Sunitinib malate (SU-11248) alone or in combination with low-dose docetaxel inhibits the growth of DU-145 prostate cancer xenografts

The aim of the study was to evaluate the activity of the antiangiogenic agent SU-11248 (sunitinib malate, Sutent^®), alone or in combination with docetaxel. To this end, animals bearing DU-145 human hormone-refractory prostate cancer (HRPC) xenografts were treated with sunitinib (40 mg/kg daily, p.o.), docetaxel (10 or 30 mg/kg/week, i.v.), a combination of sunitinib (40 mg/kg daily) and docetaxel (10 mg/kg/week) or vehicle alone. At the end of the 3-week dosing schedule, single-agent treatment induced a tumor regression of 59%, 49% and 75% for sunitinib, docetaxel 10 mg/kg, and docetaxel 30 mg/kg, respectively. The combination of sunitinib with low-dose (10 mg/kg) docetaxel produced a tumor regression comparable to that obtained with high-dose (30 mg/kg) docetaxel, but tolerability was higher as indicated by mice weight. Both sunitinib and docetaxel inhibited tumor regrowth after initial treatment with the alternate drug. These results suggest that sunitinib alone or in combination with low-dose docetaxel may have a role in the treatment of HRPC.     

Salinomycin inhibits prostate cancer growth and migration via induction of oxidative stress

Background:

We have shown that a sodium ionophore monensin inhibits prostate cancer cell growth. A structurally related compound to monensin, salinomycin, was recently identified as a putative cancer stem cell inhibitor.

Methods:

The growth inhibitory potential of salinomycin was studied in a panel of prostate cells. To get insights into the mechanism of action, a variety of assays such as gene expression and steroid profiling were performed in salinomycin-exposed prostate cancer cells.

Results:

Salinomycin inhibited the growth of prostate cancer cells, but did not affect non-malignant prostate epithelial cells. Salinomycin impacted on prostate cancer stem cell functions as evidenced by reduced aldehyde dehydrogenase activity and the fraction of CD44⁺ cells. Moreover, salinomycin reduced the expression of MYC, AR and ERG, induced oxidative stress as well as inhibited nuclear factor-κB activity and cell migration. Furthermore, profiling steroid metabolites revealed increased levels of oxidative stress-inducing steroids 7-ketocholesterol and aldosterone and decreased levels of antioxidative steroids progesterone and pregnenolone in salinomycin-exposed prostate cancer cells.

Conclusion:

Our results indicate that salinomycin inhibits prostate cancer cell growth and migration by reducing the expression of key prostate cancer oncogenes, inducing oxidative stress, decreasing the antioxidative capacity and cancer stem cell fraction.     

Post-translational regulation of COX2 activity by FYN in prostate cancer cells

While increased COX2 expression and prostaglandin levels are elevated in human cancers, the mechanisms of COX2 regulation at the post-translational level are unknown. Initial observation that COX2 forms adduct with non-receptor tyrosine kinase FYN, prompted us to study FYN-mediated post-translational regulation of COX2. We found that FYN increased COX2 activity in prostate cancer cells DU145, independent of changes in COX2 or COX1 protein expression levels. We report that FYN phosphorylates human COX2 on Tyr 446, and while corresponding phospho-mimetic COX2 mutation promotes COX2 activity, the phosphorylation blocking mutation prevents FYN-mediated increase in COX2 activity.     

E2F5和MYC癌基因在前列腺癌中的表达及其意义

目的： 检测染色体8q21~24区域E2F5和MYC癌基因在前列腺癌 (prostate cancer,PCa) 细胞系、癌组织及癌旁组织中的表达,分析其表达水平与临床病理参数的关系,并探讨其意义。方法：采用DNA qPCR检测PCa细胞系Du145、LNCap、PC3和PCa组织标本中E2F5和MYC基因DNA拷贝数,Western blot和免疫组织化学技术检测前列腺癌和癌旁组织中E2F5和MYC蛋白的表达,结合患者临床病理参数进一步验证和分析蛋白表达及其临床意义。结果：DNA qPCR结果显示,PCa组织中E2F5和MYC DNA拷贝数较癌旁组织明显增加 (P<0.05或P<0.01),但在癌细胞系中其表达与对照组比较无明显变化 (P>0.05)。Western blot显示,在PCa组织中E2F5和MYC蛋白表达水平显著高于癌旁组织 (P<0.05或P<0.01)。免疫组化染色发现,与癌旁良性组织相比,E2F5和MYC蛋白表达显著增加 (P均<0.01)。而且,E2F5的过表达与高的Gleason评分 (P<0.01)、临床分期 (P=0.01)、阳性转移 (P <0.01)、生化复发阳性 (P<0.01)相关。MYC的过表达与阳性转移 (P=0.02)、生化复发阳性 (P=0.02)相关。且PCa组织中E2F5和MYC蛋白表达两者呈正相关关系 (rs=0.5,P<0.01)。结论：E2F5和MYC的表达增加可能与PCa的发生发展密切相关,E2F5可能是PCa新的潜在候选分子标志物。     

Circulating tumor cells as a predictive biomarker in patients with hormone-sensitive prostate cancer

Introduction

Circulating tumor cell (CTC) enumeration by using the Cellsearch platform has established prognostic and predictive value in patients with metastatic castration-resistant prostate cancer (mCRPC). Limited information exists regarding the clinical utility of CTC enumeration in metastatic hormone-sensitive prostate cancer (mHSPC). The goal of this study was to prospectively determine the relative clinical utility of CTCs in mHSPC.

Patients and Methods

We analyzed serial CTC in conjunction with other classic biomarkers in 33 consecutive patients treated at the Nevada Cancer Institute with HSPC initiating androgen deprivation therapy and correlated these patients with prognostic prostate-specific antigen (PSA) endpoints and onset of CRPC.

Results

Initial CTC correlated positively with lactate dehydrogenase and alkaline phosphatase, and were unrelated to PSA and testosterone. In univariate analysis, baseline CTC, alkaline phosphatase, lactate dehydrogenase, testosterone, and follow-up CTC were individual predictors of progression to CRPC. In a multivariate Cox regression, only baseline CTC retained independent predictive value. Threshold analysis revealed the cutpoint that optimized specificity and sensitivity of the test to be 3 cells per 7.5 mL whole blood. Baseline CTC also correlated well with PSA nadir benchmarks.

Conclusions

Initial CTC values predict the duration and magnitude of response to hormonal therapy. CTC enumeration may identify patients at risk of progression to CRPC before initiation of androgen deprivation therapy.     

Prostaglandin E2 induces vascular endothelial growth factor secretion in prostate cancer cells through EP2 receptor-mediated cAMP pathway

Prostaglandin E2 (PGE2) has been shown to induce expression of vascular endothelial growth factor (VEGF) and other signaling molecules in several cancers. PGE2 elicits its functions though four G-protein coupled membrane receptors (EP1-4). In this study, we investigated the role of EP receptors in PGE2-induced molecular events in prostate cancer cells. qRT-PCR analysis revealed that PC-3 cells express a substantially higher level of EP2 and moderately higher EP4 than DU145 and LNCaP cells. LNCaP cells had virtually no detectable EP2 mRNA. EP1 and EP3 mRNAs were not detected in these cells. Treatment of prostate cancer cells with PGE2 (1 nM-10 microM) increased both VEGF secretion and cyclic adenosine monophosphate (cAMP) production. Levels of induction in PC-3 cells were greater than in DU145 and LNCaP cells. The selective EP2 agonist CAY10399 also significantly increased VEGF secretion and cAMP production in PC-3 cells, but not in DU145 and LNCaP cells. Moreover, PGE2 and CAY10399 increased mitogen activated protein kinase/extracellular signal regulated kinase (MAPK/Erk) and Akt phosphorylation in PC-3 and DU145 cells, but not in LNCaP cells. However, neither the MAPK/Erk inhibitor U0126 nor the PI3K/Akt inhibitor LY294002 abolished PGE2-induced VEGF secretion in PC-3 cells. We further demonstrated that the adenylate cyclase activator forskolin and the cAMP anologue 8-bromo-cAMP mimicked the effects of PGE2 on VEGF secretion in PC-3 cells. Meanwhile, the adenylate cyclase inhibitor 2'5'-dideoxyadenosine, at concentrations that inhibited PGE2-induced cAMP, significantly blocked PGE2-induced VEGF secretion in PC-3 cells. We conclude that PGE2-induced VEGF secretion in prostate cancer cells is mediated through EP2-, and possibly EP4-, dependent cAMP signaling pathways.     

Aldehyde dehydrogenase 3A1 associates with prostate tumorigenesis

Background:

Accumulating evidence demonstrates high levels of aldehyde dehydrogense (ALDH) activity in human cancer types, in part, because of its association with cancer stem cells. Whereas ALDH1A1 and ALDH7A1 isoforms were reported to associate with prostate tumorigenesis, whether other ALDH isoforms are associated with prostate cancer (PC) remains unclear.

Methods:

ALDH3A1 expression was analysed in various PC cell lines. Xenograft tumours and 54 primary and metastatic PC tumours were stained using immunohistochemistry for ALDH3A1 expression.

Results:

In comparison with the non-stem counterparts, a robust upregulation of ALDH3A1 was observed in DU145-derived PC stem cells (PCSCs). As DU145 PCSCs produced xenograft tumours with more advanced features compared with those derived from DU145 cells, higher levels of ALDH3A1 were detected in the former; a dramatic elevation of ALDH3A1 occurred in DU145 cell-derived lung metastasis compared with local xenograft tumours. Furthermore, while ALDH3A1 was not observed in prostate glands, ALDH3A1 was clearly present in PIN, and further increased in carcinomas. In comparison with the paired local carcinomas, ALDH3A1 was upregulated in lymph node metastatic tumours; the presence of ALDH3A1 in bone metastatic PC was also demonstrated.

Conclusions:

We report here the association of ALDH3A1 with PC progression.     

A novel polyamine analog (SL-11093) inhibits growth of human prostate tumor xenografts in nude mice

PURPOSE: We tested the polyamine analog SL-11093 (3,8,13,18-tetraaza-10,11-[(E)-1,2-cyclopropyl]eicosane tetrahydrochloride) as an effective chemotherapeutic agent against human prostate cancer grown in nude mice. METHODS: NCr-nu mice grafted with DU-145 human prostate tumor cells were treated i.p. with SL-11093 at 50 mg/kg q1dx5 for either three or five cycles separated by intervals of about 10-15 days. RESULTS: In treated animals, tumor growth remained arrested for up to 100 days with minimal animal weight loss. None of the animals died during the treatment and in one experiment two out of six animals showed no palpable tumor. SL-11093 was readily taken up by the tumors, where its levels remained elevated for about 48 h after the end of drug administration. In liver and in kidney, SL-11093 (a (alpha)N,(omega)N-bisethyl derivative) was oxidatively N-deethylated predominantly to its monoethyl and di-deethyl derivatives. In time, the monoethyl derivative was further dealkylated, with a loss of an aminobutyl chain to form an aminomethyl cyclopropyl derivative. In tumor (and in lung), N-dealkylation reactions were less evident. CONCLUSION: SL-11093 is an effective chemotherapeutic agent against a human prostate tumor xenograft grown in nude mice. The drug accumulation and slow metabolism in tumor compared to other tissues would most likely reduce systemic toxicity of the drug and contribute to a larger therapeutic window for SL-11093 as compared to other cytotoxic polyamine analogs.     

Tumorigenic potential of circulating prostate tumor cells

Circulating tumor cells (CTCs) have received intense scientific scrutiny because they travel in the bloodstream and are therefore well situated to mediate hematogenous metastasis. However, the potential of CTCs to actually form new tumors has not been tested. Popular methods of isolating CTCs are biased towards larger, more differentiated, non-viable cells, creating a barrier to testing their tumor forming potential. Without relying on cell size or the expression of differentiation markers, our objective was to isolate viable prostate CTCs from mice and humans and assay their ability to initiate new tumors. Therefore, blood was collected from transgenic adenocarcinoma of the mouse prostate (TRAMP) mice and from human patients with metastatic castration-resistant prostate cancer (PCa). Gradient density centrifugation or red cell lysis was used to remove erythrocytes, and then leukocytes were depleted by magnetic separation using CD45 immunoaffinity beads. CTCs fractions from TRAMP mice and PCa patients were verified by immunocytochemical staining for cytokeratin 8 and EpCAM, and inoculated into immunodeficient mice. TRAMP tumor growth was monitored by palpation. Human tumor growth formation was monitored up to 8 months by ultrasensitive PSA assays performed on mouse serum. We found viable tumor cells present in the bloodstream that were successfully isolated from mice without relying on cell surface markers. Two out of nine immunodeficient mice inoculated with TRAMP CTCs developed massive liver metastases. CTCs were identified in blood from PCa patients but did not form tumors. In conclusion, viable CTCs can be isolated without relying on epithelial surface markers or size fractionation. TRAMP CTCs were tumorigenic, so CTCs isolated in this way contain viable tumor-initiating cells. Only two of nine hosts grew TRAMP tumors and none of the human CTCs formed tumors, which suggests that most CTCs have relatively low tumor-forming potential. Future studies should identify and target the highly tumorigenic cells.     

Epac inhibits migration and proliferation of human prostate carcinoma cells

Background:

It was recently found that cAMP mediates protein kinase A-independent effects through Epac proteins. The aim of this study was to investigate the role of Epac in migration and proliferation of prostate carcinoma cells.

Methods:

The effect of Epac activation was determined by [³H]thymidine incorporation and scratch assays in PC-3 and DU 145 cells. Furthermore, cytoskeletal integrity was analysed by phalloidin staining. The participation of intracellular Epac effectors such as mitogen-activated protein (MAP) kinases, Rap1- and Rho-GTPases was determined by immunoblotting and pull-down assay.

Results:

The specific Epac activator 8-pCPT-2′-O-Me-cAMP (8-pCPT) interfered with cytoskeletal integrity, reduced DNA synthesis, and migration. Although 8-pCPT activated Rap1, it inhibited MAP kinase signalling and RhoA activation. These findings were translated into functional effects such as inhibition of mitogenesis, cytoskeletal integrity, and migration.

Conclusion:

In human prostate carcinoma cells, Epac inhibits proliferative and migratory responses likely because of inhibition of MAP kinase and RhoA signalling pathways. Therefore, Epac might represent an attractive therapeutic target in the treatment of prostate cancer.     

A novel BRCA2 mutation in prostate cancer sensitive to combined radiotherapy and androgen deprivation therapy

Genetic factors contribute to more than 40% of prostate cancer risk, and mutations in BRCA1 and BRCA2 are well-established risk factors. By using target capture-based deep sequencing to identify potential pathogenic germline mutations, followed by Sanger sequencing to determine the loci of the mutations, we identified a novel pathogenic BRCA2 mutation caused by a cytosine-to-guanine base substitution at position 4211, resulting in protein truncation (p.Ser1404Ter), which was confirmed by immunohistochemistry. Analysis of peripheral blood also identified benign polymorphisms in BRCA2 (c.7397T>C, p.Val2466Ala) and SRD5A2 (c.87G>C, p.Lys29Asn). Analysis of tumor tissues revealed seven somatic mutations in prostate tumor tissue and nine somatic mutations in esophageal squamous carcinoma tissue (single nucleotide polymorphisms, insertions, and deletions). Five-year follow-up results indicate that ADT combined with radiotherapy successfully treated the prostate cancer. To our knowledge, we are the first to report the germline BRCA2 mutation c.4211C>G (p.Ser1404Ter) in prostate cancer. Combined ADT and radiotherapy may be effective in treating other patients with prostate cancer caused by this or similar mutations.     

CIP2A is a candidate therapeutic target in clinically challenging prostate cancer cell populations

Residual androgen receptor (AR)-signaling and presence of cancer stem-like cells (SCs) are the two emerging paradigms for clinically challenging castration-resistant prostate cancer (CRPC). Therefore, identification of AR-target proteins that are also overexpressed in the cancer SC population would be an attractive therapeutic approach.Our analysis of over three hundred clinical samples and patient-derived prostate epithelial cultures (PPECs), revealed Cancerous inhibitor of protein phosphatase 2A (CIP2A) as one such target. CIP2A is significantly overexpressed in both hormone-naïve prostate cancer (HN-PC) and CRPC patients. CIP2A is also overexpressed, by 3- and 30-fold, in HN-PC and CRPC SCs respectively. In vivo binding of the AR to the intronic region of CIP2A and its functionality in the AR-moderate and AR-high expressing LNCaP cell-model systems is also demonstrated. Further, we show that AR positively regulates CIP2A expression, both at the mRNA and protein level. Finally, CIP2A depletion reduced cell viability and colony forming efficiency of AR-independent PPECs as well as AR-responsive LNCaP cells, in which anchorage-independent growth is also impaired.These findings identify CIP2A as a common denominator for AR-signaling and cancer SC functionality, highlighting its potential therapeutic significance in the most clinically challenging prostate pathology: castration-resistant prostate cancer.     

Metformin inhibits prostate cancer cell proliferation,migration, and tumor growth through upregulation of PEDF expression

Metformin has been reported to inhibit the growth of various types of cancers, including prostate cancer. Yet the mode of anti-cancer action of metformin and the underlying mechanisms remain not fully elucidated. We hypothesized that the antitumorigenic effects of metformin are mediated through upregulation of pigment epithelium-derived factor (PEDF) expression in prostate cancer cells. In this report, metformin treatment significantly inhibited the proliferation and colony formation of prostate cancer cells, in a dose- and time-dependent manner. Meanwhile, Metformin markedly suppressed migration and invasion and induced apoptosis of both LNCaP and PC3 cancer cells. Metformin also reduced PC3 tumor growth in BALB/c nude mice in vivo. Furthermore, metformin treatment was associated with higher PEDF expression in both prostate cancer cells and tumor tissue. Taken together, metformin inhibits prostate cancer cell proliferation, migration, invasion and tumor growth, and these activities are mediated by upregulation of PEDF expression. These findings provide a novel insight into the molecular functions of metformin as an anticancer agent.