A nucleosome sliding assay platform to screen inhibitors of SWI SNF chromatin remodeling complexes

PROJECT SUMMARY In this proposal EpiCypher Inc will develop the first commercially available assay to monitor nucleosome remodeling a major current focus for rational drug discovery that is starved of HTS tools Nucleosomes are the basic units of chromatin made up of bp DNA tightly wrapped around an octamer of histone proteins Chromatin remodeling complexes utilize ATP to disrupt DNA histone interactions and reposition nucleosomes thus regulating DNA access Aberrant nucleosome organization can severely disrupt gene expression DNA repair and cellular differentiation and is associated with cancer and disorders including schizophrenia chronic inflammation and intellectual disability The profound role of remodeling complexes in tumorigenesis and cancer progression represents a new target class for epigenetic drug discovery rapidly gathering attention from our commercial partners Of particular therapeutic interest are the SWI SNF family of chromatin remodeling complexes with of all human cancers harboring a subunit mutation SWI SNF complexes are comprised of a remodeling ATPase SMARCA BRM or SMARCA BRG three core proteins and an additional four to eight accessory proteins varying by cell type that direct genomic localization of the complex and stimulate catalysis The catalytic ATPase domains of SWI SNF complexes have been identified in multiple studies as rational targets for therapeutic development However chromatin remodeling assays amenable to high throughput screening are currently lacking a problem EpiCypher will address through this proposal We will assess the feasibility of using recombinant nucleosomes rNucs homogenously assembled on sliding DNA sequences as a high throughput biologically relevant screening platform to identify compounds that directly inhibit chromatin remodeling In Aim EpiCypher will develop an innovative rNuc based nucleosome sliding assay which capitalizes on a unique DNA template that can be radiolabeled after SWI SNF repositions the histone octamer to expose a Dam methylation site GATC In Aim we will validate the sliding rNuc substrates by assaying chromatin remodeler activity with the isolated SMARCA enzyme In Phase II we will focus on commercial production of this high throughput sliding rNuc screening platform for the identification of inhibitors that target the SWI SNF chromatin remodeling family This highly innovative research program will enable therapeutic development toward chromatin remodeling complexes a major class of drug targets that currently lack adequate screening tools PROJECT NARRATIVE Chromatin remodeling or the repositioning of nucleosomes regulates DNA access and plays major roles in human disease from inflammation and autoimmunity to cancer However the enzymes responsible for modulating nucleosome positioning are relatively inaccessible to HTS and drug development with no standard approaches to interrogate their modeling activities In this proposal EpiCypher will develop the first high throughput nucleosome sliding assay to accelerate the identification of therapeutics that target chromatin remodeling enzymes