Long-term culture of patient-derived pheochromocytoma organoids.

Introduction: Pheochromocytomas (PCCs) are rare neuroendocrine tumors with limited treatment options once metastasized. Progress toward effective therapies has been hindered by their rarity, disease heterogeneity, and lack of representative preclinical models. Organoids are three-dimensional, self-renewing structures that recapitulate key features of their tissue of origin, providing valuable platforms for disease modeling, drug screening, and personalized medicine. This study aimed to establish and characterize patient-derived organoid cultures of PCCs. Methods: Tumor tissue from two patients undergoing surgery for PCC was collected to generate organoid cultures. Organoids were expanded under growth factor-enriched conditions and characterized by histology, immunohistochemistry, immunofluorescence, gene expression via qPCR, and metanephrine quantification via LC-MS/MS. Results: Both cultures could be maintained for multiple weeks and serially passaged, with PCC1 maintained in culture for 129 days and expanded up to passage 3 (passaged every 34-53 days), and PCC2 cultured for 139 days, reaching passage 2 (passaged every 63 days). At the protein level, organoids expressed both stem/progenitor markers (nestin, vimentin, SOX10) and chromaffin differentiation markers (synaptophysin, CD56), although expression of the latter was markedly reduced compared to primary tumor tissue. Metanephrine production confirmed initial secretory activity, but hormone production declined over time. Conclusions: This pilot study demonstrates that patient-derived organoid cultures can be established from human PCCs and maintained with serial passaging. In early stages, these organoids maintain key phenotypic and functional traits of their original tumors; however, these features diminish over time, consistent with a progressive loss of chromaffin differentiation and/or expansion of less differentiated cell populations. Further optimization is required to improve their long-term proliferation and differentiation potential. Together, these findings provide a proof-of-concept for the development of patient-derived PCC organoid models, which may ultimately serve as a platform for studying PCC biology and for future exploration of personalized therapeutic approaches.