Somatic mutation (E17K) that constitutively activates the protein kinase AKT1 has been found in human cancer patients. We determined the role of the E17K mutation of AKT1 in lung cancer, through sequencing of AKT1 exon 4 in 105 resected, clinically annotated non-small cell lung cancer specimens. We detected a missense mutations G->A transition at nucleotide 49 (that results in the E17K substitution) in two squamous cell carcinoma (2/36) but not in adenocarcinoma (0/53). The activity of the endogenous kinase carrying the E17K mutation immunoprecipitated by tumour tissue was significantly higher compared with the wild-type kinase immunoprecipitated by the adjacent normal tissue as determined both by in vitro kinase assay using a consensus peptide as substrate and by in vivo analysis of the phosphorylation status of AKT1 itself (pT308, pS473) or of known downstream substrates such as GSK3 (pS9/S22) and p27 (T198). Immunostaining or immunoblot analysis on membrane-enriched extracts indicated that the enhanced...

Aberrant activation of PI3K/AKT signalling represents one of the most common molecular alterations in lung cancer, though the relative contribution of the single components of the cascade to the NSCLC development is still poorly defined. In this manuscript we have investigated the relationship between expression and genetic alterations of the components of the PI3K/AKT pathway [KRAS, the catalytic subunit of PI3K (p110α), PTEN, AKT1 and AKT2] and the activation of AKT in 107 surgically resected NSCLCs and have analyzed the existing relationships with clinico-pathologic features. Expression analysis was performed by immunohistochemistry on Tissue Micro Arrays (TMA); mutation analysis was performed by DNA sequencing; copy number variation was determined by FISH. We report that activation of PI3K/AKT pathway in Italian NSCLC patients is associated with high grade (G3–G4 compared with G1–G2; n = 83; p<0.05) and more advanced disease (TNM stage III vs. stages I and II; n = 26; p<0.05). In addition, we found that PTEN loss (41/104, 39%) and the overexpression of p110α (27/92, 29%) represent the most frequent aberration observed in NSCLCs. Less frequent molecular lesions comprised the overexpression of AKT2 (18/83, 22%) or AKT1 (17/96, 18%), and KRAS mutation (7/63, 11%). Our results indicate that, among all genes, only p110α overexpression was significantly associated to AKT activation in NSCLCs (p = 0.02). Manipulation of p110α expression in lung cancer cells carrying an active PI3K allele (NCI-H460) efficiently reduced proliferation of NSCLC cells in vitro and tumour growth in vivo. Finally, RNA profiling of lung epithelial cells (BEAS-2B) expressing a mutant allele of PIK3 (E545K) identified a network of transcription factors such as MYC, FOS and HMGA1, not previously recognised to be associated with aberrant PI3K signalling in lung cancer.

Long noncoding RNAs (lncRNAs) represent key regulators of gene transcription during the inflammatory response. Recent findings showed lncRNAs to be dysregulated in human diseases, such as inflammatory bowel disease, diabetes, allergies, asthma, and cancer. These noncoding RNAs are crucial for immune mechanism, as they are involved in differentiation, cell migration and in the production of inflammatory mediators through regulating protein–protein interactions or their ability to assemble with RNA and DNA. The last interaction can occur in cis or trans and is responsible for all the possible lncRNAs biological effects. Our proposal is to provide an overview on lncRNAs roles and functions related to immunity and immune mediated diseases, since these elucidations could be beneficial to untangle the complex bond between them.

Objective: The aim of this study was to investigate the presence of germline mutations in theCDKN1Bgene that encodes the cyclin-dependent kinase (Cdk) inhibitor p27 in multiple endocrine neoplasia 1 (MEN1)-like Spanish index patients. TheCDKN1Bgene has recently been identified as a tumor susceptibility gene for MEN4, with six germline mutations reported so far in patients with a MEN-like phenotype but negative for MEN1 mutations.Design and methods: Fifteen Spanish index cases with MEN-like symptoms were screened for mutations in theCDKN1Bgene and the mutant variant was studied functionally by transcription/translation assaysin vitroand in transiently transfected HeLa cells.Results: We report the identification of a heterozygous GAGA deletion in the 5′-UTR ofCDKN1B, NM_004064.3:c.-32_-29del, in a patient affected by gastric carcinoid tumor and hyperparathyroidism. This deletion falls inside the region that is responsible forCDKN1Btranscription and is predicted to destroy a secondary stem and loop structure that includes the GAGAGA element responsible for ribosome recruitment. Accordingly,in vitrostudies of coupled transcription/translation assays and transient transfection in HeLa cells showed that the GAGA deletion in theCDKN1B5′-UTR significantly impairs the transcription of downstream reporter luciferase (of ∼40–60%) and, possibly, the translation of the corresponding mRNA. This mutation was associated with a significant reduction in the amount ofCDKN1BmRNA in peripheral blood leukocytes from the patient, as demonstrated by quantitative real-time PCR.Conclusions: Our results confirm that germlineCDKN1Bmutations may predispose to a human MEN4 condition and add novel evidence that alteration in the transcription/translation rate ofCDKN1BmRNA might be the mechanism implicated in tumor susceptibility.