Maspin sensitizes prostate cancer cells to doxazosin-induced apoptosis

Abstract

Maspin is a mammary serine protease inhibitor or serpin with tumor suppressive and antiangiogenic activity that inhibits tumor motility, invasion and metastasis, at least by its actions on cell membrane and extracellular matrix (ECM) proteins. Previous studies documented that the quinazoline-derived a1-adrenoceptor antagonist doxazosin affects the attachment and migration of prostate cancer cells. In this study, we investigated the effect of maspin overexpression on the apoptotic/antiadhesion response of prostate cancer cells to doxazosin. The response of maspin-overexpressing clones of human prostate cancer cells DU-145 to doxazosin was evaluated by determining cell viability, apoptosis and cell proliferation on the basis of the trypan blue exclusion assay/methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay, Hoechst staining and caspase-3 activation, and [ 3 H]thymidine incorporation assay. Vascular endothelial growth factor (VEGF), transforming growth factor bRII (TGFbRII), Smad4 (a TGFb intracellular effector) and bax expression was evaluated at the mRNA and protein level using reverse transcriptase-polymerase chain reaction and Western blotting, respectively. The effect of doxazosin on cell attachment of maspin-expressing prostate cancer cells was evaluated on collagen-and fibronectin-coated plates. Cell migration was assessed using the wounding assay. In response to tumor necrosis factor-related apoptosisinducing ligand, DU-145-maspin expressing cells undergo apoptosis, via poly(ADP-ribose) polymerasecleavage and caspase-3 activation. DU-145-maspin cells exhibited higher sensitivity to doxazosin and an earlier temporal activation of caspase-3. The number of apoptotic cells detected in response to doxazosin was significantly higher compared to the neo control (Po0.0001). Doxazosin resulted in dramatic downregulation of the 189 isoform of VEGF in maspin transfectants, while a fivefold induction of Smad4 mRNA expression was detected in those cells after 24 h of treatment. Maspin overexpression in prostate cancer cells resulted in an increased ability to attach to ECM-coated plates, and doxazosin treatment considerably antagonized this effect by decreasing the attachment potential to collagen and fibronectin. The present study supports the ability of maspin to enhance the apoptotic threshold of prostate cancer cells to the quinazoline-based a1-adrenoceptor antagonist doxazosin. These findings may have therapeutic significance in the development of antiangiogenic targeting by doxazosin and derivative agents for advanced prostate cancer.

Maspin Expression and its Metastasis Suppressing Function in Prostate Cancer

“…In contrast, siRNA knockdown of maspin in prostate cancer PC-3 cells increased the basal ROS level. Tahmatzopoulos et al have shown that treatment of human prostate cancer DU145 cells with H 2 O 2 (or PMA) but not with TRAIL further increases the maspin/GST interaction and significantly attenuated H 2 O 2 -induced ROS generation and VEGF expression [33]. This study further demonstrates that maspin transfected tumor cells produced less VEGF than the transfection control cells.…”

Tumor suppressive maspin and epithelial homeostasis

“…Oxidative stress, that is, treatment of with H 2 O 2 (or PMA) but not with TRAIL, further increases the maspin/GST interaction in DU145 cells, and significantly attenuated H 2 O 2 -induced ROS generation and VEGF expression. This evidence is further supported by the evidence that maspin transfected tumor cells produced less VEGF than the transfection control cells when treated with doxazosin [Tahmatzopoulos et al, 2005]. Interestingly, a single point mutation at the RSL p 1 position of maspin (Mas R340A ) greatly reduced the affinity for GST.…”

“…Later, we found that maspin expression sensitized tumor cells to a series of apoptotic stimuli, ranging from death ligands (TRAIL and TNF-a) to brefeldin-induced endoplasmic reticulum stress [Jiang et al, 2002;Liu et al, 2004a]. We also showed that maspin also sensitized DU145 cells to apoptosis induced by doxazosin, an a1-adrenoceptor antagonist in clinical trials for prostate cancer [Tahmatzopoulos et al, 2005].…”

Maspin Expression in Prostate Tumor Cells Averts Stemness and Stratifies Drug Sensitivity

“…The interplay between maspin and the pathways targeted by MS275 and Salinomycin also requires further investigation. Maspin has been shown to sensitize cells to drug induced apoptosis (26-28). Consistently, maspin sensitized the stem cell subpopulations to MS275 and Salinomycin.…”

“…Ectopic expression of maspin in prostate tumor cells was sufficient to drive the full spectrum of progressive changes leading to acini formation in 3-dimensional collagen I (17), and in a xenograft model for prostate tumor bone metastasis (18). It is important to note that maspin also enhances the sensitivity of tumor cells to apoptosis-inducing drugs (26-28). Consistently, clinical evidence demonstrated the correlation of maspin with better differentiated phenotypes and better prognosis (29).…”