Purpose <p>Locally advanced thyroid cancer (LATC) presents significant surgical challenges, with a high risk of incomplete resection and poor prognosis. Patient-derived organoids (PDOs) are a powerful tool to assess drug sensitivity at an individual level and to suggest new treatment options or re-challenge. This study aimed to evaluate the method’s feasibility and efficacy as applied to patients with LATC.</p> Methods <p>In this single arm, phase 2 study, we enrolled 75 patients with LATC. Biopsies from the primary tumor or metastatic site were cultured using organoid models. Sensitivity testing was performed by using PDOs with a panel of drugs with proven activity in phase II or III trials. At the discretion of the investigator considering toxicity, the drug with the highest relative activity was offered. The primary endpoint was the objective response rate (ORR).</p> Results <p>Fifty-five patients received at least one dose of recommended drug and the primary endpoint, objective response was met in 18 patients with an overall ORR as 32.7% (95% CI 20.7–46.7). Based on the pre-defined subgroups of different histological subtypes, the ORR for patients with differentiated thyroid cancer, medullary thyroid cancer, anaplastic thyroid cancer were 32.6%, (95% CI 19.1–48.5), 16.7% (95% CI 0.4–64.1) and 50% (95% CI 11.8–88.2), respectively. The R0/R1 resection rate was 34.5% (19/55).</p> Conclusions <p>This study is the first to validate the feasibility of PDOs and in vitro sensitivity testing for LATC. PDO-based neoadjuvant therapy holds promise in improving prognosis and providing surgical opportunities for these patients.</p> Trial registration <p>The study was registered at ClinicalTrials.gov (NCT06482086) on 06/25/2024.</p> Graphical Abstract <p></p>