Isothiazole Dioxides: Synthesis and Inhibition of Trypanosoma brucei Protein Farnesyltransferase Francesca Clerici, a, * Maria Luisa Gelmi, a Kohei Yokoyama, b,c Donato Pocar, a Wesley C. Van Voorhis, d Frederick S. Buckner d and Michael H. Gelb b,c, * a Istituto di Chimica Organica, Facolta ` di Farmacia, Universita ` di Milano, Via Venezian 21, 20133 Milan, Italy b Department of Chemistry, University of Washington, Seattle, WA 98195, USA c Department of Biochemistry, University of Washington, Seattle, WA 98195, USA d Department of Medicine, University of Washington, Seattle, WA 98195, USA Received 9 February 2002; accepted 19 April 2002 Abstract—A series of isothiazole dioxides was synthesized and evaluated as inhibitors of protein farnesyltransferase from the parasite that causes African sleeping sickness (Trypanosoma brucei). The most potent compound in the series inhibited the parasite enzyme with an IC 50 of 2 mM and blocked the growth of the bloodstream parasite in vitro with an ED 50 of 10 mM. The same compound inhibited rat protein farnesyltransferase and protein geranylgeranyltransferase type I only at much higher concentration. # 2002 Elsevier Science Ltd. All rights reserved. Protein farnesyltransferase (PFT) has been shown to be an ideal drug target for combating infections caused by the protozoan parasite Trypanosoma brucei, the causa- tive agent of African sleeping sickness. Not only are inhibitors of T. brucei PFT (TB-PFT) able to arrest the growth of cultured parasites, 1,2 but PFT inhibitors are well developed as pharmaceutically active agents that are now in clinical development for the treatment of cancer. 3 PFT catalyzes the transfer of the farnesyl group from farnesyl pyrophosphate (FPP) to the cysteine SH of the tetrapeptide motif CaaX (C is cysteine, a is usually but not necessarily an aliphatic residue, and X is a variety of different amino acids). The active site of TB-PFT, derived from the homology model using the X-ray structure of mammalian PFT as a template, shows that the residues that contact the farnesyl pyrophosphate (FPP) substrate and the aaX portion of the CaaX sub- strate are nearly identical to those found in mammalian PFT, but five of nine residues that contact the X portion of CaaX are different in the Trypanosomatid enzyme. 2 Regions of the active site not involved in FPP and CaaX binding also differ between the two PFTs. This leads to the prediction that some inhibitors of mammalian PFT may also inhibit TB-PFT with similar potency, but that species specific inhibitors should also be obtainable. In our previous studies we have shown that CaaX mimetics including FTI-276 inhibit mamma- lian PFT and TB-PFT in the low nanomolar range. 1,2 On the other hand, the non-peptide heterocycle SCH- 44342, which binds in the CaaX binding region but also extends into a remote enzyme pocket, is highly specific for mammalian PFT. 2 In the present study, we have iden- tified a class of isothiazole dioxides that inhibit TB-PFT in preference to the mammalian enzyme. Compounds used in this study belong to two different classes in the isothiazole dioxide series. The first class includes 3-diethylamino-4-(4-methoxyphenyl)-isothiazole 1,1-dioxides unsubstituted at C-5 or substituted with different substituents ranging from simple alkyl to aryl, or heteroaryl groups. To the second class belongs a series of 5-substituted 3-diethylamino-4-(4-methoxyphenyl)- isothiazole 1,1-dioxides and the corresponding 4,5- dihydro derivatives whose main feature is an S-atom as a linker between the isothiazole moiety and the substituent. Compounds 1 and 2 were synthesized through intramo- lecular cyclization of the corresponding a-ketosulfonyl- amidines with a known methodology. 4 This methodology is very useful but severely limited by the scarce availability of the sulfonylazides used as the 0960-894X/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved. PII: S0960-894X(02)00338-4 Bioorganic & Medicinal Chemistry Letters 12 (2002) 2217–2220 *Corresponding authors. Tel.: +39-02-5031-4480; fax: +39-02-5031- 4476; e-mail: francesca.clerici@unimi.it (F. Clerici); tel.: +1-206-543- 7142; fax: +1-206-685-8665; e-mail: gelb@chem.washington.edu (M. H. Gelb).