UP-LN1 is a poorly differentiated carcinoma cell line established from a lymph node (LN) metastatic lesion of the neck of a male patient with unknown primary. The CK7/CK20+/CEA+/SCCA phenotype detected in the original metastatic lesion and cultured cells led us to believe this cell line to be originated from a primary cancer of the gastrointestinal tract. UP-LN1 exhibited unique in vitro growth characteristics with naturally occurring floating (F) and adherent (A) cells. We hypothesize that the tumor-initiating cells (TICs) with cancer stem cell (CSC) properties may have differentially distributed in F and A cells of the UP-LN1 cell line contributing to cancer metastasis. We first made comparisons of TIC properties between F and A cells in terms of tumorigenicity in NOD/SCID mice. F and A cells were further examined for the expression of selected tumor markers and genes associated with TICs by DNA-microarray, RT-PCR and cytofluorometric analyses. Comparative sensitivities of the two cell types to cytolysis of non-MHC-restricted effector cells were also determined using 51Cr release cytotoxicity assays. Results can be summarized as follows. (i) Injection of F cells at as low as 103 cells could initiate tumor formation in NOD/SCID mice, while an equivalent value for A cells was 105. (ii) A panel of drug resistant gene mRNAs were tested, of which ABCG2, ABCA7, ABCB1, ABCC1 and ABCC4 were expressed in greater amounts in F cells as compared with A cells. (iii) F and A cells exhibited different phenotypes, CD44bright/CD24dim and CD44dim/CD24bright, respectively. (iv) Surface HLA class I expression was down-regulated in F cells, which could be restored after exposure to IFN-γ as low as 1 U/ml for 48 h. (v) F cells but not A cells exhibited extreme resistance to activated NK cells (CD56dim/CD16+ cytotoxic subset) sorted out from peripheral blood mononuclear cells followed by IL-2 (100 U/ml, 72 h) activation. Moreover, F cells turned out to be highly sensitive to be modulated by IFN-γ to express CXCR4, a receptor of CXCL12. Up-regulation of surface CXCR4 expression with a concomitant down-regulation of cytoplasmic expression of CXCL12 was observed in F cells which are highly resistant to activated NK killing. Such modulation was found to be mediated via IFN-γ, a mediator which could be secreted by the immunomodulatory NK subset (CD56bright/CD16) within activated NK cell population. In summary, we have here identified resistance to activated NK killing, depressed HLA class I expression and CD44bright/CD24dim to be the phenotype of highly tumorigenic F cells, in which TICs/CSCs predominantly reside. IFN-γ-mediated induction of surface CXCR4 expression and enhancement of HLA class I expression in F cells might have occurred in the patient bearing UP-LN1 cells, resulting in invasive and metastatic events.

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 4265.