Background: Medullary thyroid carcinoma (MTC) is a neuroendocrine tumor derived from the parafollicular C-cells of the thyroid gland. Mutations in the gene encoding the Rearranged during Transfection (RET) tyrosine kinase, play a vital role in the development of MTCs. In patients with distant metastases, only targeted therapy can prolong survival. Tyrosine kinase inhibitors (TKIs) block tyrosine kinases such as RET and thereby inhibit tumor proliferation. However, it is difficult to determine the best treatment option for each individual patient. This study therefore aims to set up an MTC organoid model to study its potential for patient-tailored drug and PET tracer screening.
Methods: Cells were isolated from surgically obtained MTC biopsies, suspended in Matrigel, and plated in tissue culture plates with defined growth medium. For self-renewal potential assessment of putative MTC stem cells, the MTC-organoids (MTOs) were dissociated and replated every three weeks (passaging). To check origin, MTC-specific proteins were characterized by immunofluorescent (IF) staining. To determine the functionality of the MTOs, hormone concentrations were measured in the medium. Moreover, response to various TKIs was investigated by measuring hormone concentrations in the medium. Lastly, we evaluated whether the MTOs could bind various PET tracers (18F-FDG, 18F-DOPA and 18F-PSMA), to validate their potential for PET tracer screening and development.
Results: Nine MTC biopsies were cultured to MTOs. We found a maximum organoid forming efficiency of 6.3% in passage 1 (p1), 5.9% in p2, and 9.4% in p3 indicating self-renewal potential of MTC cells. IF staining showed expression of four MTC-specific markers: Calcitonin Related Polypeptide Alpha (CALCA), Carcinoembryonic Antigen-related Cell Adhesion Molecule (CEACAM), Thyroid Transcription Factor 1 (TTF1), and Somatostatin (SST), in both tissue and MTOs. Secretory hormones, calcitonin and CEA, were observed in the medium indicating functionality of the MTOs. MTOs from four patients - two with Cys634Arg, one with Met918Thr and one without RET mutation - were exposed to TKIs (vandetanib, cabozantinib, selpercatinib or pralsetinib) during 72 hours. Concentrations of calcitonin and CEA after TKI exposure were only reduced in patients with the Cys634Arg mutation. Exposure to PET tracers showed cell-specific binding to 18F-FDG and 18F-DOPA but not 18F-PSMA, indicating increased glucose metabolism and a neuroendocrine origin, but no PSMA target receptor, respectively and thus MTC-specific uptake.
Conclusion: MTC organoids can be successfully cultured, maintained and expanded, and show MTC-specific functionality. Moreover, the variable response to TKIs and specific binding of clinically used PET tracers may indicate the potential use of MTOs in prediction models. This however warrants further studies.