Ai Xin-Yi, Hou Xue-Feng, Feng Nian-Ping
School of Pharmacy, Shanghai University of Traditional Chinese Medicine Shanghai 201203, China.
School of Pharmacy, Shanghai University of Traditional Chinese Medicine Shanghai 201203, China School of Pharmacy, Wannan Medical College Wuhu 241002, China.
Zhongguo Zhong Yao Za Zhi. 2024 Jan;49(1):175-184. doi: 10.19540/j.cnki.cjcmm.20230810.401.
The occurrence and development of tumors are associated with the cell energy metabolism. Inhibiting energy metabolism of lung cancer cells is an important strategy to overcome drug resistance. Based on the cellular energy metabolism pathway, this study observed the effect of combination of shikonin(SKN) and gefitinib(GFB) on the drug resistance in non-small cell lung cancer and explored the underlying mechanism. The human non-small cell lung cancer line HCC827/GR resistant to gefitinib was used as the cell model in vitro. The CCK-8 assay and flow cytometry were employed to investigate the cell viability and apoptosis, respectively. The high performance liquid chromatography was employed to measure the intracellular accumulation of GFB. A Seahorse XFe96 Analyzer was used to detect the changes of cellular energy metabolism. Western blot was employed to determine the expression of the proteins involved in the drug resistance. The tumor-bearing nude mouse model was used to verify the efficacy of SKN+GFB in overcoming drug resistance in vivo. The results showed that SKN+GFB significantly reduced the IC_(50) of GFB on HCC827/GR cells, with the combination index of 0.628, indicating that the combination of the two drugs had a synergistic effect and promoted cell apoptosis. SKN increased the intracellular accumulation of GFB. SKN+GFB lowered the oxygen consumption rate(OCR) and glycolytic proton efflux rate(GlycoPER) in cell energy metabolism, and down-regulated the overexpression of PKM2, p-EGFR, P-gp, and HIF-1α in drug resistance. The results of reversing drug resistance test in vivo showed that GFB or SKN alone had no significant antitumor effect, while the combination at different doses induced the apoptosis of the tumor tissue and inhibited the expression of PKM2 and P-gp, demonstrating a significant antitumor effect. Moreover, the tumor inhibition rate in the high-dose combination group reached 64.01%. In summary, SKN+GFB may interfere with the energy metabolism to limit the function of HCC827/GR cells, thus reversing the GFB resistance in non-small cell lung cancer.
肿瘤的发生发展与细胞能量代谢相关。抑制肺癌细胞的能量代谢是克服耐药性的重要策略。基于细胞能量代谢途径，本研究观察了紫草素（SKN）与吉非替尼（GFB）联合应用对非小细胞肺癌耐药性的影响，并探讨其潜在机制。将对吉非替尼耐药的人非小细胞肺癌细胞系HCC827/GR用作体外细胞模型。采用CCK-8法和流式细胞术分别检测细胞活力和凋亡情况。采用高效液相色谱法测定细胞内GFB的蓄积量。使用Seahorse XFe96分析仪检测细胞能量代谢的变化。采用蛋白质印迹法测定与耐药相关的蛋白质表达。利用荷瘤裸鼠模型在体内验证SKN+GFB克服耐药性的疗效。结果显示，SKN+GFB显著降低了GFB对HCC827/GR细胞的IC50，联合指数为0.628，表明两种药物联合具有协同作用并促进细胞凋亡。SKN增加了细胞内GFB的蓄积量。SKN+GFB降低了细胞能量代谢中的氧消耗率（OCR）和糖酵解质子外流率（GlycoPER），并下调了耐药相关的PKM2、p-EGFR、P-gp和HIF-1α的过表达。体内逆转耐药试验结果显示，单独使用GFB或SKN均无显著抗肿瘤作用，而不同剂量联合用药可诱导肿瘤组织凋亡并抑制PKM2和P-gp的表达，显示出显著的抗肿瘤作用。此外，高剂量联合组的肿瘤抑制率达到64.01%。综上所述，SKN+GFB可能通过干扰能量代谢来限制HCC827/GR细胞的功能，从而逆转非小细胞肺癌对GFB的耐药性。
