Investigating the Mechanism of Action of Sanglifehrin A

Abstract

Macrocyclic natural products occupy a special niche in the small molecule chemical space. Compared with other classes of small molecules, their larger size and conformational flexibility confer a higher propensity to recognize relatively flat surfaces that are predominant in protein-protein interactions. They are also thought to be the smallest examples of biomolecules that display functional sub-domains. Examples in this class of compounds are the immunosuppressants cyclosporine A (CSA), FK506, rapamycin (RAP) and sanglifehrin A (SFA) – the subject of this dissertation.

Aside from their therapeutic use in the clinic, immunosuppressants CSA, FK506, and RAP have also been proven to be indispensable tools for interrogating signal transduction pathways at the molecular level. These natural products have significantly contributed to our understanding of T cell signal transduction pathways and the molecular events involved in T cell activation. SFA is another immunosuppressive macrocyclic natural product that is structurally distinct from CSA but which, like CSA, binds to an abundant intracellular protein, cyclophilin A (PPIA) with high affinity. Unlike CSA, neither RAP nor SFA affect calcium-dependent IL-2 production. However, distinct from RAP, SFA does not inhibit mTOR. The molecular target of SFA and the cellular events resulting from its effect are still poorly understood. This dissertation describes efforts to identify the molecular target of SFA and tease apart its mode of action by enlisting a variety of target identification and validation approaches.