Loss of P53 facilitates invasion and metastasis of prostate cancer cells

Prostate cancer is a lethal cancer for the invasion and metastasis in its earlier period. P53 is a tumor suppressor gene which plays a critical role on safeguarding the integrity of genome. However, loss of P53 facilitates or inhibits the invasion and metastasis of tumor is still suspended. In this study, we are going to explain whether loss of P53 affect the invasion and metastasis of prostate cancer cells. To explore whether loss of P53 influences the invasion and metastasis ability of prostate cancer cells, we first compared the invasion ability of si-P53 treated cells and control cells by wound healing, transwell assay, and adhesion assay. We next tested the activity of MMP-2, MMP-9, and MMP-14 by western blot and gelatin zymography. Moreover, we employed WB and IF to identify the EMT containing E-cad, N-cad, vimentin, etc. We also examined the expression of cortactin, cytoskeleton, and paxillin by immunofluorescence, and tested the expression of ERK and JNK by WB. Finally, we applied WB to detect the expression of FAK, Src, and the phosphorylation of them to elucidate the mechanism of si-P53 influencing invasion and metastasis. According to the inhibition rate of si-P53, we choose the optimized volume of si-P53. With the volume, we compare the invasion and metastasis ability of Du145 and si-P53 treated cells. We find si-P53 promotes the invasion and metastasis in prostate cancer cells, increases the expression and activity of MMP-2/9 and MMP-14. Also, si-P53 promotes EMT and cytoskeleton rearrangement. Further analyses explain that this effect is associated with FAK-Src signaling pathway. Loss of P53 promotes the invasion and metastasis ability of prostate cancer cells and the mechanism is correlated with FAK-Src signaling pathway. P53 is involved in the context of invasion and metastasis.     

Androgen receptor represses the neuroendocrine transdifferentiation process in prostate cancer cells

Androgen-ablation therapy is an effective method for treating prostate cancer. However, prostate tumors that survive long-term androgen-ablation therapy are classified as androgen-independent as they proliferate in the absence of androgens, and they tend to be enriched for neuroendocrine (NE) cells. Androgen withdrawal causes androgen-dependent prostate cancer cells to adopt a pronounced NE phenotype, suggesting that androgen receptor (AR) represses an intrinsic NE transdifferentiation process in prostate cancer cells. In this report we show that short interfering RNA-induced AR silencing induced a NE phenotype that manifested itself in the growth of dendritic-like processes in both the androgen-dependent LNCaP and androgen-independent LNCaP-AI human prostate cancer cells. Western blot analysis revealed that neuronal-specific enolase, a marker of the neuronal lineage, was increased by AR knockdown in LNCaP cells. The expression levels of the neuronal-specific cytoskeletal proteins beta-tubulin III, nestin, and glial acidic fibrillary protein were also characterized in AR knockdown cells. Most interestingly, AR silencing induced beta-tubulin III expression in LNCaP cells, while AR knockdown increased glial acidic fibrillary protein levels in both LNCaP and LNCaP-AI cells. Lastly, AR silencing reduced the proliferative capacity of LNCaP and LNCaP-AI cells. Our data demonstrate that AR actively represses an intrinsic NE transdifferentiation process in androgen-responsive prostate cancer cells and suggest a potential link between AR inactivation and the increased frequency of NE cells in androgen-independent tumors.     

Function and molecular mechanisms of neuroendocrine cells in prostate cancer

Benign prostate contains luminal epithelial cells, basal cells and a minor component of neuroendocrine cells whose function may be to regulate the growth, differentiation and secretory function of the prostate gland. Neuroendocrine (NE) cells are also present in prostate cancer (PC), and many studies have shown that their number increases in high-grade and high-stage tumors, particularly in hormonally treated and hormone-refractory (androgen independent) PC. Unlike the non-neuroendocrine secretory-type PC cells, NE cells lack androgen receptor and are likely androgen independent. Therefore it is conceivable that hormonal therapy for advanced or metastatic prostate cancer, which consists of inhibiting androgen production or blocking androgen function, will not eliminate NE cancer cells. Instead, these cells may be enriched after the therapy and they may establish paracrine networks to stimulate androgen-independent proliferation of PC, leading to tumor recurrence. This article reviews the major functions of NE cells in PC, including stimulation of cancer proliferation and invasion, apoptosis resistance and angiogenesis. It also discusses molecular pathways involved in NE differentiation and the effectors of the NE cells.     

Expression of core 2 beta1,6-N-acetylglucosaminyltransferase facilitates prostate cancer progression

Cell surface carbohydrates expressed on epithelial cells are thought to play an important role in tumor progression. Previously, we have shown that expression of core 2-branched O-glycans is closely correlated with vessel invasion and depth of invasion in colon and lung carcinomas. In this study, we found that expression of core 2 beta1,6-N-acetylglucosaminyltransferase-1, Core2GnT, is positively correlated with the progression of prostate cancer in human patients. Statistical analysis demonstrated that Core2GnT is an independent predictor for progressed pathological stage (pT3) and for prostate-specific antigen (PSA) relapse. To determine directly the roles of Core2GnT in prostate cancer progression, we set up an experimental tumor model using the LNCaP prostate cancer cell line. Because this line does not express Core2GnT, we established an LNCaP line stably expressing Core2GnT, LNCap-Core2GnT, by transfecting cDNA encoding Core2GnT. When mock-transfected LNCaP cells and LNCaP-Core2GnT were inoculated in the prostate of nude mice, LNCaP-Core2GnT cells produced three times heavier prostate tumors than mock-transfected LNCaP cells. Furthermore, we found that LNCaP-Core2GnT cells adhered more strongly to prostate stromal cells, type IV collagen and laminin than did LNCaP-mock cells, but LNCaP and LNCaP-Core2GnT cells grew almost at the same rate on plates coated with type IV collagen or laminin. These results indicate that Core2GnT is an extremely useful prognostic marker for prostate cancer progression. The results also suggest that acquiring Core2GnT in prostate carcinoma cells facilitates adhesion to type IV collagen and laminin, and this increased adhesion may be a cause for aggressive tumor formation by prostate cancer cells expressing Core2GnT.     

A model for the intrinsic limit of cancer therapy: Duality of treatment-induced cell death and treatment-induced stemness

Intratumor cellular heterogeneity and non-genetic cell plasticity in tumors pose a recently recognized challenge to cancer treatment. Because of the dispersion of initial cell states within a clonal tumor cell population, a perturbation imparted by a cytocidal drug only kills a fraction of cells. Due to dynamic instability of cellular states the cells not killed are pushed by the treatment into a variety of functional states, including a “stem-like state” that confers resistance to treatment and regenerative capacity. This immanent stress-induced stemness competes against cell death in response to the same perturbation and may explain the near-inevitable recurrence after any treatment. This double-edged-sword mechanism of treatment complements the selection of preexisting resistant cells in explaining post-treatment progression. Unlike selection, the induction of a resistant state has not been systematically analyzed as an immanent cause of relapse. Here, we present a generic elementary model and analytical examination of this intrinsic limitation to therapy. We show how the relative proclivity towards cell death versus transition into a stem-like state, as a function of drug dose, establishes either a window of opportunity for containing tumors or the inevitability of progression following therapy. The model considers measurable cell behaviors independent of specific molecular pathways and provides a new theoretical framework for optimizing therapy dosing and scheduling as cancer treatment paradigms move from “maximal tolerated dose,” which may promote therapy induced-stemness, to repeated “minimally effective doses” (as in adaptive therapies), which contain the tumor and avoid therapy-induced progression.     

Polo-like kinase 1 facilitates loss of Pten tumor suppressor-induced prostate cancer formation

Loss of the tumor suppressor Pten (phosphatase and tensin homolog deleted on chromosome 10) is thought to mediate the majority of prostate cancers, but the molecular mechanism remains elusive. In this study, we demonstrate that Pten-depleted cells suffer from mitotic stress and that nuclear function of Pten, but not its phosphatase activity, is required to reverse this stress phenotype. Further, depletion of Pten results in elevated expression of Polo-like kinase 1 (Plk1), a critical regulator of the cell cycle. We show that overexpression of Plk1 correlates with genetic inactivation of Pten during prostate neoplasia formation. Significantly, we find that elevated Plk1 is critical for Pten-depleted cells to adapt to mitotic stress for survival and that reintroduction of wild-type Pten into Pten-null prostate cancer cells reduces the survival dependence on Plk1. We further show that Plk1 confers the tumorigenic competence of Pten-deleted prostate cancer cells in a mouse xenograft model. These findings identify a role of Plk1 in facilitating loss of Pten-induced prostate cancer formation, which suggests that Plk1 might be a promising target for prostate cancer patients with inactivating Pten mutations.     

Loss of mir-146a function in hormone-refractory prostate cancer

The pattern of microRNA (miRNA) expression is associated with the degree of tumor cell differentiation in human prostate cancer. MiRNAs bind complementarily to either oncogenes or tumor suppressor genes, which are consequently silenced, resulting in alterations of tumorigenecity. We have detected eight down-regulated and three up-regulated known miRNAs in androgen-independent human prostate cancer cells compared to those in androgen-dependent cells, using miRNA microarray analyses. These identified miRNAs showed the same expression patterns in hormone-refractory prostate carcinomas (HRPC) compared to androgen-sensitive noncancerous prostate epithelium as determined by fluorescent in situ hybridization assays in human prostate cancer tissue arrays. One of the eight down-regulated miRNAs, mir-146a, was selected and constitutively expressed to examine its effects on suppression of prostate cancer transformation from androgen-dependent to -independent cells as determined by in vitro tumorigenecity assays. Transfection of mir-146a, which perpetually express the miRNA, suppressed >82% of the expression of the targeted protein-coding gene, ROCK1, in androgen-independent PC3 cells, consequently markedly reducing cell proliferation, invasion, and metastasis to human bone marrow endothelial cell monolayers. Given that ROCK1 is one of the key kinases for the activation of hyaluronan (HA)-mediated HRPC transformation in vivo and in PC3 cells, mir-146a may function as a tumor-suppressor gene in modulating HA/ROCK1-mediated tumorigenecity in androgen-dependent prostate cancer.     

Molecular changes associated with prostate cancer development

The epidemiologic characteristics of prostate cancer (PCa) have been recognized for several decades. It is of great importance to understand the factors responsible for prostate carcinogenesis, why some carcinomas remain "clinically silent" during life, whereas other tumors progress to present clinically and may lead to PCa-related death. A better understanding of these mechanisms in molecular genetic terms should point to more rational approaches to disease prevention, intervention and treatment. The aim of this review is to provide a comprehensive overview of the current state of knowledge regarding the molecular alterations of PCa.     

Role of estrogens in development of prostate cancer

Estrogens have previously been extensively used in prostate cancer treatment. Serious side effects, primarily in cardiovascular system have, however, limited their use. The therapeutic effect of estrogen in preventing prostate cancer growth was mainly obtained indirectly by feedback inhibition of the hypothalamic release of LRH leading to lowered serum androgen levels and castration like effects. Prostate tissue is also most probably a target for direct regulation by estrogens. Prostate contains estrogen receptor alpha (ERalpha) and beta (ERbeta), which are localized characteristically in stroma and epithelium, respectively. The physiological function of these receptors is not known but there is evidence of the role of estrogens in prostatic carcinogenesis. Developing prostate seems particularly sensitive to increased level of endogenous and/or exogenous estrogens. Perinatal or neonatal exposure of rats and mice to estrogens leads to "imprinting" of prostate associated with increased proliferation, inflammation and dysplastic epithelial changes later in life. Prolonged treatment of adult rodents with estrogens along with androgens also leads to epithelial metaplasia, PIN-like lesions and even adenocarcinoma of prostate speaking for the role of estrogen in prostate cancer development. Recent results concerning antiestrogen inhibition of prostate cancer development beyond PIN-type lesions in transgenic mouse models further suggests a role for estrogens in prostate cancer progression. These results also suggest that direct inhibition of estrogen action at the level of prostate tissue may provide an important novel principle of development of prostate cancer therapies.     

Discoidin domain receptor 2 facilitates prostate cancer bone metastasis via regulating parathyroid hormone-related protein

Prostate cancer frequently metastasizes to the skeleton but the underlying mechanism remains largely undefined. Discoidin domain receptor 2 (DDR2) is a member of receptor tyrosine kinase (RTK) family and is activated by collagen binding. This study aimed to investigate the function and detailed mechanism of DDR2 in prostate cancer bone dissemination. Herein we found that DDR2 was strongly expressed in bone-metastatic prostate cancer cells and tissues compared to that in normal controls. Enhanced expression of constitutively activated DDR2 led to elevation in motility and invasiveness of prostate cancer cells, whereas knockdown of DDR2 through specific shRNA caused a dramatic repression. Knockdown of DDR2 in prostate cancer cells resulted in significant decrease in the proliferation, differentiation and function of osteoblast. Over-expression of DDR2 in prostate cancer cells resulted in notable acceleration of osteoclast differentiation and bone resorption, whereas knockdown of DDR2 exhibited the opposite effects. An intrabone injection bone metastasis animal model demonstrated that DDR2 promoted osteolytic metastasis in vivo. Molecular evidence demonstrated that DDR2 regulated the expression, secretion, and promoter activity of parathyroid hormone-related protein (PTHrP), via modulating Runx2 phosphorylation and transactivity. DDR2 was responsive to TGF-β and involved in TGF-β-mediated osteoclast activation and bone resorption. In addition, DDR2 facilitated prostate cancer cells adhere to type I collagen. This study reveals for the first time that DDR2 plays an essential role in prostate cancer bone metastasis. The mechanism disclosure may provide therapeutic targets for the treatment of prostate cancer.     

Loss of PTEN expression in neuroendocrine pancreatic tumors

PTEN (phosphatase and tensin homologue deleted from chromosome 10) is a well established tumor suppressor gene, which was cloned to chromosome 10q23. PTEN plays an important role in controlling cell growth, apoptosis, cell adhesion, and cell migration. In various studies, a genetic change as well as loss of PTEN expression by different carcinomas has been described. To date, the role of PTEN as a differentiation marker for neuroendocrine tumors (NET) and for the loss of PTEN expression is still unknown. It is assumed that loss of PTEN expression is important for tumor progression of NETs. We hypothesize that PTEN might be used as a new prognostic marker. We report 38 patients with a NET of the pancreas. Tumor tissues were surgically resected, fixed in formalin, and embedded in paraffin. PTEN expression was evaluated by immunohistochemistry and was correlated with several clinical and pathological parameters of each individual tumor. After evaluation of our immunohistochemistry data using a modified Remmele Score, a widely accepted method for categorizing staining results for reports and statistical evaluation, staining results of PTEN expression were correlated with the clinical and pathological parameters of each individual tumor. Our data demonstrates a significant difference in survival with existence of lymph node or distant metastases. Negative patients show a significant better survival compared with positive patients. Furthermore, we show a significant difference between PTEN expression and WHO or TNM classification. Taken together, our data shows a positive correlation between WHO classification and the new TNM classification of NETs, and loss of PTEN expression as well as survival. These results strongly implicate that PTEN might be helpful as a new prognostic factor.     

Steroid hormones, polypeptide growth factors, hormone refractory prostate cancer, and the neuroendocrine phenotype

The growth, development, and differentiation of the prostate gland is largely dependent on the action of androgens and peptide growth factors that act differentially at the level of the mesenchymal and epithelial compartments. It is our premise that to understand the emergence of metastatic and hormone refractory prostate cancer we need to investigate: (1) how androgen action at the level of the mesenchyme induces the production of peptide growth factors that in turn can facilitate the growth and development of the epithelial compartment; (2) how androgen action at the level of the epithelium induces and maintains cellular differentiation, function, and replicative senescence; and (3) how transformation of the prostate gland can corrupt androgen and growth factor signaling homeostasis. To this end, we focus our discussion on how deregulation of the growth factor signaling axis can cooperate with deregulation of the androgen signaling axis to facilitate transformation, metastasis, and the emergence of the hormone refractory and neuroendocrine phenotypes associated with progressive androgen-independent prostate cancer. Finally, we suggest a working hypothesis to explain why hormone ablation therapy works to control early disease but fails to control, and may even facilitate, advanced prostate cancer.     

Phosphatidylinositol 3-kinase-AKT-mammalian target of rapamycin pathway is essential for neuroendocrine differentiation of prostate cancer

Hormonal therapy of prostate cancer, by inhibiting androgen production and/or androgen function, is the treatment of choice for advanced prostate cancer. Although most patients respond initially, the effect is only temporary, and the tumor cells will resume proliferation in an androgen-deprived environment. The mechanism for androgen-independent proliferation of cancer cells is unclear. Hormonal therapy induces neuroendocrine differentiation of prostate cancer cells, which is hypothesized to contribute to tumor recurrence by a paracrine mechanism. We studied signal transduction pathways of neuroendocrine differentiation in LNCaP cells after androgen withdrawal, and we showed that both the phosphatidylinositol 3-kinase-AKT-mammalian target of rapamycin pathway and ERK are activated, but only the former is required for neuroendocrine differentiation. A constitutively active AKT promotes neuroendocrine differentiation and a dominant negative AKT inhibits it. Activation of AKT by IGF-1 leads to neuroendocrine differentiation, and neuroendocrine differentiation induced by epinephrine requires AKT activation. We also show that the AKT pathway is likely responsible for neuroendocrine differentiation in DU145, an androgen-independent prostate cancer cell line. Therefore, our study demonstrated a novel function of the AKT pathway in prostate cancer progression and identified potential targets that may be explored for the treatment of androgen-independent cancer.     

The Notch pathway in prostate development and cancer

Abstract The Notch family of transmembrane receptors are important mediators of cell fate determination. Accordingly, Notch signaling is intimately involved in the development of numerous tissues. Recent findings have highlighted a critical role for Notch signaling in normal prostate development. Notch signaling is required for embryonic and postnatal prostatic growth and development, for proper cell lineage specification within the prostate, as well as for adult prostate maintenance and regeneration following castration and hormone replacement. Evidence for Notch as a regulator of prostate cancer development, progression, and metastasis has also emerged. This review summarizes our current understanding of the role of Notch pathway elements, including members of the Jagged, Delta-like, hairy/enhancer-of-split, and hairy/enhancer-of-split related with YRPW motif families, in prostate development and tumorigenesis. Data supporting Notch pathway elements as oncogenes and tumor suppressors in prostate tumors, as well as data implicating Notch receptors and ligands as potential markers of normal prostate stem/progenitor cells and prostate cancer stem/initiating cells, are also presented.     

Autophagy protects breast cancer cells from epirubicin-induced apoptosis and facilitates epirubicin-resistance development

Epirubicin (EPI) is one of the most effective drugs against cancer. But the acquired resistance of cancer cells to EPI is becoming a major obstacle for successful cancer therapy. Recently, some studies have revealed that macroautophagy (here referred to as autophagy) may protect the cancer cell from anticancer drug-induced death, so autophagy might be related to the development of drug resistance to these reagents. However, the relationship between autophagy and drug resistance has yet to be defined. Our study showed that EPI induced autophagy in human breast cancer MCF-7 cells. And the EPI-induced autophagy protected MCF-7 cells from EPI-induced apoptosis. Furthermore, autophagy was elevated in EPI-resistant MCF-7 cells (MCF-7er cells), and inhibition of autophagy restored the sensitivity of MCF-7er cells to EPI. Therefore, autophagy is a prosurvival factor and has a role in the development of EPI-acquired resistance in EPI-treated MCF-7 cells. Also, this finding indicates that the use of clinically applicable autophagy inhibitors might be one of the important strategies for breast cancer therapy.     

Autophagy protects breast cancer cells from epirubicin-induced apoptosis and facilitates epirubicin-resistance development

Epirubicin (EPI) is one of the most effective drugs against cancer. But the acquired resistance of cancer cells to EPI is becoming a major obstacle for successful cancer therapy. Recently, some studies have revealed that macroautophagy (here referred to as autophagy) may protect the cancer cell from anticancer drug-induced death, so autophagy might be related to the development of drug resistance to these reagents. However, the relationship between autophagy and drug resistance has yet to be defined. Our study showed that EPI induced autophagy in human breast cancer MCF-7 cells. And the EPI-induced autophagy protected MCF-7 cells from EPI-induced apoptosis. Furthermore, autophagy was elevated in EPI-resistant MCF-7 cells (MCF-7er cells), and inhibition of autophagy restored the sensitivity of MCF-7er cells to EPI. Therefore, autophagy is a prosurvival factor and has a role in the development of EPI-acquired resistance in EPI-treated MCF-7 cells. Also, this finding indicates that the use of clinically applicable autophagy inhibitors might be one of the important strategies for breast cancer therapy.     

CaV3.2 T-type calcium channels are involved in calcium-dependent secretion of neuroendocrine prostate cancer cells

Because prostate cancer is, in its early stages, an androgen-dependent pathology, treatments aiming at decreasing testosterone plasma concentration have been developed for many years now. However, a significant proportion of patients suffer a relapse after a few years of hormone therapy. The androgen-independent stage of prostate cancer has been shown to be associated with the development of neuroendocrine differentiation. We previously demonstrated that neuroendocrine prostate cancer cells derived from LNCaP cells overexpress CaV3.2 T-type voltage-dependent calcium channels. We demonstrate here using prostatic acid phosphatase as a marker of prostate secretion and FM1-43 fluorescence imaging of membrane trafficking that neuroendocrine differentiation is associated with an increase in calcium-dependent secretion which critically relies on CaV3.2 T-type calcium channel activity. In addition, we show that these channels are expressed by neuroendocrine cells in prostate cancer tissues obtained from patients after surgery. We propose that CaV3.2 T-type calcium channel up-regulation may account for the alteration of secretion during prostate cancer development and that these channels, by promoting the secretion of potential mitogenic factors, could participate in the progression of the disease toward an androgen-independent stage.