title: A novel in vitro assay of tumor-initiating cells in xenograft prostate tumors.
abstract: BACKGROUND
  The field of prostate cancer has been stymied by the difficulty of cultivating patient-derived samples in the laboratory. In order to help circumvent this challenge, we sought to develop an in vitro assay of human prostate cancer initiation employing a prostate-associated mesenchymal feeder layer.
  
  
  METHODS
  Rat seminal vesicle mesenchyme (rSVM) harvested from male neonatal rats was plated in 12-well plates and then irradiated with 30 Gy after approximately 75% confluence. Single-cell suspensions of two human non-adherent prostate cancer xenograft lines (TRPC and LAPC9) were then plated on irradiated rSVM. At 3-4 weeks, three-dimensional solid structures, termed glandoids, were harvested and analyzed or transplanted singly into the renal capsule of immunodeficient mice. Animals were assessed for tumor formation 8-12 weeks after engraftment. Finally, clonality assays were performed to determine whether glandoids usually arise from a single cell and are therefore clonal in origin.
  
  
  RESULTS
  Glandoids form with reliable frequency (1/ approximately 300 plated cells), are constituted by relevant cell types (CK8+, CK5-, PSA+) and after implantation into immunocompromised mice, give rise to tumors that recapitulate original xenograft histology and cell composition; defining a glandoid as a tumor-initiating unit. In addition, assessment of red fluorescent protein (RFP)-labeled glandoids revealed either all red or non-red structures, with few areas of fusion, suggesting glandoids are clonal in origin.
  
  
  CONCLUSIONS
  The above assay describes an adjunct technique to readily cultivate cells from prostate cancer xenografts in vitro and as such provides a platform on which tumor-initiating cell studies and high-throughput drug discovery may be performed.
authors.affiliation: Department of Urology, University of Rochester School of Medicine, Rochester, New York, USA.
authors.forename: Christopher R
authors.initials: CR
authors.lastname: Silvers
authors.forename: Karin
authors.initials: K
authors.lastname: Williams
authors.forename: Linda
authors.initials: L
authors.lastname: Salamone
authors.forename: Jiaoti
authors.initials: J
authors.lastname: Huang
authors.forename: Craig T
authors.initials: CT
authors.lastname: Jordan
authors.forename: Haijun
authors.initials: H
authors.lastname: Zhou
authors.forename: Ganesh S
authors.initials: GS
authors.lastname: Palapattu
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D006801:Humans; D008297:Male; D051379:Mice; D009368:Neoplasm Transplantation; D011467:Prostate / Q000473:pathology*; D051381:Rats; D012669:Seminal Vesicles / Q000473:pathology*; D014183:Transplantation, Heterologous; D014407:Tumor Cells, Cultured