Van Vugt Research Articles

Setting conscious perception alight

Neuroscience What are the neuronal mechanisms that enable conscious perception? Why do some images remain subliminal? Van Vugt et al. trained monkeys to detect low-contrast images and compared neuronal activity in brain areas V1, V4, and the dorsolateral prefrontal cortex. Some stimuli made it into

The cost of sarcopenia.

van Vugt and colleagues found that sarcopenia is an independent risk factor for increased hospital costs for liver transplant candidates. (1) They report that diminished muscle mass measured on CT imaging as the skeletal muscle index (SMI, cm2 muscle area/m2 patient height) is strongly associated with increased hospitalizations and cost of care. The association was significant on multivariable modeling adjusted for potential confounders. This original observation using an anatomic muscle mass measurement is consistent with work showing that physical frailty measured by gait speed indicates similar risk for increased hospitalization and healthcare costs in such patients.(2) This article is protected by copyright. All rights reserved.

EEG-based neural correlates of ACT-R model for multitasking

EEG-based neural correlates of ACT-R model for multitasking

Introduction to Volume 10, Issue 1 of topiCS.

In this issue, we lead with the topic, Coordination and Context in Cognitive Science, from Topic Editor Chris Kello (University of California, Merced). Kello's introduction begins with a quick introduction to the concepts of the dynamics of coordination and phase transitions in forms of human behavior as diverse as walking, neural activity, and dinner conversations. He then transitions to a series of short introductions to each of his six articles. His introduction provides a gentle introduction to this approach, and each of the six papers provides an update on the progress made in advancing the role of context and coordination in cognitive science. Second in this issue is a comment by Ian Apperly (University of Birmingham, England), on last October's topic, Memory and Common Ground Processes in Language Use, which was organized by Sarah Brown-Schmidt. The two next topics are both “Best Ofs”—the best papers presented at their respective conferences. The first batch was curated by Topic Editors Bill Kennedy (George Mason University, U.S.) Marieke van Vugt (University of Groningen, The Netherlands), and Adrian Banks (University of Surrey, England), and includes revised and expanded versions of the best five papers presented at the 2017 meeting of the International Conference on Cognitive Modeling. The last topic is a perennial favorite of mine and includes revised and expanded versions of three prize-winning papers presented at the 2017 Cognitive Science Conference (London): the Best Perception & Action paper, the best Higher-Level Cognition, and the Marr Prize winner for Best Student paper. In case you missed it … a brief mention of topics appearing in recent issues of topiCS A year ago, in January 2017 (Volume 9, Issue 1) we featured Cortical Color by Topic Editor Berit Brogaard (University of Miami) and two Best Ofs … the Best of the ICCM 2016 Conference and those papers from the 2016 Cognitive Science Conference which had won prizes and accepted our offer to expand their conference paper and publish it in topiCS. As of this writing (October 2016) less than a year has passed since 9:1's publication, but its papers are already garnering citations by the metrics used by the Web of Science. Specifically, the papers in the collection of five ICCM papers have gained five citations and those in Topic Editor Berit Brogaard's “Cortical Color” topic have been cited twice. Going back in time to January 2016 (Volume 8, Issue 1), we find that Topic Editor Stephan Lewandowsky's collection of 12 papers on Future Global Change and Cognition has gathered an impressive 66 citations, including an impressive 15 citations for Ranney and Clark's paper on “Climate Change Conceptual Change,” which has been designed on ISI's Web of Science as a “highly cited paper.” These designations are given to papers in the “top 1% of the academic field of Psychiatry/Psychology based on a highly cited threshold for the field and publication year.” The six papers from 2016's collection of “Best of” ICCM papers are attracting attention as well with a collected total of seven citations. All we can say is to keep reading topiCS for the greatest and latest breaking work in Cognitive Science. To would-be Topic Editors, we say, “Keep those proposals coming in!” topiCS has established itself as the “go-to” site for curated collections of papers on timely topics of interest to the broad cognitive science community. topiCS encourages letters and commentaries on all topics, as well as proposals for new topics. Letters are typically 400–1,000 words (maximum of two published pages) and will be published without abstract or references (possibly 1–2 but usually none). Commentaries are often solicited by Topic Editors prior to the publication of their topic. However, commentaries after publication are also considered and should range between 1,000 and 2,000 words. Most commentaries should not have an abstract and should not include many references. The Executive Editor and the Senior Editorial Board (SEB) members are constantly searching for new and exciting topics for topiCS. Feel free to open communications with a short note to the Executive Editor (grayw@rpi.edu) or an SEB member (SEB members are listed under the Editorial Board heading on the publisher's homepage for topiCS (http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1756-8765/homepage/EditorialBoard.html).

Diagnostic utility of appetite loss in addition to existing prediction models for community-acquired pneumonia in the elderly: a prospective diagnostic study in acute care hospitals in Japan

ObjectiveDiagnosis of community-acquired pneumonia (CAP) in the elderly is often delayed because of atypical presentation and non-specific symptoms, such as appetite loss, falls and disturbance in consciousness. The aim of...

Waarom wordt draaiduizeligheid niet vaker behandeld?

Abstract: Van Vugt VA, Diaz Nerio PM, Van der Wouden JC, Van der Horst HE, Maarsingh OR. Why is dizziness not treated more often? Huisarts Wet 2017;60(9):436-9. Background: Dizziness is an unpleasant problem that affects daily functioning, thereby giving rise to considerable economic loss. While there are cheap, safe, and effective treatment techniques, such as the Epley manoeuvre and vestibular rehabilitation, many general practitioners prefer a watchful waiting strategy. The aim of this study was to investigate to what extent Dutch GPs use these treatment techniques and the reasons why they choose not to do so. Method: In 2012, 426 Dutch GPs were asked to complete an online questionnaire about their use of the Epley manoeuvre, Brandt-Daroff exercise, and vestibular rehabilitation for patients with dizziness. Results: While about half of the respondents used the Epley manoeuvre (57.3%) or Brandt-Daroff exercises (50.2%), only 6.8% used vestibular rehabilitation. Most GPs gave these treatments themselves, with only 25.8%, 10.7%, and 34.7% of the GPs referring patients to other healthcare practitioners for these treatments, respectively. The most mentioned reason for not using these treatments was not knowing how to apply them (49.5%, 89.6%, 92.4%, respectively) or doubt about their effectiveness (29.7%, 11.8%, and 6.3%, respectively). Conclusion: Dutch GPs make too little use of repositioning manoeuvres and vestibular rehabilitation as treatment for dizziness. Because GPs appear to be uncertain how to perform these techniques, Apps and E-Health applications may help to improve the necessary knowledge and skills.

From Dominance to Leadership: Emerging Lessons from the Ancestral Past

Management researchers who study leadership and followership commonly draw upon models from a subsample of contemporary organizations or relatively recent historical cases. In contrast, the present symposium brings together four sets of studies that illustrate the benefits and potential for thinking about leadership and followership through broader and deeper evolutionary perspectives. Beyond significantly expanding the parameter space of potential models or benchmarks to include a wide range of formal and informal organizations that span from prehistory (i.e., before writing emerged) through today, the studies that we present will also highlight a set of specific preferences and biases that appear to have evolved across the course of human evolution and persist (if vestigially) in today's workplaces. First, van Vugt applies prior research concerning followers' perceptions of leaders' faces to test the “evolutionary contingency hypothesis&x201D; that people will tend to value specific types of physical features in leaders depending on environmental demands (e.g., war or peace). Second, Kniffin presents evidence that physically large men are perceived to be relatively more persuasive – a pattern that is evolutionarily sensible in light of anthropological knowledge about pre-industrial “big man&x201D; societies where (physically large) male leaders relied on influence rather than force. Third, Case introduces Dual Strategies Theory to highlight the distinction between leaders who are dominance- or prestige- oriented and presents the results of three new experiments designed to explore conditions under which prestige- oriented leadership might yield poor outcomes. Fourth, Brosnan presents new research she has conducted with groups of non-human primates (e.g., chimpanzees) that shows “inequity aversion&x201D; to be prevalent, thereby providing a deep basis for understanding human preferences for fairness – and fair leaders. Finally, Mary Uhl-Bien – a leading scholar on complexity leadership, relational leadership and followership – will offer discussion on the symposium and focus attention on the relational and contextual aspects of the presentations. Considering leadership as a relational phenomenon constructed in context, Uhl-Bien will lead a discussion addressing questions raised across the papers related to gender, environment, social-cognitive processes, and the relational co-construction of leadership and followership.

Abstract 2488: Tumor necrosis factor-alpha signaling determines cytotoxicity induced by BRCA2 deficiency

Abstract Introduction Breast cancer 2, early onset (BRCA2) is involved in DNA-damage repair via homologous recombination (HR). Defective HR leads to severe genomic instability and homozygous deletion of BRCA2 is therefore deleterious for survival of normal cells. In apparent contrast, inherited heterozygous mutations in BRCA2 increase the risk to develop cancer, which goes along with somatic loss of the wild type allele. So paradoxically, BRCA2 loss is lethal in normal cells, but can be tolerated in cancer cells. It is currently unclear how tumor cells can survive without BRCA2. To address this question, we performed a genome-wide functional genetic screen and identify Tumor Necrosis Factor-alpha (TNFα) signaling, via TNF receptor 1 (TNF-R1) and Src-associated-substrate-during-mitosis-of-68kDa (Sam68), as an important determinant of cell death induction upon BRCA2 inactivation. Material & Methods A genetic screen was performed in human near-haploid KMB-7 cells. Two monoclonal KMB-7 lines were engineered to express independent doxycycline-inducible BRCA2 short hairpin RNAs (shRNAs). Cells were randomly mutagenized using retrovirus carrying a strong splice-acceptor, resulting in gene knock-outs. Deep sequencing of surviving colonies revealed gene mutations that prevented shBRCA2-mediated cell death. Validation of TNF-R1 and Sam68 was done by shRNAs in multiple breast- and leukemic cell lines. Cell viability was determined by MTT and clonogenic assays. Competition assays with vectors expressing shRNA including IRES-driven mCherry were analyzed with flow cytometry. Biochemical analyses were assessed by western blotting and ELISA. Immunoprecipitation with FLAG-tagged TNFα was performed to study TNF-R1 complex formation. Results From our genetic screens, loss-of-function mutations involved in TNFα signaling were found to significantly rescue cell death in BRCA2-depleted cells. To validate these findings, BRCA2 was stably depleted in a panel of cancer cell lines, which decreased viability. Subsequent co-depletion of TNF-R1 or Sam68 rescued survival. Notably, BRCA2 depletion resulted in the production of TNFα. In line with this finding, the majority of tested cancer cell lines were found to be sensitive to recombinant TNFα upon BRCA2 depletion. This phenomenon was not restricted to BRCA2 inactivation, since depletion of related DNA repair genes (BRCA1, FANCD2) or hydroxyurea-induced replication stress also sensitized cells to recombinant TNFα. Mechanistically, BRCA2 depletion was induced to observe differential complex formation of TNF-R1 with downstream effector proteins upon TNFα activation. Conclusion Our data reveal that down-regulation of TNF-R1 or its downstream signaling component Sam68 rescued cytotoxicity upon BRCA2 inactivation. These findings and mechanistic follow-up experiments describe a novel mechanism by which autocrine TNFα signaling, induced by loss of BRCA2, limits tumor cell viability. Citation Format: Anne M. Heijink, Francien G. Talens, Anouk Baars, Marcel A. van Vugt. Tumor necrosis factor-alpha signaling determines cytotoxicity induced by BRCA2 deficiency [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2488. doi:10.1158/1538-7445.AM2017-2488

Abstract 1409: Analysis of 16,172 patient-derived tumor samples indicate TPX2 as being essential for survival of genomically instable cancer cells

Abstract Mutations in homologous recombination (HR) genes, including BRCA1 and BRCA2, compromise DNA repair and lead to genomic instability (GI). GI is lethal to normal cells but is a characteristic of many cancers. Apparently, these cancers are somehow re-wired to survive high levels of GI. Identification of the genetic alterations that allow viability of genomically instable tumor cells may uncover novel therapeutic targets. To elucidate how these tumor cells are rewired, we analyzed publically available mRNA expression data of 16,172 human cancer samples. Functional genomic mRNA profiling (FGmRNA-profiling) was applied on these samples to infer levels of GI and to capture the downstream effects of somatic copy number alterations on gene expression. A genome-wide association analysis was subsequently performed to assess the correlation between FGmRNA signals of individual genes with the degree of GI. From the top 250 genes with strong positive correlation with GI, 11 genes were prioritized based on a co-functionality network in which genes are co-regulated and share similar predicted biological function. The 11 genes that were identified in this cluster were: BIRC5, UBE2C, CENPA, CDCA3, DEK, SKP2, TPX2, KIF2C, RAD21, MYBL2 and WDR67. To validate these findings in genetically-defined models, we engineered a panel of 5 triple negative breast cancer (TNBC) cell lines with doxycycline-inducible shRNAs targeting BRCA2. BRCA2 depletion resulted in a failure of RAD51 foci to localize to DNA double strand breaks which generated isogenic cell line pairs proficient and deficient of HR repair. First, we depleted each of the identified 11 genes using RNA interference in BT-549 cells and observed that depletion of TPX2, a microtubule-associated protein, led to largest differential levels of cell death when comparing the BRCA2-deficient with the BRCA2-proficient context (86.6% vs 32.9% cell death in BRCA2-depleted vs controlled depleted cells respectively). Subsequently, we could replicate this decreased survival with TPX2 depletion in a BRCA2-deficient context in an additional 2 out of 4 other TNBC cell lines. Furthermore, we investigated whether BRCA2-depleted cells were also more sensitive to depletion of Aurora kinase A, a substrate of TPX2. For this purpose, mouse mammary tumor cell lines, derived from Tp53-/- or Brca2-/-;Tp53-/- mice, or a Brca2-reconstituted version thereof were treated with an Aurora A inhibitor, Alisertib. Again, we found that the BRCA2-deficient cell line was more sensitive to Aurora A inhibition than the two BRCA2-proficient cell lines. In conclusion, FGmRNA-profiling of mRNA expression data of human cancer samples identified TPX2 as an essential gene for survival of BRCA2-deficient breast cancer cells, when compared to BRCA2-proficient cells. Thus, targeting the TPX2/AURKA axis could potentially act as a novel therapeutic target in the treatment of genomically instable cancers. Citation Format: Stephanie E. van Gijn, Elles Wierenga, Anne Margriet Heijink, Rudolf S. Fehrmann, Marcel A. van Vugt. Analysis of 16,172 patient-derived tumor samples indicate TPX2 as being essential for survival of genomically instable cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1409. doi:10.1158/1538-7445.AM2017-1409

Abstract 3861: Development of testicular cancer patient derived xenograft models to test combination therapies targeting PI3K/Akt and MDM2

Abstract Metastatic testicular cancer (TC) is highly sensitive to cisplatin-based chemotherapy. However, patients with advanced disease in the poor prognosis group only have a 50% 5-year survival resulting from chemoresistance. Previous data showed that both the PI3K/Akt pathway and the MDM2/p53 axis are involved in cisplatin resistance of TC cells. In this study we aim to establish and characterize TC patient-derived xenografts (PDX) in order to investigate PI3K/Akt and/or MDM2 inhibition as possible treatment options for TC. Excised and minced (8mm3) primary TC tissue was implanted subcutaneously into male NSG mice to obtain a first generation PDX model (F1). Animals were sacrificed when tumor volume exceeded 1500 mm3. The tumor was harvested and minced. Small pieces (8mm3) were implanted in a second generation (F2), as well as stored in FCS-5% DMSO to establish a biobank. In a panel of TC cell lines, combination treatment of Akt inhibition with or without cisplatin or Nutlin-3a was performed. Cleaved caspase-3 staining was assessed to measure apoptosis, and western blot was used to determine the effect of different treatments on sub-cellular localization and phosphorylation status of MDM2. We have established 3 TC PDX models. Histology of the primary tumors included the following subtypes: mixed germ cell tumors with embryonal- and yolk sac carcinoma and teratoma components. Engraftment efficacy was 100% and tumor growth initiated within 4-5 weeks after implantation. Interestingly, we successfully implanted a biopsy taken from a metastatic lesion of a patient presenting with progressive refractory disease after receiving standard chemotherapy. Histology of the different PDX generations was comparable to the patient material, although a loss of the teratoma component was observed. In our cell line panel, Akt inhibition using MK2206 sensitized the acquired cisplatin resistant cell line TeraCP towards cisplatin treatment, whereas an additive effect was observed in other cell lines. Combination of Akt and MDM2 inhibition was highly synergistic in apoptosis induction in all cell lines. No relation was observed between the synergistic effect of Akt combined with MDM2 inhibition and sub-cellular localization or phosphorylation levels of MDM2. Taken together, we have successfully established 3 TC PDX models, including a chemo-resistant model. The biobank is currently being expanded. Combined Akt and MDM2 inhibition resulted in synergistic induction of apoptosis, and these combinations or other novel therapies will be tested in established TC PDX models. Supported by Dutch Cancer Society grant RUG 2014-6691 and CONACYT grant 381543 Citation Format: Gerda de Vries, Ximena Rosas-Plaza, Marcel A. van Vugt, Albert J.H. Suurmeijer, Jourik A. Gietema, Steven de Jong. Development of testicular cancer patient derived xenograft models to test combination therapies targeting PI3K/Akt and MDM2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3861. doi:10.1158/1538-7445.AM2017-3861

Abstract 152: Dual mTORC1/2 inhibition sensitizes testicular cancer cell lines toward cisplatin treatment

Abstract Testicular cancer (TC) patients with metastatic disease and poor prognosis have a 50% 5-year survival. It has been reported that resistant TC cell lines show hyperactivation of the PI3K/Akt/mTOR pathway. In this study we investigated the potential benefit from PI3K, Akt and mTOR inhibition in combination with cisplatin in TC. Our panel of embryonal carcinoma cell lines include the cisplatin-sensitive cells: Tera and 833KE and the cisplatin-resistant cells: Scha, TeraCP and NCCIT. Western blotting showed that the resistant TC cell line TeraCP expresses higher levels of p-Akt compared to the sensitive cell line Tera. To evaluate sensitization towards cisplatin, TC cell lines were treated with cisplatin and/or PI3K inhibitor GDC0941, Akt inhibitor MK2206, mTORC1 inhibitor everolimus and mTORC1/2 inhibitors AZD8055 and AZD2014 for 24 hours and DilC5/PI staining was performed to estimate apoptosis with flow cytometry. All TC cell lines were strongly sensitized by mTORC1/2 inhibition by increasing cisplatin induced apoptosis to 60-80% when cisplatin alone induced only 15-30%. TC cells were also sensitized by mTORC1 and PI3K inhibition but to a lesser extent. Akt inhibition did not sensitize Scha or Tera cells to cisplatin, while TeraCP was marginally sensitized. We then evaluated clonogenic capacity in cells pretreated with AZD8055 for 24 hours and seeded in the presence of different concentrations of cisplatin for 6 days. Clonogenic capacity was reduced in Tera and TeraCP after mTORC1/2 inhibition with AZD8055 compared with control cells treated with cisplatin only. Western blot done with Scha, Tera and TeraCP lysates treated for 24 hours with AZD8055, everolimus, GDC0941 and MK2206 showed that only mTOR inhibition was able to block S6 phosphorylation. In addition, we performed immunohistochemistry of p-S6 and Ki-67 in paraffin embedded tissue from TC patients. IHC showed that patient derived xenografts showed high expression of these markers compared to non-cancerous tissue, meaning that the mTOR pathway is very active in this tumor type. These data indicate that TC relies on the PI3K/Akt/mTOR pathway for survival and mTORC1/2 inhibition showed the stronger sensitizing effect towards cisplatin treatment. We consider adding mTORC1C/2 inhibition to cisplatin based treatment a potential therapeutic option for chemoresistant TC patients that warrants further in vivo investigations. Supported by CONACYT grant 381543 and Dutch Cancer Society grant RUG 2014-6691 Citation Format: Fernanda Ximena Rosas Plaza, Gerda de Vries, Albert J. Suurmeijer, Jourik A. Gietema, Marcel A.T.M. van Vugt, Steven de Jong. Dual mTORC1/2 inhibition sensitizes testicular cancer cell lines toward cisplatin treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 152. doi:10.1158/1538-7445.AM2017-152

Abstract 1406: Towards an RNA expression-based signature for oncogene-induced replication stress

Abstract Introduction Certain tumor subgroups, including triple negative breast cancers (TNBCs), are characterized by high levels of genomic instability (GI). These tumors are typically very aggressive and difficult to threat. Increasingly, we realize that GI is fueled by replication stress (RS). RS can be induced by expression of oncogenes, including CCNE1, MYC and CDC25A, which perturb initiation, elongation and/or termination phases of DNA replication. RS poses a threat to cellular viability, and tumor cells with high levels of RS will therefore increasingly depend for their survival on DNA damage checkpoint and repair pathways. Possibly, this increased dependence creates therapeutic opportunities. Unfortunately, it is currently difficult to determine levels of RS in tumors, a requirement to efficiently select patients for novel treatments. The aim of this work is to develop an RNA expression-based signature for oncogene-induced replication-stress. Material & Methods A panel of cell lines (RPE-1, RPE-1-P53-/-, MDA-MB-231, BT-549 and HCC-1806) was engineered to doxycycline-inducibly express a set of oncogenes (CDC25A, CCNE1 or MYC). Single DNA fiber replication analysis in combination with immunoblot analysis of RS markers was used to validate the cell line models. MTT assays were performed to assess sensitivity to ATR and Wee1 inhibitors. RNA-sequencing was used to uncover expression changes upon RS induction. Results Treatment with doxycycline resulted in high expression levels of CDC25A, CYCLINE or MYC. Expression was heterogeneous but present in the far majority of RPE-1 cells (40% CDC25A, 88% CCNE1 and 93% MYC). DNA fiber analysis was subsequently used to study the alteration of DNA replication dynamics. Severe reduction on ongoing DNA synthesis velocity was shown in engineered cells (RPE-1 and MDA-MB231) under doxycycline treatment, confirming prominent RS. In line with elevated levels of RS, oncogene induction resulted in elevated levels of ATR activity, as judged by phospho-CHK1 and phospho-RPA levels. Importantly, oncogene-expression enhanced sensitivity to Wee1 and ATR checkpoint inhibitors. Subsequently, RNA samples were harvested under the presence and absence of doxycycline. In parallel to oncogene-expression, hydroxyurea, a chemical agent known to increase stalled replication forks was applied as a DNA damage control. RNA-sequencing analysis will be performed to identify differentially expressed genes upon oncogene expression. Commonly identified genes over multiple cell lines and shared between the different oncogenes will be used to build a ‘RS’ profile. Conclusion Our results indicate that oncogene expression provokes RS in multiple cell line models and induces impaired replication fork stability and sensitivity to cell cycle checkpoint inhibitors. These models are therefore valid to develop an RNA expression-based qualifier in order to facilitate patient selection towards novel therapeutics. Citation Format: Sergi Guerrero, Rudolf Fehrmann, Marcel ATM van Vugt. Towards an RNA expression-based signature for oncogene-induced replication stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1406. doi:10.1158/1538-7445.AM2017-1406

Abstract 3581: How to deal with DNA damage during mitosis: a genetic interaction map of the PICH DNA translocase

Abstract Genomic instability is one of the enabling characteristics of cancer, and is thought to be driven in large part by replication stress. It is generally believed that damaged DNA needs to be repaired before entry into mitosis. Recent evidence showed that DNA damage occasionally is present in mitosis, and can be repaired either through break-induced replication during early mitosis or during anaphase at so-called ultrafine DNA bridges (UFBs). These UBFs are characterized by the presence of the PICH DNA translocase, and Rif1, which we recently identified to be involved in the resolution of UFBs (Hengeveld et al, Dev Cell, 2015). However, detailed molecular composition of UFBs associated proteins and their mechanism of action are incompletely understood. Also, whether specific cancer types are dependent on this DNA repair pathway for their survival is not known. We hypothesise that cancer cells with high levels of replication stress may increasingly depend on these mechanisms for their survival. Here, we aimed to identify the genetic make-off of cancer cells that depend for their survival on repair of damage lesion during anaphase. We used inactivation PICH or Rif1 as a model for defective resolution of anaphase bridges. To elucidate the consequences of loss of DNA repair at UFBs, we analysed the sensitivity to genotoxic agents and assessed genomic instability in HAP1 cells, in which the ERCC6L gene, which encodes the PICH DNA translocase, or the Rif1 gene were inactivated using Cas9/CRISPR-mediated mutation. ERCC6L as well as RIF1 knock-out cells showed increased levels of intrinsic DNA damage, as judged by elevated levels of H2AX phosphorylation and increased frequencies of 53BP1 bodies, known to represent unrepaired replication damage. ERCC6L as well as RIF1 knock-out cells also display increased sensitive to the topoisomerase II inhibitor ICRF193. Amplifications and loss of genomic areas was assessed using single strand sequencing of single cells. Subsequently, we wanted to determine which genetic mutations are not allowed or show competitive outgrowth in the absence of UFB resolution during anaphase. To this end, we exploited the haploid nature of the HAP1 cells, which allows insertional mutagenesis using retroviruses carrying a strong spice-acceptor to create random instantaneous knock-outs. The list of candidate genes of which depletion leads to a lethal phenotype specifically in cells lacking PICH is currently being analysed. In conclusion, our studies have shown that the inactivation of PICH or RIF1 results in a marked increase in unresolved UFBs which ultimately lead to elevated numbers of 53BP1 bodies, and increased sensitivity to topoisomerase inhibitors. These data makes both RIF1 and PICH interesting therapeutic targets in tumours with high levels of replication stress, or targets to potentiate the effects of topoisomerases. Citation Format: Inês Teles Alves, Anne-Magriet Heijink, Floris Foijer, Marcel van Vugt. How to deal with DNA damage during mitosis: a genetic interaction map of the PICH DNA translocase. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3581.

36th International Symposium on Intensive Care and Emergency Medicine : Brussels, Belgium. 15-18 March 2016.

36th International Symposium on Intensive Care and Emergency Medicine : Brussels, Belgium. 15-18 March 2016.

Breaks in the 45S rDNA Lead to Recombination-Mediated Loss of Repeats.

rDNA repeats constitute the most heavily transcribed region in the human genome. Tumors frequently display elevated levels of recombination in rDNA, indicating that the repeats are a liability to the genomic integrity of a cell. However, little is known about how cells deal with DNA double-stranded breaks in rDNA. Using selective endonucleases, we show that human cells are highly sensitive to breaks in 45S but not the 5S rDNA repeats. We find that homologous recombination inhibits repair of breaks in 45S rDNA, and this results in repeat loss. We identify the structural maintenance of chromosomes protein 5 (SMC5) as contributing to recombination-mediated repair of rDNA breaks. Together, our data demonstrate that SMC5-mediated recombination can lead to error-prone repair of 45S rDNA repeats, resulting in their loss and thereby reducing cellular viability.

External Validation of Prediction Models for Pneumonia in Primary Care Patients with Lower Respiratory Tract Infection: An Individual Patient Data Meta-Analysis.

BackgroundPneumonia remains difficult to diagnose in primary care. Prediction models based on signs and symptoms (S&S) serve to minimize the diagnostic uncertainty. External validation of these models is essential before implementation into routine practice. In this study all published S&S models for prediction of pneumonia in primary care were externally validated in the individual patient data (IPD) of previously performed diagnostic studies.Methods and FindingsS&S models for diagnosing pneumonia in adults presenting to primary care with lower respiratory tract infection and IPD for validation were identified through a systematical search. Six prediction models and IPD of eight diagnostic studies (N total = 5308, prevalence pneumonia 12%) were included. Models were assessed on discrimination and calibration. Discrimination was measured using the pooled Area Under the Curve (AUC) and delta AUC, representing the performance of an individual model relative to the average dataset performance. Prediction models by van Vugt et al. and Heckerling et al. demonstrated the highest pooled AUC of 0.79 (95% CI 0.74–0.85) and 0.72 (0.68–0.76), respectively. Other models by Diehr et al., Singal et al., Melbye et al., and Hopstaken et al. demonstrated pooled AUCs of 0.65 (0.61–0.68), 0.64 (0.61–0.67), 0.56 (0.49–0.63) and 0.53 (0.5–0.56), respectively. A similar ranking was present based on the delta AUCs of the models. Calibration demonstrated close agreement of observed and predicted probabilities in the models by van Vugt et al. and Singal et al., other models lacked such correspondence. The absence of predictors in the IPD on dataset level hampered a systematical comparison of model performance and could be a limitation to the study.ConclusionsThe model by van Vugt et al. demonstrated the highest discriminative accuracy coupled with reasonable to good calibration across the IPD of different study populations. This model is therefore the main candidate for primary care use.

Abstract P6-08-01: Inverse relationships between high somatic copy number load and immune phenotypes in breast cancer

Abstract Introduction Currently, there is no clear biomarker to predict which breast cancer patient may benefit from immunotherapy. High somatic point mutational load is thought to lead to immune activation and CD8+ T cell activation and infiltration. In addition, pre-existing CD8+ T cells distinctly located at the invasive tumor margin are associated with expression of the PD-1/PD-L1 immune inhibitory axis and may predict response to PD-1 targeted therapy. However, breast tumors are considered copy number-driven rather than mutational-driven cancers. Therefore, in order to identify which breast cancer patients may potentially benefit from immunotherapy, we investigated relations between copy number load and CD8+ T cell abundance, immune pathways and immune activation scores in a large set of publically available breast cancer expression profiles. Methods Functional genomic mRNA-profiling (Fehrmann et al., Nat Genet, 2015) was used to capture the downstream effect of somatic copy number alterations on gene expression levels. This method allowed using gene expression profiles to quantify somatic copy number load occurring in tumor samples in a univariate measurement called the copy number load index (CNL-index). Immune activation scores were calculated according to two known gene immune signatures (Teschendorff et al., Genome Biol, 2007 and Desmedt et al., Clin Cancer Res, 2008). Cibersort (Newman et al., Nat Methods, 2015) was applied to estimate CD8+ T cell abundance. Publicly available gene expression profiles of 7,270 primary breast cancer samples were used. The relation between CNL-index, immune activation scores and disease-free survival (DFS), defined as time from diagnosis until distant metastasis development, was assessed by multivariate cox-regression analysis including age, ER and HER2 status, tumor size, lymph node involvement, tumor grade and treatment regimen. Gene set enrichment analysis (GSEA) was applied to assess relations between immune pathways and CNL-index. Results In primary breast cancer, low CNL-index was found in 1,796 samples (24.7%) versus 5,474 samples with a high CNL-index (75.3%). Higher CNL-index was correlated with shorter DFS (HR 2.51, P = 1.6x10-4), whereas higher immune activation scores were associated with prolonged DFS (HR 0.199, P = 0.003 and HR 0.347, P = 0.017). CD8+ T cell abundance was negatively correlated with the CNL-index (Spearman R = -0.14, P = 8.11x10-34). GSEA showed enrichment of immune pathways amongst genes that negatively correlated with CNL-index. Subset analysis in 1,555 TNBC samples showed low CNL-index in 287 samples (18.5%) versus high CNL-index in 1,268 samples (81.5%). Similar to the entire breast cancer set, CD8+ T cell abundance and CNL-index were negatively correlated in TNBC (Spearman R = -0.11, P = 4.11x10-05). Immune pathway enrichment in genes in TNBC also negatively correlated to CNL-index. Conclusion High CNL-index (i.e. high copy number load) is inversely related to immunoactivation, immune pathways and CD8+ T cell abundance. This implies that in breast cancer, tumors with low CNL-index may be intrinsically sensitive to immune modulation, which warrants further confirmation in prospective trials. This research was supported by Dutch Cancer Society Grants RUG 2010-4739 and RUG 2013-5960. Citation Format: Bense RD, van der Vegt B, de Vries EGE, van Vugt MATM, Schröder CP, Fehrmann RSN. Inverse relationships between high somatic copy number load and immune phenotypes in breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-08-01.

Abstract LB-C23: A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity

Abstract The Wee1 cell cycle checkpoint kinase prevents premature mitotic entry by inhibiting cyclin-dependent kinases. Chemical inhibitors of Wee1 are currently being tested clinically as targeted anti-cancer drugs. Wee1 inhibition is thought to be preferentially cytotoxic in p53-defective cancer cells. However, TP53 mutant cancers do not respond consistently to Wee1 inhibitor treatment, indicating the existence of genetic determinants of Wee1 inhibitor sensitivity other than TP53 status. To optimally facilitate patient selection for Wee1 inhibitor treatment and undercover potential resistance mechanisms, identification of these currently unknown genes and underappreciated functions of Wee1 is necessary. The aim of this study was to identify gene mutations that determine Wee1 inhibitor sensitivity. We performed a genome-wide unbiased functional genetic screen in TP53 mutant near-haploid KBM-7 cells using gene-trap insertional mutagenesis. Insertion site mapping of cells that survived long-term Wee1 inhibition revealed enrichment of G1/S regulatory genes, including SKP2, CUL1 and CDK2. Stable depletion of SKP2, CUL1 and CDK2 or chemical Cdk2 inhibition rescued the γ-H2AX induction and abrogation of G2 phase as induced by Wee1 inhibition in breast and ovarian cancer cell lines. Remarkably, live cell imaging showed that depletion of SKP2, CUL1 and CDK2 did not rescue the Wee1 inhibition-induced karyokinesis and cytokinesis defects. These data indicate that the activity of the DNA replication machinery, beyond TP53 mutation status, determines Wee1 inhibitor sensitivity, and could serve as a selection criterion for Wee1-inhibitor eligible patient. Conversely, loss of the identified S-phase genes could serve as a resistance mechanism and coincides with development of severe genomic instability. Citation Format: Anne Margriet Heijink, Vincent Blomen, Fabian Degener, Felipe Yu Matsushita, Floris Foijer, Marcel A. Van Vugt. A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr LB-C23.

Abstract B19: Quantitative proteomics analysis identifies MUC1 as an effect sensor of EGFR inhibition

Abstract Introduction The epidermal growth factor receptor (EGFR) is often hyperactivated through overexpression or somatic mutation in multiple cancer types, driving tumor growth and survival. Targeted therapies against EGFR show clinical benefit, but resistance to these agents invariably develops. Thus, there is need for markers of effective treatment, termed ‘effect sensors’, to monitor the effective treatment with EGFR therapeutics during therapy. Therefore, we investigated the cellular rewiring at the proteomic level in response to EGFR inhibition, with the aim of identifying proteins that can be used as effect sensors. These effect sensors potentially constitute powerful targets for the development of tracers to monitor EGFR-directed therapies. Materials and Methods Human breast cancer cell lines were tested for cell viability in presence of the EGFR inhibitor erlotinib and sensitive cell lines (SKBR3, SUM149, and BT474) were identified. These cell lines were labeled with stable (SILAC) isotopes and subsequently control-treated or treated with erlotinib for 48 hours. Membranes were then isolated and analyzed with quantitative mass-spectrometry (MS). Results were validated in erlotinib-sensitive (SKBR3, SUM149, and BT474) and resistant breast cancer cell lines (MB-231 and BT549), as well as erlotinib-sensitive human non-small cell lung cancer (NSCLC) cell lines (H292 and HCC827). Results In total, 1,485 overlapping proteins were quantitatively determined in all cell lines, containing 256 plasma membrane proteins as determined using Gene Ontology. Of those, 8 proteins showed at least 1.5 fold up-regulation and were selected for validation. We found MUC1 to be significantly upregulated in response to erlotinib treatment. Upregulation of MUC1 was detected both at protein and mRNA levels in all erlotinib-sensitive breast cancer cell lines, but not in erlotinib-resistant cell lines. In addition, MUC1 levels were elevated upon erlotinib treatment in NSCLC cell lines H292 (EGFR-WT) and HCC827 (EGFR-exon-19 deletion). MUC1 upregulation was observed as early as 48 hours after treatment, and persisted until 14 days with continuous erlotinib treatment. Upregulation of MUC1 was also observed in response to other clinical EGFR inhibitors, including lapatinib and afatinib, as well as in response to the monoclonal EGFR-directed antibody cetuximab. Erlotinib-induced MUC1 expression was dependent on PI3K/AKT/mTOR signaling, since MUC1 expression was completely blocked when EGFR inhibition was combined with either BEZ235, MK2206, or everolimus, which inhibit PI3K, AKT, and mTOR respectively. Importantly, MUC1 upregulation could be measured by flow cytometry, indicating that elevated MUC1 levels are expressed at plasma membranes and underscoring that MUC1 is a potential target for tracer development for medical imaging. Conclusion We identified MUC1 to be specifically upregulated in response to EGFR-directed treatments in breast and lung cancer cell lines. These results warrant in vivo assessment of MUC1 as an effect sensor, to ultimately test whether this strategy can be employed to stratify patients for treatment with EGFR therapeutics. Supported by ERC grant OnQview-2011-293445 to EGE de Vries. Citation Format: Harmen R. de Boer, Esméé Joosten, Fabrizia Fusetti, Rudolf S.N. Fehrmann, Elisabeth G.E. de Vries, Marcel A.T.M. van Vugt. Quantitative proteomics analysis identifies MUC1 as an effect sensor of EGFR inhibition. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B19.

Commentary: When the brain takes a break: a model-based analysis of mind wandering.

Commentary: When the brain takes a break: a model-based analysis of mind wandering.