Tumor-initiating activity and tumor morphology of HNSCC is modulated by interactions between clonal variants within the tumor

Abstract

Tumor initiation (TI) in xenotransplantation models of head and neck squamous cell carcinoma (HNSCC) is an inefficient process. Poor TI could be due to (1) posttransplant cell loss, (2) a rare sub-population of cancer stem cells or (3) a requirement for specific cellular interactions, which rely on cell number. By tracking GFP-expressing HNSCC cells, we conclude that the posttransplant loss of cancer cells is minimal in the xenotransplant model. Furthermore, an examination of putative cancer stem cell markers (such as CD133, CD44, SP and label retention) in HNSCC cell lines revealed no correlation between marker expression and tumorigenicity. In addition, single-cell clones randomly isolated from HNSCC cell lines and then transplanted into mice were all capable of initiating tumors with efficiencies varying almost 34-fold. As the observed variation in the clones was both more and less tumorigenic than the parental cells, a combination of two clones, at suboptimal cell numbers for TI, was implanted into mice and was found to modulate the tumor-initiating activity, thus indicating that TI is dependent on a 'critical' number of cells and, for the first time, that interactions between clonal variants within tumors can modulate the overall tumor-initiating activity. Put in context with previous literature on tumorigenic activity, we believe that interactions between clonal variants within a tumor as well as (1) stromal interactions, (2) angiogenic activity, (3) immunocompetence and (4) cancer stem cells may all contribute to tumorigenic potential and the propensity for tumor growth and recurrence.

Dueling models in head and neck tumor formation

“…Single cell clones randomly isolated from HNSCC cell lines were all capable of initiating tumors after implantation into mice (Cameron et al). 3 This result provides support for the clonal evolution model. It is to be noted, however, that after subcloning, cells were propagated for two to five passages in vitro before the implantation.…”

“…In the Cameron paper, GFP labeling to trace implanted cells obviated this possibility. 3 Isolated subpopulations of tumor cells with stem cell-like features can form solid tumors in vivo. To identify solid tumor CSCs, tumor cells are fractionated using cell surface markers and implanted into immunodeficient mice, after which xenograft growth and cellular composition are assessed.…”

A Novel E2F/Sphingosine Kinase 1 Axis Regulates Anthracycline Response in Squamous Cell Carcinoma

“…Stocks of BGT226 were prepared as described previously (19). Viability was determined using trypan blue, Cell Titer 96 Aqueous One Solution Cell Proliferation Assay (Promega), or Western blot analysis for cleaved caspase-3 or PARP cleavage as described previously (20,21). Sphk1 activity and S1P levels were estimated using commercially available kits (Echelon Biosciences).…”

Dysregulation of the Repressive H3K27 Trimethylation Mark in Head and Neck Squamous Cell Carcinoma Contributes to Dysregulated Squamous Differentiation

“…Immunohistochemistry was conducted as described (31). Sections were incubated with the following primary antibodies: H3K27me3 1:100 (Upstate), BrdUrd 1:100 (Roche), Ae1/Ae3 1:600 (ICN Biomedical), involucrin 1:100 (Santa Cruz Biotechnology), Keratin 1 (Novus Biologicals), cleaved caspase III 1:500, and EZH2 1:50 (Cell Signaling).…”