Maria Joao Bugalho

Expression of iodine metabolism genes in human thyroid tissues: evidence for age and BRAFV600E mutation dependency

Context  Children present a higher susceptibility to developing thyroid cancer after radioiodine exposure and also a higher frequency of functional metastases than adults.

Identification of the first germline HRPT2 whole‐gene deletion in a patient with primary hyperparathyroidism

Objective  Germline mutations in the HRPT2 gene are associated with the hereditary hyperparathyroidism‐jaw tumour syndrome (HPT‐JT) and a subset of familial isolated hyperparathyroidism (FIHP). Somatic HRPT2 mutations are detected in sporadic parathyroid carcinomas and less frequently in cystic adenomas. The purpose of this study was to investigate the underlying HRPT2 defect in a young patient with symptomatic hyperparathyroidism due to an apparently sporadic parathyroid adenoma with cystic features.

Rac1B overexpression in Papillary Thyroid Carcinoma: a role to unravel

CONTEXT The BRAF V600E mutation is the most frequent genetic alteration in papillary thyroid carcinoma (PTC). In colorectal cancer, BRAF V600E was described to functionally cooperate with RAC1b, a hyperactive splice variant of the small GTPase RAC1, to sustain cell survival. This interplay has never been investigated in PTCs. OBJECTIVE We aimed to analyze the expression of RAC1b in PTC and correlate its expression with BRAF V600E mutational status, histopathological features, and clinical outcome. PATIENTS AND METHODS Sixty-one patients and 87 samples (61 PTCs and 26 normal thyroid tissues) were included. Patients were divided into two groups based on longitudinal evolution and final outcome. RAC1b expression levels were determined by quantitative RT-PCR and western blotting. RESULTS RAC1b was expressed in thyroid and overexpressed in 46% of PTCs. Neither RAC1b overexpression nor V600E mutation correlated with histopathological features classically associated with worse prognosis. RAC1b overexpression was significantly associated with both V600E mutation (P=0.0008) and poor clinical outcome (P=0.0029). Whereas BRAF V600E alone did not associate with patient outcome (P=0.2865), the association of RAC1b overexpression with BRAF V600E was overrepresented in the group with poorer clinical outcome (P=0.0044). CONCLUSIONS Present results document, for the first time, expression of RAC1b in normal thyroid cells as well as overexpression in a subset of PTCs. Furthermore, they suggest a possible interplay between BRAF V600E and RAC1b contributing to poor clinical outcome. Future studies are needed to clarify the oncogenic potential of RAC1b in thyroid carcinogenesis.

Occurrence of the Cys611Tyr mutation and a novel Arg886Trp substitution in the RET proto‐oncogene in multiple endocrine neoplasia type 2 families and sporadic medullary thyroid carcinoma cases originating from the central region of Portugal

Objective  Medullary thyroid carcinoma (MTC) occurs both sporadically and in the context of autosomal dominantly inherited multiple endocrine neoplasia type 2 (MEN2) syndromes: MEN2A, MEN2B, and familial medullary thyroid carcinoma (FMTC), which are caused by activating germline mutations in the RET proto‐oncogene. The aim of this study was to characterize the RET mutational spectrum in MEN2 families and apparently sporadic MTC (AS‐MTC) cases originating from the central region of Portugal.

A kindred with multiple endocrine neoplasia type 2A associated with pruritic skin lesions.

Background. A kindred affected by multiple endocrine neoplasia type 2A (MEN 2A), associated with symmetric, bilateral, scapular pruritic skin lesions (PSL), is reported.

Molecular diagnosis of multiple endocrine neoplasia Type 2.

Multiple endocrine neoplasia Type 2 is a rare familial cancer syndrome transmitted in an autosomal dominant manner. It is characterized by the association of medullary thyroid carcinoma with pheochromocytoma and hyperparathyroidism. Medullary thyroid carcinoma, present in virtually all patients, is the principal cause of death. In 1993, germline mutations in the RET proto-oncogene were identified as the underlying cause of the syndrome. Genetic screening of at-risk family members can now be performed with high specificity and sensitivity. The ability to determine gene carrier status at a preclinical stage is of great value as it allows early prophylactic thyroidectomy. The specific RET codon mutation correlates with clinical variants of the syndrome, age at onset and aggressiveness of medullary thyroid carcinoma. This review will focus on mutational spectrum, genotype–phenotype correlations and clinical decisions based on genetic information.

Calcitonin‐producing insulinoma: clinical, immunocytochemical and cytogenetical study

The case of a patient with a large goitre associated with hypercaicitoninaemia and fasting hypoglycaemia is reported. Pentagastrin (PG) test was negative. Repeated measurements of fasting glycaemia, insulin and C peptide established the diagnosis of insulinoma. After localization by endoscopic ultrasonography, a distal pancreatectomy was performed and a small insulinoma was recovered. Glycaemia and calcitonin (cT) became normal. The tumour cells a a strong immunoreactivity for insulin and CT. Cytogenetical evaluation of the tumour revealed a translocation t(1;9) (pl3;p22).

RAC1b overexpression stimulates proliferation and NF-kB-mediated anti-apoptotic signaling in thyroid cancer cells

Overexpression of tumor-associated RAC1b has been recently highlighted as one of the most promising targets for therapeutic intervention in colon, breast, lung and pancreatic cancer. RAC1b is a hyperactive variant of the small GTPase RAC1 and has been recently shown to be overexpressed in a subset of papillary thyroid carcinomas associated with unfavorable outcome. Using the K1 PTC derived cell line as an in vitro model, we observed that both RAC1 and RAC1b were able to induce a significant increase on NF-kB and cyclin D1 reporter activity. A clear p65 nuclear localization was found in cells transfected with RAC1b-WT, confirming NF-kB canonical pathway activation. Consistently, we observed a RAC1b-mediated decrease in IκBα (NF-kB inhibitor) protein levels. Moreover, we show that RAC1b overexpression stimulates G1/S progression and protects thyroid cells against induced apoptosis, the latter through a process involving the NF-kB pathway. Present data support previous findings suggesting an important role for RAC1b in the development of follicular cell-derived thyroid malignancies and point out NF-kB activation as one of the molecular mechanisms associated with the pro-tumorigenic advantage of RAC1b overexpression in thyroid carcinomas.

Ectopic cushing in a patient with medullary thyroid carcinoma: hypercortisolism control and tumor reduction with Sunitinib

Medullary thyroid carcinoma (MTC) is rarely associated with ectopic Cushing’s syndrome (CS). Management of this condition is challenging. Kinase inhibitors (KI) may play a role in this setting [1–3]. We report the case of an advanced MTC and ectopic CS successfully treated with Sunitinib. In 2005, the patient had a total thyroidectomy and bilateral modified radical neck dissection followed by radiotherapy. RET germline mutation screening was negative. In 2011, imaging studies documented mediastinal and pulmonary metastases. The patient remained asymptomatic, and no treatment was undertaken. In July 2013, at the age of 64, he developed proximal muscle weakness and weight gain in less than 1 month. Physical examination revealed plethora, moon facies, abdominal obesity, hypertension, and ankle edema. Tests were consistent with ACTH-dependent CS: plasma ACTH = 83.4 pg/mL (normal\46); morning serum cortisol = 33 lg/dL (reference range 5–25); midnight serum cortisol = 29.8 lg/dL; 24-h urine free cortisol (UFC) = 2,647 lg/24 h (reference range 21–85). There was no suppression under dexamethasone. No adenoma was seen on pituitary magnetic resonance. The known mediastinal and lung metastases had increased in number and dimensions (Fig. 1), and calcitonin was 10,065 ng/L. Ga-DOTANOCPET/CT showed low expression of somatostatin receptors in the metastatic lesions. After starting metyrapone, there were clinical and laboratorial improvements (Fig. 1). However, fatigue, nausea, and diarrhea made us to consider alternative approaches. Somatostatin analogs were excluded, considering the low expression of somatostatin receptors. Bilateral adrenalectomy was considered a treatment of last resort. Based on previous reports on the use of KI to control ectopic secretion of ACTH [1–3] and taking into account our own experience with Sunitinib, in the setting of MTC, we decided to use this KI. After starting treatment, ACTH normalized, UFC remained controlled, and calcitonin decreased (Fig. 1). The initial dose of 50 mg/day was adjusted to 25 mg/day due to complaints of fatigue, diarrhea, and myalgia. Using the lower dose, on a continuous schedule, it was possible to maintain the treatment. After 3 months, RECIST evaluation documented partial response (Fig. 1). After 20 weeks under Sunitinib, the patient remains asymptomatic with normal ACTH and UFC. In this case, ACTH secretion was likely due to MTC, considering the parallel worsening of CS and MTC. The priority was correction of the life-threatening hypercortisolism. Therefore, metyrapone was introduced, and a remarkable reduction in UFC was observed. Treatment with Sunitinib leads to a dramatic reduction in ACTH and calcitonin levels almost immediately as such not correlating with tumor response. If an additional and direct effect, on cortisol synthesis, contributed to the final results, remains unknown. The control of hypercortisolism with different KIs [1–3] is likely to suggest an antisecretory class effect yet underexplored. The antisecretory effect observed with Vandetanib [1, 2] or Sorafenib [3] was not associated with tumor reduction. On the contrary, treatment with Sunitinib resulted in control of hypercortisolism and tumor shrinkage; the low dose of 25 mg allowed a continuous regimen P. Marques (&) M. da Silva Vieira M. J. Bugalho Endocrinology Department, Instituto Português de Oncologia de Lisboa, Francisco Gentil, 1099-023 Lisbon, Portugal e-mail: pedro.miguel.sousa.marques@gmail.com

Retrospective Analysis of 255 Papillary Thyroid Carcinomas ≤2 cm: Clinicohistological Features and Prognostic Factors

Background: Papillary thyroid carcinoma (PTC) is the most common thyroid cancer. The widespread use of neck ultrasound (US) and US-guided fine-needle aspiration cytology is triggering an overdiagnosis of PTC. Objective: To evaluate clinical behavior and outcomes of patients with PTCs ≤2 cm, seeking for possible prognostic factors. Methods: Clinical records of cases with histological diagnosis of PTC ≤2 cm followed at the Endocrine Department of Instituto Português de Oncologia, Lisbon between 2002 and 2006 were analyzed retrospectively. Results: We identified 255 PTCs, 111 were microcarcinomas. Most patients underwent near-total thyroidectomy, with lymph node dissections in 55 cases (21.6%). Radioiodine therapy was administered in 184 patients. At the last evaluation, 38 (14.9%) had evidence of disease. Two deaths were attributed to PTC. Median (±SD) follow-up was 74 (±23) months. Multivariate analysis identified vascular invasion, lymph node and systemic metastases significantly associated with recurrence/persistence of disease. In addition, lymph node involvement was significantly associated with extrathyroidal extension and angioinvasion. Median (±SD) disease-free survival (DFS) was estimated as 106 (±3) months and the 5-year DFS rate was 87.5%. Univariate Cox analysis identified some relevant parameters for DFS, but multivariate regression only identified lymph node and systemic metastases as significant independent factors. The median DFS estimated for lymph node and systemic metastases was 75 and 0 months, respectively. Conclusions: In the setting of small PTCs, vascular invasion, extrathyroidal extension and lymph node and/or systemic metastases may confer worse prognosis, perhaps justifying more aggressive therapeutic and follow-up approaches in such cases.