Abstract 5318: DPC4 loss results in the activation of alternative oncogenic pathways in pancreatic cancer

Abstract

Abstract Introduction: Pancreatic cancer remains one of the most lethal forms of cancer, with a five-year survival rate of less than five percent. Many of these patients succumb to complications from metastasis. We have shown that genetic inactivation of DPC4, a major factor in TGF-β signaling, is significantly correlated with widespread metastasis in pancreatic cancer patients at autopsy. We hypothesize that DPC4 loss may be driving pancreatic cancer metastasis by forcing the TGF-β signal down an alternative pathway. Methods: DPC4 was stably restored into three cell lines that harbor a homozygous deletion of the locus (A2.1, A6L, and BxPC3), forming isogenic pairs of DPC4−/− and DPC4+ cells. Matched DPC4−/− and DPC4+ cells were treated with TGF-β1 for 24 hours prior to protein extraction. Immunoblotting was performed for total and phospho Akt (Pi3K pathway), p44-42 (ERK pathway), and Stat3 (JAK-Stat pathway), as well as nuclear and cytoplasmic p65 (NFκB pathway), as these networks have previously been identified as being activated by TGF-β. Immunohistochemistry was performed on 50 surgically resected pancreatic cancers for pAkt. Results: DPC4 was successfully introduced into DPC4−/− cells, as evidenced by semi-quantitative PCR for DPC4, as well as a TGF-β luciferase assay. DPC4+ cells displayed characteristic differences in cell behavior, such as decreased proliferation, migration, and invasion, compared to DPC4−/− cells. Western blot analysis of Smad-independent TGF-β pathways in three sets of DPC4-complemented pancreatic cancer cell lines revealed patterns of preferential activation of the Pi3K and ERK signaling pathways in cells lacking DPC4. In surgically-resected pancreatic cancers, activation of Pi3K was associated with the presence of lymph node metastases (p=0.03), while concomitant DPC4 loss and Pi3K pathway activity was associated with poor survival in pancreatic cancer patients (p=0.02). Conclusions: Restoration of DPC4 into DPC4−/− cells results in the abrogation of Pi3K and ERK signaling, and simultaneous DPC4 loss and Pi3K activation correlates to a poor prognosis of pancreatic cancer patients at surgery. Further studies will confirm whether genotype-specific activation of the ERK and Pi3K pathways plays a role in pancreatic cancer cell behavior, particularly those processes that directly impact metastasis (i.e. migration and invasion). Nevertheless, it is intriguing to consider the Pi3K and ERK pathways as therapeutic targets in pancreatic cancer patients with loss of DPC4. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5318. doi:1538-7445.AM2012-5318