Methylation profiling and evaluation of demethylating therapy in renal cell carcinoma

Christopher J. Ricketts, Mark R. Morris, Dean Gentle, Salwati Shuib, Michael Brown, Noel Clarke, Wenbin Wei, Paul Nathan, Farida Latif, Eamonn R. Maher

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

Background: Despite therapeutic advances in targeted therapy, metastatic renal cell carcinoma (RCC) remains incurable for the vast majority of patients. Key molecular events in the pathogenesis of RCC include inactivation of the VHL tumour suppressor gene (TSG), inactivation of chromosome 3p TSGs implicated in chromatin modification and remodelling and de novo tumour-specific promoter methylation of renal TSGs. In the light of these observations it can be proposed that, as in some haematological malignancies, demethylating agents such as azacitidine might be beneficial for the treatment of advanced RCC. Results: Here we report that the treatment of RCC cell lines with azacitidine suppressed cell proliferation in all 15 lines tested. A marked response to azacitidine therapy (>50% reduction in colony formation assay) was detected in the three cell lines with VHL promoter methylation but some RCC cell lines without VHL TSG methylation also demonstrated a similar response suggesting that multiple methylated TSGs might determine the response to demethylating therapies. To identify novel candidate methylated TSGs implicated in RCC we undertook a combined analysis of copy number and CpG methylation array data. Candidate novel epigenetically inactivated TSGs were further prioritised by expression analysis of RCC cell lines pre and post-azacitidine therapy and comparative expression analysis of tumour/normal pairs. Thus, with subsequent investigation two candidate genes were found to be methylated in more than 25% of our series and in the TCGA methylation dataset for 199 RCC samples: RGS7 (25.6% and 35.2% of tumours respectively) and NEFM in (25.6% and 30.2%). In addition three candidate genes were methylated in >10% of both datasets ( TMEM74 (15.4% and 14.6%), GCM2 (41.0% and 14.6%) and AEBP1 (30.8% and 13.1%)). Methylation of GCM2 (P = 0.0324), NEFM (P = 0.0024) and RGS7 (P = 0.0067) was associated with prognosis. Conclusions: These findings provide preclinical evidence that treatment with demethylating agents such as azacitidine might be useful for the treatment of advanced RCC and further insights into the role of epigenetic changes in the pathogenesis of RCC.

Original languageEnglish
Article number16
JournalClinical Epigenetics
Volume5
Issue number1
DOIs
Publication statusPublished - 2013

Fingerprint

Renal Cell Carcinoma
Methylation
Azacitidine
Therapeutics
Cell Line
Tumor Suppressor Genes
Chromatin Assembly and Disassembly
Gene Silencing
Hematologic Neoplasms
Epigenomics
Carcinogens
Genes
Neoplasms
Chromosomes
Cell Proliferation
Kidney

Keywords

  • Demethylation
  • Epigenetics
  • Methylation
  • Renal cancer
  • Renal cell carcinoma
  • Therapy
  • VHL

ASJC Scopus subject areas

  • Genetics
  • Molecular Biology
  • Developmental Biology
  • Genetics(clinical)

Cite this

Ricketts, C. J., Morris, M. R., Gentle, D., Shuib, S., Brown, M., Clarke, N., ... Maher, E. R. (2013). Methylation profiling and evaluation of demethylating therapy in renal cell carcinoma. Clinical Epigenetics, 5(1), [16]. https://doi.org/10.1186/1868-7083-5-16

Methylation profiling and evaluation of demethylating therapy in renal cell carcinoma. / Ricketts, Christopher J.; Morris, Mark R.; Gentle, Dean; Shuib, Salwati; Brown, Michael; Clarke, Noel; Wei, Wenbin; Nathan, Paul; Latif, Farida; Maher, Eamonn R.

In: Clinical Epigenetics, Vol. 5, No. 1, 16, 2013.

Research output: Contribution to journalArticle

Ricketts, CJ, Morris, MR, Gentle, D, Shuib, S, Brown, M, Clarke, N, Wei, W, Nathan, P, Latif, F & Maher, ER 2013, 'Methylation profiling and evaluation of demethylating therapy in renal cell carcinoma', Clinical Epigenetics, vol. 5, no. 1, 16. https://doi.org/10.1186/1868-7083-5-16
Ricketts, Christopher J. ; Morris, Mark R. ; Gentle, Dean ; Shuib, Salwati ; Brown, Michael ; Clarke, Noel ; Wei, Wenbin ; Nathan, Paul ; Latif, Farida ; Maher, Eamonn R. / Methylation profiling and evaluation of demethylating therapy in renal cell carcinoma. In: Clinical Epigenetics. 2013 ; Vol. 5, No. 1.
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abstract = "Background: Despite therapeutic advances in targeted therapy, metastatic renal cell carcinoma (RCC) remains incurable for the vast majority of patients. Key molecular events in the pathogenesis of RCC include inactivation of the VHL tumour suppressor gene (TSG), inactivation of chromosome 3p TSGs implicated in chromatin modification and remodelling and de novo tumour-specific promoter methylation of renal TSGs. In the light of these observations it can be proposed that, as in some haematological malignancies, demethylating agents such as azacitidine might be beneficial for the treatment of advanced RCC. Results: Here we report that the treatment of RCC cell lines with azacitidine suppressed cell proliferation in all 15 lines tested. A marked response to azacitidine therapy (>50{\%} reduction in colony formation assay) was detected in the three cell lines with VHL promoter methylation but some RCC cell lines without VHL TSG methylation also demonstrated a similar response suggesting that multiple methylated TSGs might determine the response to demethylating therapies. To identify novel candidate methylated TSGs implicated in RCC we undertook a combined analysis of copy number and CpG methylation array data. Candidate novel epigenetically inactivated TSGs were further prioritised by expression analysis of RCC cell lines pre and post-azacitidine therapy and comparative expression analysis of tumour/normal pairs. Thus, with subsequent investigation two candidate genes were found to be methylated in more than 25{\%} of our series and in the TCGA methylation dataset for 199 RCC samples: RGS7 (25.6{\%} and 35.2{\%} of tumours respectively) and NEFM in (25.6{\%} and 30.2{\%}). In addition three candidate genes were methylated in >10{\%} of both datasets ( TMEM74 (15.4{\%} and 14.6{\%}), GCM2 (41.0{\%} and 14.6{\%}) and AEBP1 (30.8{\%} and 13.1{\%})). Methylation of GCM2 (P = 0.0324), NEFM (P = 0.0024) and RGS7 (P = 0.0067) was associated with prognosis. Conclusions: These findings provide preclinical evidence that treatment with demethylating agents such as azacitidine might be useful for the treatment of advanced RCC and further insights into the role of epigenetic changes in the pathogenesis of RCC.",
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AU - Morris, Mark R.

AU - Gentle, Dean

AU - Shuib, Salwati

AU - Brown, Michael

AU - Clarke, Noel

AU - Wei, Wenbin

AU - Nathan, Paul

AU - Latif, Farida

AU - Maher, Eamonn R.

PY - 2013

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N2 - Background: Despite therapeutic advances in targeted therapy, metastatic renal cell carcinoma (RCC) remains incurable for the vast majority of patients. Key molecular events in the pathogenesis of RCC include inactivation of the VHL tumour suppressor gene (TSG), inactivation of chromosome 3p TSGs implicated in chromatin modification and remodelling and de novo tumour-specific promoter methylation of renal TSGs. In the light of these observations it can be proposed that, as in some haematological malignancies, demethylating agents such as azacitidine might be beneficial for the treatment of advanced RCC. Results: Here we report that the treatment of RCC cell lines with azacitidine suppressed cell proliferation in all 15 lines tested. A marked response to azacitidine therapy (>50% reduction in colony formation assay) was detected in the three cell lines with VHL promoter methylation but some RCC cell lines without VHL TSG methylation also demonstrated a similar response suggesting that multiple methylated TSGs might determine the response to demethylating therapies. To identify novel candidate methylated TSGs implicated in RCC we undertook a combined analysis of copy number and CpG methylation array data. Candidate novel epigenetically inactivated TSGs were further prioritised by expression analysis of RCC cell lines pre and post-azacitidine therapy and comparative expression analysis of tumour/normal pairs. Thus, with subsequent investigation two candidate genes were found to be methylated in more than 25% of our series and in the TCGA methylation dataset for 199 RCC samples: RGS7 (25.6% and 35.2% of tumours respectively) and NEFM in (25.6% and 30.2%). In addition three candidate genes were methylated in >10% of both datasets ( TMEM74 (15.4% and 14.6%), GCM2 (41.0% and 14.6%) and AEBP1 (30.8% and 13.1%)). Methylation of GCM2 (P = 0.0324), NEFM (P = 0.0024) and RGS7 (P = 0.0067) was associated with prognosis. Conclusions: These findings provide preclinical evidence that treatment with demethylating agents such as azacitidine might be useful for the treatment of advanced RCC and further insights into the role of epigenetic changes in the pathogenesis of RCC.

AB - Background: Despite therapeutic advances in targeted therapy, metastatic renal cell carcinoma (RCC) remains incurable for the vast majority of patients. Key molecular events in the pathogenesis of RCC include inactivation of the VHL tumour suppressor gene (TSG), inactivation of chromosome 3p TSGs implicated in chromatin modification and remodelling and de novo tumour-specific promoter methylation of renal TSGs. In the light of these observations it can be proposed that, as in some haematological malignancies, demethylating agents such as azacitidine might be beneficial for the treatment of advanced RCC. Results: Here we report that the treatment of RCC cell lines with azacitidine suppressed cell proliferation in all 15 lines tested. A marked response to azacitidine therapy (>50% reduction in colony formation assay) was detected in the three cell lines with VHL promoter methylation but some RCC cell lines without VHL TSG methylation also demonstrated a similar response suggesting that multiple methylated TSGs might determine the response to demethylating therapies. To identify novel candidate methylated TSGs implicated in RCC we undertook a combined analysis of copy number and CpG methylation array data. Candidate novel epigenetically inactivated TSGs were further prioritised by expression analysis of RCC cell lines pre and post-azacitidine therapy and comparative expression analysis of tumour/normal pairs. Thus, with subsequent investigation two candidate genes were found to be methylated in more than 25% of our series and in the TCGA methylation dataset for 199 RCC samples: RGS7 (25.6% and 35.2% of tumours respectively) and NEFM in (25.6% and 30.2%). In addition three candidate genes were methylated in >10% of both datasets ( TMEM74 (15.4% and 14.6%), GCM2 (41.0% and 14.6%) and AEBP1 (30.8% and 13.1%)). Methylation of GCM2 (P = 0.0324), NEFM (P = 0.0024) and RGS7 (P = 0.0067) was associated with prognosis. Conclusions: These findings provide preclinical evidence that treatment with demethylating agents such as azacitidine might be useful for the treatment of advanced RCC and further insights into the role of epigenetic changes in the pathogenesis of RCC.

KW - Demethylation

KW - Epigenetics

KW - Methylation

KW - Renal cancer

KW - Renal cell carcinoma

KW - Therapy

KW - VHL

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