**Results and Clinical Interpretation of Germline** *RET* **Analysis in a Series of Patients with Medullary Thyroid Carcinoma: The Challenge of the Variants of Uncertain Significance**

**Giovanni Innella 1,2 , Cesare Rossi 1, Maria Romagnoli 1, Andrea Repaci 3, Davide Bianchi 4, Maria Elena Cantarini 5, Davide Martorana <sup>6</sup> , Lea Godino <sup>1</sup> , Andrea Pession 2,5, Antonio Percesepe <sup>6</sup> , Uberto Pagotto 2,3 and Daniela Turchetti 1,2,\***


Received: 2 October 2020; Accepted: 3 November 2020; Published: 5 November 2020

**Simple Summary:** About 25% of Thyroid Carcinomas of Medullary type occur in carriers of hereditary alterations in the *RET* gene. Different alterations are associated with different risks (highest, high, and moderate) and management depends on risk category. We explored prevalence, clinical presentation and management of inherited *RET* variants in patients tested at our center. We found inherited *RET* variants in 31.9% of tested individuals: the vast majority of patients with Medullary Thyroid Carcinoma who had a family history positive for the disease was found to carry a *RET* change, but also 14.3% of those with no family history tested positive, supporting the recommendation to perform genetic testing in all cases of Medullary Thyroid Carcinoma. For known variants, findings in our patients were consistent with available risk classification. Besides, we obtained evidence supporting the classification of two rare variants of uncertain clinical significance (p.Ser904Phe and p.Asp631\_Leu633delinsGlu), which may help future management of carriers.

**Abstract:** Germline *RET* variants are responsible for approximately 25% of medullary thyroid carcinoma (MTC) cases. Identification of *RET* variant carriers allows for the adoption of preventative measures which are dependent on the risk associated with the specific alteration. From 2002 to 2020, at our cancer genetics clinic, *RET* genetic testing was performed in 163 subjects (102 complete gene analyses and 61 targeted analyses), 72 of whom presented with MTC. A germline *RET* variant was identified in 31.9% of patients affected by MTC (93.8% of those having positive family history and 14.3% of clinically sporadic cases). Subsequent target testing in relatives allowed us to identify 22 asymptomatic carriers, who could undertake appropriate screening. Overall, patients with germline *RET* variants differed significantly from those who tested negative by family history (*p* < 0.001) and mean age at MTC diagnosis (44.45 vs. 56.42 years; *p* = 0.010), but the difference was not significant when only carriers of moderate risk variants were considered (51.78 vs. 56.42 years; *p* = 0.281). Out of

12 different variants detected in 49 patients, five (41.7%) were of uncertain significance (VUS). For two of these, p.Ser904Phe and p.Asp631\_Leu633delinsGlu, co-segregation and genotype/phenotype analysis, matched with data from the literature, provided evidence supporting their classification in the moderate and the highest/high risk class (with a MEN2B phenotype), respectively.

**Keywords:** *RET*; medullary thyroid carcinoma; clinical management; variants of uncertain significance
