The effect of 2D and 3D cell cultures on treatment response, EMT profile and stem cell features in head and neck cancer

Abstract

BackgroundHead and Neck Squamous Cell Carcinoma (HNSCC) tumors are often resistant to therapies. Therefore searching for predictive markers and new targets for treatment in clinically relevant in vitro tumor models is essential. Five HNSCC-derived cell lines were used to assess the effect of 3D culturing compared to 2D monolayers in terms of cell proliferation, response to anti-cancer therapy as well as expression of EMT and CSC genes.MethodsThe viability and proliferation capacity of HNSCC cells as well as induction of apoptosis in tumor spheroids cells after treatment was assessed by MTT assay, crystal violet- and TUNEL assay respectively. Expression of EMT and CSC markers was analyzed on mRNA (RT-qPCR) and protein (Western blot) level.ResultsWe showed that HNSCC cells from different tumors formed spheroids that differed in size and density in regard to EMT-associated protein expression and culturing time. In all spheroids, an up regulation of CDH1, NANOG and SOX2 was observed in comparison to 2D but changes in the expression of EGFR and EMT markers varied among the cell lines. Moreover, most HNSCC cells grown in 3D showed decreased sensitivity to cisplatin and cetuximab (anti-EGFR) treatment.ConclusionsTaken together, our study points at notable differences between these two cellular systems in terms of EMT-associated gene expression profile and drug response. As the 3D cell cultures imitate the in vivo behaviour of neoplastic cells within the tumor, our study suggest that 3D culture model is superior to 2D monolayers in the search for new therapeutic targets.Electronic supplementary materialThe online version of this article (10.1186/s12935-019-0733-1) contains supplementary material, which is available to authorized users.

Modeling the Efficacy of Oncolytic Adenoviruses In Vitro and In Vivo: Current and Future Perspectives

“…Further examination of 2D cultures suggests different patterns of gene expression emerge compared to in vivo and 3D culture experiments. In a study examining head and neck cancer cell lines, cancer stem cell (CSC) genes (NANOG and SOX2) were down regulated in 2D culture compared to 3D models [20]. Similar results were found in bladder cancer with T24 cell spheres expressing higher levels of mRNA and protein levels of NANOG compared to 2D culture [21].…”

Petromurin C Induces Protective Autophagy and Apoptosis in FLT3-ITD-Positive AML: Synergy with Gilteritinib

“…Tumor cells are known to be more resistant to drugs in 3D than in 2D cultures due to reduced access to compounds and pathophysiological differences such as hypoxia, which leads to activation of cell survival genes [29]. However, some studies showed that cells can also be more sensitive in 3D cultures compared to 2D cultures [30,31]. Melissaridou et al showed head and neck squamous cell carcinoma cell line LK0902 was more sensitive to cetuximab, which is an epidermal growth factor receptor (EGFR) inhibitor in 3D conditions compared to 2D [30].…”

Multicellular tumor spheroids- an effectivein vitromodel for understanding drug resistance in head and neck cancer

“…The schematic representation of the in vitro experimental workflow used for determining the MDR profile is provided below (Scheme 1). The cell lines LK0912, LK0917, and LK1108 used in our experiments, were established from three different HNSCC patients as reported previously by our team 25,26 . As reported previously 25,26 , biopsies were excised from the tumors of gingiva, tongue, and hypopharynx and harvested immediately for establishing monolayer cell lines in vitro.…”

“…In addition, owing to their multicellular nature, MCSs spontaneously develop MDR against many chemotherapeutic drugs 23,24 , thus making them the appropriate model system for the purpose of the present study. Recently, members of our group reported a marked treatment response difference between 2D cell cultures and MCSs of head and neck cancer cells pertaining to epithelial-mesenchymal transition and stem cell characteristics, suggesting that 3D cell cultures are clinically relevant models that are superior to 2D monolayers for the investigation of new therapeutic targets 25 . However, there is no well-defined in vitro method or criteria for the identification of the resistance status of cancer cells.…”

“…Our strategy combines drug screening, real time, time-lapse fluorescence microscopy, and flow cytometry for rapid identification of drug resistance status using MCSs, so that a beneficial personalized treatment regimen can be offered to patients. The cell lines used were previously established by the members of our group 25,26 . Briefly, we have investigated the drug response profiles of LK0917 (gingiva), LK0902 (tongue), and LK1108 (hypopharynx) cells to doxorubicin, cisplatin, and methotrexate in 2D and MCSs.…”