Development of a novel cell-based assay system EPISSAY for screening epigenetic drugs and liposome formulated decitabine

Abstract

BackgroundDespite 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.MethodsA 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.ResultsFollowing 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.ConclusionsThe EPISSAY bioassay system provides a novel and rapid system to compare the efficiencies of existing and newly formulated drugs that reactivate gene expression.

Advancements in the delivery of epigenetic drugs

“…A 50% higher potency was observed when the decitabine was encapsulated in the PEGylated liposomes. The EPISSAY was also shown to be able to indicate the potency of HDAC inhibitors such as Vorinostat [95]. The promoter used in this system – a CMV promoter – had no major changes in methylation before and after treatment with decitabine.…”