1. Development of a novel cell-based assay system EPISSAY for screening epigenetic drugs and liposome formulated decitabine
- Author
-
Sue Ping Lim, Diego J. Walther, Wen Wang, Raman Kumar, Kristen Ho, Paul M. Neilsen, David F. Callen, Rachel J. Suetani, Clive A. Prestidge, Yamini Akkamsetty, Lim, Sue Ping, Kumar, Raman, Akkamsetty, Yamini, Wang, Wen, Ho, Kitty, Neilsen, PM, Walther, Diego J, Suetani, R, Prestidge, Clive, and Callen, David
- Subjects
Cancer Research ,Antimetabolites, Antineoplastic ,Cell ,Azacitidine ,Genetic Vectors ,Decitabine ,Cytomegalovirus ,Antineoplastic Agents ,CB1954 ,Pharmacology ,Biology ,Hydroxamic Acids ,Epigenesis, Genetic ,Nitroreductase ,Genes, Reporter ,Genetics ,medicine ,Bioassay ,Nanotechnology ,Humans ,Epigenetics ,Breast ,RNA, Messenger ,Promoter Regions, Genetic ,Cells, Cultured ,Unilamellar Liposomes ,Regulation of gene expression ,Liposome ,Vorinostat ,Cell-based assay system ,Liposomes ,DNA Methylation ,Nitroreductases ,Repressor Proteins ,Luminescent Proteins ,medicine.anatomical_structure ,Oncology ,Technical Advance ,Gene Expression Regulation ,DNA methylation ,Drug Screening Assays, Antitumor ,Carrier Proteins ,medicine.drug ,Plasmids - Abstract
Background: Despite the potential of improving the delivery of epigenetic drugs, the subsequent assessment of changes in their epigenetic activity is largely dependent on the availability of a suitable and rapid screening bioassay. Here, we describe a cell-based assay system for screening gene reactivation. Conclusions: The EPISSAY bioassay system provides a novel and rapid system to compare the efficiencies of existing and newly formulated drugs that reactivate gene expression. Methods: A cell-based assay system (EPISSAY) was designed based on a silenced triple-mutated bacterial nitroreductase TMnfsB fused with Red-Fluorescent Protein (RFP) expressed in the non-malignant human breast cell line MCF10A. EPISSAY was validated using the target gene TXNIP, which has previously been shown to respond to epigenetic drugs. The potency of a epigenetic drug model, decitabine, formulated with PEGylated liposomes was also validated using this assay system. Results: Following treatment with DNA methyltransferase (DNMT) and histone deacetylase (HDAC) inhibitors such as decitabine and vorinostat, increases in RFP expression were observed, indicating expression of RFP-TMnfsB. The EPISSAY system was then used to test the potency of decitabine, before and after PEGylated liposomal encapsulation. We observed a 50% higher potency of decitabine when encapsulated in PEGylated liposomes, which is likely to be due to its protection from rapid degradation. Refereed/Peer-reviewed
- Published
- 2013