| Summary: | 碩士 === 國立中正大學 === 生命科學系暨分子生物研究所暨生物醫學研究所 === 98 === Aberrant DNA methylation identified within the promoter regions is associated with the tumorigenesis and varied from person to person. There is globe methylation differences among individual and tissuses but there was no whole genome targeting method thus far was developed to efficiently target cancer stem cell-like (CSC-like) or specific tumors. We developed a new method named the total knockdown of the cancer epigenome to identify and isolate the global methylation differences between two cells, baits versus target cells, and used the differences to kill the target cells. Subtraction PCR was combined with high through-put sequencing to identify and isolate the methylation differences between the estrogen receptor (ER) expressing (MCF7, ER+) versus the nonexpressing (MDA-MB-231, ER-) breast cancer cells; or mesenchymal stem cells (MSC) versus transformed, cancer stem cell-like MSC cells. The amplified methylation differences and semi-quantitative real-time PCR was performed to validate the identified loci. The isolated DNA were then methylated in vitro and used to treat the target cells. Global modification in DNA methylation was validated by Differential Methylation Hybridization and methylation within individual locus was validated by PCR as well. Gene expression was decreased after targeted DNA methylation. Treatment with the global methylation differences loci between two cells caused the cell death of MDA-MB-231 and reduced drug-resistance of the transformed MSC. Treatment with the global methylation differences loci between two cells also reduced or arreast the tumor size in the MDA-MB-231-inoculated immuno-deficient mice. Our data suggested this new method might be an efficient way to identify and target the methylation differences between two genomes and an alternative for cancer therapy.
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