title: Release of monocyte migration signals by breast cancer cell lines after ablative and fractionated γ-irradiation.
abstract: BACKGROUND
  Radiotherapy, administered in fractionated as well as ablative settings, is an essential treatment component for breast cancer. Besides the direct tumor cell death inducing effects, there is growing evidence that immune mechanisms contribute - at least in part - to its therapeutic success. The present study was designed to characterize the type and the extent of cell death induced by fractionated and ablative radiotherapy as well as its impact on the release of monocyte migration stimulating factors by dying breast cancer cells.
  
  
  METHODS
  Cell death and senescence assays were employed to characterize the response of a panel of breast cancer cell lines with different receptor and p53 status towards γ-irradiation applied in a fractionated (daily doses of 2 Gy) or ablative setting (single dose of 20 Gy). Cell-free culture supernatants were examined for their monocyte migration stimulating potential in transwell migration and 2D chemotaxis/chemokinesis assays. Irradiation-induced transcriptional responses were analyzed by qRT-PCR, and CD39 surface expression was measured by flow cytometry.
  
  
  RESULTS
  Fast proliferating, hormone receptor negative breast cancer cell lines with defective p53 predominantly underwent primary necrosis in response to γ-irradiation when applied at a single, ablative dose of 20 Gy, whereas hormone receptor positive, p53 wildtype cells revealed a combination of apoptosis, primary, and secondary (post-apoptotic) necrosis. During necrosis the dying tumor cells released apyrase-sensitive nucleotides, which effectively stimulated monocyte migration and chemokinesis. In hormone receptor positive cells with functional p53 this was hampered by irradiation-induced surface expression of the ectonucleotidase CD39.
  
  
  CONCLUSIONS
  Our study shows that ablative radiotherapy potently induces necrosis in fast proliferating, hormone receptor negative breast cancer cell lines with mutant p53, which in turn release monocyte migration and chemokinesis stimulating nucleotides. Future studies have to elucidate, whether these mechanisms might be utilized in order to stimulate intra-tumoral monocyte recruitment and subsequent priming of adaptive anti-tumor immune responses, and which breast cancer subtypes might be best suited for such approaches.
authors.forename: Roman
authors.initials: R
authors.lastname: Hennel
authors.forename: Nikko
authors.initials: N
authors.lastname: Brix
authors.forename: Karin
authors.initials: K
authors.lastname: Seidl
authors.forename: Anne
authors.initials: A
authors.lastname: Ernst
authors.forename: Heike
authors.initials: H
authors.lastname: Scheithauer
authors.forename: Claus
authors.initials: C
authors.lastname: Belka
authors.affiliation: Department of Radiation Oncology, Ludwig-Maximilians-University, Munich, Germany. kirsten.lauber@med.uni-muenchen.de.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Lauber
mesh_terms: D015703:Antigens, CD / Q000378:metabolism; D017209:Apoptosis; D001081:Apyrase / Q000378:metabolism; D001943:Breast Neoplasms / Q000378:metabolism* / Q000532:radiotherapy*; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D002469:Cell Separation; D004252:DNA Mutational Analysis; D005260:Female; D005434:Flow Cytometry; D005720:Gamma Rays; D015972:Gene Expression Regulation, Neoplastic / Q000528:radiation effects; D006801:Humans; D009000:Monocytes / Q000166:cytology*; D009154:Mutation; D009336:Necrosis; D009711:Nucleotides / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics; D014965:X-Rays
chemical_list: D015703:Antigens, CD; D009711:Nucleotides; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D001081:Apyrase; C067869:CD39 antigen