Inhibition of L-leucine methyl ester mediated killing of THP-1, a human monocytic cell line, by a new anti-inflammatory drug, T614

T614 (3-formylamino-7-methylsulfonylamino-6-phenoxy-4H-1-benzopyran-4-one) is a member of the family of methane-sulfonanilide non-steroidal anti-infla mmatory drugs (mNSAIDs), most of which act as cyclooxygenase (COX)-2 inhibi tors. L-leucine methyl ester (Leu-OME) is a reagent which has been shown to kill phagocytes following interaction with intracellular proteases. There are two pathways whereby Leu-OME becomes cytotoxic to phagocytes. Within ly sososmes, Leu-OME is converted into free Leu, which causes disruption of th e lysosomes and subsequent cell necrosis. The other is the conversion of Le u-OME into (Leu-Leu)(n)-OME, which is associated with the induction of apop tosis. In the present study, we examined the action of T614 on Leu-OME medi ated killing of THP-1, a human monocytic cell line. We revealed that T614 a nd phenylmethyl sulfonyl fluoride (PMSF), a serine protease inhibitor, inhi bited Leu-OME mediated killing of THP-1 cells. All the other mNSAIDs, inclu ding nimesulide (NIM-03), fluosulide (CGP28238), FK3311 and NS398, also res cued THP-1 from Leu-OME mediated killing, although to a lesser degree. Of t he classical NSAIDs tested, a protective effect was observed with diclofena c at high concentration, but not with naproxen or indomethacin. Unlike conv entional lysosomal inhibitors, such as chloroquine and ammonium chloride (N H4Cl), T614 and PMSF did not raise lysosomal pH, as measured by flow cytome try using fluorescein isothiocyanate dextran (FITC-dextran). Therefore, the mechanism whereby T614 and PMSF inhibit Leu-OME killing is distinct from t hat of chloroquine or NH4Cl. Based on the similarity of T614 and PMSF, we s uggest that, besides their roles as COX-2 inhibitors, T614 and other mNSAID s may act as lysosomal proteose inhibitors. (C) 2000 Elsevier Science B.V. All rights reserved.