[Background] The treatment targeting signal addiction caused by oncogenic driver mutation has led to unprecedented results in clinical setting; however, it is still difficult to achieve cure. Tumor diversity based on both genetic and non-genetic influences give rise of survivors as treatment-resistant cells. Most of the acquired resistance reflects the selection of the cancer cells harboring stochastic resistance-conferring genetic alterations. However, the mechanisms how the cancer cells survive until acquisition of additional mutations are unclear.

We identified DEAD/H (Asp-Glu-Ala-Asp/His) box polypeptide 3, X-linked (DDX3X), an ATP-dependent RNA helicase, was preferentially expressed in a cancer stem cell (CSC)-like population of murine melanoma. DDX3X is believed to be involved in epigenetic regulation of gene expression based on metabolism of mRNA and miRNA.

[Objectives] In the current study, we examine that DDX3X affects phenotype and signal status of PC9 cells, a lung adenocarcinoma harboring EGFR exon19 deletion as an oncogenic driver mutation.

[Results] DDX3X plays a role in acquisition of CSC-like properties. PC9 A-1 cells that were engineered to overexpress DDX3X acquired ability to proliferate as tumor spheres and showed up-regulated Sox2 and ALDH expression. Epithelial mesenchymal transition with cadherin switching from E-cadherin to N-cadherin accompanied with vimentin expression was detected in non-adherent A-1 population. E-cadherin+ non-adherent cells exhibited strong CD44 expression, a putative CSC marker. Surprisingly, A-1 cells exhibited almost no EGFR phosphorylation even in the presence of EGF. Knocking-down of DDX3X restored EGFR phosphorylation. On the other hand, activated nuclear translocation of β-catenin was observed. Loss of EGFR signal addiction resulted in resistance to EGFR-TKI. On contrast, PC9-A1 cells were sensitive to ICG-001, a β-catenin signal inhibitor. Further, we identified a minor non-adherent subpopulation of parental PC9 cells strongly expressed DDX3X and that they were resistant to EGFR-TKI because they did not addict to EGFR signaling. Survivor cells of parental PC9 after long term-exposure to EGFR-TKI strongly expressed DDX3X.

[Conclusion] DDX3X plays a critical role in a novel EGFR-TKI resistance mechanism, signal switching, correlating with CSC transformation. DDX3X and β-catenin are likely promising target molecules to overcome EGFR-TKI resistance.

Citation Format: Satoshi Shoji, Hiroshi Kagamu, Koichiro Nozaki, Natsue Igarashi, Masaaki Okajima, Satoru Miura, Satoshi Watanabe, Hrohisa Yoshizawa, Ichiei Narita. DDX3X induces signal switching to stem cell-specific Wnt/β-catenin signaling, resulting in EGFR-TKI resistance in lung cancer cells harboring EGFR activating mutation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 200. doi:10.1158/1538-7445.AM2014-200