Establishment of a patient-derived organoid model and living biobank for nasopharyngeal carcinoma.

Background: Recurrent nasopharyngeal carcinoma (NPC) remains a major challenge for clinicians and scientists. Tumor organoid is a revelational disease model that highly resembled the heterogeneity and histopathological characteristics of original tumors. This study aimed to optimize the modeling process of patient-derived NPC organoids (NPCOs), and establish a living-biobank of NPCs to study the mechanism and explore the more effective treatment of the disease. Methods: Sixty-two fresh NPC tissue samples and 15 normal mucosa samples were collected for 3-dimensional (3D) organoid culture. The organoids were confirmed using hematoxylin and eosin assays. The expression levels of CD133, CD44, BMI-1, and Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) were detected by immunohistochemistry (IHC) and in situ hybridization (ISH). Recurrent NPCOs were frozen in liquid nitrogen for 6 months and then resuscitated and passaged. Results: We identified a novel two-step enzymatic strategy for the treatment of NPC and nasal mucosa specimens and an optimal medium for culturing NPCOs and nasal mucosa organoids (NMOs). Organoid cultures were generated from 34 primary NPC samples, 28 recurrent NPC samples, and 15 normal mucosa samples. The success rates for primary NPCO, recurrent NPCO, and NMO formation were 47.06%, 81.25%, and 86.5%, respectively. All the NPCOs were EBER positive and CK7 negative. Recurrent NPCOs had higher expressions of stem cell markers, including BMI-1, CD44, and CD133. Additionally, recurrent NPCOs could be cultured to passage 4 and frozen and revived repeatedly, while primary NPCOs were challenging to culture. Conclusions: In summary, we successfully established a living biobank using the NPCO model, which has enormous potential in basic and clinical research on NPC.