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, 24 (4), 217-223

Overcoming the Intrinsic Gefitinib-resistance via Downregulation of AXL in Non-small Cell Lung Cancer

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Overcoming the Intrinsic Gefitinib-resistance via Downregulation of AXL in Non-small Cell Lung Cancer

Inae Jeong et al. J Cancer Prev.

Abstract

Background: Resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), such as gefitinib, is a limited factor in the treatment of non-small-cell lung cancer (NSCLC) patients. Therefore, ongoing studies are trying to identify EGFR-TKIs-resistant mechanisms and to discover novel therapeutic strategies and targets for NSCLC treatment.

Methods: In the present study, the possibility of overcoming intrinsic gefitinib-resistance was examined by regulating the expression of AXL. A natural product-derived antitumor agent, yuanhuadine (YD) was employed to modulate the expression of AXL in the cells.

Results: Treatment with YD effectively downregulated AXL expression in AXL-overexpressed gefitinib-resistant H1299 cells. The combination of gefitinib and YD exhibited a synergistic grwoth-inhibitory activity in H1299 cells by downregulation of AXL expression.

Conclusions: Based on these findings, AXL was found to be a promising therapeutic target to overcome the intrinsic resistance to gefitinib in NSCLC. Furthermore, YD is able to effectively regulate the expression of AXL and thus it may be applicable as a potential lead compound for the treatment of gefitinib-resistant NSCLC.

Keywords: Drug resistance; Gefitinib; Non-small-cell lung cancer; Yuanhuadine; axl receptor tyrosine kinase.

Conflict of interest statement

CONFLICTS OF INTEREST No potential conflicts of interest were disclosed.

Figures

Figure 1
Figure 1
Cell proliferative activity of gefitinib in non-small-cell lung cancer cell lines. The cells were treated with gefitinib for 48 hours (A) or 48 and 72 hours (B). The cell proliferation was then determined by the sulforhodamine B assay as described in Materials and Methods.
Figure 2
Figure 2
Downregulation of AXL expression by yuanhuadine (YD) in H1299 cell line. (A) The chemical structure of YD. (B) The effect of YD on cell proliferation in H1299 cells. The cells were treated with YD for 48 or 72 hours, and the cell growth was determined by the sulforhodamine B assay. (C) The effect of YD on the expression of AXL in H1299 cells. The cells were treated with the indicated concentrations of YD for 6 hours, and cell lysates were analyzed by Western blotting. β-actin was used as a loading control. (D) The effect of YD on the expression of AXL mRNA in H1299 cells. The mRNA expression was determined by real-time PCR, and β-actin mRNA levels were used for normalization. The data are presented as the mean ± SD. **P < 0.01 by t-test.
Figure 3
Figure 3
Enhancement of AXL degradation by yuanhuadine (YD) in H1299 cells. (A, C) Cells were treated with 25 μg/mL cycloheximide (CHX) and 10 nM YD for the indicated times. Cell lysates were analyzed by Western blot with an antibody against AXL. β-actin was used as a loading control. (B, D) AXL expression levels were quantifies by densitometry using ImageJ.
Figure 4
Figure 4
Effect of yuanhuadine (YD) and gefitinib in combination on cell proliferation in H1299 cells. The cells were treated with either YD alone or in combination with gefitinib for 48 hours, and then the cell proliferation was determined by the sulforhodamine B assay. CI, combination index.
Figure 5
Figure 5
Downregulation of AXL expression in combination with yuanhuadine (YD) and gefitinib in H1299 cells. The cells were treated with YD (10 nM) and/or gefitinib (12.5 μM) for 6 hours, and the cell lysates were analyzed by Western blotting. β-actin was used as a loading control.

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