P5-06-09: Acquired Sensitivity to TRAIL Mediated Apoptosis in Lapatinib Resistant SKBR3 Cells.

Abstract

Abstract Introduction Lapatinib, a dual EGFR/HER2 tyrosine kinase inhibitor is approved for the treatment of trastuzumab-refractory HER2 positive metastatic breast cancer. However, not all HER2 positive tumors respond to lapatinib and patients who initially respond frequently relapse. Understanding the molecular alterations associated with acquisition of lapatinib resistance may lead to the identification of targets to overcome resistance. Thus, the aim of this study was to examine changes in apoptosis in a model of acquired lapatinib resistance. Methods SKBR3 cells were treated with 250 nM lapatinib twice weekly for 6 months to establish the lapatinib resistant cell line SKBR3-L. The effects of TNF-related apoptosis-inducing ligand (TRAIL) and tumour necrosis factor-α (TNF-α) on cell survival and apoptosis induction were examined in SKBR3 and SKBR3-L cells, using the acid phosphatase proliferation assay and the terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. Cleavage of poly ADP ribose polymerase (PARP) was used to confirm apoptosis. TRAIL-1 and TRAIL-2 receptor expression were measured by flow cytometry (FACS). Results In SKBR3-L cells, TRAIL treatment (25 ng/ml) significantly inhibited cell survival (94.0 ± 12.0 %) compared to the SKBR3 cells (11.0 ± 2.2%). Treatment with TNF-α (0.5 ng/ml) also resulted in significant inhibition of cell survival (68.2 ± 10.1 %) in SKBR3-L cells compared to SKBR3 cells (9.4 ± 9.0 %). TRAIL (25 ng/ml) induced a low level of apoptosis in SKBR3 cells (7.0 ± 4.0%), whereas in SKBR3-L cells, TRAIL (25 ng/ml) induced significant apoptosis (54.8 ± 2.6%, p < 0.001), as determined by the TUNEL assay. We confirmed by western blotting that TRAIL treatment for 72 hours resulted in higher levels of PARP cleavage in SKBR3-L cells compared to SKBR3 cells. Both western blotting and FACS analysis of TRAIL receptor expression showed that neither TRAIL-1 nor TRAIL-2 receptor expression was significantly elevated in SKBR3-L cells compared to SKBR3 cells. Conclusions SKBR3-L cells, a model of acquired lapatinib resistance, display significant sensitivity to TRAIL induced apoptosis compared to the parental cell line SKBR3. Thus, targeting TRAIL may represent a novel therapeutic strategy to overcome acquired lapatinib resistance. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P5-06-09.