Abstract 4314: CD24 is a functional marker that mediates liver tumor initiation via regulation of Nanog
Authors: Antonia G. Castilho, Terence Kin Wah Lee, Stephanie Ma, Kwan Ho Tang, Irene Oi Lin Ng
Journal: Cancer Research (2010)

Abstract Hepatocellular carcinoma (HCC) is a disease with poor prognosis attributed to the high recurrence rate post-resection. Chemotherapy is often ineffective due to chemoresistance, conferred by the presence of tumor-initiating cells (TICs). Hence, to improve outcome, it is imperative that markers and their pivotal pathways involved in tumor initiation. Using HCC chemoresistant nude mice model, CD24 was found highly upregulated when compared to untreated tumor by cDNA microarray analysis. CD24 expression was first examined in HCC cell lines and human clinical HCC. CD24 expression across a HCC cell line panel ranged from 6.0% to 99.8%. Notably, no expression was detected in the non-tumorigenic hepatic cell line MIHA. Expression of CD24 was also found to represent only a minority (&amp;lt;1%) of the tumor cell population in 40% human HCC by immunohistochemistry. Using antibody-based magnetic and FACS-sorting respectively, CD24- and CD24+ tumor cells were isolated from three HCC patients and two HCC cell lines. The CD24+ HCC cells displayed the phenotype of self-renewing CSCs, including enhanced sphere-forming ability, higher anchorage-independent growth ability, capability of differentiation, and preferential expression of “stem-ness’ genes. Functional roles of CD24 in self-renewal and tumor initiation were further demonstrated by stable knockdown of CD24 expression in both HCC cell lines and clinical samples using lentiviral-based shRNA. When shCD24-Huh7 cells were injected subcutaneously into SCID mice, only 7/31 (22%) mice formed tumors, as compared to 20/31 (65%) tumors of significantly greater size, for controls. In addition, CD24-knockdown cells exhibited more rapid proliferation and chemo-sensitivity towards chemotherapeutic drugs. By quantitative PCR, CD24 mRNA was implicated in poor prognosis of 36 clinical HCC samples including tumor stages (p=0.024) and tumor recurrence(p=0.047). Notably, whether by cell sorting or gene-knockdown approach, it was shown by quantitative PCR and western blotting that CD24 and Nanog, a gene important for the self-renewal of embryonic stem cells, were co-expressed in HCC cells. We therefore hypothesize that the functional role of CD24 in liver tumor initiation is mediated through regulation of Nanog expression. Upon transfection of Nanog cDNA into CD24-knockdown cells, self-renewal and tumor formation were functionally recovered, suggesting Nanog as the downstream effector of CD24. Moreover, CD24 acts through nanog via phosphorylation of Stat3, which is the main upstream regulator of Nanog in embryonic stem (ES) cells Findings from this study indicate the role of CD24 as a marker of chemoresistant TICs in HCC, and introduce a novel mechanism through which tumor initiation may be effected by this marker. This opens the window towards further studies on HCC TICs, which may have important ramifications in future therapeutics against this deadly disease. 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 4314.