Epigenetic Factors Affect Tumor Initiation, Progression and Recurrence WE are witnessing a major paradigm shift in the way that we diagnose and treat cancer. Therapies designed in the 20th century were predominantly agents that caused indiscriminate genomic damage or cellular stress, whereas an emerging understanding of tumor genetics and biology has redirected our efforts toward targeting the pathophysiology of individual tumors. This has been facilitated by exponential advances in sequencing technology. The first fully sequenced eukaryote genome, Saccharomyces cerevisiae (12.1 Mb), was celebrated in 1996,1 followed in 2003 by the first full human genome, a massive international collaborative effort spanning more than a decade. In the few short years since, we have sequenced thousands of human genomes, highlighting the heterogeneity of our species.2 As a logical extension of these efforts, The Cancer Genome Atlas Project was launched in 2005 to catalogue genetic mutations responsible for cancer. The promise of genomics was the advent of personalized or precision medicine in which molecular analysis could be used to tailor specific treatment strategies. However, genomics alone do not provide the entire picture of what occurs at the cellular level. DNA is subject to layers of complex control that ultimately orchestrate which genes are expressed and when, determining the final protein makeup of a cell. These regulatory mechanisms represent the emerging field of epigenetics and are at least as critical to tumor initiation and progression as specific genetic alterations. Epigenetic changes involve chromatin modifications that turn genes on or off without altering the DNA base pair sequence. These processes are essential to normal development and physiology, facilitating X chromosome inactivation or differentiation of the zygote into multiple cell lineages with diverse functions. The genome provides the raw data for life but the epigenetic pattern of gene expression determines the ultimate behavior of a cell. Mechanisms of epigenetic control occur at multiple levels. Some examples include histone modifications that affect chromatin remodeling, dictating
0022-5347/14/1925-1318/0 THE JOURNAL OF UROLOGY® © 2014 by AMERICAN UROLOGICAL ASSOCIATION EDUCATION
1318
j
www.jurology.com
AND
RESEARCH, INC.
where the double helix will unwind sufficiently to expose a gene for transcription. Direct chemical modifications to DNA affect polymerase binding or recruitment of transcriptional cofactors. Once transcribed, mRNAs are subject to RNA induced transcriptional silencing by specific miRNAs or siRNAs. Many epigenetic changes are environmentally induced and accumulate with time, leading to the concept of epigenetic drift seen with aging in monozygotic twins.3 A hallmark of epigenetic changes is their heritability. Once established, gene expression patterns are faithfully passed on to daughter cells to maintain the lineage. The first discovered and most studied epigenetic mechanism is promoter methylation. This involves the addition of methyl groups to cytosine residues of CpG dinucleotides. Long chains of CpG repeats are often located at or near the transcriptional start site. Accumulation of bulky methyl side chains physically blocks the binding of transcription factors, prevents transcription and functionally silences the hypermethylated gene. Cancers exploit epigenetic changes to customize cell expression patterns to suit their needs. They not only use hypermethylation to silence proteins involved in cellular defense or tumor suppression but can also demethylate physiologically silenced genes to inappropriately “turn on” oncogenes. As these changes accumulate in the cell, they enhance the malignant potential of the tumor by promoting survival, invasion or metastasis. Alterations in methylation status have been linked to environmental factors, including oxidative stress, pesticides, dietary excess or deficiencies, tobacco smoke or other pollutants,4 providing an epigenetic link between known carcinogens and the development of malignancy. Rampant hypomethylation will ultimately destabilize the genome, contributing to the accumulation of genetic abnormalities common to advanced cancers. Comprehensive characterization of the tumor “epigenome” will be critical to realizing the dream of personalized medicine and tailored strategies for the prevention, detection, stratification and treatment of individual tumors. Novel anticancer
http://dx.doi.org/10.1016/j.juro.2014.08.051 Vol. 192, 1318-1319, November 2014 Printed in U.S.A.
EPIGENETIC FACTORS AFFECT TUMOR
therapies, including methyltransferase and histone deacetylase inhibitors, are aimed at reversing epigenetic alterations. This issue of The Journal includes 2 examples of how cancer epigenetics play a role in genitourinary malignancies. In the first article Lian et al (page 1528) revisit hypermethylation inactivation of the VHL tumor suppressor gene,5 reported in 15% to 20% of sporadic clear cell renal cell carcinomas.6 Historically, these assays were slow and costly. The authors report the feasibility of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry instead of sequencing, having noted robust and reproducible detection of methylation at the base pair level. This technology will facilitate tumor interrogation as we expand personalized medicine to include epigenetics. Interestingly, they report that VHL promoter silencing can alternatively occur via hypermethylation of sites in exon 1, suggesting that future assays will have to expand the search area. They propose that VHL methylation status in serum or urine might serve as a biomarker for cancer detection. In the second article Maldonado et al (page 1542) present GSTP1 methylation as a “tissue based recurrence marker.”7 GSTP1 normally functions to detoxify environmental stressors, protecting cells from resulting DNA damage. Functional silencing is found in 90% of prostate cancers, leading to increased vulnerability to genetic aberrations. The presence of GSTP1 promoter methylation can
1319
distinguish prostate cancer from benign prostatic tissue and the extent of hypermethylation correlates with advancing tumor stage.8 To determine whether the extent of GSTP1 promoter methylation could predict men at increased risk for recurrence following radical prostatectomy Maldonado et al used quantitative methylation-specific polymerase chain reaction to compare primary tumor samples from men with recurrence and control samples from age and stage matched men without recurrence. On multivariate analysis an increased extent of GSTP1 methylation in early stage prostate cancer increased the risk of recurrence about 1.7fold. Although they conclude that this incremental improvement alone is unlikely to change clinical care, they note the potential for epigenetic markers to risk stratify otherwise indistinguishable cancers. They propose that this is a step toward the creation of a robust panel of genetic and epigenetic features that will ultimately guide clinical decision making. Cancer is a heterogeneous disease that has repeatedly shown us that it is not amenable to broad categorizations or generalizations. Although in its infancy, the field of cancer epigenetics will be critical to realize the promise of personalized medicine to identify, stratify, target and ultimately cure cancer. Jodi K. Maranchie Department of Urology University of Pittsburgh Pittsburgh, Pennsylvania
REFERENCES 1. Goffeau A, Barrell BG, Bussey H et al: Life with 6000 genes. Science 1996; 274: 546. 2. Abecasis GR, Auton A, Brooks LD et al: An integrated map of genetic variation from 1,092 human genomes. Nature 2012; 491: 56. 3. Fraga MF, Ballestar E, Paz MF et al: Epigenetic differences arise during the lifetime of monozygotic twins. Proc Natl Acad Sci U S A 2005; 102: 10604.
4. Hou L, Zhang X, Wang D et al: Environmental chemical exposures and human epigenetics. Int J Epidemiol 2012; 41: 79. 5. Lian F, Sreedharan S, Arnold RS et al: von Hippel-Lindau exonic methylation analysis using MALDI-TOF mass spectrometry. J Urol 2014; 192: 1528. 6. Herman JG, Latif F, Weng Y et al: Silencing of the VHL tumor-suppressor gene by DNA methylation
in renal carcinoma. Proc Natl Acad Sci U S A 1994; 91: 9700. 7. Maldonado L, Brait M, Loyo M et al: GSTP1 promoter methylation is associated with recurrence in early stage prostate cancer. J Urol 2014; 192: 1542. 8. Enokida H, Shiina H, Urakami S et al: Multigene methylation analysis for detection and staging of prostate cancer. Clin Cancer Res 2005; 11: 6582.
0022-5347/14/1925-1318/0 THE JOURNAL OF UROLOGY® © 2014 by AMERICAN UROLOGICAL ASSOCIATION EDUCATION
1318
j
www.jurology.com
AND
RESEARCH, INC.
where the double helix will unwind sufficiently to expose a gene for transcription. Direct chemical modifications to DNA affect polymerase binding or recruitment of transcriptional cofactors. Once transcribed, mRNAs are subject to RNA induced transcriptional silencing by specific miRNAs or siRNAs. Many epigenetic changes are environmentally induced and accumulate with time, leading to the concept of epigenetic drift seen with aging in monozygotic twins.3 A hallmark of epigenetic changes is their heritability. Once established, gene expression patterns are faithfully passed on to daughter cells to maintain the lineage. The first discovered and most studied epigenetic mechanism is promoter methylation. This involves the addition of methyl groups to cytosine residues of CpG dinucleotides. Long chains of CpG repeats are often located at or near the transcriptional start site. Accumulation of bulky methyl side chains physically blocks the binding of transcription factors, prevents transcription and functionally silences the hypermethylated gene. Cancers exploit epigenetic changes to customize cell expression patterns to suit their needs. They not only use hypermethylation to silence proteins involved in cellular defense or tumor suppression but can also demethylate physiologically silenced genes to inappropriately “turn on” oncogenes. As these changes accumulate in the cell, they enhance the malignant potential of the tumor by promoting survival, invasion or metastasis. Alterations in methylation status have been linked to environmental factors, including oxidative stress, pesticides, dietary excess or deficiencies, tobacco smoke or other pollutants,4 providing an epigenetic link between known carcinogens and the development of malignancy. Rampant hypomethylation will ultimately destabilize the genome, contributing to the accumulation of genetic abnormalities common to advanced cancers. Comprehensive characterization of the tumor “epigenome” will be critical to realizing the dream of personalized medicine and tailored strategies for the prevention, detection, stratification and treatment of individual tumors. Novel anticancer
http://dx.doi.org/10.1016/j.juro.2014.08.051 Vol. 192, 1318-1319, November 2014 Printed in U.S.A.
EPIGENETIC FACTORS AFFECT TUMOR
therapies, including methyltransferase and histone deacetylase inhibitors, are aimed at reversing epigenetic alterations. This issue of The Journal includes 2 examples of how cancer epigenetics play a role in genitourinary malignancies. In the first article Lian et al (page 1528) revisit hypermethylation inactivation of the VHL tumor suppressor gene,5 reported in 15% to 20% of sporadic clear cell renal cell carcinomas.6 Historically, these assays were slow and costly. The authors report the feasibility of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry instead of sequencing, having noted robust and reproducible detection of methylation at the base pair level. This technology will facilitate tumor interrogation as we expand personalized medicine to include epigenetics. Interestingly, they report that VHL promoter silencing can alternatively occur via hypermethylation of sites in exon 1, suggesting that future assays will have to expand the search area. They propose that VHL methylation status in serum or urine might serve as a biomarker for cancer detection. In the second article Maldonado et al (page 1542) present GSTP1 methylation as a “tissue based recurrence marker.”7 GSTP1 normally functions to detoxify environmental stressors, protecting cells from resulting DNA damage. Functional silencing is found in 90% of prostate cancers, leading to increased vulnerability to genetic aberrations. The presence of GSTP1 promoter methylation can
1319
distinguish prostate cancer from benign prostatic tissue and the extent of hypermethylation correlates with advancing tumor stage.8 To determine whether the extent of GSTP1 promoter methylation could predict men at increased risk for recurrence following radical prostatectomy Maldonado et al used quantitative methylation-specific polymerase chain reaction to compare primary tumor samples from men with recurrence and control samples from age and stage matched men without recurrence. On multivariate analysis an increased extent of GSTP1 methylation in early stage prostate cancer increased the risk of recurrence about 1.7fold. Although they conclude that this incremental improvement alone is unlikely to change clinical care, they note the potential for epigenetic markers to risk stratify otherwise indistinguishable cancers. They propose that this is a step toward the creation of a robust panel of genetic and epigenetic features that will ultimately guide clinical decision making. Cancer is a heterogeneous disease that has repeatedly shown us that it is not amenable to broad categorizations or generalizations. Although in its infancy, the field of cancer epigenetics will be critical to realize the promise of personalized medicine to identify, stratify, target and ultimately cure cancer. Jodi K. Maranchie Department of Urology University of Pittsburgh Pittsburgh, Pennsylvania
REFERENCES 1. Goffeau A, Barrell BG, Bussey H et al: Life with 6000 genes. Science 1996; 274: 546. 2. Abecasis GR, Auton A, Brooks LD et al: An integrated map of genetic variation from 1,092 human genomes. Nature 2012; 491: 56. 3. Fraga MF, Ballestar E, Paz MF et al: Epigenetic differences arise during the lifetime of monozygotic twins. Proc Natl Acad Sci U S A 2005; 102: 10604.
4. Hou L, Zhang X, Wang D et al: Environmental chemical exposures and human epigenetics. Int J Epidemiol 2012; 41: 79. 5. Lian F, Sreedharan S, Arnold RS et al: von Hippel-Lindau exonic methylation analysis using MALDI-TOF mass spectrometry. J Urol 2014; 192: 1528. 6. Herman JG, Latif F, Weng Y et al: Silencing of the VHL tumor-suppressor gene by DNA methylation
in renal carcinoma. Proc Natl Acad Sci U S A 1994; 91: 9700. 7. Maldonado L, Brait M, Loyo M et al: GSTP1 promoter methylation is associated with recurrence in early stage prostate cancer. J Urol 2014; 192: 1542. 8. Enokida H, Shiina H, Urakami S et al: Multigene methylation analysis for detection and staging of prostate cancer. Clin Cancer Res 2005; 11: 6582.
