title: Reduced 64Cu uptake and tumor growth inhibition by knockdown of human copper transporter 1 in xenograft mouse model of prostate cancer.
abstract: Copper is an element required for cell proliferation and angiogenesis. Human prostate cancer xenografts with increased (64)Cu radioactivity were visualized previously by PET using (64)CuCl2 as a radiotracer ((64)CuCl2 PET). This study aimed to determine whether the increased tumor (64)Cu radioactivity was due to increased cellular uptake of (64)Cu mediated by human copper transporter 1 (hCtr1) or simply due to nonspecific binding of ionic (64)CuCl2 to tumor tissue. In addition, the functional role of hCtr1 in proliferation of prostate cancer cells and tumor growth was also assessed.
  
  
  METHODS
  A lentiviral vector encoding short-hairpin RNA specific for hCtr1 (Lenti-hCtr1-shRNA) was constructed for RNA interference-mediated knockdown of hCtr1 expression in prostate cancer cells. The degree of hCtr1 knockdown was determined by Western blot, and the effect of hCtr1 knockdown on copper uptake and proliferation were examined in vitro by cellular (64)Cu uptake and cell proliferation assays. The effects of hCtr1 knockdown on tumor uptake of (64)Cu were determined by PET quantification and tissue radioactivity assay. The effects of hCtr1 knockdown on tumor growth were assessed by PET/CT and tumor size measurement with a caliper.
  
  
  RESULTS
  RNA interference-mediated knockdown of hCtr1 was associated with the reduced cellular uptake of (64)Cu and the suppression of prostate cancer cell proliferation in vitro. At 24 h after intravenous injection of the tracer (64)CuCl2, the (64)Cu uptake by the tumors with knockdown of hCtr1 (4.02 ± 0.31 percentage injected dose per gram [%ID/g] in Lenti-hCtr1-shRNA-PC-3 and 2.30 ± 0.59 %ID/g in Lenti-hCtr1-shRNA-DU-145) was significantly lower than the (64)Cu uptake by the control tumors without knockdown of hCtr1 (7.21 ± 1.48 %ID/g in Lenti-SCR-shRNA-PC-3 and 5.57 ± 1.20 %ID/g in Lenti-SCR-shRNA-DU-145, P < 0.001) by PET quantification. Moreover, the volumes of prostate cancer xenograft tumors with knockdown of hCtr1 (179 ± 111 mm(3) for Lenti-hCtr1-shRNA-PC-3 or 39 ± 22 mm(3) for Lenti-hCtr1-shRNA-DU-145) were significantly smaller than those without knockdown of hCtr1 (536 ± 191 mm(3) for Lenti- SCR-shRNA-PC-3 or 208 ± 104 mm(3) for Lenti-SCR-shRNA-DU-145, P < 0.01).
  
  
  CONCLUSIONS
  Overall, data indicated that hCtr1 is a promising theranostic target, which can be further developed for metabolic imaging of prostate cancer using (64)CuCl2 PET/CT and personalized cancer therapy targeting copper metabolism.
authors.affiliation: Department of Radiology, University of Texas Southwestern Medical Center, Dallas, Texas.
authors.forename: Huawei
authors.initials: H
authors.lastname: Cai
authors.forename: Jiu-sheng
authors.initials: JS
authors.lastname: Wu
authors.forename: Otto
authors.initials: O
authors.lastname: Muzik
authors.forename: Jer-Tsong
authors.initials: JT
authors.lastname: Hsieh
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Lee
authors.forename: Fangyu
authors.initials: F
authors.lastname: Peng
mesh_terms: D000818:Animals; D015153:Blotting, Western; D027682:Cation Transport Proteins / Q000235:genetics / Q000627:therapeutic use*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003300:Copper / Q000378:metabolism*; D003301:Copper Radioisotopes; D000079923:Copper Transporter 1; D005822:Genetic Vectors; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008297:Male; D051379:Mice; D049268:Positron-Emission Tomography; D011471:Prostatic Neoplasms / Q000000981:diagnostic imaging / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D019275:Radiopharmaceuticals; D014018:Tissue Distribution; D023041:Xenograft Model Antitumor Assays
chemical_list: D027682:Cation Transport Proteins; D003301:Copper Radioisotopes; D000079923:Copper Transporter 1; D034741:RNA, Small Interfering; D019275:Radiopharmaceuticals; D003300:Copper; C029892:cupric chloride
keywords: 64Cu-chloride; PET/CT; copper metabolism; human copper transporter 1; prostate cancer