Small molecule disruptors of HuR-mRNA interaction as novel cancer therapy
Xiaoqing Wu1, Lan Lan1, Rebecca Marquez1, Bryan Tsao1, Jia Yu1, Amber Smith1, Philip Gao2, Na Zhang2, David Wilson3, Scott Lovell 4, Kawaljit Kaur1, Roberto De Guzman1, Ragul Gowthaman1,5, John Karanicolas1,5, Jeffrey Aubé6, Dan Dixon7 and Liang Xu1
1Department of Molecular Biosciences, 2COBRE-PSF Protein Purification Group, 3Laboratory for Early Stage Translational Research, 4COBRE-PSF Protein Structure Core, 5Center for Bioinformatics, 6Department of Medicinal Chemistry, 7Department of Cancer Biology, The University of Kansas Medical Center, The University of Kansas.
Post-transcriptional gene regulation is essential for normal development, but when dysregulated, has many implications in disease conditions, including cancer. The RNA- binding proteins (RBPs) are critical trans factors, they associate with specific cis elements present in mRNAs to modulate the stability and translation of target mRNAs. The RBP Hu antigen R (HuR) is highly abundant in many types of cancer. HuR promotes tumorigenesis by interacting with cancer-associated mRNAs, which encode proteins that are implicated in different tumor processes including cell proliferation, cell survival, angiogenesis, invasion, and metastasis. Our hypothesis is that small molecule compounds that disrupt the HuR-mRNA interaction will block HuR function, leading to the decay and reduced translation of target mRNAs critical for cancer cell growth and progression. High throughput screening (HTS) was carried out in several chemical libraries (~ 10,000 compounds) using fluorescence polarization (FP) assay and identified a series of initial hits with sub-micromolar inhibitory constants (Ki). Those potential disruptors were then validated by Alpha assay (Amplified Luminescent Proximity Homogeneous Assay), confirmed by Surface Plasmon Resonance (SPR). We are now performing NMR and X-ray crystallization to further confirm the binding of the lead compounds with HuR. In cell-based assays, one of the top hits, ST-3, specifically destabilized HuR target mRNAs and down-regulated corresponding proteins (Bcl-2, XIAP and Msi1). ST-3 significantly inhibited cancer cell proliferation but not normal cells. More cell-based assays are carrying out to validate the target specificity and investigate the mechanism of action using cell lines with HuR Knock-in/down. In conclusion, we identified potential small molecule disrupters of HuR-mRNA interaction as novel cancer therapy that inhibited cancer with HuR overexpression.