The Identification of Senescence-Specific Genes during the Induction of Senescence in Prostate Cancer Cells

Abstract

Classic mechanisms of tumor response to chemotherapy include apoptosis and mitotic catastrophe. Recent studies have suggested that cellular senescence, a terminal proliferation arrest seen in vitro, may be invoked during the exposure of cancer cells to chemotherapeutic agents. To identify markers associated specifically with the cellular senescence phenotype, we utilized expression data from cDNA microarray experiments identifying transcripts whose expression levels increased as human prostate epithelial cells progressed to senescence. When screened against other growth-inhibitory conditions, including quiescence and apoptosis, many of these transcripts were also upregulated, indicating that similar pathways occur between apoptosis and senescence. A senescent-like phenotype was then induced in several prostate cancer cell lines using 5-aza-2'-deoxycytidine, doxorubicin, or Docetaxel. Treatment with these agents resulted in a significant increase in the induction of senescence-specific genes when compared to nonsenescent conditions. The performance of the panel was improved with fluorescence-activated cell sorting using PKH26 to isolate nonproliferating, viable, drug-treated populations, indicating that a heterogeneous response occurs with chemotherapy. We have defined an RNA-based gene panel that characterizes the senescent phenotype induced in cancer cells by drug treatment. These data also indicate that a panel of genes, rather than one marker, needs to be utilized to identify senescence.

Drug-induced senescence bystander proliferation in prostate cancer cells in vitro and in vivo

“…To monitor the bystander effect of chemically induced senescent cancer cells, GFP ( þ ) cells were cocultured with proliferating or senescent unlabelled cancer cells, collected and analysed by flow cytometry. Both DU145 and LNCaP cells treated with low-dose (25 nM) doxorubicin for 3 days develop a senescent phenotype, increased SAb-gal staining ( Figure 1A), and express previously described senescence marker genes (Schwarze et al, 2005).…”

“…No increase in expression of these genes (IGF2, BRAK, FGF11 and Wnt5a) was seen in the senescent fibroblast lines. Comparing growth-promoting gene expression data from a number of studies involving fibroblasts, epithelial cells and cancer cells (Schwarze et al, 2005;Bavik et al, 2006) reveals little overlap when fibroblasts are compared to other cell lines ( Figure 2B). In sum, our data show that senescent fibroblasts induce the proliferation of bystander cells in vitro significantly more than senescent prostate cancer cells.…”

“…Cell lines and cell culture DU145 and LNCaP prostate cancer cell lines, and human primary fibroblasts, were cultured and senescence induced by treatment with 25 nM doxorubicin as described previously (Schwarze et al, 2005). Polyclonal green fluorescence protein (GFP) ( þ ) cell lines were generated by infecting DU145 and LNCaP cells with pLS-GFP virus and repeated sorting of GFP ( þ ) cells.…”

“…Heterogeneous SAb-gal staining has been observed in sections of frozen human breast tumours after treatment with cyclophosphamide, doxorubicin and 5-fluorouracil (te Poele et al, 2002), and in lung tumours, after carboplatin and taxol (Roberson et al, 2005). Senescence develops at lower drug concentrations than apoptosis, potentially limiting treatment-related side effects (Schwarze et al, 2005).…”

“…Senescent cells express a variety of growth factors and secreted proteins that may stimulate as well as inhibit cell proliferation (Chang et al, 2002;Schwarze et al, 2002Schwarze et al, , 2005Untergasser et al, 2002;Bavik et al, 2006). In contrast to apoptosis, a programme of cellular destruction, senescent cells persist and remain viable.…”