EBV is preserved in NPC268 and can be induced into lytic infection. A, EBER staining was positive in early and late passage of NPC268, but LMP1 and BZLF1 protein expressions were lost in the late passage. Scale bar: 100 µm. B, EBV latent and lytic gene expression profiles in NPC268 in comparison with other EBV-positive NPC cell lines. C, EBV CN in NPC268 has reduced following prolonged culture but stabilized at later passages, retaining about approximately 13 copies of EBV per cell. D, EBV-FISH assay confirming presence of EBV-positive cells in late passage of NPC268. E, EBV lytic genes-encoded protein expression in NPC268 induced by different methods. EBV-negative Akata cells infected with EBV derived from NPC268 expresses Zta and Ea-D, confirming infectious EBV production. NPC43 treated with TPA for 48 hours was used as positive control. F, qPCR analysis of the EBV-negative Akata cells showed substantial upregulation of various latent and lytic genes postinfection with the NPC268-derived EBV. Data are shown as mean ± SD (n = 2 biological repeats).
ARTICLE ABSTRACT
Nasopharyngeal carcinoma (NPC), a cancer that is etiologically associated with the Epstein-Barr virus (EBV), is endemic in Southern China and Southeast Asia. The scarcity of representative NPC cell lines owing to the frequent loss of EBV episomes following prolonged propagation and compromised authenticity of previous models underscores the critical need for new EBV-positive NPC models. Herein, we describe the establishment of a new EBV-positive NPC cell line, designated NPC268 from a primary non-keratinizing, differentiated NPC tissue. NPC268 can undergo productive lytic reactivation of EBV and is highly tumorigenic in immunodeficient mice. Whole-genome sequencing revealed close similarities with the tissue of origin, including large chromosomal rearrangements, while whole-genome bisulfite sequencing and RNA sequencing demonstrated a hypomethylated genome and enrichment in immune-related pathways, respectively. Drug screening of NPC268 together with six other NPC cell lines using 339 compounds, representing the largest high-throughput drug testing in NPC, revealed biomarkers associated with specific drug classes. NPC268 represents the first and only available EBV-positive non-keratinizing differentiated NPC model, and extensive genomic, methylomic, transcriptomic, and drug response data should facilitate research in EBV and NPC, where current models are limited.
NPC268 is the first and only EBV-positive cell line derived from a primary non-keratinizing, differentiated nasopharyngeal carcinoma, an understudied but important subtype in Southeast Asian countries. This model adds to the limited number of authentic EBV-positive lines globally that will facilitate mechanistic studies and drug development for NPC.
