The role and mechanism of 2-deoxyglucose in reversing osimertinib-acquired resistance of non-small cell lung cancer cell line

Abstract

Objective: To explore the role and mechanism of 2-deoxyglucose (2-dg) in reversing osimertinib- acquired resistance of non-small cell lung cancer(NSCLC)cell line. Methods: The NSCLC line H1975 (purchased from the American Type Culture Collection) was conducted by induction method in vitro to construct the osimertinib-resistance NSCLC cell line H1975-OR. The osimertinib-resistance of H1975-OR cell line was examined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony-formation assay, Ki67 incorporation assay and the expression of apoptosis-related protein. The glycolysis level was assayed by the lactic acid production measured in the culture medium supernatant of H1975 and H1975-OR. The expression of glycolysis key enzymes (HK2, GLUT1, P-PKM2) and apoptosis-related protein (BIM, Bcl-2) were detected by Western blot. The cells were divided into control group, 2-deoxyglucose (4 mmol/L) monotherapy group, osimertinib (3 μmol/L) monotherapy group and 2-deoxyglucose (4 mmol/L)+ osimertinib (3 μmol/L) combination therapy group, then the apoptosis rate of cells was measured by flow cytometry to evaluate the pro-apoptotic ability of drugs. Date were analyzed by Independent-Samples t-test using SPSS 16.0 statistical software. Results: The glycolysis level of osimertinib-sensitive cell line H1975 was lower than that of osimertinib-resistance cell line H1975-OR [the yield of lactic acid, respectively, was (21.0±0.9) and (26.5±2.8) mmol·L(-1)·10(4)cells(-1), P<0.05]. The osimertinib- acquired resistance of H1975-OR could be reversed by 4 mmol/L 2-deoxyglucose(the IC(50) value of osimertinib in H1975-OR cell line decreased from (7.0±1.9) μmol/L to (1.4±0.1) μmol/L, which was close to the IC(50) value of osimertinib in H1975 cell line (1.0±0.2) μmol/L. The apoptosis rate of H1975-OR was significantly higher in 2-deoxyglucose + osimertinib combination therapy group (26.7±2.4)%, compared to control group (5.1±0.7)%, 2-deoxyglucose monotherapy group (6.1±2.5)% and osimertinib monotherapy group (11.4±2.7)%(all P<0.05). The expression of pro-apoptotic protein BIM in H1975-OR was significantly higher in 2-deoxyglucose+ osimertinib combination therapy group (177.8±28.1)% and the expression of anti-apoptotic protein Bcl-2 in H1975-OR was significantly lower in 2-deoxyglucose+ osimertinib combination therapy group (24.6±5.2)%, compared to control group (100±0)%, all P<0.05. Conclusion: 2-deoxyglucose can reverse the acquired resistance of NSCLC cell line to osimertinib, which may be related to the inhibition of cell glycolysis and the induction of apoptosis.