Abstract 3324: Epithelial mesenchymal transition is associated with chemoresistance of poorly differentiated hepatoma cells

Abstract

Abstract BACKGROUND: Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide. It is the most rapidly increasing type of cancers in the US. Chemotherapy has been largely ineffective for HCC treatment, especially in cases with poorly-differentiated HCC. Cancer stem cells (CSCs) are a critical subpopulation that may be responsible for the initiation, progression, metastasis and recurrence of malignancies. CD133 and aldehyde dehydrogenase (ALDH) are useful markers to identify CSCs in solid tumors, such as lung and pancreatic cancers. AIMS: To separate CD133+/ALDHhigh cells in well-defined hepatoma cell lines, and to evaluate chemosentivity based on their CSC marker profile. METHODS: Hepatoma cell lines: HepG2, Hep3B, Huh-7, HLE, HLF and SKHep1 were separated into CD133+/ALDHhigh and CD133−/ALDHlow subpopulations by FACS. The resulting subpopulations were seeded on non-attachable plates for spheroid formation or used for chemosentivity assays. Epithelial mesenchymal transition (EMT) was determined by immunocytochemical staining of E-cadherin and vimentin. Gene expression of procollagen type I (α1) (Procoll-I) and transforming growth factor β1 (TGF-β1) was determined by quantitative RT-PCR. RESULTS: In well-differentiated hepatoma cell lines, HepG2, Hep3B and Huh-7, the positivity for CD133+/ALDHhigh was 1.1, 13.4 and 15.6%; whereas, poorly-differentiated hepatoma cells were almost all negative for these two markers. The positivity for CD133/EpCAM was 2.3, 10.3 and 18.3% in HepG2, Hep3B and Huh-7 cells, whereas it was almost negative in HLE, HLF and SKHep1 cells. FACS-enriched CD133+/ALDHhigh HepG2, Huh-7 and Hep3B cells were able to form spheroids in non-attachable culture plates; whereas their negative counterparts failed. In contrast, CD133−/ALDHlow SKHep1, HLF and HLE cells formed larger spheroids with negative α-feto protein staining. More CD133−/ALDHlow HLE cells survived in the exposure to cisplatin or doxorubicin than their positive counterparts (p<0.05-0.01). Moreover, CD133+/ALDHhigh HepG2, Hep3B and Huh-7 cells were positive for E-cadherin, but negative for vimentin; whereas, their CD133−/ALDHlow cells displayed an opposite expression pattern of E-cadherin and vimentin. CD133−/ALDHlow HLE, HLF and SKHep1 cells were E-cadherin-negative and vimentin-positive. In accordance, mRNA levels of Procoll-I and TGF-β1 in CD133−/ALDHlow HLE cells were one-fold higher than their positive counterparts. CONCLUSIONS: In well-differentiated hepatoma cells, CD133+/ALDHhigh or CD133+/EpCAM+ cells appear to be a CSC/progenitor subpopulation; whereas, in poorly-differentiated hepatoma cell lines which are negative for these CSC markers, EMT may be responsible for their chemoresistance. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3324.