Abstract 1924: Discovery and characterization of EMT positive and chemo-resistant novel population of HCC like stem cells

Abstract

Abstract Increasing evidence suggests that tumor heterogeneity facilitates relapse of liver cancer in patients during the post therapeutic period. This concept of heterogeneity has gained increasing support from the liver cancer stem cells (LCSC), which have the potential of differentiation and genetic and epigenetic alterations to adapt to the tumor microenvironment. With the technological advancement, researchers were able to identify new population of LCSCs besides CD133+ clones by discovering new surface markers-CD90, EpCAM, CD13 and CD24. These studies corroborated that CD133- population is highly heterogeneous and might have some additional markers. Recently, by screening colorectal cancer patient samples we discovered a novel surface marker vimentin (Vim) on metastatic liver nodules. We further initiated screening of primary hepatocellular carcinoma murine liver tumor samples and discovered the surface expression of Vim in both species of HCC. Interestingly, we purified Vim+CD133- from primary liver tumor cell suspensions. Using sphere assays, limited dilution assays and staining of the stem cell markers Sox2, Nanog and Oct4A, we showed that Vim+CD133- cells have stem cell like properties similar to Vim-CD133+ population. Then we performed differentiation assays with Vim+CD133- cells under suitable condition and showed hepatocyte like phenotypes by upregulating protein expression of albumin, periodic acid schiff's staining and cholangiocyte type morphologies- tubular and star shaped structures. Several studies correlated that cells undergo epithelial-mesenchymal transition (EMT)phenotypes when vimentin is expressed on their surface. Our investigation revealed that Vim+CD133- population supported EMT like phenotypes by presence of the twist and β-catenin inside the nucleus and absence of E-cadherin on cell surface. Contrary to this Vim-CD133+ population did not show EMT like phenotypes. In next step of our characterization of Vim+CD133- cells, we investigated its chemoresistance properties as LCSCs are well known to cause relapse in patients. We treated these cells with highly toxic doses of 5 Fluorouracil and cisplatin for 4 days and interestingly observed significantly more viability compared to Vim-CD133+ population. We also validated multidrug resistance properties such as expression of multidrug resistance protein 1 (MDR1) and anti-apoptotic markers in Vim+CD133- cells. Finally, we validated their tumorigenic potential using anchorage independent assays and in vivo inoculation in immunocompromised NOD-SCID mice. Collectively our studies characterized a novel population Vim+CD133- in heterogeneous LCSCs, which showed EMT like phenotypes and higher chemo-resistance compared to Vim-CD133+ cells. In the era of personalized cancer therapy, this population should be considered as superior druggable target compared to our canonical Vim-CD133+ cells to obtain greater therapeutic outcome. Citation Format: Abhisek Mitra, Arun Satelli, Xueqing Xia, Shulin Li. Discovery and characterization of EMT positive and chemo-resistant novel population of HCC like stem cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1924. doi:10.1158/1538-7445.AM2014-1924