Abstract 4787: The role of CaMK2A in reactive oxygen species homeostasis in lung adenocarcinoma

The role of Calcium/calmodulin-dependent protein kinase II alpha (CaMK2A) in reactive oxygen species homeostasis in lung adenocarcinoma Lung cancer is the most common cancer worldwide and patients have poor prognosis and low survival rate. The unsatisfactory outcomes of lung cancer treatment are related to multiple factors including tumor recrudescence, metastasis, cancer drug resistance, etc. Cancer stem cells are cell subsets within a cancer that excel in tumor properties, especially survival, renewal and resistance to treatment. Reactive oxygen species (ROS) are high energy molecules produced during cellular hyperproliferation and cell death. Studies have shown though a low level of ROS is required to support stem cell functions but excessive ROS damages DNA and predisposes to cell death. Therefore, the ability to attenuate oxidative damage from ROS and maintain redox homeostasis is an important property of aggressive and resilient cancer cells. The multifunctional serine/threonine kinase calcium/calmodulin- dependent protein kinase II alpha (CaMK2A) is a calcium signaling molecule involved in cell growth and stress signals integration. It has been reported to play a tumor-supportive role in osteosarcoma, prostate cancer and leukemia. However, little is known about the impact and mechanisms of CaMK2A on lung cancer and cancer redox homeostasis. This study aims to investigate the role of CaMK2A in ROS regulation in lung cancer stem cells. We measured ROS levels using H 2 DCFDA probes in cancer stem cells isolated from two lung cancer cell lines by the known lung cancer stem cell marker CD44 and CD166. Compared to CD44 - and CD166 - non-stem cells, ROS levels were lower in both CD44 + and CD166 + cancer stem cells. In cancer cells with stable CaMK2A knockdown by shRNA (CaMK2A-KD), ROS was suppressed while CaMK2A overexpression led to ROS elevation, suggesting a potential role of CaMK2A on ROS regulation in lung cancer stem cells. To understand the mechanism, activated NRF2 levels, the master transcription factor mediating ROS regulation, were measured in total nuclear protein extracts. By Western blot, nuclear NRF2 protein showed upregulation in CaMK2A overexpression cells and decreased levels in CaMK2A-KD cells, implying NRF2 might be a mediator of CaMK2A in ROS regulation. To further identify downstream regulators, mRNA expression analysis of five ROS-regulatory NRF2 target genes (GSTP1, NQO1, GCLC, HMOX1, TRXR1) was performed. Quantitative PCR results showed significant corresponding changes of some these gene expressions in cell lines, which upregulated in CaMK2A overexpression and downregulated in both CaMK2A-KD cells and CaMK2A pharmacological inhibition cells. Together, the results indicate CaMK2A might regulate ROS status through mediating downstream target genes of NRF2. Citation Format: Siqi Wang, Zhijie Xiao, Vicky Tin, Ian K. Lam, Xuyuan Gao, Maria Wong. The role of CaMK2A in reactive oxygen species homeostasis in lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4787. doi:10.1158/1538-7445.AM2017-4787