Pancreatic cancer continues to be a devastating tumor (28,000 new cases/year; 5%-5 year survival). Pancreatic tumors frequently (90%) overexpress fibroblast growth factor ligands (FGF-1, FGF-2) and alternatively spliced high affinity receptors (FGFR-1b). (FGFR-1a previously found in normal pancreatic tissue).
Methods: In vitro: to study the significance of this observation, PANC-1 cells were stably transfected via the pMEXneo vector containing FGFR-1a (PANC-1a) or FGFR-1b (PANC-1b) isoforms. Cells were treated with 150 mM peroxynitrite, 25 and 100 mg/ml of 5-FU. Cells were evaluated for growth inhibition, apoptosis (by fluorescein diacetate/propidium iodide and annexin V staining, caspase 3 activation) and for Bcl-XL/BAX expression (by Western Blot). In vivo: 5x106 cells of each isoform were injected into nude Balb/c mice for xerograft formation (N=8).
Results: In vitro, both PANC-1a and PANC-1b exhibited similar growth rates. Compared to PANC-1b (31%), pro-oxidant (peroxynitrite)-induced apoptosis was significantly (p<0.05) increased in PANC-1a (50%). PANC-1a (70%) demonstrated significantly (p<0.05) increased sensitivity to the cytotoxic effects of 5-FU when compared to PANC-1b (32%). Increased cell death in PANC-1a correlated with increased cell-surface staining for annexin V, activation of caspase-3, and decreased expression of Bcl-XL/BAX. In vivo, PANC-1b readily demonstrated formation of tumor xerograft at 2 weeks; whereas, PANC-1a did not form tumors.
Conclusions: Alternative splicing of FGFR-1 to the b isoform appears to correlate with pancreatic cancer cell growth in vivo and resistance to chemotherapy. Inhibition of FGFR-1 splicing or overexpression of FGFR-1a inhibits pancreatic cancer cell growth in vivo and restores cytotoxic responses to chemotherapy, thereby suggesting the basis of rational interventional strategies for this devastating pathology.
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