Purpose: Cytosine arabinoside induces apoptosis and this cell death process is influenced by protein kinase C signaling events in leukemic cells. We present findings that extend these observations to include another deoxycytidine analog, gemcitabine, which is more potent in solid tumors.
Methods and results: Gemcitabine induced programmed cell death in BG-1 human ovarian cancer cells based on biochemical and morphologic analyses. The DNA was fragmented in BG-1 cells exposed to gemcitabine (0.5 microM, 1.0 microM and 10 microM) for 8 h, but gemcitabine treatment did not induce internucleosomal DNA degradation. Scanning and transmission electron microscopy of BG-1 cells showed morphologic changes associated with apoptosis in response to gemcitabine: membrane blebbing, the formation of apoptotic bodies and chromatin condensation. Thus, BG-1 cells undergo programmed cell death in response to gemcitabine treatment without internucleosomal DNA fragmentation. Furthermore, gemcitabine (10 microM) activated protein kinase C in BG-1 cells and the phosphorylation of the endogenous protein kinase C substrate, myristoylated alanine-rich C kinase substrate, was increased following exposure of BG-1 cells to gemcitabine for up to 6 h. Clonogenicity studies with gemcitabine in combination with various protein kinase C-modulating agents demonstrated that gemcitabine cytotoxicity was influenced by protein kinase C signaling events in BG-1 cells. Short-term (1 h) exposure to TPA (1 or 10 nM) followed by gemcitabine (0.5 microM for 4 h) did not alter the response to gemcitabine. However, a 24-h exposure to TPA followed by gemcitabine resulted in synergistic cytotoxicity, while coincubation of TPA with a PKC inhibitor (e.g. bisindolylmaleimide or calphostin-C) in this regimen abrogated the synergistic response.
Conclusions: Based on our findings, it is plausible that gemcitabine therapy could be improved by modulating PKC signaling events linked to drug-induced apoptosis/cytotoxicity.