Use of zidovudine in the preparation of a medicament for the prevention and / or treatment of chronic heart damage
Zidovudine improved cardiomyocyte death and calcium regulation disorders caused by the anticancer drug doxorubicin, resolved chronic cardiac damage caused by doxorubicin, restored myocardial structure and function, and provided a new treatment option.
Patent Information
- Application Number
- CN202311232600.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-09-22
- Publication Date
- 2026-02-24
- Estimated Expiration
- 2043-09-22
AI Technical Summary
In the current technology, there is a lack of effective prevention and treatment methods for chronic cardiotoxicity caused by anticancer drugs such as doxorubicin, especially the problems of damage to cardiomyocytes and calcium regulation disorders have not been effectively solved.
Using zidovudine as the drug, cell and animal experiments were conducted to verify its ability to improve cardiomyocyte death and calcium regulation disorders induced by the anticancer drug doxorubicin, restore myocardial structure, reverse the decline in cardiac function, and provide a solution for the prevention and treatment of chronic heart damage.
Zidovudine significantly improved cardiomyocyte death and calcium regulation disorders induced by the anticancer drug doxorubicin, restored myocardial structure and function, and provided a novel, effective and non-toxic treatment option.
Smart Images
Figure CN117243958B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of small molecule pharmaceutical technology, and in particular to the use of zidovudine in the preparation of drugs for the prevention and / or treatment of chronic heart injury. Background Technology
[0002] Doxorubicin, as a chemotherapy drug, has broad-spectrum antitumor effects and good clinical efficacy, and is generally an important component of many chemotherapy regimens. However, it exhibits strong dose-limiting cardiotoxicity during clinical use. Common clinical manifestations include arrhythmias, pericardial disease, and decreased left ventricular ejection fraction. With increasing dosage, it can gradually develop into heart failure. Doxorubicin-induced cardiotoxicity is a pressing issue that needs to be addressed. Currently, clinical drugs for preventing doxorubicin cardiotoxicity are very limited and have certain limitations. Some pediatric patients still experience cardiotoxicity when using dextromethorphan as a protective agent. High doses or long-term use result in strong bone marrow suppression, and its combination with doxorubicin significantly increases the risk of bone marrow suppression. Furthermore, there are concerns in clinical use about its impact on the antitumor efficacy of doxorubicin or the occurrence of secondary tumors. Therefore, developing highly effective protective drugs against doxorubicin cardiotoxicity and elucidating the protective mechanism will help improve the prevention and treatment of cardiotoxicity of anthracyclines, represented by doxorubicin, and has significant clinical application value and practical significance. Summary of the Invention
[0003] The purpose of this invention is to overcome the shortcomings of the prior art and provide the use of zidovudine in the preparation of drugs for the prevention and / or treatment of chronic heart injury.
[0004] To achieve the above objectives, the technical solution adopted by the present invention is as follows:
[0005] In a first aspect, the present invention provides the use of zidovudine in the preparation of medicaments for the prevention and / or treatment of chronic cardiac injury caused by anticancer drugs.
[0006] This invention is the first to discover, at both the cellular and animal experimental levels, that zidovudine can effectively prevent chronic cardiac damage caused by anticancer drugs. Cell experiments showed that zidovudine can improve myocardial cell death induced by the anticancer drug doxorubicin, and animal experiments showed that zidovudine can improve decreased cardiac function and myocardial structural loosening and fibrosis induced by the anticancer drug doxorubicin, indicating that zidovudine has a therapeutic effect on chronic cardiac damage caused by anticancer drugs.
[0007] In a preferred embodiment of the application described in this invention, the effective content of zidovudine in the drug for chronic cardiac injury caused by the anticancer drug is 3-10 μM. Cell experiments in this application revealed no significant difference in the effect of high-dose (10 μM) and low-dose (3 μM) zidovudine on improving cardiomyocyte death induced by doxorubicin, indicating that zidovudine at concentrations of 3-10 μM can improve cardiomyocyte death induced by doxorubicin, and that even a lower dose of zidovudine (3 μM) also shows an effect on improving cardiomyocyte death induced by doxorubicin.
[0008] In a preferred embodiment of the application described in this invention, the anticancer drug is an anthracycline anticancer drug.
[0009] In a preferred embodiment of the application described in this invention, the anthracycline anticancer drug is doxorubicin.
[0010] As a preferred embodiment of the application described in this invention, the zidovudine improves cardiomyocyte death induced by anticancer drugs.
[0011] As a preferred embodiment of the application described in this invention, the zidovudine improves the disordered calcium regulation of cardiomyocytes caused by anticancer drugs.
[0012] As a preferred embodiment of the application described in this invention, the dosage form of the drug for preventing and / or treating chronic heart damage caused by anticancer drugs is at least one of capsules, tablets, oral preparations, microcapsule preparations, injections, suppositories, sprays, ointments, gels, solutions, powders, lotions, tinctures, oils, creams, and aerosols.
[0013] As a preferred embodiment of the application described in this invention, the dosage form of the drug for preventing and / or treating chronic heart damage caused by anticancer drugs is an injection.
[0014] In a preferred embodiment of the application described in this invention, the zidovudine content in the injectable drug for the prevention and / or treatment of chronic heart damage caused by anticancer drugs is 0.3 mg / mL. During experiments, this invention found that 20 mg of zidovudine could reverse the decline in cardiac function induced by the anticancer drug doxorubicin, and that it reduced myocardial structure and fibrosis levels.
[0015] Secondly, the present invention provides the use of zidovudine in the preparation of drugs for the prevention and / or treatment of cardiotoxicity caused by doxorubicin.
[0016] Compared with the prior art, the beneficial effects of the present invention are as follows:
[0017] This invention demonstrates at the cellular and animal levels that zidovudine can reverse the decline in cardiac function induced by doxorubicin in mice by improving cardiomyocyte death and calcium regulation caused by the anticancer drug doxorubicin, restoring normal myocardial structure, and alleviating cardiotoxicity caused by doxorubicin. This provides a novel, effective, and non-toxic drug for the clinical treatment of chronic cardiac injury caused by doxorubicin. Attached Figure Description
[0018] Figure 1 The images show the Calcein / PI / Hoechst staining results for each experimental group in Experiment Example 1.
[0019] Figure 2 The results of live cell statistics for each experimental group in Experiment Example 1;
[0020] Figure 3 The images show the TUNEL staining results for each experimental group in Experiment Example 1.
[0021] Figure 4 The statistical results of apoptotic cells in each experimental group in Experiment Example 1;
[0022] Figure 5 This is a representative waveform of the intracellular calcium ion concentration change during calcium transient in cardiomyocytes of each experimental group in Experiment Example 2.
[0023] Figure 6 The statistical results of intracellular calcium ion reserves (amplitude) in cardiomyocytes of each experimental group in Experiment Example 2;
[0024] Figure 7 The statistical results of the time changes of intracellular calcium transient at different stages in each experimental group in Experiment Example 2;
[0025] Figure 8 This is the timeline of drug treatment in the doxorubicin-induced chronic heart injury mouse model in Experiment Example 3;
[0026] Figure 9 These are echocardiograms of the mice in each experimental group in Experiment Example 3.
[0027] Figure 10 This is a statistical graph showing the ejection fraction and fractional shortening of mice in each experimental group in Experiment Example 3;
[0028] Figure 11 The results of HE staining and Masson staining of the cut edges of mouse heart samples from each experimental group in Experiment Example 3;
[0029] Figure 12 The statistical results of cardiac tissue fibrosis in mice in each experimental group in Experiment Example 3;
[0030] In the above figure, "ns" indicates no statistical difference, "*" indicates p < 0.05, "**" indicates p < 0.01, "***" indicates p < 0.001, and "****" indicates p < 0.0001. Detailed Implementation
[0031] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.
[0032] Unless otherwise specified, all reagents and consumables used in the following experimental examples are commercially available.
[0033] Human cardiomyocytes were obtained by differentiating human embryonic stem cells and purifying them in sugar-free DMEM medium containing 10 mM sodium lactate. The human embryonic stem cells were obtained from the WiCell Institute in Wisconsin, USA.
[0034] C57BL / 6 mice were provided by the Experimental Animal Center of Sun Yat-sen University.
[0035] In the experimental case, doxorubicin (DOX) was used as an anticancer drug.
[0036] Experimental Example 1
[0037] To verify the effects of zidovudine (ZIDO) on human cardiomyocytes, the efficacy of zidovudine-based external therapy for DOX-induced cardiomyocyte death was evaluated using cell experiments. The specific protocol is as follows:
[0038] (1) Experimental materials and treatment
[0039] Human cardiomyocytes were selected as the experimental subject and cultured at 37°C with 5% CO2 in DMEM medium.
[0040] This experiment consisted of five experimental groups, including a control group and a treatment group, as detailed in Table 1. Cardiac cardiomyocytes were injected at a rate of 8 × 10⁻⁶. 3 After seeding cells at a density of 100 cells / well into 384-well plates coated with matrix gel and culturing for 24 hours, DOX was added to a final concentration of 1 μM. ZidO or dexrazoxane (DRZ) was then added for 2-3 days, and various indicators were measured.
[0041] Table 1 Treatment of each experimental group
[0042] Group DOX processing Drug treatment Blank control group 0 none negative control group 1μM 1μL DMSO Positive control group 1μM 10μM DRZ low-dose group 1μM 3μM ZIDO High-dose group 1μM 10μM ZIDO
[0043] (2) Observe the apoptosis of myocardial cells in each experimental group.
[0044] Cardiac cells from each experimental group were stained with Calcein / PI / Hoechst and TUNEL.
[0045] The specific procedure for Calcein / PI / Hoechst staining is as follows: Calcein-AM, propidium iodide (PI), and Hoechst were diluted with DMEM basal medium to achieve working concentrations of 2 μM for Calcein-AM, 4.5 μM for PI, and 1 μg / mL for Hoechst. After culturing cardiomyocytes for 3 days, the diluted Calcein-AM, PI, and Hoechst were added, and the cells were incubated at 37°C for 15 min. Calcein / PI / Hoechst was detected and imaged using a high-content imaging analysis system. Nine replicates were used for each treatment. The imaging results are shown in the figure below. Figure 1 The results of live cell statistics and analysis are shown in [link to relevant documentation]. Figure 2 .
[0046] The specific procedures for TUNEL staining are as follows: Cardiac cardiomyocytes from each experimental group were fixed with 4% paraformaldehyde after 2 days of culture. The TUNEL apoptosis detection kit was followed, and the cardiomyocytes were then labeled with green fluorescent protein cTnT. Imaging was performed using a high-content imaging system. Nine replicates were used for each treatment. The imaging results are shown below. Figure 3 The statistical analysis results of apoptotic cells are shown in [the table below]. Figure 4 .
[0047] In the Calcein / PI / Hoechst staining experiment, green indicates live cells, orange indicates dead cells, and blue indicates the cell nucleus. For example... Figure 1 , 2 As shown, compared with the blank control group, the number of surviving cells in the negative control group was significantly reduced, while the proportion of surviving cells in the low-dose group and the high-dose group with added ZIDO was significantly higher than that in the negative control group, showing a clear dose-escalation effect. The proportion of surviving cells in the high-dose group was comparable to that in the positive control group.
[0048] In the TUNEL staining experiment, cells appearing red are apoptotic cells, green are cardiomyocytes, and blue are cell nuclei. For example... Figure 3 , 4 As shown, compared with the blank control group, the number of apoptotic cells in the negative control group was significantly increased; the number of apoptotic cells in the low-dose group and the high-dose group with added ZIDO was significantly reduced compared with the negative control group, and showed a clear dose-reduction effect. The proportion of apoptotic cells in the high-dose group was close to that in the positive control group.
[0049] These results indicate that ZIDO can significantly improve DOX-induced cardiomyocyte death. Furthermore, since different doses of ZIDO showed little difference in their effectiveness in improving DOX-induced cardiomyocyte death, subsequent experiments all used low-dose ZIDO.
[0050] Experimental Example 2
[0051] To verify the effect of Zidovudine on calcium regulation in human cardiomyocytes, the effect of Zidovudine external treatment with DOX on cardiomyocyte calcium transients was evaluated through cell experiments. The specific protocol is as follows:
[0052] (1) Experimental materials and treatment
[0053] The experimental subjects and their culture conditions were the same as those in Experiment Example 1.
[0054] This experiment consisted of six experimental groups, including a control group and a treatment group, as detailed in Table 2. Cardiac cardiomyocytes were cultured at 8 × 10⁻⁶ cells / day. 3 Cells were seeded at a density of 100 cells / well into wells of a laser confocal glass-bottomed culture dish coated with matrix gel and cultured for 24 hours. DOX was added to bring the final concentration to 0.75 μM, and then ZidO or DRZ was added for 2 days. Various indicators were then measured.
[0055] Table 2 Treatment of each experimental group
[0056] Group DOX processing Drug treatment Blank control group 0 none negative control group 0.75μM Same volume of DMSO Positive control group 0.75μM 10μM DRZ low-dose group 0.75μM 3μM ZIDO
[0057] (2) Detection of calcium transients in myocardial cells of each experimental group
[0058] The calcium transients of cardiomyocytes in each experimental group were observed under electric field stimulation. The specific steps are as follows:
[0059] Cardiomyocytes cultured for 2 days in each experimental group were stained with 1 μM fura-2 dye at 37℃ for 20 min, transferred to working solution, and then calcium transient signal changes were recorded using a 1 Hz or 2 Hz electric field stimulation system for simultaneous cell contraction and calcium ion measurement (Ionoptix). Each treatment had 9 replicates. Results are shown below. Figure 5-7 The working solution was prepared by mixing DMEM medium containing 10 v / v% fetal bovine serum with the benchtop solution at a volume ratio of 1:1. The benchtop solution contained 140.0 mM NaCl, 5.0 mM KCl, 2 mM MgCl2, 10 mM HEPES, 1.8 mM CaCl2, 10 mM glucose, and pH 7.4.
[0060] like Figure 5-7As shown, compared with the blank control group, DOX causes calcium overload in cardiomyocytes, leading to a significant increase in calcium signal amplitude. Simultaneously, the low-dose ZIDO treatment group significantly reversed this process, while the positive control group did not. Furthermore, the time for calcium ions to be released from the endoplasmic reticulum to the cytoplasm was longer in the negative control group than in the blank control group, while data from the low-dose ZIDO treatment group showed a significantly shorter time for calcium ion release compared to the negative control group, comparable to the positive control group. These results indicate that ZIDO can significantly improve the calcium regulation capacity of cardiomyocytes induced by DOX and significantly reduce the toxicity of DOX to cardiomyocytes.
[0061] Experimental Example 3
[0062] To verify whether ZIDO has the effect of improving DOX-induced chronic cardiac damage in vivo, an animal experiment was conducted to evaluate the effect of ZIDO on DOX-induced cardiac damage. The specific protocol is as follows:
[0063] (1) Experimental materials and establishment of an animal model of DOX-induced chronic heart injury
[0064] The subjects of the experiment were male C57BL / 6 mice aged 8-10 weeks. There were 4 experimental groups, with 7 mice in each group. The experimental groups included a control group and a treatment group. The treatments for each group are shown in Table 3.
[0065] Table 3 Treatment of each experimental group
[0066] Group DOX processing ZIDO or DRZ processing Blank control group The same volume of physiological saline The same volume of physiological saline negative control group 5mg / kg The same volume of physiological saline Positive control group 5mg / kg 50mg / kg DRZ Processing group 5mg / kg 15mg / kg ZIDO
[0067] Except for the blank control group, the other experimental groups received intraperitoneal injections of DOX once a week for four weeks, while the blank control group received the same volume of saline. One hour before each DOX injection, the mice were intraperitoneally injected with Zido, DRZ, or saline according to the groupings in Table 3. The drug administration pattern diagram is shown below. Figure 8 .
[0068] (2) Assess cardiac function in mice of each experimental group
[0069] On day 35 of administration of ZiDO or DRZ, the hearts of mice in each experimental group were examined by ultrasound using a Vevo3100 ultrasound instrument, and heart samples were collected for staining.
[0070] The specific procedure for echocardiography is as follows: Mice were anesthetized with 1%-1.5% isoflurane. Echocardiography was performed using a Vevo 3100 40MHz high-resolution imaging system. The obtained M-mode echocardiographic images were used to calculate the left ventricular ejection fraction (LVEF) and left ventricular fractional shortening (LVFS). The results are shown in [Figure number missing]. Figure 9-10 .
[0071] The collected mouse heart samples were fixed with 4% PFA for 1 hour at room temperature, and then dehydrated with 10wt%-30wt% sucrose solution for 3 hours at room temperature. All samples were embedded in OCT embedding medium and sectioned to 8 μm according to standard cryosectioning procedures. HE and Masson staining were performed according to the kit instructions. The results are shown in the figure. Figure 11-12 .
[0072] like Figure 9 , 10 As shown, mice in the negative control group had significantly reduced LVEF and LVFS, while mice treated with ZIDO had the same LVEF and LVFS levels as mice in the blank control group, which was comparable to the effect of the positive control group, indicating that ZIDO can reverse DOX-induced decline in cardiac function.
[0073] like Figure 11 , 12 As shown, the myocardial structure of the negative control group was loose and the level of fibrosis was increased, while the level of myocardial fibrosis in mice treated with ZIDO was reduced and the normal and intact myocardial tissue structure was restored, indicating that ZIDO can reverse DOX-induced myocardial tissue damage.
[0074] The above results indicate that ZIDO can improve DOX-induced cardiac function and restore myocardial tissue structure to normal, thereby treating DOX-induced cardiotoxicity and can be used to prevent and treat DOX-induced chronic cardiac damage.
[0075] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.
Claims
1. The use of zidovudine in the preparation of drugs for the prevention and / or treatment of chronic cardiac injury caused by anticancer drugs; The effective content of zidovudine in the drug for chronic cardiac injury caused by anticancer drugs is 3-10 μM; The anticancer drug mentioned is doxorubicin.
2. The application as described in claim 1, characterized in that, Zidovudine improves myocardial cell death induced by anticancer drugs.
3. The application as described in claim 1, characterized in that, Zidovudine improves the disordered calcium regulation of myocardial cells caused by anticancer drugs.
4. The application as described in claim 1, characterized in that, The dosage form of the drug for preventing and / or treating chronic heart damage caused by anticancer drugs is an oral formulation.
5. The application as described in claim 1, characterized in that, The dosage form of the drug for preventing and / or treating chronic heart damage caused by anticancer drugs is an injection.
6. The application as described in claim 5, characterized in that, The injectable form of the drug for the prevention and / or treatment of chronic heart damage caused by anticancer drugs contains 0.3 mg / mL of zidovudine.