Withaferin A

Withaferin A is a steroidal lactone, derived from Acnistus arborescens, Withania somnifera and other members of family Solanaceae. It is the first member of the withanolide class of ergostane type product to be discovered.

Structure
Withanolides are a group of naturally occurring C28- steroidal lactones. They contain four cycloalkane ring structures, three cyclohexane rings and one cyclopentane ring. Withaferin A is highly reactive because of the ketone-containing unsaturated A ring, the epoxide in the B ring, and the unsaturated lactone ring. The double bond in ring A and the epoxide ring are mainly responsible for the cytotoxicity. The 22nd and 26th carbons of the ergostane skeleton in withaferin A and related steroidal compounds are oxidized to form a six-membered delta lactone unit. NMR spectral analysis identifies C3 in the unsaturated A ring as the main nucleophilic target site for ethyl mercaptan, thiophenol and L-cysteine ethyl ester in vitro. A library of 2, 3-dihydro-3β-substituted derivatives are synthesized by regio/stereoselective Michael addition to ring A.

Transcription factor NF-κB in vitro
NF-κB is a transcription factor that regulates many genes involved in cell survival, growth, immune response and angiogenesis. Withaferin A inhibits NF-κB at a very low concentration by targeting the ubiquitin-mediated proteasome pathway (UPP) in endothelial cells. In vitro experiments demonstrated that withaferin A inhibits other transcription factors including Ap1 and Sp1.

Biosynthesis
In the Withania somnifera plant, the withaferin A is present in the leaves. Withanolides are terpenoids, which are synthesized in plants using isoprenoids as precursors. Isoprenoids can be synthesized through mevalonate or 1-deoxy-D-xylulose 5-phosphate pathways. Isoprenogenesis significantly governs withanolide synthesis.

Isoprenoids form squalene, which then goes through a variety of intermediate steps to form 24-methylenecholesterol - the sterol precursor of the withanolides.

The biosynthesis of withaferin A uses enzymes such as squalene epoxidase (SQE), cycloartenol synthase (CAS), sterol methyl transferase (SMT), obtusifoliol-14 –demethylase (ODM). To produce withaferin A from 24-methylene cholesterol, the molecule undergoes several functional changes including formation of a ketone, epoxide, 2 hydroxyl groups, and lactone ring.