Screening natural products for chemopreventive agents based on binding RXRalpha and alkylation of Keap 1.

摘要

Although there is no 'magic bullet' that can completely conquer cancer, many types of the disease might be avoidable. The chemoprevention of cancer, which uses pharmacological agents to impede, arrest or reverse carcinogenesis at its earliest stages, has great scientific promise. Natural products including plant and marine organisms are important sources of new chemopreventive agents but pose significant analytical challenges. The development of new mass spectrometric capabilities provide opportunities to speed up this natural product screening and drug discovery process for cancer chemoprevention agents. The advantage of mass spectrometry based screening method is that it can provide structural information of the hit compound in the screening process.;Mass spectrometry is ideal for studying interactions between small molecules and proteins. Cancer chemopreventive and treatment agents can function by interacting with functional proteins either through non-covalent binding or covalent modification which could both be studies using mass spectrometry. In this dissertation, two mass spectrometry-based assays were employed for screening botanical extracts for potential chemoprevention agents targeting functional proteins by either interaction.;(1) Screening marine extracts for ligands binding to human retinoid X receptor alpha (hRXRalpha) using ultrafiltration LC-MS-MS. A novel assay was developed and optimized. Structures of potential ligands of hRXRalpha-LBD from marine organism were characterized.;(2) Screening marine extracts for phase II inducers targeting human kelch-like ECH-associated protein 1 (hKeap1) by covalent modification. Ligustilide and its oxidative product from Angelica sinensis were confirmed as phase II enzyme inducers targeting hKeap1. To support mechanistic studies of how a phase II enzyme inducers up-regulate phase II genes, the relative reactivities of the cysteine residues in hKeap1 were profiled.