Abstract 5000: The functional stem cell marker aldehyde dehydrogenase enhances pancreatic cancer stem cell isolation and correlates with clinical prognosis

Abstract

Abstract Pancreatic adenocarcinoma is a highly aggressive cancer that is largely resistant to current therapies and portends a median survival for patients of 9-12 months. Resistance may be due to a subpopulation of cells that are chemoresistant and able to support tumor propagation, such as cancer stem cells (CSCs). CSCs have emerged as attractive therapeutic targets, but the development of effective therapies requires the isolation of highly purified cell populations. Surface antigen expression has been used to identify CSC in most systems, however, methods based on functional properties, such as Hoechst dye exclusion or aldehyde dehydrogenase (ALDH) activity, can enhance stem cell isolation from normal and malignant tissues. We compared CSCs from pancreatic adenocarcinomas isolated by cell surface phenotype, ALDH activity, or both. Initially, we studied ALDH expression in human pancreatic cancer cell lines (CAPAN-1, DAN-G, CFPAC-1) using the Aldefluor assay. We found that each line contained a small population of ALDH+ cells (0.9-6.5% of total cells) that demonstrated increased clonogenic growth potential both in vitro and in vivo compared to ALDH- cells. Next, we compared ALDH expression with the published pancreatic CSC phenotype (CD24+CD44+) in cell lines and low-passage xenografts derived from primary human tumor specimens. Interestingly, we found that the putative CSC populations were only partially overlapping since 7-18.2% of the ALDH+ cells were CD44+CD24+ and only 4.2-36.2% of the CD44+CD24+ cells expressed ALDH. We compared CD44+CD24+ and CD44+CD24+ALDH+ cells isolated from 5 unique human pancreatic cancer xenografts and found that the CD44+CD24+ALDH+ cells were more tumorgenic in NOD/SCID mice. We also evaluated ALDH expression in primary pathologic specimen from 268 patients with pancreatic cancer by immunohistochemistry and found small, intensely positive cells with round or oval nuclei near the base of cancerous glands or at the invasive front in 90 (34%) of those patients. Furthermore, the presence of these cells was significantly associated with poorer overall survival (14 vs. 18 mos, p=0.02). We have begun to evaluate the mechanism by which pancreatic CSCs may be involved in more aggressive disease and found that CD44+CD24+ALDH+ cells isolated from tumor xenografts have significantly lower expression of E-cadherin and higher expression of Slug, a transcriptional repressor of E-cadherin, compared to ALDH+ or ALDH- cells. Therefore, ALDH appears to mark highly clonogenic pancreatic CSCs. Moreover, increased features associated with epithelial-to-mesenchymal transition may play a role in the correlation of ALDH+ cells with long-term outcomes.