1alpha,25-dihydroxyvitamin D3 Agonist and Histone Deacetylase Inhibitor Bifunctional Ligand Discovery Virtual Docking, Synthesis, Fluorescence Polarization-Based Screening, and Molecular Dynamics Simulations

The vitamin D receptor (VDR) and histone deacetylases (HDACs) are important chemotherapeutic targets. 1,25-dihydroxyvitamin D3 (calcitriol, or 1,25D) is the natural ligand of the VDR, stimulates immune responses, and inhibits cellular proliferation in a number of cancer cell lines. Small molecule HDAC inhibitors such as suberoylanilide hydroxamic acid (SAHA, Zolinza; Merck), block angiogenesis and promote cell apoptosis and differentiation and are being investigated as clinical treatments for a wide range of cancers. This thesis describes work on three projects focusing on the VDR and HDAC. In the first project, a hybrid molecule incorporating deacetylase activity into the structure of a 1,25D aromatic analog was identified in a virtual screen using FITTED, a docking program, synthesized, and tested. Surprisingly, it was found to be inactive as an agonist of the VDR. An in silico retrospective analysis of the novel hybrid compound and other non-steroidal VDR ligands revealed several general structural requirements for VDR activity. In a second project, computational modeling techniques including molecular dynamics simulations were employed to understand the differences between two ortho-aminoanilide hybrid compounds, which led one to act as a VDR agonist while the other acted as a VDR antagonist. Finally, as the mechanisms of anticancer activity of HDAC inhibitors are not well understood and current assays for HDAC inhibitors are cumbersome and not generally applicable to all HDAC isoforms, a fluorescence polarization assay for high-throughput screening of HDAC competitive inhibitors was developed. HDAC ligands combining the structures of SAHA and fluorescein were synthesized and determined to be well-suited as a fluorescence polarization assay probe.