Dalin He

Genome-wide association study in Chinese men identifies two new prostate cancer risk loci at 9q31.2 and 19q13.4

Prostate cancer risk–associated variants have been reported in populations of European descent, African-Americans and Japanese using genome-wide association studies (GWAS). To systematically investigate prostate cancer risk–associated variants in Chinese men, we performed the first GWAS in Han Chinese. In addition to confirming several associations reported in other ancestry groups, this study identified two new risk-associated loci for prostate cancer on chromosomes 9q31.2 (rs817826, P = 5.45 × 10−14) and 19q13.4 (rs103294, P = 5.34 × 10−16) in 4,484 prostate cancer cases and 8,934 controls. The rs103294 marker at 19q13.4 is in strong linkage equilibrium with a 6.7-kb germline deletion that removes the first six of seven exons in LILRA3, a gene regulating inflammatory response, and was significantly associated with the mRNA expression of LILRA3 in T cells (P < 1 × 10−4). These findings may advance the understanding of genetic susceptibility to prostate cancer.

Role of Wnt/β‐catenin signaling pathway in epithelial‐mesenchymal transition of human prostate cancer induced by hypoxia‐inducible factor‐1α

Objectives:  Epithelial‐mesenchymal transition (EMT) is an important process in tumor development, and several studies suggest that the Wnt/β‐catenin signal pathway may play an important role in EMT. However, there is no direct evidence showing that the Wnt/β‐catenin pathway actually determines the EMT induced by an exogenous signal. Our previous research has successfully proved that overexpression of hypoxia‐inducible factor‐1α (HIF‐1α) could induce EMT in LNCaP cells, but not in PC‐3. The present study aims to determine whether the signal of HIF‐1α for inducing prostate cancer cells to undergo EMT might possibly pass through the Wnt/β‐catenin pathway.

Silibinin inhibits prostate cancer invasion, motility and migration by suppressing vimentin and MMP-2 expression.

AbstractAim:Silibinin is known to exert growth inhibition and cell death together with cell cycle arrest and apoptosis in human prostate cancer cells. Whether silibinin could inhibit the invasion, motility and migration of prostate cancer cells remains largely unknown. This study was designed to evaluate this efficacy and possible mechanisms using a novel highly bone metastatic ARCaPM cell model.Methods:Four prostate cancer cell lines, LNCaP, PC-3, DU145, and ARCaPM, were used in this study. These cells were treated with increasing concentrations of silibinin (50, 100, and 200 μmol/L) for different periods of time. After treatment, cell viabilities of four prostate cancer cells were compared by MTT assay. Alterations of ARCaPM cell invasion, motility and migration were assessed by cell invasion, motility and wound healing assays. The changes of vimentin expression were observed by Western blotting and immunofluorescence staining, and the expression of MMP-2, MMP-9, and uPA was analyzed by reverse transcription-polymerase chain reaction (RT-PCR).Results:ARCaPM cells showed less sensitivity to the growth inhibition of pharmacological doses of silibinin than LNCaP, PC-3, and DU145 cells. However, silibinin exerted significant dose- and time-dependent inhibitory effects on the invasion, motility and migration of ARCaPM cells. Furthermore, the expression of vimentin and MMP-2, but not MMP-9 or uPA, was down-regulated in a dose- and time-dependent manner after treatment of silibinin.Conclusion:This study shows that silibinin could inhibit the invasion, motility and migration of ARCaPM cells via down-regulation of vimentin and MMP-2 and therefore may be a promising agent against prostate cancer bone metastasis.

Silibinin inhibits cell growth and induces apoptosis by caspase activation, down-regulating survivin and blocking EGFR-ERK activation in renal cell carcinoma.

Silibinin as an effective anti-cancer and chemopreventive agent in various epithelial cancer models has been reported inhibition of cancer cell growth through mitogenic signaling pathways. However, whether it could inhibit renal cell carcinoma growth and what are the underlying mechanisms is still not well elucidated. Since EGFR-MAPK and apoptosis pathways play important roles in renal cell carcinoma survival. Here, for the first time we evaluated the inhibitory proliferation effects of silibinin in renal cell carcinoma growth and examined whether silibinin modulates EGFR-MAPK and tumor apoptosis cascades signals. Our results indicated that silibinin effectively inhibits the renal cancer carcinoma Caki-1 cell proliferation and induces apoptosis through inhibiting the activation of EGFR and ERK and the expression of survivin, up-regulating the expression of p53 and triggering the cascades of caspase pathways. Our results suggested silibinin might be as one of the candidate chemopreventive agents for renal cell carcinoma therapy.

Knockdown of β-Catenin Through shRNA Cause a Reversal of EMT and Metastatic Phenotypes Induced by HIF-1α

Obective: Wnt/β-catenin signaling pathway regulates pattern formation during embryogenesis as well as tumor progression. Numbers of studies suggest that this signaling pathway may play an important role in Epithelial-Mesenchymal transition (EMT), however, there was no evidence that Wnt/β-catenin signaling pathway directly controlled the EMT occurrence. Our previous research has successfully proved that overexpression of hypoxia inducible factor-1α (HIF-1α) could induce EMT in LNCaP cells, but not in PC-3. Consistently, the expression of β-catenin protein increased in LNCaP/HIF-1α cells, but not in PC-3/HIF-1α. This study mainly aimed at exploring the essentiality and importance of Wnt/β-catenin signaling pathway in HIF-1α-induced EMT. Methods: Human prostate cancer cells (LNCaP) were stably transfected by recombinant plasmid pcDNA3.1(-)/HIF-1α. The positive clones were selected by G418 and confirmed through western blot analysis, reverse transcription-polymerase chain reaction (RT-PCR), and indirect immunofluoesence. Then LNCaP/HIF-1α was transiently transfected with β-catenin shRNA (shRNA1 and shRNA2) and negative shRNA (shRNA-scr). The epithelial markers, mesenchymal markers, and critical proteins in Wnt/β-catenin signaling pathway were separately detected by western blot analysis. Finally, the invasive potency of cells in different transfection group was examined by Matrgel transwell assay. Result: We successfully established prostate cancer cell line LNCaP/HIF-1α and LNCaP/HIF-1α/β-catenin(-). LNCaP/HIF-1α displayed high expression of mesenchymal markers and low expression of epithelial markers. However, compared with LNCaP/HIF-1α, the epithelial marker E-cadherin was increased in LNCaP/HIF-1α/β-catenin(-), whereas the expression of mesenchymal marker N-cadherin, vimentin, MMP-2 were significantly decreased. Inhibition of Wnt signal activity through β-catenin shRNA cause a reversal of EMT induced by HIF-1α in human prostate cancer. Conclusion: Overexpression of HIF-1α stimulates the invasion potency of human prostate carcinoma cells through EMT pathway and Wnt/β-catenin signaling pathway played a vital role in this process. Wnt/β-catenin signaling pathway might be a necessary endogenous signal that directly controlled the EMT occurrence induced by HIF-1α.

Targeting silibinin in the antiproliferative pathway.

Importance of the field: Due to the failure and severe toxicity of cancer chemotherapy, silibinin, a natural flavonoid from the seeds of milk thistle, has recently received more attention for its potential anticancer and nontoxic roles in animals and humans. Silibinin has clearly demonstrated inhibition of multiple cancer cell signaling pathways, including growth inhibition, inhibition of angiogenesis, chemosensitization, and inhibition of invasion and metastasis. Cumulative evidence implicates that silibinin is a potential agent for cancer chemoprevention and chemotherapy. Areas covered in this review: Our aim is to discuss the recent progress of silibinin in regulating multiple anticancer proliferative signaling pathways; the review covers literature mainly from the past 3 – 5 years. What the reader will gain: One of the strategies for tumor therapy is eradication of cancer cells through targeting specific cell-proliferative pathways. This review highlights the current knowledge of silibinin in regulating multiple cellular proliferative pathways in cancer cells, including receptor tyrosine kinases, androgen receptor, STATs, NF-κB, cell cycle regulatory and apoptotic signaling pathways. Take home message: The molecular mechanisms of silibinin-mediated antiproliferative effects are mainly via receptor tyrosine kinases, androgen receptor, STATs, NF-κB, cell cycle regulatory and apoptotic signaling pathways in various cancer cells. Targeting inhibition of proliferative pathways through silibinin treatment may provide a new approach for improving chemopreventive and chemotherapeutic effects.

Tumorspheres derived from prostate cancer cells possess chemoresistant and cancer stem cell properties

PurposeProstate cancer (PCa) becomes lethal when cancer cells develop into castration-resistant PCa, which remains incurable because of the poor understanding of their cell origin and characteristics. We aim to investigate the potential role of cancer stem cells (CSCs) in PCa progression.MethodsHuman PCa cell lines (LNCaP, 22RV1, DU145 and PC-3) were plated in serum-free suspension culture system allowed for tumorsphere forming. To evaluate the CSC characteristics of tumorspheres, the self-renewal, chemoresistance, tumorigenicity of the PCa tumorsphere cells, and the expression levels of stemness-related proteins in the PCa tumorsphere cells were assessed, comparing with the parental adherent cells.ResultsTumorsphere cells from PCa cell lines displayed enhanced self-renewal, chemoresistance and tumor-initiating capacity when compared with the adherent cells. Additionally, these cells overexpressed CSC marker CD44. Also, the tumorsphere cells expressed high levels of “stemness” genes Gli1, ABCG2 and Bmi-1.ConclusionsCollectively, these data demonstrated that tumorspheres derived from PCa cells possess chemoresistant and CSC properties. Our study suggests that the identification of PCa CSCs could provide new insight into the lethal phenotype of PCa and therapeutic implications.

Tetrandrine induces apoptosis and triggers caspase cascade in human bladder cancer cells.

BACKGROUND Tetrandrine is known to exert anti-tumor effects, however, little is known about its effect on human bladder carcinoma. In this study, employing two different human bladder cancer cell lines, 5637 and T24, which represent high-risk superficial bladder cancer (5637) and high-grade bladder cancer (T24), we tested tetrandrine-induced apoptosis and growth inhibition in bladder carcinoma cell lines and investigated the possible mechanisms. MATERIALS AND METHODS Growth inhibition and apoptosis induction was determined by MTT assay and flow cytometry analysis, respectively. Activation of caspases were analyzed by Western blotting and caspase colorimetric assay. The collapse of mitochondrial membrane potential (ΔΨ(m)) and subcellular distribution of cytochrome c was determined by JC-1 staining and Western blotting, respectively. RESULTS Tetrandrine treatment showed strong growth inhibitory and apoptotic effects on human bladder cancer 5637 and T24 cells in a concentration-dependent manner. Additionally, induction of apoptosis by tetrandrine was associated with a very strong and prominent caspase-9, caspase-8, and caspase-3 activation as well as PARP cleavage. Flow cytometric studies revealed that tetrandrine induced a dose-dependent loss of ΔΨ(m), which was accompanied by the release of cytochrome c from mitochondria to the cytosol. CONCLUSION Taken together, this study provided the first evidence that tetrandrine imparted inhibitory and apoptotic activity in human bladder cancer cells. The tetrandrine-induced apoptosis might be related to the activation of the caspase cascade and mitochondrial pathway. Our results suggest that tetrandrine merits further in vivo investigation as a novel bladder cancer chemopreventive and chemotherapeutic agent in the clinical setting.

A novel anti-cancer effect of genistein: reversal of epithelial mesenchymal transition in prostate cancer cells

AbstractAim:The aim of the present study was to investigate whether low dose genistein affects the invasion and epithelial mesenchymal transition (EMT) of prostate cancer (PCa) cells.Methods:Human PCa cell lines, IA8-ARCaP and LNCaP/HIF-1a, were used in this study. The cell lines were found to process EMT in our previous study. The PCa cells were treated with increasing concentrations, from 0.1 to 75μmol/L. Proliferation was assessed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. EMT was proven by cell morphological transition and the expression changes of EMT-related markers, which were confirmed by RT-PCR, Western blotting, and indirect immunofluorescence labeling. Transwell invasion assay was used to analyze the invasive potency.Results:The addition of genistein to the medium reduced the IA8-ARCaP and LNCaP/HIF-1a viable cell number in a dose-dependent manner (with increasing concentrations from 15 to 75 μmol/L). Less than 15 μmol/L genistein was selected as the low dose concentration, which did not affect cell proliferation. The treatment of cells with low-dose genistein induced the reversal of EMT, which was confirmed by cell morphological transition and the expression changes of EMT-related markers. The reversal of EMT in the PCa cells by low-dose genistein was in a dose-dependent manner. Moreover, low-dose genistein effectively inhibited invasion of the PCa cells in vitro.Conclusion:These results showed that treatment with low-dose genistein may be a potential strategy for the suppression of invasive growth through the reversal of EMT in cancer cells, which justifies the potential use of soybean foods as a practical chemopreventive approach for patients with PCa.

Chemopreventive and Chemotherapeutic Effects of Intravesical Silibinin against Bladder Cancer by Acting on Mitochondria

Intravesical chemotherapy is often used to prevent the recurrence of superficial bladder cancer after transurethral resection. A search for more effective and less toxic intravesical agents is urgently needed. We previously found the in vitro apoptotic effects of silibinin, a natural flavonoid, on high-risk bladder carcinoma cells. Here, we further explored the underlying mechanisms and examined the intravesical efficacy in the prevention and treatment of bladder cancer. Human bladder carcinoma cell line 5637, which has the same molecular features of high-risk superficial bladder cancer, was used as the model system in vitro and in vivo. Autochthonous rat model of bladder cancer induced by intravesical N-methyl-N-nitrosourea (MNU) was used to investigate its intravesical efficacy. Exposure of 5637 cells to silibinin resulted in growth inhibition and induction of caspase-dependent and -independent apoptosis, which was associated with disruption of mitochondrial membrane potential and selective release of cytochrome c, Omi/HtrA2, and apoptosis-inducing factor (AIF) from mitochondria. Silibinin also downregulated survivin and caused nuclear translocation of AIF. Oral silibinin suppressed the growth of 5637 xenografts, which was accompanied with the activation of caspase-3, downregulation of survivin, and increased translocation of AIF. Furthermore, intravesical silibinin effectively inhibited the carcinogenesis and progression of bladder cancer in rats initiated by MNU by reducing the incidence of superficial and invasive bladder lesions without any side effects, which was accompanied with proapoptotic effects. These findings identify the in vitro and in vivo antitumor efficacy of silibinin, and suggest silibinin as an effective and novel intravesical agent for bladder cancer. Mol Cancer Ther; 10(1); 104–16. ©2011 AACR.