The anthracycline doxorubicin induces cathepsin B via activation of the transcription factor NFκB.

2260 Anthracyclines like doxorubicin are widely used for the treatment of many human neoplasms. However, their therapeutic index is compromised by side effects like the irreversible cardiotoxicity leading to heart failure. Although antineoplastic action and cardiac toxicity seem to be closely related, the underlying mechanisms are not entirely understood. Therefore, cDNA microarray analysis of the cervical carcinoma cell line HeLa after treatment with doxorubicin was performed. Out of 22283 genes on the chip, 3979 genes were regulated by a 24 h-incubation with doxorubicin. Among the 2004 genes that were upregulated, one group of genes encoding for the lysosomal protein family of cathepsins was strongly induced by doxorubicin treatment. Cathepsin B is of particular interest since this cystein protease is highly expressed in tumor cells and involved in intracellular protein degradation, angiogenesis, metastasis and apoptosis. Incubation of HeLa cells with doxorubicin resulted in an induction of mRNA and protein expression as well as enzyme activity in a time and concentration dependent manner as determined by Real Time PCR, Western Blot Analysis and enzyme activity assay. Usage of NFκB inhibitors HNE, CAPE, and dexamethason, IκBα immunoblotting and electrophoretic mobility shift assay (EMSA) revealed that cathepsin B induction in response to doxorubicin treatment resulted from activation of the transcription factor NFκB. In addition, inhibition of PI3K and PKC with the specific inhibitors LY294002 and bisindolylmaleimide resulted in a significant reduction of doxorubicin-mediated cathepsin B induction. Since it is known that cathepsin B is involved in apoptosis we further investigated the effect of NFκB mediated cathepsin B induction by doxorubicin on apoptosis by measuring activities of caspases 3 and 9 as early markers of apoptosis. After preincubation of cells with the cathepsin B inhibitor CA-074Me, activities of caspases 3 and 9 were strongly reduced in HeLa cells. These data are relevant since cathepsin B can be used as activating enzyme for specific tumor targeted therapies. Moreover, induction of cathepsin B by doxorubicin could increase the therapeutic efficiency of tumor targeted drugs.