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

Inferring structural variant cancer cell fraction.

We present SVclone, a computational method for inferring the cancer cell fraction of structural variant (SV) breakpoints from whole-genome sequencing data. SVclone accurately determines the variant allele frequencies of both SV breakends, then simultaneously estimates the cancer cell fraction and SV copy number. We assess performance using in silico mixtures of real samples, at known proportions, created from two clonal metastases from the same patient. We find that SVclone's performance is comparable to single-nucleotide variant-based methods, despite having an order of magnitude fewer data points. As part of the Pan-Cancer Analysis of Whole Genomes (PCAWG) consortium, which aggregated whole-genome sequencing data from 2658 cancers across 38 tumour types, we use SVclone to reveal a subset of liver, ovarian and pancreatic cancers with subclonally enriched copy-number neutral rearrangements that show decreased overall survival. SVclone enables improved characterisation of SV intra-tumour heterogeneity

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

<p>Abstract</p> <p>Background</p> <p>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.</p> <p>Methods</p> <p>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.</p> <p>Results</p> <p>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.</p> <p>Conclusion</p> <p>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.</p

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 clinical pathological covariates was found. A metanalysis of the mutations occurring in the same regions 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

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

DNA methylation patterns identify subgroups of pancreatic neuroendocrine tumors with clinical association

Here we report the DNA methylation profile of 84 sporadic pancreatic neuroendocrine tumors (PanNETs) with associated clinical and genomic information. We identified three subgroups of PanNETs, termed T1, T2 and T3, with distinct patterns of methylation. The T1 subgroup was enriched for functional tumors and ATRX, DAXX and MEN1 wild-type genotypes. The T2 subgroup contained tumors with mutations in ATRX, DAXX and MEN1 and recurrent patterns of chromosomal losses in half of the genome with no association between regions with recurrent loss and methylation levels. T2 tumors were larger and had lower methylation in the MGMT gene body, which showed positive correlation with gene expression. The T3 subgroup harboured mutations in MEN1 with recurrent loss of chromosome 11, was enriched for grade G1 tumors and showed histological parameters associated with better prognosis. Our results suggest a role for methylation in both driving tumorigenesis and potentially stratifying prognosis in PanNETs

Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Funder: NCI U24CA211006Abstract: The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts

Molecular characterization of Pancreatic NeuroEndocrine Tumors (PanNETs)

Scopo: identificare le alterazioni molecolari clinicamente rilevanti associate ai tumori neuroendocrini del pancreas (PanNETs). Materiali e Metodi: 204 pazienti sono stati studiati per la ricerca mutazionale in 8 geni frequentemente alterati in PanNETs utilizzando Tecnologia Ion Torrent. Tali geni si dividono tra MEN1, ATRX e DAXX coinvolti nel rimodellamento della cromatina, PIK3CA,TSC2,PTEN,MTOR implicati nel mTOR pathway e infine ATM presente come ruolo centrale nel mantenimento della stabilit\ue0 e riparazione del DNA. Inoltre sono stati valutati: i) le variazioni del numero di copie (CNV) tramite dati CGH array disponibili in 90 casi, ii) l\u2019allungamento alternativo dei telomeri (ALT) tramite FISH in un numero selezionato di casi iii) l\u2019espressione proteica tramite immunoistochimica per Menina, ATRX, Daxx, Atm e Pten in 171PanNETs. Risultati: l\u2019analisi mutazionale mirata tramite next generation sequencing di 204 PanNETs ha dimostrato la presenza di mutazioni in 112/204 (55%), mentre in 92 (45%) risultano non alterati. E\u2019 stato inoltre rilevato i) le mutazioni a carico dei geni Daxx e ATRX risultano essere mutuamente esclusive e pi\uf9 frequentemente presenti pari a 53/204 (26%) e associate con mutazioni di MEN1 o ATM a loro volta mutuamente esclusive; ii) mutazioni DAXX/ATRX sono associate a un sottoinsieme di PanNETs che presentano perdite cromosomiche ; iii ) le mutazioni PTEN e TSC2 si escludono a vicenda e sono state trovate in 31/204 (15%), mentre l'attivazione mTOR \ue8 associato con una migliore sopravvivenza libera da malattia. Conclusioni : in questo studio abbiamo dimostrato che i PanNETs possono essere suddivisi tra due principali sottogruppi molecolari. Un primo gruppo mostra una simultanea perdita cromosomica nelle neoplasie con mutazioni a carico di MEN1 e la contemporanea alterazione nei geni DAXX , ATRX , PTEN e TSC2 . Il secondo gruppo di casi presenta mutazioni di MEN1 senza alterazioni cromosomiche rincorrenti il cui significato deve essere ancora scoperto.Purpose: To identify clinically relevant molecular alterations associated to pancreatic endocrine tumors (PanNETs) discovered by recent exome sequencing data. Patients and Methods: A series of 204 PanNETs were explored for intragenic mutations in 8 genes reported as recurrently altered in PanNETs using Ion Torrent technology. The 8 genes were MEN1, ATM, ATRX, DAXX, PIK3CA, TSC2, PTEN, MTOR. Their relation with chromosomal copy number aberrations were assessed in 90 cases having CGH array data available, and in a selected number of cases alternative lengthening of telomeres (ALT) was investigated by FISH. A total of 171 PanNETs were also investigated by immunohistochemistry for Menin, Atrx, Daxx, Atm and Pten. Results: Our targetted next-generation mutational analysis of 204 PanNETs for three chromatin remodelling genes (MEN1, ATRX, DAXX), MTOR pathway genes (PTEN, TSC2, PIK3CA, MTOR) and the ATM DNA repair gene assessed that 112/204 (55%) PanNETS had mutations, while 92 (45%) lacked mutations, and: i) the mutually exclusive mutations in DAXX and ATRX chromatin remodelling genes were the most frequent, accounting for 53/204 (26%) cases, and were associated with MEN1 or ATM mutations that were in turn mutually exclusive; ii) DAXX/ATRX mutations are associated with a subset of PanNETS showing a peculiar pattern of recurrent chromosomal losses; iii) the mutually exclusive mutations in genes belonging to mTOR pathways were found in 31/204 (15%), and mTOR pathway activation is associated with shorter disease-free survival. Conclusions: We report that PanNET can be subdived into two main molecular subgroups. The first showing the peculiar simultaneous loss of twelve chromosomes, which includes the vast majority of neoplasms showing MEN1 mutations and concurrent mutations in DAXX, ATRX, PTEN and TSC2. The second group, where only a small number of cases shows MEN1 mutations, lacks recurrent chromosomal anomalies and the gene alterations responsible for their development remain to be discovered

HER-2/neu assessment in breast cancer using the original FDA and new ASCO/CAP guideline recommendations: impact on selecting patients for herceptin therapy

We evaluated HER-2/neu status in 100 consecutive ductal breast carcinomas by using the Food and Drug Administration (FDA) and American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) scoring systems. With the FDA system, scores were 3+ in 23.0\%, 2+ in 25.0\%, and 0 or 1+ in 52.0\% of cases. With the ASCO/CAP system, scores were 3+ in 16.0\%, 2+ in 34.0\%, and 0 or 1+ in 50.0\%. With the FDA and ASCO/CAP systems, respectively, 3+ cases (n = 23 and 16, respectively) showed high-grade, granular HER-2/neu amplification in 15 (65\%) and 14 (88\%); low-grade, borderline amplification in 7 (30\%) and 1 (6\%); and chromosome 17 polysomy without amplification in 1 (4\%) and 1 (6\%).Concordance between schemes was higher for cases with high-grade, granular HER-2/neu amplification (concordance coefficient, 0.74). Cases with low-grade, borderline HER-2/neu amplification showed poor concordance (concordance coefficient, 0.20).The FDA and ASCO/CAP schemes for HER-2/neu evaluation select patients differently for trastuzumab therapy. Major discordance is present for low-grade, borderline HER-2/neu amplification. FDA low-grade, borderline tumors would be reclassified as without HER-2/neu amplification or as polysomic. The ASCO/CAP scheme has a great concordance coefficient between strong 3+ immunohistochemical cases and cases with high-grade, granular HER-2/neu amplification