Abstract
Hypermethylation of tumor suppressor genes (TSGs) promoters by DNA methyltransferase (DNMT) can be observed in almost all cancers which represent a hallmark of carcinogenesis, including lung cancer. DNMT inhibitors (e.g.5-Aza-CR/CdR) reactivate TSGs to exert anti-cancer activity and have been applied into the clinical. However, it is cytotoxic even at low concentrations, which might be not directly related to DNA methylation. We here investigated an alternative strategy in the lung cancer therapy and aimed to estimate and compare its efficiency and side effects of knockdown of DNMT1 in vitro and in vivo. Lung cancer tissues (n=20) showed enhanced expression of DNMT1 than corresponding non-neoplastic tissues. Similar results were found in lung cancer cell lines A549 and H538. The treatment of 5-Aza-CR or knockdown of DNMT1 in vitro could inhibit the expressions of DNMT1 but restore the TSGs expressions including the Ras association domain family 1A (RASSF1A) and the adenomatous polyposis coli (APC) via the demethylation of its promoter region, which results in the decreased proliferation, increased apoptosis and impaired ability of migration. Importantly, knockdown of DNMT1 by siRNA in vivo also effectively demethylated the RASSF1A and APC promoter, elevated their expressions and limited tumor growth, which functioned like 5-Aza-CR but with alleviated side effects, suggesting that knockdown of DNMT1 might be potential strategy for the treatment of lung cancer with better tolerability.
Highlights
As a global health burden, lung cancer remains the leading cause of cancer-related deaths worldwide with 221,200 estimated new cases, and accounting for 1.59 million deaths annually [1]
To test the possibility that siRNA-DNMT1 demethylated the promoters of Ras association domain family 1A (RASSF1A) and adenomatous polyposis coli (APC), we estimated the status of methylation of two lung cancer cells and found that RASSF1A and APC were demethylated by DNMT1 knockdown for 48h (Figure 3E and 3F), which suggested that loss-of-function of DNMT1 by siRNA could effectively reactivate the tumor suppressive genes RASSF1A and APC via demethylation
Retrospective data showed that the prognosis for advanced-stage lung cancer is very poor, with a 5-year overall survival rate of 17%
Summary
As a global health burden, lung cancer remains the leading cause of cancer-related deaths worldwide with 221,200 estimated new cases, and accounting for 1.59 million deaths annually [1]. Results in vitro showed that knockdown of DNMT1 by siRNA, similar to the 5-Aza-CR, could re-express RASSF1A and APC by demethylating their promoter regions, and could promote apoptosis, inhibit proliferation and migration of two lung cancer cell lines. We demonstrated that knockdown of DNMT1 in vivo could enhance the RASSF1A and APC expression via demethylation, and delayed the lung cancer growth with reduced adverse side effects. These findings implicated that siRNA-DNMT1 could be a potential treatment for lung cancer therapy
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