title: Suppression of autophagy enhances preferential toxicity of paclitaxel to folliculin-deficient renal cancer cells.
abstract: BACKGROUND
  Paclitaxel, a widely used chemotherapeutic drug, can induce apoptosis in variety of cancer cells. A previous study has shown preferential toxicity of paclitaxel to FLCN-deficient kidney cancer cell line, UOK257. In this report, we investigate the cellular and molecular mechanism of paclitaxel-induced autophagy and apoptosis in renal cancer cells with and without FLCN expression.
  
  
  METHODS
  Two pairs of cell lines were used: FLCN siRNA-silenced ACHN cell line (ACHN-5968) and scrambled ACHN cell line (ACHN-sc); FLCN-null UOK257 cell line and UOK257-2 cell line restored with ectopic expression of FLCN. Autophagy was examined by western blot, GFP-LC3, transmission electron microscopy, and MDC assay. Cell viability and apoptosis were detected using MTT assay, DAPI stain and TUNEL assay. After inhibition of autophagy with 3-Methyladenine (3-MA) or Beclin 1 siRNA, cell viability and apoptosis were measured by MTT assay and TUNEL assay.
  
  
  RESULTS
  After paclitaxel treatment, a dose-dependent decrease in cell viability and increase in apoptosis were observed in FLCN-deficient UOK257 and ACHN-5968 cells compared to their FLCN-expressing counterparts, suggesting that renal cancer cells without FLCN were more sensitive to paclitaxel. Enhanced autophagy was found to be associated with paclitaxel treatment in FLCN-deficient RCC cells. The MAPK pathway was also identified as a key pathway for the activation of autophagy in these kidney cancer cells. Inhibition of phosphorylated ERK with ERK inhibitor U0126 showed a significant decrease in autophagy. Furthermore, after inhibition of autophagy with 3-Methyladenine (3-MA) or Beclin 1 siRNA, apoptosis induced by paclitaxel was significantly increased in FLCN-deficient UOK257 and ACHN-5968 cells.
  
  
  CONCLUSIONS
  Preferential toxicity of paclitaxel to FLCN-deficient kidney cancer cells is associated with enhanced autophagy. Suppression of autophagy further enhances paclitaxel-induced apoptosis in FLCN-deficient renal cancer cells. Our results suggest that paclitaxel combined with an autophagy inhibitor might be a potentially more effective chemotherapeutic approach for FLCN-deficient renal cancer.
authors.forename: Qi
authors.initials: Q
authors.lastname: Zhang
authors.forename: Shuhui
authors.initials: S
authors.lastname: Si
authors.forename: Sue
authors.initials: S
authors.lastname: Schoen
authors.forename: Jindong
authors.initials: J
authors.lastname: Chen
authors.affiliation: Minimally Invasive Urology Center, Provincial Hospital Affiliated to Shandong University, Jinan 250021, China. jsbsdu@gmail.com.
authors.forename: Xun-Bo
authors.initials: XB
authors.lastname: Jin
authors.forename: Guan
authors.initials: G
authors.lastname: Wu
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D001343:Autophagy / Q000187:drug effects* / Q000235:genetics; D000071186:Beclin-1; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004970:Estrone / Q000172:deficiency* / Q000378:metabolism; D055785:Gene Knockdown Techniques; D006801:Humans; D007680:Kidney Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008565:Membrane Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D017239:Paclitaxel / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D051017:Apoptosis Regulatory Proteins; C491997:BECN1 protein, human; D000071186:Beclin-1; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D004970:Estrone; D017239:Paclitaxel