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Context: The clinical response after surgery is a determinant in the management of patients with medullary thyroid carcinoma (MTC). In case of excellent or structural incomplete response, the follow-up strategies are well designed. Conversely, in case of biochemical incomplete response (BiR) the management is not clearly defined., Objective: This work aimed to evaluate the overall and per-site prevalence of structural disease detection in sporadic MTC patients with BiR and to assess the predictive value of various clinical, biochemical, and genetic features., Methods: We evaluated data of 599 consecutive patients surgically treated for sporadic MTC (2000-2018) and followed-up at the endocrine unit of the University Hospital of Pisa., Results: After a median of 5 months from surgery, 145 of 599 (24.2%) patients were classified as BiR. Structural disease was detected in 64 of 145 (44.1%), after a median time of 3.3 years. In 73.6%, structural disease was detected at a single site, prevalently cervical lymph nodes. Among several others, at the time of first evaluation after surgery, only basal calcitonin (bCTN) and stage IVa/b were independent predictive factors. Also, structural disease was more frequent in patients with shorter CTN doubling time and somatic RET mutation., Conclusion: In sporadic MTC patients with BiR, the risk of detection of structural disease was about 50% at 10 years. Higher bCTN levels and staging predicted the risk of detecting structural disease. According to these findings, stricter follow-up should be reserved for MTC with BiR and elevated values of bCTN and to those with an advanced stage. Long follow-up should be considered for all BiR patients since 50% of them develop structural disease within 10 years., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Context: Serum thyroglobulin (Tg) is a highly sensitive and specific tumor marker, employed in post-operative management of patients with differentiated thyroid carcinomas. Tumor shrinkage of radioiodine-refractory thyroid cancer (RAIR-DTC) treated with multitarget kinase inhibitors as lenvatinib, expressed according to the Response Evaluation Criteria in Solid Tumors (RECIST), is also associated with a drastic reduction of Tg levels. However, interference caused by circulating thyroglobulin autoantibodies (TgAb) represents the main limitation in the clinical use of Tg., Objective: To evaluate if in RAIR-DTC TgAb could be considered a surrogate marker of Tg in monitoring response to treatment with lenvatinib., Design: We retrospectively evaluated patients who had started lenvatinib and correlated serum Tg and TgAb with the radiological response across visits., Setting: University of Pisa, Italy., Patients: We selected 9/97 RAIR-DTC patients with detectable TgAb., Intervention: None., Main Outcome Measures: None., Results: Tg values correlated neither with TgAb title nor with radiological response across visits. Greater decreases in TgAb titer correlated with favorable radiological response to lenvatinib after 1 month (Spearman's correlation = 0.74, P = .021) and 6 months (correlation = 0.61, P = .079). According to RECIST, patients with partial response showed a ∼10-fold greater decrease in TgAb compared to those with stable disease at 1 month (median TgAb decrease: -142 vs -14 IU/mL, P = .01) and those with progressive disease at 6 months (median TgAb decrease: -264 vs-24 IU/mL, P = .04)., Conclusion: TgAb evaluation may represent a reliable surrogate marker for Tg trend in evaluating response of RAIR-DTC to treatment with lenvatinib. A multicentric study would be useful to confirm our results., (Published by Oxford University Press on behalf of the Endocrine Society 2023.)

Currently, the differentiated thyroid cancer (DTC) management is shifted toward a tailored approach based on the estimated risks of recurrence and disease-specific mortality. While the current recommendations on the management of metastatic and progressive DTC are clear and unambiguous, the management of slowly progressive or indeterminate disease varies according to different centers and different physicians. In this context, active surveillance (AS) becomes the main tool for clinicians, allowing them to plan a personalized therapeutic strategy, based on the risk of an unfavorable prognosis, and to avoid unnecessary treatment. This review analyzes the main possible scenarios in treated DTC patients who could take advantage of AS., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Campopiano, Ghirri, Prete, Lorusso, Puleo, Cappagli, Agate, Bottici, Brogioni, Gambale, Minaldi, Matrone, Elisei and Molinaro.)

Objective: Selpercatinib is a highly selective RET-inhibitor drug, approved for the treatment of RET-altered lung and thyroid cancers. So far, RET-altered medullary thyroid cancer (MTC) patients treated with selpercatinib showed a remarkable objective response rate and safety profile. However, new treatment emerging adverse events (TEAEs) have been recently reported. The aim of this study was to evaluate the prevalence, features, and clinical management of effusions that are one of these TEAEs., Design: Around 10 of 11 patients with advanced MTC enrolled in the LIBRETTO-201 clinical trial at Endocrinology Unit of the Pisa University Hospital were evaluated for the presence and management of effusions., Methods: We retrospectively evaluated MTC patients treated with selpercatinib. The presence of pleural, pericardial, abdominal, and/or pelvic effusions was evaluated by reviewing the computerized tomography scan performed during the study protocol and up to 24 months of observation., Results: All but one MTC patient experienced previous multikinase inhibitors treatment. Three patients already had effusions before starting selpercatinib treatment. New effusions appeared in eight of ten (80%) patients during the treatment. A chylous nature was documented in patients who underwent fluid aspiration. Whenever a dose reduction was performed, a significant positive effect was observed., Conclusions: Chylous effusions are a new TEAE of selpercatinib treatment. They can appear or worsen at any time during the treatment. For cases with asymptomatic and mild effusions, active surveillance may be appropriate and safe. In symptomatic and/or moderate/severe cases, aspiration of the fluid and a dose reduction can improve this AE, strongly supporting a cause-effect correlation with selpercatinib., Significance Statement: Effusions, particularly of chylous nature, represent emergent and quite frequent adverse events in the management of patients affected by advanced MTC on treatment with the highly selective inhibitor selpercatinib. In this study, we evaluated, in a series of MTC patients treated with selpercatinib, the prevalence of pleural, pericardial, abdominal, and/or pelvic effusions. Insights into the diagnosis and treatment of the effusions are provided as well as suggestions for clinical management.

Objectives: Liver metastases occur in 45% of patients with advanced metastatic medullary thyroid cancer (MTC). Transarterial radioembolization (TARE) has been proposed to treat liver metastases (LM), especially in neuroendocrine tumors. The aim of this study was to investigate the biochemical (calcitonin and carcino-embryonic antigen) and objective response of liver metastases from MTC to TARE., Methods: TARE is an internal radiotherapy in which microspheres loaded with β-emitting yttrium-90 (90Y) are delivered into the hepatic arteries that supply blood to LM. Eight patients with progressive multiple LM underwent TARE and were followed prospectively. They were clinically, biochemically and radiologically evaluated at 1, 4, 12 and 18 months after TARE., Results: Two patients were excluded from the analysis due to severe liver injury and death due to extrahepatic disease progression, respectively. One month after TARE, a statistically significant (P = 0.02) reduction of calcitonin was observed in all patients and remained clinically relevant during follow-up; reduction of CEA, although not significant, was found in all patients. Significant reduction of liver tumor mass was observed 1, 4 and 12 months after TARE (P = 0.007, P = 0.004, P = 0.002, respectively). After 1 month, three of six patients showed partial response (PR) and three of six stable disease (SD) according to RECIST 1.1, while five of six patients had a PR and one of six a SD according to mRECIST. The clinical response remained relevant 18 months after TARE. Excluding one patient, all others showed only a slight and transient increase in liver enzymes., Conclusions: TARE is effective in LM treatment of MTC. The absence of severe complications and the good tolerability make TARE a valid therapeutic strategy when liver LM are multiple and progressive.

Osteonecrosis of the jaw (ONJ) is a rare but very serious disease that can affect both jaws. It is defined as exposed bone in the maxillofacial region that does not heal within 8 weeks after a health care provider identification. ONJ can occur spontaneously or can be due to drugs like bisphosphonates (BPS) and anti-RANK agents, in patients with no history of external radiation therapy in the craniofacial region. Although in phase 3 trials of tyrosine kinase inhibitors (TKIs) used in thyroid cancer (TC) the ONJ was not reported among the most common side effects, several papers reported the association between ONJ and TKIs, both when they are used alone and in combination with a bisphosphonate. The appearance of an ONJ in a patient with metastatic radio-iodine refractory differentiated TC, treated with zoledronic acid and sorafenib, has put us in front of an important clinical challenge: when a ONJ occurred during TKIs treatment, it really worsens the patients' quality of life. We should consider that in the case of ONJ a TKI discontinuation becomes necessary, and this could lead to a progression of neoplastic disease. The most important aim of this review is to aware the endocrinologists/oncologists dealing with TC to pay attention to this possible side effect of BPS and TKIs, especially when they are used in association. To significantly reduced the risk of ONJ, both preventive measures before initiating not only antiresorptive therapy but also antiangiogenic agents, and regular dental examinations during the treatment should always be proposed., (© 2021. The Author(s).)

Multiple Endocrine Neoplasia 2 (MEN2) is a hereditary cancer syndrome for developing medullary thyroid cancer (MTC) due to germline mutations of RET gene. Subjects harboring a germline RET mutation without any clinical signs of MTC are defined as gene carriers (GCs), for whom guidelines propose a prophylactic thyroid surgery. We evaluate if active surveillance of GCs, pursuing early thyroid surgery, can be safely proposed and if it allows safely delaying thyroid surgery in children until adolescence/adulthood. We prospectively followed 189 GCs with moderate or high risk germline RET mutation. Surgery was planned in case of: elevated basal calcitonin (bCT) and/or stimulated CT (sCT); surgery preference of subjects (or parents, if subject less than 18 years old); other reasons for thyroid surgery. Accordingly, at RET screening, we sub-grouped GCs in subjects who promptly were submitted to thyroid surgery (Group A, n = 67) and who were not (Group B, n = 122). Group B was further sub-grouped in subjects who were submitted to surgery during their active surveillance (Group B1, n = 22) and who are still in follow-up (Group B2, n = 100). Group A subjects presented significantly more advanced age, bCT and sCT compared to Group B. Mutation RET V804M was the most common variant in both groups but it was significantly less frequent in Group A than B. Analyzing age, bCT, sCT and genetic landscape, Group B1 subjects differed from Group B2 only for sCT at last evaluation. Group A subjects presented more frequently MTC foci than Group B1. Moreover, Group A MTCs presented more aggressive features (size, T and N) than Group B1. Accordingly, at the end of follow-up, all Group B1 subjects presented clinical remission, while 6 and 12 Group A MTC patients had structural and biochemical persistent disease, respectively. Thank to active surveillance, only 13/63 subjects younger than 18 years at RET screening have been operated on during childhood and/or adolescence. In Group B1, three patients, while actively surveilled, had the possibility to reach the age of 18 (or older) and two patients the age of 15, before being submitted to thyroid surgery. In Group B2, 12 patients become older than 18 years and 17 older than 15 years. In conclusion, we demonstrated that an active surveillance pursuing an early thyroid surgery could be safely recommended in GCs. This patient-centered approach permits postponing thyroid surgery in children until their adolescence/adulthood. At the same time, we confirmed that genetic screening allows finding hidden MTC cases that otherwise would be diagnosed much later.

Background: Serum Ca19.9 positivity is a prognostic factor for mortality in patients with advanced medullary thyroid cancer (aMTC), independently from calcitonin doubling time (DT). However, it is unknown whether aMTC patients who become positive for Ca19.9 also have progressive disease (PD) according to response evaluation criteria in solid tumors (RECIST) and whether Ca19.9 DT has a role in the management of aMTC patients. The aims of this study were to evaluate whether in aMTC, when serum Ca19.9 becomes positive, PD develops, and to determine the role of Ca19.9 DT in predicting mortality and PD. Patients and Methods: Serum Ca19.9 was periodically measured in 107 aMTC patients, and the DTs were calculated. Restaging of the disease was radiologically performed in 104 of 107 patients and PD was evaluated according to RECIST. Results: At the end of follow-up, 25 of 107 patients were Ca19.9 positive and PD was identified in 30 of 104 patients. No significant association was found between Ca19.9 positivity and PD, while there was a significant association between Ca19.9 positivity and mortality ( p  < 0.0001). Ca19.9 DTs <6 months and <1 year were not associated with PD but were associated with mortality ( p  < 0.0001 and p  < 0.0001, respectively). In particular, 3 patients who had a Ca19.9 DT <6 months with no evidence of PD according to RECIST died of their disease after 6, 5, and 3 months, respectively. Conclusions: Serum Ca19.9 positivity and DTs <6 months and <1 year are prognostic factors for mortality but not for PD. Serum Ca19.9 positivity and DTs <6 months and <1 year should be considered in the decision-making process of whether to initiate systemic therapy even if there is no evidence of PD according to RECIST.

Introduction: Tyrosine kinase inhibitors represent a better treatment in patients with radioiodine-refractory differentiated thyroid cancer (RAI-R DTC). Lenvatinib is usually well-tolerated, but sometimes, it is associated with serious and even life-threatening side effects. The aim of this study was to evaluate the prevalence of and the potential risk factors for fistula and/or organ perforation in RAI-R DTC patients treated with lenvatinib., Methods: This study included data from advanced and progressive RAI-R DTC patients treated with lenvatinib from February 2011 to February 2020 who were followed up at a single center. The clinical-pathological features and the biochemical and morphological results of the patients were collected at the time of starting lenvatinib and during the follow-up., Results: Fourteen of 95 (14.7%) locally advanced or metastatic RAI-R DTC patients treated with lenvatinib developed a fistula or organ perforation. Nine of 14 (64.3%) patients had tumor infiltration of the trachea, bronchus, esophagus, pleura, or bladder. Five of 14 (35.7%) had a bowel perforation, but only 2 had preexisting diverticulosis. Evaluation of the risk factors for developing a fistula or organ perforation showed that the presence of tumor infiltration and the tumor histology (papillary and poorly differentiated vs. follicular and Hurthle thyroid cancer) were significantly correlated with the development of a fistula or organ perforation ( p = 0.003 and p = 0.02, respectively). In the subgroup of patients with tumor infiltration, we found that the papillary thyroid cancer histotype was the only potential predictor of fistula development. External beam radiation therapy (EBRT), the starting dose of lenvatinib, and the duration of treatment were not relevant for the development of fistula., Conclusions: In metastatic thyroid cancer patients treated with lenvatinib, the presence of tumor infiltration and histological type should be considered as potential risk factors for the development of fistula or organ perforation, although they do not represent an absolute contraindication. Although EBRT and the presence of diverticulosis were not significantly associated with the development of fistula and organ perforation, they should be regarded as potential additional reasons for the development of these complications. According to our findings, there is no reason to start lenvatinib at a lower daily dose when tumor infiltration is present., Competing Interests: R.E. is a consultant for EISAI; however, the present study was not conditioned by this activity. L.V., C.G., L.A., E.M., D.V., V.B., A.M., L.P., L.L., V.C., and A.R. have nothing to disclose., (Copyright © 2021 by S. Karger AG, Basel.)

Purpose: Severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) is challenging health systems all over the world. Cancer patients have a higher risk of being infected by SARS-Cov-2 and higher coronavirus disease 2019 (COVID-19) severity and mortality. Up to date, there were no data about COVID-19 in patients with thyroid cancer (TCs). The aim of the study was to describe the prevalence of COVID-19 in a well-characterized series of TC patients evaluated for the persistence of the neoplastic disease from March to September 2020; as secondary objective, we looked for the COVID-19 disease severity in a subgroup of multimetastatic TC patients., Methods: We evaluated 1464 patients affected by persistent TC: 67 patients who were taking multikinase inhibitors (MKIs) and 1397 under active surveillance for a persistent but stable disease. During the clinical evaluation, all patients were specifically investigated about a positive history of Sars-Cov-2 infection., Results: SARS-Cov-2 infection was identified in 4/1464 (0.3%) cases of patients affected by TC. We identified three cases among patients under active surveillance (0.2%), and one case among patients treated with MKI systemic therapy (1/67, 1.5%). This patient was taking vandetanib for metastatic medullary thyroid cancer (MTC), when he came to our attention referring severe fatigue, dyspnea for light physical activities. He presented a mild COVID-19 and he received exclusively supportive care. After a multidisciplinary consultation, we decided against the discontinuation of vandetanib. After 2 months from the infection, he did not present any signs of active infection, and the MTC metastatic disease was stable., Conclusions: We showed that COVID-19 is not more frequent in TC patients than in general population, although a relatively higher prevalence in the group of TC patients treated with MKIs. A single patient with advanced TC and SARS-Cov-2 infection during MKIs treatment had a mild COVID-19 and did not require the discontinuation of MKI therapy. In cases of more severe COVID-19, an accurate evaluation from a multidisciplinary team would consider risks and benefits in taking the decision to continue or stop MKI treatment.

Differentiated thyroid cancers (DTC) are commonly and successfully treated with total thyroidectomy plus/minus radioiodine therapy (RAI). Medullary thyroid cancer (MTC) is only treated with surgery but only intrathyroidal tumors are cured. The worst prognosis is for anaplastic (ATC) and poorly differentiated thyroid cancer (PDTC). Whenever a local or metastatic advanced disease is present, other treatments are required, varying from local to systemic therapies. In the last decade, the efficacy of the targeted therapies and, in particular, tyrosine kinase inhibitors (TKIs) has been demonstrated. They can prolong the disease progression-free survival and represent the most important therapeutic option for the treatment of advanced and progressive thyroid cancer. Currently, lenvatinib and sorafenib are the approved drugs for the treatment of RAI-refractory DTC and PDTC while advanced MTC can be treated with either cabozantinib or vandetanib. Dabrafenib plus trametinib is the only approved treatment by FDA for BRAF V600E mutated ATC. A new generation of TKIs, specifically for single altered oncogenes, is under evaluation in phase 2 and 3 clinical trials. The aim of this review was to provide an overview of the current and future treatments of thyroid cancer with regards to the advanced and progressive cases that require systemic therapies that are becoming more and more targeted on the molecular identity of the tumor.

Purpose: The use of tyrosine kinase inhibitors (TKIs) in thyroid cancer patients is often limited by toxicities. Some have a long-term onset and potentially could impact patients' survival. Among them, there is the nephrotoxicity, mainly represented by proteinuria. The aim of the study was to evaluate the prevalence of proteinuria in medullary thyroid cancer patients treated with cabozantinib, to examine whether it could be a marker for treatment monitoring and to evaluate histological kidney alterations., Methods: We collected data of 31 medullary thyroid cancer patients enrolled in the EXAM trial. Proteinuria was defined and evaluated using the National Cancer Institute's Common Terminology Criteria for Adverse Events. In two symptomatic cases with high-grade proteinuria, a kidney biopsy was performed., Results: Proteinuria was observed in 4/18 patients (22.2%) and occurred after a mean period of 38 months (median: 35.5 months). It was significantly associated with previous chemotherapy (p = 0.005) and/or treatment with other TKIs (p = 0.04), a prolonged use of cabozantinib (p = 0.0004), and a better radiological response at the end of follow-up (p = 0.002). The kidney biopsy showed pathognomonic features of thrombotic microangiopathy in both cases and a focal amyloid deposit in one., Conclusion: Proteinuria is a quite frequent adverse event during cabozantinib treatment. It is relatively well manageable with the early detection and correction of risk factors, the temporary discontinuation of cabozantinib and/or its dose reduction, and the use of anti-proteinuric and renoprotective drugs in patient with hypertension. The histological findings confirmed some typical features of the anti-VEGF inhibition injury, already described for other TKIs.

Sporadic medullary thyroid carcinoma (MTC) is a rare malignancy with a heterogeneous clinical course. Several potential prognostic factors have been investigated, but the impact of some of these is controversial, such as age at diagnosis. We evaluated the data of 432 sporadic MTC patients followed-up for a median of 7.4 years. Patients were divided and compared according to their age at diagnosis in group A (<65 years-n = 338, 78.2%) and group B (≥65 years-n = 94, 21.8%). No differences were detected between the two groups. Median follow-up time was significantly longer in patients <65 than ≥65 years. We observed 41 (9.5%) cancer-related death events. The death rate was similar between the two age groups. However, the Kaplan Meier curve showed a longer survival time for younger patients compared to older patients [HR 2.5 (CI 95%: 1.27-4.94), p < 0.01]. Nevertheless, no differences in the aggressiveness of the disease at presentation and in the number and type of treatments performed were found in the two subgroups of dead patients. In patients with sporadic MTC, age at diagnosis did not correlate with any clinical and pathological features. Cancer-related death events are similar in older and younger patients, but survival time is longer in the younger.

Copy number variations (CNV) of the RET gene have been described in 30% of Medullary Thyroid Cancer (MTC), but no information is available about their role in this tumor. This study was designed to clarify RET gene CNV prevalence and their potential role in MTC development. RET gene CNV were analyzed in 158 sporadic MTC cases using the ION Reporter Software (i.e., in silico analysis) while the multiplex ligation-dependent probe amplification assay (i.e., in vitro analysis) technique was performed in 78 MTC cases. We identified three categories of RET ploidy: 137 in 158 (86.7%) cases were diploid and 21 in 158 (13.3%) were aneuploid. Among the aneuploid cases, five out of 21 (23.8%) showed an allelic deletion while 16 out of 21 (76.2%) had an allelic amplification. The prevalence of amplified or deleted RET gene cases (aneuploid) was higher in RET positive tumors. Aneuploid cases also showed a higher allelic frequency of the RET driver mutation. The prevalence of patients with metastatic disease was higher in the group of aneuploid cases while the higher prevalence of disease-free patients was observed in diploid tumors. A statistically significant difference was found when comparing the ploidy status and mortality. RET gene CNVs are rare events in sporadic MTC and are associated with RET somatic mutation, suggesting that they could not be a driver mechanism of tumoral transformation per se. Finally, we found a positive correlation between RET gene CNV and a worse clinical outcome.

The long-term survival of differentiated thyroid cancer (DTC) patients and the need to perform several treatments with radioiodine (131-I) lead to the question if the lifetime risk of developing a nonthyroidal second primary cancer (NTSPC) is increased in these patients. In our study, we assessed the prevalence of NTSPCs in thyroid cancer population and evaluated the possible causative role of 131-I treatment. We analyzed 1096 consecutive patients followed at our institution from 1964 to 1998. A total of 101 NTSPCs were observed in 92/1096 patients (8.4%) among which 17/101 (16.8%) diagnosed before DTC and 84/101 (83.2%) diagnosed after. The most frequent tumor sites observed were breast and bladder/urinary tract in the post-DTC group and breast and hematological system in the pre-DTC group. Regarding 131-I treatment, we did not observe any significant differences regarding either the number of treatments or the cumulative activity. The only significant parameter associated with an increased incidence of NTSPC was follow-up (P = .02): a longer follow-up period was associated with a higher number of NTSPCs. The mean latency between 131-I and NTSPC was 10.52 ± 7.69 years. Comparing with the general Italian population, independent of radioiodine treatment, the standard incidence ratio in our cohort was similar to that of the general population (SIR 1.07) and this result was confirmed by analyzing only the treated group. In conclusion, these results show that the risk of NTSPCs in the DTC patients' population is similar to that in the general population and 131-I treatment was not associated with an increased risk., (© 2020 UICC.)

Objective: At present, recombinant TSH cannot be used for the treatment of metastatic differentiated thyroid cancer patients. The aim of this study was to evaluate if the type of TSH stimulation, recombinant or endogenous, had an impact on the outcome of these patients., Design and Methods: We compared the outcome of two propensity score-matched groups of metastatic patients, stimulated by either only recombinant TSH (n = 43) or only endogenous TSH (n = 34)., Results: As expected from the matching procedure, the clinical-pathological features and the cumulative 131-I activities administered to the two groups were very similar. After 4 years of follow-up, 4% of patients were cured, 3% had biochemical disease and 93% had structural disease. However, 91% of patients obtained a clinical benefit from this therapy in terms of stabilization of the disease or complete remission or partial response. When considering the two groups separately, we did not find any difference in their outcome. When considering the response to 131-I therapy of the single type of metastases, 8% of lymph node metastases and 8% of lung metastases disappeared but none of the bone metastases. The response to 131-I therapy of the single type of metastases was similar when we looked at the two groups separately., Conclusions: This study shows (i) an overall clinical benefit of the 131-I therapy, since the majority of patients remained affected but with a stable disease, and (ii) that the preparation with either recombinant or endogenous TSH has no impact on the 131-I therapy efficacy and the outcome of our two groups of patients.

Context: The dramatic rise in the incidence of thyroid cancer over the last 30 years is largely attributable to the increasing diagnosis of papillary microcarcinomas (mPTCs). Current guidelines endorse an observational management approach in properly selected cases., Objective: To evaluate the feasibility of active surveillance in mPTC in Italy, its impact on real life, and to identify risk factors of progression., Design and Setting: In 2014 we started a prospective-observational study of active surveillance in mPTC patients., Patients: Included patients demonstrated a single Thy4 or Thy5 thyroid nodule, with largest diameter ≤1.3 cm, and no suspicious laterocervical lymph nodes by neck ultrasonography. Of 185 eligible subjects, 50.3% (93/185) enrolled in the observational management protocol while the others opted for surgery and were excluded from this analysis., Intervention: Enrolled patients were followed with neck ultrasound at 6- to 12-month intervals. Disease progression was defined as the appearance of abnormal lymph nodes or nodule enlargement during follow-up. In these cases, patients were directed to surgery., Results: Three patients (3/93, 3%) showed clinical progression and required surgery. Another 19 patients (19/93, 20%) decided to transition to surgical intervention even though there was no evidence of disease progression. All operated patients had excellent response to initial treatment despite the delayed surgery., Conclusions: Within an Italian medical context, active surveillance appears to be a feasible and safe alternative to immediate surgery in healthy mPTC patients. Only 3% of mPTC demonstrated disease progression during a median follow-up of 19 months (range 6-54) and importantly demonstrated excellent outcomes after surgical intervention in a short-term follow-up., (© Endocrine Society 2019. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Vandetanib is an important treatment option for advanced metastatic medullary thyroid cancer. The aims of this study were to evaluate the predictors of both a longer response to vandetanib and the outcome. Medical records of 79 medullary thyroid cancer patients treated with vandetanib at our center were analysed. Twenty-five patients were treated for <12 months, 54 were treated for ≥12 months and 24 of these latter were treated for ≥48 months (short-, long- and very long-term). The median progression free survival of the long and very long-term treated patients was significantly longer than in the ZETA trial. When comparing the groups of short - and long-term treated patients the only significant difference was that these latter were less frequently previously treated with a tyrosine kinase inhibitor. However, the long-term treated patients had a younger age, both at diagnosis and enrolment, which was statistically significant in the very long-term treated patients. In the long-term treated group, younger age, enrolment for symptoms and development of adverse events were significantly correlated with a better outcome. The enrolment for symptoms remained the only statistically significant predictor of a good outcome in the very long-term treated patients. In conclusion, early treatment with vandetanib, when patients are younger, with a good ECOG performance status and symptomatic disease, not necessarily progressing for RECIST, seem to be the best predictors of a longer and durable response. Further studies are needed to confirm these results

Sporadic Medullary Thyroid Carcinoma (sMTC) is a rare but aggressive thyroid tumor. RET and RAS genes are present in about 50%-80% of cases, but most of the remaining cases are still orphan of a genetic driver. We studied the largest series of sMTC by deep sequencing to define the mutational landscape. With this methodology we greatly reduced the number of RET- or RAS-negative cases and we confirmed the central role of RET and RAS mutations. Moreover, we highlighted the bad prognostic role of RET mutations in sMTC and consolidated the favorable prognostic role of RAS mutations. For the first time, we showed that the variant allele frequency represents an additional prognostic marker inside the group of RET-mutated sMTC., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Pathogenic germline mutations affecting the RET proto-oncogene underlie the development of hereditary medullary thyroid carcinoma (MTC). The aims of this study were to evaluate the prevalence of germline RET mutations in a large series of MTC, collected over the last 25 years, and to reappraise their clinical significance., Methods: We performed RET genetic screening in 2031 Italian subjects: patients who presented with sporadic (n = 1264) or hereditary (n = 117) MTC, plus 650 relatives., Results: A RET germline mutation was found in 115/117 (98.3%) hereditary and in 78/1264 (6.2%) apparently sporadic cases: in total, 42 distinct germline variants were found. The V804M mutation was the most prevalent in our cohort, especially in cases that presented as sporadic, while mutations affecting cysteine residues were the most frequent in the group of clinically hereditary cases. All M918T mutations were "de novo" and exclusively associated with MEN2B. Several variants of unknown significance (VUS) were also found., Conclusions: a) RET genetic screening is informative in both hereditary and sporadic MTC; b) the prevalence of different mutations varies with V804M being the most frequent; c) the association genotype-phenotype is confirmed; d) by RET screening, some VUS can be found but their pathogenic role must be demonstrated before screening the family.

Background: Multiple endocrine neoplasia type 2B (MEN 2B) is a very rare syndrome characterized by a very peculiar phenotype with mucosal neuromas, marfanoid habitus, and bumpy lips associated with medullary thyroid cancer (MTC) and pheochromocytoma (PHEO). Although the syndrome was first described 50 years ago, it is still diagnosed too late, when the MTC is metastatic and frequently when the PHEO has already developed., Case Presentations: We report on two cases of MEN 2B that were diagnosed too late, preventing a cure. The cases involve two females who were 25 and 12 years old. Both were previously treated for congenital skeletal abnormalities; however, despite their bumpy lips and mucosal neuromas, MEN 2B syndrome was not recognized. When they arrived at our center for both the presence of thyroid nodules and elevated serum calcitonin values, the MTC was already metastatic, and the older patient had already developed a bilateral PHEO. After 3 years and 1 year of follow-up, the two patients are still alive but with persistent structural and biochemical disease., Discussion: These two cases show that knowledge of this syndrome is still insufficient and that the lack of knowledge impairs the ability to obtain an early diagnosis and cure. Because most patients with MEN 2B have no familial history, the only way to ensure a timely diagnosis is to recognize the MEN 2B phenotype on a clinical basis., (Copyright © 2019 Endocrine Society.)

Medullary thyroid cancer (MTC) is a rare thyroid tumor whose prevalence is 3-5% among all thyroid tumors. The pathogenesis of MTC is mainly related to germline or somatic RET activating point mutations that are causative of hereditary and sporadic cases, respectively. Hereditary MTC can occur as multiple endocrine neoplasia type 2A (MEN 2A), type 2B (MEN 2B) and familial MTC (FMTC) that differ for the association with other endocrine neoplasia. Germline RET point mutations are prevalently localized in exons 5, 8, 10-11, 13-16 and a significant genotype-phenotype correlation has been observed. RET genetic screening is mandatory in all patients with a diagnosis of MTC regardless from their apparent sporadic origin. The identification of RET germline mutation in an apparently sporadic case is of great clinical utility because it allows the identification of those subjects who will develop the tumor. RET positive relatives must undergo clinical and biochemical tests to verify if the MTC is already present and, according to the type of RET mutation, they have to be screened for the presence of pheochromocytoma and/or hyperparathyroidism. If a MTC is already present patients must be surgically treated. If MTC is not yet present subjects will be followed up with basal and stimulated calcitonin serum measurement, which is the serum marker of MTC. Indeed, RET negative subjects can be reassured that they do not run any risk to develop the disease as well as their children. In conclusions RET genetic screening allows the identification of the hereditary/sporadic nature of MTC and of a relevant percentage of hidden familial MTC. Furthermore, it favors the early diagnosis of MTC in RET positive subjects. RET positive patients and no clinical evidence of MTC can be followed and surgical treatment can be delayed. Finally RET negative relatives do not need to be further monitored., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

Background: Recently, there has been a trend to reduce the use of radioiodine remnant ablation (RRA) in patients with low-risk (LR) and intermediate-risk (IR) differentiated thyroid cancer (DTC)., Objectives: The aim of this paper was to evaluate the diagnostic role of whole-body scan (ptWBS) performed after RRA in LR and IR DTC patients., Methods: We analyzed 545 DTC patients treated with total thyroidectomy and RRA in hypothyroidism followed by a ptWBS. Neck ultrasound (US) and serum thyroglobulin measurement were performed. According to the American Thyroid Association guidelines, patients were classified as LR ( n = 345) and IR ( n = 200)., Results: In addition to the thyroid remnant, the ptWBS showed the presence of further areas of 131 I uptake in 16/545 (2.9%) cases. ptWBS showed laterocervical lymph node metastases in 11/16 patients (10/11 were also detected by US), mediastinal uptake in 1/16, lung metastases in 3/16, and bone metastases in 1/16. Only 6/545 (1.1%) metastases were detected by ptWBS alone. After 7.8 years, 8/16 patients were free of disease, and 8 had persistent disease: 4 "biochemical" and 4 "structural." Remission was achieved in 3 cases after one single 131 I course, in 1 case after surgery, and in the last 4 cases after several 131 I courses., Conclusions: The ptWBS diagnostic role was clinically relevant for the therapeutic strategies of our patients only in 1.1% of the cases. The cost-effectiveness of performing RRA and ptWBS in all LR and IR patients to find 1-2% of the cases with distant metastases remains controversial.

Introduction: Papillary thyroid cancer (PTC) is the most common endocrine malignancy. More than 98% of patients achieve an excellent response with no evidence of clinical, biochemical, or structural disease after initial treatment. In these patients structural recurrence is rare, more frequently diagnosed in the first 5 years from initial treatment and almost invariably localized in neck lymph nodes. Patient: We report the case of a woman affected by PTC who presented with rapidly rising anti-thyroglobulin antibodies (TgAb) level after 10 years from clinical, morphological and biochemical remission. Diagnosis and Treatment: In 2003, a 56 year old patient was treated with total thyroidectomy and radioiodine remnant ablation (RRA) for a PTC (2 cm) with minimal extrathyroidal extension (T3N1aM0 according to the 6th AJCC TNM staging system) associated with diffuse lymphocytic thyroiditis. In 2004 the patient was free of disease defined as undetectable Tg after recombinant human TSH administration in the absence of TgAb and structural disease. Since February 2012 the appearance and progressive increase of TgAb titer was observed and in 2014 a 18 FDG-PET scan documented three hypermetabolic lesions suggestive of lung micrometastases. The lung lesions were cytologically confirmed as PTC metastases. Both the primary tissue and the lung metastasis were positive for BRAF V600E mutation. The patient was treated with 131-radioiodine that showed radioiodine avid lung lesions that lose the ability to take up iodine at the following treatment. The patient is still alive and the lung lesions are growing slowly. Conclusions: Structural recurrence in patients that demonstrated an excellent response after initial treatment for PTC is extremely rare, and distant metastases exceptional but possible. This case is peculiar because recurrence was early identified after 10 years from initial treatment for the presence of detectable TgAb in a patient that had an histological diagnosis of lymphocytic thyroiditis but with an atypical clinical presentation (normal thyroid at neck ultrasound and undetectable TgAb and anti-thyroid peroxidase antibodies). For this reason TgAb should be tested with Tg in patients with a history of lymphocytic thyroiditis, either histological or humoral, also when TgAb is in the normal range and not suggestive of autoimmune thyroiditis.

The last ten years have been characterized by the introduction in the clinical practice of new drugs named tyrosine kinase inhibitors for the treatment of several human tumors. After the positive conclusion of two international multicentric, randomized phase III clinical trials, two of these drugs, sorafenib and lenvatinib, have been recently approved and they are now available for the treatment of advanced and progressive radioiodine refractory thyroid tumors. We have been involved in most clinical trials performed with different tyrosine kinase inhibitors in different histotypes of thyroid cancer thus acquiring a lot of experience in the management of both drugs and their adverse events. Aim of this review is to give an overview of both the rationale for the use of these inhibitors in thyroid cancer and the major results of the clinical trials. Some suggestions for the management of treated patients in the real life are also provided., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Thyroid cancer typically has a good outcome following standard treatments, which include surgery, radioactive iodine ablation for differentiated tumours and treatment with thyrotropine hormone-suppressive levothyroxine. Thyroid cancers that persist or recur following these therapies have a poorer prognosis. Cytotoxic chemotherapy or external beam radiotherapy has a low efficacy in these patients. 'Target therapy' with tyrosine kinase inhibitors (TKIs) represent an important therapeutic option for the treatment of advanced cases of radioiodine refractory (RAI-R) differentiated thyroid cancer (DTC), medullary thyroid cancer (MTC) and possibly for cases of poorly differentiated (PDTC) and anaplastic thyroid cancer (ATC). In the last few years, several TKIs have been tested for the treatment of advanced, progressive and RAI-R thyroid cancers and some of them have been recently approved for use in clinical practice: sorafenib and lenvatinib for DTC and PDTC; vandetanib and cabozantinib for MTC. The objective of this overview is to present the current status of the treatment of advanced DTC, MTC, PDTC and ATC with the use of TKIs by describing the benefits and the limits of their use. A comprehensive analysis and description of the molecular basis of these drugs and the new therapeutic perspectives are also reported. Some practical suggestions are also given for the management to the potential side-effects of these drugs., (Copyright © 2017 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.)

Context: There is much debate surrounding the choice of which patient should be submitted to postsurgical remnant radioiodine remnant ablation (RRA), particularly in low-risk (LR) and intermediate-risk (IR) differentiated thyroid cancer (DTC)., Objective: The aim of this study was to evaluate the role of postoperative high-sensitive thyroglobulin (Tg) on L-thyroxine (LT4-HSTg) and postoperative neck ultrasound (US) in risk restratification and decision to perform RRA., Patients: We evaluated 505 patients with LR or IR DTC 3 to 4 months after total thyroidectomy (TTx). All patients underwent RRA and a posttherapeutic whole body scan (ptWBS)., Results: After TTx, 29.7% DTC patients had LT4-HSTg <0.1 ng/mL (Group A) and could be restratified as cured: 1 of 150 had lymph node metastases (LN mets) detected by neck US but negative at ptWBS. 56.8% DTC patients had LT4-HSTg between 0.1 and ≤1 ng/mL (Group B) and could be restratified either as cured or not cured. In this group, 15 of 287 (5.2%) had metastases but only 7 were detected by ptWBS; 13.5% DTC patients had LT4-HSTg >1 ng/mL (Group C) and could not be considered as cured by definition. LN mets were present in 11 of 68(16.2%) cases, all detected by neck US. No correlation was found with the presence of metastases and serum LT4-HSTg values or with the level of risk., Conclusions: LT4-HSTg measured 3 to 4 months after TTx is important in the risk restratification of DTC patients but is less relevant than neck US in the decision to perform RRA., (Copyright © 2017 by the Endocrine Society)

Background: Papillary (PTC) and medullary (MTC) thyroid carcinomas represent two distinct entities, but quite frequently, they may occur simultaneously., Aim: To provide genetic analysis of PTC and MTC occurring in the same patient (PTC/MTC) to elucidate their origin., Methods: Sequencing analysis of RAS, BRAF and RET oncogenes hot spots mutations in tumoral and normal tissues of 24 PTC/MTC patients., Results: Two of 24 patients (8.3 %) were affected by familial MTC (FMTC) harboring RET germline mutations in all tissues. Eight of 22 (36.4 %) sporadic cases did not show any somatic mutation in the three tissue components. Considering the MTC component, 10/22 (45.4 %) patients did not show any somatic mutation, 7 of 22 (31.8 %) harbored the M918T RET somatic mutation and 4/22 (18.2 %) presented mutations in the H-RAS gene. In an additional case (1/22, 4.6 %), H-RAS and RET mutations were simultaneously present. Considering the PTC component, 1 of 24 (4.2 %) patients harbored the V600E BRAF mutation, 1 of 24 (4.2 %) the T58A H-RAS mutation and 1 of 24 (4.2 %) the M1T K-RAS mutation, while the remaining PTC cases did not show any somatic mutation. In one case, the MTC harbored a RET mutation and the PTC a BRAF mutation. None of the mutations found were present in both tumors., Conclusions: To our knowledge, this is the first study analyzing a possible involvement of RET, BRAF and RAS oncogene mutations in PTC/MTC. These data clearly suggest that the classical activating mutations of the oncogenes commonly involved in the pathogenesis of PTC and MTC may not be responsible for their simultaneous occurrence.

Background: The RET proto-oncogene is responsible for the pathogenesis of hereditary (98%) and sporadic (40%) medullary thyroid carcinoma (MTC). In sporadic MTC, somatic RET mutations are associated with a poor prognosis., Objectives: We looked at the genetic profile of patients with advanced and metastatic MTC. The correlation between these mutations and outcome was also investigated., Methods: 70 patients with advanced and metastatic sporadic MTC were studied. Exons 10-11 and 13-16 of RET were analysed by direct sequencing. All cases were studied for RAS and the majority also for TERT mutations. RET/RAS -negative cases were analysed for other oncogene mutations., Results: 64/70 cases (91.4%) showed a somatic mutation, while 6 (8.6%) were negative. Among the mutated cases, RET mutations, mainly M918T, were the most prevalent (93.8%). K- or H-RAS mutations were present in 6.2% of cases and were mutually exclusive with RET. No other mutations were found. Four tumours showed two RET somatic mutations. We found a complex somatic RET alteration in 6/60 (10%) RET -positive sporadic MTC cases. A positive correlation between a poor prognosis and the multiple number of RET mutations was found., Conclusions: This study showed a high prevalence of somatic RET mutations in advanced and metastatic MTCs. RAS mutations were present in a small percentage of cases and mutually exclusive with RET mutations. In a small number of cases, more than one RET mutation was present in the same tissue. RET double mutations and, to a lesser extent, also complex mutations showed a worse outcome., Competing Interests: Competing interests: None declared., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.)

Thyroid cancer is rare, but it is the most frequent endocrine malignancy. Its prognosis is generally favorable, especially in cases of well-differentiated thyroid cancers (DTCs), such as papillary and follicular cancers, which have survival rates of approximately 95% at 40 years. However, 15-20% of cases became radioiodine refractory (RAI-R), and until now, no other treatments have been effective. The same problems are found in cases of poorly differentiated (PDTC) and anaplastic (ATC) thyroid cancers and in at least 30% of medullary thyroid cancer (MTC) cases, which are very aggressive and not sensitive to radioiodine. Tyrosine kinase inhibitors (TKIs) represent a new approach to the treatment of advanced cases of RAI-R DTC, MTC, PDTC, and, possibly, ATC. In the past 10 years, several TKIs have been tested for the treatment of advanced, progressive, and RAI-R thyroid tumors, and some of them have been recently approved for use in clinical practice: sorafenib and lenvatinib for DTC and PDTC and vandetanib and cabozantinib for MTC. The objective of this review is to present the current status of the treatment of advanced thyroid cancer with the use of innovative targeted therapies by describing both the benefits and the limits of their use based on the experiences reported so far. A comprehensive analysis and description of the molecular basis of these therapies, as well as new therapeutic perspectives, are reported. Some practical suggestions are given for both the choice of patients to be treated and their management, with particular regard to the potential side effects., (© 2016 Society for Endocrinology.)

Background: Medullary thyroid cancer (MTC) is capable of secreting several proteins, such as calcitonin (Ct), carcinoembryonic antigen (CEA), chromogranin and others. Recently, we observed an aggressive MTC with high levels of serum carbohydrate antigen 19.9 (Ca 19.9) and a rapid evolution to death., Objective: The aim of this study was to evaluate whether high levels of serum Ca 19.9 could be a prognostic factor of death in patients with advanced MTC., Patients and Methods: We measured Ca 19.9, CEA and Ct in 100 advanced structural recurrent/persistent MTC patients and in 100 cured or biochemically affected MTC patients. Clinical and pathological data were also collected., Results: Sixteen percent of the patients with advanced MTC had high levels of Ca 19.9. The group with abnormal Ca 19.9 levels had significantly higher levels of CEA and Ct compared with the group with normal values of Ca 19.9 (P<0.0001 for both Ct and CEA). At variance, all 100 patients in the MTC control group showed normal levels of Ca 19.9. Moreover, among the advanced cases, the Ca 19.9-positive group showed a higher mortality rate than the group with normal levels. A logistic regression analysis demonstrated that an elevated level of Ca 19.9 is a predictor of mortality (OR=3.78, P=0.04), independent from Ct doubling time., Conclusions: These results demonstrated that an elevated value of serum Ca 19.9 appears to be a predictive factor of poor prognosis in advanced MTC patients and identifies those cases with a higher risk of mortality in the short term., (© 2015 European Society of Endocrinology.)

Objective: Medullary thyroid carcinoma (MTC) is a rare disease that can be inherited or sporadic; its pathogenesis is related to activating mutations in the RET gene., Design: This study describes our 20-year experience regarding RET genetic screening in MTC., Patients and Methods: We performed RET genetic screening in 1556 subjects, 1007 with an apparently sporadic MTC, 95 with a familial form and 454 relatives of RET-positive patients with MTC., Results: A germline RET mutation was found in 68 of 1007 (6·7%) patients with sporadic MTC, while 939 patients with MTC were negative for germline RET mutations. We then identified a total of 137 gene carriers (GC). These subjects initiated a clinical evaluation for the diagnosis of MEN 2. A total of 139 MEN 2 families have been followed: 94 FMTC, 33 MEN 2A and 12 MEN 2B. Thirty-three different germline RET mutations were identified. Codon 804 was the most frequently altered codon particularly in FMTC (32/94, 34%), while codon 634 was the most frequently altered codon in MEN 2A (31/33, 94%); MEN 2B cases were exclusively associated with an M918T mutation at exon 16., Conclusions: Our 20-year study demonstrated that RET genetic screening is highly specific and sensitive, and it allows the reclassification as hereditary of apparently sporadic cases and the identification of GC who require an adequate follow-up. We confirmed that FMTC is the most prevalent MEN 2 syndrome and that it is strongly correlated with noncysteine RET mutations. According to these findings, a new paradigm of follow-up of hereditary MTC cases might be considered in the next future., (© 2014 John Wiley & Sons Ltd.)

Introduction: DTC patients having detectable Tg and negative post-therapeutic (131)I-WBS have to be investigated by different imaging techniques to detect metastases., Purpose: Comparison of neck US, CT and [18F]-FDG PET scan., Methods: In 49 DTC patients with biochemical disease, neck was examined by US, CT and [18F]-FDG PET. FNA was performed and Tg was determined by FNA-Tg in selected cases of suspicious lymph nodes. Thorax was examined by CT and PET. Serum Tg was measured on LT4 therapy (basal Tg) and after the stimulation with recombinant human TSH (peak Tg)., Results: A thyroid remnant was seen by US, CT and PET in eight patients; recurrences were seen by US, CT and PET in six, five and five patients, respectively. Two metastatic nodes were identified by US and CT but not by PET. Lung micronodules were detected by CT in 7/49 (14.3 %) patients and by FDG PET in three of them. Basal Tg ranged from 0.5-1,725 ng/ml while peak Tg ranged from 0.5 to 2,135 ng/ml: the distribution between positive and negative patients was similar. Bone scan was negative in all cases., Conclusions: In DTC patients with detectable Tg and negative I-131 post-therapy WBS, imaging examination revealed remnant or metastases in 43 % of cases. Remnant and recurrences were equally detected by the three techniques; US was better than [18F]-FDG PET for lymph node metastases since this latter method can give false both positive and negative results; chest examination is best made by CT versus FDG PET due to its higher spatial resolution.

Background: During follow-up for patients with medullary thyroid cancer (MTC), the levels of serum calcitonin and carcinoembryonic antigen are important, and the doubling time of these biomarkers significantly correlates with disease progression. Other antigens are present in tumor tissue and the sera of patients with MTC, but there are scarce published data on the serum levels of carbohydrate antigen 19-9 (Ca 19-9), a tumor marker primarily used for the diagnosis and follow-up of pancreatic and gastrointestinal neoplasias. Recently, the case of a 56-year-old woman with multiple endocrine neoplasia type 2B with high serum levels of Ca 19-9 was reported; this patient experienced rapid disease progression that led to her death., Case Presentation: A 28-year-old man was referred to the Department of Endocrinology of the University Hospital of Pisa with suspected MTC with laterocervical lymph node metastasis, a single liver lesion (10 mm), several bone metastases, and bilateral pheochromocytomas. RET genetic testing revealed a germline Cys634Arg mutation. During the hospitalization, the carcinoembryonic antigen and Ca 19-9 levels increased while the calcitonin concentration remained stable; despite the apparent stability of the lesions, the condition of the patient worsened rapidly and resulted in death., Conclusions: High levels of serum Ca 19-9 could be considered a marker of the dedifferentiation of MTC and disease aggressiveness, but additional data on the association between Ca 19-9 and advanced MTC are required to confirm this hypothesis.

Background: Approximately 60% of sporadic medullary thyroid carcinomas (sMTC) remain orphan of a recognized genetic cause. Recently, a high percentage of RAS point mutations have been described in RET-negative sMTC. The aim of this study was to assess the prevalence of RAS point mutations in a large series of MTC collected in four Italian centers., Methods: For this purpose, we studied codons 12, 13, and 61 of H-, K-, and N-RAS genes in 188 MTC samples, either hereditary or sporadic, by direct sequencing. Correlations between the RAS mutational status and the clinical-pathological features of MTC patients as well as a meta-analysis of all published data were performed., Results: The prevalence of RAS mutations in the present series of MTC was 10.1%, and 17.6% when considering only RET-negative cases. RAS mutations were found in MTC tumoral tissue, but not in peripheral blood indicating their somatic origin. A novel mutation in codon 72 (M72I) was found, but with a low or null transforming potential. No association was found between the presence of RAS mutations and the clinical-pathological features of the patients. Although not statistically significant, a positive association between the presence of RAS mutations and a better outcome was observed. The meta-analysis of all published studies confirmed a prevalence of 8.8% for RAS mutations in MTC., Conclusions: The prevalence of RAS mutations in our MTC series was relatively low and consistent with the meta-analysis data. Only somatic RAS mutations were found and only in RET-negative sMTC. Likewise, MTCs that harbor a RAS mutation identify a subgroup of tumors with less aggressive behavior. To our knowledge, this is the largest series of MTCs studied for the presence of mutations in RAS genes and the first meta-analysis on this specific topic.

We analyzed the in vitro effects of celecoxib, a COX-2 inhibitor, and determined if celecoxib can sensitize a human MTC-derived cell line (TT) to chemotherapeutics. We found that celecoxib induced apoptosis in TT cells and decreased drug efflux by reducing the expression of MDR-1 mRNA, which codes for the drug efflux pump P-gp. We also observed that TT cells were 10-fold more resistant to doxorubicin than to vinorelbine, mimicking what can be observed in clinical practice. In addition, we found that the combination of celecoxib and vinorelbine, but not doxorubicin, induced a significant reduction in cell viability and a significant increase in apoptosis. In conclusion, we showed that celecoxib was able to enhance the chemotherapeutic effect of vinorelbine. A clinical trial exploring the in vivo activities of celecoxib in MTC patients who cannot benefit from available treatments would be desirable, taking into account the possible risks of cardiovascular effects of this drug., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Background: Medullary thyroid carcinoma (MTC) is a calcitonin (CT)-producing C-cell tumor. In hereditary cases, a germline RET mutation is found in 98% of families. Because MTC is cured only if intrathyroidal, prophylactic thyroidectomy is recommended in the gene carrier (GC)., Aims: The aim was to determine whether thyroidectomy performed when stimulated CT becomes detectable is as safe as prophylactic thyroidectomy and to identify the serum CT cutoff able to distinguish intrathyroidal from extrathyroidal MTC., Patients: Eighty-four GC were prospectively enrolled; 53 of the 84 underwent total thyroidectomy, one refused surgery, and 30 with normal basal and stimulated CT were under surveillance. The follow-up ranged from 2 to 18 yr., Results: GC operated on for elevated stimulated CT included 27 GC with a positive peak CT at the screening and four cases who became positive after 4 yr. All of them had intrathyroidal MTC and no node metastases; all were cured after a mean follow-up of 7.5 yr. Among those operated on for detectable basal CT, intrathyroidal tumors were found when CT was below 60 pg/ml, whereas either node metastases or larger tumors were observed when CT was above 60 pg/ml. No correlation among serum CT, age, and type of RET mutation was observed. Thirty GC were still biochemically negative at the annual control., Conclusions: The time of thyroidectomy in GC with negative CT could be personalized and safely planned when stimulated CT becomes positive, independent of the type of RET mutation and patient's age. In this series, a basal CT below 60 pg/ml was always associated to an intrathyroidal localization of MTC.

About 30% of hereditary Medullary Thyroid Carcinoma (MTC) have been demonstrated to harbour imbalance between mutant and wild-type RET alleles. We studied the RET copy number alterations (RET CNA) in 65 MTC and their correlation with RET mutation and patients' outcome. Fluorescence in situ Hybridization and Real-time PCR revealed RET CNA in 27.7% MTC but only in a variable percentage of cells. In sporadic MTC, RET CNA were represented by chromosome 10 aneuploidy while in hereditary MTC by RET amplification. A significant higher prevalence of RET CNA was observed in RET mutated MTC (P=0.003). RET CNA was also associated to a poorer outcome (P=0.005). However, the multivariate analysis revealed that only RET mutation and advanced clinical stage correlated with the worst outcome. In conclusion, 30% MTC harbour RET CNA in variable percentage of cells suggesting cell heterogeneity. RET CNA can be considered a poor prognostic factor potentiating the poor prognostic role of RET mutation., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Germline and somatic RET oncogene mutations are found in 98% hereditary and 40% sporadic medullary thyroid carcinomas. Our aim was to analyse by in silico and in vitro assays the transforming activity of six rare RET mutations (T338I, V648I, M918V, A883T, S904F and M848T). Six known RET mutations were used as controls. The in silico analysis showed the highest score value (i.e. 65) for S904F, M848T, M918T and C634R, whereas L790F, G691S, T338I and V648I had 0 score. Intermediate score values were obtained by A883T (score=55), M918V, V804M and Y791F (score=15). The in vitro focus formation assay showed that cells transfected with S904F, M918T, M848T or C634R generated the largest number of focus formation units (FFU). Intermediate numbers of FFU were observed in cells transfected with M918V, V804M, Y791F or A883T, while cells transfected with L790F, G691S, T338I or V648I showed a number of FFU similar to control cells. A positive correlation between the in silico score and in vitro FFU was found (P=0.0005). Only cells transfected with M918T or C634R grew faster and generated higher number of colonies in soft agar than control cells. However, the cells that were transfected with V804M produced an intermediate number of colonies. In conclusion, two of the six rare RET mutations, S904F and M848T possessed a relatively high transforming activity but a low aggressiveness; the other four mutations T338I, V648I, M918V and A883T were low or non-transforming, and their ability to induce tumoural transformation might be related to particular genetic conditions.

Objective: This study was aimed to demonstrate the clinical benefits of rearranged during transfection (RET) genetic screening in patients with apparently sporadic medullary thyroid cancer (MTC) not only to identify the hereditary nature of the disease in the index case but also to discover family members harbouring the same germline mutations (i.e. gene carriers) who are unaware of their condition., Context: RET genetic screening allowed the identification of germline RET mutations in apparently sporadic MTC resulting in their re-classification as hereditary forms., Patients and Measurements: RET genetic screening was performed in 729 apparently sporadic MTC patients by direct sequencing RET exons 5, 8, 10, 11 and 13-16. Clinical and biochemical evaluation of gene carriers was also performed., Results: We discovered an unsuspected germline RET mutation in 47 of 729 (6·5%) apparently sporadic MTC who were re-classified as hereditary. We found 60 of 146 (41·1%) gene carriers, 35 of whom had biochemical or clinical evidence of MTC. Thirty gene carriers underwent total thyroidectomy and 27 of 30 (90%) were persistently cured after a mean follow-up of 6·0 years. As a further result of RET genetic screening, we observed a significantly higher prevalence of familial medullary thyroid cancer (FMTC) in our series with respect to the largest series of the International RET Consortium (P = 0·0002)., Conclusions: RET genetic screening of patients with apparently sporadic MTC represents a major tool for the preclinical diagnosis and early treatment of unsuspected affected family members and allows the identification of a relevant percentage of hidden FMTC., (© 2011 Blackwell Publishing Ltd.)

Objective: Multiple endocrine neoplasia type 2 (MEN 2) is a genetic disease characterized by medullary thyroid carcinoma (MTC) associated (MEN 2A and 2B) or not familial MTC (FMTC) with other endocrine neoplasia due to germline RET gene mutations. The prevalence of these rare genetic diseases and their corresponding RET mutations are unknown due to the small size of the study population., Methods: We collected data on germline RET mutations of 250 families with hereditary MTC followed in 20 different Italian centres., Results and Conclusions: The most frequent RET amino acid substitution was Val804Met (19.6%) followed by Cys634Arg (13.6%). A total of 40 different germline RET mutations were present. Six families (2.4%) were negative for germline RET mutations. The comparison of the prevalence of RET germline mutations in the present study with those published by other European studies showed a higher prevalence of Val804Met and Ser891Ala mutations and a lower prevalence of Leu790Phe and Tyr791Phe (P<0.0001). A statistically significant higher prevalence of mutations affecting non-cysteine codons was also found (P<0.0001). Furthermore, the phenotype data collection showed an unexpected higher prevalence of FMTC (57.6%) with respect to other MEN 2 syndromes (34% MEN 2A and 6.8% of MEN 2B). In conclusion, we observed a statistically significant different pattern of RET mutations in Italian MEN 2 families with respect to other European studies and a higher prevalence of FMTC phenotype. The different ethnic origins of the patients and the particular attention given to analysing apparently sporadic MTC for RET germline mutations may explain these findings.

Background: In the last decades, a marked increased prevalence of differentiated thyroid cancer (DTC) has been observed worldwide. The aim of this study was to evaluate the changing features of DTC referred to our institution between 1969 and 2004., Methods: Clinical and pathological features and prognostic factors were analyzed in 4187 DTC patients, subdivided into two groups: group 1 (n = 1215) and group 2 (n = 2972) diagnosed before and after 1990, respectively., Results: Group 2 showed an increased proportion of micropapillary carcinoma and a concomitant decrease of follicular histotype. Male percentage was greater in group 2, whereas median age at diagnosis was unchanged. DTC of group 2 were more frequently associated with multinodular goiter or autoimmune thyroiditis, but many were unexpected findings. Features of aggressiveness were significantly less frequent in group 2, and the survival rate was greater (98.7 vs. 91.4%, P < 0.0001). Gender, age, histotype, tumor size, extrathyroidal macroinvasion, and lymph node and/or distant metastases were found to be poor prognostic factors in both groups using univariate analysis, but with multivariate analysis, only advanced age (odds ratio = 22.52 for older patients) and advanced stage (odds ratio = 53.54 for more advanced cases) were independently correlated with a lower survival., Conclusions: DTC patients diagnosed after 1990 have smaller tumors with less advanced stage and a better prognosis. The question of whether this is related to the finding of tumors with a low clinical penetrance or to the anticipation of diagnosis remains to be clarified. Despite these significant differences, both advanced stage and older age still represent the most important poor prognostic factors for survival.

A low sodium iodide symporter (NIS) expression has been shown in papillary thyroid carcinomas (PTCs) harboring the BRAFV600E mutation. In the present study, we analyzed the mRNA expression of thyroid differentiation genes, glucose transporter (GLUT)-1 and GLUT-3, in 78 PTCs according to the presence of BRAFV600E or RET/PTC rearrangements. We found BRAFV600E and RET/PTC rearrangements in 35.8 and 19.4% of PTCs respectively. The mRNA expression of NIS and thyroperoxidase (TPO) genes were significantly lower (P<0.0001 and P=0.004 respectively) in BRAFV600E-positive PTC with respect to non-mutated samples. In support of this result, immunohistochemistry showed that the percentage of NIS-positive cells was significantly lower (P=0.005) in BRAFV600E-mutated PTC (mean 53.5%) than in negative cases (mean 72.6%). In contrast, no difference either in NIS or in any other thyroid differentiation genes' mRNA expression was found in PTC with or without RET/PTC rearrangements. When GLUT-1 and GLUT-3 mRNA expression was considered, no correlation was found either in BRAFV600E- nor in RET/PTC-mutated cases. In conclusion, this study confirmed the presence of a genetic alteration of BRAF and/or RET oncogenes in 64% of PTC cases and revealed a significant correlation of BRAFV600E mutation with a lower expression of both NIS and TPO. This latter finding could indicate that an early dedifferentiation process is present at the molecular level in BRAFV600E-mutated PTC, thus suggesting that the previously demonstrated poor prognostic significance of BRAFV600E mutation could be related to the dedifferentiation process more than to a more advanced stage at diagnosis.

Background: Medullary thyroid carcinoma (MTC) is a well-differentiated thyroid tumor that maintains the typical features of C cells. An advanced stage and the presence of lymph node metastases at diagnosis have been demonstrated to be the most important bad prognostic factors. Somatic RET mutations have been found in 40-50% of MTCs. Although a relationship between somatic mutations and bad prognosis has been described, data are controversial and have been performed in small series with short-term follow ups. The aim of this study was to verify the prognostic value of somatic RET mutations in a large series of MTCs with a long follow up., Methods: We studied 100 sporadic MTC patients with a 10.2 yr mean follow-up. RET gene exons 10-11 and 13-16 were analyzed. The correlation between the presence/absence of a somatic RET mutation, clinical/pathological features, and outcome of MTC patients was evaluated., Results: A somatic RET mutation was found in 43 of 100 (43%) sporadic MTCs. The most frequent mutation (34 of 43, 79%) was M918T. RET mutation occurrence was more frequent in larger tumors (P=0.03), and in MTC with node and distant metastases (P<0.0001 and P=0.02, respectively), thus, a significant correlation was found with a more advanced stage at diagnosis (P=0.004). A worse outcome was also significantly correlated with the presence of a somatic RET mutation (P=0.002). Among all prognostic factors found to be correlated with a worse outcome, at multivariate analysis only the advanced stage at diagnosis and the presence of a RET mutation showed an independent correlation (P<0.0001 and P=0.01, respectively). Finally, the survival curves of MTC patients showed a significantly lower percentage of surviving patients in the group with RET mutations (P=0.006)., Conclusions: We demonstrated that the presence of a somatic RET mutation correlates with a worse outcome of MTC patients, not only for the highest probability to have persistence of the disease, but also for a lower survival rate in a long-term follow up. More interestingly, the presence of a somatic RET mutation correlates with the presence of lymph node metastases at diagnosis, which is a known bad prognostic factor for the definitive cure of MTC patients.

Background: Germline RET gene mutations are causative of multiple endocrine neoplasia (MEN) 2 and may be identified by genetic screening. Three different syndromes are distinguished: MEN 2A, when medullary thyroid carcinoma (MTC) is associated with pheochromocytoma and/or parathyroid adenomas; MEN 2B, when accompanied by a marfanoid habitus and/or pheochromocytoma; and familial medullary thyroid carcinoma (FMTC), when only MTC is present., Patients and Methods: During the last 13 yr, we performed RET genetic screening in 807 subjects: 481 with apparently sporadic MTC, 37 with clinical evidence of MEN 2, and 289 relatives. Genomic DNA was extracted from the blood of all subjects, and exons 10, 11, 13, 14, 15, and 16 were analyzed by direct sequencing after PCR., Results: We unexpectedly discovered a germline RET mutation in 35 of 481 (7.3%) apparently sporadic MTC patients. A germline RET mutation was also found in 36 of 37 patients with clinical evidence of hereditary MTC. The distribution of RET mutations in cysteine and noncysteine encoding codons was significantly different in the two groups of patients, with the prevalence of RET mutations in noncysteine codons being higher in MTC that presented as apparently sporadic (P < 0.0001). A total of 34 FMTCs (75.5% of all FMTC) arrived with apparent sporadic MTC, with no familial history of other MTC cases. According to genetic screening and clinical data, our 72 families were classified as follows: 45 FMTC (62.5%), 22 MEN 2A (30.5%), and five MEN 2B (7%)., Conclusions: In this large series of MTC, hereditary forms, mainly FMTC, were clinically unsuspected in 7.3% of apparently sporadic cases. As a consequence, the prevalence of FMTC in our series is higher than that previously reported (60 vs. 10%). In these cases, RET mutations were more prevalently located in noncysteine codons. Data derived from our series helped elucidate the role of RET genetic screening for the identification of all forms of MEN 2, and especially for FMTC, which are frequently clinically misdiagnosed as nonheritable, sporadic cases.

Recently, a somatic point mutation of the B-RAF gene (V600E) has been identified as the most common genetic event in papillary thyroid carcinoma (PTC), with a prevalence variable among different series. Since discordant data on the clinico-pathologic features of B-RAF mutated PTC are present in the literature, the aim of the present co-operative study was to establish the prevalence of this genetic alteration and to perform a genotype-phenotype correlation in a large cohort of patients with PTC. To this purpose, a series of 260 sporadic PTCs with different histological variants were included in the study. The mutational analysis of the B-RAF gene was performed either by RT-PCR followed by single-stranded conformational polymorphism or by PCR and direct sequencing. Statistical analyses were obtained by means of chi2/Fisher's exact test and t-test. Overall, a heterozygous T > A transversion at nucleotide 1799 (V600E) was found in 99 out of 260 PTCs (38%). According to the histological type of the tumor, the B-RAF (V600E) mutation was present in 48.3% of cases of classic PTCs (85 out of 176), in 17.6% (nine out of 51) of follicular variants of PTCs, in 21.7% (five out of 23) in other PTC variants and in none of the ten poorly differentiated tumors. B-RAF (V600E) was significantly associated with the classic variant of PTC (P = 0.0001) and with an older age at diagnosis (P = 0.01). No statistically significant correlation was found among the presence of B-RAF (V600E) and gender, tumor node metastasis (TNM), multicentricity of the tumor, stage at diagnosis and outcome. In conclusion, the present study reports the prevalence of B-RAF (V600E) (38%) in the largest series of sporadic PTCs, including 260 cases from three different Italian referring centers. This prevalence is similar to that calculated by pooling together all data previously reported, 39.6% (759 out of 1914 cases), thus indicating that the prevalence of this genetic event lies around 38-40%. Furthermore, B-RAF (V600E) was confirmed to be associated with the papillary growth pattern, but not with poorer differentiated PTC variants. A significant association of B-RAF mutation was also found with an older age at diagnosis, the mutation being very rare in childhood and adolescent PTCs. Finally, no correlation was found with a poorer prognosis and a worse outcome after a median follow-up of 72 months.