Gene transcription mechanisms play critical roles in organismal development, cellular differentiation, and human pathophysiology. Selective pharmacological targeting of gene transcription has remained elusive. Recent evidence has implicated a transcription pathway mediated by RhoA and RhoC which modulates actin dynamics to induce the nuclear accumulation of the Myocardin-Related Transcription Factors (MRTF-A and MRTF-B). This activates transcription of a subset of serum-responsive genes that are targets of the Serum Response Factor (SRF) transcription factor. This transcription pathway plays key roles in cancer cell migration and metastasis as well as in pathological fibrosis in scleroderma, pulmonary fibrosis, and other diseases. We have identified, through a phenotypic screen, a novel series of oxadiazole-thioether compounds that disrupt this Rho/MRTF/SRF transcription pathway at single-digit nanomolar concentrations. They also block cancer cell migration and myofibroblast differentiation in vitro at nanomolar concentrations and prevent skin fibrosis in vivo. However, the direct molecular target of the compounds is not known. In this project, we will examine the mechanisms of action of these compounds to block Rho/MRTF/SRF-mediated transcriptional regulation. The high potency and well-defined structure-activity relationship (SAR) suggest that these compounds act at a novel druggable target which could be of broad significance. The immediate goal of this work will be to identify the molecular target of the compounds which will enhance our understanding of the regulation of the important Rho/MRTF/SRF transcription pathway. Knowing the molecular target would also facilitate structure-based design and optimization of these novel agents for eventual therapeutic use.
The specific aims of this project are: 1) To develop affinity reagents with which to label or isolate the molecular target of our highly potent oxadiazole-thioether inhibitor of MRTF/SRF-regulated gene transcription; 2) To undertake labeling and isolation of the molecular target followed by proteomic candidate identification; and 3) To validate candidate molecular targets by knock-down and overexpression approaches. A long-term goal of this research program will be to utilize this information to facilitate structure- based design to accelerate compound optimization and to conduct target-based high-throughput screening for novel chemical scaffolds. Successfully accomplishing the aims of this grant should reveal novel targets for control of a complex, regulated gene transcription mechanism. It will also greatly facilitate novel therapeutics discovery and development for critical clinical conditions such as scleroderma and pulmonary fibrosis as well as melanoma and breast cancer metastasis.
There is a critical need for new therapies for lethal diseases such as scleroderma and lung fibrosis as well as to prevent the spread of cancers. In this project, we will determine how new drug-like compounds work to block mechanisms involved in these diseases. This should speed their development as therapeutics for these important conditions.
