Organoids of patient‐derived medullary thyroid carcinoma: The first milestone towards a new in vitro model in dogs

Abstract

Organoid cultures could constitute a valuable in vitro model to explore new treatments for canine (c) medullary thyroid carcinoma (MTC). The study's objectives were to establish and characterize 3D organoid cultures of cMTC using histology and immunohistochemistry (IHC) and to evaluate the effect of antitumor drugs on organoids' viability.Five cMTC tissue samples were used to develop organoid cultures of which one organoid line, named cMTC N 2, could be passaged for an extended period. This cMTC N 2 organoid line was further compared to the primary tumour regarding morphology and IHC expression of thyroid transcription factor-1 (TTF-1), thyroglobulin, calcitonin, synaptophysin, vimentin, Ki-67, cyclooxygenase-2 (COX-2), P-glycoprotein and vascular endothelial growth factor (VEGF). Quality control of the cMTC N 2 organoid line was achieved by a single nucleotide polymorphism (SNP) array of the organoids, primary tumour and healthy blood cells of the same dog. The effect of carboplatin, meloxicam and toceranib phosphate (TOC) on cMTC N 2 organoids' viability was evaluated. The cMTC N 2 organoid line was cultured for 94 days and showed similar histological features with the primary tumour. Immunolabelling for TTF-1, thyroglobulin, calcitonin and VEGF was similar between the primary tumour and cMTC N 2 organoids. Compared to the primary tumour, organoids showed higher immunolabelling for vimentin and Ki-67, and lower immunolabelling for synaptophysin, COX-2 and P-glycoprotein. The SNP genotype was similar for each chromosome between healthy blood cells, primary tumour and cMTC N 2 organoids. Carboplatin, meloxicam and TOC had no effect on cMTC N 2 organoid cell viability within achievable in vivo concentration range. In conclusion, the cMTC N 2 organoid line is a promising first milestone towards an established in vitro organoid model to explore pathophysiology and new treatment modalities in cMTC.

Advances in organoid technology for veterinary disease modeling

“…Organoids derived from canine follicular cell thyroid carcinoma highly expressed thyrotropin receptor, sodium iodide symporter, pendrin, and thyroid peroxidase, the expressions of which are similar or higher compared to the primary tumors ( 45 ). Organoids derived from canine medullary thyroid carcinoma ( 46 ) showed similar histological features with the primary tumor after a long-term culture. Canine lung cancer organoids recapitulated the tissue architecture of canine lung adenocarcinoma, and expressed TTF1, a lung adenocarcinoma marker ( 48 ).…”

“…Organoid models of animal cancer, which might be derived from a small number of stem cells, enabled a better understanding of the molecular characteristics and neoplastic behavior of the disease in animals and acted as preliminary models for drug screening ( 78 – 81 ). For example, canine medullary thyroid carcinoma, studies on its biological behavior are scarce but treatment outcomes are always disappointing, so the development of a canine organoid model should help elucidate the best therapeutic approaches ( 46 ). Organoids could be formed from limited numbers of cells such as urine-derived stem cells, which are successfully used in the study of organoids of canine prostatic and urinary tract cancer ( 40 – 43 ).…”

“…Due to better preservation of tumor microenvironment than conventional 2D cell line models, organoids of canine prostate cancer ( 40 ), canine bladder cancer ( 41 – 43 ), canine follicular cell and medullary thyroid carcinoma ( 45 , 46 ), canine lung adenocarcinoma ( 48 ), and canine mammary tumor ( 47 ), accurately retain original morphological characteristics, genomic structures, and/or mutational profiles and recapitulate the genetic and phenotypic heterogeneity of the tumor cells. Organoids of canine prostate cancer ( 40 ) expressed epithelial markers (E-cadherin, CK5 and CK8), a myofibroblast marker (a-SMA) and a cancer stem cell marker (CD44).…”