The Identification and Characterization of Hydrazinyl Urea-Based Antibacterial Agents through Combinatorial Chemistry LawrenceJ.Wilson,*TimothyW.Morris,QiminWu,PaulJ.Renick, Christian N. Parker, y MichaelC.Davis,HelanaD.McKeever,PaulM.Hershberger, A.GregSwitzer,GaryShrum,ShyamSunder,DavidR.Jones,ShariS.Soper, RoyL.M.Dobson,ThomasBurt,KennethL.MorandandMarkStella Procter & Gamble Pharmaceuticals, Healthcare Research Center, 8700 Mason-Montgomery Road, Mason, OH 45040, USA Received 10 January 2001; accepted 26 February 2001 Abstract—An effort to identify novel inhibitors of peptidoglycan synthesis with antibacterial activity resulted in the discovery of a series of biaryl urea-based antibacterial agents through isolation of a by-product from a mixture-based combinatorial library of semi-carbazones and subsequent parallel synthesis efforts. The compounds were shown to possess broad spectrum antibacterial activity against gram-positive drug resistant pathogens, and showed apparent specificity for disruption of the bacterial cell wall biosynthesis pathway. # 2001ElsevierScienceLtd.Allrightsreserved. The rapid emergence of drug resistant bacterial patho- gens emphasizes the need for new classes of anti- bacterial agents. 1 Historically, new types of compounds have been identified from the diverse pool of natural products, for which there are many proven clinically efficaciousagents.Overthepastfewdecades,onlyafew new classes have been identified (quinolones, oxazolidi- nones, etc.), indicating the great degree of technical dif- ficulty in such an effort. 1,2 The chemical diversity of compounds being generated using novel methods, espe- cially combinatorial chemistry, coupled with rapid screening programs offers the opportunity to identify new classes of antibacterials. Many existing anti- bacterial drugs act by direct inhibition of one or more key steps in the metabolic pathway for synthesis of bacterial cell wall peptidoglycan. These agents include b-lactams (penicillin and methicillin), glycopeptides (vancomycin), and a variety of other natural products (bacitracin, D-cycloserine, and fosfomycin). 1 In our program, we sought to identify cell wall active agents through screening and deconvolution of mixture-based combinatorial libraries, similar to the screening of nat- ural products via plant and animal extracts. The cell wallassayweimplementedwasonedesignedtomeasure the final product of over 20 different interdependent enzyme reactions that create peptidoglycan. Also, part of the strategy was to simultaneously screen against bacterial growth inhibition to identify chemicals which act as both inhibitors of the cell wall biosynthesis and have antibacterial properties. This report describes the serendipitous discovery of aryl hydrazine urea-based antibacterial agents identified from a 10,000 member indexed library of semi-carbazones. Solution-based combinatorial chemistry has been uti- lized to make mixture-based libraries of smaller pro- portions. 3 Identification of the active components can be addressed through synthesizing the library in a par- ticular format (indexed, scanning, etc.) and subsequent guideddeconvolution. 4 Inourprogram,abroadvariety of combinatorial libraries were screened in parallel for inhibition of growth, bacterial histidine kinases, 5 and peptidoglycan biosynthesis in whole cells of Staphylo- coccus aureus. Among the 20 plus libraries that were tested, only an indexed library of 10,000 semi-carba- zones (4, Scheme 1), showed activity in both cell wall- peptidoglycanbiosynthesisandgrowthinhibitionassays that led to successful isolation and follow-up of an active component. Semi-carbazones (4) and their deri- vatives (i.e., acyl hydrazones) exhibit diverse pharma- cology, and the aza linkage resident in this class has 0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved. PII:S0960-894X(01)00160-3 Bioorganic & Medicinal Chemistry Letters 11 (2001) 1149–1152 *Corresponding author at current address: The R.W. Johnson Par- maceutical Research Institute, 1000 Route 202, PO Box 300, Raritan, NJ08869,USA.Fax:+908-203-8109;e-mail:lwilson3@prius.jnj.com y Current address: Pharmacia, Discovery Technologies, 7000 Portage Road,Kalamazoo,MI49001,USA.