Gastrointestinal Toxicity, Antiinflammatory Activity, and Superoxide Dismutase Activity of Copper and Zinc Complexes of the Antiinflammatory Drug Indomethacin Carolyn T. Dillon, † Trevor W. Hambley, † Brendan J. Kennedy, † Peter A. Lay,* ,† Qingdi Zhou, † Neal M. Davies, ‡ J. Ray Biffin, § and Hubert L. Regtop § Centre for Heavy Metals Research, School of Chemistry, University of Sydney, NSW, 2006, Australia, Faculty of Pharmacy, University of Sydney, NSW, 2006, Australia, and Biochemical Veterinary Research, Pty Ltd, Braemar, NSW, 2575, Australia Received August 16, 2002 Gastrointestinal (GI) toxicity is one of the major problems associated with antiinflammatory drugs. The complexation of the powerful antiinflammatory drug (IndoH) by metal ions, as a means of reducing GI toxicity, has been studied. The in vitro superoxide dismutase (SOD) activity, in vivo antiinflammatory activity, and gastrointestinal ulcerogenic properties of IndoH, [Cu 2 (Indo) 4 (DMF) 2 ], and [Zn 2 (Indo) 4 (DMA) 2 ] are reported. No SOD activity was observed for IndoH or [Zn 2 (Indo) 4 (DMA) 2 ], but [Cu 2 (Indo) 4 (DMF) 2 ] inhibited the reduction of nitroblue tetrazolium (NBT) at an IC 50 value of 0.23 μM. All three compounds exhibited antiinflammatory activity in male Sprague-Dawley rats at an equivalent Indo dose of 10 mg/kg following oral administration of the drugs in 2% CMC solution. The severity of the toxicity (macroscopic ulcerations) in the stomach following oral dosing with [Zn 2 (Indo) 4 (DMF) 2 ] was not significantly lower than that induced by IndoH (P ) 0.78). Gastric ulcerations induced by [Cu 2 (Indo) 4 (DMF) 2 ] were significantly lower than those induced by IndoH or [Zn 2 (Indo) 4 (DMA) 2 ](P ) 0.0012 and P ) 0.0175, respectively) but significantly greater than the control (P ) 0.0013). The intestinal ulcerations induced by [Cu 2 (Indo) 4 (DMF) 2 ] or [Zn 2 (Indo) 4 (DMA) 2 ] were approximately 15 times lower than those of IndoH. A further indicator of gastrointestinal toxicity, caecal haemoglobin, increased in the following order: control < [Cu 2 (Indo) 4 (DMF) 2 ] < [Zn 2 (Indo) 4 (DMA) 2 ] < IndoH. [Cu 2 (Indo) 4 (DMF) 2 ] exhibited the most promising results of the Indo complexes assayed, in that it exhibited SOD activity and the lowest gastrointestinal damage while also exhibiting antiinflammatory activity that was comparable to that for IndoH. Low-temperature EPR analyses also showed that the formulation used for [Cu 2 (Indo) 4 (DMF) 2 ] administration was crucial to the integrity of the complex. Introduction Indomethacin [IndoH, 1 I, N-(p-chlorobenzoyl)-5-meth- oxy-2-methyl-1H-indole-3 acetic acid] is a commonly used and powerful nonsteroidal antiinflammatory drug that is believed to act by inhibiting cyclooxygenase and, consequently, prostaglandin synthesis (1). There are two established isoforms of cyclooxygenase (COX), COX-1 and COX-2, that play different roles in the inflammatory process (1-3). It is believed that COX-2 primarily produces the prostaglandins involved in inflammation and mitogenesis while COX-1 acts to maintain normal physiological function by its control of the renal paren- chyma, gastric mucosa, platelets, and most other mam- malian tissues (1-3). The inhibition of COX-1 leads to gastrointestinal ulcerogenic toxicity, and as such, it is perceived that a drug that displays selective inhibition of COX-2 over COX-1 would be a safer and more effective * To whom correspondence should be addressed. E-mail: lay_p@ chem.usyd.edu.au. Telephone: +61-2-9351 4269. Fax: +61-2-9351 3329. † Centre for Heavy Metals Research. ‡ Faculty of Pharmacy. § Biochemical Veterinary Research. 1 Abbreviations: CMC, carboxymethyl cellulose; COX, cyclooxyge- nase (oCOX, ovine-cyclooxygenase, hCOX, human-cyclooxygenase); DMA, N,N-dimethylacetamide; DMF, N,N-dimethylformamide; DMSO, dimethyl sulfoxide; EPR, electron paramagnetic resonance; Hbpzbiap, 1,5-bis(1-pyrazolyl)-3-[bis(2-imidazolyl)methyl]azapentane); IC 50, con- centration at which 50% inhibition occurs; IndoH, N-[p-chlorobenzoyl)- 5-methoxy-2-methyl-1H-indole-3 acetic acid; MEM-PR, minimum es- sential medium without phenol red; NBT, nitroblue tetrazolium; NSAID, nonsteroidal antiinflammatory drugs; RPMI, Roswell Park Memorial Institute; SEM, standard error of the mean; SOD, superoxide dismutase. 28 Chem. Res. Toxicol. 2003, 16, 28-37 10.1021/tx020078o CCC: $25.00 © 2003 American Chemical Society Published on Web 12/10/2002