729. C-Met Antisense Plasmid Liposome Complex Increases Radiation Sensitivity of Lung Tumor Cells In Vivo or In Vitro

Abstract

Lung cancer cells have an increased expression of hepatocyte growth factor (HGF) receptor. Stimulation of the HGF receptor may be responsible for the rapid growth of lung tumors. Transfection of lung tumor cells with a plasmid containing the c-Met antisense transgene decreases expression of hepatocyte growth factor (HGF) receptor and increases apoptosis. To determine whether the increased expression of the c-Met antisense sensitize cells to irradiation, three lung tumor cell lines (3LL murine cell line, 201T human cell line, and A549 human cell line) were transfected with either a control plasmid or a plasmid containing the c-Met antisense transgene. Irradiation survival curves were performed 24 hours later. Seven days later colonies were stained with crystal violet and colonies of greater than 50 cells were counted. There was no statistical difference in the irradiation survival curves for the control cells compared to the cells transfected with the control plasmid by the Do or the shoulder on the survival curve (n). In contrast in both A549 and 201T cell lines transfected with the c-Met antisense plasmid there was an increased irradiation sensitivity as seen by a decreased shoulder on the survival curve compared to the non-transfected irradiated cells (n = 3.05 |[plusmn]| 0.45 compared to 6.43 |[plusmn]| 1.03 and 3.00 |[plusmn]| 1.31 compared to 5.95 |[plusmn]| 0.21, respectively). Cells from the 3LL cell line transfected with the c-Met antisense plasmid demonstrated radiation sensitivity by a decreased Do compared to the control irradiated but nontransfected cells (1.40 |[plusmn]| 0.18 Gy compared to 1.73 |[plusmn]| 0.09 Gy, respectively). In a separate experiment, cells were grown on cover slips, irradiated to 10 Gy and 48 hours later, the percent apoptotic cells were determined using a tunnel assay. For the 3LL, A549 and 201T cell lines there was increased apoptosis following c-Met antisense transfection of the tumor cells (9.3 |[plusmn]| 0.9%, 68.1 |[plusmn]| 2.0%, 4.6 |[plusmn]| 0.5%, respectively, compared to 0% in the control nonirradiated cells). There was an additional increase in the percent of apoptotic cells following irradiation in the c-Met antisense transfected 3LL, A549, and 201T cells compared to the irradiated control non-transfected cells (91.6 |[plusmn]| 1.4%, 94.2 |[plusmn]| 0.8%, or 95.9 |[plusmn]| 1.1%, respectively, compared to 0% in the control irradiated cells). To determine the effect of c-Met antisense gene therapy in the in vivo orthotopic tumor model, C57BL/6J mice were injected intratracheally with 3LL lung tumor cells. Beginning one day later and continuing for the next three days, the mice were injected I.V. with c-Met antisense plasmid liposome complexes. The mice were irradiated 48 hours after tumor implantation to 18 Gy to the pulmonary cavity. Mice injected with the antisense c-Met plus 18 Gy had improved survival compared to mice injected with the 3LL cells alone plus 18 Gy of irradiation. Optimization of the delivery of the c-Met antisense plasmid liposome complexes may facilitate radiosensitization of lung cancer.