Global demethylation of rat chondrosarcoma cells after treatment with 5-aza-2'-deoxycytidine results in increased tumorigenicity

• Published in: PloS one 2009-12, Vol.4 (12), p.e8340
• Main Authors: Hamm, Christopher A, Xie, Hehuang, Costa, Fabricio F, Vanin, Elio F, Seftor, Elisabeth A, Sredni, Simone T, Bischof, Jared, Wang, Deli, Bonaldo, Maria F, Hendrix, Mary J C, Soares, Marcelo B
• Format: Article
• Language: English
• Subjects: Animals; Azacitidine - analogs & derivatives; Azacitidine - pharmacology; Brain cancer; Cancer; Cancer genetics; Cell Biology/Gene Expression; Cell Line, Tumor; Chondrosarcoma; Chondrosarcoma - genetics; Chondrosarcoma - pathology; CpG islands; Cytokines - genetics; Cytokines - metabolism; Decitabine; Demethylation; Deoxyribonucleic acid; DNA; DNA methylation; DNA Methylation - drug effects; DNA microarrays; DNA sequencing; Epigenesis, Genetic - drug effects; Epigenetics; Gene expression; Gene Expression Profiling; Gene Expression Regulation, Neoplastic - drug effects; Gene sequencing; Gene silencing; Genes; Genetic aspects; Genetic engineering; Genetics and Genomics/Epigenetics; Genetics and Genomics/Functional Genomics; Genetics and Genomics/Gene Expression; Genomes; Genomics; Invasiveness; Long Interspersed Nucleotide Elements - genetics; Methylation; Mice; Microsatellite Repeats - genetics; Midkine; Molecular Biology/DNA Methylation; Neoplasm Invasiveness - pathology; Nucleotide sequence; Oligonucleotide Array Sequence Analysis; Oncology/Sarcomas; Rats; Satellite DNA; Sequence Analysis, DNA; SOXB1 Transcription Factors - genetics; SOXB1 Transcription Factors - metabolism; Stem cells; Tumorigenesis; Tumorigenicity; Xenograft Model Antitumor Assays
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0008340

Abnormal patterns of DNA methylation are observed in several types of human cancer. While localized DNA methylation of CpG islands has been associated with gene silencing, the effect that genome-wide loss of methylation has on tumorigenesis is not completely known. To examine its effect on tumorigenesis, we induced DNA demethylation in a rat model of human chondrosarcoma using 5-aza-2-deoxycytidine. Rat specific pyrosequencing assays were utilized to assess the methylation levels in both LINEs and satellite DNA sequences following 5-aza-2-deoxycytidine treatment. Loss of DNA methylation was accompanied by an increase in invasiveness of the rat chondrosarcoma cells, in vitro, as well as by an increase in tumor growth in vivo. Subsequent microarray analysis provided insight into the gene expression changes that result from 5-aza-2-deoxycytidine induced DNA demethylation. In particular, two genes that may function in tumorigenesis, sox-2 and midkine, were expressed at low levels in control cells but upon 5-aza-2-deoxycytidine treatment these genes became overexpressed. Promoter region DNA analysis revealed that these genes were methylated in control cells but became demethylated following 5-aza-2-deoxycytidine treatment. Following withdrawal of 5-aza-2-deoxycytidine, the rat chondrosarcoma cells reestablished global DNA methylation levels that were comparable to that of control cells. Concurrently, invasiveness of the rat chondrosarcoma cells, in vitro, decreased to a level indistinguishable to that of control cells. Taken together these experiments demonstrate that global DNA hypomethylation induced by 5-aza-2-deoxycytidine may promote specific aspects of tumorigenesis in rat chondrosarcoma cells.