Phylogenetic analysis of thyroid cancer metastatic cell clones via barcode technology

Anaplastic thyroid cancer (ATC), the most aggressive form of thyroid carcinoma, is characterized by a highly metastatic phenotype, poor prognosis, and low survival rates. The mechanisms underlying tumour progression and metastasis remain poorly understood. For this reason, there are limited treatment strategies for ATC metastasis, and therapeutic strategies that are effective for primary tumours may not be equally effective for metastatic tumours. In this study, we investigated the complex biological and molecular mechanisms driving thyroid cancer tumourigenesis, progression and metastasis, focusing on the evolutionary forces that underlie these processes.Here, we developed an innovative thyroid tumourigenesis model starting from human embryonic stem cells upon CRISPR-based genetic editing. We identified that only thyroid cancer progenitor cells, obtained at day 22 of differentiation protocol toward thyroid lineage, were able to generate tumours. Specifically, we identified BRAF and NRAS mutations as drivers of papillary and follicular thyroid cancers, respectively, and with the co-occurrence of TP53 mutations leading to the aggressive phenotype of ATC. This model was then used to investigate the dynamics of thyroid cancer cellular clones and the molecular mechanisms underlying thyroid cancer promotion and progression. Barcode-mediated clonal tracking revealed the selection and enrichment of a small subpopulation of aggressive cellular clones that demonstrated increased migratory and invasive abilities, as well as upregulation of genes associated with epithelial-mesenchymal transition during TC progression. We decided to investigate the adaptive mechanism governing thyroid cancer progression and we characterized a CD44v6/Gal-3 feedback loop crucial for promoting a dormant state in metastatic TC cells, influencing stemness, EMT, and the expression of dormancy-associated genes. Finally, we investigated the metabolic reprogramming of metastatic ATC cells, highlighting the hyperactivation of oxidative phosphorylation and cholesterol metabolism as potential drivers of the metastatic phenotype. These findings offer significant insights into the complex mechanisms governing TC progression and metastasis, paving the way for targeted therapeutic strategies. Moreover, our results suggest that while based on Darwinian selection model, TC cells also undergo adaptive changes to overcome selective pressures during progression.