Abstract
Gemcitabine (GEM) is an agent commonly used in the treatment of non small cell lung cancer (NSCLC). GEM induces apoptosis in NSCLC cells indirectly by increasing functionally active Fas expression. To further explore the mechanisms involved in the activation by GEM of apoptosis extrinsic pathway in lung cancer cells, we evaluated the ability of GEM to upregulate the expression of FasL in NSCLC H292 (mucoepidermoid carcinoma) cell line and to increase the sensitivity of these cells to Fas-mediated killing of cytotoxic lymphocytes. Cells were cultured with and without GEM (0.05 μM) for 24, 48, 72 and 96 hrs, and FasL mRNA and protein were evaluated by real-time PCR, and by western blot and flow cytometry, respectively. Apoptosis of cells expressing FasL was evaluated by flow cytometry. Cytotoxicity of LAK and malignant pleural fluid (PF) lymphocytes against H292 cells was analyzed in presence and absence of neutralizing anti-Fas antibody (clone ZB4), by a flow cytometry-based assay.
Expression of FasL mRNA and protein in H292 cells after incubation with GEM was increased at all time-points and this increase was higher after 72 hrs. Accordingly, the percentage of apoptotic H292 cells expressing FasL was higher after 72 hrs. Cytotoxicity of LAK and PF lymphocytes was significantly increased after incubation of H292 cells with GEM and was partially inhibited by neutralizing anti-Fas antibody.
These data demonstrate that: 1) GEM induces an up-regulation of FasL in NSCLC cells triggering cell apoptosis via an autocrine/paracrine loop; 2) GEM is able to increase the sensitivity of NSCLC cells to cytotoxic activity of LAK and PF lymphocytes by activation of Fas/FasL signalling system.
- © 2011 ERS