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Oncogenic potential of CYP24A1 in Lung Adenocarcinoma

Research paper by Hiroe Shiratsuchi, Zhuwen Wang, Guoan Chen, Paramita Ray, Jules Lin, Zhuo Zhang, Lili Zhao, David Beer, Dipankar Ray, Nithya Ramnath

Indexed on: 27 Oct '16Published on: 26 Oct '16Published in: Journal of Thoracic Oncology



Abstract

We have previously demonstrated that a subset of lung cancer cells express higher CYP24A1 mRNA, a metabolizing enzyme for 1,25-D3, compared to benign tumors or surrounding normal lung and that high CYP24A1 mRNA expression is associated with poor prognosis in resected lung adenocarcinoma (AC). We hypothesized that CYP24A1 has oncogenic potential and increased CYP24A1 expression may contribute to tumor growth, whereas, CYP24A1 targeting may reduce tumor burden. Two low CYP24A1 expressing human lung cancer cell lines (SK-LU-1 and Calu-6) were stably transfected either with an empty lentiviral vector or with the CYP24A1 expressing vector. Over-expression of mRNA and protein levels of CYP24A1 in SK-LU-1 and Calu-6, confirmed using qRT-PCR and immunoblotting respectively, correlated with accelerated cell growth. Furthermore, significantly higher cell invasion was observed in SK-LU-1 cells overexpressing CYP24A1 compared to control vector-transfected cells. CYP24A1 over-expression also increased RAS protein expression. Next, effects of targeting CYP24A1 were examined in lung cancer cells (A549 and H441), which express higher basal levels of CYP24A1. Knockdown of CYP24A1 using either si- or shRNA reduced CYP24A1 mRNA and protein expression and significantly decreased cell proliferation (30-60%) and reduced mitochondrial DNA content compared to non-targeting (NT) si-/shRNA transfected/transduced cells. Transfection with CYP24A1 siRNA also decreased total RAS protein, thus reducing phosphorylated AKT. Importantly, stable knockdown of CYP24A1 in A549 and H441 lung tumor xenograft models resulted in tumor growth delay and smaller tumor size as evident from tumor bioluminescence and tumor volume measurement studies. Such observations were correlated with decreased tumor cell proliferation as evidenced by reduced Ki67 and Cyclin D staining. Together, our data suggest that CYP24A1 has oncogenic properties mediated by increasing RAS signaling, targeting of which may provide an alternate strategy to treat a subset of lung AC.