89P - Epstein-Barr virus-positive and Epstein-Barr virus-negative nasopharyngeal carcinoma in multicellular spheroid model

Background

Nasopharyngeal carcinoma (NPC) is an epithelial malignancy of the nasopharynx that raises public health concern in its endemic region due to its late diagnosis. The preclinical study of NPC and the bench-to-bed transition is challenged by the lack of appropriate NPC models. Conventional two-dimensional (2D) cell culture models fail to accurately replicate the complex three-dimensional (3D) structure and microenvironment of solid tumour while available 3D spheroid models for NPC do not take into account the interaction between different cell types.

Methods

In this study, we aimed to generate 3D multicellular tumour spheroids (MCTSs) to mimic the in vivo tumour architecture and microenvironment. Homotypic MCTSs were generated using Epstein-Barr virus (EBV)-positive (C666-1, NPC43) and EBV-negative NPC (HK-1) cell lines, respectively, while heterotypic MCTSs involve co-culturing EBV-positive and EBV-negative cells with stromal cells including fibroblasts (MRC-5) and HUVEC). Spheroids were constructed using the scaffold-based technique, and morphological examination was performed using light microscopy and fluorescence microscopy to characterize the spheroid size, shape and cellular organization.

Results

Homotypic and heterotypic MCTSs generated from EBV-positive and EBV-negative cell lines form spherical aggregates. H&E staining of the MCTSs demonstrated in vivo tumour characteristics, including 3D organisation, scanted cytoplasm, irregular and enlarged nuclei, and necrotic regions. EBV+ cell lines generally formed homotypic MCTSs with more compact and spherical morphology, while homotypic MCTSs generated from EBV- cell lines are less compact and displayed alveolar-like shape. The presence of NPC cells, fibroblasts, and endothelial cells in the heterotypic MCTSs were confirmed using immunofluorescence staining.

Conclusions

Homotypic and heterotypic MCTSs were generated from both EBV- and EBV+ NPC cells. The MCTSs generated replicates the in vivo tumour features in vitro. This model would serve as a platform for the study of intercellular interactions and evaluation of potential therapeutic responses.

Editorial acknowledgement

Clinical trial identification

Legal entity responsible for the study

The authors.

Funding

Malaysia Ministry of Higher Education.

Disclosure

All authors have declared no conflicts of interest.