CD74-ROS1 L2026M mutant enhances autophagy through the MEK/ERK pathway to promote invasion, metastasis and crizotinib resistance in non-small cell lung cancer cells

The treatment of non-small cell lung Cancer (NSCLC) patients harboring a proto-oncogene tyrosine-protein kinase c-ros oncogene 1 (ROS1) fusion gene has greatly benefited from the use of crizotinib. However, drug resistance inevitably occurs after 1 year of treatment. Clinical studies have shown that patients with an L2026M mutation in the ROS1 kinase domain account for about 6% of the total number of crizotinib-resistant cases, which is an important group that cannot be ignored. To explore the mechanism involved, we constructed the HLA class II histocompatibility antigen gamma chain (CD74)-ROS1 L2026M mutant gene by fusion polymerase chain reaction (PCR) and transfected it into H460 and A549 cells. We found that the invasion and metastasis abilities of drug-resistant cells were increased. The results of monodansylcadaverine (MDC) staining, Acridine orange (AO) staining and western blot indicated that the Autophagy level of CD74-ROS1 L2026M mutant NSCLC cells was increased compared with the CD74-ROS1 group, and the inhibition of Autophagy could reverse the increased invasion and metastasis abilities caused by the L2026M mutation. In addition, the L2026M mutation led to excessive activation of the MEK/ERK pathway, and MEK inhibitors could reduce the Autophagy level, invasion and metastasis abilities of cells; additionally, this process could be blocked by rapamycin, an activator of Autophagy. Furthermore, crizotinib treatment activated expression of Src homology region 2 domain-containing phosphatase-2 (SHP2; also known as PTPN11) to upregulate the MEK/ERK pathway, and the combination of MEK inhibitors and crizotinib increased Apoptosis compared with crizotinib alone. In conclusion, our results indicate that the MEK/ERK pathway mediates the induction of invasion, metastasis and crizotinib resistance through Autophagy caused by CD74-ROS1 L2026M mutation in NSCLC cells, and targeting MEK could reverse these processes.

CD74-ROS1 L2026M; MEK/ERK; crizotinib resistance; invasion; migration.