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UNLABELLED: Ovarian carcinosarcoma (OCS) is an aggressive and rare tumor type with limited treatment options. OCS is hypothesized to develop via the combination theory, with a single progenitor resulting in carcinomatous and sarcomatous components, or alternatively via the conversion theory, with the sarcomatous component developing from the carcinomatous component through epithelial-to-mesenchymal transition (EMT). In this study, we analyzed DNA variants from isolated carcinoma and sarcoma components to show that OCS from 18 women is monoclonal. RNA sequencing indicated that the carcinoma components were more mesenchymal when compared with pure epithelial ovarian carcinomas, supporting the conversion theory and suggesting that EMT is important in the formation of these tumors. Preclinical OCS models were used to test the efficacy of microtubule-targeting drugs, including eribulin, which has previously been shown to reverse EMT characteristics in breast cancers and induce differentiation in sarcomas. Vinorelbine and eribulin more effectively inhibited OCS growth than standard-of-care platinum-based chemotherapy, and treatment with eribulin reduced mesenchymal characteristics and N-MYC expression in OCS patient-derived xenografts. Eribulin treatment resulted in an accumulation of intracellular cholesterol in OCS cells, which triggered a downregulation of the mevalonate pathway and prevented further cholesterol biosynthesis. Finally, eribulin increased expression of genes related to immune activation and increased the intratumoral accumulation of CD8+ T cells, supporting exploration of immunotherapy combinations in the clinic. Together, these data indicate that EMT plays a key role in OCS tumorigenesis and support the conversion theory for OCS histogenesis. Targeting EMT using eribulin could help improve OCS patient outcomes. SIGNIFICANCE: Genomic analyses and preclinical models of ovarian carcinosarcoma support the conversion theory for disease development and indica

Background & Aims: Continuing recalcitrance to therapy cements pancreatic cancer (PC) as the most lethal malignancy, which is set to become the second leading cause of cancer death in our society. The study aim was to investigate the association between DNA damage response (DDR), replication stress, and novel therapeutic response in PC to develop a biomarker-driven therapeutic strategy targeting DDR and replication stress in PC. Methods: We interrogated the transcriptome, genome, proteome, and functional characteristics of 61 novel PC patient–derived cell lines to define novel therapeutic strategies targeting DDR and replication stress. Validation was done in patient-derived xenografts and human PC organoids. Results: Patient-derived cell lines faithfully recapitulate the epithelial component of pancreatic tumors, including previously described molecular subtypes. Biomarkers of DDR deficiency, including a novel signature of homologous recombination deficiency, cosegregates with response to platinum (P <.001) and PARP inhibitor therapy (P <.001) in vitro and in vivo. We generated a novel signature of replication stress that predicts response to ATR (P <.018) and WEE1 inhibitor (P <.029) treatment in both cell lines and human PC organoids. Replication stress was enriched in the squamous subtype of PC (P <.001) but was not associated with DDR deficiency. Conclusions: Replication stress and DDR deficiency are independent of each other, creating opportunities for therapy in DDR-proficient PC and after platinum therapy.

Sex differences have been observed in multiple facets of cancer epidemiology, treatment and biology, and in most cancers outside the sex organs. Efforts to link these clinical differences to specific molecular features have focused on somatic mutations within the coding regions of the genome. Here we report a pan-cancer analysis of sex differences in whole genomes of 1983 tumours of 28 subtypes as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium. We both confirm the results of exome studies, and also uncover previously undescribed sex differences. These include sex-biases in coding and non-coding cancer drivers, mutation prevalence and strikingly, in mutational signatures related to underlying mutational processes. These results underline the pervasiveness of molecular sex differences and strengthen the call for increased consideration of sex in molecular cancer research.

OBJECTIVE: We aimed to define preoperative clinical and molecular characteristics that would allow better patient selection for operative resection. BACKGROUND: Although we use molecular selection methods for systemic targeted therapies, these principles are not applied to surgical oncology. Improving patient selection is of vital importance for the operative treatment of pancreatic cancer (pancreatic ductal adenocarcinoma). Although surgery is the only chance of long-term survival, 80% still succumb to the disease and approximately 30% die within 1 year, often sooner than those that have unresected local disease. METHOD: In 3 independent pancreatic ductal adenocarcinoma cohorts (total participants = 1184) the relationship between aberrant expression of prometastatic proteins S100A2 and S100A4 and survival was assessed. A preoperative nomogram based on clinical variables available before surgery and expression of these proteins was constructed and compared to traditional measures, and a postoperative nomogram. RESULTS: High expression of either S100A2 or S100A4 was independent poor prognostic factors in a training cohort of 518 participants. These results were validated in 2 independent patient cohorts (Glasgow, n = 198; Germany, n = 468). Aberrant biomarker expression stratified the cohorts into 3 distinct prognostic groups. A preoperative nomogram incorporating S100A2 and S100A4 expression predicted survival and nomograms derived using postoperative clinicopathological variables. CONCLUSIONS: Of those patients with a poor preoperative nomogram score, approximately 50% of patients died within a year of resection. Nomograms have the potential to improve selection for surgery and neoadjuvant therapy, avoiding surgery in aggressive disease, and justifying more extensive resections in biologically favorable disease.

OBJECTIVE: Extensive molecular heterogeneity of pancreatic ductal adenocarcinoma (PDA), few effective therapies and high mortality make this disease a prime model for advancing development of tailored therapies. The p16-cyclin D-cyclin-dependent kinase 4/6-retinoblastoma (RB) protein (CDK4) pathway, regulator of cell proliferation, is deregulated in PDA. Our aim was to develop a novel personalised treatment strategy for PDA based on targeting CDK4. DESIGN: Sensitivity to potent CDK4/6 inhibitor PD-0332991 (palbociclib) was correlated to protein and genomic data in 19 primary patient-derived PDA lines to identify biomarkers of response. In vivo efficacy of PD-0332991 and combination therapies was determined in subcutaneous, intrasplenic and orthotopic tumour models derived from genome-sequenced patient specimens and genetically engineered model. Mechanistically, monotherapy and combination therapy were investigated in the context of tumour cell and extracellular matrix (ECM) signalling. Prognostic relevance of companion biomarker, RB protein, was evaluated and validated in independent PDA patient cohorts (>500 specimens). RESULTS: Subtype-specific in vivo efficacy of PD-0332991-based therapy was for the first time observed at multiple stages of PDA progression: primary tumour growth, recurrence (second-line therapy) and metastatic setting and may potentially be guided by a simple biomarker (RB protein). PD-0332991 significantly disrupted surrounding ECM organisation, leading to increased quiescence, apoptosis, improved chemosensitivity, decreased invasion, metastatic spread and PDA progression in vivo. RB protein is prevalent in primary operable and metastatic PDA and may present a promising predictive biomarker to guide this therapeutic approach. CONCLUSION: This study demonstrates the promise of CDK4 inhibition in PDA over standard therapy when applied in a molecular subtype-specific context.

Background: The return of research results (RoR) remains a complex and well-debated issue. Despite the debate, actual data related to the experience of giving individual results back, and the impact these results may have on clinical care and health outcomes, is sorely lacking. Through the work of the Australian Pancreatic Cancer Genome Initiative (APGI) we: (1) delineate the pathway back to the patient where actionable research data were identified; and (2) report the clinical utilisation of individual results returned. Using this experience, we discuss barriers and opportunities associated with a comprehensive process of RoR in large-scale genomic research that may be useful for others developing their own policies. Methods: We performed whole-genome (n = 184) and exome (n = 208) sequencing of matched tumour-normal DNA pairs from 392 patients with sporadic pancreatic cancer (PC) as part of the APGI. We identified pathogenic germline mutations in candidate genes (n = 130) with established predisposition to PC or medium-high penetrance genes with well-defined cancer associated syndromes or phenotypes. Variants from candidate genes were annotated and classified according to international guidelines. Variants were considered actionable if clinical utility was established, with regard to prevention, diagnosis, prognostication and/or therapy. Results: A total of 48,904 germline variants were identified, with 2356 unique variants undergoing annotation and in silico classification. Twenty cases were deemed actionable and were returned via previously described RoR framework, representing an actionable finding rate of 5.1%. Overall, 1.78% of our cohort experienced clinical benefit from RoR. Conclusion: Returning research results within the context of large-scale genomics research is a labour-intensive, highly variable, complex operation. Results that warrant action are not infrequent, but the prevalence of those who experience a clinical difference as a result of returning

BACKGROUND: Pancreatic cancer has a five-year survival rate of ~8%, with characteristic molecular heterogeneity and restricted treatment options. Targeting metabolism has emerged as a potentially effective therapeutic strategy for cancers such as pancreatic cancer, which are driven by genetic alterations that are not tractable drug targets. Although somatic mitochondrial genome (mtDNA) mutations have been observed in various tumors types, understanding of metabolic genotype-phenotype relationships is limited. METHODS: We deployed an integrated approach combining genomics, metabolomics, and phenotypic analysis on a unique cohort of patient-derived pancreatic cancer cell lines (PDCLs). Genome analysis was performed via targeted sequencing of the mitochondrial genome (mtDNA) and nuclear genes encoding mitochondrial components and metabolic genes. Phenotypic characterization of PDCLs included measurement of cellular oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) using a Seahorse XF extracellular flux analyser, targeted metabolomics and pathway profiling, and radiolabelled glutamine tracing. RESULTS: We identified 24 somatic mutations in the mtDNA of 12 patient-derived pancreatic cancer cell lines (PDCLs). A further 18 mutations were identified in a targeted study of ~1000 nuclear genes important for mitochondrial function and metabolism. Comparison with reference datasets indicated a strong selection bias for non-synonymous mutants with predicted functional effects. Phenotypic analysis showed metabolic changes consistent with mitochondrial dysfunction, including reduced oxygen consumption and increased glycolysis. Metabolomics and radiolabeled substrate tracing indicated the initiation of reductive glutamine metabolism and lipid synthesis in tumours. CONCLUSIONS: The heterogeneous genomic landscape of pancreatic tumours may converge on a common metabolic phenotype, with individual tumours adapting to increased anabolic demands via different ge

Pancreatic cancer has a devastating prognosis, with an overall 5-year survival rate of ∼8%, restricted treatment options and characteristic molecular heterogeneity. SerpinB2 expression, particularly in the stromal compartment, is associated with reduced metastasis and prolonged survival in pancreatic ductal adenocarcinoma (PDAC) and our genomic analysis revealed that SERPINB2 is frequently deleted in PDAC. We show that SerpinB2 is required by stromal cells for normal collagen remodelling in vitro, regulating fibroblast interaction and engagement with collagen in the contracting matrix. In a pancreatic cancer allograft model, co-injection of PDAC cancer cells and SerpinB2 -/- mouse embryonic fibroblasts (MEFs) resulted in increased tumour growth, aberrant remodelling of the extracellular matrix (ECM) and increased local invasion from the primary tumour. These tumours also displayed elevated proteolytic activity of the primary biochemical target of SerpinB2 - urokinase plasminogen activator (uPA). In a large cohort of patients with resected PDAC, we show that increasing uPA mRNA expression was significantly associated with poorer survival following pancreatectomy. This study establishes a novel role for SerpinB2 in the stromal compartment in PDAC invasion through regulation of stromal remodelling and highlights the SerpinB2/uPA axis for further investigation as a potential therapeutic target in pancreatic cancer.

The contributions of coding mutations to tumorigenesis are relatively well known; however, little is known about somatic alterations in noncoding DNA. Here we describe GECCO (Genomic Enrichment Computational Clustering Operation) to analyze somatic noncoding alterations in 308 pancreatic ductal adenocarcinomas (PDAs) and identify commonly mutated regulatory regions. We find recurrent noncoding mutations to be enriched in PDA pathways, including axon guidance and cell adhesion, and newly identified processes, including transcription and homeobox genes. We identified mutations in protein binding sites correlating with differential expression of proximal genes and experimentally validated effects of mutations on expression. We developed an expression modulation score that quantifies the strength of gene regulation imposed by each class of regulatory elements, and found the strongest elements were most frequently mutated, suggesting a selective advantage. Our detailed single-cancer analysis of noncoding alterations identifies regulatory mutations as candidates for diagnostic and prognostic markers, and suggests new mechanisms for tumor evolution.

© 2016 The Author(s). Immunotherapy approaches for pancreatic ductal adenocarcinoma (PDAC) have met with limited success. It has been postulated that a low mutation load may lead to a paucity of T cells within the tumor microenvironment (TME). However, it is also possible that while neoantigens are present, an effective immune response cannot be generated due to an immune suppressive TME. To discern whether targetable neoantigens exist in PDAC, we performed a comprehensive study using genomic profiles of 221 PDAC cases extracted from public databases. Our findings reveal that: (a) nearly all PDAC samples harbor potentially targetable neoantigens; (b) T cells are present but generally show a reduced activation signature; and (c) markers of efficient antigen presentation are associated with a reduced signature of markers characterizing cytotoxic T cells. These findings suggest that despite the presence of tumor specific neoepitopes, T cell activation is actively suppressed in PDAC. Further, we identify iNOS as a potential mediator of immune suppression that might be actionable using pharmacological avenues.

Pancreatic cancer is a major cause of cancer-associated mortality, with a dismal overall prognosis that has remained virtually unchanged for many decades. Currently, prevention or early diagnosis at a curable stage is exceedingly difficult; patients rarely exhibit symptoms and tumours do not display sensitive and specific markers to aid detection. Pancreatic cancers also have few prevalent genetic mutations; the most commonly mutated genes are KRAS, CDKN2A (encoding p16), TP53 and SMAD4-none of which are currently druggable. Indeed, therapeutic options are limited and progress in drug development is impeded because most pancreatic cancers are complex at the genomic, epigenetic and metabolic levels, with multiple activated pathways and crosstalk evident. Furthermore, the multilayered interplay between neoplastic and stromal cells in the tumour microenvironment challenges medical treatment. Fewer than 20% of patients have surgically resectable disease; however, neoadjuvant therapies might shift tumours towards resectability. Although newer drug combinations and multimodal regimens in this setting, as well as the adjuvant setting, appreciably extend survival, ∼80% of patients will relapse after surgery and ultimately die of their disease. Thus, consideration of quality of life and overall survival is important. In this Primer, we summarize the current understanding of the salient pathophysiological, molecular, translational and clinical aspects of this disease. In addition, we present an outline of potential future directions for pancreatic cancer research and patient management.

Background:Pancreatic stellate cells (PSCs, which produce the stroma of pancreatic cancer (PC)) interact with cancer cells to facilitate PC growth. A candidate growth factor pathway that may mediate this interaction is the HGF-c-MET pathway.Methods:Effects of HGF inhibition (using a neutralising antibody AMG102) alone or in combination with gemcitabine were assessed (i) in vivo using an orthotopic model of PC, and (ii) in vitro using cultured PC cells (AsPC-1) and human PSCs.Results:We have shown that human PSCs (hPSCs) secrete HGF but do not express the receptor c-MET, which is present predominantly on cancer cells. HGF inhibition was as effective as standard chemotherapy in inhibiting local tumour growth but was significantly more effective than gemcitabine in reducing tumour angiogenesis and metastasis. HGF inhibition has resulted in reduced metastasis; however, interestingly this antimetastatic effect was lost when combined with gemcitabine. This suggests that gemcitabine treatment selects out a subpopulation of cancer cells with increased epithelial-mesenchymal transition (EMT) and stem-cell characteristics, as supported by our findings of increased expression of EMT and stem-cell markers in tumour sections from our animal model. In vitro studies showed that hPSC secretions induced proliferation and migration, but inhibited apoptosis, of cancer cells. These effects were countered by pretreatment of hPSC secretions with a HGF-neutralising antibody but not by gemcitabine, indicating a key role for HGF in PSC-PC interactions.Conclusions:Our studies suggest that targeted therapy to inhibit stromal-tumour interactions mediated by the HGF-c-MET pathway may represent a novel therapeutic approach in PC that will require careful modelling for optimal integration with existing treatment modalities.

Pancreatic cancer (PC) is a lethal disease which is characterized by chemoresistance. Components of the cell cytoskeleton are therapeutic targets in cancer. βIV-tubulin is one such component that has two isotypes—βIVa and βIVb. βIVa and βIVb isotypes only differ in two amino acids at their C-terminus. Studies have implicated βIVa-tubulin or βIVb-tubulin expression with chemoresistance in prostate, breast, ovarian and lung cancer. However, no studies have examined the role of βIV-tubulin in PC or attempted to identify isotype specific roles in regulating cancer cell growth and chemosensitivity. We aimed to determine the role of βIVa- or βIVb-tubulin on PC growth and chemosensitivity. PC cells (MiaPaCa-2, HPAF-II, AsPC1) were treated with siRNA (control, βIVa-tubulin or βIVb-tubulin). The ability of PC cells to form colonies in the presence or absence of chemotherapy was measured by clonogenic assays. Inhibition of βIVa-tubulin in PC cells had no effect chemosensitivity. In contrast, inhibition of βIVb-tubulin in PC cells sensitized to vinca alkaloids (Vincristine, Vinorelbine and Vinblastine), which was accompanied by increased apoptosis and enhanced cell cycle arrest. We show for the first time that βIVb-tubulin, but not βIVa-tubulin, plays a role in regulating vinca alkaloid chemosensitivity in PC cells. The results from this study suggest βIVb-tubulin may be a novel therapeutic target and predictor of vinca alkaloid sensitivity for PC and warrants further investigation.

Recent advances in biotechnologies have led to the development of multiplex genomic and proteomic analyses for clinical use. Nevertheless, guidelines are currently lacking to determine which molecular assays should be implemented in metastatic cancers. The first MAP conference was dedicated to exploring the use of genomics to better select therapies in the treatment of metastatic cancers. Sixteen consensus items were covered. There was a consensus that new technologies like next-generation sequencing of tumors and ddPCR on circulating free DNA have convincing analytical validity. Further work needs to be undertaken to establish the clinical utility of liquid biopsies and the added clinical value of expanding from individual gene tests into large gene panels. Experts agreed that standardized bioinformatics methods for biological interpretation of genomic data are needed and that precision medicine trials should be stratified based on the level of evidence available for the genomic alterations identified., SCOPUS: ar.j, info:eu-repo/semantics/published

Pancreatic cancer is a leading cause of cancer-related deaths in Western societies. This poor prognosis is due to chemotherapeutic drug resistance and metastatic spread. Evidence suggests that microtubule proteins namely, β-tubulins are dysregulated in tumor cells and are involved in regulating chemosensitivity. However, the role of β-tubulins in pancreatic cancer are unknown. We measured the expression of different β-tubulin isotypes in pancreatic adenocarcinoma tissue and pancreatic cancer cells. Next, we used RNAi to silence βIII-tubulin expression in pancreatic cancer cells, and measured cell growth in the absence and presence of chemotherapeutic drugs. Finally, we assessed the role of βIII-tubulin in regulating tumor growth and metastases using an orthotopic pancreatic cancer mouse model. We found that βIII-tubulin is highly expressed in pancreatic adenocarcinoma tissue and pancreatic cancer cells. Further, we demonstrated that silencing βIII-tubulin expression reduced pancreatic cancer cell growth and tumorigenic potential in the absence and presence of chemotherapeutic drugs. Finally, we demonstrated that suppression of βIII-tubulin reduced tumor growth and metastases in vivo. Our novel data demonstrate that βIII-tubulin is a key player in promoting pancreatic cancer growth and survival, and silencing its expression may be a potential therapeutic strategy to increase the long-term survival of pancreatic cancer patients.

Myc oncoproteins and histone deacetylases (HDACs) exert oncogenic effects by modulating gene transcription. Paradoxically, N-Myc induces p53 gene expression. Tumor protein 53-induced nuclear protein 1 (TP53INP1) phosphorylates p53 protein at serine 46, leading to enhanced p53 activity, transcriptional activation of p53 target genes and programmed cell death. Here we aimed to identify the mechanism through which N-Myc overexpressing p53 wild-type neuroblastoma cells acquired resistance to apoptosis. TP53INP1 was found to be one of the genes most significantly repressed by HDAC2 and N-Myc according to Affymetrix microarray gene expression datasets. HDAC2 and N-Myc reduced TP53INP1 gene expression by direct binding to the TP53INP1 gene promoter, leading to transcriptional repression of TP53INP1, p53 protein de-phosphorylation at serine 46, neuroblastoma cell proliferation and survival. Moreover, low levels of TP53INP1 expression in human neuroblastoma tissues correlated with high levels of N-Myc expression and poor patient outcome, and the BET bromodomain inhibitors JQ1 and I-BET151 reduced N-Myc expression and reactivated TP53INP1 expression in neuroblastoma cells. These findings identify TP53INP1 repression as an important co-factor for N-Myc oncogenesis, and provide further evidence for the potential application of BET bromodomain inhibitors in the therapy of N-Myc-induced neuroblastoma.

OBJECTIVE: Pancreatic cancer is a leading cause of cancer-related death in the Western world. Current chemotherapy regimens have modest survival benefit. Thus, novel, effective therapies are required for treatment of this disease. DESIGN: Activating KRAS mutation almost always drives pancreatic tumour initiation, however, deregulation of other potentially druggable pathways promotes tumour progression. PTEN loss leads to acceleration of Kras(G12D)-driven pancreatic ductal adenocarcinoma (PDAC) in mice and these tumours have high levels of mammalian target of rapamycin (mTOR) signalling. To test whether these KRAS PTEN pancreatic tumours show mTOR dependence, we compared response to mTOR inhibition in this model, to the response in another established model of pancreatic cancer, KRAS P53. We also assessed whether there was a subset of pancreatic cancer patients who may respond to mTOR inhibition. RESULTS: We found that tumours in KRAS PTEN mice exhibit a remarkable dependence on mTOR signalling. In these tumours, mTOR inhibition leads to proliferative arrest and even tumour regression. Further, we could measure response using clinically applicable positron emission tomography imaging. Importantly, pancreatic tumours driven by activated KRAS and mutant p53 did not respond to treatment. In human tumours, approximately 20% of cases demonstrated low PTEN expression and a gene expression signature that overlaps with murine KRAS PTEN tumours. CONCLUSIONS: KRAS PTEN tumours are uniquely responsive to mTOR inhibition. Targeted anti-mTOR therapies may offer clinical benefit in subsets of human PDAC selected based on genotype, that are dependent on mTOR signalling. Thus, the genetic signatures of human tumours could be used to direct pancreatic cancer treatment in the future.

BACKGROUND: Disclosure of individual results to participants in genomic research is a complex and contentious issue. There are many existing commentaries and opinion pieces on the topic, but little empirical data concerning actual cases describing how individual results have been returned. Thus, the real life risks and benefits of disclosing individual research results to participants are rarely if ever presented as part of this debate. METHODS: The Australian Pancreatic Cancer Genome Initiative (APGI) is an Australian contribution to the International Cancer Genome Consortium (ICGC), that involves prospective sequencing of tumor and normal genomes of study participants with pancreatic cancer in Australia. We present three examples that illustrate different facets of how research results may arise, and how they may be returned to individuals within an ethically defensible and clinically practical framework. This framework includes the necessary elements identified by others including consent, determination of the significance of results and which to return, delineation of the responsibility for communication and the clinical pathway for managing the consequences of returning results. RESULTS: Of 285 recruited patients, we returned results to a total of 25 with no adverse events to date. These included four that were classified as medically actionable, nine as clinically significant and eight that were returned at the request of the treating clinician. Case studies presented depict instances where research results impacted on cancer susceptibility, current treatment and diagnosis, and illustrate key practical challenges of developing an effective framework. CONCLUSIONS: We suggest that return of individual results is both feasible and ethically defensible but only within the context of a robust framework that involves a close relationship between researchers and clinicians.

Background: Pancreatic cancer is one of the most lethal and molecularly diverse malignancies. Repurposing of therapeutics that target specific molecular mechanisms in different disease types offers potential for rapid improvements in outcome. Although HER2 amplification occurs in pancreatic cancer, it is inadequately characterized to exploit the potential of anti-HER2 therapies. Methods: HER2 amplification was detected and further analyzed using multiple genomic sequencing approaches. Standardized reference laboratory assays defined HER2 amplification in a large cohort of patients (n = 469) with pancreatic ductal adenocarcinoma (PDAC). Results: An amplified inversion event (1 MB) was identified at the HER2 locus in a patient with PDAC. Using standardized laboratory assays, we established diagnostic criteria for HER2 amplification in PDAC, and observed a prevalence of 2%. Clinically, HER2- amplified PDAC was characterized by a lack of liver metastases, and a preponderance of lung and brain metastases. Excluding breast and gastric cancer, the incidence of HER2-amplified cancers in the USA is >22,000 per annum. Conclusions: HER2 amplification occurs in 2% of PDAC, and has distinct features with implications for clinical practice. The molecular heterogeneity of PDAC implies that even an incidence of 2% represents an attractive target for anti-HER2 therapies, as options for PDAC are limited. Recruiting patients based on HER2 amplification, rather than organ of origin, could make trials of anti-HER2 therapies feasible in less common cancer types. © 2013 Chou et al.; licensee BioMed Central Ltd.

All cancers are caused by somatic mutations; however, understanding of the biological processes generating these mutations is limited. The catalogue of somatic mutations from a cancer genome bears the signatures of the mutational processes that have been operative. Here we analysed 4,938,362 mutations from 7,042 cancers and extracted more than 20 distinct mutational signatures. Some are present in many cancer types, notably a signature attributed to the APOBEC family of cytidine deaminases, whereas others are confined to a single cancer class. Certain signatures are associated with age of the patient at cancer diagnosis, known mutagenic exposures or defects in DNA maintenance, but many are of cryptic origin. In addition to these genome-wide mutational signatures, hypermutation localized to small genomic regions, ‘kataegis’, is found in many cancer types. The results reveal the diversity of mutational processes underlying the development of cancer, with potential implications for understanding of cancer aetiology, prevention and therapy, SCOPUS: ar.j, info:eu-repo/semantics/published

Large-scale cancer genome studies are unveiling significant complexity and heterogeneity even in histopathologically indistinguishable cancers. Differentiating 'driver' mutations that are functionally relevant from 'passenger' mutations is a major challenge in cancer genomics. While recurrent mutations in a gene provides supporting evidence of 'driver' status, novel computational methods and model systems are greatly improving our ability to identify genes important in carcinogenesis. Reimand and Bader have recently shown that driver gene discovery in discrete gene classes (in this case the kinome) is possible across multiple cancer types and has the potential to yield new druggable targets and clinically relevant leads.

Somatic mutation calling from next-generation sequencing data remains a challenge due to the difficulties of distinguishing true somatic events from artifacts arising from PCR, sequencing errors or mis-mapping. Tumor cellularity or purity, sub-clonality and copy number changes also confound the identification of true somatic events against a background of germline variants. We have developed a heuristic strategy and software (http://www.qcmg.org/bioinformatics/qsnp/) for somatic mutation calling in samples with low tumor content and we show the superior sensitivity and precision of our approach using a previously sequenced cell line, a series of tumor/normal admixtures, and 3,253 putative somatic SNVs verified on an orthogonal platform.

Overexpression of the antiapoptotic factor BCL-2 is a frequent feature of malignant disease and is commonly associated with poor prognosis and resistance to conventional chemotherapy. In breast cancer, however, high BCL-2 expression is associated with favorable prognosis, estrogen receptor (ER) positivity, and low tumor grade, whereas low expression is included in several molecular signatures associated with resistance to endocrine therapy. In the present study, we correlate BCL-2 expression and DNA methylation profiles in human breast cancer and in multiple cell models of acquired endocrine resistance to determine whether BCL-2 hypermethylation could provide a useful biomarker of response to cytotoxic therapy. In human disease, diminished expression of BCL-2 was associated with hypermethylation of the second exon, in a region that overlapped a CpG island and an ER-binding site. Hypermethylation of this region, which occurred in 10% of primary tumors, provided a stronger predictor of patient survival (P = 0.019) when compared with gene expression (n = 522). In multiple cell models of acquired endocrine resistance, BCL-2 expression was significantly reduced in parallel with increased DNA methylation of the exon 2 region. The reduction of BCL-2 expression in endocrine-resistant cells lowered their apoptotic threshold to antimitotic agents: nocodazole, paclitaxel, and the PLK1 inhibitor BI2536. This phenomenon could be reversed with ectopic expression of BCL-2, and rescued with the BCL-2 inhibitor ABT-737. Collectively, these data imply that BCL-2 hypermethylation provides a robust biomarker of response to current and next-generation cytotoxic agents in endocrine-resistant breast cancer, which may prove beneficial in directing therapeutic strategy for patients with nonresectable, metastatic disease.

Background Current staging methods for pancreatic cancer (PC) are inadequate, and biomarkers to aid clinical decision making are lacking. Despite the availability of the serum marker carbohydrate antigen 19.9 (CA19.9) for over two decades, its precise role in the management of PC is yet to be defined, and as a consequence, it is not widely used. Methods We assessed the relationship between perioperative serum CA19.9 levels, survival and adjuvant chemotherapeutic responsiveness in a cohort of 260 patients who underwent operative resection for PC. Results By specifically assessing the subgroup of patients with detectable CA19.9, we identified potential utility at key clinical decision points. Low postoperative CA19.9 at 3 months (median survival 25.6 vs 14.8 months, P = 0.0052) and before adjuvant chemotherapy were independent prognostic factors. Patients with postoperative CA 19.9 levels >90 U/ml did not benefit from adjuvant chemotherapy (P = 0.7194) compared with those with a CA19.9 of 5 U/ml) and have potential clinical utility in predicting outcome and response to adjuvant chemotherapy. Future clinical trials should prioritize incorporation of CA19.9 measurement at key decision points to prospectively validate these findings and facilitate implementation.

Pancreatic ductal adenocarcinoma (PDA) remains a lethal malignancy despite much progress concerning its molecular characterization. PDA tumours harbour four signature somatic mutations1, 2, 3, 4 in addition to numerous lower frequency genetic events of uncertain significance5. Here we use Sleeping Beauty (SB) transposon-mediated insertional mutagenesis6, 7 in a mouse model of pancreatic ductal preneoplasia8 to identify genes that cooperate with oncogenic KrasG12D to accelerate tumorigenesis and promote progression. Our screen revealed new candidate genes for PDA and confirmed the importance of many genes and pathways previously implicated in human PDA. The most commonly mutated gene was the X-linked deubiquitinase Usp9x, which was inactivated in over 50% of the tumours. Although previous work had attributed a pro-survival role to USP9X in human neoplasia9, we found instead that loss of Usp9x enhances transformation and protects pancreatic cancer cells from anoikis. Clinically, low USP9X protein and messenger RNA expression in PDA correlates with poor survival after surgery, and USP9X levels are inversely associated with metastatic burden in advanced disease. Furthermore, chromatin modulation with trichostatin A or 5-aza-2Œ-deoxycytidine elevates USP9X expression in human PDA cell lines, indicating a clinical approach for certain patients. The conditional deletion of Usp9x cooperated with KrasG12D to accelerate pancreatic tumorigenesis in mice, validating their genetic interaction. We propose that USP9X is a major tumour suppressor gene with prognostic and therapeutic relevance in PDA.

BACKGROUND: The receptor tyrosine kinase RON exhibits increased expression during pancreatic cancer progression and promotes migration, invasion and gemcitabine resistance of pancreatic cancer cells in experimental models. However, the prognostic significance of RON expression in pancreatic cancer is unknown. METHODS: RON expression was characterized in several large cohorts, including a prospective study, totaling 492 pancreatic cancer patients and relationships with patient outcome and clinico-pathologic variables were assessed. RESULTS: RON expression was associated with outcome in a training set, but this was not recapitulated in the validation set, nor was there any association with therapeutic responsiveness in the validation set or the prospective study. CONCLUSIONS: Although RON is implicated in pancreatic cancer progression in experimental models, and may constitute a therapeutic target, RON expression is not associated with prognosis or therapeutic responsiveness in resected pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDA) remains a lethal malignancy despite much progress concerning its molecular characterization. PDA tumours harbour four signature somatic mutations in addition to numerous lower frequency genetic events of uncertain significance. Here we use Sleeping Beauty (SB) transposon-mediated insertional mutagenesis in a mouse model of pancreatic ductal preneoplasia to identify genes that cooperate with oncogenic Kras(G12D) to accelerate tumorigenesis and promote progression. Our screen revealed new candidate genes for PDA and confirmed the importance of many genes and pathways previously implicated in human PDA. The most commonly mutated gene was the X-linked deubiquitinase Usp9x, which was inactivated in over 50% of the tumours. Although previous work had attributed a pro-survival role to USP9X in human neoplasia, we found instead that loss of Usp9x enhances transformation and protects pancreatic cancer cells from anoikis. Clinically, low USP9X protein and messenger RNA expression in PDA correlates with poor survival after surgery, and USP9X levels are inversely associated with metastatic burden in advanced disease. Furthermore, chromatin modulation with trichostatin A or 5-aza-2'-deoxycytidine elevates USP9X expression in human PDA cell lines, indicating a clinical approach for certain patients. The conditional deletion of Usp9x cooperated with Kras(G12D) to accelerate pancreatic tumorigenesis in mice, validating their genetic interaction. We propose that USP9X is a major tumour suppressor gene with prognostic and therapeutic relevance in PDA.

The Protein Interaction Network Analysis (PINA) platform is a comprehensive web resource, which includes a database of unified protein-protein interaction data integrated from six manually curated public databases, and a set of built-in tools for network construction, filtering, analysis and visualization. The second version of PINA enhances its utility for studies of protein interactions at a network level, by including multiple collections of interaction modules identified by different clustering approaches from the whole network of protein interactions ('interactome') for six model organisms. All identified modules are fully annotated by enriched Gene Ontology terms, KEGG pathways, Pfam domains and the chemical and genetic perturbations collection from MSigDB. Moreover, a new tool is provided for module enrichment analysis in addition to simple query function. The interactome data are also available on the web site for further bioinformatics analysis. PINA is freely accessible at http://cbg.garvan.unsw.edu.au/pina/.

Tumour cellularity, the relative proportion of tumour and normal cells in a sample, affects the sensitivity of mutation detection, copy number analysis, cancer gene expression and methylation profiling. Tumour cellularity is traditionally estimated by pathological review of sectioned specimens; however this method is both subjective and prone to error due to heterogeneity within lesions and cellularity differences between the sample viewed during pathological review and tissue used for research purposes. In this paper we describe a statistical model to estimate tumour cellularity from SNP array profiles of paired tumour and normal samples using shifts in SNP allele frequency at regions of loss of heterozygosity (LOH) in the tumour. We also provide qpure, a software implementation of the method. Our experiments showed that there is a medium correlation 0.42 ([Formula: see text]-value=0.0001) between tumor cellularity estimated by qpure and pathology review. Interestingly there is a high correlation 0.87 ([Formula: see text]-value [Formula: see text] 2.2e-16) between cellularity estimates by qpure and deep Ion Torrent sequencing of known somatic KRAS mutations; and a weaker correlation 0.32 ([Formula: see text]-value=0.004) between IonTorrent sequencing and pathology review. This suggests that qpure may be a more accurate predictor of tumour cellularity than pathology review. qpure can be downloaded from https://sourceforge.net/projects/qpure/.

The N-Myc oncoprotein is a critical factor in neuroblastoma tumorigenesis which requires additional mechanisms converting a low-level to a high-level N-Myc expression. N-Myc protein is stabilized when phosphorylated at Serine 62 by phosphorylated ERK protein. Here we describe a novel positive feedback loop whereby N-Myc directly induced the transcription of the class III histone deacetylase SIRT1, which in turn increased N-Myc protein stability. SIRT1 binds to Myc Box I domain of N-Myc protein to form a novel transcriptional repressor complex at gene promoter of mitogen-activated protein kinase phosphatase 3 (MKP3), leading to transcriptional repression of MKP3, ERK protein phosphorylation, N-Myc protein phosphorylation at Serine 62, and N-Myc protein stabilization. Importantly, SIRT1 was up-regulated, MKP3 down-regulated, in pre-cancerous cells, and preventative treatment with the SIRT1 inhibitor Cambinol reduced tumorigenesis in TH-MYCN transgenic mice. Our data demonstrate the important roles of SIRT1 in N-Myc oncogenesis and SIRT1 inhibitors in the prevention and therapy of N-Myc-induced neuroblastoma.

The International Cancer Genome Consortium (ICGC) was launched to coordinate large-scale cancer genome studies in tumours from 50 different cancer types and/or subtypes that are of clinical and societal importance across the globe. Systematic studies of more than 25,000 cancer genomes at the genomic, epigenomic and transcriptomic levels will reveal the repertoire of oncogenic mutations, uncover traces of the mutagenic influences, define clinically relevant subtypes for prognosis and therapeutic management, and enable the development of new cancer therapies.

Background & Aims: Current methods of preoperative staging and predicting outcome following pancreatectomy for pancreatic cancer (PC) are inadequate. We evaluated the utility of multiple biomarkers from distinct biologic pathways as potential predictive markers of response to pancreatectomy and patient survival. Methods: We assessed the relationship of candidate biomarkers known, or suspected, to be aberrantly expressed in PC, with disease-specific survival and response to therapy in a cohort of 601 patients. Results: Of the 17 candidate biomarkers examined, only elevated expression of S100A2 was an independent predictor of survival in both the training (n = 162) and validation sets (n = 439; hazard ratio [HR], 2.19; 95% confidence interval [CI]: 1.48-3.25; P < .0001) when assessed in a multivariate model with clinical variables. Patients with high S100A2 expressing tumors had no survival benefit with pancreatectomy compared with those with locally advanced disease, whereas those without high S100A2 expression had a survival advantage of 10.6 months (19.4 vs 8.8 months, respectively) and a HR of 3.23 (95% CI: 2.39-4.33; P < .0001). Of significance, patients with S100A2-negative tumors had a significant survival benefit from pancreatectomy even in the presence of involved surgical margins (median, 15.7 months; P = .0007) or lymph node metastases (median, 17.4 months; P = .0002). Conclusions: S100A2 expression is a good predictor of response to pancreatectomy for PC and suggests that high S100A2 expression may be a marker of a metastatic phenotype. Prospective measurement of S100A2 expression in diagnostic biopsy samples has potential clinical utility as a predictive marker of response to pancreatectomy and other therapies that target locoregional disease. © 2009 AGA Institute.

Identification of a biomarker of prognosis and response to therapy that can be assessed preoperatively would significantly improve overall outcomes for patients with pancreatic cancer. In this study, patients whose tumours exhibited high LMO4 expression had a significant survival advantage following operative resection, whereas the survival of those patients whose tumours had low or no LMO4 expression was not significantly different when resection was compared with operative biopsy alone.