High-Throughput Fluorescence Anisotropy Screen for Inhibitors of VICKZ1 RNA Binding

The VICKZ protein family consists of RNA binding proteins (RBPs) that have important roles during development, helping to stabilize, localize, and translationally regulate mRNAs in many embryonic cell types. After birth, VICKZ1 expression is dramatically down-regulated and is almost non-detectable. In many types of cancers, however, VICKZ1 is expressed and has been correlated with many pro-oncogenic processes, including enhanced cell proliferation, suppression of apoptosis, resistance to anticancer drugs, tumor progression, and metastasis. VICKZ1 binds a number of specific mRNAs that encode proteins associated with transformed phenotypes. One such pro-oncogenic target is K-ras; VICKZ1 binding can lead to stabilization of the mRNA, elevated protein levels, and increased downstream signaling. Inhibition of VICKZ1 expression in mouse models inhibits tumor metastases.

Because VICKZ1 expression is tightly correlated with tumors and tumor progression in adults, it is a very good candidate for drug therapy. In collaboration with the Israel National Center for Personalized Medicine, we have performed a high-throughput fluorescence anisotropy screen for small molecule inhibitors of VICKZ1 RNA binding. By comparing 5’ fluorescently-labeled fragments of K-ras mRNA, we identified a sequence in its 3’UTR that binds VICKZ1. Using this fragment as a probe, we scanned over 60,000 compounds for those that would prevent VICKZ1 binding. This highly robust assay has yielded approximately 50 reproducible hits. These hits are currently being validated with orthogonal assays. Using computational methods and comparing the molecules that do and do not inhibit binding, we are also trying to model both the docking site in the protein and the molecular descriptions of the inhibiting compounds in the hopes of predicting additional, more active molecules that might inhibit VICKZ1 binding.