Isoquinoline‐1,3,4‐trione and its derivatives attenuate β‐amyloid‐induced apoptosis of neuronal cells

Abstract

Caspase‐3 is a programmed cell death protease involved in neuronal apoptosis during physiological development and under pathological conditions. It is a promising therapeutic target for treatment of neurodegenerative diseases. We reported previously that isoquinoline‐1,3,4‐trione and its derivatives inhibit caspase‐3. In this report, we validate isoquinoline‐1,3,4‐trione and its derivatives as potent, selective, irreversible, slow‐binding and pan‐caspase inhibitors. Furthermore, we show that these inhibitors attenuated apoptosis induced by β‐amyloid(25–35) in PC12 cells and primary neuronal cells.

Isoquinoline-1,3,4-trione Derivatives Inactivate Caspase-3 by Generation of Reactive Oxygen Species

“…Consistently, structural results presented here indicate that only the catalytic cysteine but not the other cysteines in caspase-3 are oxidized to sulfonic acid in the crystal structures of caspase-3 treated with isoquinoline-1,3,4-trione derivatives and DTT. It is likely that the differences among the microenvironment of the active site of the caspases and other cysteine proteases surrounding the catalytic cysteine might render it with varied sensitivities to oxidation, which could explain the phenomena we found previously that these compounds have different selectivity against caspase-3, other caspases, and other proteases (22,23).…”

“…The activity of caspase-3, monitored continuously by the production of pNA from Ac-DEVD-pNA, decreased in a pseudo-first order process. Volume dilution and dialysis methods indicated that the inhibition is irreversible (23). The observed rate of inactivation increased nearly linearly with increased concentration of compound III, yielding an apparent inactivation rate constant (k obs /I) of 20,098 Ϯ 867 s Ϫ1 M Ϫ1 within the linear response zone (Fig.…”

“…Expression and Purification of Recombinant Caspase-3 and Its Mutants-The catalytic domain of human caspase-3 was expressed according to the procedure described previously (23). The cells that contained the His 6 -tagged caspase-3 were lysed on ice by sonication and the insoluble cell debris was removed by centrifugation at 15,000 ϫ g at 4°C.…”

“…Previously we have identified isoquinoline-1,3,4-trione as a novel caspase-3 inhibitor through high-throughput screening of a small molecule library using recombinant caspase-3 (22,23). A series of isoquinoline-1,3,4-trione derivatives developed through structural modification exhibit nanomolar potency against caspase-3 in vitro and are validated as selective, irreversible, slow-binding and pancaspase inhibitors.…”

“…A series of isoquinoline-1,3,4-trione derivatives developed through structural modification exhibit nanomolar potency against caspase-3 in vitro and are validated as selective, irreversible, slow-binding and pancaspase inhibitors. Furthermore, we have found that these inhibitors could attenuate apoptosis induced by ␤-amyloid (25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35) in PC12 cells and primary neuronal cells (23) and decrease in infarct volume in the transient middle cerebral artery occlusion stroke model (22). These caspase-3 inhibitors with novel structures have provided a new therapeutic strategy directed against diseases involving abnormally up-regulated apoptosis.…”