Bronchial carcinoids (BCs) are rare neuroendocrine tumors that are still orphans of medical treatment. Human BC primary cultures may display resistance to everolimus, an inhibitor of the mammalian target of rapamycin (mTOR), in terms of cell viability reduction. Our aim was to assess whether the novel dual phosphatidylinositol 3-kinase (PI3K)/mTOR inhibitor NVP-BEZ235 is effective in everolimus-resistant human BC tissues and cell lines. In addition, we searched for possible markers of the efficacy of mTOR inhibitors that may help in identifying the patients who may benefit from treatment with mTOR inhibitors, sparing them from ineffective therapy. We found that NVP-BEZ235 is twice as potent as everolimus in reducing cell viability and activating apoptosis in human BC tissues that display sensitivity to mTOR inhibitors, but is not effective in everolimus-resistant BC tissues and cell lines that bypass cyclin D1 downregulation and escape G0/G1 blockade. Rebound AKT activation was not observed in response to treatment with either mTOR inhibitor in the 'resistant' BC cells. In addition to total mTOR levels, putative markers of the sensitivity of BCs to mTOR inhibitors are represented by AKT, p70S6K (RPS6KB2), and ERK1/2 (MAPK3/1) protein levels. Finally, we validated these markers in an independent BC group. These data indicate that the dual PI3K/mTOR inhibitor NVP-BEZ235 is more potent than everolimus in reducing the proliferation of human BC cells. 'Resistant' cells display lower levels of mTOR, p70S6K, AKT, and ERK1/2, indicating that these proteins may be useful as predictive markers of resistance to mTOR and PI3K/mTOR inhibitors in human BCs.

Genetic alterations frequently are involved in the development of a pituitary adenoma in young age. We here characterize the functional role of a deletion in CDKN1B 5'-UTR region (c.-29_-26delAGAG) identified in an acromegalic patient that developed a growth hormone in pituitary adenoma during childhood. Our results show that the identified novel heterozygous deletion in the CDKN1B 5'-UTR region associates with a reduction in CDKN1B mRNA levels, a predicted altered secondary mRNA structure, and a reduced CDKN1B 5'-UTR transcriptional activity in vitro. The patient displayed loss of heterozygosity in the same CDKN1B 5'-UTR region at tissue level and the 5'UTR region containing the deleted sequence encompasses a GRE. These findings indicate that the identification of functional alterations of newly discovered genetic derangements need to be fully characterized and always correlated with the clinical manifestations.

Background: Bronchial carcinoids (BCs) are rare neuroendocrine tumors that are still orphan of medical treatment. Human BC primary cultures may display resistance to everolimus, an inhibitor of the mammalian target of rapamycin (mTOR), in terms of cell viability reduction.\ud \ud Aim: To assess whether the novel dual PI3K/mTOR inhibitor, NVP-BEZ235, may be effective in everolimus-resistant human BC tissues and cell lines. In addition, we search for possible markers of mTOR inhibitors efficacy, that may help in identifying the patients that may benefit from mTOR inhibitors treatment, sparing them from ineffective therapy.\ud \ud Results: NVP-BEZ235 is twice as potent as everolimus in reducing cell viability and activating apoptosis in human BC tissues that display sensitivity to mTOR inhibitors, but is not effective in everolimus-resistant BC tissues and cell lines, that by-pass cyclin D1 down-regulation and escape G0/G1 blockade. Rebound AKT activation was not observed in response to treatment with either mTOR inhibitor in ‘resistant’ BC cells. We also show that, in addition to total mTOR levels, putative markers of BC sensitivity to mTOR inhibitors are represented by higher AKT, p70S6K and ERK1/2 protein levels.\ud \ud Conclusion: These data indicate that the dual PI3K/mTOR inhibitor NVP-BEZ235 is more potent than everolimus in reducing human BC cell proliferation. ‘Resistant’ cells display lower levels of mTOR, AKT, p70S6K and ERK1/2, indicating that these proteins may be useful as predictive markers of resistance to mTOR and PI3K/mTOR inhibitors in human BC

Our data confirm that mTOR inhibitors may be useful to reduce pituitary adenoma cell viability, while Erk 1/2 pathway may be considered as a useful therapeutic target to control GH secretion. Our results open the field for further studies searching for effective drugs to control GH hyper-secretion.

Bronchial carcinoids (BC) are rare tumors originating from endocrine cells dispersed in the respiratory epithelium. Currently, the main BC treatment is surgery, that can be curative in most of the cases, but is not feasible for large, infiltrating and metastatic disease. In these settings, medical therapy is often tried, being mainly represented by chemotherapy and radiation in the attempt to reduce tumor mass, while somatostatin analogues are employed for symptomatic control. Therefore it is important to identify new therapeutic targets and new molecules capable of providing adequate medical treatment for patients with BC for which surgical removal is not feasible. Growth factors which are important in experimental models of neuroendocrine tumors include epidermal growth factor (EGF), transforming growth factor (TGF) α, TGFβ. EGF and TGFα bind to the EGF receptor to stimulate the PI3K/RAS/RAF/MAPK pathway, leading to the transcription of genes associated with cell proliferation, invasion and metastasis. Our aim is to evaluate the effects of Sunitinib, a multi-targeted receptor tyrosine kinase (RTK) inhibitor, and NVP-BEZ235, a PI3K/mTOR inhibitor, on human primary BC cells cultures in order to verify the involvement of the EGF pathway in regulating crucial cellular processes. Human BC primary cultures were treated with Sunitinib or NVP-BEZ235, alone or in combination with EGF. EGFR expression, cell viability and caspase 3/7 activation were evaluated. By immunofluorescences we found that EGFR is expressed in all primary cultures. In addition, 100 nM NVP-BEZ235 and 10 μM Sunitinib inhibit cell viability by 30 and 20% (P<0.01), respectively. Both NVP-BEZ 235 and Sunitinib promote apoptosis (100%). 100 ng/ml EGF impairs the antiproliferative and pro-apoptotic effects of both Sunitinib and NVP-BEZ 235. These data suggest a possible role for EGFR pathway as molecular target in the medical treatment of BC. Further studies are necessary to understand the molecular basis of this mechanism