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Background: Pancreatic ductal adenocarcinoma (PDA) is a lethal cancer with complex genomes and dense fibrotic stroma. This study was designed to identify clinically relevant somatic aberrations in pancreatic cancer genomes of patients with primary and metastatic disease enrolled and treated in two clinical trials.Methods: Tumour nuclei were flow sorted prior to whole genome copy number variant (CNV) analysis. Targeted or whole exome sequencing was performed on most samples. We profiled biopsies from 68 patients enrolled in two Stand Up to Cancer (SU2C)-sponsored clinical trials. These included 38 resected chemoradiation naïve tumours (SU2C 20206-003) and metastases from 30 patients who progressed on prior therapies (SU2C 20206-001). Patient outcomes including progression-free survival (PFS) and overall survival (OS) were observed.Results: We defined: (a) CDKN2A homozygous deletions that included the adjacent MTAP gene, only its' 3' region, or excluded MTAP; (b) SMAD4 homozygous deletions that included ME2; (c) a pancreas-specific MYC super-enhancer region; (d) DNA repair-deficient genomes; and (e) copy number aberrations present in PDA patients with long-term (⩾ 40 months) and short-term (⩽ 12 months) survival after surgical resection.Conclusions: We provide a clinically relevant framework for genomic drivers of PDA and for advancing novel treatments. [ABSTRACT FROM AUTHOR]

Liposarcoma is the most common soft tissue sarcoma, but little is known about the genomic basis of this disease. Given the low cell content of this tumor type, we utilized flow cytometry to isolate the diploid normal and aneuploid tumor populations from a well-differentiated liposarcoma prior to array comparative genomic hybridization and whole genome sequencing. This work revealed massive highly focal amplifications throughout the aneuploid tumor genome including MDM2, a gene that has previously been found to be amplified in well-differentiated liposarcoma. Structural analysis revealed massive rearrangement of chromosome 12 and 11 gene fusions, some of which may be part of double minute chromosomes commonly present in well-differentiated liposarcoma. We identified a hotspot of genomic instability localized to a region of chromosome 12 that includes a highly conserved, putative L1 retrotransposon element, LOC100507498 which resides within a gene cluster (NAV3, SYT1, PAWR) where 6 of the 11 fusion events occurred. Interestingly, a potential gene fusion was also identified in amplified DDR2, which is a potential therapeutic target of kinase inhibitors such as dastinib, that are not routinely used in the treatment of patients with liposarcoma. Furthermore, 7 somatic, damaging single nucleotide variants have also been identified, including D125N in the PTPRQ protein. In conclusion, this work is the first to report the entire genome of a well-differentiated liposarcoma with novel chromosomal rearrangements associated with amplification of therapeutically targetable genes such as MDM2 and DDR2. [ABSTRACT FROM AUTHOR]

Sarcomas are cancers that arise in soft tissues or bone and make up a small percentage of malignancies. In an effort to identify potential genetic targets for therapy, this study explores the genomic landscape of a metastatic undifferentiated pleomorphic sarcoma (UPS) with spindle cell morphology. Thick sections (50 μm) of formalin-fixed, paraffinembedded tissue from a primary, recurrent, and metastatic tumor were collected and processed from a single patient for DNA content- based flow-sorting and analyses. Nuclei of diploid and aneuploid populations were sorted from the malignant tissues and their genomes interrogated with array comparative genomic hybridization. The third sample was highly degraded and did not contain any intact ploidy peaks in our flow assays. A 2.5N aneuploid population was identified in the primary and recurrent sample. We detected a series of shared and unique genomic aberrations in the sorted aneuploid populations. The patterns of aberrations suggest that two similar but independent clonal populations arose during the clinical history of this rare tumor. None of these aberrations were detected in the matching sorted diploid samples. The targeted regions of interest might play a role in UPS and may lead to clinical significance with further investigation. [ABSTRACT FROM AUTHOR]

This article traces Romania's development to a low-tax country among the member states of the European Union by analysing the major tax law changes in corporate taxation since 1992. We compute and compare the effective company tax burdens of a European model company at different points of time which constitute milestones in the development of corporate taxation in Romania. We find that the significant reduction of the corporate income tax rate from 45% in 1992 to 16% since 2005 has not been accompanied by a comprehensive broadening of the corporate income tax base as it has been in many longstanding member states of the EU and the OECD. Since 1992 the average company tax burden of the underlying model company has dropped by almost 65%. As a result, Romania holds second position among the group of Central and Eastern European EU member states. [ABSTRACT FROM AUTHOR]

Formalin fixed paraffin embedded (FFPE) tissues are a vast resource of annotated clinical samples. As such, they represent highly desirable and informative materials for the application of high definition genomics for improved patient management and to advance the development of personalized therapeutics. However, a limitation of FFPE tissues is the variable quality of DNA extracted for analyses. Furthermore, admixtures of non-tumor and polyclonal neoplastic cell populations limit the number of biopsies that can be studied and make it difficult to define cancer genomes in patient samples. To exploit these valuable tissues we applied flow cytometry-based methods to isolate pure populations of tumor cell nuclei from FFPE tissues and developed a methodology compatible with oligonucleotide array CGH and whole exome sequencing analyses. These were used to profile a variety of tumors (breast, brain, bladder, ovarian and pancreas) including the genomes and exomes of matching fresh frozen and FFPE pancreatic adenocarcinoma samples. [ABSTRACT FROM AUTHOR]

So far, the field of cross-border taxation of dividends paid to individual investors in the European Union (EU) Member States has not been subject to harmonization. The differences in the Member States' tax systems give rise to obstacles to the internal market. As a consequence, individual investors may be discouraged from investing in shares of foreign corporations. Hence, the benefits of international risk diversification are not fully exploited. The discussion of possible solutions to these issues has recently received a new impetus as the European Commission held a consultation on the issues of cross-border dividend taxation in Spring 2011. In this article, we discuss the alternative solutions pointed out by the European Commission and present our genuine proposal. We show that dividends paid to individual investors should not be taxed at source by drawing on a set of normative criteria comprising conformity with European Law, efficiency, simplicity, and equity considerations. We furthermore argue that the abolition of withholding taxes should be accompanied by introducing automatic exchange of information. The analysis of alternative reform options is preceded by a brief, yet comprehensive summary of the taxation of dividends in the EU Member States highlighting the shortcomings of the current dividend taxation. [ABSTRACT FROM AUTHOR]

Pancreatic ductal adenocarcinoma (PDAC) can be divided into transcriptomic subtypes with two broad lineages referred to as classical (pancreatic) and squamous. We find that these two subtypes are driven by distinct metabolic phenotypes. Loss of genes that drive endodermal lineage specification, HNF4A and GATA6 , switch metabolic profiles from classical (pancreatic) to predominantly squamous, with glycogen synthase kinase 3 beta (GSK3β) a key regulator of glycolysis. Pharmacological inhibition of GSK3β results in selective sensitivity in the squamous subtype; however, a subset of these squamous patient-derived cell lines (PDCLs) acquires rapid drug tolerance. Using chromatin accessibility maps, we demonstrate that the squamous subtype can be further classified using chromatin accessibility to predict responsiveness and tolerance to GSK3β inhibitors. Our findings demonstrate that distinct patterns of chromatin accessibility can be used to identify patient subgroups that are indistinguishable by gene expression profiles, highlighting the utility of chromatin-based biomarkers for patient selection in the treatment of PDAC. • HNF4A loss upregulates GSK3β and drives a squamous-like metabolic profile • GSK3β targeting inhibits glycolysis in squamous patient-derived cell lines (PDCLs) • A subset of squamous PDCLs acquires GSK3β drug tolerance • ATAC-seq analysis reveals an accessible WNT gene program in drug-tolerant PDCLs Brunton et al. demonstrate that differential chromatin accessibility can predict responsiveness and tolerance to GSK3β inhibitors in the squamous subtype of PDAC. This study provides an important proof of concept that chromatin accessibility can be used to identify additional PDAC subgroups with potential therapeutic utility. [ABSTRACT FROM AUTHOR]

Introduction We sought to identify the genomic abnormalities in squamous cell carcinomas (SCC) arising in ovarian mature cystic teratoma (MCT, also known as dermoid cyst), a rare gynaecological malignancy of poor prognosis. Materials /Patients and methods We performed copy number, mutational state and zygosity analysis of 151 genes in SCC arising in MCT (n=25) using next-generation sequencing. The presence of high/intermediate risk HPV genotypes was assessed by quantitative PCR. Whole genome sequencing (WGS) was completed on 2 MCT cases and all genomic events were correlated with clinical features and outcome. Results MCT had a low mutation burden with a mean of only 1 mutation per case. WGS on 2 MCT cases revealed no driver mutations or rearrangements. Zygosity analyses of MCT indicated four separate patterns, suggesting that MCT can arise from errors at various stages of oogenesis. A total of 244 abnormalities were identified in 79 genes in MCTassociated SCC, and the overall mutational burden was high (mean 10.2 mutations per megabase). No SCC was positive for HPV. The most frequently altered genes in SCC were TP53 (20/25 cases, 80%), PIK3CA (13/25 cases, 52%) and CDKN2A (11/25 cases, 44%). Mutation in TP53 was associated with improved overall survival. In 8/20 cases with TP53 mutations, two or more variants were identified, which were bi-allelic. Conclusions Ovarian MCT has low mutation burden. By contrast, ovarian SCC arising in MCT has a high mutational burden with TP53 mutation the most common abnormality. The presence TP53 mutation is a good prognostic factor. SCC arising in MCT share similar mutation profiles to other SCC. Given their rarity, they should be included in basket studies that recruit patients with SCC of other organs. [ABSTRACT FROM AUTHOR]

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., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: Glioblastoma is the most frequent and lethal primary brain tumor. Development of novel therapies relies on the availability of relevant preclinical models. We have established a panel of 96 glioblastoma patient-derived xenografts (PDX) and undertaken its genomic and phenotypic characterization., Experimental Design: PDXs were established from glioblastoma, IDH-wildtype ( n = 93), glioblastoma, IDH-mutant ( n = 2), diffuse midline glioma, H3 K27M-mutant ( n = 1), and both primary ( n = 60) and recurrent ( n = 34) tumors. Tumor growth rates, histopathology, and treatment response were characterized. Integrated molecular profiling was performed by whole-exome sequencing (WES, n = 83), RNA-sequencing ( n = 68), and genome-wide methylation profiling ( n = 76). WES data from 24 patient tumors was compared with derivative models., Results: PDXs recapitulate many key phenotypic and molecular features of patient tumors. Orthotopic PDXs show characteristic tumor morphology and invasion patterns, but largely lack microvascular proliferation and necrosis. PDXs capture common and rare molecular drivers, including alterations of TERT, EGFR, PTEN, TP53, BRAF , and IDH1 , most at frequencies comparable with human glioblastoma. However, PDGFRA amplification was absent. RNA-sequencing and genome-wide methylation profiling demonstrated broad representation of glioblastoma molecular subtypes. MGMT promoter methylation correlated with increased survival in response to temozolomide. WES of 24 matched patient tumors showed preservation of most genetic driver alterations, including EGFR amplification. However, in four patient-PDX pairs, driver alterations were gained or lost on engraftment, consistent with clonal selection., Conclusions: Our PDX panel captures the molecular heterogeneity of glioblastoma and recapitulates many salient genetic and phenotypic features. All models and genomic data are openly available to investigators., (©2019 American Association for Cancer Research.)

Next-generation sequencing is enabling molecularly guided therapy for many cancer types, yet failure rates remain relatively high in pancreatic cancer (PC). The aim of this study is to investigate the feasibility of genomic profiling using endoscopic ultrasound (EUS) biopsy samples to facilitate personalised therapy for PC. Ninty-five patients underwent additional research biopsies at the time of diagnostic EUS. Diagnostic formalin-fixed (FFPE) and fresh frozen EUS samples underwent DNA extraction, quantification and targeted gene sequencing. Whole genome (WGS) and RNA sequencing was performed as proof of concept. Only 2 patients (2%) with a diagnosis of PC had insufficient material for targeted sequencing in both FFPE and frozen specimens. Targeted panel sequencing (n=54) revealed mutations in PC genes (KRAS, GNAS, TP53, CDKN2A, SMAD4) in patients with histological evidence of PC, including potentially actionable mutations (BRCA1, BRCA2, ATM, BRAF). WGS (n=5) of EUS samples revealed mutational signatures that are potential biomarkers of therapeutic responsiveness. RNA sequencing (n=35) segregated patients into clinically relevant molecular subtypes based on transcriptome. Integrated multi-omic analysis of PC using standard EUS guided biopsies offers clinical utility to guide personalized therapy and study the molecular pathology in all patients with PC.

Purpose: We sought to identify the genomic abnormalities in squamous cell carcinomas (SCC) arising in ovarian mature cystic teratoma (MCT), a rare gynecological malignancy of poor prognosis. Experimental design: We performed copy number, mutational state, and zygosity analysis of 151 genes in SCC arising in MCT ( n = 25) using next-generation sequencing. The presence of high-/intermediate-risk HPV genotypes was assessed by quantitative PCR. Genomic events were correlated with clinical features and outcome. Results: MCT had a low mutation burden with a mean of only one mutation per case. Zygosity analyses of MCT indicated four separate patterns, suggesting that MCT can arise from errors at various stages of oogenesis. A total of 244 abnormalities were identified in 79 genes in MCT-associated SCC, and the overall mutational burden was high (mean 10.2 mutations per megabase). No SCC was positive for HPV. The most frequently altered genes in SCC were TP53 (20/25 cases, 80%), PIK3CA (13/25 cases, 52%), and CDKN2A (11/25 cases, 44%). Mutation in TP53 was associated with improved overall survival. In 8 of 20 cases with TP53 mutations, two or more variants were identified, which were bi-allelic. Conclusions: Ovarian SCC arising in MCT has a high mutational burden, with TP53 mutation the most common abnormality. The presence of TP53 mutation is a good prognostic factor. SCC arising in MCT share similar mutation profiles to other SCC. Given their rarity, they should be included in basket studies that recruit patients with SCC of other organs. Clin Cancer Res; 23(24); 7633-40. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Liposarcoma is the most common soft tissue sarcoma, but little is known about the genomic basis of this disease. Given the low cell content of this tumor type, we utilized flow cytometry to isolate the diploid normal and aneuploid tumor populations from a well-differentiated liposarcoma prior to array comparative genomic hybridization and whole genome sequencing. This work revealed massive highly focal amplifications throughout the aneuploid tumor genome including MDM2, a gene that has previously been found to be amplified in well-differentiated liposarcoma. Structural analysis revealed massive rearrangement of chromosome 12 and 11 gene fusions, some of which may be part of double minute chromosomes commonly present in well-differentiated liposarcoma. We identified a hotspot of genomic instability localized to a region of chromosome 12 that includes a highly conserved, putative L1 retrotransposon element, LOC100507498 which resides within a gene cluster (NAV3, SYT1, PAWR) where 6 of the 11 fusion events occurred. Interestingly, a potential gene fusion was also identified in amplified DDR2, which is a potential therapeutic target of kinase inhibitors such as dastinib, that are not routinely used in the treatment of patients with liposarcoma. Furthermore, 7 somatic, damaging single nucleotide variants have also been identified, including D125N in the PTPRQ protein. In conclusion, this work is the first to report the entire genome of a well-differentiated liposarcoma with novel chromosomal rearrangements associated with amplification of therapeutically targetable genes such as MDM2 and DDR2.

Tumors frequently arise as a result of an acquired genomic instability and the subsequent evolution of neoplastic populations with variable genomes. A barrier to the study of the somatic genetics of human solid tumors in vivo is the presence of admixtures of non-neoplastic cells with normal genomes in patient samples. These can obscure the presence of somatic aberrations including mutations, homozygous deletions, and breakpoints in biopsies of interest. Furthermore, clinical samples frequently contain multiple neoplastic populations that cannot be distinguished by morphology. Consequently, it is difficult to determine whether mutations detected in a sample of interest are concurrent in a single clonal population or if they occur in distinct cell populations in the same sample. The advent of targeted therapies increases the selection for preexisting populations. However the asymmetric distribution of therapeutic targets in clonal populations provides a mechanism for the rapid evolution of resistant disease. Thus, there is a need to not only isolate tumor from normal cells, but to also enrich distinct populations of clonal neoplastic cells in order to apply genome technologies to identify clinically relevant genomic aberrations that drive disease in patients in vivo. To address this we have applied single and multiparameter DNA content based flow assays to the study of solid tumors. Our work has identified examples of clonal resistance to effective therapies. This includes androgen withdrawal in advanced prostate cancer. In addition we demonstrate examples of co-existing clonal populations with highly aberrant genomes and ploidies in a wide variety of solid tumors. We propose that clonal analysis of tumors, based on flow cytometry and high resolution genome analyses of purified neoplastic populations, provides a unique approach to the study of therapeutic responses and the evolution of resistance.

It is widely accepted that many cancers arise as a result of an acquired genomic instability and the subsequent evolution of tumor cells with variable patterns of selected and background aberrations. The presence and behaviors of distinct neoplastic cell populations within a patient's tumor may underlie multiple clinical phenotypes in cancers. A goal of many current cancer genome studies is the identification of recurring selected driver events that can be advanced for the development of personalized therapies. Unfortunately, in the majority of rare tumors, this type of analysis can be particularly challenging. Large series of specimens for analysis are simply not available, allowing recurring patterns to remain hidden. In this paper, we highlight the use of DNA content-based flow sorting to identify and isolate DNA-diploid and DNA-aneuploid populations from tumor biopsies as a strategy to comprehensively study the genomic composition and behaviors of individual cancers in a series of rare solid tumors: intrahepatic cholangiocarcinoma, anal carcinoma, adrenal leiomyosarcoma, and pancreatic neuroendocrine tumors. We propose that the identification of highly selected genomic events in distinct tumor populations within each tumor can identify candidate driver events that can facilitate the development of novel, personalized treatment strategies for patients with cancer.