Fulvio Basolo

KRAS codon 61, 146 and BRAF mutations predict resistance to cetuximab plus irinotecan in KRAS codon 12 and 13 wild-type metastatic colorectal cancer

Background:KRAS codons 12 and 13 mutations predict resistance to anti-EGFR monoclonal antibodies (moAbs) in metastatic colorectal cancer. Also, BRAF V600E mutation has been associated with resistance. Additional KRAS mutations are described in CRC.Methods:We investigated the role of KRAS codons 61 and 146 and BRAF V600E mutations in predicting resistance to cetuximab plus irinotecan in a cohort of KRAS codons 12 and 13 wild-type patients.Results:Among 87 KRAS codons 12 and 13 wild-type patients, KRAS codons 61 and 146 were mutated in 7 and 1 case, respectively. None of mutated patients responded vs 22 of 68 wild type (P=0.096). Eleven patients were not evaluable. KRAS mutations were associated with shorter progression-free survival (PFS, HR: 0.46, P=0.028). None of 13 BRAF-mutated patients responded vs 24 of 74 BRAF wild type (P=0.016). BRAF mutation was associated with a trend towards shorter PFS (HR: 0.59, P=0.073). In the subgroup of BRAF wild-type patients, KRAS codons 61/146 mutations determined a lower response rate (0 vs 37%, P=0.047) and worse PFS (HR: 0.45, P=0.023). Patients bearing KRAS or BRAF mutations had poorer response rate (0 vs 37%, P=0.0005) and PFS (HR: 0.51, P=0.006) compared with KRAS and BRAF wild-type patients.Conclusion:Assessing KRAS codons 61/146 and BRAF V600E mutations might help optimising the selection of the candidate patients to receive anti-EGFR moAbs.

BRAF(V600E) mutation and outcome of patients with papillary thyroid carcinoma: a 15-year median follow-up study.

BACKGROUND The BRAF(V600E) mutation is the most frequent genetic alteration in papillary thyroid carcinoma (PTC). The role of BRAF(V600E) mutation as a poor prognostic factor has been controversially reported in series with short-term follow-ups. In this study we verified the prognostic value of the BRAF(V600E) mutation in PTC patients with a long-term follow-up. METHODS We studied 102 PTC patients with a median follow-up of 15 yr. The BRAF(V600E) mutation was analyzed by PCR-single-strand conformational polymorphism and sequencing. The correlation between the presence/absence of the BRAF(V600E) mutation, clinicopathological features, and outcome of PTC patients were evaluated. RESULTS The BRAF(V600E) mutation was found in 38 of 102 (37.3%) PTC patients, and was significantly more frequent in patients older than 60 yr (P = 0.02), in advanced stages (P = 0.03), and in cases with vascular invasion (P = 0.02). At univariate analysis the worst outcome for PTC patients was significantly correlated with clinicopathological features (i.e. age, tumor size, extrathyroid extension, lymph node and distant metastases, advanced stage, vascular endothelial growth factor expression, and vascular invasion) and the BRAF(V600E) mutation (P < 0.002). However, at multivariate analysis only the BRAF(V600E) mutation showed an independent correlation with the worst outcome (P = 0.03). Moreover, the survival curves of PTC patients showed a lower percentage of survivors in the BRAF(V600E)-mutated group (P = 0.015). CONCLUSIONS In this study the BRAF(V600E) mutation correlated with the worst outcome for PTC patients, who were not only at a higher risk not to be cured but also for death. In particular, the BRAF(V600E) mutation was demonstrated to be a poor prognostic factor independent from other clinicopathological features.

The RET/PTC-RAS-BRAF linear signaling cascade mediates the motile and mitogenic phenotype of thyroid cancer cells

In papillary thyroid carcinomas (PTCs), rearrangements of the RET receptor (RET/PTC) and activating mutations in the BRAF or RAS oncogenes are mutually exclusive. Here we show that the 3 proteins function along a linear oncogenic signaling cascade in which RET/PTC induces RAS-dependent BRAF activation and RAS- and BRAF-dependent ERK activation. Adoptive activation of the RET/PTC-RAS-BRAF axis induced cell proliferation and Matrigel invasion of thyroid follicular cells. Gene expression profiling revealed that the 3 oncogenes activate a common transcriptional program in thyroid cells that includes upregulation of the CXCL1 and CXCL10 chemokines, which in turn stimulate proliferation and invasion. Thus, motile and mitogenic properties are intrinsic to transformed thyroid cells and are governed by an epistatic oncogenic signaling cascade.

Bcl-2 protein: a prognostic factor inversely correlated to p53 in non-small-cell lung cancer.

Non-small-cell lung cancer (NSCLC) prognosis is strictly related to well-established clinicopathological parameters which have unfortunately become insufficient in the prognostic evaluation of this type of cancer. As p53 and bcl-2 gene deregulations are frequently involved in several types of epithelial malignancies, we investigated the Bcl-2 and p53 protein expression in 91 and 101 cases of NSCLC respectively. The expression was then compared with established indicators of prognosis and biological behaviour of the tumours. No relationship was observed between Bcl-2 and either clinicopathological or biological parameters such as histology, grading, tumour status, nodal metastasis and proliferative activity evaluated by scoring proliferating cell nuclear antigen expression and Ki-67 immunoreactivity. However, the mean Bcl-2 expression was significantly lower in patients who developed metastasis during follow-up or died of metastatic disease (P = 0.006 and P = 0.01 respectively). Moreover, survival probability was higher in patients who expressed the Bcl-2 protein (P = 0.0002). In contrast with this, p53 protein accumulation was observed in tumours with metastatic nodal involvement (P = 0.02) or in patients who developed metastasis during follow-up (P = 0.01), although no correlation was found between p53 expression and overall survival. An inverse relationship was also found between Bcl-2 and the anti-oncogene protein product p53 (P = 0.01). Thus, a high proportion of NSCLCs express p53 and Bcl-2 proteins and their expression may have prognostic importance.

Are the Clinical and Pathological Features of Differentiated Thyroid Carcinoma Really Changed over the Last 35 Years? Study on 4187 Patients from a Single Italian Institution to Answer this Question

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.

Lower levels of TSH are associated with a lower risk of papillary thyroid cancer in patients with thyroid nodular disease: thyroid autonomy may play a protective role.

Higher TSH values, even within normal ranges, have been associated with a greater risk of thyroid malignancy. The relationship between TSH and papillary thyroid cancer (PTC) has been analyzed in 10 178 patients submitted to fine needle aspiration of thyroid nodules with a cytology of PTC (n=497) or benign thyroid nodular disease (BTND, n=9681). In 942 patients, submitted to surgery (521 from BTND and 421 from PTC), the histological diagnosis confirmed an elevated specificity (99.6%) and sensitivity (98.1%) of cytology. TSH levels were significantly higher in PTC than in BTND both in the cytological and histological series and also in patients with a clinical diagnosis of multinodular goiter (MNG) and single/isolate nodule (S/I). A significant age-dependent development of thyroid autonomy (TSH <0.4 microU/ml) was observed in patients with benign thyroid disease, but not in those with PTC, diagnosed both on cytology and histology. In patients with MNG, the frequency of thyroid autonomy was higher and the risk of PTC was lower compared to those with S/I. In all patients, the presence of thyroid auto-antibodies (TAb) was associated with a significant increase of TSH. However, both in TAb positive and TAb negative patients TSH levels were significantly higher in PTC than in BTND. Our data confirm a direct relationship between TSH levels and risk of PTC in patients with nodular thyroid diseases. Thyroid autonomy conceivably protects against the risk of PTC, while thyroid autoimmunity does not play a significant role.

Real-Time Elastosonography: Useful Tool for Refining the Presurgical Diagnosis in Thyroid Nodules with Indeterminate or Nondiagnostic Cytology

BACKGROUND Indeterminate and nondiagnostic patterns represent the main limitation of fine-needle aspiration (FNA) cytology of thyroid nodules, clinical and echographic features being poorly predictive of malignancy. The newly developed real-time ultrasound elastography (USE) has been previously applied to differentiate malignant from benign lesions. The aim of this study was to get further insights into the role of USE in the presurgical diagnosis of nodules with indeterminate or nondiagnostic cytology. PATIENTS The study included 176 patients who had one (n=138) or multiple (n=38) nodules with indeterminate or nondiagnostic cytology on FNA, for whom histology was available after thyroidectomy. A total of 195 nodules (142 indeterminate, 53 nondiagnostic) were submitted to USE, and elasticity was scored as 1 (high), 2 (intermediate), or 3 (low). RESULTS In indeterminate lesions, the score 1, describing high elasticity, was strongly predictive of benignity, being found in 102 of 111 benign nodules and in only one of 31 carcinomas (P<0.0001). By combining the scores 2 and 3, USE had a sensitivity of 96.8% and a specificity of 91.8%. In nodules with nondiagnostic cytology, score 1 was found in 39 of 45 benign nodules and in only one of eight carcinomas (P<0.0001). By combining the scores 2 and 3, USE had a sensitivity of 87.5% and a specificity of 86.7%. CONCLUSIONS USE may represent an important tool for the diagnosis of thyroid cancer in nodules with indeterminate or nondiagnostic cytology and may prove useful in selecting patients who are candidates for surgery.

Correlation between the BRAF V600E Mutation and Tumor Invasiveness in Papillary Thyroid Carcinomas Smaller than 20 Millimeters: Analysis of 1060 Cases

CONTEXT Evaluation of the degree of neoplastic infiltration beyond the thyroid capsule remains a unique parameter that can be evaluated by histopathological examination to label a papillary thyroid carcinoma (PTC) of 20 mm or less in size as a pT1 or pT3 tumor. OBJECTIVE We correlated the BRAF V600E mutation with both clinical-pathological features and the degree of neoplastic infiltration to redefine the reliability of the actual system of risk stratification in a large selected group of PTCs smaller than 20 mm. DESIGN The presence of BRAF mutations was examined in 1060 PTCs less than 20 mm divided into four degrees of neoplastic infiltration: 1) totally encapsulated; 2) not encapsulated without thyroid capsule invasion; 3) thyroid capsule invasion; and 4) extrathyroidal extension. RESULTS The overall frequency of the BRAF V600E mutation was 44.6%. In both univariate and multivariate analyses, BRAF mutations showed a strong association with PTC variants (classical and tall cell), tumor size (11-20 mm), multifocality, absence of tumor capsule, extrathyroidal extension, lymph node metastasis, higher American Joint Commission on Cancer stage, and younger patient age. In PTCs staged as pT1 with thyroid capsule invasion, the frequency of BRAF mutations was significantly higher than in pT1 tumors that did not invade the thyroid capsule (67.3 vs. 31.8%, respectively; P < 0.0001). No statistically significant difference in BRAF alterations was found between pT1 tumors with thyroid capsule invasion and pT3 tumors (67.3 and 67.5%, respectively). CONCLUSION We suggest that evaluation of BRAF status would be useful even in pT1 tumors to improve risk stratification and patient management, although follow-up data are necessary to confirm our speculations.

The BRAF(V600E) mutation is an independent, poor prognostic factor for the outcome of patients with low-risk intrathyroid papillary thyroid carcinoma: single-institution results from a large cohort study

BACKGROUND The BRAF(V600E) mutation, the most frequent genetic alteration in papillary thyroid carcinoma (PTC), was demonstrated to be a poor prognostic factor. The aim of this study was to evaluate its prognostic significance in a large cohort of low-risk intrathyroid PTC. METHODS Among the 431 consecutive PTC patients, we selected 319 patients with an intrathyroid tumor and no metastases (T1-T2, N0, M0). The BRAF(V600E) mutation was analyzed by PCR-single-strand conformation polymorphism analysis and direct genomic sequencing. The correlation between the presence/absence of the mutation, the clinical-pathological features, and the outcome of the PTC patients was investigated. RESULTS The BRAF(V600E) mutation was present in 106 of 319 PTC patients (33.2%). Its prevalence was also the same in subgroups identified according to the level of risk. The BRAF(V600E) mutation correlated with multifocality, aggressive variant, absence, or infiltration of the tumoral capsule. BRAF(V600E)-mutated PTC also required a higher number of radioiodine courses to obtain disease-free status. The BRAF(V600E) mutation was the only prognostic factor predicting the persistence of the disease in these patients after 5 yr of follow-up. CONCLUSIONS The BRAF(V600E) mutation was demonstrated to be a poor prognostic factor for the persistence of the disease independent from other clinical-pathological features in low-risk intrathyroid PTC patients. It could be useful to search for the BRAF(V600E) mutation in the workup of low-risk PTC patients to distinguish those who require less or more aggressive treatments. In particular, the high negative predictive value of the BRAF(V600E) mutation could be useful to identify, among low-risk PTC patients, those who could avoid 131-I treatment.

Neoangiogenesis and p53 protein in lung cancer: their prognostic role and their relation with vascular endothelial growth factor (VEGF) expression.

Following up-regulation of an angiogenesis inhibitor by the wild-type p53 protein proven recently, we have analysed on the one hand the prognostic impact of microvessel count (MC) and p53 protein overexpression in non-small-cell lung carcinoma (NSCLC) progression and, on the other hand, the inter-relation between the microvascular pattern and the p53 protein expression. Moreover, we assessed the expression of vascular endothelial growth factor (VEGF), one of the pivotal mediators of tumour angiogenesis, in order to investigate its relation to p53 protein expression and MC. Tumours from 73 patients resected for NSCLC between March 1991 and April 1992 (median follow-up 47 months, range 32-51 months) were analysed using an immunohistochemical method. In univariate analysis, MC and p53 accumulation were shown to affect metastatic nodal involvement, recurrence and death significantly. Multiple logistic regression analysis showed an important prognostic influence of MC and nodal status on overall (P = 0.0009; P = 0.01) and disease-free survival (P = 0.0001; P = 0.03). Interestingly, a strong statistical association was observed between p53 nuclear accumulation and MC (P = 0.0003). The same inter-relationship was found in non-squamous histotype (P = 0.002). When we analysed the concomitant influence of MC and p53 expression on overall survival, we were able to confirm a real predominant role of MC in comparison with p53. With regard to VEGF expression, p53-negative and lowly vascularized tumours showed a mean VEGF expression significantly lower than p53-positive and highly vascularized cancers (P = 0.02). These results underline the prognostic impact of MC and p53 protein accumulation in NSCLC and their reciprocal inter-relationship, supporting the hypothesis of a wild-type p53 regulation on the angiogenetic process through a VEGF up-regulation.