Wengang Cao

Dichloroacetate (DCA) sensitizes both wild-type and over expressing Bcl-2 prostate cancer cells in vitro to radiation.

Bcl‐2 protects cells from apoptosis and provides a survival advantage to cells over‐expressing this oncogene. In addition, over expression of Bcl‐2 renders cell resistant to radiation therapy. Recently, dichloroacetate (DCA) was proven to potentiate the apoptotic machinery by interacting with Bcl‐2. In this study, we investigated whether treating human prostate cancer cells with DCA could modulate Bcl‐2 expression and if the modulation in Bcl‐2 expression could render the Bcl‐2 over expressing cells more susceptible to cytotoxicity effects of radiation.

Effects of CXCR4 antagonist CTCE-9908 on prostate tumor growth†

Recent reports have linked the survival‐promoting effect of CXCR4 to the up regulation of Bcl‐2 protein expression.

Design of a carbonic anhydrase IX active-site mimic to screen inhibitors for possible anticancer properties

Recently, a convincing body of evidence has accumulated suggesting that the overexpression of carbonic anhydrase isozyme IX (CA IX) in some cancers contributes to the acidification of the extracellular matrix, which in turn promotes the growth and metastasis of the tumor. These observations have made CA IX an attractive drug target for the selective treatment of certain cancers. Currently, there is no available X-ray crystal structure of CA IX, and this lack of availability has hampered the rational design of selective CA IX inhibitors. In light of these observations and on the basis of structural alignment homology, using the crystal structure of carbonic anhydrase II (CA II) and the sequence of CA IX, a double mutant of CA II with Ala65 replaced by Ser and Asn67 replaced by Gln has been constructed to resemble the active site of CA IX. This CA IX mimic has been characterized kinetically using (18)O-exchange and structurally using X-ray crystallography, alone and in complex with five CA sulfonamide-based inhibitors (acetazolamide, benzolamide, chlorzolamide, ethoxzolamide, and methazolamide), and compared to CA II. This structural information has been evaluated by both inhibition studies and in vitro cytotoxicity assays and shows a correlated structure-activity relationship. Kinetic and structural studies of CA II and CA IX mimic reveal chlorzolamide to be a more potent inhibitor of CA IX, inducing an active-site conformational change upon binding. Additionally, chlorzolamide appears to be cytotoxic to prostate cancer cells. This preliminary study demonstrates that the CA IX mimic may provide a useful model to design more isozyme-specific CA IX inhibitors, which may lead to development of new therapeutic treatments of some cancers.

Docetaxel and bortezomib downregulate Bcl-2 and sensitize PC-3-Bcl-2 expressing prostate cancer cells to irradiation

BackgroundCurrently, docetaxel is used to treat hormone-refractory metastatic prostate cancer. Docetaxel not only inhibits microtubule formation but can also downregulate expression of Bcl-2, a known antiapoptotic oncogene. Furthermore, the 26S proteasome inhibitor bortezomib can downregulate Bcl-2 expression. Previously, we demonstrated that overexpression of Bcl-2 renders cells resistant to radiation therapy. In this study, we investigated whether treating human prostate cancer cells with docetaxel, bortezomib, or both could modulate Bcl-2 expression and whether such modulation could render Bcl-2-overexpressing cells more susceptible to radiation.MethodsPC-3-Bcl-2 and PC-3-Neo human prostate cancer cells treated with docetaxel and/or bortezomib in addition to irradiation were analyzed in vitro for proliferation, clonogenic survival, cell cycle phase distribution, and expression of Bcl-2 and Bcl-xL proteins.ResultsDocetaxel and bortezomib alone had significant cytotoxic effects. In addition, docetaxel, bortezomib, or radiation resulted in a G2M phase arrest in PC-3-Bcl-2, whereas only docetaxel or radiation did so in PC-3-Neo cells. Both cell lines were more sensitized to radiation’s killing effects when treated with the combination of docetaxel and bortezomib than when treated with either agent alone. Furthermore, docetaxel and bortezomib-treated cells exhibited marked changes in the expression of Bcl-2 and Bcl-xL.ConclusionsThis is the first study to demonstrate that docetaxel and bortezomib in combination can effectively sensitize Bcl-2-overexpressing human prostate cancer cells to radiation effects by modulating the expression of key members of the Bcl-2 family. Together, these findings warrant further evaluation of the combination of docetaxel and bortezomib in prostate cancer.

Dual targeting of Bcl-2 and VEGF: A potential strategy to improve therapy for prostate cancer

We previously demonstrated that Bcl-2 overexpression stimulates angiogenesis in PC-3 human prostate cancer cells, thus giving these tumors a growth advantage. To further elucidate the relationship between Bcl-2 and vascular endothelial growth factor (VEGF) in PC-3-Bcl-2 cells, tumorigenicity and angiogenesis were evaluated in our in vitro and in vivo model treated with antisense Bcl-2 oligodeoxynucleotide (ASO) and bevacizumab. In vitro and in vivo angiogenesis assays, as well as a xenograft tumor model of the human prostate cancer cell line PC-3-Bcl-2, were subjected to ASO alone, bevacizumab alone, or the combination of ASO and bevacizumab. Protein-based assays (e.g., immunohistochemical staining and enzyme-linked immunosorbent assay [ELISA]) were utilized to detect molecular changes. Interestingly, targeting Bcl-2 with ASO resulted in the inhibition of in vitro tube formation and inhibition of angiogenesis in Matrigel plugs similar to treatment with bevacizumab. In our PC-3-Bcl-2 xenograft model, ASO alone resulted in 41% reduction in tumor size, bevacizumab alone resulted in a 50% reduction in tumor size, whereas the combination of ASO with bevacizumab was associated with >95% reduction in tumor volume. Reduction in tumor size in all groups was associated with reduction in Bcl-2 and VEGF expression, induction of apoptosis, and inhibition of angiogenesis and its associated chemokine production. These findings confirm that Bcl-2 is a pivotal target for cancer therapy and thus, further study of this novel combination of Bcl-2 reduction and angiogenic targeting in human tumors is warranted.

Bcl-2 Mediated Modulation of Vascularization in Prostate Cancer Xenografts

We previously demonstrated that Bcl‐2 overexpression enhances the radiation resistance of PC‐3 human prostate cancer cells and xenografts by inhibiting apoptosis, increasing proliferation, and promoting angiogenesis. To further elucidate the relationship between Bcl‐2 expression and the angiogenic potential of PC‐3‐Bcl‐2 cells, tumorigenicity, angiogenesis, and lymphangiogenesis were evaluated and compared in a Bcl‐2 overexpressing clone in vitro and in vivo.

VEGF induces expression of Bcl-2 and multiple signaling factors in microvascular endothelial cells in a prostate cancer model.

PurposeWe have previously demonstrated that prostate tumors that highly express Bcl-2 are not only more tumorigenic, but also more angiogenic than low Bcl-2 expressing tumors. Observed increased rates of angiogenesis are likely due to the secretion of multiple factors from the tumor cells.Experimental designHuman endothelial cells were subjected to exogenous VEGF or conditioned media from PC-3 cells and assayed by several in vitro systems to better characterize the effects of tumor microenvironment on endothelial cells.ResultsVEGF stimulation increased Bcl-2 expression in human microvascular endothelial cells (HMVECs), at least partially through stabilization of Bcl-2 mRNA transcripts, and protected these cells from apoptosis. These effects were mimicked by treatment of HMVECs with conditioned media from cultured PC-3 prostate tumor cells manipulated to overexpress Bcl-2. Through the use of kinase inhibitors and molecular profiling, several distinct pathways were implicated in the regulation of Bcl-2 in HMVECs, including those involving PI3K/AKT, PKC, mTOR, STAT-1, and IL-8, factors associated with tumor survival and growth.ConclusionsThis study identifies molecular elements of a link between Bcl-2 expression in distinct cell types within a tumor and reaffirms that strategies designed to target Bcl-2 are desirable as they might enhance treatment response through dual effects.

BCL-2 MEDIATED MODULATION OF VASCULARIZATION IN PROSTATE CANCER XENOGRAFTS

PURPOSE. We previously demonstrated that Bcl-2 overexpression enhances the radiation resistance of PC-3 human prostate cancer cells and xenografts by inhibiting apoptosis, increasing proliferation, and promoting angiogenesis. To further elucidate the relationship between Bcl-2 expression and the angiogenic potential of PC-3-Bcl-2 cells, tumorigenicity, angiogenesis, and lymphangiogenesis were evaluated and compared in a Bcl-2 overexpressing clone in vitro and in vivo. EXPERIMENTAL DESIGN. Human prostate cancer cells over expressing Bcl-2 were studied in vitro and in vivo to determine the angiogenic and lymphangiogenic properties of these cells. RESULTS. Increased Bcl-2 expression enhanced the tumorigenicity of prostate cancer xenografts. It also enhanced the expression and secretion of key angiogenic and lymphangiogenic factors that stimulated the synthesis of CD31-positive blood vessels and LYVE-1 positive lymphatics. Specifically, the increased angiogenic and lymphangiogenic potential correlated with increased serum levels of basic fibroblast growth factor (bFGF), interleukin 8 (CXCL8), and matrix metalloproteinase (MMP 9). In vitro analysis demonstrated that Bcl-2 expressing tumor cells secreted bFGF and vascular endothelial growth factor (VEGF) into culture supernatants. Microarray analysis of Bcl-2 expressing PC-3 cells demonstrated increased transcription of genes involved in metabolism, such as interleukins, growth factors, tumor necrosis factors (TNF) family members, and peptidases. CONCLUSIONS. Together, these results demonstrate that Bcl-2 can regulate tumoral angiogenesis and lymphangiogenesis and suggest that therapy targeted at Bcl-2 expression, angiogenesis, and lymphangiogenesis may synergistically modulate tumor growth and confirm that Bcl-2 is a pivotal target for cancer therapy. Prostate 69: 459–470, 2009. # 2008 Wiley-Liss, Inc.