Evaluation of the cytotoxic effect of recombinant hexameric FasL protein (MegaFasL) on cultured thoracic cancers with emphasis on malignant pleural mesothelioma.

5144 Introduction. Direct targeting of apoptosis-inducing receptors using recombinant death ligands such as TRAIL or FasL has significant translational value as molecularly targeted therapeutics. Activation of Fas to induce apoptosis of cancer cells has been achieved by either agonistic antibodies or the recombinant FasL/enhancer complex, none of which is clinically applicable. In this study, we evaluated the anticancer property of MegaFasL (Apoxis, SA) in cultured thoracic cancer cells in vitro either alone or in combination with cytotoxic chemotherapeutic drugs. Materials and Methods. Twelve Fas-positive cancer cells (2 lung cancer, 3 esophageal cancer and 7 malignant pleural mesothelioma) or primary human normal cells (umbilical endothelial cells and epithelial keratinocytes) were treated with MegaFasL or with recombinant soluble FasL/enhancer complex (sFasL) at 5 to 100 ng/ml x 48 hours). In combination therapy, cancer cells were either pre-treated with cisplatin (0.25 to 3 microg/ml x 24hrs), paclitaxel (25 to 100 nM) x 3hours or concurrently treated with histone deacetylase inhibitors Trichostatin A (50 to 1000 nM) or MS275 (2.5 to 10 microM) x 12 hours. Cell viability and apoptosis were determined by MTT and TUNEL-based ApoBrdU assays respectively. Experiments were performed in quadruplicates. Results. 8/12 cultured cancer cells are intrinsically sensitive (defined as IC50 20-fold reduction of MegaFasL IC50 values. This is observed in both MegaFasL-sensitive and -resistant cells. While either chemotherapy alone or MegaFasL alone (10 or 25 ng/ml) mediates less than 30% cell death in representative MPM cells H513, H2052 or H211, >90% of cancer cells treated with drug combinations undergo apoptosis. More importantly, primary normal cells are refractory to combinations of MS275 (5 microM) + MegaFasL (10 ng/ml) or cisplatin (1 microg/ml) + MegaFasL that induce > 80% reduction of cell viability in multiple cancer cells. Conclusion. The recombinant hexameric MegFasL induces significant cytotoxicity in cultured thoracic cancer cells in vitro and this process is profoundly enhanced by different classes of chemotherapeutic drugs. Combinations of chemotherapy + MegaFasL at low drug concentrations are selectively cytotoxic to cancer cells and not primary normal cells. In vivo evaluation of the antitumor effect of cisplatin + MegaFasL or MS275 + MegaFasL on subcutaneous MPM H513 tumors in nude mice are ongoing.