Owing to the heterogeneity of neuroendocrine neoplasms (NENs), the availability of reliable circulating markers is critical for improving diagnostics, prognostic stratification, follow-up and definition of treatment strategy. This review is focused on chromogranin A (CgA), a hydrophilic glycoprotein present in large dense core vesicles of neuroendocrine cells. Despite being long identified as the most useful NEN-related circulating marker, clinical application of CgA is controversial. CgA assays still lack standardization, thus hampering not only clinical management but also the comparison between different analyses. In the diagnostic setting, clinical utility of CgA is limited as hampered by (a) the variety of oncological and non-oncological conditions affecting marker levels, which impairs specificity; (b) the fact that 30–50% of NENs show normal CgA, which impairs sensitivity. Regarding the prognostic phase, there is prospective evidence which demonstrates that advanced NENs secreting CgA have poorer outcome, as compared with those showing non-elevated marker levels. Although the identification of cut-offs allowing a proper risk stratification of CgA-secreting patients has not been performed, this represents the most important clinical application of the marker. By contrast, based on prospective studies, the trend of elevated circulating CgA does not represent a valid indicator of morphological evolution and has therefore no utility for the follow-up phase. Ultimately, current knowledge about the role of the marker for the definition of treatment strategy is poor and is limited by the small number of available studies, their prevalent retrospective nature and the absence of control groups of untreated subjects.

BRAF(V600E) is considered a primary event, a negative prognostic marker, and a site for pharmacological intervention in papillary thyroid carcinoma (PTC). We asked whether BRAF(V600E) can occur as a subclonal event in PTC and whether this and other oncogenes can coexist in the same tumor. We determined by pyrosequencing the percentage of mutant BRAF, NRAS, and KRAS alleles in a series of conventional PTC. We also analyzed the BRAF mutation status in PTC cell clones in culture. BRAF(V600E) alleles were present in 41 of 72 PTC (56.9%) in the range 44.7 to 5.1% of total BRAF alleles. In four PTC samples, mutant BRAF alleles were about 50%, being therefore compatible with a clonal heterozygous mutation. In 27 PTC samples, BRAF(V600E) alleles were in the range of 25 to 5.1%. This finding was confirmed after exclusion of the presence of a large contamination by lymphoreticular cells and by the analysis of PTC cells selected by laser capture. Analysis of clones derived from a single cell confirmed the presence of two distinct PTC populations with wild-type or mutated BRAF. Simultaneous subclonal BRAF and KRAS mutations were demonstrated in two PTC. These data demonstrate that clonal BRAF(V600E) is a rare occurrence in PTC, although frequently this cancer consists of a mixture of tumor cells with wild-type and mutant BRAF. These results suggest that BRAF mutation in PTC is a later subclonal event, its intratumoral heterogeneity may hamper the efficacy of targeted pharmacotherapy, and its association with a more aggressive disease should be reevaluated.

Context: BRAFV600E is considered a negative prognostic marker in papillary thyroid carcinoma (PTC), but unexplained conflicting results are present in the literature. In light of the new finding that most PTC consist of a mixture of tumor cells with wild-type and mutant BRAF, we examined the associations between the percentage of BRAFV600E alleles and both the clinicopathological parameters at time of diagnosis and the disease outcome in a large series of PTCs. Study Design: Tumors from 168 patients with PTC were genotyped for BRAFV600E using BigDye Terminator sequencing and pyrosequencing, and the clinical parameters were analyzed. The associations between clinicopathological characteristics, including disease recurrence at follow-up (median 5.1 yr) and the percentage of mutant BRAF alleles were assessed. Results: The observed prevalence of BRAFV600E was higher when using pyrosequencing then when using BigDye Terminator sequencing (53.6 vs. 36.9%). In the PTC positive for BRAFV600E, the percentage of mutant alleles ranged from 5.1 to 44.7% of the total BRAF alleles, with a median of 20.6%. The presence or the percentage of BRAFV600E alleles did not correlate significantly with sex, multicentricity, lymph node metastasis, or tumor stage. The percentage of BRAFV600E alleles directly correlated with age at diagnosis and tumor volume (R2 = 0.223, P = 0.039, and R2 = 0.166, P < 0.001, respectively). The percentage of BRAFV600E alleles (P = 0.014), tumor volume (P = 0.012), and lymph node metastasis (P = 0.008) predicted the disease outcome. The odds ratio of recurrence for PTC with BRAFV600E alleles of 30% or greater, compared with that for PTC with BRAFV600E alleles of less than 30%, was 5.31 (P = 0.002). Conclusions: A high percentage of BRAFV600E alleles defines a PTC molecular subtype and predicts a poorer disease outcome. The analysis of BRAF mutations by pyrosequencing is useful to refine the risk stratification of patients with PTC.

Purpose: Percutaneous radiofrequency thermal ablation (RTA) was reported as an effective tool for the management of thyroid nodules (TNs). The aim of this study was to investigate the effects of RTA and to establish whether they were treatment-related by comparison with a matched, untreated control group. Patients and Methods: The study population included 40 patients with compressive TNs: 22 had nontoxic TNs, and 18 had toxic TNs and were treated with methimazole. In all patients, a fine-needle aspiration cytology was performed to exclude a thyroid malignancy. Study Design: Twenty patients were treated with RTA (group A), and 20 others did not receive any treatment (group B). At baseline, age, gender, and TN features did not differ significantly between groups. All patients were clinically, biochemically, and morphologically evaluated at baseline and after 1, 3, 6, and 12 months. Results: TN volume significantly decreased in group A (1.8 ± 0.3 ml at 12 months vs. 13.3 ± 1.8 ml at baseline; P < 0.0001) and remained stable in group B [11.7 ± 1.5 ml at 12 months vs. 11.2 ± 1.5 ml at baseline; P = not significant (NS)]. At 3-, 6-, and 12-month evaluations, TN volume was significantly lower in group A than in group B (P < 0.005). At the end of the follow-up, pressure symptoms were improved in all patients in group A but persisted unchanged in group B. In group A, hyperthyroidism completely recovered in 40% and improved in 40% of patients with toxic TNs, whereas it persisted in all patients with toxic TNs in group B. RTA was safe and well tolerated in all patients. Conclusions: RTA induced a marked TN volume shrinkage resulting in parallel improvement of pressure symptoms. In most patients with toxic TNs, hyperthyroidism significantly improved as well. RTA may represent a valid therapeutic approach in patients with TNs not receiving conventional treatments.