Liquid Biopsy-Based RET Mutation Profiling to Guide RET Inhibitor Treatment in Sporadic Medullary Thyroid Carcinoma May Be Useful in Cases with High Tumor Burden and Progressive Disease

Background: One of the challenges in tumor molecular profiling for therapeutic decisions is the unavailability of tumor tissue or its inadequacy to provide high-quality nucleic acids. Although tissue biopsy remains the "gold-standard," analysis of circulating tumor DNA (ctDNA) may offer an alternative to characterize mutations necessary to initiate systemic therapy with selective inhibitors in eligible patients. This study aimed to identify cases of sporadic medullary thyroid carcinoma (sMTC) in which plasma ctDNA analysis may be useful for RET gene testing when tumor tissue is unavailable. Methods: We conducted a retrospective cohort study analyzing plasma from 36 patients affected by RET-mutated sMTC. Patients were divided into three cohorts (1) 18 patients with progressive sMTC; (2) nine patients with stable disease under treatment with a multikinase inhibitor; and (3) nine patients with metastatic but stable disease without treatment. For patients in cohort 1, we studied plasma collected at the time of progression just before the initiation of systemic therapy, while in cohorts 2 and 3, we studied last plasma available at follow-up. The RET driver mutation was analyzed in ctDNA using specific digital droplet PCR assays. Results: The RET driver mutation was detected in 16/36 (44.4%, CI 27.9-61.9) ctDNA samples, with a statistically significant difference among the three cohorts 16/18 (88.9%, CI 65.3-98.6) in cohort 1 and 0/9 (0%, CI 0-33.6) in cohorts 2 and 3 (p < 0.001) with a mean variant allele frequency of 5.4%. The presence of detectable RET-mutated ctDNA was associated with disease progression (p < 0.001), higher percentage of metastatic sites with > five lesions (i.e., tumor burden; p = 0.001), and higher levels of serum calcitonin (Ct) (p = 0.009), and carcinoembryonic antigen (p = 0.038). Conclusions: Our study demonstrates that ctDNA analysis may be a valid approach to genotype sMTC patients. Thus, liquid biopsy-based RET mutation profiling may be beneficial in advanced and progressive sMTC cases when primary tumor tissue is unavailable or inadequate for high-quality nucleic acid extraction, enabling RET-inhibitor therapy, if needed, according to specific indications. However, to obtain reliable results, ctDNA molecular profiling should be performed when tumor burden is high and the disease is progressing.