ITH4012 (ethyl 5-amino-6,7,8,9-tetrahydro-2-methyl-4-phenylbenzol[1,8]naphthyridine-3-carboxylate), a novel acetylcholinesterase inhibitor with "calcium promotor" and neuroprotective properties

Ethyl 5-amino-6,7,8,9-tetrahydro-2-methyl-4-phenylbenzol[ 1,8] naphthyridine-3-carboxylate (ITH4012) is a novel tacrine derivative that can reduce cell death induced by various compounds with different mechanisms of action, such as thapsigargin ( reticular stress), H2O2 ( free radicals), and veratridine ( calcium overload), in bovine chromaffin cell. Cell viability, quantified as lactic dehydrogenase release, was significantly reduced by ITH4012 at concentrations ranging from 0.01 to 3 muM. In the human neuroblastoma cell line SH-SY5Y, ITH4012 also reduced amyloid beta(25-35)-induced apoptosis, determined by flow cytometry. ITH4012 caused a slight elevation in the cytosolic concentration of Ca2+ in fura 2-loaded bovine chromaffin cells, which could be related to the induction of protein synthesis relevant for cell survival. Blockade of protein synthesis by cycloheximide or blockade of Bcl-2' s active site with HA14-1 ( ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)- 4H-chromene-3-carboxylate) reversed the cytoprotective action of ITH4012. Furthermore, exposure of bovine chromaffin cells for 24 or 48 h to neuroprotective concentrations of this compound enhanced, nearly 3-fold, the expression of the antiapoptotic protein Bcl-2. In conclusion, ITH4012 is a tacrine derivative that maintains acetylcholinesterase-inhibiting activity (IC50 = 0.8 muM) but has the additional property of acting as a calcium promotor, a property leading to neuroprotection through the induction of antiapoptotic proteins.