Cyclopamine DOI: 10.1002/anie.200902520 Synthesis of Cyclopamine Using a Biomimetic and Diastereoselective Approach** Athanassios Giannis,* Philipp Heretsch, Vasiliki Sarli, and Anne Stößel Dedicated to Philip A. Beachy, LynnF. James, and RichardF. Keeler The hedgehog signaling (Hh) pathway is a key regulator in embryogenesis and in the maintenance and generation of cells in adults. [1] Since aberrant activation of the Hh pathway leads to malignancies including basal cell carcinoma, medulloblas- toma, rhabdomyosarcoma, and prostate, pancreatic and breast cancers, inhibition of Hh signaling provides a route to novel anticancer therapies. [2, 3] The Veratrum alkaloid cyclopamine was the first known inhibitor of Hh signaling; it influences the balance between active and inactive forms of the protein Smoothened. Though high demand exists for cyclopamine as well as for metabolically more stable and potent analogues, an efficient chemical synthesis of this steroidal alkaloid is still missing. We report herein an expeditious synthesis of cyclopamine starting from commercially available dehydroepiandroster- one and making use of biomimetic and diastereoselective transformations. While one total synthesis strategy can be envisioned based on previous publications, [4–6] this approach is tedious, non-stereoselective, and low yielding. Our synthesis, which entails a copper-mediated regioselective C–H activa- tion/hydroxylation, cationic ring contraction/expansion, and finally an Alder-ene reaction, enables the rapid construction of this unusual molecule. This strategy also provides access to various cyclopamine analogues that do not exist in nature which might be useful in investigations of the biological activity of this unique natural product. Cyclopamine (1; Scheme 1) was first reported in 1957, when sheepherders in Idaho, USA, were alarmed by the unsettling observation of newborn lambs with a single eye located, cyclops-like, in the middle of their foreheads. [7] Intense investigation led to the finding that the ingestion of the corn lily (Veratrum californicum) by pregnant sheep caused the birth of lambs with cyclopia. The Veratrum alkaloid cyclopamine was identified as the causative agent. The malformation cyclopia has been known in humans since ancient times. It has been observed in severe cases of holoprosencephaly (HPE), a malformation of the brain in which the prosencephalon fails to cleave, resulting in undi- vided or incompletely divided cerebral hemispheres, telen- cephalon and diencephalon, and olfactory and optic nerves. [8] Associated with the brain abnormalities is a spectrum of midline craniofacial anomalies, including ocular hypotelorism and interorbital proboscis (ethmocephaly). [9] The etiology of HPE is heterogeneous and can include genetic and/or teratogenic factors. Only recently was the cellular target of cyclopamine elucidated. This molecule binds to the plasma membrane protein smoothened, induces a conformational change, and inhibits the hedgehog signaling pathway. [3] Cyclopamine (1) displays a plethora of unusual structural features, and therefore its synthesis poses significant chal- lenges. Among them are the C-nor -D-homo steroid substruc- ture and a highly substituted furan ring, which is attached through a spiro linkage to the D ring and also fused to a piperidine unit having a basic secondary nitrogen atom. Additionally, cyclopamine is quite acid sensitive: at pH < 3 or upon treatment of 1 with Lewis acids the furan ring is cleaved and the D ring aromatizes to form the toxic veratramine. Scheme 1. Structure and retrosynthetic analysis of cyclopamine (1). Bn = benzyl. [*] Prof. Dr. A. Giannis, Dipl.-Chem. P. Heretsch, Dipl.-Chem. A. Stößel Institut für Organische Chemie Fakultät für Chemie und Mineralogie, Universität Leipzig Johannisallee 29, 04103 Leipzig (Germany) Fax: (+ 49) 341-973-6599 E-mail: giannis@uni-leipzig.de Dr. V. Sarli Laboratory of Organic Chemistry, Department of Chemistry, University of Thessaloniki, 54124 Thessaloniki (Greece) [**] We thank Dr. Lothar Hennig for recording NMR spectra and for his help in interpreting the 2D NMR spectra. The Deutsche For- schungsgemeinschaft (DFG) is acknowledged for partial financial support. P.H. is a fellow of Fonds der Chemischen Industrie. Supporting information for this article (including the syntheses and spectroscopic data for the compounds shown) is available on the WWW under http://dx.doi.org/10.1002/anie.200902520. Angewandte Chemie 7911 Angew. Chem. Int. Ed. 2009, 48, 7911 –7914  2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim