DA-125, a novel anthracycline derivative showing high-affinity DNA binding and topoisomerase II inhibitory activities, exerts cytotoxicity via c-Jun N-terminal kinase pathway

Abstract.

Purpose: DA-125 [(8S,10S)-8-(3-Aminopropanoyloxyacetyl)-10- [(2,6-dideoxy-2-fluoro-α-L-talopyranosyl)oxy]- 7,8,9,10-tetrahydro-6,8,11-trihydroxy- 1-methoxy-5,12-naphthacene-dione hydrochloride] is a novel anthracycline derivative with anticancer activity. In the present study, we compared the cytotoxicity of DA-125 with that of doxorubicin in H4IIE rat hepatoma cells and investigated the mechanistic basis. Because activation of MAP kinases, in particular c-Jun N-terminal kinase (JNK), is implicated in apoptotic cell death, the signaling pathways responsible for DA-125-induced apoptosis were studied. Methods: Cytotoxicity and apoptosis were measured in H4IIE cells and cells were stably transfected with a dominant-negative mutant of JNK1 (JNK1^–) by MTT and TUNEL assays. Inhibition of topoisomerase II activity was determined in vitro. Drug accumulation and DNA binding affinity were determined by fluorescence spectroscopy. Results: The cytotoxicity of DA-125 was greater than that of doxorubicin (IC₅₀ 11.5 vs 70 µM). DA-125 induced apoptosis with 30-fold greater potency than doxorubicin. Inhibition of topoisomerase II by DA-125 was fourfold greater. The presence of excess β-alanine, a DA-125 moiety, failed to alter cytotoxicity and accumulation of DA-125, indicating that the improved cytotoxicity of DA-125 did not result from the β-alanine moiety. Greater cellular accumulation of DA-125 correlated with its high-affinity DNA binding. Although neither PD98059 nor SB203580 altered the degree of cytotoxicity induced by DA-125, JNK1^– cells exhibited about a twofold greater viability than control cells. DA-125-induced apoptosis was also decreased in JNK1^–-transfected cells. Conclusions: DA-125 potently inhibited topoisomerase II activity and induced apoptosis by a high rate of prooxidant production. DA-125 exhibited high-affinity DNA binding with improved cellular drug accumulation. Apoptosis induced by DA-125 involved the pathway of JNK1, but not ERK1/2 or p38 kinase