Efficient Loading of Sulfonamide Safety-Catch Linkers by Fmoc Amino Acid Fluorides Raffaele Ingenito,* Dijana Drez ˇnjak, Stefan Guffler, and Holger Wenschuh Jerini AG, Rudower Chaussee 29, 12489 Berlin, Germany ingenito@jerini.com Received January 28, 2002 ABSTRACT Fmoc-protected amino acid fluorides were found to be excellent reagents for the acylation of sulfonamide safety-catch linkers (SCL) suitable for the subsequent preparation of peptide C-terminal thioesters. High loadings were obtained on different types of resins with low levels of epimerization. The efficient acylation of sulfonamide type linkers 1 by means of Fmoc-protected amino acids with minimal epimerization at the chiral R-carbon is critical for the successful Fmoc- based solid-phase synthesis of thioesters by the safety-catch linker strategy. 2 Recently, it was reported that the reactivity of the sulfonamide function is comparable to that of an alcohol. 3 Accordingly, it was demonstrated that acylation of solid- support-bound alkanesulfonamide safety-catch linkers with Fmoc amino acids proceeds best under conditions previously described for effective acylation of alcohols (PyBOP, DIEA, DCM, or CHCl 3 , -20 °C). 4 However, long reaction times (8 or 16 h) and careful selection of reaction conditions were needed in order to obtain adequate loadings. 3 These results prompted a systematic study on the potential application of Fmoc-protected amino acid fluorides, 5 recently demonstrated to be highly reactive reagents for the acylation of very hindered amino acids and hydroxy functions on solid supports, 6,7 to the acylation of the sulfonamide safety-catch linker (SCL). Initially, model studies were carried out on the acylation of a commercial 4-sulfamylbutyryl SCL-PS resin (loading: 1.12 mmol/g). Fmoc-Ser(tBu)-OH, known to racemize readily under conditions 8 commonly used for solid-phase peptide synthesis, was used as a sensitive model system. The acid fluoride was synthesized via cyanuric fluoride 5 and used in the presence of various bases (DMAP, DIEA, NMI, TMP, and DABCO) and solvent systems (DMF, DCM, and CHCl 3 ). The highest acylation yields were obtained when DCM was used as solvent (for 2 equiv of DIEA; 0.5 M final concentration; reaction time 60 min; DCM 86%, CHCl 3 44%, DMF 38%). 9 Therefore DCM was chosen as the solvent of choice, and the efficiency of loading was examined in the presence of varying amounts of different bases. It was found that DMAP and DIEA (2 and 3 equiv) gave the highest loadings (Table 1). Racemization was tested for all bases (1.5, 2, or 3 equiv) via assembly of the dipeptide L-Ser-Phe- OMe and HPLC-based comparison of the peak area with (1) Backes, J. A.; Virgilio, A. A.; Ellman, J. A. J. Am. Chem. Soc. 1996, 118, 3055. (2) Ingenito, R.; Bianchi, E.; Fattori, D.; Pessi, A. J. Am. Chem. Soc. 1999, 121, 11369. (3) Backes, B. J.; Ellman J. A. J. Org. Chem. 1999, 64, 2322. (4) Kim, M. H.; Patel, D. V. Tetrahedron Lett. 1994, 35, 5603. See however: Coste, J.; Campagne, J.-M. Tetrahedron Lett. 1995, 36, 4253. (5) Carpino, L. A.; Sadat-Aalaee, D.; Chao, H. G.; DeSelms, R. H. J. Am. Chem. Soc. 1990, 112, 9651. (6) Wenschuh, H.; Beyermann, M.; Krause, E.; Brudel, M.; Winter, R.; Schu ¨mann, M.; Carpino, L. A.; Bienert, M. J. Org. Chem. 1994, 59, 3275. (7) Granitza, D.; Beyermann, M.; Wenschuh, H.; Haber, H.; Carpino, L. A.; Truran, G. A.; Bienert, M. J. Chem. Soc., Chem. Commun. 1995, 2223. (8) Di Fenza, A.; Tancredi, M.; Galoppini, C.; Rovero, P. Tetrahedron Lett. 1998, 39, 8529. (9) In general, loadings were determined by cleavage of the Fmoc group from a defined amount of resin using 20% piperidine in DMF for 20 min and UV measurement of the resulting piperidine-dibenzofulvene adduct at 301 nm. ORGANIC LETTERS 2002 Vol. 4, No. 7 1187-1188 10.1021/ol0256320 CCC: $22.00 © 2002 American Chemical Society Published on Web 03/05/2002