title: A role for the copper transporter Ctr1 in the synergistic interaction between hyperthermia and cisplatin treatment.
abstract: OBJECTIVE
  Hyperthermia enhances cytotoxic effects of chemotherapeutic agents such as cisplatin. However, the underlying molecular mechanisms remain unclear. We hypothesised that hyperthermia increases cisplatin accumulation and efficacy by modulating function of copper transport protein 1 (Ctr1), a major regulator of cellular cisplatin uptake. We examined the significance of Ctr1 in the synergistic interaction between hyperthermia and cisplatin. We assessed the importance of cisplatin- and hyperthermia-induced Ctr1 multimerisation in sensitising cells to cisplatin cytotoxicity.
  
  
  METHODS
  Ctr1 protein levels and cisplatin sensitivities were assessed in bladder cancer cell lines with immunoblotting and clonogenic survival assays. Using Myc-tagged-Ctr1 HEK293 cells, we assessed the effect of hyperthermia on Ctr1 multimerisation with immunoblotting. The effect of hyperthermia on cisplatin sensitivity and accumulation was assessed in wild-type (WT) and Ctr1 knockout (Ctr1-/-) mouse embryonic fibroblasts (MEFs) with clonogenic assays and inductively coupled plasma-mass spectrometry (ICP-MS).
  
  
  RESULTS
  Increased Ctr1 protein expression was observed for the most cisplatin-sensitive bladder cancer cell lines and MEFs. Heat-induced increase in Ctr1 multimerisation with cisplatin was observed in Myc-tagged Ctr1 cells. Hyperthermia enhanced cisplatin-mediated cytotoxicity in WT more than Ctr1-/- cells (dose modifying factors 1.75 versus 1.4, respectively). WT cells accumulated more platinum versus Ctr1-/- cells; this was further increased by hyperthermia in WT cells.
  
  
  CONCLUSIONS
  Hyperthermia enhanced cisplatin uptake and cytotoxicity in WT cells. Heat increased Ctr1 activity by increasing multimerisation, enhancing drug cytotoxicity. Furthermore, Ctr1 protein profiles of bladder tumours, as well as other tumour types, may predict their response to cisplatin and overall efficacy of treatment.
authors.affiliation: Department of Pathology, Duke University, Durham, NC, USA.
authors.forename: Chelsea D
authors.initials: CD
authors.lastname: Landon
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Benjamin
authors.forename: Kathleen A
authors.initials: KA
authors.lastname: Ashcraft
authors.forename: Mark W
authors.initials: MW
authors.lastname: Dewhirst
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage*; D027682:Cation Transport Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002945:Cisplatin / Q000008:administration & dosage*; D000079923:Copper Transporter 1; D005347:Fibroblasts / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D006979:Hyperthermia, Induced; D051379:Mice; D012333:RNA, Messenger / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000378:metabolism* / Q000628:therapy
chemical_list: D000970:Antineoplastic Agents; D027682:Cation Transport Proteins; D000079923:Copper Transporter 1; D012333:RNA, Messenger; D002945:Cisplatin