Prostatic adenocarcinoma CNS parenchymal and dural metastases: alterations in ERG, CHD1 and MAP3K7 expression.

BACKGROUND: Prostatic carcinoma metastatic to dura is commonly encountered at autopsy, but presenting as a dural or, especially parenchymal, brain metastasis during life is far less common. Our group has been interested in two immunohistochemical (IHC) markers previously shown to be downregulated in particularly aggressive primary prostatic carcinomas: CHD1 and MAP3K7. Here we assess protein expression in clinically-relevant CNS metastases. We also assessed how these two markers correlated with the most common genetic alteration in prostate cancer: TMPRSS2 fusion to ERG (40-60% of carcinomas at the primary site), which places ERG expression under the control of the androgen-regulated TMPRSS2 gene, increasing expression. DESIGN: Database query, 2000-2016, identified 16 metastases to dura, 5 to brain parenchyma. RESULTS: Four of five intraparenchymal metastases and 15/16 informative dural-based metastases were ERG-negative (90.5% overall). There was reduced expression of CHD1 in 8/21 and reduced MAP3K7 in 17/21 cases; 7/19 (37%) ERG-negative metastases had dual low expression of CHD1/MAP3K7. ERG-positive cases had high expression of one or both markers. CONCLUSION: Metastatic prostatic carcinoma to CNS demonstrates expression patterns consistent with particularly aggressive behavior. Lower ERG expression in dural and intraparenchymal metastases suggests a possibility that ERG-negative tumors with loss of MAP3K7 may become resistant to standard therapies and diffusely metastasize.

Changes in Susceptibility to Oncolytic Vesicular Stomatitis Virus during Progression of Prostate Cancer.

UNLABELLED: A major challenge to oncolytic virus therapy is that individual cancers vary in their sensitivity to oncolytic viruses, even when these cancers arise from the same tissue type. Variability in response may arise due to differences in the initial genetic lesions leading to cancer development. Alternatively, susceptibility to viral oncolysis may change during cancer progression. These hypotheses were tested using cells from a transgenic mouse model of prostate cancer infected with vesicular stomatitis virus (VSV). Primary cultures from murine cancers derived from prostate-specific Pten deletion contained a mixture of cells that were susceptible and resistant to VSV. Castration-resistant cancers contained a higher percentage of susceptible cells than cancers from noncastrated mice. These results indicate both susceptible and resistant cells can evolve within the same tumor. The role of Pten deletion was further investigated using clonal populations of murine prostate epithelial (MPE) progenitor cells and tumor-derived Pten(-/-) cells. Deletion of Pten in MPE progenitor cells using a lentivirus vector resulted in cells that responded poorly to interferon and were susceptible to VSV infection. In contrast, tumor-derived Pten(-/-) cells expressed higher levels of the antiviral transcription factor STAT1, activated STAT1 in response to VSV, and were resistant to VSV infection. These results suggest that early in tumor development following Pten deletion, cells are primarily sensitive to VSV, but subsequent evolution in tumors leads to development of cells that are resistant to VSV infection. Further evolution in castration-resistant tumors leads to tumors in which cells are primarily sensitive to VSV. IMPORTANCE: There has been a great deal of progress in the development of replication-competent viruses that kill cancer cells (oncolytic viruses). However, a major problem is that individual cancers vary in their sensitivity to oncolytic viruses, even when these cancers arise from the same tissue type. The experiments presented here were to determine whether both sensitive and resistant cells are present in prostate cancers originating from a single genetic lesion in transgenic mice, prostate-specific deletion of the gene for the tumor suppressor Pten. The results indicate that murine prostate cancers are composed of both cells that are sensitive and cells that are resistant to oncolytic vesicular stomatitis virus (VSV). Furthermore, androgen deprivation led to castration-resistant prostate cancers that were composed primarily of cells that were sensitive to VSV. These results are encouraging for the use of VSV for the treatment of prostate cancers that are resistant to androgen deprivation therapy.

Antenatal endotoxin disrupts lung vitamin D receptor and 25-hydroxyvitamin D 1α-hydroxylase expression in the developing rat.

Vitamin D [vit D; 1,25-(OH)2D] treatment improves survival and lung alveolar and vascular growth in an experimental model of bronchopulmonary dysplasia (BPD) after antenatal exposure to endotoxin (ETX). However, little is known about lung-specific 1,25-(OH)2D3 regulation during development, especially regarding maturational changes in lung-specific expression of the vitamin D receptor (VDR), 1α-hydroxylase (1α-OHase), and CYP24A1 during late gestation and the effects of antenatal ETX exposure on 1,25-(OH)2D3 metabolism in the lung. We hypothesized that vit D regulatory proteins undergo maturation regulation in the late fetal and early neonatal lung and that prenatal exposure to ETX impairs lung growth partly through abnormal endogenous vit D metabolism. Normal fetal rat lungs were harvested between embryonic day 15 and postnatal day 14. Lung homogenates were assayed for VDR, 1α-OHase, and CYP24A1 protein contents by Western blot analysis. Fetal rats were injected on embryonic day 20 with intra-amniotic ETX, ETX + 1,25-(OH)2D3, or saline and delivered 2 days later. Pulmonary artery endothelial cells (PAECs) from fetal sheep were assessed for VDR, 1α-OHase, and CYP24A1 expression after treatment with 25-(OH)D3, 1,25-(OH)2D3, ETX, ETX + 25-(OH)D3, or ETX + 1,25-(OH)2D3. We found that lung VDR, 1α-OHase, and CYP2741 protein expression dramatically increase immediately before birth (P < 0.01 vs. early fetal values). Antenatal ETX increases CYP24A1 expression (P < 0.05) and decreases VDR and 1α-OHase expression at birth (P < 0.001), but these changes are prevented with concurrent vit D treatment (P < 0.001). ETX-induced reduction of fetal PAEC growth and tube formation and lung 1α-OHase expression are prevented by vit D treatment (P < 0.001). We conclude that lung VDR, 1α-OHase, and CYP24A1 protein content markedly increase before birth and that antenatal ETX disrupts lung vit D metabolism through downregulation of VDR and increased vit D catabolic enzyme expression, including changes in developing endothelium. We speculate that endogenous vitamin D metabolism modulates normal fetal lung development and that prenatal disruption of vit D signaling may contribute to impaired postnatal lung growth at least partly through altered angiogenic signaling.

Silibinin prevents prostate cancer cell-mediated differentiation of naïve fibroblasts into cancer-associated fibroblast phenotype by targeting TGF ß2.

Tumor microenvironment (TM) is an essential element in prostate cancer (PCA), offering unique opportunities for its prevention. TM includes naïve fibroblasts that are recruited by nascent neoplastic lesion and altered into 'cancer-associated fibroblasts' (CAFs) that promote PCA. A better understanding and targeting of interaction between PCA cells and fibroblasts and inhibiting CAF phenotype through non-toxic agents are novel approaches to prevent PCA progression. One well-studied cancer chemopreventive agent is silibinin, and thus, we examined its efficacy against PCA cells-mediated differentiation of naïve fibroblasts into a myofibroblastic-phenotype similar to that found in CAFs. Silibinin's direct inhibitory effect on the phenotype of CAFs derived directly from PCA patients was also assessed. Human prostate stromal cells (PrSCs) exposed to control conditioned media (CCM) from human PCA PC3 cells showed more invasiveness, with increased alpha-smooth muscle actin (α-SMA) and vimentin expression, and differentiation into a phenotype we identified in CAFs. Importantly, silibinin (at physiologically achievable concentrations) inhibited α-SMA expression and invasiveness in differentiated fibroblasts and prostate CAFs directly, as well as indirectly by targeting PCA cells. The observed increase in α-SMA and CAF-like phenotype was transforming growth factor (TGF) ß2 dependent, which was strongly inhibited by silibinin. Furthermore, induction of α-SMA and CAF phenotype by CCM were also strongly inhibited by a TGFß2-neutralizing antibody. The inhibitory effect of silibinin on TGFß2 expression and CAF-like biomarkers was also observed in PC3 tumors. Together, these findings highlight the potential usefulness of silibinin in PCA prevention through targeting the CAF phenotype in the prostate TM.

ProstaMine: a bioinformatics tool for identifying subtype-specific co-alterations associated with aggressiveness in prostate cancer.

Background: Prostate cancer is a leading cause of cancer-related deaths among men, marked by heterogeneous clinical and molecular characteristics. The complexity of the molecular landscape necessitates tools for identifying multi-gene co-alteration patterns that are associated with aggressive disease. The identification of such gene sets will allow for deeper characterization of the processes underlying prostate cancer progression and potentially lead to novel strategies for treatment. Methods: We developed ProstaMine to systematically identify co-alterations associated with aggressiveness in prostate cancer molecular subtypes defined by high-fidelity alterations in primary prostate cancer. ProstaMine integrates genomic, transcriptomic, and clinical data from five primary and one metastatic prostate cancer cohorts to prioritize co-alterations enriched in metastatic disease and associated with disease progression. Results: Integrated analysis of primary tumors defined a set of 17 prostate cancer alterations associated with aggressive characteristics. We applied ProstaMine to NKX3-1-loss and RB1-loss tumors and identified subtype-specific co-alterations associated with metastasis and biochemical relapse in these molecular subtypes. In NKX3-1-loss prostate cancer, ProstaMine identified novel subtype-specific co-alterations known to regulate prostate cancer signaling pathways including MAPK, NF-kB, p53, PI3K, and Sonic hedgehog. In RB1-loss prostate cancer, ProstaMine identified novel subtype-specific co-alterations involved in p53, STAT6, and MHC class I antigen presentation. Co-alterations impacting autophagy were noted in both molecular subtypes. Conclusion: ProstaMine is a method to systematically identify novel subtype-specific co-alterations associated with aggressive characteristics in prostate cancer. The results from ProstaMine provide insights into potential subtype-specific mechanisms of prostate cancer progression which can be formed into testable experimental hypotheses. ProstaMine is publicly available at: https://bioinformatics.cuanschutz.edu/prostamine.

Genetic markers associated with early cancer-specific mortality following prostatectomy.

BACKGROUND: This study sought to identify novel effectors and markers of localized but potentially life-threatening prostate cancer (PCa), by evaluating chromosomal copy number alterations (CNAs) in tumors from patients who underwent prostatectomy and correlating these with clinicopathologic features and outcome. METHODS: CNAs in tumor DNA samples from 125 patients in the discovery cohort who underwent prostatectomy were assayed with high-resolution Affymetrix 6.0 single-nucleotide polymorphism microarrays and then analyzed using the Genomic Identification of Significant Targets in Cancer (GISTIC) algorithm. RESULTS: The assays revealed 20 significant regions of CNAs, 4 of them novel, and identified the target genes of 4 of the alterations. By univariate analysis, 7 CNAs were significantly associated with early PCa-specific mortality. These included gains of chromosomal regions that contain the genes MYC, ADAR, or TPD52 and losses of sequences that incorporate SERPINB5, USP10, PTEN, or TP53. On multivariate analysis, only the CNAs of PTEN (phosphatase and tensin homolog) and MYC (v-myc myelocytomatosis viral oncogene homolog) contributed additional prognostic information independent of that provided by pathologic stage, Gleason score, and initial prostate-specific antigen level. Patients whose tumors had alterations of both genes had a markedly elevated risk of PCa-specific mortality (odds ratio = 53; 95% CI = 6.92-405, P = 1 × 10(-4)). Analyses of 333 tumors from 3 additional distinct patient cohorts confirmed the relationship between CNAs of PTEN and MYC and lethal PCa. CONCLUSIONS: This study identified new CNAs and genes that likely contribute to the pathogenesis of localized PCa and suggests that patients whose tumors have acquired CNAs of PTEN, MYC, or both have an increased risk of early PCa-specific mortality.

Management of Advanced Prostate Cancer in the Precision Oncology Era.

Prostate cancer (PC) is the second leading cause of cancer death in men in the United States. While diversified and improved treatment options for aggressive PC have improved patient outcomes, metastatic castration-resistant prostate cancer (mCRPC) remains incurable and an area of investigative therapeutic interest. This review will cover the seminal clinical data supporting the indication of new precision oncology-based therapeutics and explore their limitations, present utility, and potential in the treatment of PC. Systemic therapies for high-risk and advanced PC have experienced significant development over the past ten years. Biomarker-driven therapies have brought the field closer to the goal of being able to implement precision oncology therapy for every patient. The tumor agnostic approval of pembrolizumab (a PD-1 inhibitor) marked an important advancement in this direction. There are also several PARP inhibitors indicated for patients with DNA damage repair deficiencies. Additionally, theranostic agents for both imaging and treatment have further revolutionized the treatment landscape for PC and represent another advancement in precision medicine. Radiolabeled prostate-specific membrane antigen (PSMA) PET/CT is rapidly becoming a standard of care for diagnosis, and PSMA-targeted radioligand therapies have gained recent FDA approval for metastatic prostate cancer. These advances in precision-based oncology are detailed in this review.

Effect of dietary polyunsaturated fatty acids on castration-resistant Pten-null prostate cancer.

A common treatment of advanced prostate cancer involves the deprivation of androgens. Despite the initial response to hormonal therapy, eventually all the patients relapse. In the present study, we sought to determine whether dietary polyunsaturated fatty acid (PUFA) affects the development of castration-resistant prostate cancer. Cell culture, patient tissue microarray, allograft, xenograft, prostate-specific Pten knockout and omega-3 desaturase transgenic mouse models in conjunction with dietary manipulation, gene knockdown and knockout approaches were used to determine the effect of dietary PUFA on castration-resistant Pten-null prostate cancer. We found that deletion of Pten increased androgen receptor (AR) expression and Pten-null prostate cells were castration resistant. Omega-3 PUFA slowed down the growth of castration-resistant tumors as compared with omega-6 PUFA. Omega-3 PUFA decreased AR protein to a similar extent in tumor cell cytosolic and nuclear fractions but had no effect on AR messenger RNA level. Omega-3 PUFA treatment appeared to accelerate AR protein degradation, which could be blocked by proteasome inhibitor MG132. Knockdown of AR significantly slowed down prostate cancer cell proliferation in the absence of androgens. Our data suggest that omega-3 PUFA inhibits castration-resistant prostate cancer in part by accelerating proteasome-dependent degradation of the AR protein. Dietary omega-3 PUFA supplementation in conjunction with androgen ablation may significantly delay the development of castration-resistant prostate cancer in patients compared with androgen ablation alone.

Hydroxyproline metabolism in mouse models of primary hyperoxaluria.

Primary hyperoxaluria type 1 (PH1) and type 2 (PH2) are rare genetic diseases that result from deficiencies in glyoxylate metabolism. The increased oxalate synthesis that occurs can lead to kidney stone formation, deposition of calcium oxalate in the kidney and other tissues, and renal failure. Hydroxyproline (Hyp) catabolism, which occurs mainly in the liver and kidney, is a prominent source of glyoxylate and could account for a significant portion of the oxalate produced in PH. To determine the sensitivity of mouse models of PH1 and PH2 to Hyp-derived oxalate, animals were fed diets containing 1% Hyp. Urinary excretions of glycolate and oxalate were used to monitor Hyp catabolism and the kidneys were examined to assess pathological changes. Both strains of knockout (KO) mice excreted more oxalate than wild-type (WT) animals with Hyp feeding. After 4 wk of Hyp feeding, all mice deficient in glyoxylate reductase/hydroxypyruvate reductase (GRHPR KO) developed severe nephrocalcinosis in contrast to animals deficient in alanine-glyoxylate aminotransferase (AGXT KO) where nephrocalcinosis was milder and with a lower frequency. Plasma cystatin C measurements over 4-wk Hyp feeding indicated no significant loss of renal function in WT and AGXT KO animals, and significant and severe loss of renal function in GRHPR KO animals after 2 and 4 wk, respectively. These data suggest that GRHPR activity may be vital in the kidney for limiting the conversion of Hyp-derived glyoxylate to oxalate. As Hyp catabolism may make a major contribution to the oxalate produced in PH patients, Hyp feeding in these mouse models should be useful in understanding the mechanisms associated with calcium oxalate deposition in the kidney.

MIRO2 Regulates Prostate Cancer Cell Growth via GCN1-Dependent Stress Signaling.

There is a continued need to identify novel therapeutic targets to prevent the mortality associated with prostate cancer. In this context, mitochondrial Rho GTPase 2 (MIRO2) mRNA was upregulated in metastatic prostate cancer compared with localized tumors, and higher MIRO2 levels were correlated with poor patient survival. Using human cell lines that represent androgen-independent or -sensitive prostate cancer, we showed that MIRO2 depletion impaired cell growth, colony formation, and tumor growth in mice. Network analysis of MIRO2's binding partners identified metabolism and cellular responses to extracellular stimuli as top overrepresented pathways. The top hit on our screen, General Control Nonderepressible 1 (GCN1), was overexpressed in prostate cancer, and interacted with MIRO2 in prostate cancer cell lines and in primary prostate cancer cells. Functional analysis of MIRO2 mutations present in patients with prostate cancer led to the identification of MIRO2 159L, which increased GCN1 binding. Importantly, MIRO2 was necessary for efficient GCN1-mediated GCN2 kinase signaling and induction of the transcription factor activating transcription factor 4 (ATF4) levels. Further, MIRO2's effect on regulating prostate cancer cell growth was mediated by ATF4. Finally, levels of activated GCN2 and ATF4 were correlated with MIRO2 expression in prostate cancer xenografts. Both MIRO2 and activated GCN2 levels were higher in hypoxic areas of prostate cancer xenografts. Overall, we propose that targeting the MIRO2-GCN1 axis may be a valuable strategy to halt prostate cancer growth. IMPLICATIONS: MIRO2/GCN1/GCN2 constitute a novel mitochondrial signaling pathway that controls androgen-independent and androgen-sensitive prostate cancer cell growth.

Regulation of Skp2 levels by the Pim-1 protein kinase.

The Pim-1 protein kinase plays an important role in regulating both cell growth and survival and enhancing transformation by multiple oncogenes. The ability of Pim-1 to regulate cell growth is mediated, in part, by the capacity of this protein kinase to control the levels of the p27, a protein that is a critical regulator of cyclin-dependent kinases that mediate cell cycle progression. To understand how Pim-1 is capable of regulating p27 protein levels, we focused our attention on the SCF(Skp2) ubiquitin ligase complex that controls the rate of degradation of this protein. We found that expression of Pim-1 increases the level of Skp2 through direct binding and phosphorylation of multiple sites on this protein. Along with known Skp2 phosphorylation sites including Ser(64) and Ser(72), we have identified Thr(417) as a unique Pim-1 phosphorylation target. Phosphorylation of Thr(417) controls the stability of Skp2 and its ability to degrade p27. Additionally, we found that Pim-1 regulates the anaphase-promoting complex or cyclosome (APC/C complex) that mediates the ubiquitination of Skp2. Pim-1 phosphorylates Cdh1 and impairs binding of this protein to another APC/C complex member, CDC27. These modifications inhibit Skp2 from degradation. Marked increases in Skp2 caused by these mechanisms lower cellular p27 levels. Consistent with these observations, we show that Pim-1 is able to cooperate with Skp2 to signal S phase entry. Our data reveal a novel Pim-1 kinase-dependent signaling pathway that plays a crucial role in cell cycle regulation.

Fine mapping association study and functional analysis implicate a SNP in MSMB at 10q11 as a causal variant for prostate cancer risk.

A single nucleotide polymorphism (SNP) at 10q11 (rs10993994) in the 5' region of the MSMB gene was recently implicated in prostate cancer risk in two genome-wide association studies. To identify possible causal variants in the region, we genotyped 16 tagging SNPs and imputed 29 additional SNPs in approximately 65 kb genomic region at 10q11 in a Swedish population-based case-control study (CAncer of the Prostate in Sweden), including 2899 cases and 1722 controls. We found evidence for two independent loci, separated by a recombination hotspot, associated with prostate cancer risk. Among multiple significant SNPs at locus 1, the initial SNP rs10993994 was most significant. Importantly, using an MSMB promoter reporter assay, we showed that the risk allele of this SNP had only 13% of the promoter activity of the wild-type allele in a prostate cancer model, LNCaP cells. Curiously, the second, novel locus (locus 2) was within NCOA4 (also known as ARA70), which is known to enhance androgen receptor transcriptional activity in prostate cancer cells. However, its association was only weakly confirmed in one of the three additional study populations. The observations that rs10993994 is the strongest associated variant in the region and its risk allele has a major effect on the transcriptional activity of MSMB, a gene with previously described prostate cancer suppressor function, together suggest the T allele of rs10993994 as a potential causal variant at 10q11 that confers increased risk of prostate cancer.

Androgen Receptor Signaling in Prostate Cancer Genomic Subtypes.

While many prostate cancer (PCa) cases remain indolent and treatable, others are aggressive and progress to the metastatic stage where there are limited curative therapies. Androgen receptor (AR) signaling remains an important pathway for proliferative and survival programs in PCa, making disruption of AR signaling a viable therapy option. However, most patients develop resistance to AR-targeted therapies or inherently never respond. The field has turned to PCa genomics to aid in stratifying high risk patients, and to better understand the mechanisms driving aggressive PCa and therapy resistance. While alterations to the AR gene itself occur at later stages, genomic changes at the primary stage can affect the AR axis and impact response to AR-directed therapies. Here, we review common genomic alterations in primary PCa and their influence on AR function and activity. Through a meta-analysis of multiple independent primary PCa databases, we also identified subtypes of significantly co-occurring alterations and examined their combinatorial effects on the AR axis. Further, we discussed the subsequent implications for response to AR-targeted therapies and other treatments. We identified multiple primary PCa genomic subtypes, and given their differing effects on AR activity, patient tumor genetics may be an important stratifying factor for AR therapy resistance.

MAP3K7 Loss Drives Enhanced Androgen Signaling and Independently Confers Risk of Recurrence in Prostate Cancer with Joint Loss of CHD1.

Prostate cancer genomic subtypes that stratify aggressive disease and inform treatment decisions at the primary stage are currently limited. Previously, we functionally validated an aggressive subtype present in 15% of prostate cancer characterized by dual deletion of MAP3K7 and CHD1. Recent studies in the field have focused on deletion of CHD1 and its role in androgen receptor (AR) chromatin distribution and resistance to AR-targeted therapy; however, CHD1 is rarely lost without codeletion of MAP3K7. Here, we show that in the clinically relevant context of co-loss of MAP3K7 and CHD1 there are significant, collective changes to aspects of AR signaling. Although CHD1 loss mainly impacts the expansion of the AR cistrome, loss of MAP3K7 drives increased AR target gene expression. Prostate cancer cell line models engineered to cosuppress MAP3K7 and CHD1 also demonstrated increased AR-v7 expression and resistance to the AR-targeting drug enzalutamide. Furthermore, we determined that low protein expression of both genes is significantly associated with biochemical recurrence (BCR) in a clinical cohort of radical prostatectomy specimens. Low MAP3K7 expression, however, was the strongest independent predictor for risk of BCR over all other tested clinicopathologic factors including CHD1 expression. Collectively, these findings illustrate the importance of MAP3K7 loss in a molecular subtype of prostate cancer that poses challenges to conventional therapeutic approaches. IMPLICATIONS: These findings strongly implicate MAP3K7 loss as a biomarker for aggressive prostate cancer with significant risk for recurrence that poses challenges for conventional androgen receptor-targeted therapies.

MicroRNA-101 negatively regulates Ezh2 and its expression is modulated by androgen receptor and HIF-1alpha/HIF-1beta.

BACKGROUND: In prostate cancer (PCa), the common treatment involving androgen ablation alleviates the disease temporarily, but results in the recurrence of highly aggressive and androgen-independent metastatic cancer. Therefore, more effective therapeutic approaches are needed. It is known that aberrant epigenetics contributes to prostate malignancy. Unlike genetic changes, these epigenetic alterations are reversible, which makes them attractive targets in PCa therapy to impede cancer progression. As a histone methyltransferase, Ezh2 plays an essential role in epigenetic regulation. Since Ezh2 is overexpressed and acts as an oncogene in PCa, it has been proposed as a bona fide target of PCa therapy. MicroRNAs (miRNAs) regulate gene expression through modulating protein translation. Recently, the contribution of miRNAs in cancer development is increasingly appreciated. In this report, we present our study showing that microRNA-101 (miR-101) inhibits Ezh2 expression and differentially regulates prostate cancer cells. In addition, the expression of miR-101 alters upon androgen treatment and HIF-1alpha/HIF-1beta induction. RESULT: In our reporter assays, both miR-101 and miR-26a inhibit the expression of a reporter construct containing the 3'-UTR of Ezh2. When ectopically expressed in PC-3, DU145 and LNCaP cells, miR-101 inhibits endogenous Ezh2 expression in all three cell lines, while miR-26a only decreases Ezh2 in DU145. Ectopic miR-101 reduces the invasion ability of PC-3 cells, while restored Ezh2 expression rescues the invasiveness of PC-3 cells. Similarly, miR-101 also inhibits cell invasion and migration of DU145 and LNCaP cells, respectively. Interestingly, ectopic miR-101 exhibits differential effects on the proliferation of PC-3, DU-145 and LNCaP cells and also causes morphological changes of LNCaP cells. In addition, the expression of miR-101 is regulated by androgen receptor and HIF-1alpha/HIF-1beta. While HIF-1alpha/HIF-1beta induced by deferoxamine mesylate (DFO) decreases miR-101 levels, the overall effects of R-1881 on miR-101 expression are stimulatory. CONCLUSIONS: This study indicates that miR-101 targets Ezh2 and decreases the invasiveness of PCa cells, suggesting that miR-101 introduction is a potential therapeutic strategy to combat PCa. MiR-101 differentially regulates prostate cell proliferation. Meanwhile, the expression of miR-101 is also modulated at different physiological conditions, such as androgen stimulation and HIF-1alpha/HIF-1beta induction.

1,25-dihydroxyvitamin D(3) and PI3K/AKT inhibitors synergistically inhibit growth and induce senescence in prostate cancer cells.

BACKGROUND: 1-Alpha, 25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) inhibits proliferation of multiple cancer cell types including prostate cells and upregulates p21 and/or p27, while loss of Pten and PI3K/AKT activation stimulates survival and downregulates p21 and p27. We hypothesized that inhibition of the PI3K/AKT pathway synergizes with the antiproliferative signaling of 1,25(OH)(2)D(3). METHODS: Viability, cell cycle and senescence of cells were evaluated upon combinational treatment with 1,25(OH)(2)D(3) and pharmacological PI3K/AKT inhibitors. RESULTS: Pharmacological inhibitors of PI3K or Akt and 1,25(OH)(2)D(3) synergistically inhibited growth of DU145, LNCaP, primary human prostate cancer cell strains and Pten null mouse prostatic epithelial cells (MPEC). The inhibitors used included API-2 (Triciribine) and GSK690693 which are currently in clinical trials for treatment of cancer. A novel mechanism for antiproliferative effects of 1,25(OH)(2)D(3) in prostate cells, induction of senescence, was discovered. Combination of 1,25(OH)(2)D(3) and AKT inhibitor cooperated to induce G(1) arrest, senescence, and p21 levels in prostate cancer cells. As AKT is commonly activated by PTEN loss, we evaluated the role of Pten in responsiveness to 1,25(OH)(2)D(3) using shRNA knockdown and by in vitro knockout of Pten. MPEC that lost Pten expression remained sensitive to the antiproliferative action of 1,25(OH)(2)D(3), and showed higher degree of synergism between AKT inhibitor and 1,25(OH)(2)D(3) compared to Pten-expressing counterparts. CONCLUSIONS: These findings provide the rationale for the development of therapies utilizing 1,25(OH)(2)D(3) or its analogs combined with inhibition of PI3K/AKT for the treatment of prostate cancer.

The interplay of autophagy and non-apoptotic cell death pathways.

Autophagy, the process of macromolecular degradation through the lysosome, has been extensively studied for the past decade or two. Autophagy can regulate cell death, especially apoptosis, through selective degradation of both positive and negative apoptosis regulators. However, multiple other programmed cell death pathways exist. As knowledge of these other types of cell death expand, it has been suggested that they also interact with autophagy. In this review, we discuss the molecular mechanisms that comprise three non-apoptotic forms of cell death (necroptosis, pyroptosis and ferroptosis) focusing on how the autophagy machinery regulates these different cell death mechanisms through (i) its degradative functions, i.e., true autophagy, and (ii) other non-degradative functions of the autophagy machinery such as serving as a signaling scaffold or by participating in other autophagy-independent cellular processes.

MAP3K7 and CHD1 Are Novel Mediators of Resistance to Oncolytic Vesicular Stomatitis Virus in Prostate Cancer Cells.

A key principle of oncolytic viral therapy is that many cancers develop defects in their antiviral responses, making them more susceptible to virus infection. However, some cancers display resistance to viral infection. Many of these resistant cancers constitutively express interferon-stimulated genes (ISGs). The goal of these experiments was to determine the role of two tumor suppressor genes, MAP3K7 and CHD1, in viral resistance and ISG expression in PC3 prostate cancer cells resistant to oncolytic vesicular stomatitis virus (VSV). MAP3K7 and CHD1 are often co-deleted in aggressive prostate cancers. Silencing expression of MAP3K7 and CHD1 in PC3 cells increased susceptibility to the matrix (M) gene mutant M51R-VSV, as shown by increased expression of viral genes, increased yield of progeny virus, and reduction of tumor growth in nude mice. Silencing MAP3K7 alone had a greater effect on virus susceptibility than did silencing CHD1. Silencing MAP3K7 and CHD1 decreased constitutive expression of ISG mRNAs and proteins, whereas silencing MAP3K7 alone decreased expression of ISG proteins, but actually increased expression of ISG mRNAs. These results suggest a role for the protein product of MAP3K7, transforming growth factor ß-activated kinase 1 (TAK1), in regulating translation of ISG mRNAs and a role of CHD1 in maintaining the transcription of ISGs.

Association of reported prostate cancer risk alleles with PSA levels among men without a diagnosis of prostate cancer.

BACKGROUND: Prostate specific antigen (PSA) is widely used for prostate cancer screening but its levels are influenced by many non cancer-related factors. The goal of the study is to estimate the effect of genetic variants on PSA levels. METHODS: We evaluated the association of SNPs that were reported to be associated with prostate cancer risk in recent genome-wide association studies with plasma PSA levels in a Swedish study population, including 1,722 control subjects without a diagnosis of prostate cancer. RESULTS: Of the 16 SNPs analyzed in control subjects, significant associations with PSA levels (P < or = 0.05) were found for six SNPs. These six SNPs had a cumulative effect on PSA levels; the mean PSA levels in men were almost twofold increased across increasing quintile of number of PSA associated alleles, P-trend = 3.4 x 10(-14). In this Swedish study population risk allele frequencies were similar among T1c case patients (cancer detected by elevated PSA levels alone) as compared to T2 and above prostate cancer case patients. CONCLUSIONS: Results from this study may have two important clinical implications. The cumulative effect of six SNPs on PSA levels suggests genetic-specific PSA cutoff values may be used to improve the discriminatory performance of this test for prostate cancer; and the dual associations of these SNPs with PSA levels and prostate cancer risk raise a concern that some of reported prostate cancer risk-associated SNPs may be confounded by the prevalent use of PSA screening.

Late diagnosis of primary hyperoxaluria type 2 in the adult: effect of a novel mutation in GRHPR gene on enzymatic activity and molecular modeling.

PURPOSE: Genetic causes of nephrolithiasis are underestimated. Primary hyperoxaluria type 2 is a rare autosomal recessive disease caused by mutations in the GRHPR gene, leading to an accumulation of oxalate and L-glycerate with recurrent kidney stone formation and nephrocalcinosis, and the later development of renal failure and systemic oxalate depositions. We studied the effects of a novel GRHPR mutation on GRHPR enzymatic activity and molecular modeling. MATERIALS AND METHODS: Genomic DNA from a 50-year-old male with a late diagnosis of primary hyperoxaluria type 2 was extracted, analyzed and compared with the established human GRHPR gene sequence. Restriction enzyme analysis of the patient, 30 healthy controls and 30 patients with nephrolithiasis of various causes was done to confirm the presence of the mutation. GRHPR activity was analyzed by site directed mutagenesis of WT and mutant clones. We studied the effects of the mutation on enzymatic molecular modeling. RESULTS: We found the novel homozygous single missense mutation A975G in exon 9, creating an amino acid change from asparagine to aspartic acid in position 312. No mutations were detected in restriction enzyme analysis in all 30 healthy controls and 30 patients with nephrolithiasis of various causes. Transfected cells with the mutant clone showed abolished GRHPR activity. Molecular modeling studies revealed that the mutation was likely to disrupt the correct folding of the GRHPR substrate binding domain, hence affecting the enzyme active site. CONCLUSIONS: Primary hyperoxaluria type 2 should be considered in patients at adult stone clinics who have had a history of nephrolithiasis since childhood, especially in those with consanguineous parents. Biochemical analysis followed by mutation identification should be the approach for making the definitive diagnosis of primary hyperoxaluria type 2.