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Background/aim: As an increasing number of drugs are approved for targeted cancer therapy, comprehensive genomic profiling of cancer patients is frequently conducted to identify potentially relevant genetic variants. Different sequencing technologies are used for this purpose; targeted gene panels cover a selected set of biomarker genes and hotspot regions, while whole genome sequencing (WGS) delivers genome-wide data. This comparison study aimed at evaluating whether one method performs superiorly to the other regarding the detection of targetable variants., Patients and Methods: To evaluate the performance of the two sequencing technologies, we compared the results of targeted sequencing using the Ion Torrent Oncomine Comprehensive Assay Plus (OCA-Plus) panel to Illumina WGS reports in 11 patients diagnosed with pancreatic cancer (PC). All pathogenic and likely pathogenic variants, including those relevant for targeted therapy, reported by WGS and OCA-Plus, were included in the final comparison., Results: Both techniques identified common driver mutations implicated in PC with high concordance (81%) across all variants. For variants relevant to targeted therapy, a 100% concordance between the technologies was observed., Conclusion: A comparable number of variants were reported by WGS and OCA-Plus, and all genetic variants relevant for targeted therapy were identified by both technologies. Thus, WGS does not provide substantial additional information in this patient group., (Copyright © 2025 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background: Small Bowel Adenocarcinoma (SBA) is a rare gastrointestinal cancer with a limited understanding of the molecular pathology. This study aims to bridge the knowledge gap, providing a robust molecular foundation for SBA and addressing the clinical challenges inherent in treating this orphan disease. The study proposes to redefine the clinical management for SBA patients through advanced molecular profiling techniques to improve potential precision medicine., Methods/design: This National multicenter, observational cohort study combines retrospective and prospective analyses across Danish University Hospitals. The study enrolls patients diagnosed with SBA, retrospectively from 2009 and prospectively from 2022 onwards. Molecular profiling, including DNA, RNA, and T-cell receptor sequencing, will be conducted on SBA tissue samples. The primary outcome is to categorize SBA into consensus molecular-guided subgroups. Secondary outcomes include correlating these subgroups with clinical features, treatment responses, and patient outcomes. Machine learning algorithms will be employed for bioinformatic analyses to interpret molecular data. Ethical approval has been obtained, and patient consent will be secured for the retrospective study component., Discussion: The molecular and clinical characterization of SBA is expected to add novel insights into the heterogeneity of this rare disease. By identifying molecular subgroups, the research could enable the development of personalized treatment strategies, a paradigm shift within SBA. The study acknowledges the challenges of working with orphan diseases, including limited patient numbers and diverse clinical presentations. However, its findings will have the potential to substantially impact future clinical practices and guide targeted therapies for SBA patients., Trial Registration: ClinicalTrials.gov NCT06234306., Competing Interests: Declarations. Ethics approval and consent to participate: This study is approved by the Danish National Ethics Committee as of 23rd October 2023 (case number: 2312342). Signed informed consent will be required from all prospectively included patients. The study has received dispensation from signed informed consent to include patients treated at the participating departments from 1st January 2010. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

The ovarian oncobiome is subject to increasing scientific focus, but a potential link between bacterial dysbiosis and ovarian carcinogenesis remains controversial. Our primary aim was to characterize the bacterial microbiota in epithelial ovarian cancer samples. Secondarily, we aimed to compare results from the bacterial microbiota in formalin-fixed, paraffin-embedded ovarian tissue samples from 194 patients with epithelial ovarian cancer, fallopian tube tissue samples from 16 patients with serous tubal intraepithelial carcinomas and in benign fallopian tube tissue samples from 25 patients. Bacterial abundance was investigated by means of 16S rDNA Next Generation Sequencing. The 16S rDNA sequencing run produced a very low number of bacterial reads. Only seven samples displayed bacterial reads above 5000. Validation of detected bacterial reads by qPCR was negative and indicative of results being background amplification. Our results indicate no amount of bacterial biomass in the ovarian cancer, benign fallopian tube and in the samples with serous tubal intraepithelial carcinomas. The findings underline the importance of validating results from bacterial microbiota studies with additional technical assays before any conclusion may be drawn., (© 2025 Scandinavian Societies for Pathology, Medical Microbiology and Immunology.)

Background/aim: Adult granulosa cell tumor (aGCT) is a rare and challenging ovarian tumor due to its unpredictable recurrence and its associated increased risk of breast and endometrial cancer. Identifying and describing molecular alterations in tumors has become common with the advent of high-throughput sequencing. However, DNA sequencing in rare tumors, such as aGCT, often lacks statistical power due to the limited number of cases in each study, thereby clinical implications of DNA alterations are difficult to interpretate. This scoping review aims to systematically describe somatic and germline DNA alterations identified in women with aGCT., Materials and Methods: Search terms (granulosa cell tumour AND molecular alterations) were searched in May 2024 in the following databases: MEDLINE (Ovid), Embase (Ovid), Web of Science Core Collection and Google Scholar. Screening, full-text review and data extraction were performed by two independent reviewers., Results: Twenty-four publications were identified. Eighteen reported on somatic DNA alterations of patholgenic mutations identified in total 1,226 tissues being sequenced. FOXL2 (c.402C>G; p.C134W) was present in 97% of aGCTs. Other pathogenic mutations in the tissues investigated were TERT promoter mutation (41%), truncating KMT2D mutations (14%) and TP53 pathogenic variant (4%). TERT promoter mutation was reported more frequently in recurrent tumors (p<0.01), whereas comparing truncating KMT2D and TP53 mutations reported in primary and recurrent tumors revealed no difference (p=0.15 and p=0.26 respectively). Tumor mutational burden (TMB) was reported in five studies and all showed a low TMB. None of the somatic mutations were candidate targets for biological treatment. Six publications reported germline variants and no shared germline pathogenic variants were described in the published literature., Conclusion: The FOXL2 missense mutation was the only common somatic DNA alteration in aGCT. TERT promoter mutations were reported more frequently in recurrent aGCT but their clinical relevance remains uncertain. In contrast to previous reports, truncating KMT2D mutations were not found to be associated with recurrent aGCT. Evidence on common germline variants in aGCT is sparse. The role of somatic and germline DNA alterations in the development of other malignancies in women with aGCT remains uncertain. Further research involving matched primary and recurrent tumors, as well as other primary malignancies, is essential to better understand the mutations that drive tumor development., (Copyright © 2025 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Introduction: Testing for homologous recombination deficiency (HRD) as a biomarker in relation to poly (ADP-ribose) polymerase inhibitor (PARPi) treatment in ovarian cancer is done by sequencing of the BRCA1/2 genes and/or by assessing a genomic instability signature. Here we present data obtained with two different methods for genomic instability testing: the Oncomine™ Comprehensive Assay Plus (OCA Plus) NGS panel and the OncoScan CNV assay., Methods: The retrospective analytical study included 80 ovarian cancer samples of patients previously referred to clinical Myriad testing (reference cohort), and 50 ovarian cancer samples from patients collected as part of the Pelvic Mass study. OCA Plus NGS libraries were sequenced with the Ion S5™XL Sequencer and analyzed with the Ion Reporter™ Software v5.20 for calculation of the genomic instability metric (GIM). In addition, all samples were tested with the OncoScan CNV FFPE Assay and analyzed with a previously published R-algorithm for generation of an in-house genomic instability score (in-house GIS)., Results: The OCA Plus assay had a concordance to the reference of 89% on samples with a tumor fraction ≥ 30% (auto-calculated or via molecular estimation). A total of 15 samples in the reference cohort had a calculated tumor fraction < 30% in the OCA Plus assay. In these, the concordance to reference was only 60%. For the OncoScan CNV in-house GIS a local cutoff point of ≥ 50 was calculated. This gave a concordance to the reference of 85%, with 91% of the samples in the reference cohort passing quality control (QC) on tumor fraction. Both assays had a high sensitivity for the detection of genomic instability in samples with pathogenic or likely pathogenic BRCA1/2 mutations, with 12/13 being GIM positive (OCA Plus assay) and 13/13 being in-house GIS positive (OncoScan CNV assay)., Conclusions: The OCA Plus assay and the OncoScan CNV assay show a high but not complete concordance to reference standard homologous recombination deficiency (HRD) detection. The main reason for QC failure or non-concordance in our study was a low tumor fraction estimated in the assay, despite the selection of material by a pathologist with an inclusion criterion of > 30% tumor. QC steps should include careful tumor content evaluation, and results on samples with < 30% tumor should not be reported., Competing Interests: Declarations. Funding: Open access funding provided by Copenhagen University. Conflicts of Interest/Competing Interests (include appropriate disclosures): L.S., T.S.P., L.K.V., I.J.C., E.H.: None. Ethics Approval: This retrospective study was conducted on already available data or biological material and approved by The Danish National Committee on Health Research Ethics (Case ID 2212528) and the Danish Capital Region Knowledge Center for Data Reviews (Case ID P-2023-13). Consent for Publication/Participation: Not applicable. Availability of Data and Material (Data Transparency): The datasets generated during and/or analyzed during the current study are not publicly available, which is in accordance with the rules concerning the processing of personal data set out in the EU General Data Protection Regulation (GDPR) and the Danish Data Protection Act. Nonetheless, may a researcher have an interest in our data, they are welcome to contact us for collaborations. The data that support the findings of this study can be requested from The National Secretariat for Bio- and Genome Bank Denmark, RBGB.sekretariat.herlev-og-gentofte-hospital@regionh.dk, Herlev Hospital, Borgmester Ib Juuls Vej 73, 2730 Herlev, Denmark. Author Contributions: Estrid Høgdall, Lone Schejbel, and Tim Svenstrup Poulsen contributed to the study conception and design. Next generation sequencing and data collection and analysis were performed by Lone Schejbel and Lau Kræsing Vestergaard. Statistical analyses were performed by Ib Jarle Christensen. The first draft of the manuscript was written by Lone Schejbel and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Code Availability (Software Application or Custom Code): Not applicable., (© 2024. The Author(s).)

Coronary CT angiography (CTA) derived fractional flow reserve (FFR CT ) is recommended for physiological assessment in intermediate coronary stenosis for guiding referral to invasive coronary angiography (ICA). In this study, we report real-world data on the feasibility of implementing a CTA/FFR CT test algorithm as a gatekeeper to ICA at referral hospitals. Retrospective all-comer study of patients with new onset stable symptoms and suspected coronary stenosis (30-89%) by CTA. Evaluation of CTA datasets, interpretation of FFR CT analysis, and decisions on downstream testing were performed by skilled CT-cardiologists. CTA was performed in 3974 patients, of whom 381 (10%) were referred directly to ICA, whereas 463 (12%) to non-invasive functional testing: FFR CT 375 (81%) and perfusion imaging 88 (19%). FFR CT analysis was rejected in 8 (2%) due to inadequate CTA image quality. Number of patients deferred from ICA after FFR CT was 267 (71%), while 100 (27%) were referred to ICA. Obstructive coronary artery disease (CAD) was confirmed in 62 (62%) patients and revascularization performed in 53 (53%). Revascularization rates, n (%), were higher in patients undergoing FFR CT -guided versus CTA-guided referral to ICA: 30-69% stenosis, 28 (44%) versus 8 (21%); 70-89% stenosis, 39 (69%) versus 25 (46%), respectively, both p < 0.05. Implementation of FFR CT at referral hospitals was feasible, reduced the number of invasive procedures, and increased the revascularization rate., (© 2024. The Author(s).)

Background/aim: The emergence of novel DNA damage repair (DDR) pathways in molecular-target therapy drugs (MTTD) has shown promising outcomes in treating patients with metastatic castration-resistant prostate cancer (mCRPC). About 25% of mCRPC patients have actionable deleterious aberrations in DDR genes, primarily in the homologous recombination (HR) pathway. However, the response rate in patients with BRCA1/2 or mutations in HRR-related genes is only 45%-55%, when exposed to poly ADP ribose polymerase (PARP) inhibitor-based therapy (PARPi). A frequent characteristic feature of prostate cancer (PC) is the occurrence of genomic rearrangement that affects the transmembrane protease serine 2 (TMPRSS2) and E26 transformation-specific (ETS)- transcription factor-related gene (ERG)., Materials and Methods: In this study, a total of 114 patients with mCRPC had their RNA and DNA sequenced using next-generation sequencing., Results: Based on their genetic profile of deleterious gene alterations of BRCA1/2 or ATM, six patients were selected for PARPi. Patients with TMPRSS2:ERG gene fusion and homozygous alteration in ATM or BRCA2 (n=2) or heterozygous alterations (BRCA1 or BRCA2) and lack of TMPRSS2:ERG gene fusion (n=2) did not show clinical benefit from PARPi (treatment duration <16 weeks). In contrast, patients (n=2) without TMPRSS2:ERG gene fusion and homozygous deleterious alterations in ATM or BRCA2 all had clinical benefit from PARPi (treatment duration ≥16 weeks)., Conclusion: The TMPRSS2:ERG transcript product might be used as a PARPi resistance biomarker., (Copyright © 2024 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Purpose: The purpose of this study was to evaluate the clinical benefits of nivolumab with/without ipilimumab combined with stereotactic body radiotherapy (SBRT) in patients with pretreated metastatic biliary tract cancer (mBTC)., Patients and Methods: The study was a phase 2 randomized trial with Simon's optimal two-stage design requiring 36 evaluable patients per group after second stage. Sixty-one patients were included from September 2018 to January 2022 and randomized (1:1) to receive SBRT (15 Gy × 1 on day 1 to a primary or metastatic lesion) and nivolumab (3 mg/kg intravenously on day 1 and every 2 weeks) with/without ipilimumab (1 mg/kg intravenously on day 1 and every 6 weeks). Primary endpoint was clinical benefit rate (CBR), defined as the percentage of patients with complete response, partial response, or stable disease. Decision to continue accrual into the second stage depended on the CBR from the first stage., Results: Forty-two patients received SBRT/nivolumab/ipilimumab with a CBR of 31.0% [95% confidence interval (CI), 17.6-47.1]. Five patients (11.9%) achieved partial response with median duration of 4.4 months (range, 1.1-21.5). Nineteen patients received SBRT/nivolumab. This group was closed after the initial stage based on a CBR of 10.5% (95% CI, 1.3-33.1). Adverse events were graded with National Cancer Institute Common Terminology Criteria for Adverse Events version 4.0. Grade ≥3 treatment-related adverse events occurred in 13 (31%) and 3 (16%) patients in the SBRT/nivolumab/ipilimumab and SBRT/nivolumab groups, respectively. One patient died from immune-related hepatitis in the SBRT/nivolumab/ipilimumab group., Conclusions: Combining SBRT, nivolumab, and ipilimumab is well tolerated, feasible, and shows response in a subgroup of patients with mBTC., (©2024 American Association for Cancer Research.)

Background: Next-generation sequencing (NGS) has been implemented in clinical oncology as a personalized medicine tool to identify targetable genetic alterations and to guide treatment decisions. However, the optimal NGS test strategy and target genes for clinical use are still being discussed. The aim was to compare the performance of the Oncomine™ Comprehensive Assay v3 (OCAv3) (targeted gene panel) and whole-exome sequencing (WES) to investigate somatic single and multiple nucleotide variants and small indels in ovarian cancer patients., Methods and Results: Genomic DNA was isolated from fresh frozen samples of five high-grade serous (HGSC) and three clear cell ovarian (oCCC) cancer patients. Exome sequencing libraries were prepared by using the Ion AmpliSeq Exome RDY kit, whereas libraries for OCAv3 were prepared using by Ion AmpliSeq™ Library Kit Plus. Sequencing was performed using the Ion S5XL System (Thermo Fisher Scientific). When including only variants classified as pathogenic, likely pathogenic or unknown significance based on ClinVar database verdicts and comparing overlapping regions covered both by the OCAv3 assay and WES, 23 variants were detected by both assays. However, OCAv3 detected additionally two variants: ARID1A: p.Gln563Ter and TP53: p.Ser261ValfsTer84 that have not passed WES filtering criteria due to low coverage., Conclusions: With the present treatment possibilities, OCAv3 panel testing provided higher diagnostic yield due to better coverage. Our study emphasizes that WES, although offering the potential to identify novel findings in genes not covered by OCAv3, might overlook variants in genes relevant for OC., (© 2024. The Author(s).)

Background: Identifying and describing molecular alterations in tumors has become common with the development of high-throughput sequencing. However, DNA sequencing in rare tumors, such as ovarian adult granulosa cell tumor (aGCT), often lacks statistical power due to the limited number of cases in each study. Questions regarding personalized treatment or prognostic biomarkers for recurrence or other malignancies therefore still need to be elucidated. This scoping review protocol aims to systematically map the current evidence and identify knowledge gaps regarding DNA alterations, actionable variations and prognostic biomarkers in aGCT., Methods: This scoping review will be conducted based on Arksey and O'Malley's methodological framework and later modifications by JBI Evidence Synthesis. The protocol complies with Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for scoping reviews. All original publications describing molecular alterations of aGCT will be included. The search will be performed in May 2024 in the following databases: MEDLINE (Ovid), Embase (Ovid), Web of Science Core Collection and Google Scholar (100-top ranked)., Discussion: This scoping review will identify knowledge and gaps in the current understanding of the molecular landscape of aGCT, clinical trials on actionable variations and priorities for future research. As aGCT are rare, a possible limitation will be the small sample sizes and heterogenic study settings., Scoping Review Registration: The review protocol is registered at Open Science Framework under https://doi.org/10.17605/OSF.IO/PX4MF., Competing Interests: The authors have declared that no competing interests exist, (Copyright: © 2024 Karstensen et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Differential diagnosis of inflammatory bowel disease (IBD) to Crohn's disease (CD) or ulcerative colitis (UC) is crucial for treatment decision making. With the aim of generating a clinically applicable molecular-based tool to classify IBD patients, we assessed whole transcriptome analysis on endoscopy samples. A total of 408 patient samples were included covering both internal and external samples cohorts. Whole transcriptome analysis was performed on an internal cohort of FFPE IBD samples (CD, n = 16 and UC, n = 17). The 100 most significantly differentially expressed genes (DEG) were tested in two external cohorts. Ten of the DEG were further processed by functional enrichment analysis from which seven were found to show consistent significant performance in discriminating CD from UC: PI3, ANXA1, VDR, MTCL1, SH3PXD2A-AS1, CLCF1, and CD180. Differential expression of PI3, ANXA1, and VDR was reproduced by RT-qPCR, which was performed on an independent sample cohort of 97 patient samples (CD, n = 44 and UC, n = 53). Gene expression levels of the three-gene profile, resulted in an area under the curve of 0.84 (P = 0.02) in discriminating CD from UC, and therefore appear as an attractive molecular-based diagnostic tool for clinicians to distinguish CD from UC., (© 2023. The Author(s).)

Background/aim: Several clinical trials have investigated homologous recombination deficiency and BRCA1/2 status to select ovarian cancer patients for treatment with poly(ADP-ribose) polymerase-inhibitors (PARPi), but less attention has been given to other DNA-damage response (DDR) pathways. Therefore, we investigated somatic single/multiple nucleotide variants and small insertions/deletions in exonic and splice-site regions of 356 DDR genes to examine whether genes other than BRCA1/2 are altered., Materials and Methods: Whole-exome sequencing data from eight high-grade serous adenocarcinoma (HGSC) and four clear cell carcinoma (oCCC) patients were analyzed., Results: Forty-two variants (pathogenic, likely pathogenic or variants of uncertain significance) in 28 genes from DDR pathways were identified. Seven out of nine TP53 variants were previously described in The Cancer Genome Atlas Ovarian Cancer; other variants were found in 23 out of 28 unique genes, whereas no variants were reported in FAAP24, GTF2H4, POLE4, RPA3, and XRCC4., Conclusion: As the identified variants were not only limited to well-known TP53, BRCA1/2, and HR-associated genes, our study might contribute to the better understanding of which DDR pathways potentially influence disease progression. Moreover, they may display a potential role as biomarkers to predict platinum-based chemotherapy or PARPi treatment response or disease progression, as differences in disrupted DDR pathways were observed between patients with long and short overall survival in HGSC and oCCC groups., (Copyright © 2023 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background/aim: Gastrointestinal stromal tumours (GISTs) harbour genetic aberrations in receptor tyrosine kinase KIT (KIT) or platelet-derived growth factor receptor A (PDGFRA) in 85-90% of the patients. Circulating tumour DNA (ctDNA) is a potential biomarker in patients with GIST. Previous studies investigating ctDNA around surgery in patients with GIST presented divergent results regarding the proportion of patients with detectable ctDNA. This study aimed to 1) investigate the feasibility of detecting and monitoring ctDNA pre-and postoperative, 2) compare two different circulating free DNA (cfDNA) extraction methods, and validate results obtained by next-generation sequencing (NGS) using Real-Time PCR technology., Patients and Methods: Eight patients planned for immediate surgery or surgery after neoadjuvant oncological treatment were included in the study, from whom blood collection was performed pre- and postoperatively for ctDNA analysis. Furthermore, blood samples from six patients with GIST harbouring a point mutation in KIT or PDGFRA in tissues from primary tumours were used for comparison and validation sub-study., Results: In this explorative study, none of the patients with very low to intermediate risk GIST harboured KIT, or PDGFRA mutated ctDNA in pre-or postoperative blood samples. The methods used for cfDNA extraction gave similar output, and the two methods for ctDNA analysis gave identical results., Conclusion: There is no benefit in analysing ctDNA around surgery in very low to intermediate-risk GIST patients. Larger studies investigating ctDNA in patients with high-risk GIST around surgery are warranted., (Copyright © 2022 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background: We aimed to identify somatic pathogenic and likely pathogenic mutations using next-generation sequencing (NGS). The mutational findings were held against clinically well-described data to identify potential targeted therapies in Danish patients diagnosed with high-grade serous ovarian cancer (HGSC)., Methods: We characterized the mutational profile of 128 HGSC patients. Clinical data were obtained from the Danish Gynecological Database and tissue samples were collected through the Danish CancerBiobank. DNA was analyzed using NGS., Results: 47 (37%) patients were platinum-sensitive, 32 (25%) partially platinum-sensitive, 35 (27%) platinum-resistant, and three (2%) platinum-refractory, while 11 (9%) patients did not receive chemotherapy. Overall, 27 (21%) had known druggable targets. Twelve (26%) platinum-sensitive patients had druggable targets for PARP inhibitors: one for tyrosine kinase inhibitors and one for immunotherapy treatment. Eight (25%) partially platinum-sensitive patients had druggable targets: seven were eligible for PARP inhibitors and one was potentially eligible for alpesilib and hormone therapy. Seven (20%) platinum-resistant patients had druggable targets: six (86%) were potentially eligible for PARP inhibitors, one for immunotherapy, and one for erdafitinib., Conclusions: PARP inhibitors are the most frequent potential targeted therapy in HGSC. However, other targeted therapies remain relevant for investigation according to our mutational findings.

Background: Metastatic colorectal cancer (mCRC) is a complex and heterogeneous disease with few standard and targeted treatment options. Next-generation sequencing of tumor tissue was performed to identify cancer driver mutations to discover possible personalized treatment options, as targeted treatment possibilities are limited for this patient population. Results of genomic sequencing in patients with treatment-refractory mCRC are described in this retrospective analysis., Material and Methods: Clinico-pathological characteristics and genomic sequence results of consecutive patients with refractory mCRC, referred to the Experimental Cancer Therapy Unit (ECTU) at Department of Oncology, Herlev & Gentofte Hospital in the period from 1 October 2015 to 14 December 2018 were reviewed in this retrospective analysis. Tumor tissue from the patients was analyzed by next-generation sequencing using the Oncomine Comprehensive primer panel to detect actionable variants of cancer driver mutations and microsatellite instability status. From August 2018 tumor mutational burden was also analyzed., Results: A total of 80 patients with treatment-refractory mCRC and in a fairly good performance were referred to the ECTU during this period. Genomic sequencing of tumor tissue was performed for all 80 patients and a cancer driver mutation was identified in 90% ( n  = 72) of the patients. A total of 31.3% ( n  = 25) of the patients received therapy either as targetable therapy outside an available trial ( n  = 2), FDA approved therapy ( n  = 2), or treatment in phase 1 or 2 trials, independent of the genomic signature 26.3% ( n  = 21)., Conclusion: Most mCRC patients refractory to standard anti-neoplastic therapies, presented with a cancer driver mutation, however, only a few of these mutations gave rise to matched therapies as only 2.5% of the patients from this period received targeted therapy.

Gastrointestinal stromal tumour (GIST) is the most common sarcoma and can be seen in any part of the gastrointestinal tract. The effect of tyrosine kinase inhibitors varies with mutation status in receptor tyrosine kinase KIT and in platelet-derived growth factor receptor A ( PDGFRA ). This case presents a 61-year-old man, diagnosed with an 11-cm GIST located at the stomach with a high risk of recurrence. The patient showed intolerance to imatinib shortly after introduction and subsequently progressed on sunitinib and nilotinib. The patient started fourth-line treatment with sorafenib with an impressive response to a point at which metastases intra-abdominally and in the liver could be resected. After surgery, sorafenib was restarted. Due to toxicity, sorafenib dose was reduced over time. The dose was insufficient to control the disease since a new recurrence was detected. Mutation analyses revealed a GIST harbouring a deletion of codon p.I843&#95;D846del, located at PDGFRA exon 18, right next to the codon D842 where mutations are known leading to imatinib resistance. In this case, the GIST was highly sensitive to sorafenib, and the response was dose related. It is mandatory to perform mutation analyses on primary tumour and at recurrence in the decision-making of the correct treatment for the patient. In March 2021, the patient had been in treatment with sorafenib for 12.5 years and was still without signs of recurrence. A multidisciplinary approach was essential for the long-term survival of the patient in this case., Competing Interests: The authors have no conflicts of interest to declare., (Copyright © 2021 by S. Karger AG, Basel.)

Ovarian clear cell carcinoma (OCCC) is characterized by dismal prognosis, partially due to its low sensitivity to standard chemotherapy regimen. It is also well-known for presenting unique molecular features in comparison to other epithelial ovarian cancer subtypes. Here, we aim to identify potential subgroups of patients in order to (1) determine their molecular features and (2) characterize their mutational signature. Furthermore, we sought to perform the investigation based on a potentially clinically relevant setting. To that end, we assessed the mutational profile and genomic instability of 55 patients extracted from the Gynecologic Cancer Database (DGCD) by using a panel comprised of 409 cancer-associated genes and a microsatellite assay, respectively; both are currently used in our routine environment. In accordance with previous findings, ARID1A and PIK3CA were the most prevalent mutations, present in 49.1% and 41.8%, respectively. From those, the co-occurrence of ARID1A and PIK3CA mutations was observed in 36.1% of subjects, indicating that this association might be a common feature of OCCC. The microsatellite instability frequency was low across samples. An unbiased assessment of signatures identified the presence of three subgroups, where "PIK3CA" and "Double hit" (with ARID1A and PIK3CA double mutation) subgroups exhibited unique signatures, whilst "ARID1A" and "Undetermined" (no mutations on ARID1A nor PIK3CA ) subgroups showed similar profiles. Those differences were further indicated by COSMIC signatures. Taken together, the current findings suggest that OCCC presents distinct mutational landscapes within its group, which may indicate different therapeutic approaches according to its subgroup. Although encouraging, it is noteworthy that the current results are limited by sample size, and further investigation on a larger group would be crucial to better elucidate them.

The usage of next generation sequencing in combination with targeted gene panels has enforced a better understanding of tumor compositions. The identification of key genomic biomarkers underlying a disease are crucial for diagnosis, prognosis, treatment and therapeutic responses. The Oncomine™ Comprehensive Assay v3 (OCAv3) covers 161 cancer-associated genes and is routinely employed to support clinical decision making for a therapeutic course. An improved version, Oncomine™ Comprehensive Assay Plus (OCA-Plus), has been recently developed, covering 501 genes (144 overlapping with OCAv3) in addition to microsatellite instability (MSI) and tumor mutational burden (TMB) assays in one workflow. The validation of MSI and TMB was not addressed in the present study. However, the implementation of new assays must be validated and confirmed across multiple samples before it can be introduced into a clinical setting. Here, we report the comparison of DNA sequencing results from 50 ovarian cancer formalin-fixed, paraffin-embedded samples subjected to OCAv3 and OCA-Plus. A validation assessment of gene mutations identified using OCA-Plus was performed on the 144 overlapping genes and 313,769 intersecting nucleotide positions of the OCAv3 and the OCA-Plus. Our results showed a 91% concordance within variants classified as likely-pathogenic or pathogenic. Moreover, results showed that a region of PTEN is poorly covered by the OCA-Plus assay, hence, we implemented rescue filters for those variants. In conclusion, the OCA-Plus can reflect the mutational profile of genomic variants compared with OCAv3 of 144 overlapping genes, without compromising performance.

The MAPK signalling genes KRAS, NRAS and BRAF and the PIK3CA gene are routinely investigated for mutations in the diagnostic routine of colorectal cancer. Few studies have reported co-existing mutations in these genes with clinical relevance, while some have been previously regarded as mutually exclusive. We set to investigate the frequency and co-occurrent mutations in these targets, and the occurrence of mismatch repair deficiency (dMMR) in a large cohort of Danish colorectal cancers. 1000 colorectal tumours were sequenced as part of our diagnostic workflow for KRAS, NRAS, BRAF and PIK3CA mutations using next-generation sequencing (NGS) and analysed by immunohistochemistry (IHC) for loss of the MMR proteins, MLH1, PMS2, MSH2 and MSH6. Co-existing mutations in 12 patients (1.2%) occurred as multiple mutations in the same gene or spread across several genes (KRAS, NRAS and/or BRAF). The frequency of single mutations in the genes occurred with a frequency similar to previously reported, except for a higher frequency of BRAF mutations (18.0%). We found dMMR in 14.6% of the cases with a majority lacking expression of both MLH1 and PMS2. BRAF mutations were only present in dMMR cases involving MLH1 and/or PMS2. Our findings suggest that co-existing mutations occur, except for the hotspot BRAF V600E, which is mutually exclusive with KRAS/NRAS mutations. Therefore, instead of single gene alterations from the MAPK signalling, assessing co-occurrence of mutations within one or more of those genes should also be accounted. This may impact future oncological treatments and should be considered in the diagnostic workflow., (© 2020 APMIS. Published by John Wiley & Sons Ltd.)

In this study we investigated the use of cancer cell protein expression of ABCG2 to predict efficacy of systemic first-line irinotecan containing therapy in patients with metastatic colorectal cancer (mCRC). From a Danish national cohort, we identified 119 mCRC patients treated with irinotecan containing therapy in first-line setting. Among these, 108 were eligible for analyses. Immunohistochemistry (IHC) analyses were performed on the primary tumor tissue in order to classify samples as high or low presence of ABCG2 protein. Data were then associated with patient outcome (objective response (OR), progression free survival (PFS) and overall survival (OS)). ABCG2 protein expression in the basolateral membrane was high (score 3+) in 33% of the patients. Exploratory analyses revealed a significant interaction between ABCG2 score, adjuvant treatment and OR ( p = 0.041) in the 101 patients with evaluable disease. Patients with low ABCG2 (score 0-2) and no prior adjuvant therapy had a significantly higher odds ratio of 5.6 (Confidence Interval (CI) 1.68-18.7; p = 0.005) for obtaining OR. In contrast, no significant associations between ABCG2 expression and PFS or OS were found. These results suggest that measurement of the ABCG2 drug efflux pump might be used to select patients with mCRC for irinotecan treatment. However, additional studies are warranted before conclusions regarding a clinical use can be made. Moreover, patients with high ABCG2 immunoreactivity could be candidates for specific ABCG2 inhibition treatment in combination with irinotecan.

Background and Objectives: EUS-FNA is inconclusive in up to 10%-15% of patients with solid pancreatic lesions (SPLs). We aimed to investigate whether supplementary genetic analyses with whole-exome sequencing add diagnostic value in patients with SPLs suspicious of malignancy but inconclusive EUS-FNA., Patients and Methods: Thirty-nine patients, who underwent EUS-FNA of an SPL were retrospectively included. Three groups were defined: 16 (41.0%) had suspected malignancy on EUS confirmed by cytology (malignant), 13 (33.3%) had suspected malignancy on EUS but benign cytology (inconclusive), and 10 (25.6%) had benign EUS imaging and cytology (benign). Areas with the highest epithelial cell concentrations were macro-dissected from the FNA smears from each patient, and extracted DNA was used for whole-exome sequencing by next-generation sequencing of a selected gene panel including 19 genes commonly mutated in cancer., Results: Pathogenic mutations in K-RAS, TP53, and PIK3CA differed significantly between the three groups (P < 0.001, P = 0.018, and P = 0.026, respectively). Pathogenic mutations in KRAS and TP53 were predominant in the inconclusive (54% and 31%, respectively) and malignant groups (81.3% and 50%, respectively) compared to the benign group (0%). Malignant and inconclusive diagnoses correlated strongly with poor overall survival (P < 0.001)., Conclusion: Whole-exome sequencing of genes commonly mutated in pancreatic cancer may be an important adjunct in patients with SPLs suspicious for malignancy on EUS but with uncertain cytological diagnosis., Competing Interests: None

Objective: In patients with large B-cell lymphoma (LBCL) according to WHO, the prognostic significance of MYC translocation is still not sufficiently clarified. We therefore aimed to investigate whether prognostication could be improved in patients with MYC translocation positive LBCL by additional stratification according to MYC and BCL2 protein expression levels or MYC translocation partner gene as well as concurrent BCL2 and/or BCL6 translocation (DH)., Methods: From an unselected consecutive cohort of >600 patients with LBCL investigated with fluorescent in situ hybridization (FISH), 64 patients were diagnosed with MYC translocation positive LBCL and included in the study. They were further investigated for supplemental translocations with FISH and MYC and BCL2 protein expression with immunohistochemistry (IHC)., Results: MYC expression >75% was associated with both reduced progression-free survival (PFS) and overall survival (OS) (PFS: HR 6.8 (95% CI 1.5-31), P = 0.004. OS: HR 4.3 (95% CI 0.9-21), P = 0.05). Immunoglobulin (IG) MYC translocation partner gene was related to high MYC protein expression (P = 0.047) but was not prognostic for PFS (P = 0.8) or OS (P = 0.6). DH did not confer a worse outcome compared to MYC single hit (SH). These findings were confirmed in a comparable, independent validation cohort of 28 patients with MYC translocation positive LBCL. All patients included in the survival analyses were treated with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) or R-CHOEP (R-CHOP + etoposide)., Conclusion: These findings suggest that in patients with LBCL stratification by MYC protein expression level significantly improves the prognostic impact associated with MYC translocation., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Several studies have investigated correlations between metastatic pattern and mutation status in patients with colorectal cancer (CRC). However, most of the studies were small and heterogeneously designed and further research is needed to confirm previous results. In this study, we investigated the association between RAS (KRAS or NRAS), BRAF, PIK3CA mutations and metastatic pattern in patients with metastatic (m) CRC., Material and Methods: This study reviewed Danish biobank and database of patients with mCRC who received cetuximab and irinotecan, independent of RAS mutation status, after fluoropyrimidine, oxaliplatin and irinotecan treatment failure. The database contained information regarding tumor mutation status of KRAS, NRAS, BRAF and PIK3CA genes., Results: Totally, 448 patients were included. On multivariate analyses, RAS mutations were significantly associated with increased odds of having lung metastases at diagnosis of mCRC (odds ratio (OR) = 2.04; 95% confidence interval (CI) = 1.32-3.17), and PIK3CA mutations with decreased odds of peritoneal metastases at diagnosis of mCRC (OR = 0.10; 95%CI = 0.01-0.79). On multivariate analyses of the hazard of developing metastases at any time during follow-up, RAS mutations were significantly associated with increased hazard of lung (hazard ratio (HR) = 1.34; 95%CI = 1.04-1.72) and ovarian metastases (HR = 3.12; 95%CI = 1.05-9.24), BRAF V600E mutation was associated with increased hazard of skin metastases (HR = 6.82; 95%CI = 1.86-25.02) and PIK3CA mutations with decreased hazard of peritoneal metastases (HR = 0.31; 95%CI = 0.11-0.86)., Conclusions: This study indicated that in patients with mCRC, RAS mutations are associated with increased risk of lung and ovary metastases. BRAF V600E is associated with increased risk of skin metastases, and PIK3CA mutation with decreased risk of peritoneal metastases.

Background: Data outlining the mortality and the causes of death in patients with type 1 myocardial infarction, type 2 myocardial infarction, and those with myocardial injury are limited., Methods: During a 1-year period from January 2010 to January 2011, all hospitalized patients who had cardiac troponin I measured on clinical indication were prospectively studied. Patients with at least one cardiac troponin I value >30 ng/L underwent case ascertainment and individual evaluation by an experienced adjudication committee. Patients were classified as having type 1 myocardial infarction, type 2 myocardial infarction, or myocardial injury according to the criteria of the universal definition of myocardial infarction. Follow-up was ensured until December 31, 2014. Data on mortality and causes of death were obtained from the Danish Civil Registration System and the Danish Register of Causes of Death., Results: Overall, 3762 consecutive patients were followed for a mean of 3.2 years (interquartile range 1.3-3.6 years). All-cause mortality differed significantly among categories: Type 1 myocardial infarction 31.7%, type 2 myocardial infarction 62.2%, myocardial injury 58.7%, and 22.2% in patients with nonelevated troponin values (log-rank test; P < .0001). In patients with type 1 myocardial infarction, 61.3% died from cardiovascular causes, vs 42.6% in patients with type 2 myocardial infarction (P = .015) and 41.2% in those with myocardial injury (P < .0001). The overall mortality and the causes of death did not differ substantially between patients with type 2 myocardial infarction and those with myocardial injury., Conclusions: Patients with type 2 myocardial infarction and myocardial injury exhibit a significantly higher long-term mortality compared with patients with type 1 myocardial infarction . However, most patients with type 1 myocardial infarction die from cardiovascular causes in contrast to patients with type 2 myocardial infarction and myocardial injury, in whom noncardiovascular causes of death predominate., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Brain metastases (BM) from colorectal cancer (CRC) are rare, but the incidence is suspected to rise as treatment of metastatic (m) CRC improves. The aim of this study was to identify possible biological and clinical characteristics at initial presentation of mCRC that could predict later risk of developing BM. Furthermore, we wished to estimate the incidence of BM in long-term surviving patients., Material and Methods: We conducted a retrospective study on a Danish multicenter cohort of patients with mCRC who received cetuximab and irinotecan (CetIri) as third-line treatment. All patients had previously progression on 5-FU, irinotecan and oxaliplatin containing regimens and received CetIri treatment independent of RAS mutations status. We subsequently performed KRAS, NRAS, BRAF, PIK3CA, PTEN, ERBB2 and EGFR sequencing of DNA extracted from primary tumor tissue., Results: Totally, 480 patients were included in our study. BM were diagnosed in 42 [8.8%; 95% confidence interval (CI) 6.4-11.6%] patients. Patients with BM had a significantly longer survival from mCRC diagnosis than non-BM patients (median = 32 versus 28 months, p = 0.001). On univariate cox regression analysis, the risk of developing BM was significantly increased in patients with rectal cancer (HR = 3.9; 95% CI = 1.2-13.3), metachronous metastatic disease (HR = 2.3; 95% CI = 1.2-4.4) and lung metastases (HR = 4.2; 95% CI = 2.2-7.9). On multivariate cox regression analysis only lung metastases were significantly associated BM (HR = 3.5; 95% CI = 1.8-6.8). None of the investigated mutations were associated with BM., Conclusion: The incidence of BM was 8.8% in patients with mCRC who received third-line therapy. The most important risk factor for developing BM was lung metastases. Furthermore, rectal cancer, metachronous metastatic disease and long survival were linked to BM development.

Background: No biomarker exists to guide the optimal choice of chemotherapy for patients with metastatic colorectal cancer. We examined the copy numbers (CN) of topoisomerase I (TOP1) as well as the ratios of TOP1/CEN-20 and TOP1/CEN-2 as biomarkers for irinotecan efficacy in patients with metastatic colorectal cancer., Methods: From a national cohort, we identified 163 patients treated every third week with irinotecan 350 mg/m 2 as second-line therapy. Among these 108 were eligible for analyses and thus entered the study. Primary tumors samples were collected and tissue microarray (TMA) blocks were produced. FISH analysis was performed using two probe-mixes: TOP1/CEN-20 and TOP1/CEN-2. Only samples harboring all three signals (TOP1, CEN-20 and CEN-2) using FISH were included in the analyses., Results: In the TOP1/CEN-20 probe-mix the median TOP1- and CEN-20 CN were 4.46 (range: 1.5-9.5) and 2.00 (range: 0.55-4.55), respectively. The median TOP1- and CEN-2 CN in the TOP1/CEN-2 probe-mix, were 4.57 (range: 1.82-10.43) and 1.98 (range: 1.22-6.14), respectively. The median TOP1/CEN-20 ratio and TOP1/CEN-2 ratio were 1.25 (range: 0.92-2.90) and 2.05 (range: 1.00-6.00), respectively. None of the markers TOP1 CN, TOP1/CEN-20-ratio or TOP1/CEN-2-ratio were associated with progression free survival, overall survival or baseline characteristics. Yet, we observed a borderline association for a stepwise increase of the TOP1 CN in relation to objective response as hazard ratio were 1.35 (95% CI 0.96-1.90; p = 0.081)., Conclusions: We verified a borderline significant association between increasing TOP1 CN and objective response as previously reported. Applying the probes representing CEN-20 and CEN-2, in order to investigate the ratios of TOP1/CEN-20 and TOP1/CEN-2 provided no further information in search of a biomarker driven patient stratification. Other biomarkers to be paired with TOP1 CN are therefore highly warranted.

The role of c-myc in multiple myeloma (MM) is controversial. We conducted a retrospective study of 117 patients with MM diagnosed between 2004 and 2010 at Herlev Hospital. Immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) were performed on tissue microarrays (TMAs) made from diagnostic bone marrow aspirates. Clinical data were obtained from the Danish Multiple Myeloma Database (DMMD). Overexpression of c-myc was found in 40% of patients. MYC translocation was found in 10% of patients. Overexpression of c-myc was not associated with MYC translocation. Overexpression of c-myc was associated with hypercalcemia (p = 0.02) and extramedullary myeloma (p < 0.01). Overexpression of c-myc was associated with shorter overall survival (OS) by multivariable analysis of the entire patient cohort [HR 1.92 (1.06-3.45), p = 0.03] and univariable analysis of high-dose-therapy (HDT)-ineligible patients [HR 2.01 (1.05-3.86), p = 0.04]. Further studies of c-myc overexpression in larger cohorts of patients with MM are warranted.

Background: Elevated cardiac troponins in clinical conditions other than myocardial infarction are well known. For such occurrences, the term "myocardial injury" has been proposed. The long-term outcome in patients with myocardial injury related to various cardiac and noncardiac clinical disorders is unknown., Methods: During January 2010 to January 2011, we prospectively studied hospitalized patients who had cardiac troponin I measured on clinical indication. Patients with cardiac troponin I values >30 ng/L and no evidence of myocardial ischemia were diagnosed as having myocardial injury. Patients were classified into 5 categories of plausible related conditions: cardiac ischemic, cardiac nonischemic, noncardiac, multifactorial, or indeterminate. Follow-up was a minimum of 3 years, with all-cause mortality as the single end-point., Results: A total of 3762 patients were considered, of whom 1089 (29%) had myocardial injury. The most common associated conditions were noncardiac (n = 346) or multifactorial (n = 359). Cardiac ischemic (n = 183) and cardiac nonischemic (n = 134) conditions occurred less frequently. After a median of 3.2 years, 645 patients (59%) had died. A multivariate Cox regression analysis showed no difference in mortality between patients with cardiac ischemic and cardiac nonischemic conditions (hazard ratio [HR] 0.75; 95% confidence interval [CI], 0.50-1.13; P = .2). Patients with noncardiac or multifactorial disorders, however, had significantly higher mortality than those with associated cardiac ischemic conditions (HR 1.39; 95% CI, 1.06-1.80; P = .02, and HR 1.94; 95% CI, 1.50-2.51; P <.001), respectively., Conclusions: In patients with myocardial injury, the most common associated conditions were noncardiac or multifactorial. Of notice, these patients had significantly higher long-term mortality when compared with those with associated cardiac conditions., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Cardiac troponins have emerged as the preferred biomarkers for detecting myocardial necrosis and diagnosing myocardial infarction. However, current cardiac troponin assays do not discriminate between ischemic and nonischemic causes of myocardial cell death. Thus, when an increased troponin value is encountered in the absence of obvious myocardial ischemia, a careful search for other clinical conditions is crucial., Methods: In 2010 to 2011, we prospectively studied hospitalized patients who had cardiac troponin I measured on clinical indication. An acute myocardial infarction was diagnosed in cases of a cardiac troponin I increase or decrease pattern with at least 1 value >30 ng/L (99th percentile) together with myocardial ischemia. Myocardial injury was defined as cardiac troponin I values >30 ng/L, but without signs or symptoms indicating overt cardiac ischemia. Patients with peak values ≤30 ng/L were classified as nonelevated cardiac troponin I. Follow-up was at least 3 years with all-cause mortality as the sole clinical end point., Results: A total of 3762 patients were included. Of these, 488 (13%) had acute myocardial infarction, 1089 (29%) had myocardial injury, and 2185 (58%) had nonelevated cardiac troponin I values. Patients with myocardial injury frequently presented with dyspnea, were older, and had more comorbidity than patients in the 2 other groups. During a median follow-up of 3.2 years, 1342 patients died. Mortality differed significantly between groups: 39% in those with myocardial infarction, 59% in those with myocardial injury, and 23% in those with nonelevated cardiac troponin I (log-rank test; P < .0001). No significant difference in mortality between patients with type 2 myocardial infarction and patients with myocardial injury was observed (63% and 59%, respectively)., Conclusions: Patients with myocardial injury are older and have more comorbidity than those with acute myocardial infarction. Both groups exhibit a poorer prognosis than patients with nonelevated cardiac troponin I values. Of note, a very high long-term mortality is observed in patients with type 2 myocardial infarction and patients with myocardial injury., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Topoisomerase-1 (Top1) targeting drugs have shown promising efficacy in patients with metastatic breast cancer (BC). However, these drugs are rather toxic calling for development and validation of predictive biomarkers to increase the therapeutic index. As these drugs are targeting the Top1 protein and since no validated anti-Top1 antibodies for immunohistochemistry have been reported, we raised the hypothesis that TOP1 gene amplifications may serve as a proxy for the Top1 protein and thereby a biomarker of response to treatment with Top1 inhibitors in BC. The aim was to determine the prevalence of TOP1 gene copy gain in BC. The prevalence of TOP1 gene copy gain was investigated by fluorescence in situ hybridization with a TOP1/CEN-20 probemix in normal breast tissue (N = 100) and in tissue from patients with metastatic BC in a discovery (N = 100) and a validation cohort (N = 205). As amplification of 20q including CEN-20 is common in BC a TOP1/CEN-2 probemix was applied to the validation cohort. More than 30% of the patients had gene copy numbers of ≥ 4 and ∼20% of the patients had TOP1/CEN-20 ratios ≥ 1.5. The CEN-2 probe did not add any information. Gain of the TOP1 gene appears to be common in BC making the gene a potential biomarker for response to treatment with Top1 inhibitors. As 20q amplification is a common finding in BC and as no other suitable reference gene has yet been identified, TOP1 copy number may be a more valid method of detecting gain than using a gene/centromere ratio., (© 2015 UICC.)

Background: Since the arrival of the universal definition of myocardial infarction more sensitive troponin assays have been developed. How these occurrences have influenced the proportions and clinical features of the components of acute coronary syndrome have not been studied prospectively in unselected hospital patients., Methods: During 2010 we evaluated all patients in whom cardiac troponin I had been measured at a single university hospital. The diagnosis of acute myocardial infarction (ST-elevation myocardial infarction [STEMI] or non-ST-elevation myocardial infarction [NSTEMI]) was established in cases of a rise and/or fall of cardiac troponin I together with cardiac ischemic features. Patients with unstable chest discomfort and cardiac troponin I values below the decision limit of myocardial infarction were diagnosed as having unstable angina pectoris. The definition of acute coronary syndrome included unstable angina pectoris, NSTEMI, and STEMI. Mortality data were obtained from the Danish Civil Personal Registration System., Results: Of 3762 consecutive patients, 516 had acute coronary syndrome. Unstable angina pectoris was present in 7%, NSTEMI in 67%, and STEMI in 26%. The NSTEMI patients were older, more frequently women, and had more comorbidities than patients with unstable angina pectoris and STEMI. At median follow-up of 3.2 years 195 patients had died: 14% of unstable angina pectoris, 45% of NSTEMI, and 25% of STEMI patients. Age-adjusted log-rank statistics revealed differences in mortality: NSTEMI vs unstable angina pectoris (P = .0091) and NSTEMI vs STEMI (P = .0045)., Conclusions: The application of the universal definition together with the use of a contemporary troponin assay seems to have reduced the proportion of patients with unstable angina pectoris to the benefit of patients with NSTEMI. Despite this, NSTEMI patients have a sustained higher mortality than patients with STEMI., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Topoisomerase (TOP) gene copy number changes may predict response to treatment with TOP-targeting drugs in cancer treatment. This was first described in patients with breast cancer and is currently being investigated in other malignant diseases. TOP-targeting drugs may induce TOP gene copy number changes at relapse, with possible implications for relapse therapy efficacy. TOP gene alterations in lymphoma are poorly investigated. In this study, TOP1 and TOP2A gene alterations were investigated in patients with de novo diffuse large B-cell lymphoma (DLBCL) (n = 33) and relapsed DLBCL treated with chemotherapy regimens including TOP2-targeting drugs (n = 16). No TOP1 or TOP2A copy number changes were found. Polysomy of chromosomes 20 and 17 was seen in 3 of 25 patients (12%) and 2 of 32 patients (6%) with de novo DLBCL. Among relapsed patients, chromosome polysomy was more frequently observed in 5 of 13 patients (38%) and 4 of 16 patients (25%) harboring chromosome 20 and 17 polysomy, respectively; however, these differences only tended to be significant (p = 0.09 and p = 0.09, respectively). The results suggest that TOP gene copy number changes are very infrequent in DLBCL and not likely induced by TOP2-targeting drugs. Increased polyploidy of chromosomes 17 and 20 among patients with relapsed DLBCL may reflect genetic compensation in the tumor cells after TOP2 inhibition, but is more likely due to the increased genetic instability often seen in progressed cancers. Therefore, it is unlikely that TOP1 and TOP2A gene alterations can be used as predictive markers for response to treatment with TOP2-targeting drugs in patients with DLBCL., (Copyright © 2015 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.)

Introduction: Topoisomerase 1 (TOP1) and 2A (TOP2A) are potential predictive biomarkers for irinotecan and anthracycline treatment, respectively, in colorectal cancer (CRC), and we have recently reported a high frequency of gene gain of the TOP1 and TOP2A genes in CRC. Furthermore, Mismatch Repair (MMR) subtypes of CRC have been associated with benefit from adjuvant chemotherapy of primary CRC. Given the involvement of the topoisomerase enzymes in DNA replication and repair, we raised the hypothesis that an association may exist between TOP gene copy numbers and MMR proficiency/deficiency in CRC., Material and Methods: Test cohort: FISH analysis with an in-house TOP1/CEN20 probe mix and a commercially available TOP2A/CEN17 (Dako, Glostrup, Denmark) probe mix was performed on archival formalin fixed paraffin embedded (FFPE) tissue samples from 18 patients with proficient MMR (pMMR) CRC and 18 patients with deficient MMR (dMMR) CRC. TOP1 and TOP2A gene copy numbers and their ratios per nucleus were correlated with MMR status using the Mann-Whitney test. Validation cohort: FFPE samples from 154 patients with primary stage III CRC (originally included in the RANX05 study) were classified according to MMR status by immunohistochemical analysis using validated antibodies for MLH1, MLH2, MSH6 and PMS2, and information on TOP1, CEN20, TOP2A and CEN17 status was previously published for this cohort., Results: The observed TOP1 gene copy numbers in the 36 CRC test cohort were significantly greater (p < 0.01) in the pMMR subgroup (mean: 3.84, SD: 2.03) than in the dMMR subgroup (mean: 1.50, SD: 0.12). Similarly, the TOP2A copy numbers were significantly greater (p < 0.01) in the pMMR subgroup (mean: 1.99, SD: 0.52) than in the dMMR subgroup (mean: 1.52, SD: 0.10). These findings were confirmed in the validation cohort, where in the pMMR subgroup 51% had ≥2 extra TOP1 copies per cell, while all tumors classified as dMMR had diploid TOP1 status and mean TOP2A copy numbers were 2.30 (SD: 1.36) and 1.80 (SD: 0.31) (p = 0.01) in the pMMR subgroup vs. dMMR subgroup, respectively., Discussion and Conclusion: Our results show that TOP1 and TOP2A gene copy numbers are increased in the pMMR subgroup. We propose that this preference may reflect a selective pressure to gain and/or maintain the gained extra copies of topoisomerase genes whose products are required to cope with high replication stress present in the pMMR tumors, thereby providing a survival advantage selectively in pMMR tumors. Future studies should test this concept and explore potential differences between pMMR and dMMR tumors in response to Top1 and Top2 inhibitors., (Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

The aim of this study was to prospectively investigate the clinical characteristics including symptoms and long-term mortality in patients with acute myocardial infarction (AMI) accidentally admitted to non-cardiology departments (NCDs). For comparison, similar observations in patients admitted to the coronary care unit (CCU) were collected. During a 1-year period, consecutive patients having cardiac troponin I measured at the Odense University Hospital were considered. The hospital has 27 clinical departments. Patients were classified as having an AMI if the diagnostic criteria of the universal definition were met. Follow-up was at least 1 year with mortality as the clinical end point. Of 3,762 consecutive patients, an AMI was diagnosed in 479, of whom 114 patients (24%) were hospitalized in NCDs and 365 (76%) in the CCU. Chest pain or chest discomfort more frequently occurred in patients from the CCU (83%) than in patients from the NCDs (45%, p <0.0001). At median follow-up of 2.1 years, 150 patients had died: 73 (64%) of patients from the NCDs and 77 (21%) of the patients from the CCU. In the multivariable Cox regression analysis, the adjusted hazard ratio of mortality for patients from the NCDs versus CCU was 2.0 (95% confidence interval 1.3 to 3.2). In conclusion, chest pain/discomfort was absent in more than half of the patients with AMI admitted to NCDs, and admission to NCDs was an independent predictor of a 2 times higher long-term mortality in comparison with admission to the CCU., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Purpose: The biomarker Osteoprotegerin (OPG) is associated with coronary artery disease (CAD). The main purpose of this study was to evaluate the diagnostic value of OPG in healthy subjects and in patients with suspected angina pectoris (AP)., Methods: A total of 1805 persons were enrolled: 1152 healthy subjects and 493 patients with suspected AP. For comparison 160 patients with acute myocardial infarction (MI) were included. To uncover subclinical coronary atherosclerosis, a non-contrast cardiac-CT scan was performed in healthy subjects; while in patients with suspected AP a contrast coronary angiography was used to detect significant stenosis. OPG concentrations were analyzed and compared between groups. ROC-analyses were performed to estimate OPG cut-off values., Results: OPG concentrations increased according to disease severity with the highest levels found in patients with acute MI. No significant difference (p = 0.97) in OPG concentrations was observed between subgroups of healthy subjects according to severity of coronary calcifications. A significant difference (p < 0.0001) in OPG concentrations was found between subgroups of patients with suspected stable AP according to severity of CAD. ROC-analysis showed an AUC of 0.62 (95% CI: 0.57-0.67). The optimal cut-off value of OPG (<2.29 ng/mL) had a sensitivity of 56.2% (95% CI: 49.2-63.0%) and a specificity of 62.9% (95% CI: 57.3-68.2%)., Conclusion: OPG cannot be used to differentiate between healthy subjects with low versus high levels of coronary calcifications. In patients with suspected AP a single OPG measurement is of limited use in the diagnosis of CAD., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Trastuzumab is established as standard care for patients with HER2-positive breast cancer both in the adjuvant and metastatic setting. However, 50% of the patients do not respond to the trastuzumab therapy, and therefore new predictive biomarkers are highly warranted. MicroRNAs (miRs) constitute a new group of biomarkers and their cellular expression can be determined in tumor samples by in situ hybridization (ISH) analysis. miR-21 is highly prevalent and up-regulated in breast cancer and has been linked to drug resistance in clinical and in vitro settings. To determine expression patterns of miR-21 in high-grade breast cancers, we examined miR-21 expression in 22 HER2-positive tumors and 15 HER2-negative high-grade tumors by ISH. The histological examination indicated that patient samples could be divided into three major expression patterns: miR-21 predominantly in tumor stroma, predominantly in cancer cells, or in both stromal and cancer cells. There was no obvious difference between the HER2-positive and HER2-negative tumors in terms of the miR-21 expression patterns and intensities. To explore the possibility that miR-21 expression levels and/or cellular localization could predict resistance to adjuvant trastuzumab in HER2-positive breast cancer patients, we analyzed additional 16 HER2-positive tumors from patients who were treated with trastuzumab in the adjuvant setting. Eight of the 16 patients showed clinical recurrence and were considered resistant. Examination of the miR-21 expression patterns and intensities revealed no association between the miR-21 scores in the cancer cell population (p = 0.69) or the stromal cells population (p = 0.13) and recurrent disease after adjuvant trastuzumab. Thus, our findings show that elevated miR-21 expression does not predict resistance to adjuvant trastuzumab.

Objectives: The clinical classification of myocardial infarction (MI) into five types was introduced in 2007 as a component of the universal definition. A Type 5 MI was defined as a MI related to coronary artery bypass surgery. In a setting of patients undergoing elective coronary artery bypass grafting, we set out (i) to describe the pattern of multiple serial cardiac troponin I (cTnI) measurements within 72 h postoperatively and (ii) to determine the optimal cardiac troponin I cut-off value in ruling in or ruling out a Type 5 MI., Methods: In 2011-2012, patients with two- and three-vessel disease scheduled for elective on-pump coronary artery bypass grafting were considered. Samples for cTnI were drawn before and 0, 2, 4, 6, 12, 24, 48 and 72 h after surgery. Analysis for cardiac troponin I was performed by use of the Abbott Architect c16000 system with an upper reference limit (URL) of 30 ng/l. The diagnosis of a Type 5 MI was prospectively made by a consultant cardiologist and was based on clinical, electrocardiographic and imaging data together with routine sampling and measurements of cTnI, but without knowledge of the results of serial study cTnI measurements., Results: Of the 141 eligible patients, 99 (70%) qualified for final enrollment. In 8 patients (8%), the clinical diagnosis of a Type 5 MI was made. Patients without Type 5 MI (n = 91) had a median cTnI peak value of 7675 ng/l compared with 20 500 ng/l in Type 5 MI patients (P = 0.01). By use of receiver operating characteristic curves, optimal cut-off values for identifying Type 5 MI were defined as 7970 ng/l (corresponding to 266 times the URL) 12 h postoperatively and 9950 ng/l (corresponding to 331 times the URL) 24 h postoperatively. These cut-off values resulted in negative predictive values of 0.99 (12 h) and 0.99 (24 h). Positive predictive values were 0.23 (12 h) and 0.35 (24 h)., Conclusions: In clinically stable patients undergoing elective coronary artery bypass grafting, measurements of cTnI are useful in ruling out a Type 5 MI.

Background: The classification of myocardial infarction into 5 types was introduced in 2007. The prognostic impact of this universal definition, with particular focus on type 2 myocardial infarction, has not been studied prospectively in unselected hospital patients., Methods: During a 1-year period, all hospitalized patients having cardiac troponin I measured were considered. The diagnosis of a myocardial infarction was according to the universal definition, and specified criteria were used in the classification of type 2 myocardial infarction. Follow-up was at least 1 year, with mortality as the end point., Results: A total of 3762 consecutive patients were studied, of whom 488 (13%) had a myocardial infarction. In 119 patients a type 2 myocardial infarction was diagnosed. After a median of 2.1 years (interquartile range, 1.6-2.5 years), 150 patients had died, with a mortality rate of 49% (58/119) in those with type 2 myocardial infarction and 26% (92/360) in those with type 1 myocardial infarction (P < .0001). In a multivariable Cox regression analysis the following variables were independently associated with mortality: current or prior smoker, high age, prior myocardial infarction, type 2 myocardial infarction, hypercholesterolemia, high p-creatinine, and diabetes mellitus. The multivariable-adjusted hazard ratio for type 2 myocardial infarction was 2.0 (95% confidence interval, 1.3-3.0). With shock as the only exception, mortality was independent of the triggering conditions leading to type 2 myocardial infarction., Conclusions: Mortality in patients with type 2 myocardial infarction is high, reaching approximately 50% after 2 years. Further descriptive and survival studies are needed to improve the scientific evidence on which treatment of type 2 myocardial infarction is based., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objective: Over the last decades Lipocalin-type prostaglandin D synthase (L-PGDS), Osteoprotegerin (OPG), Osteopontin (OPN) and Pregnancy associated plasma protein A (PAPP-A) have been reported to be associated with coronary artery disease, and L-PGDS has been proposed as a potential new diagnostic tool in the setting of stable coronary artery disease. We set out to investigate if measurement of concentrations of these biomarkers could be used to differentiate between four groups of individuals with different atherosclerotic manifestations., Methods: A total of 120 individuals from four equal gender- and age-matched groups were studied: (i) no previous cardiovascular disease (CVD) and no coronary calcifications [CAC-negative group], (ii) no previous CVD but evidence of severe coronary calcifications [CAC-positive group], (iii) acute coronary syndrome [ACS-group], and (iv) clinical stable patients with CVD, who were referred for cardiovascular surgery [CVD-group]. Concentrations of L-PGDS, OPG, OPN and PAPP-A were analyzed and compared between the four groups., Results: We did not find any significant differences in L-PGDS concentrations between the four groups (p = 0.32). OPG concentrations differed significantly (p = 0.003), with the highest concentration observed in ACS patients. Considering OPN (p = 0.12) and PAPP-A (p = 0.53) their concentrations between groups did not differ significantly., Conclusion: The main message from this study is the observation that L-PGDS based on a single blood test appears to be less valuable than previously proposed in identification of patients with coronary artery disease. However, ACS patients have higher OPG concentrations than patients with different manifestations of stable atherosclerosis. Neither OPN nor PAPP-A concentrations differed between groups.

In large B-cell lymphoma (LBCL) MYC- and MYC/BCL2 double-hit (DH) translocations have been associated with inferior survival. We hypothesised that the negative prognostic impact of MYC translocation was determined by an immunoglobulin MYC translocation partner gene (IG-MYC), as opposed to a non-immunoglobulin partner gene (nonIG-MYC). In a prospective, unselected cohort of 237 LBCL patients MYC and BCL2 translocations were identified by fluorescent in situ hybridisation (FISH) with split probes. MYC translocation partner gene was identified by IGH/MYC fusion probes and/or kappa/lambda split probes. Clinical data were collected from patient files. MYC translocation was identified in 28/225 patients. IG-MYC translocation partner gene was identified in 12/24 patients. DH translocation was identified in 23/228 patients. IG-MYC translocation partner gene was identified in 9/19 DH patients. Neither MYC-nor DH translocation showed correlation with survival. However, MYC translocation with IG-MYC translocation partner gene was associated with worse OS compared with both MYC translocation with nonIG-MYC translocation partner gene (P = 0.02) as well as absence of MYC translocation (P = 0.03). In patients with DH a similar, however, stronger correlation was seen (P = 0.003 and P = 0.0004 respectively). MYC - or DH translocation with nonIG-MYC translocation partner gene was not associated with worse overall survival (P = 0.2 and P = 0.3 respectively). Most patients received Rituximab (86%) and CHOP/CHOP-like chemotherapy regimes (81%). We suggest that prognostic stratification of LBCL patients by MYC and/or DH translocations should include identification of MYC translocation partner gene because approximately half of the cases harbour nonIG-MYC translocation partner genes with no or minor influence on survival., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Microfibrillar-associated protein 4 (MFAP4) is located in the extracellular matrix (ECM). We sought to identify tissues with high levels of MFAP4 mRNA and MFAP4 protein expression. Moreover, we aimed to evaluate the significance of MFAP4 as a marker of cardiovascular disease (CVD) and to correlate MFAP4 with other known ECM markers, such as fibulin-1, osteoprotegerin (OPG), and osteopontin (OPN). Quantitative real-time PCR demonstrated that MFAP4 mRNA was more highly expressed in the heart, lung, and intestine than in other elastic tissues. Immunohistochemical studies demonstrated high levels of MFAP4 protein mainly at sites rich in elastic fibers and within blood vessels in all tissues investigated. The AlphaLISA technique was used to determine serum MFAP4 levels in a clinical cohort of 172 patients consisting of 5 matched groups with varying degrees of CVD: 1: patients with ST elevation myocardial infarction (STEMI), 2: patients with non-STEMI, 3: patients destined for vascular surgery because of various atherosclerotic diseases (stable atherosclerotic disease), 4: apparently healthy individuals with documented coronary artery calcification (CAC-positive), and 5: apparently healthy individuals without signs of coronary artery calcification (CAC-negative). Serum MFAP4 levels were significantly lower in patients with stable atherosclerotic disease than CAC-negative individuals (p<0.05). Furthermore, lower serum MFAP4 levels were present in patients with stable atherosclerotic disease compared with STEMI and non-STEMI patients (p<0.05). In patients with stable atherosclerotic disease, positive correlations between MFAP4 and both fibulin-1 (ρ = 0.50; p = 0.0244) and OPG (ρ = 0.62; p = 0.0014) were found. Together, these results indicate that MFAP4 is mainly located in elastic fibers and is highly expressed in blood vessels. The present study suggests that serum MFAP4 varies in groups of patients with different cardiovascular conditions. Further studies are warranted to describe the role of serum MFAP4 as a biomarker of stable atherosclerotic disease.

Background: The classification of myocardial infarction into 5 types was introduced in 2007 as an important component of the universal definition. In contrast to the plaque rupture-related type 1 myocardial infarction, type 2 myocardial infarction is considered to be caused by an imbalance between demand and supply of oxygen in the myocardium. However, no specific criteria for type 2 myocardial infarction have been established., Methods: We prospectively studied unselected hospital patients who had cardiac troponin I measured on clinical indication. The diagnosis and classification of myocardial infarction were established, and the frequency and features of type 2 myocardial infarction were investigated by use of novel developed criteria., Results: From January 2010 to January 2011, a total of 7230 consecutive patients who had cardiac troponin I measured were evaluated, and 4499 patients qualified for inclusion. The diagnosis of myocardial infarction was established in 553 patients, of whom 386 (72%) had a type 1 myocardial infarction and 144 (26%) had a type 2 myocardial infarction. Patients in the group with type 2 myocardial infarction were older and more likely to be female, and had more comorbidities. The proportion of patients without significant coronary artery disease was higher in those with type 2 myocardial infarction (45%) than in those with type 1 myocardial infarction (12%) (P < .001). Tachyarrhythmias, anemia, and respiratory failure were the most prevalent mechanisms causing type 2 myocardial infarction., Conclusions: In a cohort of patients with myocardial infarction who were admitted consecutively through 1 year, the category of type 2 myocardial infarction comprised one fourth when diagnosed by the use of newly developed criteria. Approximately half of patients with type 2 myocardial infarction had no significant coronary artery disease., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background: Von Willebrand factor (VWF) is pivotal in arterial thrombosis, and osteoprotegerin (OPG) is besides being a bone protein also related to cardiovascular diseases. OPG can bind VWF, but the significance of this interaction is not known., Objectives: The aim was to develop an assay for measurement of von Willebrand factor-osteoprotegerin complex (VWF:OPG) in human plasma. Furthermore, the significance of VWF:OPG complex as a marker of cardiovascular disease (CVD) was evaluated., Patients/methods: A sandwich ELISA for quantification of VWF:OPG was developed using a polyclonal rabbit anti-human VWF capturing antibody and a monoclonal anti-human OPG detecting antibody. Samples were quantified relative to a standard curve obtained from dilutions of a plasma pool from healthy individuals. The assay was evaluated in two groups of patients with CVD and two groups of asymptomatic individuals with and without documented coronary calcification (total n=118)., Results and Conclusions: The assay detected VWF:OPG complexes in human plasma, while no significant signal was observed when testing solutions containing VWF or recombinant OPG alone. Importantly, the ELISA assay was able to detect in vitro formed complexes between human VWF and recombinant OPG in a dose-dependent manner. There was a large inter-individual variation in plasma VWF:OPG levels, but we found no significant differences in the level of VWF:OPG complexes between the four groups. Thus, we conclude that increasing OPG plasma levels in atherosclerotic CVD are not derived from increased levels of complexed form of VWF and OPG, but are more likely due to increased amounts of OPG secreted into the circulation., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Forecast-based flow prediction in drainage systems can be used to implement real-time control of drainage systems. This study compares two different types of rainfall forecast - a radar rainfall extrapolation-based nowcast model and a numerical weather prediction model. The models are applied as input to an urban runoff model predicting the inlet flow to a waste water treatment plant. The modelled flows are auto-calibrated against real-time flow observations in order to certify the best possible forecast. Results show that it is possible to forecast flows with a lead time of 24 h. The best performance of the system is found using the radar nowcast for the short lead times and the weather model for larger lead times.