LAUSR

Thyroid cancer (CT) is the most common endocrine organs malignancy and is classified into:well-differentiated carcinomas, characterized by favourable prognosis and sensitive to standardtreatments, and undifferentiated carcinomas (UTC), the most lethal and highly refractoryagainst current therapies. UTCs have invasive and highly metastatic phenotype. The metastaticcascade model described distant metastasis origin deriving from cancer cells that seed to lymphnode and then disseminate to distant organs. According to this, lymphadenectomy was definedas an elective therapy for TC patients. However, recent TC trials discredit the improvement inpatient outcome. We aim to unveil the mechanism underlying tumor promotion, primary tumorclonal selection and TC metastatic origin determining which specific subclone is able to colonizedistant organs. Material and method:Thyroid carcinogenesis model was previously generated starting from human embryonic stemcells which underwent to a differentiation protocol. At day 22 of differentiation, thyroidprogenitor cells (TPCs) were engineered using CRISPR CAS-9 technology to introduce mutationin BRAFV600E/TP53R248Q and NRASQ61R/TP53R248Q gene in order to recapitulate, wheninjected in vivo, UTC. Double mutated TPCs were transduced with Barcode-mediated clonaltracking technology and subcutaneously/orthotopically injected in immunocompromisedmice. Results:We trasduced D22 double-mutated TPCs using highly complex lentiviral barcode library to mapclonal architecture in serial subcutaneous xenograft and multiple metastatic sites.Immunohistochemistry analysis obtained from P1 and P3 engraftment revealed conservation ofanaplastic thyroid cancer histologic features. Moreover, tumor-derived cells were in vitrocharacterized showing that P3 retains high migratory, invasion capacity and high MMP-9expression level, compared to P1. Transcriptomic analysis of P3 versus P1 derived cells showalso a critical metabolic switch. Cell derived from P3 engraftment were used to generateorthotopic model using Barcode mediated-clonal tracking technology. Conclusion:Given its impact on patient care, understanding the evolutionary history of anaplastic thyroidcarcinoma is critical in cancer treatment. Phylogenetic studies on the primary and metastatictumor should determine the emergence of one or more subclones, which have a crucial roleduring all the phases of tumor promotion, progression and metastatic onset. A phylogeneticanalysis performed via the barcode-mediated clonal tracking technology, will allow us to trackprimary tumor cellular clones selection and the arising progenity, determining their role in themetastatic cascade. NGS Analysis and RNA-Seq technology will define a specific primary tumor vs metastatic signature. The identification of cellular clones enriched in the metastasis willprovide new prognostic markers for metastatic thyroid carcinoma. Citation Format: Vincenzo Davide Pantina. Intratumor heterogeneity in thyroid cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5924.