DNA Methylation Markers for Prostate Cancer with a Stem Cell Twist

Published OnlineFirst August 10, 2010; DOI: 10.1158/1940-6207.CAPR-10-0131 Published OnlineFirst on August 10, 2010 as 10.1158/1940-6207.CAPR-10-0131

Perspective Cancer Prevention Perspective on Ibragimova et al., p. XXX Research DNA Methylation Markers for Prostate Cancer with a Stem Cell Twist

Carmen Jerónimo1,2 and Manel Esteller3,4

Abstract This perspective on Ibragimova et al. (beginning on page XXX in this issue of the journal) highlights the potential role of DNA methylation–based markers in the early detection of prostate cancer (PCa), with a focus on the global reactivation of expression of genes epigenetically silenced in PCa cell lines. Novel findings of these investigators identified four genes methylated specifically in PCa, including the stem cell marker TACSTD2, which seems to discriminate PCa (methylated TACSTD2) from prostatic intraepithelial neoplasia (unmethylated). These genes add significantly to the list of epigenetic markers showing promise for clinical early detection of PCa in the near future. Cancer Prev Res; 3(9); OF1–3. ©2010 AACR.

Prostate cancer (PCa) is one of the most prevalent number of men to unnecessary therapy-related morbid- malignant neoplasms in men living in Western countries. ity (3). Therefore, research devoted to finding new PCa In the United States, it accounted for 25% of all non- biomarkers that could replace or be an adjunct to serum cutaneous cancer cases in men and 9% of cancer-related PSA as a screening tool has flourished in recent years. mortality, with an estimated 192,280 new cases and The last decade has witnessed the progressive acknowl- 27,360 deaths in 2009 (1). Locally advanced or metastatic edgment of the role of epigenetics in the development disease is associated with considerable morbidity and and progression of human cancers in general and PCa mortality. Therefore, efficient early detection could greatly in particular. Indeed, there is growing evidence that epi- alleviate the burden of PCa. genetic alterations are early events in tumorigenesis and Current efforts to identify individuals with early-stage may even precede better-known genetic alterations (4). PCa, who are most amenable to curative treatments, re- Instances of aberrant DNA methylation at CpG-rich sites lies on screening methods based mainly on putative in the regulatory regions of several genes are currently the PCa-specific biomarkers. Measuring prostate-specific anti- strongest candidates among cancer-related epigenetic al- gen (PSA) in the serum of at-risk men has become the terations for becoming epigenetic-based cancer biomar- mainstay of PCa screening programs over the past kers because this methylation is relatively easy to screen 20 years. Although the introduction of PSA in routine for, even in samples with low tumor DNA content such clinical practice has increased the rate of detecting as body fluids (blood, urine; refs. 4, 5). Moreover, the organ-confined PCa and is associated with a decrease emergence of high-throughput techniques allows for the in mortality (2, 3), the role of serum PSA has been chal- simultaneous analysis of multiple gene promoters in a lenged mainly because of its relatively low specificity, large number of clinical samples. especially at the lower end of its spectrum of levels, Initial efforts to find specific DNA methylation markers where most curable PCa occurs. It is important to note for PCa followed a “candidate gene” approach. Genes with that there is growing concern in the medical community a lost or reduced expression in PCa tissue or cell lines be- that PSA-detected PCa is overtreated, subjecting a large came candidates for assessing epigenetic silencing through aberrant promoter methylation. This approach discovered some of the most robust methylation-based biomarkers for Authors' Affiliations: 1Cancer Epigenetics Group, Research Center of the GSTP1 Portuguese Oncology Institute-Porto; 2Department of Pathology and PCa detection, of which is one of the best-known Molecular Immunology, Institute of Biomedical Sciences Abel Salazar examples (6, 7). GSTP1 is hypermethylated in 80% to (ICBAS), University of Porto, Porto, Portugal; 3Cancer Epigenetics and 90% of primary prostate tumors and at much lower fre- Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL); and 4Institucio Catalana de Recerca i Estudis Avançats quencies in liver, kidney, and breast neoplasms (20-30%; (ICREA), Barcelona, Catalonia, Spain ref. 8). Extended studies of PCa methylation profiling iden- Corresponding Author: Manel Esteller, Cancer Epigenetics and Biology tified other genes that, when added to GSTP1 in panels, Program (PEBC), 3rd Floor, Hospital Duran I Reynalds, Avinguda Gran Via augmented the sensitivity of testing for GSTP1 methylation s/n Km 2.7, E-08907 L'Hospitalet de Llobregat, Barcelona, Catalonia, Spain. Phone: 34-93-2607253; Fax: 34-93-2607219; E-mail: mesteller@ alone, even in urine samples (9). The candidate gene strat- iconcologia.net. egy is very laborious, however, requiring the full screening doi: 10.1158/1940-6207.CAPR-10-0131 of a large number of genes, of which only a minority will ©2010 American Association for Cancer Research. show promise as specific cancer biomarkers.

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Jerónimo and Esteller

Genome-wide analysis of the cancer methylome cell lines may be misleading and require an extensive recently has become possible and provides interesting analysis, as performed by Ibragimova et al., of tumor results that are tested in tissues and other clinical tissues to ascertain the validity of the biomarkers in a samples to identify potential cancer biomarkers. One “real-life” setting. method that provides valuable information about epige- Another interesting finding of this study is the differen- netically regulated genes is based on the dynamics of tial methylation of TACSTD2 between PCa and PIN epigenetic alteration, which can be pharmacologically lesions. Indeed, most of the genes previously known to modulated and reversed. As reported in this issue of be methylated in PCa were also methylated in PIN, the journal, Ibragimova and colleagues (10) exposed although less frequently (11, 12). If confirmed in a larger PCa cell lines to both 5-aza-2-deoxycytidine (a DNA- series, the differential TACSTD2 finding will be an impor- demethylating agent) and trichostatin A (an inhibitor tant step toward discriminating cells originating in PCa of histone deacetylases), thus allowing for the reexpres- from cells originating in PIN lesions (e.g., in urine), thus sion of epigenetically silenced genes detected in an ex- increasing the specificity of methylation-based assays for pression-array platform. They initially identified a set early detection of PCa. Also, with expression in a subset of almost 3,000 upregulated genes, from which they of prostate basal cells with stem cell features (13) and selected a panel of 45 genes following comparative silencing by promoter methylation in PCa but not in expression analysis of normal and cancerous prostate PIN, TACSTD2 might represent an “epigenetic switch” GSTP1 cells. After removing 15 genes (including ) already that renders preinvasive neoplastic cells prone to becom- reported in the literature to be methylated in PCa, they ing invasive, thus advancing our biological understanding conducted novel work that identified and validated of prostate carcinogenesis. the previously unknown methylation of seven other A limitation of the Ibragimova et al. study is its lack KRT7,TACSTD2,SLC15A3,GADD45d,IFI30, genes ( of testing in body fluids (plasma and/or urine), where ANXA2 AQP3 ,and ) in PCa by direct bisulfite sequen- detection of CpG island hypermethylation could have cing. These genes were extensively methylated in PCa a clear clinical value (14) and would add to the novelty cell lines, representing less than 0.3% of all the genes of the findings. It also would be important to extend that surfaced from the array analysis. This result clearly the number of specimens in the tissue validation set shows the power of global methylation analysis to to ascertain whether all PIN lesions indeed are un- uncover and pinpoint potential biomarkers among a methylated at the TACSTD2 promoter. Last, it would very large number of putative candidates. Of impor- be interesting to determine whether these novel epige- tance, the validity of the methods and data analysis of netic markers are complementary, and whether their GSTP1 Ibragimova et al. is supported by the presence of combined use might increase the rate (sensitivity) of in their data set as one of the most frequently methy- PCa detection. lated genes, as would be predicted from previously In conclusion, the data provided by this novel study add published data. significantly to our knowledge of the PCa methylome, In an obvious next step, these investigators validated revealing some promising PCa biomarkers that deserve the seven newly identified genes in tissue samples, further exploratory studies in body fluids to assess their which came from 35 PCa and 19 lesions of prostatic in- power to detect early PCa in the clinic. traepithelial neoplasia (PIN), a putative PCa precursor. Including PIN in this setting provided some interesting insight into the timing of the gain of methylation of the studied genes during prostate carcinogenesis. The vali- Disclosure of Potential Conflicts of Interest dation analyses found that three (KRT7, TACSTD2, and SLC15A3) of the seven genes identified in vitro were No potential conflicts of interest were disclosed. frequently methylated (17-66%) in PCa tissues. These results highlight the differences between cancer cell lines Received 06/14/2010; accepted 07/15/2010; published OnlineFirst and primary cancer tissues. Indeed, findings in cancer 08/10/2010.

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DNA Methylation Markers for PCa with a Stem Cell Twist

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www.aacrjournals.org Cancer Prev Res; 3(9) September 2010 OF3

DNA Methylation Markers for Prostate Cancer with a Stem Cell Twist

Carmen Jerónimo and Manel Esteller

Cancer Prev Res Published OnlineFirst August 10, 2010.

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