Application of CTPI-2 / Orlistat / Etomoxir in the preparation of drugs to reverse the resistance of third-generation EGFR-TKI osimertinib in lung cancer

Abstract

The present invention relates to the field of medical technology, and proposes a drug for reversing the third-generation EGFR-TKI osimertinib resistance in lung cancer, wherein the drug is one of the fatty pathway inhibitors CTPI-2, GL, Orlistat and Etomoxir. This scheme proves through in vitro cell experiments that fatty acid pathway inhibitors can effectively reverse the third-generation EGFR-TKI osimertinib resistance, among which orlistat, a patent medicine that has been used for clinical obesity treatment, has a significant effect. In vivo experiments on mice prove that orlistat combined with osimertinib orally has no obvious toxic reaction and can effectively overcome osimertinib resistance.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a drug for reversing the drug resistance of the third-generation EGFR-TKI osimertinib in lung cancer. Background Art

[0002] The 5-year survival rate of patients with advanced lung cancer is less than 5%. For lung adenocarcinoma with EGFR gene sensitive mutations or ALK fusion genes, the objective response rate of targeted therapy is 55%-90%, which significantly improves the prognosis of lung cancer. For patients with advanced non-small cell lung cancer (NSCLC), especially those with tumors with driver gene mutations, EGFR mutations (EGFRm) are the most common, especially affecting young women and non- or light smokers. Classic EGFR mutations include exon 19 deletion (del19) and exon 21 point mutation (L858R), accounting for 80%-85% of EGFR mutations. The remaining 10%-15% of mutations are mainly located in exons 18-21 and have poor efficacy.

[0003] For patients with EGFRm advanced NSCLC and del19 or L858R mutations, first-line treatment with EGFR tyrosine kinase inhibitors (TKIs) has become the standard. The third-generation TKI osimertinib is superior to the first-generation TKIs (erlotinib, gefitinib) due to longer progression-free survival (PFS, average 18.9 months), overall survival (OS, average 38.6 months), higher intracranial response rate and less toxicity. Recently studied combination therapy strategies, such as chemotherapy combined with TKI or novel EGFR-directed therapies, are expected to surpass the effects of single-agent TKIs. Despite some success, acquired treatment resistance and central nervous system (CNS) progression remain major challenges.

[0004] Acquired resistance to osimertinib is inevitable and diverse, with resistance mechanisms including: (1) acquisition of second-site mutations in EGFR; (2) upregulation of certain pathways that promote cancer signaling even in the absence of EGFR; (3) transformation to small-cell lung cancer; and (4) insufficient drug penetration in sanctuary sites such as the central nervous system. Resistance to first-generation EGFR TKIs is usually caused by the EGFR T790M mutation, but resistance to the third-generation EGFR TKI osimertinib is more diverse. Because actionable resistance mechanisms are rare, the standard second-line option for most patients who progress after osimertinib treatment is usually empirical platinum-based doublet chemotherapy. Multiple trials have been designed to address patients who progress after osimertinib treatment and lack a clear resistance mechanism, including EGFR / MET combination chemotherapy and combination studies of immune checkpoint inhibitors (ICIs) with chemotherapy and vascular endothelial growth factor inhibitors (VEGF-i).

[0005] In the MARIPOSA-2 trial, 657 patients with disease progression after osimertinib were randomly assigned to platinum-doublet chemotherapy with or without amivantamab (ervantamab, a bispecific antibody for EGFR-MET) or to a third arm of platinum-doublet chemotherapy with amivantamab and lazertinib (a third-generation EGFR TKI). The addition of amivantamab (with or without lazertinib) to chemotherapy significantly improved progression-free survival compared with chemotherapy alone: ​​median PFS was 6.3 months (hazard ratio [HR], 0.48; 95% CI, 0.36-0.64) in the amivantamab plus chemotherapy group and 8.3 months (HR, 0.44; 95% CI, 0.35-0.56) in the amivantamab plus lazertinib plus chemotherapy group, compared with a median PFS of 4.2 months with chemotherapy alone. Common side effects in the amivantamab group included infusion reactions (chills, flushing, dyspnea, chest pain, nausea / vomiting), EGFR- and MET-mediated mucocutaneous toxicities, and venous thromboembolism, which raised concerns about the tolerability of this regimen.[9] Importantly, in cohort E of the CHRYSALIS study, the objective response rate with amivantamab plus lazertinib was only 29% (95% CI, 13-49) and the median PFS was 4.1 months (95% CI, 1.4-9.5) in patients with MET-enhanced tumors who had not received chemotherapy. Therefore, when resistance mechanisms that cannot be identified or exploited exist, platinum-doublet chemotherapy remains a core part of treatment after osimertinib.

[0006] In the MARIPOSA-2 trial, 657 patients with disease progression after osimertinib were randomly assigned to platinum-doublet chemotherapy without or with amivantamab (eivantamab, a bispecific antibody for EGFR-MET) or platinum-doublet chemotherapy plus amivantamab and lazertinib (a third-generation EGFR TKI). Adding amivantamab (with or without lazertinib) to chemotherapy significantly improved progression-free survival: median PFS was 6.3 months (HR, 0.48; 95% CI, 0.36-0.64) in the amivantamab + chemotherapy group and 8.3 months (HR, 0.44; 95% CI, 0.35-0.56) in the amivantamab + lazertinib + chemotherapy group, respectively, compared with 4.2 months in chemotherapy alone. Common side effects of the amivantamab group included infusion reactions (such as chills, flushing, dyspnea, etc.), mucocutaneous toxicity, and venous thrombosis, which raised concerns about tolerability. In cohort E of the CHRYSALIS study, the objective response rate of amivantamab plus lazertinib in patients with MET-enhanced tumors who had not received chemotherapy was only 29% (95% CI, 13-49), and the median PFS was 4.1 months (95% CI, 1.4-9.5). Therefore, platinum-doublet chemotherapy remains the core treatment after osimertinib when no resistance mechanism can be identified or exploited.

[0007] However, platinum-based double-drug chemotherapy has the problems of rapid drug resistance and severe toxic side effects. Therefore, it is urgent to conduct research on the mechanism of resistance to third-generation TKIs in non-small cell lung cancer and explore new ways to overcome resistance to third-generation TKIs. Summary of the invention

[0008] In view of this, the present invention proposes a drug for reversing the drug resistance of the third-generation EGFR-TKI osimertinib in lung cancer, thereby overcoming the drug resistance of the third-generation EGFR-TKI osimertinib in non-small cell lung cancer.

[0009] The technical solution of the present invention is implemented as follows: On the one hand, the present invention provides a drug for reversing the drug resistance of the third-generation EGFR-TKI osimertinib in lung cancer, wherein the drug is one of the fat pathway inhibitors CTPI-2, GL, Orlistat and Etomoxir.

[0010] On the basis of the above technical solution, preferably, the dosage form of the drug is one of tablets, granules, capsules, oral liquid and injection.

[0011] On the basis of the above technical solution, preferably, the drug is the fat pathway inhibitor Orlistat.

[0012] On the other hand, the present invention also provides the use of Orlistat combined with Osimertinib in the preparation of a drug for treating small cell lung cancer.

[0013] The drug of the present invention for reversing the third-generation EGFR-TKI osimertinib resistance in lung cancer has the following beneficial effects compared with the prior art: This scheme proves through in vitro cell experiments that fatty acid pathway inhibitors can effectively reverse the third-generation EGFR-TKI osimertinib resistance, among which orlistat, a patent medicine that has been used for the clinical treatment of obesity, has a significant effect. In vivo experiments on mice prove that orlistat combined with osimertinib has no obvious toxic reaction and can effectively overcome osimertinib resistance. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0015] Figure 1 These are graphs showing the effects of fatty acid pathway inhibitors CTPI-2, GL, Orlistat, and Etomoxir on improving the sensitivity of PC9 wild-type cells and H358 wild-type cells to osimertinib. Graph A shows CTPI-2, Graph B shows GL, Graph C shows Orlistat, and Graph D shows Etomoxir.

[0016] Figure 2 The effect of CTPI-2, GL, Orlistat, and Etomoxir on improving the sensitivity of osimertinib-resistant PC9-OsiR cells to osimertinib is shown in Figure A, CTPI-2, GL, Orlistat, and Etomoxir.

[0017] Figure 3 This is a graph showing the effect of the combination of orlistat and osimertinib on the body weight of mice. In the figure, Control is the solvent control; Olistat is orlistat; Osimertinib is osimertinib; Combined is the combination of orlistat and osimertinib; Cisplatin is cisplatin.

[0018] Figure 4This is a line graph showing the effect of the combination of orlistat and osimertinib on the tumor volume of mice. In the figure, the wild-type cell group: WT-Control is the solvent control, WT-Olistat is orlistat, WT-Osimertinib is osimertinib, WT-Combined is the combination of orlistat and osimertinib, and WT-Cisplatin is cisplatin; the osimertinib-resistant cell group: OsiR-Control is the solvent control, OsiR-Olistat is orlistat, OsiR-Osimertinib is osimertinib, OsiR-Combined is the combination of orlistat and osimertinib, and OsiR-Cisplatin is cisplatin.

[0019] Figure 5 This is a graph showing the effect of the combination of orlistat and osimertinib on the tumor volume in mice. DETAILED DESCRIPTION

[0020] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0021] Example 1 Cell IC50 drug sensitivity test

[0022] The commonly used NSCLC cell line PC9 wild-type cells (PC9-WT), the constructed osimertinib-resistant PC9-OsiR cells, and the commonly used NSCLC cell line H358 wild-type cells (H358-WT) were used respectively. PC9 cells are more sensitive to osimertinib, while H358 cells are less sensitive.

[0023] The cells in good growth state and in the logarithmic growth phase were selected and inoculated in a 96-well plate at a density of 5000 cells / well (6 replicate wells were set for each group). An experimental control group (containing culture medium and cells) and a blank control group (only normal culture medium was added without cells) were set up. After the cells were plated, they were placed in a 37°C, 5% CO 2 Incubate in the incubator for 24 hours. After 24 hours, the control group was cultured according to 10 -9 M, 10 -8 M, 10 -7 M, 10 -6 M, 10 -5 M, 10 -4 Osimertinib (Osi) was added to the cells at a final concentration of 10 M. -6The corresponding inhibitors (Selleck Inhibitor Company) were added to the cells at medium concentrations. After 48 h of action, MTT working solution (Solebo Company) was added. After incubation at 37 °C for 4 h, the cells were dissolved in formazan solution for 10 min. The absorbance was measured at a wavelength of 490 nm using an ELISA reader. The relative cell viability was calculated according to the formula and statistical analysis was performed. The results are shown in Figure 1 .

[0024] The formula is: relative cell viability (%) = (absorbance of experimental group - absorbance of blank group) / (absorbance of control group - absorbance of blank group) × 100%.

[0025] Figure 1 The results showed that with the increasing concentration of osimertinib, the viability of the cell lines PC9-WT and H358-WT gradually decreased. The viability of PC9-WT and H358-WT cells treated with fatty acid pathway inhibitors CTPI-2, GL, Orlistat, and Etomoxir was significantly reduced, indicating that the four fatty acid pathway inhibitors (CTPI-2, GL, Orlistat, and Etomoxir) can effectively increase the sensitivity of the two wild-type cells to osimertinib (Osi).

[0026] akin, Figure 2 The results showed that the viability of resistant PC9-OsiR cells was significantly reduced after treatment with fatty acid pathway inhibitors CTPI-2, GL, Orlistat, and Etomoxir, indicating that the four fatty acid pathway inhibitors can effectively reverse the drug resistance of resistant PC9-OsiR cells. Among them, Orlistat, as the only clinically approved drug, has the greatest impact on the sensitivity of osimertinib (see Table 1).

[0027] Table 1 Drug sensitivity of cells IC50

[0028]

[0029] Example 2 Toxicity test in mice

[0030] BALB / c mice aged 5 to 6 weeks were selected as research subjects and randomly divided into 5 groups: Control (solvent control), Olistat, Osimertinib, Combined (orlistat combined with osimertinib), and Cisplatin, with 10 mice in each group.

[0031] A tumor-bearing model was established by using subcutaneous wild-type cells and osimertinib-resistant cells in mice. One week after tumor bearing, the mice were treated with drugs. The administration method was as follows: orlistat and osimertinib were given to mice by oral gavage, with each mouse receiving a dose of 0.05 mg each time, 5 days randomly per week. Cisplatin was given to mice by intraperitoneal injection, with each mouse receiving a dose of 0.05 mg each time, 3 days randomly per week. The drug was administered for 2 to 3 weeks, and the weight and tumor volume of the mice were observed. The results are as follows: Figure 3 shown.

[0032] Figure 3 As shown in the figure, the weight of mice did not change significantly during the administration period. Orlistat alone did not significantly slow down the tumor growth rate of wild-type cells and resistant cells, while osimertinib alone slowed down the tumor growth of wild-type cells. Orlistat combined with osimertinib significantly reduced the tumor volume of wild-type cells and resistant cells ( Figure 4-5 The above results suggest that orlistat combined with osimertinib has no obvious toxicity reaction, and orlistat combined with osimertinib can overcome osimertinib resistance.

[0033] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. Application of CTPI-2 / Orlistat / Etomoxir in the preparation of drugs to reverse the resistance of third-generation EGFR-TKI osimertinib in lung cancer.

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