibiting the growth of triple-negative breast cancer cells and reducing tumor volume; however, 10-gingerol does not increase the toxic side effects of paclitaxel, demonstrating good safety. This invention provides important support for the clinical translation of 10-gingerol as a low-toxicity and effective chemotherapy enhancer and creates greater economic value, while also providing a broader market prospect for the development of new drugs based on natural plants.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biological medicine, and particularly relates to application of 10-gingerol in preparation of a medicine for increasing antitumor activity of paclitaxel. BACKGROUND

[0002] Breast cancer is the highest incidence of cancer in the world, and is one of the main causes of cancer death in women. Triple-negative breast cancer lacks HER2, PR and ER expression, and is the most invasive subtype of breast cancer. The incidence of triple-negative breast cancer in China is as high as 126,000 per year. Triple-negative breast cancer has the clinical characteristics of high recurrence rate and high mortality. Since it lacks effective drug targets, chemotherapy is still the main treatment. Among them, taxane (paclitaxel) is a classic chemotherapeutic drug. However, these chemotherapeutic drugs have serious side effects, and large-dose chemotherapy often leads to disease recurrence and drug resistance. Paclitaxel is prone to drug resistance (drug resistance rate is as high as 50%), which leads to treatment failure. Therefore, exploring methods to reverse the drug resistance of chemotherapeutic drugs, finding chemotherapeutic "synergists", improving the efficacy of paclitaxel, and improving the prognosis of patients are of great significance for the treatment of triple-negative breast cancer. However, there is still no low-toxicity drug worldwide that can reverse drug resistance while not reducing the clinical efficacy of paclitaxel. SUMMARY

[0003] Therefore, the purpose of the present application is to provide application of 10-gingerol in preparation of a medicine for increasing antitumor activity of paclitaxel. 10-gingerol can effectively improve the sensitivity of triple-negative breast cancer cells to paclitaxel, and can be used as a chemotherapeutic "synergist" to improve the chemotherapeutic effect of paclitaxel.

[0004] To achieve the above-mentioned purpose, the present application adopts the following technical solutions.

[0005] Application of 10-gingerol in preparation of a medicine for increasing antitumor activity of paclitaxel.

[0006] In some embodiments, the tumor is breast cancer.

[0007] In some embodiments, the breast cancer is triple-negative breast cancer.

[0008] In some embodiments, the medicine can improve the ability of paclitaxel to inhibit tumor cell proliferation.

[0009] In some embodiments, the medicine can promote paclitaxel-induced tumor cell apoptosis.

[0010] The present application also provides an antitumor composition comprising 10-gingerol and paclitaxel.

[0011] In some embodiments, the tumor is breast cancer. Preferably, the tumor is triple-negative breast cancer.

[0012] The present application also provides an anti-tumor drug comprising the above anti-tumor composition and a pharmaceutically acceptable carrier.

[0013] In some embodiments, the tumor is breast cancer. Preferably, the tumor is triple negative breast cancer.

[0014] The present application provides the use of 10-gingerol in paclitaxel anti-tumor drugs. The inventors innovatively found that 10-gingerol can effectively improve the ability of paclitaxel to inhibit the proliferation of triple negative breast cancer cells and promote the apoptosis of triple negative breast cancer cells induced by paclitaxel, thereby effectively inhibiting the growth of triple negative breast cancer cells and reducing the volume of tumors in vitro and in vivo experiments. However, 10-gingerol does not increase the side effects of paclitaxel and has good safety. 10-gingerol is an extract of natural plant ginger, and there is a commercial extraction process for component extraction at present. It has high safety and good development and utilization prospect. The present application provides important support for the clinical transformation of 10-gingerol as a low-toxicity effective chemotherapy synergist and creates more economic value. It also provides a broader market prospect for the development of new drugs based on natural plants. BRIEF DESCRIPTION OF DRAWINGS

[0015] Figure 1 Figure 10 is a result graph of the effect of 10-gingerol combined with paclitaxel on the proliferation and colony formation of triple negative breast cancer cells; wherein A: cell proliferation results; B: cell colony formation results.

[0016] Figure 2 Figure 11 is a result graph of the effect of 10-gingerol combined with paclitaxel on the apoptosis of triple negative breast cancer cells; wherein A: flow cytometry detection results; B: statistical graph; C: WB detection results of apoptosis proteins.

[0017] Figure 3 Figure 12 is a result graph of the in vivo chemosensitization effect of 10-gingerol on breast cancer xenograft tumors; wherein A: tumor size; B: tumor volume and mouse weight detection results; C: tumor cell apoptosis detection results.

[0018] Figure 4 Figure 13 is a toxicity evaluation result of 10-gingerol combined with paclitaxel on mouse liver and kidney; wherein A: liver H&E staining results; B: kidney H&E staining results. DETAILED DESCRIPTION

[0019] The experimental methods not specified in the following examples of the present application are usually carried out under conventional conditions, or under conditions recommended by the manufacturer. Various common chemical reagents used in the examples are commercially available products.

[0020] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used in the description of the application herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application.

[0021] 10-Gingerol (10-G for short), molecular formula is C 21 H 34 O4, is the main stimulating component in ginger.

[0022] Taxol, English name is Taxol; alias is Tai Su, Zi Su, Te Su; chemical name is 5β, 20-epoxy-1, 2α, 4, 7β, 10β, 13α-hexahydroxy taxane-11-ene-9-ketone-4, 10-diacetate-2-benzoate-13 [(2'R, 3'S)-N-benzoyl-3-phenyl isothiocyanate]; molecular weight is 853.92; molecular formula is C 47 H 51 NO 14 .

[0023] The following will be described in conjunction with specific examples.

[0024] Example 1

[0025] The triple negative breast cancer cells were treated with 10-gingerol and taxol in combination, and the proliferation, toxicity and apoptosis of breast cancer cells were detected.

[0026] I. Cell proliferation and toxicity

[0027] Experimental grouping: Taxol group and Taxol + 10-G group. The triple negative breast cancer cell lines MDA-MB-231 and SUM-159 were treated with Taxol and Taxol + 10-G, respectively, while the non-tumor breast epithelial cell MCF-10A was used as a control. MDA-MB-231, SUM-159 and MCF-10A cells were seeded into culture plates, and after 12 h of culture, different concentrations of Taxol and Taxol + 10-G were administered to the three types of cells, respectively. The final concentrations of the drugs in the cell culture system were as follows: Taxol group: 0 nM, 25 nM, 50 nM, 100 nM, 200 nM, 400 nM; Taxol + 10-G group: Taxol 0 nM + 10-G 0 uM, Taxol 25 nM + 10-G 25 uM, Taxol 50 nM + 10-G 50 uM, Taxol 100 nM + 10-G 100 uM, Taxol 200 nM + 10-G 150 uM, Taxol 400 nM + 10-G 200 uM. Among them, Taxol 0 nM and Taxol 0 nM + 10-G 0 uM added culture medium containing no drugs (containing 10 v / v% FBS) to treat cells.

[0028] After 48 h of drug administration, the cells in each group were collected, and the cell activity was detected using the CCK8 kit, following the instructions strictly.

[0029] The median drug efficacy (chou-talalay) combined index method was used to calculate the combined index (CI) of 10-gingerol and paclitaxel, and the calculation formula was as follows:

[0030]

[0031] Among them, (d)1 and (d)2 are the dosages of 10-gingerol and paclitaxel when used in combination, respectively; (D)1 and (D)2 are the dosages of 10-gingerol and paclitaxel that produce an equivalent effect when used in combination.

[0032] The judgment criteria are: CI > 1.1, CI = 0.9-1.1 and CI < 0.9, which represent antagonism, addition and synergy, respectively.

[0033] The results are as follows: Figure 1As shown, the inhibitory effect of 10-gingerol combined with low-dose paclitaxel (Taxol + 10-G group) on cell proliferation was significantly enhanced compared with the simple use of paclitaxel (Taxol group); CI synergistic effect analysis showed that 10-gingerol significantly increased the cytotoxicity of paclitaxel on triple-negative breast cancer cells; in addition, 10-gingerol neither had a cytotoxic effect on non-tumor breast epithelial cells MCF-10A nor enhanced the cytotoxicity of paclitaxel Figure 1 A). In addition, the colony formation experiment showed that after 48h of drug intervention, the culture medium was replaced with a culture medium without drugs and continued to be cultured for 14 days, and the colony formation of MDA-MB-231 and SUM-159 cells treated with paclitaxel alone was reduced; and the number of colonies of MDA-MB-231 and SUM-159 cells treated with 10-gingerol and paclitaxel was significantly less than that of the paclitaxel single group Figure 1 B). These results showed that 10-gingerol could increase the sensitivity of MDA-MB-231 and SUM-159 cells to paclitaxel and had high safety.

[0034] II. Apoptosis

[0035] Experimental grouping: (1) MDA-MB-231 cells: control group (Control), Taxol 100nM group, Taxol 100nM + 10-G 50uM group. (2) SUM-159 cells: control group (Control), Taxol 100nM group, Taxol 100nM + 10-G 100uM group. MDA-MB-231 and SUM-159 cells were inoculated into culture plates, and the cells reached 70% confluence, and the cells were respectively given control medium (containing 10v / v% FBS), Taxol or Taxol 100nM + 10-G. The cells were collected after 24h of drug treatment, and the apoptosis of the cells was detected by flow cytometry, and the expression of apoptosis-related proteins (cleaved-PARP, Bax, Bcl-2) was detected by western blot.

[0036] The results showed that compared with paclitaxel alone, 10-gingerol and paclitaxel combined treatment significantly increased the apoptosis rate of MDA-MB-231 and SUM-159 cells Figure 2 A-B). The results of apoptosis-related protein detection showed that compared with paclitaxel alone, the cells treated with 10-gingerol and paclitaxel, the levels of cleaved-PARP and Bax proteins were significantly increased, and the expression of Bcl-2 was reduced Figure 2 C). The above results showed that 10-gingerol could synergistically promote paclitaxel-induced apoptosis.

[0037] Example 2

[0038] 10-Gingerol, in combination with paclitaxel, can inhibit the growth of triple-negative breast cancer in mice.

[0039] Fifteen 6-week-old female NOD / SCID mice were selected and divided into the following three groups: control group (Vehicle), paclitaxel group (Taxol), and paclitaxel combined with 10-gingerol group (Taxol+10-G), with 5 NOD / SCID mice in each group. Subcutaneous xenograft tumor models were constructed: (1) Triple-negative breast cancer cells MDA-MB-231 were resuspended in PBS buffer and mixed with matrix gel at a ratio of 1:1. Each mouse was subcutaneously injected with 2×10 6 (2) Resuspended cells were injected subcutaneously into mice using a microinjector with a 25G needle. (3) Treatment: Four days after subcutaneous injection, the mice were administered the drugs via gavage or intraperitoneal injection. The control group was administered PBS (0.25 mL / mouse) via gavage once a day. The paclitaxel group was administered paclitaxel solution via intraperitoneal injection every 3 days. The paclitaxel combined with 10-gingerol group was administered paclitaxel solution via gavage at 40 mg / kg once a day and paclitaxel solution via intraperitoneal injection every 3 days. After tumor implantation, the tumor volume and mouse weight were measured every 3 days. Thirty-three days after tumor implantation, the mice were euthanized and the tumors were removed. TUNEL staining and immunohistochemistry were used to detect tumor growth and apoptosis at the molecular level. HE staining was used to detect morphological changes in the liver and kidneys. The operation was strictly performed according to the instructions.

[0040] like Figure 3 As shown, compared with the control group and the paclitaxel treatment group, the paclitaxel combined with 10-gingerol treatment group showed a significant tumor growth inhibition effect, and the tumor volume was significantly reduced; after 29 days of drug treatment, the mice treated with paclitaxel alone showed a slight weight loss, but when 10-gingerol was used in combination with paclitaxel, the weight loss induced by paclitaxel was not aggravated. Figure 3 (AB). Furthermore, during the treatment period, the mice did not exhibit adverse reactions such as skin ulcers, diarrhea, or death from poisoning.

[0041] TUNEL assay and immunohistochemical staining indicated that the combination of paclitaxel and 10-gingerol significantly increased the apoptosis rate of tumor cells and reduced the expression of the growth factor Ki67. Figure 3 C).

[0042] like Figure 4 As shown, no pathological changes were observed in the liver and kidney tissues during the pathological H&E staining analysis of each group. Figure 4 AB). Furthermore, there were no significant differences in liver and kidney indices between groups ( ). Figure 4 AB).

[0043] The above results suggest that 10-gingerol has a potential auxiliary effect in paclitaxel chemotherapy of triple-negative breast cancer and is safe.

[0044] The technical features of the above-described embodiments can be combined in any manner. To make the description concise, all possible combinations of the technical features in the above embodiments have not been described, but as long as the combinations of the technical features do not exist in contradiction, they shall be considered within the scope of the present disclosure.

[0045] The above-described embodiments only express several implementation manners of the present application, and the description is relatively specific and detailed, but it shall not be understood as a limitation on the patent scope of the present application. It should be noted that, for those skilled in the art, several modifications and improvements can be made without departing from the concept of the present application, and these shall be within the protection scope of the present application. Therefore, the protection scope of the present application shall be subject to the appended claims.

Claims

1. Use of 10-gingerol for the preparation of a medicament for increasing the antitumour activity of paclitaxel, characterised in that, The tumor is triple negative breast cancer.

2. Use according to claim 1, wherein The drug can improve the ability of paclitaxel to inhibit tumor cell proliferation.

3. The use according to claim 1, wherein The drug can promote paclitaxel-induced tumor cell apoptosis.

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