FULL PAPER DOI: 10.1002/ejoc.200900443 Synthesis and Hormonal Activity of the (25S)-Cholesten-26-oic Acids – Potent Ligands for the DAF-12 Receptor in Caenorhabditis elegans René Martin, [a] Eugeni V. Entchev, [b] Frank Däbritz, [a] Teymuras V. Kurzchalia, [b] and Hans-Joachim Knölker* [a] Keywords: Diastereoselectivity / Hormones / Oxidation / Protecting groups / Steroids Using a highly stereoselective Evans aldol reaction for the introduction of the stereogenic center at C-25, we describe an efficient synthesis of the orthogonally diprotected (25S)- 26-hydroxycholesterol 11. In a few synthetic steps, this cru- cial intermediate 11 has been converted into the four (25S)- cholesten-26-oic acids 1–4, which have been obtained in 12– 15 steps and 19–53 % overall yield based on commercially available 3β-hydroxychol-5-en-24-oic acid (5). Our biological studies of the compounds 1–4 reveal that (25S)-Δ 7 -dafach- Introduction Reproductive development of nematodes such as Caeno- rhabditis elegans and Pristionchus pacificus is controlled by steroidal ligands, called dafachronic acids (Figure 1). [1,2] In C. elegans , the biosynthesis of these steroids requires ac- tivity of the cytochrome P450 DAF-9. [3] Dafachronic acids are ligands which inactivate the nuclear hormone receptor DAF-12 and thus, lead to reproductive development of worms. In daf-9 mutant worms, incapable of dafachronic acid biosynthesis, DAF-12 is active and worms enter the diapause state generating dauer larvae. Another ligand known to bind at DAF-12 is (25S)-cholestenoic acid (4). [4] Mangelsdorf and colleagues prepared (25S)-Δ 4 -dafach- ronic acid (2) and its C-25 epimer from the corresponding 26-hydroxycholesterols. [1a] In 2007, the structure of the other ligand, (25S)-Δ 7 -dafachronic acid (1), has been con- firmed by a synthesis from Corey and Giroux. [5] Moreover, it has been shown that (25S)-Δ 7 -dafachronic acid (1) repre- sents the most active ligand known so far. [1a,5] Interestingly, the synthesis of both C-25 epimers of 2 and 4 was described previously by Khripach et al. [6] Our investigations on the synthesis and biological activity of cholesterol derivatives, [7] led us to an elegant and concise synthesis of the 25R-dia- stereoisomers of 1, 2 and 4 starting from commercially [a] Department Chemie, Technische Universität Dresden, Bergstraße 66, 01069 Dresden, Germany Fax: +49-351-463-37030 E-mail: hans-joachim.knoelker@tu-dresden.de [b] Max-Planck-Institut für Molekulare Zellbiologie und Genetik, Pfotenhauerstraße 108, 01307 Dresden, Germany Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/ejoc.200900443. Eur. J. Org. Chem. 2009, 3703–3714 © 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim 3703 ronic acid (1) represents the most active steroidal ligand for the hormonal receptor DAF-12 in Caenorhabditis elegans. Moreover, the saturated (25S)-dafachronic acid (3) represents a new ligand for this receptor and the (25S)-steroidal acids are more active as compared to their corresponding (25R)- counterparts. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2009) Figure 1. Hormonally active steroidal acids 1–4. available diosgenin. [8,9] Also in the 25R-series, the Δ 7 - dafachronic acid exhibited the highest hormonal activity. [9] Since yamogenin, the C-25 epimer of diosgenin, was not available from commercial sources, we devised a novel stereoselective construction of the side chain for the synthe- sis of all three (25S)-cholesten-26-oic acids 1, 2 and 4, as well as the saturated (25S)-dafachronic acid (3). [10] The syn- thesis of (25S)-dafachronic acid (3) was also reported by Corey and co-workers. [11] Results and Discussion The Evans aldol reaction represents a powerful synthetic method for the enantioselective construction of stereogenic carbon centers. [12,13] However, applications to stereoselec- tive synthesis of steroid side chains are rare. [14] For our pur-