Identification of hypermethylated genes associated with cisplatin resistance in human cancers.

Cisplatin is among the most widely used cytotoxic anticancer agents in solid tumors; however, the development of secondary resistance remains a major obstacle to clinical efficacy. Treatment-related DNA hypermethylation may play a role in creating drug-resistant phenotypes by inactivating genes that are required for cytotoxicity. We applied a pharmacologic unmasking approach to detect hypermethylated genes whose inactivation contributes to cisplatin resistance. Using three pairs of isogeneic, cisplatin-sensitive, and cisplatin-resistant cell lines derived from two parental cell lines (KB-3-1 and SCC25), we identified several hundred genes that were downregulated in each resistant cell line and reactivated by the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine. Among them, 30 genes were common to two or more cell lines and/or reported to be downregulated in previous studies. Bisulfite sequencing confirmed that 14 genes were hypermethylated in resistant cell lines but not in the sensitive parental cell lines. Six of 14 genes (SAT, C8orf4, LAMB3, TUBB, G0S2, and MCAM) were cisplatin inducible in sensitive but not in resistant cell lines. Small interfering RNA knockdown of two genes, SAT and S100P, increased cell viability with cisplatin treatment in sensitive parental cell lines. S100P knockdown significantly decreased the S-phase fraction of parental sensitive cell lines and slowed cell proliferation, which was associated with decreased sensitivity to cisplatin. Based on these findings, we conclude that DNA methylation is a frequent event in cells that are chronically exposed to cisplatin and that methylation-induced gene silencing may play a role in the development of resistance to cytotoxic chemotherapeutic agents.

[1]  David E. Misek,et al.  Gene-expression profiles predict survival of patients with lung adenocarcinoma , 2002, Nature Medicine.

[2]  G. Giaccone.  Clinical Perspectives on Platinum Resistance , 2012, Drugs.

[3]  P. Johnston,et al.  The role of spermidine/spermine N1-acetyltransferase in determining response to chemotherapeutic agents in colorectal cancer cells , 2007, Molecular Cancer Therapeutics.

[4]  Min Liu,et al.  Establishment and gene analysis of an oxaliplatin-resistant colon cancer cell line THC8307/L-OHP , 2007, Anti-cancer drugs.

[5]  M. Gottesman,et al.  Mislocalization of membrane proteins associated with multidrug resistance in cisplatin-resistant cancer cell lines. , 2003, Cancer research.

[6]  Chun Xing Li,et al.  Epigenetic inactivation of a RAS association domain family protein from the lung tumour suppressor locus 3p21.3 , 2000, Nature Genetics.

[7]  J. Herman,et al.  Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[8]  S. Dahrouge,et al.  Chemotherapy dose--response relationships in non-small cell lung cancer and implied resistance mechanisms. , 2007, Cancer treatment reviews.

[9]  K. Franssila,et al.  Do DNA ploidy and S-phase fraction in primary tumour predict the response to chemotherapy in metastatic breast cancer? , 1995, British Journal of Cancer.

[10]  Jason L. Townson,et al.  Ineffectiveness of Doxorubicin Treatment on Solitary Dormant Mammary Carcinoma Cells or Late-developing Metastases , 2003, Breast Cancer Research and Treatment.

[11]  W. Schmitt,et al.  S100A2-S100P expression profile and diagnosis of non-small cell lung carcinoma: impairment by advanced tumour stages and neoadjuvant chemotherapy. , 2007, European journal of cancer.

[12]  J. Herman,et al.  Aberrant patterns of DNA methylation, chromatin formation and gene expression in cancer. , 2001, Human molecular genetics.

[13]  Michael Goggins,et al.  Identification of maspin and S100P as novel hypomethylation targets in pancreatic cancer using global gene expression profiling , 2004, Oncogene.

[14]  G. Kruh.  Introduction to resistance to anticancer agents , 2003, Oncogene.

[15]  Scar,et al.  Inactivation of the DNA-repair gene MGMT and the clinical response of gliomas to alkylating agents. , 2000, The New England journal of medicine.

[16]  A. Dueñas-González,et al.  Journal of Translational Medicine Global Dna Hypermethylation-associated Cancer Chemotherapy Resistance and Its Reversion with the Demethylating Agent Hydralazine , 2022 .

[17]  O. Kovalchuk,et al.  Epigenetic profiling of multidrug-resistant human MCF-7 breast adenocarcinoma cells reveals novel hyper- and hypomethylated targets , 2007, Molecular Cancer Therapeutics.

[18]  H. Magdelenat,et al.  Prognostic Value of the S‐Phase Fraction of Breast Cancers Treated by Primary Radiotherapy or Neoadjuvant Chemotherapy , 1993, Annals of the New York Academy of Sciences.

[19]  Rork Kuick,et al.  Molecular profiling of pancreatic adenocarcinoma and chronic pancreatitis identifies multiple genes differentially regulated in pancreatic cancer. , 2003, Cancer research.

[20]  C. Pérez-Plasencia,et al.  A phase II study of epigenetic therapy with hydralazine and magnesium valproate to overcome chemotherapy resistance in refractory solid tumors , 2007 .

[21]  F. Sato,et al.  Pharmacologic unmasking of epigenetically silenced tumor suppressor genes in esophageal squamous cell carcinoma. , 2002, Cancer cell.

[22]  D. Kramer,et al.  Polyamine catabolism in platinum drug action: Interactions between oxaliplatin and the polyamine analogue N1,N11-diethylnorspermine at the level of spermidine/spermine N1-acetyltransferase. , 2004, Molecular cancer therapeutics.

[23]  R. Weinberg,et al.  Estrogen-dependent cyclin E-cdk2 activation through p21 redistribution , 1997, Molecular and cellular biology.

[24]  A. Schmidt,et al.  S100P Stimulates Cell Proliferation and Survival via Receptor for Activated Glycation End Products (RAGE)* , 2004, Journal of Biological Chemistry.

[25]  D. Ferguson,et al.  A model of quiescent tumour microregions for evaluating multicellular resistance to chemotherapeutic drugs , 2005, British Journal of Cancer.

[26]  Shraddha S. Nigavekar,et al.  RAGE Activation by S100P in Colon Cancer Stimulates Growth, Migration, and Cell Signaling Pathways , 2007, Diseases of the colon and rectum.

[27]  J. Herman,et al.  SOCS-1, a negative regulator of the JAK/STAT pathway, is silenced by methylation in human hepatocellular carcinoma and shows growth-suppression activity , 2001, Nature Genetics.

[28]  J. Winstanley,et al.  Induction of metastasis by S100P in a rat mammary model and its association with poor survival of breast cancer patients. , 2006, Cancer research.

[29]  Ram Varma,et al.  Platinum drug effects on the expression of genes in the polyamine pathway: time-course and concentration-effect analysis based on Affymetrix gene expression profiling of A2780 ovarian carcinoma cells , 2007, Cancer Chemotherapy and Pharmacology.

[30]  A. Yokomizo,et al.  Inositol 1,4,5-trisphosphate (IP3) receptor type1 (IP3R1) modulates the acquisition of cisplatin resistance in bladder cancer cell lines , 2005, Oncogene.

[31]  C. Mathew,et al.  Disruption of the Fanconi anemia–BRCA pathway in cisplatin-sensitive ovarian tumors , 2003, Nature Medicine.

[32]  M. Gottesman,et al.  Reduced expression of small GTPases and hypermethylation of the folate binding protein gene in cisplatin-resistant cells , 2004, British Journal of Cancer.

[33]  R. Poon,et al.  Differential responses of proliferating versus quiescent cells to adriamycin. , 1999, Experimental cell research.

[34]  T van Doorn,et al.  Tumour resistance to cisplatin: a modelling approach , 2005, Physics in medicine and biology.

[35]  S. Itohara,et al.  Causal Relationship between the Loss of RUNX3 Expression and Gastric Cancer , 2002, Cell.

[36]  P. Kantoff,et al.  Regulation of S100P expression by androgen , 1996, The Prostate.

[37]  M. Santoro,et al.  Highly selective toxic and proapoptotic effects of two dimeric ribonucleases on thyroid cancer cells compared to the effects of doxorubicin , 2004, British Journal of Cancer.

[38]  D. Quaglino,et al.  N1,N12‐bis(ethyl)spermine effect on growth of cis‐diamminedichloroplatinum(II)‐sensitive and ‐resistant human ovarian‐carcinoma cell lines , 1998, International journal of cancer.

[39]  A. Pegg,et al.  Spermidine/spermine N1-acetyltransferase transient overexpression restores sensitivity of resistant human ovarian cancer cells to N1,N12-bis(ethyl)spermine and to cisplatin. , 2005, Carcinogenesis.

[40]  Robert Brown,et al.  The Acquisition of hMLH1 Methylation in Plasma DNA after Chemotherapy Predicts Poor Survival for Ovarian Cancer Patients , 2004, Clinical Cancer Research.