Gabriella Pellegriti

BRAF(V600E) mutation and the biology of papillary thyroid cancer

BRAF((V600E)) mutation is the most frequent genetic alteration in papillary thyroid carcinomas (PTCs) that are 80-90% of all thyroid cancers. We evaluated the relationship between BRAF((V600E)) and tumor, host, and environmental factors in PTCs from all geographical areas of Sicily. By PCR, BRAF((V600E)) was investigated in a series of 323 PTCs diagnosed in 2002-2005. The correlation between clinicopathological tumor, host, and environmental characteristics and the presence of BRAF((V600E)) were evaluated by both univariate and multivariate analyses. BRAF((V600E)) was found in 38.6% PTCs, with a 52% frequency in the classical PTCs and 26.4% in the tall cell variant. Univariate analysis indicated that BRAF((V600E)) was associated with greater tumor size (P=0.0048), extra-thyroid invasion (P<0.0001), and cervical lymph nodal metastases (P=0.0001). Multivariate logistic regression analysis confirmed that BRAF((V600E)) was an independent predictor of extra-thyroid invasion (P=0.0001) and cervical lymph nodal metastasis (P=0.0005). The association between BRAF((V600E)) and extra-thyroid invasion was also found in micro-PTCs (P=0.006). In 60 classical PTCs, BRAF((V600E)) was positively correlated with matrix metalloproteinase-9 expression (P=0.0047), suggesting a possible mechanism for BRAF((V600E)) effect on PTC invasiveness. No association was found between BRAF((V600E)) and patient age, gender, or iodine intake. In contrast, a strong association was found with residency in Eastern Sicily (P<0.0001 compared with Western Sicily). These results indicate that BRAF((V600E)) mutation is a marker of aggressive disease in both micro- and macro-PTCs. Moreover, for the first time, a possible link between BRAF((V600E)) mutation and environmental carcinogens is suggested.

Papillary Thyroid Cancer Incidence in the Volcanic Area of Sicily

BACKGROUND The steadily increasing incidence of thyroid cancer has been attributed mostly to more sensitive thyroid nodule screening. However, various environmental factors, such as those associated with volcanic areas, cannot be excluded as risk factors. We evaluated thyroid cancer incidence in Sicily, which has a homogenous population and a province (Catania) that includes the Mt Etna volcanic area. METHODS In a register-based epidemiological survey, we collected all incident thyroid cancers in Sicily from January 1, 2002, through December 31, 2004. The age-standardized incidence rate for the world population (ASR(w)) was calculated and expressed as the number of thyroid cancer diagnoses per 100 000 residents per year. The association of thyroid cancer incidence rate with sex, age, tumor histotype, and various environmental factors was evaluated by modeling the variation of the ASR(w). All statistical tests were two-sided. RESULTS In 2002-2004, 1950 incident thyroid cancers were identified in Sicily (among women, ASR(w) = 17.8, 95% confidence interval [CI] = 16.9 to 18.7; and among men, ASR(w) = 3.7, 95% CI = 3.3 to 4.1). Although the percentage of thyroid cancers that were microcarcinomas (ie, < or = 10 mm) and ratio of men to women with thyroid cancer were similar in all nine Sicilian provinces, thyroid cancer incidence was statistically significantly higher in the province of Catania (among women, ASR(w) = 31.7, 95% CI = 29.1 to 34.3; and among men, ASR(w) = 6.4, 95% CI = 5.2 to 7.5) than in the rest of Sicily (among women, ASR(w) = 14.1, 95% CI = 13.2 to 15.0; and among men, ASR(w) = 3.0, 95% CI = 2.6 to 3.4) (all P values < .001). Incidence of papillary, but not follicular or medullary, cancers was statistically significantly increased in Catania province, and papillary tumors from patients in Catania more frequently carried the BRAF V600E gene mutation (55 [52%] of 106 tumors) than tumors from patients elsewhere in Sicily (68 [33%] of 205 tumors) (relative risk = 1.7, 95% CI = 1.0 to 2.8, P = .02). Cancer incidence was statistically significantly lower in rural areas than in urban areas of Sicily (P = .003). No association with mild iodine deficiency or industrial installations was found. Levels of many elements (including boron, iron, manganese, and vanadium) in the drinking water of Catania province often exceeded maximum admissible concentrations, in contrast to water in the rest of Sicily. CONCLUSION Residents of Catania province with its volcanic region appear to have a higher incidence of papillary thyroid cancer than elsewhere in Sicily.

Risk-Adapted Management of Differentiated Thyroid Cancer Assessed by a Sensitive Measurement of Basal Serum Thyroglobulin

CONTEXT Treatment and follow-up of patients thyroidectomized for differentiated thyroid carcinoma (DTC) mainly depends on the identification of the patients risk of recurrence. Thyroglobulin (Tg) is the most important marker of persistent/recurrent disease. The recent introduction of a new, more sensitive Tg measurement allows for the early detection of the disease by measuring the basal (under L-T(4) therapy) serum Tg level without TSH stimulation. OBJECTIVE The goal of this study is to identify the basal serum Tg threshold value that indicates recurrent disease by using a second-generation Tg assay. DESIGN AND PATIENTS A continuous series of 425 DTC patients, all thyroidectomized and treated with (131)I after surgery and having basal Tg of no more than 1.0 ng/ml, negative anti-Tg antibodies, and a recombinant human TSH-stimulated Tg measurement was retrospectively analyzed. SETTING The study took place at an academic hospital. RESULTS The most accurate basal Tg value for predicting the presence of recurrent/residual disease was more than 0.15 ng/ml (sensitivity 87%, specificity 91%, negative predictive value 98.6%, and positive predictive value 47.8%). When the basal Tg level was no more than 0.15 ng/ml, the risk of disease presence was very low, even in patients classified at an intermediate or high risk. In contrast, when the basal Tg level was more than 0.15 ng/ml, the percentage of recurrent disease was relatively high (12.5% or one in eight cases) in low-risk patients. CONCLUSIONS Basal Tg, measured using a second-generation Tg assay allows for the identification of DTC patients who are likely to remain disease free with great accuracy. This simple measurement, therefore, may be sufficient to assess the risk-adapted management of DTC patients.

The influence of the environment on the development of thyroid tumors: a new appraisal

Most epidemiological studies concerning differentiated thyroid cancers (DTC) indicate an increasing incidence over the last two decades. This increase might be partially explained by the better access to health services worldwide, but clinicopathological analyses do not fully support this hypothesis, indicating that there are carcinogenetic factors behind this noticeable increasing incidence. Although we have undoubtedly understood the biology and molecular pathways underlying thyroid carcinogenesis in a better way, we have made very little progresses in identifying a risk profile for DTC, and our knowledge of risk factors is very similar to what we knew 30-40 years ago. In addition to ionizing radiation exposure, the most documented and established risk factor for DTC, we also investigated the role of other factors, including eating habits, tobacco smoking, living in a volcanic area, xenobiotics, and viruses, which could be involved in thyroid carcinogenesis, thus, contributing to the increase in DTC incidence rates observed.

Update on thyroid cancer treatment

Surgery and radioiodine therapy are usually effective for most patients with differentiated thyroid cancer. However, poorly differentiated and anaplastic thyroid carcinomas represent a challenge to physicians on the basis of the current cancer treatment modalities. These cancer subtypes are often lethal and refractory to radioiodine therapy as well as most of the common chemotherapy drugs. Several kinase inhibitors are promising targeted therapies for these malignancies; however, clinical trials involving these drugs have provided controversial results and their clinical use is still under debate. Advanced medullary thyroid carcinomas may also be refractory to conventional therapies and novel kinase inhibitors may also be useful to control tumor progression in certain patients. Novel evidence is emerging that thyroid cancer is a stem cell disease, thereby implying that the driving force of thyroid cancers is a subset of undifferentiated cells (thyroid cancer stem cells) with unlimited growth potential and resistance to conventional therapeutic regimens. Thyroid cancer stem cells have been proposed as responsible for tumor invasiveness, metastasis, relapse and differentiation. Therefore, drugs that selectively target these cells could serve as a cornerstone in the treatment of poorly differentiated thyroid cancer.

Descriptive Epidemiology of Human Thyroid Cancer: Experience From a Regional Registry and The “Volcanic Factor”

Thyroid cancer (TC), the most common endocrine tumor, has steadily increased worldwide due to the increase of the papillary histotype. The reasons for this spread have not been established. In addition to more sensitive thyroid nodule screening, the effect of environmental factors cannot be excluded. Because high incidences of TC were found in volcanic areas (Hawaii and Iceland), a volcanic environment may play a role in the pathogenesis of TC. In January 2002, the Regional Register for TC was instituted in Sicily. With a population of approximately five million inhabitants with similar genetic and lifestyle features, the coexistence in Sicily of rural, urban, industrial, moderate-to-low iodine intake, and volcanic areas provides a conducive setting for assessing the environmental influences on the etiology of TC. In Sicily, between 2002 and 2004, 1,950 new cases of TC were identified, with an age-standardized rate (world) ASR(w) = 17.8/105 in females and 3.7/105 in males and a high female/male ratio (4.3:1.0). The incidence of TC was heterogeneous within Sicily. There were 2.3 times more cases in the Catania province (where most of the inhabitants live in the volcanic area of Mt. Etna): ASR(w) = 31.7/105 in females and 6.4/105 in males vs. 14.1 in females and 3.0 in males in the rest of Sicily. Multivariate analysis documented that residents in the volcanic area of Mt. Etna had a higher risk of TC, compared to the residents in urban, industrial, and iodine deficient areas of Sicily. An abnormally high concentration of several chemicals was found in the drinking water of the Mt. Etna aquifer, which provides water to most of the residents in the Catania province. Our data suggest that environmental carcinogen(s) of volcanic origin may promote papillary TC. Additional analyses, including cancer biological and molecular features, will allow a better understanding of risk factors and etiopathogenetic mechanisms.

The BRAFV600E Mutation Influences the Short- and Medium-Term Outcomes of Classic Papillary Thyroid Cancer, But Is Not an Independent Predictor of Unfavorable Outcome

INTRODUCTION The prognostic usefulness of BRAF(V600E) evaluation in papillary thyroid cancer (PTC) has been analyzed in many studies, with controversial conclusions. AIM To analyze the clinical relevance of BRAF(V600E) measurement in a homogenous series of PTC patients followed in a single institution. METHODS One hundred three classical variant PTC patients who underwent total thyroidectomy in the 3-year period between 2005 and 2008 were retrospectively selected, and BRAF(V600E) assessment was performed using paraffin-embedded archival specimens in 2013. All patients were actively followed at our medical center, with an average follow-up of 55±13 months. RESULTS BRAF(V600E) mutation-positive cancers (55.3%) were more frequently associated with lymph node metastasis (p=0.01) and advanced TNM stage (III-IV) (p=0.03). These findings were also confirmed in the subset of 42 microcarcinomas. BRAF(V600E)-positive patients were also at a higher risk of persistent disease (OR 3.5 [95% confidence interval {CI} 1.2-10.3], p=0.03) in univariate but not multivariate analysis (OR 2.8 [CI 0.7-11.8], p=0.2). Lymph node involvement was an independent predictor of persistent disease (OR 30.9 [CI 6.0-159.0], p<0.0001). Kaplan-Meier curves confirmed a higher percentage of persistent/recurrent disease in BRAF(V600E)-positive patients (p=0.02). However, the BRAF(V600E) mutation did not change the recurrence rate of PTC in subgroup analyses on the basis of other established risk factors (p=0.2). CONCLUSIONS BRAF(V600E)-positive tumors were at higher risk of developing more aggressive behavior and were associated with less favorable outcomes in the short and medium term, but the BRAF(V600E) mutation was not an independent predictor of unfavorable outcome. Therefore, its use as a prognostic marker in clinical practice is not advisable.

Cardiac Arrest After Intravenous Calcium Administration for Calcitonin Stimulation Test

Medullary thyroid cancer (MTC) is an aggressive tumor, deriving from the parafollicular C-cells of the thyroid. Screening, diagnosis, and follow-up of MTC require evaluation of serum calcitonin (CT), a highly sensitive and specific marker of this cancer. When basal CT levels are nondiagnostic (mildly elevated), a stimulation test is useful to differentiate a neoplastic (MTC) or preneoplastic (C-cell hyperplasia) hypercalcitoninemia from nonmalignant conditions. Measurement of a stimulated CT is also helpful to decide the timing of prophylactic thyroidectomy in RET gene mutation carriers in familial MTC and in postsurgical follow-up of MTC patients (1,2). CT stimulation can be obtained by either pentagastrin injection or by short intravenous calcium gluconate infusion (2– 2.5 mg/kg of elemental calcium administered at 10 mL/min) (1,3). The calcium stimulation test is preferred to pentagastrin (which is currently unavailable) because it is causing less patient discomfort. The main reported adverse effects are paresthesia of the extremities and/or lips, feeling of warmth, flushing, and a chalky taste sensation (1,3). We describe a severe adverse event that occurred in a healthy volunteer during a pilot study designed to assess the reference ranges for calcium-stimulated serum CT levels in our Medical Center. A 28-year-old man in good health, without any known chronic or acute illness, cardiac disease, or thyroid abnormality volunteered for a calcium stimulation test. He did not smoke and took no medication, and his weight was 60 kg. Pretest evaluation indicated that electrolytes, liver enzymes, glucose, creatinine, blood pressure, and ECG were normal. The subject gave written informed consent. Two mg/kg of elemental calcium (calcium gluconate 10%; B. Braun, 10 mL vials; B. Braun, Melsungen, Germany) was infused intravenously at 10 mL/minute while the subject was in a supine position. A few seconds after completion of the infusion, the subject became unresponsive, his pulse was not present, and an electrocardiogram (ECG) revealed asystolia. Cardiopulmonary resuscitation was immediately executed, and the heart rate was restored. The patient quickly regained consciousness, was able to answer questions, and did not complain of any discomfort. Further evaluation did not reveal any cardiac abnormality (QT interval was in normal range and an echocardiogram showed no pathological findings) and after three hours of observation, he was discharged. This clinical case indicates that the CT stimulation test with calcium gluconate infusion, performed according to the currently indicated procedures, may have severe adverse effects. Previously, a case of atrial fibrillation after combined administration of calcium gluconate (2 mg/kg in 1 min) and pentagastrin (0.5 lg/kg over 5 sec) has been reported. The patient was converted to normal sinus rhythm with digoxin. In that case, the agent responsible for the adverse reaction could not be determined (4). More recently, persistent atrial fibrillation has been observed in a patient after a calcium stimulation test (5). In our case, the calcium infusion caused a severe adverse effect (cardiac arrest) in a healthy young man. The adverse event could have caused more severe consequences if the test had not been carried out under continuous cardiac monitoring inside a medical center. It is well known that rapid intravenous calcium injection may cause vasodilatation, decreased blood pressure, bradycardia, cardiac arrhythmias, syncope, and cardiac arrest (6). Even if rare, these possible adverse effects of this diagnostic test must be considered because they are potentially life-threatening. Therefore, appropriate procedures (including continuous cardiac monitoring) should be followed to guarantee rapid intervention in case of an adverse cardiovascular event. A slower calcium infusion procedure should be evaluated and the calcium stimulation test use should be limited to conditions such as the evaluation of RET gene mutation carriers (2) and suspicious MTC patients with CT values mildly elevated. Its use in the postoperative follow-up of MTC patients with undetectable basal serum CT should be avoided, as suggested in the ATA guidelines (7). In these patients, the risk of persistent or recurrent disease is low and the improved sensitivity of basal serum CT measurements has reduced the role of stimulation testing (7).

Familial Non-Medullary Thyroid Cancer Represents an Independent Risk Factor for Increased Cancer Aggressiveness: A Retrospective Analysis of 74 Families.

Objectives To assess whether familial non-medullary thyroid cancer (FNMTC) represents an independent risk factor for increased aggressiveness of the tumor, as concern as the clinical presentation and the long-term follow-up in respect of sporadic differentiated thyroid cancer (SDTC). Design Retrospective study; 1976–2014. Patients and Methods Seventy-four FNMTC families (151 affected individuals): family relationship and number of affected family members were evaluated. Clinical and histopathological features and outcome were compared to that of 643 SDTC patients followed in the same period according to the same institutional protocols. Median follow-up was 57.7 months (range 12–136) in FNMTC and 59.7 (range 15–94.6) in SDTC patients. Results Three cases occurred in 3 families and 2 cases in the other 71. F:M was 3.7:1 in FNMTC and 4.3:1 in SDTC (NS). The family relationship was siblings in 62.2%. Mean age at diagnosis was lower in FNMTC than in SDTC (p < 0.005). Papillary/follicular histotype distribution was similar (86%). Papillary tumors were more frequently multifocal in FNMTC (p = 0.004) and with lymph-node metastases (p = 0.016). Disease-free survival (DFS) was shorter in FNMTC vs. SDTC (p < 0.0001) with 74.8 vs. 90.8% patients free of disease at the last control (p < 0.005). Three patients died in FNMTC group vs. 1 in SDTC (p = 0.02). Conclusion Familial non-medullary thyroid cancer displays distinct characteristics as earlier age of onset and increased aggressiveness at diagnosis and a higher rate of persistent/recurrent disease and mortality with a shorter DFS in respect with SDTC. FNMTC patients, therefore, should be followed accurately. As the specific gene (or genes) responsible for susceptibility for FNMTC has not yet been identified, a low frequency periodic screening of relatives DTC patients may be useful to identify FNMTC patients at early stage of disease.

A novel RET gene mutation in a patient with apparently sporadic pheochromocytoma

Pheochromocytoma (Pheo) is a chromaffin tumor arising from the adrenal medulla. The recent discovery of new germline mutations in RET, SDHA, SDHB, SDHC, SDHD, VHL, NF1, TMEM127, MAX genes, increased the rate of genetic disease from 10% to 28% in patients with apparently sporadic tumor. RET germline mutations cause multiple endocrine neoplasia type 2 syndrome (MEN 2A) characterized by complete penetrance of medullary thyroid cancer (MTC), and lower prevalence of Pheo and hyperparathyroidism. We describe the genetic etiology of an apparently sporadic case of monolateral Pheo in a 42-year-old male patient. A new (not previously reported) MEN 2A-associated germline RET mutation located in exon 11 (Glu632Gly, caused by an A>G point mutation at position 1895 of the RET cDNA) was found in the patient but not in his living first-degree relatives. This observation increases the number of possible germline RET mutations. Genotype-phenotype correlation of this new genetic alteration is unknown, but this rare mutation is probably associated with a low risk for MTC (usually the first tumor diagnosed in MEN 2A syndrome) and with the development of Pheo before the onset of MTC. Since we expect MTC to occur in our patient, strict follow-up is mandatory. Our findings emphasize the relevance of genetic testing in patients with Pheo, especially when the clinical presentation (family history, young age at diagnosis, multiple locations, malignant lesions, and bilateralism) is suggestive.