CN110974829A - Application of hydroxychloroquine linolenic acid ester in improving sensitivity of 5-Fu and evaluation method - Google Patents
Application of hydroxychloroquine linolenic acid ester in improving sensitivity of 5-Fu and evaluation method Download PDFInfo
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Abstract
The invention discloses application of hydroxychloroquine linolenic acid ester in improving fluorouracil sensitivity in colorectal cancer. The invention also discloses an in vivo evaluation method for improving fluorouracil sensitivity of hydroxychloroquine linolenic acid ester in colorectal cancer. The invention also discloses an in-vitro evaluation method for improving fluorouracil sensitivity of hydroxychloroquine linolenic acid ester in colorectal cancer. The invention discovers for the first time that the combined use of 5-Fu and AHQ can improve the sensitivity of fluorouracil chemotherapy and the treatment effect, not only can improve the effectiveness and compliance of the drug for colorectal cancer patients, but also can relieve the pain of the patients.
Description
Technical Field
The invention relates to the technical field of medicines, and particularly relates to an application and evaluation method for improving 5-Fu sensitivity of hydroxychloroquine linolenic acid ester.
Background
Colorectal cancer (CRC) is one of the leading causes of cancer death, while the second most common cancer in women and the third most common cancer in men. And in recent years, the incidence of colorectal cancer is increasing in China, 19.1 ten thousand patients die in China only in 2015, and the death rate is ranked fifth in all cancers at present. Clinical treatment the treatment means for CRC is mainly surgical resection of the primary tumor; and when the disease progresses or metastasizes, treatment with a chemotherapeutic regimen. Chemotherapy has been the standard treatment for clinical CRC. According to the National Comprehensive Cancer Network (NCCN) guidelines (2018 edition), first-line treatment of advanced colorectal cancer is based on 5-fluorouracil (5-fluorouracil, 5-Fu) based chemotherapy in combination with other cytotoxic drugs such as calcium folinate, oxaliplatin, etc. Although 5-Fu is the primary drug for CRC first-line therapy and is recommended for combination therapy, it is still limited by the low sensitivity and susceptibility to chemotherapy.
Disclosure of Invention
In order to solve the problems in the prior art, the invention provides an application and an evaluation method of hydroxychloroquine linolenic acid ester for improving the sensitivity of 5-Fu, and the application and the evaluation method have the advantages that tumor cell autophagy induced by fluorouracil in the chemotherapy process is inhibited through hydroxychloroquine linolenic acid ester, so that fluorouracil resistance is overcome, treatment sensitivity is improved, the non-progressive survival time of colorectal cancer patients is prolonged, and treatment effectiveness and compliance are improved.
In order to achieve the purpose, the invention adopts the technical scheme that:
application of hydroxychloroquine linolenic acid ester in improving fluorouracil sensitivity in colorectal cancer
An in vivo evaluation method of hydroxychloroquine linolenic acid ester for increasing sensitivity to fluorouracil in colorectal cancer, comprising:
s1, culturing the human colorectal cancer HT-29 cells, and collecting the cells when the cells grow to 80% of the culture dish;
s2, centrifuging and washing the cells for 3 times by using double non-culture medium;
s3, resuspension in physiological saline to a concentration of 4X 107Cell suspension per ml;
s4, establishing an HT-29 cell transplantation ectopic tumor model;
s5, selecting a model group, an in vivo first administration group, an in vivo second administration group and an experiment group,
wherein the model group is administered solvent without therapeutic drug as a control;
wherein the first administration group in vivo refers to the group for fluorouracil administration, and fluorouracil 25mg/kg is injected into the abdominal cavity of the first group of mice once every two days;
wherein the second administration group in vivo is a group for administration of hydroxychloroquine linolenic acid ester, and hydroxychloroquine linolenic acid ester 50mg/kg is injected into abdominal cavity of the second group of mice once a day;
wherein the experimental group is characterized in that fluorouracil and hydroxychloroquine linolenic acid ester are simultaneously given, 25mg/kg of fluorouracil is injected into the abdominal cavity of a third group of mice once every two days, and 50mg/kg of hydroxychloroquine linolenic acid ester is injected into the abdominal cavity of the third group of mice once a day;
s6, weighing the four groups of mice, and recording the weight data of the mice according to the following formula: tumor Volume (TV) ═ L × W2And/2, wherein L and W represent the longest diameter and the shortest diameter of the tumor, respectively, and the tumor volume of each group of mice was calculated.
An in vitro evaluation method of hydroxychloroquine linolenic acid ester for improving the sensitivity of fluorouracil in colorectal cancer, comprising: taking fluorouracil as a first in-vitro administration group, wherein the single drug concentration of the fluorouracil is 10 mu mol/L, and adding HT-29 cells into the fluorouracil to culture and observe the fluorouracil on a microplate;
taking hydroxychloroquine linolenic acid ester as an in-vitro second administration group, wherein the single medicine concentration of the hydroxychloroquine linolenic acid ester is 10 mu mol/L, and culturing and observing on a micro-culture plate after adding HT-29 cells;
the fluorouracil and hydroxychloroquine linolenic acid ester are taken as an in-vitro experimental group, the combined concentration is (10+10) mu mol/L, HT-29 cells are added, culture and observation are carried out on a micro-culture plate, and after comparison, the effect that the addition of hydroxychloroquine linolenic acid ester can obviously improve the chemotherapy sensitivity of fluorouracil on the HT-29 cells is obtained, so that the anti-tumor activity is improved.
Preferably, when the combined concentration of fluorouracil and hydroxychloroquine linolenic acid ester is (10+10) mu mol/L, HT-29 cell apoptosis is induced to block the cell division G0/G1In addition, the sensitivity of 5-Fu chemotherapy is improved, thereby improving the anti-tumor activity.
The invention has the beneficial effects that:
the AHQ inhibits the autophagy of tumor cells induced by 5-Fu in the chemotherapy process, thereby overcoming the drug resistance of 5-Fu, improving the chemotherapy sensitivity, improving the treatment effective rate, prolonging the non-progress survival time of colorectal cancer patients, and improving the treatment effectiveness and compliance.
Drawings
FIG. 1 is a schematic representation of the change in body weight of individual mice during treatment;
FIG. 2 is a graph showing the change in tumor volume in groups of mice during dosing;
FIG. 3 is a graph showing the weights of tumor tissues in each group after the experiment;
FIG. 4 is a schematic representation of tumor tissue entities in each group after the experiment was completed;
FIG. 5 is a schematic representation of tumor tissue apoptosis and proliferation assays after treatment is complete;
FIG. 6 is a graph of data showing the effect of 5-Fu, AHQ alone and 48h in combination on HT-29 cell cycle in human colon cancer cells.
Detailed Description
Embodiments of the present invention will be described in detail below with reference to the accompanying drawings.
Example 1:
application of hydroxychloroquine linolenic acid ester in improving fluorouracil sensitivity in colorectal cancer is provided.
Example 2:
an in vivo evaluation method of hydroxychloroquine linolenic acid ester for increasing sensitivity to fluorouracil in colorectal cancer, comprising:
s1, culturing the human colorectal cancer HT-29 cells, and collecting the cells when the cells grow to 80% of the volume of the culture dish;
s2, centrifuging and washing the cells for 3 times by using double non-culture medium;
s3, resuspension in physiological saline to a concentration of 4X 107Cell suspension per ml;
s4, establishing an HT-29 cell transplantation ectopic tumor model;
s5, selecting a model group, an in vivo first administration group, an in vivo second administration group and an experiment group,
wherein the model group is administered solvent without therapeutic drug as a control;
wherein the first administration group in vivo refers to the group for fluorouracil administration, and fluorouracil 25mg/kg is injected into the abdominal cavity of the first group of mice once every two days;
wherein the second administration group in vivo is a group for administration of hydroxychloroquine linolenic acid ester, and hydroxychloroquine linolenic acid ester 50mg/kg is injected into abdominal cavity of the second group of mice once a day;
wherein the experimental group is characterized in that fluorouracil and hydroxychloroquine linolenic acid ester are simultaneously given, 25mg/kg of fluorouracil is injected into the abdominal cavity of a third group of mice once every two days, and 50mg/kg of hydroxychloroquine linolenic acid ester is injected into the abdominal cavity of the third group of mice once a day;
s6, weighing the four groups of mice, and recording the weight data of the mice according to the following formula: tumor Volume (TV) ═ L × W2And/2, wherein L and W represent the longest diameter and the shortest diameter of the tumor, respectively, and the tumor volume of each group of mice was calculated.
Example 3:
an in vitro evaluation method of hydroxychloroquine linolenic acid ester for improving fluorouracil sensitivity in colorectal cancer, comprising: taking fluorouracil as a first in-vitro administration group, wherein the single drug concentration of the fluorouracil is 10 mu mol/L, and adding HT-29 cells into the fluorouracil to culture and observe the fluorouracil on a microplate;
taking hydroxychloroquine linolenic acid ester as an in-vitro second administration group, wherein the single medicine concentration of the hydroxychloroquine linolenic acid ester is 10 mu mol/L, and culturing and observing on a micro-culture plate after adding HT-29 cells;
taking fluorouracil combined hydroxychloroquine linolenic acid ester as an in-vitro experimental group, wherein the combined concentration is (10+10) mu mol/L, adding HT-29 cells, culturing and observing on a micro-culture plate, and comparing to obtain that the anti-swelling activity of fluorouracil on HT-29 cells can be obviously improved by adding hydroxychloroquine linolenic acid ester.
When the combined concentration of fluorouracil and hydroxychloroquine linolenic acid ester is (10+10) mu mol/L, HT-29 cell apoptosis is induced to be blocked in cell division G0/G1In stage, the sensitivity of 5-Fu chemotherapy is improved.
Example 4:
in vivo evaluation method:
1. culturing sufficient HT-29 cells, collecting cells when they grow to 80% of the culture flask, centrifuging and washing the cells with double non-culture medium for 3 times, and resuspending the cells in physiological saline to 4X 107Cell suspension per ml. The cell suspension was injected subcutaneously in the right dorsal back of nude mice in an amount of 100. mu.L per mouse to establish a HT-29 cell transplantation ectopic tumor model.
The administration scheme is as follows:
every 5 mice were divided into one group, named model group, first administration group in vivo, second administration group in vivo and experimental group, respectively, wherein:
model group: intraperitoneal injection is carried out once a day without a medicine solvent (the solvent is a mixed solvent of 25% of PEG400, 5% of dimethyl sulfoxide and 70% of purified water in volume ratio);
first administration group in vivo (group 5-Fu): intraperitoneal injection of 5-Fu 25mg/kg is carried out once every two days;
in vivo second dose group (AHQ group): injecting AHQ 50mg/kg into abdominal cavity once a day;
experimental group (5-Fu + AHQ group): intraperitoneal injection of AHQ 50mg/kg is carried out once a day, and intraperitoneal injection of 5-Fu 25mg/kg is carried out once every two days.
In the course of the experiment,each group of mice was weighed every 3 days and the body weight of each group of mice was recorded. The longest and shortest diameters of the transplanted tumors were measured every 3 days with a vernier caliper, and the tumor volume of each group of mice was calculated. The tumor volume formula was calculated according to the following formula: tumor Volume (TV) ═ L × W2And/2, wherein L and W represent the longest diameter and the shortest diameter of the tumor respectively.
The experimental result shows that the average weight average of the nude mice of each group has no obvious change (P is more than 0.05) compared with the model group, which indicates that the combination of the two drugs does not obviously enhance the toxic and side effect. The body weight change data of each group of nude mice during the administration period is shown in figure 1, and the calculation formula is as follows: x ± sd, n ═ 5.
Mouse tumor volume was calculated according to the following formula: tumor Volume (TV) ═ L × W2And/2 (L, W represent the longest and shortest tumor diameters, respectively). The experimental results show that compared with the model group, the average tumor volume of the tumor-bearing mice of each administration group and the experimental group is increased slowly, which indicates that each administration group and the experimental group have the anti-tumor curative effect, and the tumor volume of the experimental group is increased most slowly. It was shown that the 5-Fu combined AHQ group had a more significant effect of inhibiting tumor volume growth than the first administration group in vivo and the second administration group in vivo, as shown in fig. 2.
After the treatment experiment was completed, the mice were sacrificed and tumor tissues were weighed and photographed. It can be seen that compared with the model group, the tumor weight of each administration group and experiment group is obviously lighter (P is less than 0.05 or P is less than 0.01), the tumor weight of the 5-Fu combined AHQ group is less than that of each single drug group (the first administration group and the second administration group in vivo), and the difference has statistical significance (P is less than 0.05), which suggests that the AHQ combined 5-Fu can increase the chemotherapy sensitivity of 5-Fu and improve the effect of inhibiting tumor growth. The tumor weight after the treatment is shown in FIG. 3 and the photograph of the tumor volume in FIG. 4.
The TUNEL method is a common method for detecting apoptosis. The Ki-67 protein is a cell proliferation nuclear antigen, is closely related to the cell division cycle, and is known as a reliable index for judging the tumor proliferation activity. And researches show that the expression of Ki-67 is positively correlated with the proliferation and metastasis degree of tumor cells in colorectal adenocarcinoma tissues. After the experiment is finished, tumor tissues of all groups of mice are taken to carry out TUNEL and Ki-67 immunohistochemical staining analysis, and the experimental result shows that the 5-Fu and AHQ combined group realizes the sensitization of 5-Fu chemotherapy and improves the tumor treatment effect by promoting the apoptosis of tumor cells and inhibiting the proliferation of the tumor cells. The experimental results are shown in FIG. 5.
In FIG. 5, a and b show the apoptosis of tumor tissues. c, d tumor tissue proliferation. Compared to the model set: p < 0.05, P < 0.01; the combination group compared to the 5-Fu group: # P < 0.05, # P < 0.01; combination group compared to AHQ group: p is less than 0.05 and P is less than 0.01; AHQ group compared to 5-Fu group: & P & lt 0.05 and & P & lt 0.01.
Example 5:
in vitro evaluation methods:
1.5-Fu single medicine group concentration is 10 mu mol/L; the concentration of the AHQ single medicine group is 10 mu mol/L; when the concentration of the 5-Fu combined AHQ group is (10+10) mu mol/L and the action time is 24 hours, 48 hours and 72 hours, the sensitivity of the 5-Fu on HT-29 cells can be obviously improved, so that the anti-tumor activity is increased. The results are shown in tables 2-1 (24 hours of action time), 2-2 (48 hours of action time) and 2-3 (72 hours of action time).
TABLE 2-15-Fu, AHQ and 5-Fu combined AHQ inhibition of HT-29 cells for 24h (%)
(
n-3) administration group compared to control group: p < 0.05, P < 0.01; the combination group compared to the 5-Fu group: # P < 0.05, # P < 0.01; combination group compared to AHQ group: p is less than 0.05 and P is less than 0.01; AHQ group compared to 5-Fu group:&P＜0.05、&&P＜0.01。
TABLE 2-25-Fu, AHQ and 5-Fu combined AHQ inhibition of HT-29 cells for 48h (%)
(
n-3) administration group compared to control group: p < 0.05, P < 0.01; the combination group compared to the 5-Fu group: # P < 0.05, # P < 0.01; combination group compared to AHQ group: p is less than 0.05 and P is less than 0.01; AHQ group compared to 5-Fu group:&P＜0.05、&&P＜0.01。
TABLE 2-35-Fu, AHQ and 5-Fu combined AHQ inhibition of HT-29 cells for 72h (%)
(
n-3) administration group compared to control group: p < 0.05, P < 0.01; the combination group compared to the 5-Fu group: # P < 0.05, # P < 0.01; combination group compared to AHQ group: p is less than 0.05 and P is less than 0.01; AHQ group compared to 5-Fu group:&P＜0.05、&&P＜0.01。
2. the 5-Fu combined AHQ group concentration is (10+10) mu mol/L, HT-29 cell apoptosis is induced, and the HT-29 cell apoptosis is blocked in cell division G0/G1Thereby increasing the sensitivity of fluorouracil in the treatment of colorectal cancer. The results of the experiment are shown in FIG. 6.
Figure 6 representative micrographs of percentage cell cycle distribution for each group analyzed by flow cytometry: (control) control group, (5-Fu)5-Fu single drug group, (AHQ) AHQ single drug group, (5-Fu + AHQ)5-Fu combined AHQ group; cell cycle profile as determined by flow cytometry. (
n-3, compared to control: p < 0.05, P < 0.01; the combination group compared to the 5-Fu group: # P < 0.05, # P < 0.01; combination group compared to AHQ group: p is less than 0.05 and P is less than 0.01; AHQ group compared to 5-Fu group:&P＜0.05、&&P＜0.01)。
in the invention, 5-Fu is short for fluorouracil, AHQ is short for hydroxychloroquine linolenic acid ester, wherein the hydroxychloroquine linolenic acid ester is a medicine disclosed in patent 201310744461.5; vehicle in the figure is the administration of a drug-free solvent, wherein the solvent composition is: the volume ratio is 25% PEG400+ 5% dimethyl sulfoxide + 70% purified water mixed solvent.
The above-mentioned embodiments only express the specific embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the present invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention.
Claims (4)
1. Application of hydroxychloroquine linolenic acid ester in improving sensitivity of fluorouracil in colorectal cancer is provided.
2. An in vivo evaluation method of hydroxychloroquine linolenate to increase sensitivity to fluorouracil in colorectal cancer comprising:
s1, culturing HT-29 cells of human colon cancer cells, and collecting the cells when the cells grow to 80% of the culture dish;
s2, centrifuging and washing the cells for 3 times by using double non-culture medium;
s3, resuspension in physiological saline to a concentration of 4X 107Cell suspension per ml;
s4, establishing an HT-29 cell transplantation ectopic tumor model;
s5, selecting a model group, an in vivo first administration group, an in vivo second administration group and an experiment group,
wherein the model group is administered solvent without therapeutic drug as a control;
wherein the first administration group in vivo refers to the group for fluorouracil treatment, and fluorouracil 25mg/kg is injected into the abdominal cavity of the first group of mice once every two days;
wherein the second administration group in vivo is a group for administration of hydroxychloroquine linolenic acid ester, and hydroxychloroquine linolenic acid ester 50mg/kg is injected into abdominal cavity of the second group of mice once a day;
wherein the experimental group is characterized in that fluorouracil and hydroxychloroquine linolenic acid ester are simultaneously given, 25mg/kg of fluorouracil is injected into the abdominal cavity of a third group of mice once every two days, and 50mg/kg of hydroxychloroquine linolenic acid ester is injected into the abdominal cavity of the third group of mice once a day;
s6, weighing the four groups of mice, and recording weight data of the mice; while according to the following formula: tumor Volume (TV) ═ L × W2And/2, wherein L and W represent the longest diameter and the shortest diameter of the tumor, respectively, and the tumor volume of each group of mice is calculated.
3. An in vitro evaluation method of hydroxychloroquine linolenate to increase the sensitivity of fluorouracil in colorectal cancer, comprising: taking fluorouracil as a first in-vitro administration group, wherein the single drug concentration of the fluorouracil is 10 mu mol/L, and adding HT-29 cells into the fluorouracil to culture and observe the fluorouracil on a microplate;
taking hydroxychloroquine linolenic acid ester as an in-vitro second administration group, wherein the single medicine concentration of the hydroxychloroquine linolenic acid ester is 10 mu mol/L, and culturing and observing on a micro-culture plate after adding HT-29 cells;
taking fluorouracil combined hydroxychloroquine linolenic acid ester as an in-vitro experimental group, wherein the combined concentration is (10+10) mu mol/L, adding HT-29 cells, culturing and observing on a micro-culture plate, and comparing to obtain that the sensitivity of fluorouracil on the HT-29 cells can be obviously improved by adding hydroxychloroquine linolenic acid ester.
4. The method for in vitro evaluation of hydroxychloroquine linolenic acid ester sensitivity of fluorouracil in colorectal cancer according to claim 3, wherein HT-29 apoptosis is induced when the combined concentration of fluorouracil and hydroxychloroquine linolenic acid ester is (10+10) μmol/L, blocking it at cell division G0/G1And (4) period.
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU1466001A (en) * | 1999-11-05 | 2001-05-14 | Phase-1 Molecular Toxicology | Methods of determining individual hypersensitivity to an agent |
| CN102068435A (en) * | 2011-01-10 | 2011-05-25 | 齐建新 | Composite fluorouracil anti-tumor medicament |
| CN102713606A (en) * | 2009-11-13 | 2012-10-03 | 无限制药股份有限公司 | Compositions, kits, and methods for identification, assessment, prevention, and therapy of cancer |
| CN103772277A (en) * | 2013-12-31 | 2014-05-07 | 四川省医学科学院（四川省人民医院） | Hydroxychloroquine linolenate and synthesis method thereof |
| CN104165976A (en) * | 2014-08-13 | 2014-11-26 | 四川省人民医院 | In vivo evaluation method for effectiveness of treating tumors through hydroxychloroquine linolenate |
| US20160193211A1 (en) * | 2013-08-02 | 2016-07-07 | Bionsil S.R.L. In Liquidazione | Combinations of a btk inhibitor and fluorouracil for treating cancers |
-
2019
- 2019-10-21 CN CN201911000917.0A patent/CN110974829A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU1466001A (en) * | 1999-11-05 | 2001-05-14 | Phase-1 Molecular Toxicology | Methods of determining individual hypersensitivity to an agent |
| CN102713606A (en) * | 2009-11-13 | 2012-10-03 | 无限制药股份有限公司 | Compositions, kits, and methods for identification, assessment, prevention, and therapy of cancer |
| CN102068435A (en) * | 2011-01-10 | 2011-05-25 | 齐建新 | Composite fluorouracil anti-tumor medicament |
| US20160193211A1 (en) * | 2013-08-02 | 2016-07-07 | Bionsil S.R.L. In Liquidazione | Combinations of a btk inhibitor and fluorouracil for treating cancers |
| CN103772277A (en) * | 2013-12-31 | 2014-05-07 | 四川省医学科学院（四川省人民医院） | Hydroxychloroquine linolenate and synthesis method thereof |
| CN104165976A (en) * | 2014-08-13 | 2014-11-26 | 四川省人民医院 | In vivo evaluation method for effectiveness of treating tumors through hydroxychloroquine linolenate |
Non-Patent Citations (9)
| Title |
|---|
| CAITLIN A SCHONEWOLF等: "Autophagy inhibition by chloroquine sensitizes HT-29 colorectal cancer cells to concurrent chemoradiation", 《WORLD JOURNAL OF GASTROINTESTINAL ONCOLOGY》 * |
| RAVI K. AMARAVADI等: "Principles and current strategies for targeting autophagy for cancer treatment", 《CLINICAL CANCER RESEARCH》 * |
| 刘阅文等: "氯喹通过诱导凋亡增强5-氟尿嘧啶的抗结肠癌效果", 《湖北医药学院学报》 * |
| 孙岩: "硫酸羟氯喹在氟尿嘧啶诱导的胃肠炎模型中的治疗作用及分子机制", 《中国优秀硕士学位论文全文数据库 医药卫生科技辑》 * |
| 戴伟钢等: "细胞自噬与肿瘤关系研究进展", 《消化肿瘤杂志(电子版)》 * |
| 肖洪涛等: "羟氯喹亚麻酸酯在大鼠体内的药代动力学和组织分布的研究", 《中国临床药理学杂志》 * |
| 苏红等: "羟基氯喹对小鼠结直肠癌细胞增殖、凋亡、迁移、超微结构及相关炎性因子表达的影响", 《华中科技大学学报（医学版）》 * |
| 蒋倩等: "自噬与肿瘤关系的研究与进展", 《现代生物医学进展》 * |
| 魏霞等: "自噬在肿瘤治疗中的研究进展", 《河南医学研究》 * |
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