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

Purpose: DA-125[(8S, 10S)-8-(3-Aminopropa- noyloxyacetyl)-10-[(2,6-dideoxy- 2-fluoro-alpha -L-talopyranosyl) oxy]-7,8,9, 10-tetrahydro-6,8,11-trihydrox y-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 ce ll death, the signaling pathways responsible for DA-125-induced apoptosis w ere studied. Methods: Cytotoxicity and apoptosis were measured in H4IIE cel ls and cells were stably transfected with a dominant-negative mutant of JNK 1 (JNK1) by MTT and TUNEL assays. Inhibition of topoisomerase II activity was determined in vitro. Drug accumulation and DNA binding affinity were de termined by fluorescence spectroscopy. Results: The cytotoxicity of DA-125 was greater than that of doxorubicin (IC50 11.5 vs 70 muM). DA-125 induced apoptosis with 30-fold greater potency than doxorubicin. Inhibition of topo isomerase II by DA-125 was fourfold greater. The presence of excess beta -a lanine, a DA-125 moiety, failed to alter cytotoxicity and accumulation of D A-125, indicating that the improved cytotoxicity of DA-125 did not result f rom the p-alanine moiety. Greater cellular accumulation of DA-125 correlate d 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-indnced apopt osis was also decreased in JNK1(-)-transfected cells. Conclusions: DA-125 p otently 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 DA125 involv ed the pathway of JNK1, but not ERK1/2 or p38 kinase.