Pancreatic endocrine tumours (PETs) are rare and ‘indolent’ neoplasms that usually develop metastatic lesions and exhibit poor response to standard medical treatments. Few studies have investigated pathways responsible for PET cell growth and invasion and no alternative therapeutic strategies have been proposed. In a recent microarray analysis for genes up-regulated in PETs, we have described the up-regulation of soluble Src family tyrosine kinases in this neoplasia, which may represent potentially promising candidates for therapy. Herein, we have investigated the expression and function of Src family kinases in PETS and PET cell lines. Western blot analysis indicated that Src is highly abundant in the PET cell lines CM and QGP-1. Immunohistochemistry and Western blot analyses showed that Src is up-regulated also in human PET lesions. Pharmacological inhibition of Src family kinases by the specific inhibitor PP2 strongly interfered with adhesion, spreading and migration of PET cell lines. Accordingly, the actin cytoskeleton was profoundly altered after inhibition of Src kinases, whereas even prolonged incubation with PP2 exerted no effect on cell cycle progression and/or apoptosis of PET cells. A transient increase in tyrosine phosphorylation of a subset of proteins was observed in QGP-1 cells adhering to the plate, with a peak at 75 min after seeding, when approximately 80% of cells were attached. Inhibition of Src kinases caused a dramatic reduction in the phosphorylation of proteins with different molecular weight that were isolated from the cell extracts by anti-phosphotyrosine immunoprecipitation or pull-down with the SH2 domain of Src. Among them, the docking protein p130Cas interacted with Src and is a major substrate of the Src kinases in QGP-1 cells undergoing adhesion. Our results suggest that Src kinases play a specific role during adhesion, spreading and migration of PET cells and may indicate therapeutical approaches directed to limiting the metastatic potential of these cells.
Alessia Di Florio, Gabriele Capurso, Massimo Milione, Francesco Panzuto, Raffaele Geremia, Gianfranco Delle Fave, and Claudio Sette
Francesco Panzuto, Silvia Nasoni, Massimo Falconi, Vito Domenico Corleto, Gabriele Capurso, Sara Cassetta, Michela Di Fonzo, Valentina Tornatore, Massimo Milione, Stefano Angeletti, Maria Sofia Cattaruzza, Vincenzo Ziparo, Cesare Bordi, Paolo Pederzoli, and Gianfranco Delle Fave
Since gastro-entero-pancreatic endocrine tumors are rare and heterogeneous diseases, their prognosis and long-term survival are not well known. This study aimed at identifying prognostic factors and assessing long-term survival in gastro-entero-pancreatic endocrine tumors. A total of 156 patients enrolled. Prognostic factors were determined by univariate/multivariate analysis; survival rates were assessed by the Kaplan–Meier method. The tumors were non-functioning in 59.6% of patients, and originated from the pancreas in 42.9%. At diagnosis, 64.3% of patients had metastases. The tumors were well differentiated in 89.6% of patients. Ki67 was >2% in 39.6% of patients. Primary tumor size was >3 cm in 49.6% of cases studied. For the univariate analysis, the negative prognostic factors were: pancreatic origin (rate ratio 4.64, P = 0.0002), poorly differentiated tumor (rate ratio 7.70, P = 0.0001), primary tumor size >3 cm (rate ratio 4.26, P = 0.0009), presence of distant metastases (liver: rate ratio 5.88, P = 0.01; distant extra-hepatic: rate ratio 13.41, P = 0.0008). The pancreatic site, the poor degree of differentiation and the distant metastases were confirmed as negative prognostic factors at multivariate analysis. Overall 5-year survival rate was 77.5%. Survival rates differed according to: primary tumor site (62% for pancreatic vs 89.9% for gastrointestinal tract, P = 0.0001) and size (65.7% for >3 cm vs 88.8% for ≤ 3 cm, P = 0.0003), degree of differentiation (22% for poor vs 86.8% for good, P<0.0001), Ki67 (53.5% for > 2% vs 90.1% for ≤ 2%, P = 0.003), metastases (96.1, 77, 73.3 and 50.1% for absent, local, liver and distant extra-hepatic metastases respectively), age at diagnosis (85.3% for ≤ 50 years vs 70.3% for > 50 years, P = 0.03). Although 64.3% of gastro-entero-pancreatic endocrine tumors present metastases at diagnosis, the 5-year survival rate is 77.5%. Pancreatic site, a poor degree of tumor cell differentiation and distant extra-hepatic metastases are the major negative prognostic factors.
Sara Pusceddu, Antonio Facciorusso, Luca Giacomelli, Natalie Prinzi, Francesca Corti, Monica Niger, Massimo Milione, Jorgelina Coppa, Tommaso Cascella, Iolanda Pulice, Lavinia Biamonte, Simonetta Papa, Maria Di Bartolomeo, Aashni Shah, Rodolfo Sacco, and Filipppo de Braud
Although combination therapy is not recommended in patients with gastro-entero-pancreatic (GEP) neuroendocrine tumors (NETs), this strategy is widely used in clinical practice. This network meta-analysis of randomized trials evaluates targeted therapies and somatostatin analogues in GEP advanced NETs, either alone or in combination, comparing the efficacy of different single or combined treatment strategies in terms of progression-free survival (PFS). Interventions were grouped as analogues, everolimus, everolimus plus SSAs, sunitinib and placebo. In a secondary analysis, we also assessed the efficacy of individual specific pharmacological treatments versus placebo or each other. From 83 studies identified, 8 randomized controlled trials were selected, with a total of 1849 patients with either functioning or non-functioning NETs. The analysis confirmed the superiority of all treatments over placebo (HR ranging from 0.34, 95% CI: 0.24–0.37 with the combination of everolimus plus SSAs to 0.42, 0.31–0.57 with the analogues; moderate quality of evidence). On ranking analysis, the combination of everolimus plus SSA (P score=0.86) and then everolimus alone (P score=0.65) ranked highest in increasing PFS. On comparative evaluation of different interventions, pasireotide (P score=0.96) and everolimus+octreotide (P score=0.82) ranked as the best pharmacological treatment options. Our findings support the use of combination therapy in the treatment of functioning and non-functioning GEP NETs. The role of pasireotide should be explored in selected subgroups of patients. Last, the combination of everolimus and octreotide appears promising and should be more widely considered in clinical practice.
Massimo Milione, Patrick Maisonneuve, Alessio Pellegrinelli, Federica Grillo, Luca Albarello, Paola Spaggiari, Alessandro Vanoli, Giovanna Tagliabue, Eleonora Pisa, Luca Messerini, Giovanni Centonze, Frediano Inzani, Aldo Scarpa, Mauro Papotti, Marco Volante, Fausto Sessa, Nicola Fazio, Giancarlo Pruneri, Guido Rindi, Enrico Solcia, Stefano La Rosa, and Carlo Capella
Mixed adenoneuroendocrine carcinomas (MANECs) are composed of a poorly differentiated neuroendocrine carcinoma (NEC) and a non-neuroendocrine (non-NEC) neoplastic epithelial component, each representing at least 30% of the tumor. At present, prognostic factors for MANECs remain largely unexplored. We investigated the clinical-pathologic features of a large multicenter series of digestive system MANECs. Surgical specimens of 200 MANEC candidates were centrally reviewed; diagnosis was confirmed in 160 cases. While morphology, proliferation (mitotic count (MC), Ki67 index) and immunophenotype (p53, SSTR2a, beta-Catenin, Bcl-2, p16, Rb1, ALDH, mismatch repair proteins and CD117) were investigated separately in both components, genomic (TP53, KRAS, BRAF) alterations were searched for on the entire tumor. Data were correlated with overall survival (OS). MANEC sites were: 92 colorectal, 44 gastroesophageal and 24 pancreatobiliary. Median OS was 13.2 months. After adjustment for primary site, Ki67 index of the NEC component (but not of the non-NEC component) was the most powerful prognostic marker. At multivariable analysis, patients with Ki67 ≥ 55% had an 8-fold risk of death (hazard ratio (HR) 7.83; 95% confidence interval (CI) 4.17–14.7; P < 0.0001) and a median OS of 12.2 months compared to those with Ki67 < 55% (median OS 40.5 months). MC (HR 1.51; 95% CI 1.03–2.20, P = 0.04) was a weaker prognostic index. Colorectal primary site (HR 1.60; 95% CI 1.11–2.32; P = 0.01) was significantly associated with poorer survival. No single immunomarker, in either component, was statistically significant. This retrospective analysis of a large series of digestive system MANECs, showed that the NEC component, particularly its Ki67 index, was the main prognostic driver.
Sara Pusceddu, Francesco Barretta, Annalisa Trama, Laura Botta, Massimo Milione, Roberto Buzzoni, Filippo De Braud, Vincenzo Mazzaferro, Ugo Pastorino, Ettore Seregni, Luigi Mariani, Gemma Gatta, Maria Di Bartolomeo, Daniela Femia, Natalie Prinzi, Jorgelina Coppa, Francesco Panzuto, Lorenzo Antonuzzo, Emilio Bajetta, Maria Pia Brizzi, Davide Campana, Laura Catena, Harry Comber, Fiona Dwane, Nicola Fazio, Antongiulio Faggiano, Dario Giuffrida, Kris Henau, Toni Ibrahim, Riccardo Marconcini, Sara Massironi, Maja Primic Žakelj, Francesca Spada, Salvatore Tafuto, Elizabeth Van Eycken, Jan Maaten Van der Zwan, Tina Žagar, Luca Giacomelli, Rosalba Miceli, and NEPscore Working Group
No validated prognostic tool is available for predicting overall survival (OS) of patients with well-differentiated neuroendocrine tumors (WDNETs). This study, conducted in three independent cohorts of patients from five different European countries, aimed to develop and validate a classification prognostic score for OS in patients with stage IV WDNETs. We retrospectively collected data on 1387 patients: (i) patients treated at the Istituto Nazionale Tumori (Milan, Italy; n = 515); (ii) European cohort of rare NET patients included in the European RARECAREnet database (n = 457); (iii) Italian multicentric cohort of pancreatic NET (pNETs) patients treated at 24 Italian institutions (n = 415). The score was developed using data from patients included in cohort (i) (training set); external validation was performed by applying the score to the data of the two independent cohorts (ii) and (iii) evaluating both calibration and discriminative ability (Harrell C statistic). We used data on age, primary tumor site, metastasis (synchronous vs metachronous), Ki-67, functional status and primary surgery to build the score, which was developed for classifying patients into three groups with differential 10-year OS: (I) favorable risk group: 10-year OS ≥70%; (II) intermediate risk group: 30% ≤ 10-year OS < 70%; (III) poor risk group: 10-year OS <30%. The Harrell C statistic was 0.661 in the training set, and 0.626 and 0.601 in the RARECAREnet and Italian multicentric validation sets, respectively. In conclusion, based on the analysis of three ‘field-practice’ cohorts collected in different settings, we defined and validated a prognostic score to classify patients into three groups with different long-term prognoses.