title: Oncogenic Potential of CYP24A1 in Lung Adenocarcinoma.
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 were confirmed using qRT-PCR and immunoblotting respectively. Next, effects of targeting CYP24A1 were examined in lung cancer cells (A549 and H441), which express higher basal levels of CYP24A1. Finally, we studied the effects of stable knockdown of CYP24A1 in xenograft models.
  
  
  
  Over-expression of CYP24A1 correlated with accelerated cell growth and invasion compared to control vector-transfected cells. CYP24A1 over-expression also increased RAS protein expression. 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.
  
  
  
  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.
authors: Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan.
authors: Hiroe
authors: H
authors: Shiratsuchi
authors: Section of Thoracic Surgery, Department of Surgery, University of Michigan, Ann Arbor, Michigan.
authors: Zhuwen
authors: Z
authors: Wang
authors: Section of Thoracic Surgery, Department of Surgery, University of Michigan, Ann Arbor, Michigan.
authors: Guoan
authors: G
authors: Chen
authors: Department of Radiation Oncology, University of Michigan, Ann Arbor, Michigan.
authors: Paramita
authors: P
authors: Ray
authors: Section of Thoracic Surgery, Department of Surgery, University of Michigan, Ann Arbor, Michigan.
authors: Jules
authors: J
authors: Lin
authors: Department of Chemical Engineering, University of Michigan, Ann Arbor, Michigan.
authors: Zhuo
authors: Z
authors: Zhang
authors: Department of Biostatistics, University of Michigan, Ann Arbor, Michigan.
authors: Lili
authors: L
authors: Zhao
authors: Section of Thoracic Surgery, Department of Surgery, University of Michigan, Ann Arbor, Michigan.
authors: David
authors: D
authors: Beer
authors: Department of Radiation Oncology, University of Michigan, Ann Arbor, Michigan.
authors: Dipankar
authors: D
authors: Ray
authors: Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan; Ann Arbor Veterans Administration Medical Center, Ann Arbor, Michigan. Electronic address: nithyar@umich.edu.
authors: Nithya
authors: N
authors: Ramnath
mesh_terms: D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D000818:Animals; D017209:Apoptosis; D063646:Carcinogenesis / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D049109:Cell Proliferation; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D034741:RNA, Small Interfering / Q000235:genetics; D047368:Tumor Burden; D014407:Tumor Cells, Cultured; D065668:Vitamin D3 24-Hydroxylase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays
chemical_list: D034741:RNA, Small Interfering; C585549:CYP24A1 protein, human; D065668:Vitamin D3 24-Hydroxylase
keywords: CYP24A1; Lung cancer; Oncogenic