Michael J. Stower

York Hospital, New York, New York, United States

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Publications (35)163.33 Total impact

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    ABSTRACT: Prostate cancer (CaP) is mostly composed of luminal-like differentiated cells, but contains a small subpopulation of basal cells (including stem-like cells), which can proliferate and differentiate into luminal-like cells. In cancers, CpG island hypermethylation has been associated with gene downregulation, but the causal relationship between the two phenomena is still debated. Here we clarify the origin and function of CpG island hypermethylation in CaP, in the context of a cancer cell hierarchy and epithelial differentiation, by analysis of separated basal and luminal cells from cancers. For a set of genes (including GSTP1) that are hypermethylated in CaP, gene downregulation is the result of cell differentiation and is not cancer specific. Hypermethylation is however seen in more differentiated cancer cells and is promoted by hyperproliferation. These genes are maintained as actively expressed and methylation-free in undifferentiated CaP cells, and their hypermethylation is not essential for either tumour development or expansion. We present evidence for the causes and the dynamics of CpG island hypermethylation in CaP, showing that, for a specific set of genes, promoter methylation is downstream of gene downregulation and is not a driver of gene repression, while gene repression is a result of tissue-specific differentiation.Cell Death and Differentiation advance online publication, 24 January 2014; doi:10.1038/cdd.2013.202.
    No preview · Article · Jan 2014 · Cell death and differentiation
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    ABSTRACT: IL-6 overexpression and constitutive STAT3 activation occur in many cancers including prostate cancer. However, their contribution to prostate stem and progenitor cells has not been explored. In this study, we show that stem-like cells from prostate cancer patients secrete higher levels of IL-6 than their counterparts in non-neoplastic prostate. Tumor grade did not influence the levels of expression or secretion. Stem-like and progenitor cells expressed the IL-6 receptor gp80 with concomitant expression of pSTAT3. Blockade of activated STAT3, by either anti-IL-6 antibody siltuximab (CNTO 328) or LLL12, a specific ρSTAT3 inhibitor, suppressed the clonogenicity of the stem-like cells in patients with high grade disease. In a murine xenograft model utilized to determine the in vivo effects of ρSTAT3 suppression, LLL12 treatment effectively abolished outgrowth of a patient-derived castrate-resistant tumor. Our results indicate that the most primitive cells in prostate cancer require ρSTAT3 for survival, rationalizing STAT3 as a therapeutic target to treat advanced prostate cancer.
    Full-text · Article · Aug 2013 · Cancer Research
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    ABSTRACT: The outcome for patients with advanced metastatic and recurrent prostate cancer is still poor. Therefore, new chemotherapeutics are required, especially for killing cancer stem cells that are thought to be responsible for disease recurrence. In this study, we screened the effect of a novel palladium-based anticancer agent (Pd complex) against six different prostate cancer cell lines, and primary cultures from seven Gleason 6/7 prostate cancer, three Gleason 8/9 prostate cancer and four benign prostate hyperplasia patient samples, as well as cancer stem cells selected from primary cultures. MTT and ATP viability assays were used to assess cell growth and flow cytometry to assess cell cycle status. In addition, immunofluorescence was used to detect γH2AX nuclear foci, indicative of DNA damage, and Western blotting to assess the induction of apoptosis and autophagy. The Pd complex showed a powerful growth-inhibitory effect against both cell lines and primary cultures. More importantly, it successfully reduced the viability of cancer stem cells as first reported in this study. The Pd complex induced DNA damage and differentially induced evidence of cell death, as well as autophagy. In conclusion, this novel agent may be promising for use against the bulk of the tumour cell population as well as the prostate cancer stem cells, which are thought to be responsible for the resistance of metastatic prostate cancer to chemotherapy. This study also indicates that the combined use of the Pd complex with an autophagy modulator may be a more promising approach to treat prostate cancer. In addition, the differential effects observed between cell lines and primary cells emphasise the importance of the model used to test novel drugs including its genetic background, and indeed the necessity of using cells cultured from patient samples.
    Full-text · Article · May 2013 · PLoS ONE
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    ABSTRACT: The mouse haematopoietic stem cell (SC) regulator Latexin (LXN) is the only known homologue of the retinoic acid receptor responder 1 (RARRES1) gene. Both genes lie adjacent on chromosome 3 and differ mostly by the presence of a transmembrane domain in RARRES1. Despite their homology, it is not known whether they possess similar regulatory mechanisms, cellular localization and function. Here, we identified RARRES1 and LXN as highly significantly downregulated genes in human prostate SCs, whose expression was induced by the pro-differentiation agent all-trans retinoic acid (atRA). AtRA induced expression in the most differentiated cells compared with the SC fraction, suggesting that this subpopulation was less responsive to atRA. Small interfering RNA suppression of RARRES1 and LXN enhanced the SC properties of primary prostate cultures, as shown by a significant increase in their colony-forming ability. Expression of both RARRES1 and LXN was co-ordinately repressed by DNA methylation in prostate cancer cell lines and inhibition of RARRES1 and LXN increased the invasive capacity of primary prostate cultures, which also fully rescued an inhibitory effect induced by atRA. Moreover, we showed that RARRES1 and LXN reside within different sub-cellular compartments, providing evidence that RARRES1 is not a plasma membrane protein as previously supposed but is located primarily in the endoplasmic reticulum; whereas LXN was detected in the nucleus of prostate epithelial cells. Thus, LXN and RARRES1 are potential tumour suppressor genes, which are co-ordinately regulated, SC-silenced genes functioning to suppress invasion and colony-forming ability of prostate cancer cells; yet the proteins reside within different sub-cellular compartments.
    Full-text · Article · Apr 2013 · Oncogenesis
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    ABSTRACT: While chromosomal translocations have a fundamental role in the development of several human leukaemias, their role in solid tumour development has been somewhat more controversial. Recently, it was shown that up to 80% of prostate tumours harbour at least one such gene fusion, and that the most common fusion event, between the prostate-specific TMPRSS2 gene and the ERG oncogene, is a critical, and probably early factor in prostate cancer development. Here we demonstrate the presence and expression of this significant chromosomal rearrangement in prostate cancer stem cells. Moreover, we show that in the prostate epithelial hierarchy from both normal and tumour tissues, TMPRSS2 transcription is subjected to tight monoallelic regulation, which is retained upon asymmetric division and relaxed during epithelial cell differentiation. The presence and expression of TMPRSS2/ERG in prostate stem cells would provide ERG-driven survival advantages, allowing maintenance of this mutated genotype.
    No preview · Article · Mar 2013 · Nature Communications
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    ABSTRACT: While chromosomal translocations have a fundamental role in the development of several human leukaemias, their role in solid tumour development has been somewhat more controversial. Recently, it was shown that up to 80% of prostate tumours harbour at least one such gene fusion, and that the most common fusion event, between the prostate-specific TMPRSS2 gene and the ERG oncogene, is a critical, and probably early factor in prostate cancer development. Here we demonstrate the presence and expression of this significant chromosomal rearrangement in prostate cancer stem cells. Moreover, we show that in the prostate epithelial hierarchy from both normal and tumour tissues, TMPRSS2 transcription is subjected to tight monoallelic regulation, which is retained upon asymmetric division and relaxed during epithelial cell differentiation. The presence and expression of TMPRSS2/ERG in prostate stem cells would provide ERG-driven survival advantages, allowing maintenance of this mutated genotype.
    No preview · Article · Mar 2013 · Nature Communications

  • No preview · Article · Jul 2012 · European Journal of Cancer

  • No preview · Article · Jun 2012 · Cancer Research
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    ABSTRACT: The aim of present investigation is to elucidate the complex stem cell dynamics within prostate cancer, which can be utilized to design novel diagnostic and therapeutic strategies for the management of prostate cancer. In order to determine precise transcriptional and microRNA regulatory mechanisms modulating stem cell self-renewal and differentiation, unique cellular assays have been developed in our lab that utilize homogeneous fractionated cell populations enriched from primary patient prostate cultures. Using a prospective bioinformatic analysis of gene expression data from Birnie et. al., 2008, we have identified LCN2, CEACAM6, and S100p as candidate genes for regulation of prostate stem cell differentiation. Their over-expression in differentiated cells, as compared to stem cells, was validated in respective cells enriched from cultures obtained from BPH, cancer and castration resistant prostate cancer samples and from primary human prostate cancer xenografts. Interestingly, the analysis of 25,000 published human Affymetrix microarray chips revealed that LCN2, CEACAM6, and S100p have a more similar expression pattern than that of any other genes in the entire human genome, suggesting that they may have common function and are co-regulated. Indeed, the promoter analysis showed that the promoters (1kb from TSS) of all these genes have common binding sites for 40 transcription factors with very high affinity and P < 0.001. Most of these transcription factors have a well-documented role in cell differentiation (RA, AR, and NANOG) and prostate carcinogenesis (NF-kB). Significant up-regulation in the expression of these genes in prostate cell lines after treatment with all-trans retinoic acid and androgen analogue R1881 further suggested the role of AR and RA in prostate differentiation. Along with transcriptional regulation, Agilent v3 miRNA microarray data revealed obviously distinct miRNA expression profiles in stem and differentiated prostate epithelial cells, confirming crucial role of miRNA in main taining epithelial hierarchies, in prostate. We anticipate that evaluation of integrative transcriptional (LCN2-CEACAM6-S100p)-microRNA regulatory network, with further functional studies, will comprehensively establish a detailed knowledge base for potential regulatory mechanisms involved in prostate stem cell and prostate cancer stem cell differentiation. These insights will be valuable to formulate efficient ‘differentiation therapy’ for the management of prostate cancer.
    No preview · Conference Paper · Apr 2012
  • Joby Taylor · Michael J. Stower

    No preview · Article · Sep 2011 · British Journal of Medical and Surgical Urology
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    ABSTRACT: Objective: To quantify the proportion of patients attending for urological investigation of non-visible haematuria who meet criteria for referral to specialist renal services under current UK guidelines. Patients and methods: Data were collected from 377 consecutive patients attending a dedicated non-visible haematuria clinic. Details were recorded for clinic outcomes and indicators of chronic kidney disease (eGFR, dipstick proteinuria, protein:creatinine ratio). The proportion of patients meeting referral criteria was calculated, including the increase in service need and costs. Results: 8.7% of patients had a positive urological diagnosis, with 2.7% having malignancies. 20.7% of patients had CKD stage 3 on the basis of their eGFR readings. 16.7% of patients had dipstick proteinuria, but only 4.4% had a raised protein:creatinine ratio. Overall, up to 22% of patients met current criteria for referral to nephrology, although this varied according to guideline. At this level, it equates to a potential increase of 20-30% in new patient referrals. Conclusion: Features of CKD are common in patients undergoing investigation for non-visible haematuria, with high numbers meeting current UK criteria for referral to nephrology. Appropriate investigation of these risk factors should form an integral part of the haematuria clinic.
    Preview · Article · Sep 2011 · British Journal of Medical and Surgical Urology
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    ABSTRACT: Figure S2: Quantification of expression of the stem cell marker CD133 in prostate cell lines. Live cells were stained with CD133/2(293C)-APC antibody (Miltenyi Biotec) (CD133-APC) or without any antibody (No Antibody) and analysed by FACs. For each dot plot, X axes: FITC channel fluorescence (not stained); Y axes: APC channel fluorescence (CD133 or No Antibody control); Percent of CD133+ cells is indicated.
    Preview · Dataset · Jul 2011
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    ABSTRACT: Table S1: List of the cell lines used. Table indicating the cell lines used in this manuscript, their origin and culture conditions.
    Preview · Dataset · Jul 2011
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    ABSTRACT: Additional analysis of CD133 promoter methylation in prostate primary epithelial cultures and prostate tissues. Figure S3: MSP analysis of the CD133 promoter with two different primer sets (MSP1 and MSP2) in prostate primary epithelial cultures (A) and in selected populations from primary epithelial cultures (B) (SC = stem cells; TA = transit amplifying cells; CB = committed basal cells; Negative Control = water; NO CELLS = negative control for whole bisulfitome amplification). Pyrosequencing methylation assay of CD133 promoter in prostate tissues (panel C: PYRO 2 assay; panel D: PYRO 3 assay; DNA extracted from glass slides of snap frozen tissue sections which were pooled together) and in prostate primary epithelial cultures (panel E: PYRO 2 assay; each bar represent a single CpG site; n = 3 technical replicas; ± SD).
    Preview · Dataset · Jul 2011
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    ABSTRACT: Table S2: List of the primers used Table indicating the primers used in this manuscript for PCR amplification and pyrosequencing.
    Preview · Dataset · Jul 2011
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    ABSTRACT: Additional analysis of CD133 promoter methylation in prostate cell lines. Figure S1: Methylation specific PCR (MSP) analysis of the CD133 promoter with two different primer sets (MSP1 and MSP2) in a panel of prostate cell lines (A). PCR products recognizing methylated (M) and unmethylated (U) CpG sites were analyzed on 2% agarose gels (Positive Control = RC165 DNA methylated in vitro with SssI methylase; Negative Control = water). Pyrosequencing methylation assay of CD133 CpG island in a panel of prostate cell lines (panel B: PYRO 2 assay; panel C: PYRO 3 assay; each bar represents a single CpG site; n = 3 technical replicas; ± SD; 100% Meth = Methylated human control DNA - QIAGEN; 0% Meth = Unmethylated human control DNA - QIAGEN; 50% Meth = 1:1 mixture of 100% Meth and 0% Meth). (D) Dot-plot showing CD133 promoter methylation (PYRO 1 assay, average of 10 CpG sites) in the same cell lines shown in Figure 1B grouped on the basis of their origin (benign or cancer) and their culture conditions (low FCS = 0%-2%; high FCS = 5%-10%; * = p < 0.05; ** = p < 0.01; *** = p < 0.001 in an unpaired t-test).
    Preview · Dataset · Jul 2011
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    ABSTRACT: Epigenetic control is essential for maintenance of tissue hierarchy and correct differentiation. In cancer, this hierarchical structure is altered and epigenetic control deregulated, but the relationship between these two phenomena is still unclear. CD133 is a marker for adult stem cells in various tissues and tumour types. Stem cell specificity is maintained by tight regulation of CD133 expression at both transcriptional and post-translational levels. In this study we investigated the role of epigenetic regulation of CD133 in epithelial differentiation and cancer. DNA methylation analysis of the CD133 promoter was done by pyrosequencing and methylation specific PCR; qRT-PCR was used to measure CD133 expression and chromatin structure was determined by ChIP. Cells were treated with DNA demethylating agents and HDAC inhibitors. All the experiments were carried out in both cell lines and primary samples. We found that CD133 expression is repressed by DNA methylation in the majority of prostate epithelial cell lines examined, where the promoter is heavily CpG hypermethylated, whereas in primary prostate cancer and benign prostatic hyperplasia, low levels of DNA methylation, accompanied by low levels of mRNA, were found. Moreover, differential methylation of CD133 was absent from both benign or malignant CD133+/α2β1integrinhi prostate (stem) cells, when compared to CD133-/α2β1integrinhi (transit amplifying) cells or CD133-/α2β1integrinlow (basal committed) cells, selected from primary epithelial cultures. Condensed chromatin was associated with CD133 downregulation in all of the cell lines, and treatment with HDAC inhibitors resulted in CD133 re-expression in both cell lines and primary samples. CD133 is tightly regulated by DNA methylation only in cell lines, where promoter methylation and gene expression inversely correlate. This highlights the crucial choice of cell model systems when studying epigenetic control in cancer biology and stem cell biology. Significantly, in both benign and malignant prostate primary tissues, regulation of CD133 is independent of DNA methylation, but is under the dynamic control of chromatin condensation. This indicates that CD133 expression is not altered in prostate cancer and it is consistent with an important role for CD133 in the maintenance of the hierarchical cell differentiation patterns in cancer.
    Full-text · Article · Jul 2011 · Molecular Cancer
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    ABSTRACT: DNA methylation analysis of CD133 promoter using pyrosequencing. Figure S4: (A) Diagram depicting a typical histogram generated by pyrosequencing methylation analysis of CD133 promoter with PYRO 1 assay and the position of each CpG site on the genomic sequence (Exon 1 A is shown in yellow). (B-C) Example of typical pyrograms generated with PYRO 1 assay using human (B) or mouse (C) bisulfite converted DNA as a template for PCR. No PCR product or sequence was generated with mouse DNA
    Preview · Dataset · Jul 2011
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    W Mahmalji · S Jain · M Stower
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    ABSTRACT: A 72-year-old Caucasian male who presented with haematuria in July of 2000 was found to have a large left-sided bladder tumour. He underwent a transurethral resection of the tumour and surveillance program. In October 2008 he underwent a transurethral resection of the prostate (TURP). Histology of the prostatic chippings showed poorly differentiated TCC with prostatic invasion. A CT of his chest abdomen and pelvis revealed no lymph node involvement or metastatic spread. He therefore underwent a cystoprostato-urethrectomy with ileal conduit formation, in December 2008. In May 2010 the decision was made to perform a left inguinal orchidectomy as he presented with a craggy mass of his left testis, and there were clinical concerns that this was a tumour. Histology revealed that the left testis had been wholly replaced by a tumour. Taking into account his previous urological history, the features of this tumour are consistent with metastatic TCC, which is very rare.
    Preview · Article · Apr 2011 · Advances in Urology
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    ABSTRACT: Expression of protein kinase C alpha (PKCα) is elevated in prostate cancer (PCa); thus, we have studied whether the development of tumourigenesis in prostate epithelial cell lines modifies the normal pattern of choline (Cho) metabolite release on PKC activation. Normal and tumourigenic human prostate epithelial cell lines were incubated with [(3)H]-Cho to label choline phospholipids. Protein kinase C was activated with phorbol ester and blocked with inhibitors. Choline metabolites were resolved by ion-exchange chromatography. Phospholipase D (PLD) activity was measured by transphosphatidylation. Protein expression was detected by western blotting and/or RT-PCR. Choline uptake was measured on cells in monolayers over 60 min. Normal prostate epithelial cell lines principally released phosphocholine (PCho) in contrast to tumourigenic lines, which released Cho. In addition, only with normal cell lines did PKC activation stimulate Cho metabolite release. Protein kinase C alpha expression varied between normal and tumourigenic cell lines but all showed a PKCα link to myristoylated alanine-rich C kinase substrate (MARCKS) protein. The five cell lines differed in Cho uptake levels, with normal PNT2C2 line cells showing highest uptake over 60 min incubation. Normal and tumourigenic cell lines expressed mRNA for PLD1 and PLD2, and showed similar levels of basal and PKC-activated PLD activity. The transition to tumourigenesis in prostate epithelial cell lines results in major changes to Cho metabolite release into the medium and PKC signalling to phosphatidylcholine turnover. The changes, which reflect the metabolic and proliferative needs of tumourigenic cells compared with untransformed cells, could be significant for both diagnosis and treatment.
    Preview · Article · Feb 2011 · British Journal of Cancer

Publication Stats

3k Citations
163.33 Total Impact Points

Institutions

  • 1996-2014
    • York Hospital
      New York, New York, United States
  • 2012
    • York Teaching Hospital NHS Foundation Trust
      York, England, United Kingdom
  • 1995-2006
    • The University of York
      • • Yorkshire Cancer Research Unit
      • • Department of Biology
      York, ENG, United Kingdom