Abstract A202: A role of AXL in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarinoma (PDAC) is the 4 th leading cause of cancer-related death in the US. Due to a combination of late diagnosis with highly metastatic nature and therapeutic resistance, it is a near lethal malignancy. The identification of genes forming the molecular basis of these attributes is essential for the development of improved therapies. AXL is a receptor tyrosine kinase that is implicated in many common epithelial malignancies and has been shown to play a role in invasion, migration and innate immunity. In this study, we investigated the clinical relevance and oncogenic potential of AXL in PDAC progression to assess its potential as a therapeutic target. We performed IHC analysis on a total of 306 human primary PDAC samples and found that AXL is highly expressed in 50% of samples tested, suggestive of a role in PDAC. Subsequent screening of cell lines by immunoblot demonstrated expression and activation of AXL protein in 17 of 19 PDAC cell lines. Subsequently, we selected 4 cell lines with varying levels of AXL expression, and generated derivatives of these lines with constitutive knockdown of AXL using shRNA technology. Two shRNAs were selected based on their effect on AXL expression. We found significant impairment of proliferation, the ability to form colonies in soft agar as well as on migration and invasion in a dose-dependent manner. These results suggest that AXL may be a contributing factor to the metastatic phenotype of PDAC. Targeted inhibition of AXL expression in vitro led to 5-fold reduction in ERK and 2-fold reduction in AKT activation compared to control cells, demonstrating that these signaling pathways mediate the effects of AXL on tumor growth and invasion. In order to address the role of AXL on tumor initiation in vivo, shRNA derivatives of Capan-2 and PaTu-8988T cells were injected subcutaneously into nude mice. We found that knockdown of AXL resulted in significant reduction of tumor growth in both models. However, the most striking observation was in diminished invasion in AXL-knockdown tumors compared to control xenograft tumors. Histopathological analyzes revealed extensive invasion of tumor cells beyond the cutaneous muscle in control xenograft tumors, whereas AXL-knockdown xenograft tumors lacked such invasion but had formed a fibrous capsule which separated the tumor mass from the skin. Furthermore, these tumors exhibited reduced degradation of matrigel matrix used in injection of tumor cells, suggesting that inhibition of AXL may lead to impairment of matrix-degrading enzymes. To assess if targeting of AXL might have potential therapeutic implications, we tested the effect of AXL inhibition on maintenance and progression of existing tumors. We used the doxycycline inducible shRNA system to knockdown expression once tumors had reached 120 mm 3 in size in Capan-2 and SW1990 xenograft tumors. Inducible inhibition of AXL greatly reduced tumor growth resulting in 36% and 60% reduction respectively, compared to control xenograft tumors, demonstrating the significance of continued AXL expression in tumor maintenance. Our data suggest that inhibition of AXL in patients could lead to reduced tumor growth and invasion warranting further investigation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A202.