Next-Generation Sequencing in Differentiated Thyroid Cancer Patients Treated with Lenvatinib: Results and Challenges in Real-Life Practice

Objective: Our objectives were to describe molecular profiling in a real-life cohort of patients with radioiodine-resistant (RAI-R) differentiated or poorly differentiated thyroid cancer (DTC or PDTC) treated with lenvatinib and to focus on factors potentially influencing the quality of tissue samples for molecular analysis, including the impact of storage time, defined as the interval between tissue collection and molecular testing. Design: We retrospectively included all lenvatinib-treated RAI-R DTC or PDTC patients tested with DNA- and/or RNA-based next-generation sequencing (NGS) in our center, also analyzing the results of fluorescence in situ hybridization (FISH) for RET fusions if the sample did not satisfy quality criteria for RNA-based NGS analysis. We investigated differences in terms of histotype, biopsy site, or storage time between adequate and inadequate samples for RNA-based NGS. Results: At least one gene alteration was detected in 50% of the cohort (18 out of 36 patients); RAS and BRAF were the most frequent mutations, while gene fusions accounted for 5.6% of cases. Tissue samples were more frequently adequate for DNA-based NGS compared to RNA-NGS analysis (93.9% vs. 58.3%, p < 0.001). The median storage time was significantly longer in the case of inadequate samples for RNA-based NGS compared with adequate specimens (41.5 vs. 9.5 months, p = 0.016); samples archived for ≥3 years led more frequently to an inadequate result. Conclusions: Advanced RAI-R TC candidates for systemic therapy often harbor gene alterations. An adequate result was less frequently achieved in cases of RNA-based NGS than in DNA-based NGS, especially if the interval between tissue collection and molecular analysis was longer; nevertheless, the limited cohort size precludes definitive conclusions.