Use of lumacaftor to improve paclitaxel resistance in triple negative breast cancer

Abstract

The application discloses application of lumacaftor in improving paclitaxel resistance of triple-negative breast cancer, and belongs to the technical field of biological medicine. The application provides application of a drug composition in preparation of a drug for preventing and / or treating paclitaxel-resistant triple-negative breast cancer; and an active ingredient of the drug composition is lumacaftor and paclitaxel. It is found for the first time that lumacaftor can significantly enhance the killing effect of paclitaxel on TNBC drug-resistant cells, and the mechanism may be related to inhibition of glycolysis pathway, recovery of cell cycle arrest and promotion of apoptosis signal. Experimental results prove that, as a GTM1 agonist, lumacaftor can inhibit the AMPK-glycolysis pathway and reduce lactic acid generation; in combination with paclitaxel in paclitaxel-resistant cells, cell proliferation is significantly inhibited, but the effect is not significant in parent cells. Therefore, lumacaftor can be used as a chemosensitizer for paclitaxel-resistant triple-negative breast cancer, and has a clear mechanism and good application prospect.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to the application of rumacato in improving paclitaxel resistance in triple-negative breast cancer. Background Technology

[0002] Triple-negative breast cancer (TNBC) is a subtype of malignant breast cancer that lacks expression of estrogen receptor (ER), progesterone receptor (PR), and HER2. It accounts for approximately 15-20% of all breast cancers and is characterized by its high invasiveness, rapid metastasis, and poor prognosis. Due to the lack of a clear molecular target, TNBC primarily relies on chemotherapy with cytotoxic drugs such as paclitaxel. However, with prolonged treatment, approximately 40-60% of TNBC patients gradually develop paclitaxel resistance, resulting in decreased treatment efficacy, continued tumor progression, or recurrence, severely impacting patient survival and quality of life.

[0003] The primary mechanism of action of paclitaxel is to stabilize microtubules, block mitosis, and induce apoptosis. However, the development of drug resistance is often accompanied by multiple mechanisms, including: upregulation of drug efflux proteins such as P-gp leading to accelerated drug clearance; abnormal expression of microtubule-associated proteins reducing drug binding capacity; blockage of apoptosis signaling pathways; and tumor metabolic reprogramming, particularly enhanced glycolysis, which provides energy support to drug-resistant cells and maintains their survival advantage. Of particular note is the abnormal activation of the AMPK signaling pathway, a core regulator of cellular metabolism, in paclitaxel-resistant cells. Its downstream glycolytic enzyme ENO1 is also significantly upregulated, forming an "AMPK-ENO1 axis," further promoting tumor glucose metabolism and enhancing drug resistance.

[0004] In existing technologies, various strategies have been attempted to reverse paclitaxel resistance, including the combined use of AMPK inhibitors, targeting the P-gp efflux pathway, or direct intervention in glycolytic enzymes. However, these methods have the following shortcomings: (1) the targets are mostly universal metabolic pathways with poor selectivity and significant toxic side effects; (2) there is a lack of stable and controllable small molecule agonists to regulate metabolic pathways from upstream; (3) the AMPK signaling pathway exhibits dual regulatory effects in different cancer types, and direct inhibition of AMPK may lead to systemic metabolic disorders. Therefore, it is urgent to find new, more specific, and less toxic therapeutic approaches to overcome the paclitaxel resistance dilemma in TNBC. Summary of the Invention

[0005] The purpose of this invention is to provide the application of rumacato in improving paclitaxel resistance in triple-negative breast cancer, thereby overcoming paclitaxel resistance in breast cancer.

[0006] The application provides a use of a pharmaceutical composition in preparation of a drug for preventing and / or treating triple-negative breast cancer resistant to paclitaxel.

[0007] The active ingredients of the pharmaceutical composition are lumarabstatin and paclitaxel.

[0008] In the use, the molar ratio of lumarabstatin and paclitaxel is 1:0.1-1:100, preferably 1:0.5-1:5, and more preferably 1:1-1:2 or 1:1.5.

[0009] The application also provides a use of a pharmaceutical composition in preparation of a product for inhibiting proliferation of paclitaxel-resistant cells.

[0010] The active ingredients of the pharmaceutical composition are lumarabstatin and paclitaxel.

[0011] In the use, the molar ratio of lumarabstatin and paclitaxel is 1:0.1-1:100, preferably 1:0.5-1:5, and more preferably 1:1-1:2.

[0012] Thirdly, the application provides a paclitaxel treatment sensitizer for triple-negative breast cancer, and the active ingredient of the sensitizer is lumarabstatin.

[0013] Finally, the application provides a drug for preventing and / or treating triple-negative breast cancer resistant to paclitaxel, and the active ingredients of the drug are lumarabstatin and paclitaxel.

[0014] In the drug, the molar ratio of lumarabstatin and paclitaxel is 1:0.1-1:100, preferably 1:0.5-1:5, and more preferably 1:1-1:2.

[0015] In the application, the drug resistance is caused by an endocrine treatment drug paclitaxel.

[0016] The present application first identifies GTM1 (Glycolytic Transition Modulator 1) as a negative regulator in paclitaxel resistance, and the expression of which is inhibited to cause the activity of an AMPK-ENO1 axis to be enhanced, and glycolysis metabolism to be increased, and the recovery of the expression of GTM1 can inhibit the above metabolic processes and restore drug sensitivity. Therefore, the development of a small molecule drug capable of agonizing GTM1 becomes a new strategy for reversing paclitaxel resistance.

[0017] Lumacaftor is a small molecule drug approved by FDA for the treatment of cystic fibrosis, which has good oral bioavailability and safety. It is found for the first time that lumacaftor can significantly enhance the killing effect of paclitaxel on TNBC drug-resistant cells, and the mechanism may be related to the inhibition of glycolysis pathway, the recovery of cell cycle arrest and the promotion of apoptosis signal. As a low-toxicity, safe and orally available clinical drug, lumacaftor is expected to become an effective sensitizer in the treatment of paclitaxel-resistant TNBC, expand its clinical indications, and significantly improve the survival outcome of patients.

[0018] As a GTM1 agonist, lumacaftor can inhibit the AMPK-glycolysis pathway and reduce lactic acid production. In paclitaxel-resistant cells (MDA-MB-231PR, BT549PR), it can significantly inhibit cell proliferation (CCK-8, EdU) when combined with paclitaxel, but has no significant effect on parent cells. GTM1 knockdown can eliminate the above effects, and rescue can restore them, proving that the sensitization effect depends on GTM1. Therefore, lumacaftor can be used as a chemosensitizer for paclitaxel-resistant triple-negative breast cancer, with a clear mechanism and good application prospect. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1 Molecular docking screening diagram for GTM1 agonists.

[0020] Figure 2 IC50 determination diagram of midostaurin, nilotinib, alectinib and lumacaftor on triple-negative breast cancer cells. 50

[0021] Figure 3 Measurement results of lactic acid production of triple-negative breast cancer cells treated with midostaurin, venetoclax, nilotinib, alectinib and lumacaftor.

[0022] Figure 4 Western blot (WB) detection results of the influence of midostaurin, nilotinib, alectinib and lumacaftor on GTM1 protein expression.

[0023] Figure 5 Western blot (WB) results of lumacaftor recovery experiment in shGTM1 background.

[0024] Figure 6 Influence of lumacaftor on parent cells MDA-MB-231 and BT549 and paclitaxel-resistant cells MDA-MB-231 and BT549. PR PR

[0025] Figure 7 ​​​For the treatment of MDA-MB-231 cells and paclitaxel-resistant MDA-MB-231 cells by combining rumacata and paclitaxel. PR It has a proliferative inhibitory effect.

[0026] Figure 8 Rumacata combined with paclitaxel for the treatment of BT549 cells and paclitaxel-resistant BT549 cells PR It has a proliferative inhibitory effect. Detailed Implementation

[0027] The present invention will be further described in detail below with reference to specific embodiments. The embodiments given are only for illustrating the present invention and are not intended to limit the scope of the present invention.

[0028] Unless otherwise specified, the experimental methods described in the following examples are conventional methods.

[0029] Unless otherwise specified, the quantitative experiments in the following examples are all repeated three times, and the results are averaged.

[0030] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0031] The biological materials, reagents, and kits used in the following examples are all commercially available, and the experimental techniques used are all standard procedures in the field or performed in accordance with the instructions of the corresponding products.

[0032] The English name of paclitaxel used in the following examples is Paclitaxel.

[0033] The English name of the Lumacaftor used in the following embodiments is Lumacaftor, CAS number: 936727-05-8.

[0034] The MDA-MB-231 and BT549 cell lines used in the following examples were obtained from the Key Laboratory of Breast Cancer Diagnosis and Treatment Research, Affiliated Cancer Hospital of Shantou University Medical College. Paclitaxel-resistant MDA-MB-231 PR and BT549 PRThe cells were constructed as follows: in 10% (v / v) carbon adsorbed fetal bovine serum (Biological Industries), 1640 culture solution (Biological Industries) without phenol red / glutamine, first, the logarithmic growth phase of the parent cells was inoculated in a 25T culture flask, 0.25 μM paclitaxel was added for 48-72 h, when the cells appeared 70%-80% round shrinkage or the survival rate detected by trypan blue was ≤20%-30%, the drug solution was discarded and replaced with drug-free medium for continuous culture, and the next round of induction was carried out when the cells recovered to 70%-80% confluence. This was repeated for 3-6 rounds until the cells could be continuously passaged and stably grown under the condition of 0.25 μM. Subsequently, the drug concentration was gradually increased to 0.5, 1, 2 and 3 μM, respectively, and each concentration was screened according to the same "induction-recovery" cycle method until the cells could be stably passaged for ≥2 weeks under this concentration. Finally, when the cells could be continuously passaged for ≥8-12 weeks and stably grown in 3 μM paclitaxel medium, single clones were isolated by limiting dilution or cloning ring method, and clones with uniform morphology and stable proliferation were screened and expanded in 3 μM paclitaxel medium to obtain paclitaxel-resistant cell lines. The whole process of induction-recovery-increase-cloning lasted about 12 months. To maintain the drug resistance, 1 μM paclitaxel was used to maintain the pressure in daily culture, and the drug was stopped 7-14 days before the experiment. The above culture was carried out in a culture box at 37°C, 5.0% CO2.

[0035] The culture, incubation or treatment described in the following examples were carried out in a culture box at 37°C, 5.0% CO2 unless otherwise specified.

[0036] GTM1 in the following examples refers to the human RSRP1 gene (Gene ID: 57035) retrieved from the NCBI gene database.

[0037] Example 1

[0038] Molecular docking screening experiment of GTM1 agonist: first, download FDA small molecule drugs from ZINC database, optimize small molecule structure using openbable and UFF force field, prepare protein GTM1 and receptor using ADFRsuite1.0, and perform docking using vina1.2.5. The top five small molecules in the docking binding energy are screened out, and the results are as follows Figure 1 , which are midostaurin, nilotinib, venetoclax, alectinib and lumacaftor. Among them, lumacaftor has better comprehensive binding energy and conformational stability, suggesting that it has the potential to become a GTM1 agonist.

[0039] Example 2

[0040] CCK-8 method drug IC 50Assay experiment: To evaluate the inhibitory ability of different small molecule drugs on the proliferation of triple-negative breast cancer cells and determine their IC 50 values, MDA-MB-231 and BT549 cells were inoculated in 96-well plates, 100 μL of cell suspension (culture medium was 1640 complete culture medium) was added to each well, the density was about 5000 cells / well, and different concentrations of drugs were given after the cells adhered. Five groups of drugs were set in the experiment: control group (DMSO), Midostaurin, Nilotinib, Alectinib and Lumacaftor, five concentration gradients were set for each drug: 0 (DMSO control), 100 nM, 200 nM, 1 μM and 2 μM, and 6 replicate wells were set for each group. Drug treatment lasted for 0, 24, 48 and 72 hours. At each time point, the original culture solution was removed, 100 μL of complete culture medium containing 10% CCK-8 reagent (CCK-8 and culture medium were prepared at a volume ratio of 1:9) was added to each well, and after incubation in a 37°C, 5% CO2 incubator for 2 hours, the absorbance (OD 450 ) was measured at 450 nm wavelength using a microplate reader. According to the OD values at different concentrations and time points, the cell activity curve was drawn, and the 72-hour data was used for dose-response fitting to calculate the IC 50 values of each drug in MDA-MB-231 and BT549 cells, thereby evaluating their sensitivity and inhibitory effect on cell growth, and the results are shown in Figure 2 . Among them, Venetoclax was used according to the literature reference concentration of 2 μM.

[0041] As Figure 2 can be seen, in MDA-MB-231 and BT549 cells, CCK-8 detection results showed that five drugs could effectively inhibit cell proliferation, among which Lumacaftor showed stable and significant inhibitory activity in both cell lines, with a half-inhibitory concentration (IC 50 ) in the low micromolar level. This result shows that Lumacaftor has a strong anti-proliferative effect on triple-negative breast cancer cells, providing experimental evidence for its use as a candidate drug and subsequent mechanism research and combination drug strategy.

[0042] Example 3

[0043] Lactic acid content detection experiment: MDA-MB-231 and BT549 cells were inoculated in 6 cm dishes, about 2 x 10 5cells were added with 2 mL 1640 complete medium, and after the cells adhered, candidate drug Midostaurin (100 nM), Venetoclax (2 μM), Nilotinib (1 μM), Alectinib (100 nM), Lumacaftor (2 μM) and DMSO control group were added respectively, and 3 replicate wells were set for each group. After 72 hours of drug treatment, the cells were trypsinized and collected into centrifuge tubes, washed with cold PBS, centrifuged, and the supernatant was discarded. The cells were broken by ice bath ultrasonic wave for 5 min (power 200 W, ultrasonic wave for 3 s, interval 7 s, repeated 30 times) at a ratio of 1 mL / 5 million with Lactate Assay Buffer, then centrifuged at 12,000 g, 4℃ for 5 min, and the supernatant was taken and measured on ice. The lactic acid content was determined by using Abbkine CheKine TM Lactic acid content detection kit (item number KTB1100) was used, and working solution (Lactate Assay Buffer, Cofactor, WST-8, Enhancer and Lactate Dehydrogenase) was prepared according to the instructions. 50 μL of cell sample was added to each well and mixed with 50 μL of working solution. After incubation at 37℃ for 30 min in the dark, the absorbance value was read on a microplate reader at 450 nm wavelength. Before the experiment, a lactic acid standard curve (0-2 mM) was prepared, and the lactic acid production level in each group of samples was calculated according to the standard curve. The results are shown in Figure 3 .

[0044] As can be seen from Figure 3 compared with the DMSO control group, only Lumacaftor treatment significantly reduced cell lactic acid production, suggesting that Lumacaftor was more effective in inhibiting glycolysis metabolic pathway; Lumacaftor was the only candidate drug that could down-regulate lactic acid, and the rest of the drugs all up-regulated lactic acid.

[0045] Example 4

[0046] Western blot mechanism verification experiment: to further verify the regulation of different small molecule drugs on GTM1 and its downstream signal AMPK pathway, MDA-MB-231 cells were inoculated in 6-well plates at 1×10 6Cells were seeded in 6-well plates at a density of 1 x 105 cells per well in 2 ml of complete 1640 medium. After the cells adhered, they were treated with DMSO (control group), Midostaurin (100 nM), Nilotinib (1 μM), Alectinib (100 nM), and Lumacaftor (2 μM) for 72 hours. After the cells were collected, total protein was extracted using RIPA lysis buffer, and equal amounts of the protein were loaded for SDS-PAGE (8% separating gel) electrophoresis and PVDF membrane transfer. GTM1, p-AMPK (Thr172), AMPK (total), ENO1, and β-Tubulin antibodies were used for incubation, and the Bio-Rad imaging system was used for chemiluminescence detection to record the protein expression signal. Results Figure 4 It was shown that Lumacaftor significantly up-regulated GTM1 protein levels and inhibited p-AMPK (Thr172) and ENO1 expression in MDA-MB-231 cells; Midostaurin, Nilotinib, and Alectinib did not show similar changes; thus, only the Lumacaftor group significantly up-regulated GTM1 expression and simultaneously inhibited p-AMPK and ENO1 expression, suggesting that it has a unique mechanism of action of "activating GTM1, inhibiting AMPK, and down-regulating glycolysis." No similar regulation trend was observed in the other drug groups.

[0047] To further verify the regulation of GTM1 on the AMPK signaling pathway under the action of Lumacaftor and whether it depends on the functional integrity of GTM1, five groups were set up in MDA-MB-231 and BT549 two kinds of triple-negative breast cancer cells: shScramble + DMSO group (negative control), shScramble + Lumacaftor group (drug treatment control), shGTM1 + DMSO group (GTM1 knockdown), shGTM1 + Lumacaftor group (knockdown background drug treatment), and shGTM1 + Lumacaftor + recovery group. Each group was treated with 2 μM Lumacaftor for 72 hours. After cell lysis, protein was extracted using RIPA lysis buffer, and equal amounts of the protein were loaded for 8% SDS-PAGE electrophoresis separation and transfer to PVDF membrane. GTM1, p-AMPK (Thr172), AMPK, ENO1, and β-Tubulin antibodies were used for immunoblotting. The Bio-Rad imaging system was used for chemiluminescence detection. The results are shown in Figure 5As shown, in both MDA-MB-231 and BT549 cells, the shScramble + VX-809 group showed up-regulation of GTM1 and down-regulation of p-AMPK and ENOl, indicating that GTM1 was stimulated by the drug and AMPK-glycolysis axis was inhibited; the trend disappeared in the shGTM1 + lumacaftor group; and in the Rescue group, after the nonsense mutation of GTM1 was restored, the effect of lumacaftor was effective again, further proving that the regulation of lumacaftor on the AMPK pathway and glycolysis depends on the functional integrity of GTM1.

[0048] The shScramble + DMSO group (negative control) is a shScramble stable strain obtained by infecting cells with an shRNA lentivirus carrying an irrelevant sequence and screening with puromycin; an equal volume of DMSO (final concentration ≤0.1%) is added under conventional culture conditions, without adding drugs, as a baseline reference;

[0049] The shScramble + lumacaftor group (drug treatment control) is a shScramble stable strain background, with 2 μM lumacaftor added during the logarithmic growth phase, for evaluating the effect of lumacaftor under normal GTM1 expression conditions;

[0050] The shGTM1 + DMSO group (GTM1 knockdown) is a shGTM1 stable strain obtained by infecting cells with an shRNA lentivirus carrying a shRNA against GTM1 and screening with puromycin; an equal volume of DMSO is added, without adding drugs, for evaluating the effect of continuous knockdown of GTM1 on related proteins / pathways;

[0051] The shGTM1 + lumacaftor group (drug treatment under knockdown background) is a shGTM1 stable strain background, with 2 μM lumacaftor added, for detecting changes in the effect of lumacaftor under GTM1 deletion / low expression conditions;

[0052] The shGTM1 + lumacaftor + rescue (GTM1-rescue) group is a further exogenous restoration of GTM1 based on the shGTM1 stable strain and 2 μM lumacaftor, using a GTM1 cDNA resistant to shRNA, i.e., introducing a synonymous mutation at the shRNA action site to avoid being silenced again, after stable restoration of expression (such as 48-72 h), for detecting whether restoring GTM1 reverses the change in the effect of lumacaftor due to knockdown, thereby proving the necessity of GTM1 in the mechanism of drug action.

[0053] Example 5

[0054] CCK-8 method drug sensitivity experiment: MDA-MB-231 and BT549 and their corresponding paclitaxel-resistant cells were seeded in 96-well plates as described above, about 3000 cells per well, about 100 μL of suspension (culture medium was 1640 complete culture medium). Each cell was divided into four groups: carrier control group, paclitaxel group, lumacaftor group and combined use (paclitaxel + lumacaftor) group. Each group was made into 6-8 replicate wells. After the cells adhered, the carrier control group was added with the corresponding drug solvent control, the paclitaxel group was added with 3 μM paclitaxel, the lumacaftor group was added with 2 μM lumacaftor, and the combined use group was added with 3 μM paclitaxel and 2 μM lumacaftor, respectively. 0, 24, 48 and 72 hours of treatment were performed. The original culture solution in the 96-well plate was aspirated, 100 μL of culture solution containing 10% CCK-8 working solution (Biyun Tian) was added to each well (CCK-8 and culture solution were prepared in advance according to the required total volume, the volume ratio of CCK-8 to culture solution was 1:9), and it was incubated in the incubator for 2 hours. The absorbance at 0, 24, 48 and 72 hours was measured. The wavelength of the microplate reader was set to 450 nm, and the absorbance was measured. The average value of the replicate wells was obtained to obtain the reading. The time (h) was taken as the abscissa, and the cell activity (450 nm absorbance) was taken as the ordinate, and the growth curve was drawn. The results are shown in Figure 6 Figure 6. Lumacaftor had no significant effect on the paclitaxel sensitivity of the parent cells (MDA-MB-231 and BT549), but lumacaftor could significantly enhance the paclitaxel sensitivity of the paclitaxel-resistant cells (MDA-MB-231 PR and BT549 PR ).

[0055] Example 6

[0056] EdU method cell proliferation detection experiment: as described before, MDA-MB-231 and BT549 and their corresponding paclitaxel-resistant cells, each cell was inoculated in a 24-well plate, and when the confluence reached about 70%, each cell was divided into four groups: vehicle control group, paclitaxel group, lumacaftor group and combined drug (paclitaxel + lumacaftor) group. The vehicle control group was added with the corresponding drug solvent control, the paclitaxel group was added with 3 μM paclitaxel, the lumacaftor group was added with 2 μM lumacaftor, and the combined drug group was added with 3 μM paclitaxel and 2 μM lumacaftor, and the treatment time was 0 or 72 hours. Prepare 2x EdU working solution (Bi Yun Tian), preheat to 37°C, for a 24-well plate, the culture volume of each well is 500 μL, and the final concentration of EdU is 10 μM (1x). After adding the preheated working solution, continue to incubate the cells for 2 hours. Remove the culture medium, add 500 μL of fixing solution (Bi Yun Tian) to each well, and fix at room temperature for 15 minutes. Remove the fixing solution, wash each well with 500 μL of washing solution (Bi Yun Tian) for 3 times, each time for 3-5 minutes. Remove the washing solution, add 500 μL of permeabilization solution (Bi Yun Tian) to each well, and incubate at room temperature for 10-15 minutes. Remove the permeabilization solution, wash each well with 500 μL of washing solution for 2 times, each time for 3-5 minutes. Prepare the reaction solution according to the instructions, for a 24-well plate, the reaction solution system is 100 μL, and the reaction solution needs to be used within 15 minutes after preparation. After removing the washing solution, add 100 μL of prepared reaction solution to each well, gently shake the culture plate to ensure that the reaction mixture can uniformly cover the sample, and incubate at room temperature for 30 minutes. Remove the reaction solution, wash with washing solution for 3 times, each time for 3-5 minutes. Use Hoechst 33342 (Bi Yun Tian) to stain the cell nucleus, dilute Hoechst 33342 (1000x) with PBS at a volume ratio of 1:1000. After removing the washing solution, add 1x 500 μL Hoechst 33342 to each well, and incubate at room temperature for 10 minutes. Remove the staining solution, wash with washing solution for 3 times, each time for 3-5 minutes, and then perform fluorescence detection. Hoechst 33342 is blue fluorescence, the maximum excitation wavelength is 346 nm, and the maximum emission wavelength is 460 nm. EdU is labeled with Alexa Fluor 488, which is green fluorescence, the maximum excitation wavelength is 495 nm, and the maximum emission wavelength is 519 nm. The results were analyzed by Image J software as shown in Figure 7 and Figure 8 In parent cells (MDA-MB-231 and BT549), lumacaftor combined with paclitaxel had no significant difference in the inhibition of breast cancer cell proliferation compared with paclitaxel alone; in paclitaxel-resistant cells (MDA-MB-231 PR and BT549 PR ), lumacaftor combined with paclitaxel had a significant inhibitory effect on breast cancer cell proliferation compared with paclitaxel alone.

Claims

1. The use of a pharmaceutical composition in the preparation of a drug for the prevention and / or treatment of paclitaxel-resistant triple-negative breast cancer; The active ingredients of the pharmaceutical composition are rumacato and paclitaxel.

2. The application according to claim 1, characterized in that: The molar ratio of rumacato to paclitaxel is 1:0.1 to 1:

100.

3. The use of a pharmaceutical composition in the preparation of a product that inhibits the proliferation of paclitaxel-resistant cells; The active ingredients of the pharmaceutical composition are rumacato and paclitaxel; The drug-resistant cells are triple-negative breast cancer drug-resistant cells.

4. The application according to claim 3, characterized in that: The molar ratio of rumacato to paclitaxel is 1:0.1 to 1:

100.

5. The application according to claim 4, characterized in that: The molar ratio of rumacato to paclitaxel is 1:0.5 to 1:

5.

6. The application according to claim 5, characterized in that: The molar ratio of rumacato to paclitaxel is 1:1 to 1:2.

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