Non-functional pancreatic neuroendocrine tumours: ATRX/DAXX and alternative lengthening of telomeres (ALT) are prognostically independent from ARX/PDX1 expression and tumour size.

OBJECTIVE: Recent studies have found aristaless-related homeobox gene (ARX)/pancreatic and duodenal homeobox 1 (PDX1), alpha-thalassemia/mental retardation X-linked (ATRX)/death domain-associated protein (DAXX) and alternative lengthening of telomeres (ALT) to be promising prognostic biomarkers for non-functional pancreatic neuroendocrine tumours (NF-PanNETs). However, they have not been comprehensively evaluated, especially among small NF-PanNETs (≤2.0 cm). Moreover, their status in neuroendocrine tumours (NETs) from other sites remains unknown. DESIGN: An international cohort of 1322 NETs was evaluated by immunolabelling for ARX/PDX1 and ATRX/DAXX, and telomere-specific fluorescence in situ hybridisation for ALT. This cohort included 561 primary NF-PanNETs, 107 NF-PanNET metastases and 654 primary, non-pancreatic non-functional NETs and NET metastases. The results were correlated with numerous clinicopathological features including relapse-free survival (RFS). RESULTS: ATRX/DAXX loss and ALT were associated with several adverse prognostic findings and distant metastasis/recurrence (p<0.001). The 5-year RFS rates for patients with ATRX/DAXX-negative and ALT-positive NF-PanNETs were 40% and 42% as compared with 85% and 86% for wild-type NF-PanNETs (p<0.001 and p<0.001). Shorter 5-year RFS rates for ≤2.0 cm NF-PanNETs patients were also seen with ATRX/DAXX loss (65% vs 92%, p=0.003) and ALT (60% vs 93%, p<0.001). By multivariate analysis, ATRX/DAXX and ALT status were independent prognostic factors for RFS. Conversely, classifying NF-PanNETs by ARX/PDX1 expression did not independently correlate with RFS. Except for 4% of pulmonary carcinoids, ATRX/DAXX loss and ALT were only identified in primary (25% and 29%) and NF-PanNET metastases (62% and 71%). CONCLUSIONS: ATRX/DAXX and ALT should be considered in the prognostic evaluation of NF-PanNETs including ≤2.0 cm tumours, and are highly specific for pancreatic origin among NET metastases of unknown primary.

Molecular characterization of extrahepatic cholangiocarcinoma: perihilar and distal tumors display divergent genomic and transcriptomic profiles.

BACKGROUND: Extrahepatic cholangiocarcinoma (ECC) is classified into two subtypes based on anatomic origin: distal extrahepatic (DECC) and perihilar (PHCC) cholangiocarcinoma. This study aimed to shed light on its genomic and transcriptomic profiles. RESEARCH DESIGN AND METHODS: The genomic alterations of 99 ECC (47 PHCC and 52 DECC) were investigated by next-generation sequencing of 96 genes. A subgroup of cases, representative of each subtype, was further investigated using transcriptomic analysis. Bioinformatics tools were applied for clustering and pathway analysis and defining the immune composition of the tumor microenvironment. RESULTS: PHCC had more frequent KRAS mutations (p = 0.0047), whereas TP53 mutations were more common in DECC (p = 0.006). Potentially actionable alterations included high-tumor mutational burden and/or microsatellite instability (7.1%), PI3KCA mutations (8.1%), and MYC (10.1%) and ERBB2 amplification (5.1%). The transcriptomic profiles showed the presence of three distinct clusters, which followed the anatomic origin and differed in immune microenvironment. DECC appeared to contain two distinct tumor subgroups, one enriched for druggable alterations and one lacking actionable opportunities. CONCLUSIONS: This study provides new insights into the molecular landscape and the actionable alterations of ECC. Our findings represent a step toward improved ECC molecular taxonomy and therapeutic strategies for precision oncology.

Alternative Lengthening of Telomeres (ALT) in Pancreatic Neuroendocrine Tumors: Ready for Prime-Time in Clinical Practice?

PURPOSE OF REVIEW: Alternative lengthening of telomeres (ALT) is a telomerase-independent mechanism used by some types of malignancies, including pancreatic neuroendocrine tumors, to overcome the issue of telomere shortening, thus supporting tumor growth and cell proliferation. This review is focused on the most important achievements and opportunities deriving from ALT assessment in PanNET onco-pathology, highlighting the most promising fields in which such biomarker could be implemented in clinical practice. RECENT FINDINGS: In pancreatic neuroendocrine tumors (PanNET), ALT is strongly correlated with the mutational status of two chromatin remodeling genes, DAXX and ATRX. Recent advances in tumor biology permitted to uncover important roles of ALT in the landscape of PanNET, potentially relevant for introducing this biomarker into clinical practice. Indeed, ALT emerged as a reliable indicator of worse prognosis for PanNET, helping in clinical stratification and identification of "high-risk" patients. Furthermore, it is a very specific marker supporting the pancreatic origin of neuroendocrine neoplasms and can be used for improving the diagnostic workflow of patients presenting with neuroendocrine metastasis from unknown primary. The activation of this process can be determined by specific FISH analysis. ALT should be introduced in clinical practice for identifying "high-risk" PanNET patients and improving their clinical management, and as a marker of pancreatic origin among neuroendocrine tumors.

Immune landscape, evolution, hypoxia-mediated viral mimicry pathways and therapeutic potential in molecular subtypes of pancreatic neuroendocrine tumours.

OBJECTIVE: A comprehensive analysis of the immune landscape of pancreatic neuroendocrine tumours (PanNETs) was performed according to clinicopathological parameters and previously defined molecular subtypes to identify potential therapeutic vulnerabilities in this disease. DESIGN: Differential expression analysis of 600 immune-related genes was performed on 207 PanNET samples, comprising a training cohort (n=72) and two validation cohorts (n=135) from multiple transcriptome profiling platforms. Different immune-related and subtype-related phenotypes, cell types and pathways were investigated using different in silico methods and were further validated using spatial multiplex immunofluorescence. RESULTS: The study identified an immune signature of 132 genes segregating PanNETs (n=207) according to four previously defined molecular subtypes: metastasis-like primary (MLP)-1 and MLP-2, insulinoma-like and intermediate. The MLP-1 subtype (26%-31% samples across three cohorts) was strongly associated with elevated levels of immune-related genes, poor prognosis and a cascade of tumour evolutionary events: larger hypoxic and necroptotic tumours leading to increased damage-associated molecular patterns (viral mimicry), stimulator of interferon gene pathway, T cell-inflamed genes, immune checkpoint targets, and T cell-mediated and M1 macrophage-mediated immune escape mechanisms. Multiplex spatial profiling validated significantly increased macrophages in the MLP-1 subtype. CONCLUSION: This study provides novel data on the immune microenvironment of PanNETs and identifies MLP-1 subtype as an immune-high phenotype featuring a broad and robust activation of immune-related genes. This study, with further refinement, paves the way for future precision immunotherapy studies in PanNETs to potentially select a subset of MLP-1 patients who may be more likely to respond.

Corrigendum: Whole-genome landscape of pancreatic neuroendocrine tumours.

This corrects the article DOI: 10.1038/nature21063.

Whole-genome landscape of pancreatic neuroendocrine tumours.

The diagnosis of pancreatic neuroendocrine tumours (PanNETs) is increasing owing to more sensitive detection methods, and this increase is creating challenges for clinical management. We performed whole-genome sequencing of 102 primary PanNETs and defined the genomic events that characterize their pathogenesis. Here we describe the mutational signatures they harbour, including a deficiency in G:C > T:A base excision repair due to inactivation of MUTYH, which encodes a DNA glycosylase. Clinically sporadic PanNETs contain a larger-than-expected proportion of germline mutations, including previously unreported mutations in the DNA repair genes MUTYH, CHEK2 and BRCA2. Together with mutations in MEN1 and VHL, these mutations occur in 17% of patients. Somatic mutations, including point mutations and gene fusions, were commonly found in genes involved in four main pathways: chromatin remodelling, DNA damage repair, activation of mTOR signalling (including previously undescribed EWSR1 gene fusions), and telomere maintenance. In addition, our gene expression analyses identified a subgroup of tumours associated with hypoxia and HIF signalling.

A Cross-Species Analysis in Pancreatic Neuroendocrine Tumors Reveals Molecular Subtypes with Distinctive Clinical, Metastatic, Developmental, and Metabolic Characteristics.

UNLABELLED: Seeking to assess the representative and instructive value of an engineered mouse model of pancreatic neuroendocrine tumors (PanNET) for its cognate human cancer, we profiled and compared mRNA and miRNA transcriptomes of tumors from both. Mouse PanNET tumors could be classified into two distinctive subtypes, well-differentiated islet/insulinoma tumors (IT) and poorly differentiated tumors associated with liver metastases, dubbed metastasis-like primary (MLP). Human PanNETs were independently classified into these same two subtypes, along with a third, specific gene mutation-enriched subtype. The MLP subtypes in human and mouse were similar to liver metastases in terms of miRNA and mRNA transcriptome profiles and signature genes. The human/mouse MLP subtypes also similarly expressed genes known to regulate early pancreas development, whereas the IT subtypes expressed genes characteristic of mature islet cells, suggesting different tumorigenesis pathways. In addition, these subtypes exhibit distinct metabolic profiles marked by differential pyruvate metabolism, substantiating the significance of their separate identities. SIGNIFICANCE: This study involves a comprehensive cross-species integrated analysis of multi-omics profiles and histology to stratify PanNETs into subtypes with distinctive characteristics. We provide support for the RIP1-TAG2 mouse model as representative of its cognate human cancer with prospects to better understand PanNET heterogeneity and consider future applications of personalized cancer therapy.

Mixed adenoneuroendocrine carcinomas of the gastrointestinal tract: targeted next-generation sequencing suggests a monoclonal origin of the two components.

BACKGROUND: Mixed adenoneuroendocrine carcinomas (MANECs) of the gastrointestinal tract are rare neoplasms characterized by coexisting exocrine and neuroendocrine neoplastic components. MANECs' histogenetic classification and molecular characterization remain unclear, significantly affecting the identification of innovative therapeutic options for these tumors. METHODS: The exocrine and neuroendocrine components of 6 gastrointestinal MANECs were microdissected and subjected to the simultaneous mutation assessment in selected regions of 54 cancer-associated genes using Ion Torrent semiconductor-based next-generation sequencing. Sanger sequencing and immunohistochemistry were used as validation of the mutational status. RESULTS: A total of 20 driver gene somatic mutations were observed among the 12 neoplastic components investigated. In 11 of 12 (91.7%) samples, at least one mutation was detected; 7 samples (58.3%) were found to have multiple mutations. TP53 gene mutations were the most frequent genetic alterations observed in the series, occurring in 11/12 samples (91.7%). Somatic mutations in other genes were detected at lower frequencies: ATM, CTNNB1, ERBB4, JAK3, KDR, KRAS, RB1. CONCLUSIONS: Five of the 6 MANECs presented an overlapping mutational profile in both components, suggesting a monoclonal origin of the two MANEC components.

Reporting tumor molecular heterogeneity in histopathological diagnosis.

BACKGROUND: Detection of molecular tumor heterogeneity has become of paramount importance with the advent of targeted therapies. Analysis for detection should be comprehensive, timely and based on routinely available tumor samples. AIM: To evaluate the diagnostic potential of targeted multigene next-generation sequencing (TM-NGS) in characterizing gastrointestinal cancer molecular heterogeneity. METHODS: 35 gastrointestinal tract tumors, five of each intestinal type gastric carcinomas, pancreatic ductal adenocarcinomas, pancreatic intraductal papillary mucinous neoplasms, ampulla of Vater carcinomas, hepatocellular carcinomas, cholangiocarcinomas, pancreatic solid pseudopapillary tumors were assessed for mutations in 46 cancer-associated genes, using Ion Torrent semiconductor-based TM-NGS. One ampulla of Vater carcinoma cell line and one hepatic carcinosarcoma served to assess assay sensitivity. TP53, PIK3CA, KRAS, and BRAF mutations were validated by conventional Sanger sequencing. RESULTS: TM-NGS yielded overlapping results on matched fresh-frozen and formalin-fixed paraffin-embedded (FFPE) tissues, with a mutation detection limit of 1% for fresh-frozen high molecular weight DNA and 2% for FFPE partially degraded DNA. At least one somatic mutation was observed in all tumors tested; multiple alterations were detected in 20/35 (57%) tumors. Seven cancers displayed significant differences in allelic frequencies for distinct mutations, indicating the presence of intratumor molecular heterogeneity; this was confirmed on selected samples by immunohistochemistry of p53 and Smad4, showing concordance with mutational analysis. CONCLUSIONS: TM-NGS is able to detect and quantitate multiple gene alterations from limited amounts of DNA, moving one step closer to a next-generation histopathologic diagnosis that integrates morphologic, immunophenotypic, and multigene mutational analysis on routinely processed tissues, essential for personalized cancer therapy.

Autoantibodies to Ezrin are an early sign of pancreatic cancer in humans and in genetically engineered mouse models.

BACKGROUND: Pancreatic Ductal Adenocarcinoma (PDAC) is a highly aggressive malignancy with only a 5% 5-year survival rate. Reliable biomarkers for early detection are still lacking. The goals of this study were (a) to identify early humoral responses in genetically engineered mice (GEM) spontaneously developing PDAC; and (b) to test their diagnostic/predictive value in newly diagnosed PDAC patients and in prediagnostic sera. METHODS AND RESULTS: The serum reactivity of GEM from inception to invasive cancer, and in resectable or advanced human PDAC was tested by two-dimensional electrophoresis Western blot against proteins from murine and human PDAC cell lines, respectively. A common mouse-to-human autoantibody signature, directed against six antigens identified by MALDI-TOF mass spectrometry, was determined. Of the six antigens, Ezrin displayed the highest frequency of autoantibodies in GEM with early disease and in PDAC patients with resectable disease. The diagnostic value of Ezrin-autoantibodies to discriminate PDAC from controls was further shown by ELISA and ROC analyses (P < 0.0001). This observation was confirmed in prediagnostic sera from the EPIC prospective study in patients who eventually developed PDAC (with a mean time lag of 61.2 months between blood drawing and PDAC diagnosis). A combination of Ezrin-autoantibodies with CA19.9 serum levels and phosphorylated α-Enolase autoantibodies showed an overall diagnostic accuracy of 0.96 ± 0.02. CONCLUSIONS: Autoantibodies against Ezrin are induced early in PDAC and their combination with other serological markers may provide a predictive and diagnostic signature.

Histomolecular phenotypes and outcome in adenocarcinoma of the ampulla of vater.

PURPOSE: Individuals with adenocarcinoma of the ampulla of Vater demonstrate a broad range of outcomes, presumably because these cancers may arise from any one of the three epithelia that converge at that location. This variability poses challenges for clinical decision making and the development of novel therapeutic strategies. PATIENTS AND METHODS: We assessed the potential clinical utility of histomolecular phenotypes defined using a combination of histopathology and protein expression (CDX2 and MUC1) in 208 patients from three independent cohorts who underwent surgical resection for adenocarcinoma of the ampulla of Vater. RESULTS: Histologic subtype and CDX2 and MUC1 expression were significant prognostic variables. Patients with a histomolecular pancreaticobiliary phenotype (CDX2 negative, MUC1 positive) segregated into a poor prognostic group in the training (hazard ratio [HR], 3.34; 95% CI, 1.69 to 6.62; P < .001) and both validation cohorts (HR, 5.65; 95% CI, 2.77 to 11.5; P < .001 and HR, 2.78; 95% CI, 1.25 to 7.17; P = .0119) compared with histomolecular nonpancreaticobiliary carcinomas. Further stratification by lymph node (LN) status defined three clinically relevant subgroups: one, patients with histomolecular nonpancreaticobiliary (intestinal) carcinoma without LN metastases who had an excellent prognosis; two, those with histomolecular pancreaticobiliary carcinoma with LN metastases who had a poor outcome; and three, the remainder of patients (nonpancreaticobiliary, LN positive or pancreaticobiliary, LN negative) who had an intermediate outcome. CONCLUSION: Histopathologic and molecular criteria combine to define clinically relevant histomolecular phenotypes of adenocarcinoma of the ampulla of Vater and potentially represent distinct diseases with significant implications for current therapeutic strategies, the ability to interpret past clinical trials, and future trial design.

Irrelevance of microsatellite instability in the epidemiology of sporadic pancreatic ductal adenocarcinoma.

BACKGROUND AND AIMS: Pancreatic cancer risk is increased in Lynch syndrome (LS) patients with mismatch repair gene defects predisposing to colonic and extracolonic cancers with microsatellite instability (MSI). However, the frequency of MSI pancreatic cancers has never been ascertained in consecutive, unselected clinical series, and their contribution to the sporadic and inherited burden of pancreatic cancer remains to be established. Aims of the study were to determine the prevalence of MSI in surgically resected pancreatic cancers in a multicentric, retrospective study, and to assess the occurrence of pancreatic cancer in LS. METHODS: MS-status was screened by a panel of 5 mononucleotide repeats (Bat26, Bat25, NR-21, NR-24 and NR-27) in 338 consecutive pancreatic ductal adenocarcinoma (PDAC), resected at two Italian and one German referral centres. The personal history of pancreatic cancer was assessed in an independent set of 58 probands with LS and in 138 first degree relatives who had cancers. RESULTS: Only one PDAC (0.3%) showed MSI. This was a medullary type cancer, with hMLH1-deficiency, and no identified germ-line mutation but methylation of hMLH1. Pancreatic cancer occurred in 5 (2.5%) LS patients. Histological sampling was available for 2 cases, revealing PDAC in one case and an ampullary cancer in the other one. CONCLUSIONS: MSI prevalence is negligible in sporadic, resected PDAC. Differently, the prevalence of pancreatic cancer is 2.5% in LS patients, and cancers other than PDAC may be encountered in this setting. Surveillance for pancreatic cancer should be advised in LS mutation carriers at referral centers.

Expression of the antiapoptotic protein BAG3 is a feature of pancreatic adenocarcinoma and its overexpression is associated with poorer survival.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most deadly cancers, being the fourth leading cause of cancer-related deaths. Long-term survival reaching 15% is achieved in less than 5% of patients who undergo surgery, and median survival is only 6 months in those with inoperable lesions. A deeper understanding of PDAC biologic characteristics as well as novel prognostic markers are therefore required to improve outcomes. Herein we report that BAG3, a protein with recognized anti-apoptotic activity, was expressed in 346 PDACs analyzed, but was not expressed in the surrounding nonneoplastic tissue. In a cohort of 66 patients who underwent radical resection (R0), survival was significantly shorter in patients with high BAG3 expression (median, 12 months) than in those with low BAG3 expression (median, 23 months) (P = 0.001). Furthermore, we report that BAG3 expression in PDAC-derived cell lines protects from apoptosis and confers resistance to gemcitabine, offering a partial explanation for the survival data. Our results indicate that BAG3 has a relevant role in PDAC biology, and suggest that BAG3 expression level might be a potential marker for prediction of patient outcome.

Classical lobular breast carcinoma consistently lacks topoisomerase-IIα gene amplification: implications for the tailored use of anthracycline-based chemotherapies.

AIMS: There is consistent lack of data focusing the topoisomerase-IIα gene status in lobular breast carcinoma, a subtype that usually shows poor responsiveness to chemotherapies including those using anthracycline drugs. METHODS AND RESULTS: Forty-six infiltrative lobular carcinomas, 13 with matched metastases, were used. Topoisomerase-IIα gene amplification was evaluated by chromogenic in situ hybridization (CISH) and fluorescence in situ hybridization (FISH). We also assessed Her2/neu status by CISH, FISH and silver in situ hybridization (SISH). HER2 immunoexpression was assessed by the HercepTest. Forty-four of 46 (95%) cases revealed no topoisomerase-IIα amplification, whereas two of 46 (5%) cases were amplified by all three techniques. Eleven of the 13 metastatic sites showed no amplification either in the primary or in the metastases (85%); the remaining two were amplified (15%). Her2/neu was not amplified in 44 of 46 (95%) cases nor was it amplified in 11 of 13 (95%) metastatic tissues. The two cases showing Her2/neu and topoisomerase-IIα amplification scored 3+; the remaining non-amplified cases scored 0 or 1+ in 40 and 2+ in four cases. CONCLUSIONS: In the era of personalized and tailored therapies, we suggest that patients affected by the classical lobular subtype of breast carcinoma constantly lack the ad hoc predictive rationale for receiving common chemotherapy that includes anthracyclines.

FISH scoring on paraffin sections versus single-cell suspension for chromophobe renal carcinoma and renal oncocytoma.

BACKGROUND: Sectioning of the nuclei on tissue sections may give an overestimate of monosomy, a feature diagnostic of chromophobe renal cell carcinoma versus renal oncocytoma. The aim of the study was to assess whether or not nuclear sectioning may distort the results obtained from interphase fluorescence in situ hybridization (FISH) comparing the data obtained from analysis of isolated nuclei derived from formalin-fixed, paraffin-embedded sections with histological sections from the adjacent sections from the same tumors. PATIENTS AND METHODS: Five chromophobe renal cell carcinomas and five renal oncocytomas were recruited. Sections of 5 µm and 30 µm were cut for FISH to investigate chromosomes 1, 2, 6 10 and 17. RESULTS: FISH of isolated nuclei from renal oncocytomas showed a mean increase of 3.0% for nuclei with two signals when compared to tissue sections. For chromosomes 2, 6, 10 and 17, isolated nuclei showed a mean increase of 4.9% of fluorescent signals over nuclei from tissue sections. FISH analysis of isolated nuclei from chromophobe renal cell carcinoma showed a similar counts. CONCLUSION: When a tumor section exhibits a borderline percentage of nuclei with single signals around the cut-off level on tissue sections, the test should be repeated on isolated nuclei to confirm chromosomal loss, diagnostic of chromophobe renal carcinoma.

Chromosome 3p alterations in pancreatic endocrine neoplasia.

Pancreatic endocrine tumors (PET) are rare neoplasms classified as functioning (F-PET) or non-functioning (NF-PET) according to the presence of a clinical syndrome due to hormonal hypersecretion. PETs show variable degrees of clinical aggressiveness and loss of chromosome 3p has been suggested to be associated with an advanced stage of disease. We assessed chromosome 3p copy number in 113 primary PETs and 32 metastases by fluorescence in situ hybridization (FISH) using tissue microarrays. The series included 56 well-differentiated endocrine tumors (WDET), 62 well-differentiated endocrine carcinomas (WDEC), and 6 poorly differentiated endocrine carcinomas (PDEC). Chromosome 3p alterations were found in 23/113 (20%) primary tumors, with losses being predominant over gains (14% vs. 6%). Loss of 3p was found in 5/55 (9%) WDET, 11/52 (21%) WDEC, and never in PDEC. Gains of 3p were detected in 4/55 (7%) WDET, no WDEC, but notably in 3/6 (50%) PDEC (OR 23.6; P = 0.003). Metastases were more frequently monosomic for 3p compared to primary tumors (OR 3.6; P = 0.005). Monosomy was significantly associated with larger tumor size, more advanced tumor stage, and metastasis. No association was found with survival. Chromosome 3p copy number alterations are frequent events in advanced stage PET, with gains prevailing in PDEC while losses are more frequent in WDEC, supporting the view that a specific pattern of alterations are involved in these diverse disease subtypes.

MEN1 in pancreatic endocrine tumors: analysis of gene and protein status in 169 sporadic neoplasms reveals alterations in the vast majority of cases.

Pancreatic endocrine tumors (PETs) may be part of hereditary multiple endocrine neoplasia type 1 (MEN1) syndrome. While MEN1 gene mutation is the only ascertained genetic anomaly described in PETs, no data exist on the cellular localization of MEN1-encoded protein, menin, in normal pancreas and PETs. A total of 169 PETs were used to assess the i) MEN1 gene mutational status in 100 clinically sporadic PETs by direct DNA sequencing, ii) immunohistochemical expression of menin in normal pancreas and 140 PETs, including 71 cases screened for gene mutations, and iii) correlation of these findings with clinical-pathological parameters. Twenty-seven PETs showed mutations that were somatic in 25 patients and revealed to be germline in 2 patients. Menin immunostaining showed strong nuclear and very faint cytoplasmic signal in normal islet cells, whereas it displayed abnormal location and expression levels in 80% of tumors. PETs harboring MEN1 truncating mutations lacked nuclear protein, and most PETs with MEN1 missense mutations showed a strong cytoplasmic positivity for menin. Menin was also misplaced in a significant number of cases lacking MEN1 mutations. In conclusion, the vast majority of PETs showed qualitative and/or quantitative alterations in menin localization. In 30% of cases, this was associated with MEN1 mutations affecting sequences involved in nuclear localization or protein-protein interaction. In cases lacking MEN1 mutations, the alteration of one of the menin interactors may have prevented its proper localization, as suggested by recent data showing that menin protein shuttles between the nucleus and cytoplasm and also affects the subcellular localization of its interactors.

The analysis of PIK3CA mutations in gastric carcinoma and metanalysis of literature suggest that exon-selectivity is a signature of cancer type.

BACKGROUND: PIK3CA is one of the genes most frequently mutated in human cancers and it is a potential target for personalized therapy. The aim of this study was to assess the frequency and type of PIK3CA mutations in gastric carcinoma and compare them with their clinical pathological correlates. METHODS: We analysed 264 gastric cancers, including 39 with microsatellite instability (MSI), for mutations in the two PIK3CA hotspots in exons 9 and 20 by direct sequencing of DNA obtained from microdissected cancer cells. RESULTS: The cases harbouring mutations were 42 (16%). All were heterozygous missense single base substitutions; the most common was H1047R (26/42; 62%) in exon 20 and the second was Q546K (4/42; 9.5%) in exon 9. All the mutated MSI cases (8/39) carried the H1047R mutation. No other association between PI3KCA mutations and their clinical pathological covariates was found. A metanalysis of the mutations occurring in the same regions presented in 27 publications showed that ratio between exon 20 and exon 9 prevalences was 0.6 (95% CI: 0.5 -0.8) for colon, 1.6 (95% CI: 1.1 -2.3) for breast, 2.7 (95% CI: 1.6 -4.9) for gastric and 4.1 (95% CI: 1.9 -10.3) for endometrial cancer. CONCLUSIONS: The overall prevalence of PIK3CA mutations implies an important role for PIK3CA in gastric cancer. The lack of association with any clinical-pathological condition suggests that mutations in PIK3CA occur early in the development of cancer. The metanalysis showed that exon-selectivity is an important signature of cancer type reflecting different contexts in which tumours arise.

Anti-viral state segregates two molecular phenotypes of pancreatic adenocarcinoma: potential relevance for adenoviral gene therapy.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) remains a leading cause of cancer mortality for which novel gene therapy approaches relying on tumor-tropic adenoviruses are being tested. METHODS: We obtained the global transcriptional profiling of primary PDAC using RNA from eight xenografted primary PDAC, three primary PDAC bulk tissues, three chronic pancreatitis and three normal pancreatic tissues. The Affymetrix GeneChip HG-U133A was used. The results of the expression profiles were validated applying immunohistochemical and western blot analysis on a set of 34 primary PDAC and 10 established PDAC cell lines. Permissivity to viral vectors used for gene therapy, Adenovirus 5 and Adeno-Associated Viruses 5 and 6, was assessed on PDAC cell lines. RESULTS: The analysis of the expression profiles allowed the identification of two clearly distinguishable phenotypes according to the expression of interferon-stimulated genes. The two phenotypes could be readily recognized by immunohistochemical detection of the Myxovirus-resistance A protein, whose expression reflects the activation of interferon dependent pathways. The two molecular phenotypes discovered in primary carcinomas were also observed among established pancreatic adenocarcinoma cell lines, suggesting that these phenotypes are an intrinsic characteristic of cancer cells independent of their interaction with the host's microenvironment. The two pancreatic cancer phenotypes are characterized by different permissivity to viral vectors used for gene therapy, as cell lines expressing interferon stimulated genes resisted to Adenovirus 5 mediated lysis in vitro. Similar results were observed when cells were transduced with Adeno-Associated Viruses 5 and 6. CONCLUSION: Our study identified two molecular phenotypes of pancreatic cancer, characterized by a differential expression of interferon-stimulated genes and easily recognized by the expression of the Myxovirus-resistance A protein. We suggest that the detection of these two phenotypes might help the selection of patients enrolled in virally-mediated gene therapy trials.