Abstract
Lung cancer cells are sensitive to 5-aza-2'-deoxycytidine (decitabine) or midostaurin (PKC412), because decitabine restores the expression of methylation-silenced tumor suppressor genes, whereas PKC412 inhibits hyperactive kinase signaling, which is essential for cancer cell growth. Here, we demonstrated that resistance to decitabine (decitabine(R)) or PKC412 (PKC412(R)) eventually results from simultaneously remethylated DNA and reactivated kinase cascades. Indeed, both decitabine(R) and PKC412(R) displayed the up-regulation of DNA methyltransferase DNMT1 and tyrosine-protein kinase KIT, the enhanced phosphorylation of KIT and its downstream effectors, and the increased global and gene-specific DNA methylation with the down-regulation of tumor suppressor gene epithelial cadherin CDH1. Interestingly, decitabine(R) and PKC412(R) had higher capability of colony formation and wound healing than parental cells in vitro, which were attributed to the hyperactive DNMT1 or KIT, because inactivation of KIT or DNMT1 reciprocally blocked decitabine(R) or PKC412(R) cell proliferation. Further, DNMT1 knockdown sensitized PKC412(R) cells to PKC412; conversely, KIT depletion synergized with decitabine in eliminating decitabine(R). Importantly, when engrafted into nude mice, decitabine(R) and PKC412(R) had faster proliferation with stronger tumorigenicity that was caused by the reactivated KIT kinase signaling and further CDH1 silencing. These findings identify functional cross-talk between KIT and DNMT1 in the development of drug resistance, implying the reciprocal targeting of protein kinases and DNA methyltransferases as an essential strategy for durable responses in lung cancer.
Highlights
The DNA methyltransferase DNMT1 and tyrosine-protein kinase KIT are crucial for lung tumorigenesis, and the resistance to their inhibitors invariably develops
It is reported that DNMT1 and KIT are highly expressed in certain chemotherapeutic resistance [34, 36], and because aberrant DNA methylation appears in resistance to imatinib (Gleevec) in leukemia [37], we proposed that KIT and DNMT1 may work in concert during lung tumor progression to a drugresistant phenotype
Specific KIT knockdown led to a decrease of Sp1 and DNMT1 expression followed by DNA demethylation (Fig. 3C); enforced KIT expression caused the up-regulation of DNMT1 and Sp1, leading to an increase of DNA methylation (Fig. 3D). These findings suggest that DNMT1 and KIT pathways have a reciprocal regulation in lung cancer cells
Summary
The DNA methyltransferase DNMT1 and tyrosine-protein kinase KIT are crucial for lung tumorigenesis, and the resistance to their inhibitors invariably develops. When engrafted into nude mice, decitabineR and PKC412R had faster proliferation with stronger tumorigenicity that was caused by the reactivated KIT kinase signaling and further CDH1 silencing These findings identify functional cross-talk between KIT and DNMT1 in the development of drug resistance, implying the reciprocal targeting of protein kinases and DNA methyltransferases as an essential strategy for durable responses in lung cancer. Our findings offer mechanistic insight into decitabine and PKC412 resistance, and they illustrate how reciprocal application of inhibitors for DNMT1 and KIT oncogenic pathways may improve the anticancer responses of decitabine and PKC412, and potentially, other types of DNA methylation and RTK inhibitors in lung cancer therapy
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