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K. Takeuchi
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MA07 - ALK-ROS1 in Advanced NSCLC (ID 385)
- Event: WCLC 2016
- Type: Mini Oral Session
- Track: Advanced NSCLC
- Presentations: 1
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MA07.10 - HDAC Inhibition Overcomes Crizotinib-Resistance by Mesenchymal-Epithelial Transition (MET) in EML4-ALK Lung Cancer Cells (ID 4367)
12:06 - 12:12 | Author(s): K. Takeuchi
- Abstract
- Presentation
Background:
ALK rearrangement, most commonly EML4-ALK, is detected in approximately 3–7% of non-small cell lung cancer (NSCLC). Crizotinib, an ALK tyrosine kinase inhibitor (TKI), shows dramatic clinical efficacy in ALK-rearranged NSCLC patients. However, almost all patients acquire resistance after only 1 to 2 years. A variety of mechanisms, including ALK-secondary mutations, ALK amplification, and activation of alternative pathway, have been reported to mediate acquired resistance to crizotinib. While epithelial–mesenchymal transition (EMT) was recently reported to be associated with resistance to crizotinib in EML4-ALK lung cancer cells in vitro, the underlying mechanism has not been defined and no optimal therapy to overcome EMT-associated resistance has been identified.
Methods:
We continuously gave crizotinib treatment to SCID mice inoculated with EML4-ALK lung cancer cell line A925L into thoracic cavity and established crizotinib resistant A925LCR cells. After the limiting dilution of A925LCR cells, we obtained several single cell clones. The effects of the HDAC inhibitor quisinostat on the EMT state and the growth of the cells were examined in vitro and in vivo.
Results:
We found that some clones acquired EMT phenotypes, such as spindle shape morphology, expression of EMT-related proteins, and increased cell motility. Interestingly, Histone deacetylase (HDAC) inhibitor, quisinostat, induced mesenchymal-epithelial transition (MET) of A925LCR clones in vitro. Quisinostat reduced ZEB1 expression, induced MET, and thus restored sensitivity to crizotinib. Knockdown of ZEB1 expression in the A925LCR clones by si-RNA also induced MET and restored sensitivity to crizotinib, suggesting that quisinostat-induced MET depends on ZEB-1 suppression. MicroRNA profile analysis revealed that the A925LCR clones expressed significantly lower levels of miR-200 family including miR-200c which targets ZEB1, compared with parental A925L cells. Furthermore, quisinostat recovered miR-200c expression and antago-miR-200c abrogated quisinostat-induced MET in the A925LCR clone cells. These results indicate that quisinostat induced MET by up-regulating miR-200c expression which target ZEB1 and thereby re-sensitizing to crizotinib. In a pleural carcinomatosis model with A925LCR clone cells, quisinostat induced MET and caused remarkable tumor regression during the subsequent crizotinib re-challenge. Furthermore, we analyzed tumor tissue obtained at autopsy from an ALK-rearranged NSCLC patient who acquired resistance to crizotinib. We found that EMT was induced in both primary and metastasis lesions after crizotinib treatment, indicating that EMT is associated with crizotinib resistance in clinical therapy.
Conclusion:
Our findings suggest that EMT is possibly occurred in acquired resistance to crizotinib and intermittent use of HDAC inhibitor could be a novel therapeutic strategy for overcoming EMT-associated crizotinib-resistance in EML4-ALK lung cancer.
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