1806 J. zyxwvuts Org. Chem. zyxwvu 1995,60, zyxwvu 1806-1813 12-Oxo-LTB4, a Key Pivotal Intermediate in LTB4 Metabolism Subhash P. Khanapure, Sukumar Manna, and Joshua Rokach" Claude Pepper Institute and Department zyxwvuts of Chemistry, Florida Znstitute of Technology, 150 West University Boulevard, Melbourne, Florida 32901 Robert C. Murphy and Pat Wheelan Department of Pediatrics, National Jewish Center for Immunology and Respiratory Medicine, University of Colorado School of Medicine, Denver, Colorado 80206 William S. Powell Meakins-Christie Laboratories, McGill University, 3626 St-Urbain St., Montreal, Quebec H2X 2P2, Canada Received December 14, 1994@ The first total synthesis of the very unstable and elusive biochemical intermediate 5(S)-hydroxy- zy 12-oxo-6(Z),8(E),10(E),14(Z)-eicosatetraenoic acid (12-oxo-LTB4) (2) has been accomplished. Incuba- tion of the synthetic 12-oxo-LTB4 (2) with human keratinocytes produced 5(S),12-dihydroxy-6- glutathionyl-7(E),9(E),14(Z)-eicosatrienoic acid (c-LTB3) and 5(S),12-dihydroxy-6-cysteinylglycyl- 7(E),9(E),14(Z)-eicosatrienoic acid (d-LTBz), two new intriguing mediators which have been identified recently during the incubations of LTB, with keratinocytes. We have also confirmed the identity of 12-oxo-LTB4 prepared from porcine leukocytes by comparison with the synthetic material and identified the formation in these incubates of the 6-trans-isomer of 12-oxo-LTB4. Introduction Scheme 1 The 5-LO-mediated transformation of arachidonic acid yields two products, 5-hydroperoxyeicosatetraenoic acid (5-HPETE) and 5(5'),6(R)-truns zyxwvut 5,6-epoxy-7,9-truns-11,- 14-cis-eicosatetraenoic acid (LT&). The enzyme is un- usual in the sense that it does mediate the two steps, the oxygenation of arachidonic acid at the 5-position and the dehydration to yield the LT&, Scheme 1. A great deal of work has been done by us and other groups on the 5-LO and its translocation mechanism.' LT& is a substrate to two enzymes. The first, LT& hydrolase, produces 5(S),12(R)-dihydroxy-6,14-cis-8,10- trans-eicosatetraenoic acid (LTB,) a potent chemotactic factor for proinflammatory cells such as polymorpho- nuclear leukocytes (PMNL) and is implicated in the pathology of psoriasis, inflammatory bowel diseases, and arthritis; the second, LTC4 synthase, only very recently identified, isolated, sequenced and cloned,2produces 56')- hydroxy-6(R)-glutathionyl-7,9,1l-truns-14-cis-eicosatet- raenoic acid (LTCJ, 5(S)-hydroxy-6(R)-cysteinylglycyl- 7,9,1l-truns-14-cis-eicosatetraenoic acid (LTD,), and 5(S)- hydroxy-6(R)-cysteinyl-7,9,1l-truns-14-cis-eicosa- tetraenoic acid (LTE4), which are potent smooth muscle constrictors in the lung and vasculature. The peptido G m " -c - - _ Arachidonic acid S(S)-HPETE 1 LTA4 LTA, hydrolaw/ synthase LTD4 LTE4 '' To whom correspondence should be addressed. Fax No.: 1 407 @ Abstract published in Advance ACS Abstracts, March 1, 1995. (1) (a) Samuelsson, B.; Rouzer, C. A,; Matsumoto, T. Adu. Prostag- 952-1818. Email: rokach@roo.fit.edu. landzn. Thromboxane Leukotriene Res. 1987. 17. 1. (b) Miller. D. K.: leukotrienes are implicated in asthma as the main tion of the microvasculature. LTBA and LTCA exert their b~onchoconstrictive agents and rhinitis by vasoconstric- Gillard, J. W.; Vickers, P. J.; Sadowski, S.; 'Leveille, C.; Mancini, J: A,; Charleson, P.; Dixon, R. A. F.; Ford-Hutchinson, A. W.; Fortin, R.; Gauthier, J. Y.; Rodkey, J.; Rosen, R.; Rouzer, C.; Sigal, I. S.; Strader, C. D.; Evans, J. F. Nature 1990, 343, 278. (c) Dixon, R. A. F.; Diehl, R. E.; Opas, E.; Rands, E.; Vickers, P. J.; Evans, J. F.; Gillard, J. W.; Miller, D. K. Nature 1990, 343, 282. tdl Gillard, J. A,; Ford- Hutchinson, A. W.; Chan, C.; Charlson, s.; Denis, D.; Foster, A,; Fortin, R.; Leger, S.; McFarlane, C. S.; Morton, H.; Piechuta, D.; Riendeau, D.; Rouzer, C. A,; Rokach, J.; Young, R.; MacIntyre, D. E.; Peterson, L.; Bach, T.; Eiermann, G.; Hopple, S.; Humes, J.; Hupe, L.; Luell, S.; Metzger, J.; Meurer, R.; Miller, D. K.; Opas, E.; Pacholok, S. Can. J. Physiol. Pharmacol. 1989,67,456. le) Rouzer, C. A,; Ford-Hutchinson, A. W.; Morton, H. E.; Gillard, J. W. J. Bid. Chrrn. 1990, 1436. biological effects through dedicated high affinity recep- tors. Drugs inhibiting the formation of LTC4 and LTBI (2) (a)Nicholson, D. W.; Klemba, M. W.; Rasper, D. M.; Metters, K. M.; Zamboni, R. J.; Ford-Hutchinson, A. W. Eur. J. Biochem. 1992, 209, 725. (b) Nicholson, D. W.; Ali, A.; Vaillancourt, J. P.; Calaycay, J. R.; Mumford, R. A,; Zamboni, R. J.; Ford-Hutchinson, A. W. Proc. Natl. Acad. Sci. USA. 1993,90, 2015. (c) Lam, B. K.; Penrose, J. F.; Freeman, G. J.: Austen, K. F. Proc. Natl. Acad. Sei. USA. 1994, 91, 7663. td) Welsch, D. J.; Creely, D. P.; Hauser, S. D.; Mathis, K. J.; Isakson, P. C. Proc. Natl. Acad. Sci. U.S.A. 1994, 91, 9745. 0022-3263/95/1960-1806$09.00/0 0 1995 American Chemical Society