Preclinical Identification Research Articles

Coronary Calcium Score Using Multi-Slice Computed Tomography

Coronary atherosclerosis and coronary artery disease (CAD) are the most common causes of death and disability in the most of industrialized countries. The problem of early diagnosis of CAD detection is extremely relevant all over the world. Preclinical identification of patients with risk factors of CAD is one of the most important goal in routine clinical practice. Multislice computed tomography (MSCT) of the heart with the determination of the calcium index (CI) is a noninvasive screening of coronary arteries assessment. The severity of coronary calcification has a close relationship with the severity of coronary atherosclerosis and the risk of acute coronary events according to numerous studies. Initially, studies targeted on assessment of coronary calcification and determination of normative parameters were performed using electron-beam scanners (EBS). Subsequently multislice computed tomography (MSCT) replaced the electronic beam tomography (EBCT). The results of studies that performed with EBS have become the basis for a methodology of coronary calcification assessment with MSCT. The reproducibility of coronary calcium score is important for assessment of atherosclerosis for dynamic monitoring. The inter- observer and intra-observer reproducibility of this method is quite high, the reproducibility according to repeated studies of the same patient is lower. In recent years, a new generation of volumetric CT- scanners has been introduced into clinical practice. Reducing the radiation dose for coronary calcium screening to the minimum values (less than 1 mSv) is one of the important advantages of volumetric computed tomography. Currently, new studies are needed to assess the reproducibility of coronary calcium index calculation using a new generation of volumetric CT-scanners.

Human brown adipose tissue as a target for obesity management; beyond cold-induced thermogenesis.

Elevating energy expenditure via adaptive thermogenesis in brown adipose tissue (BAT) is a potential strategy to reverse obesity. Much early enthusiasm for this approach, based on rodent studies, was tempered by the belief that BAT was relatively inconsequential in healthy adult humans. Interest was reinvigorated a decade ago when a series of studies re-identified BAT, primarily in upper thoracic regions, in adults. Despite the ensuing explosion of pre-clinical investigations and identification of an extensive list of potential target molecules for BAT recruitment, our understanding of human BAT physiology remains limited, particularly regarding interventions which might hold therapeutic promise. Cold-induced BAT thermogenesis (CIT) has been well studied, although is not readily translatable as an anti-obesity approach, whereas little is known regarding the role of BAT in human diet-induced thermogenesis (DIT). Furthermore, human studies dedicated to translating known pharmacological mechanisms of adipose browning from animal models are sparse. Several lines of recent evidence suggest that molecular regulation and physiology of human BAT differ to that of laboratory rodents, which form the majority of our knowledge base. This review will summarize knowledge on CIT and expand upon the current understanding and evidence gaps related to human adaptive thermogenesis via mechanisms other than cold.

Use of Rat Primary Mesenteric Cells for the Prediction of PDE4 Inhibitor Drug-Induced Vascular Injury.

Drug-induced vascular injury (DIVI) in preclinical studies can delay, if not terminate, a drug development program. Clinical detection of DIVI can be very difficult as there are no definitive biomarkers known to reliably detect this disorder in all instances. The preclinical identification of DIVI requires detailed microscopic examination of a wide range of tissues although one of the most commonly affected areas in rats is the mesenteric vasculature. The reason for this predisposition of mesenteric arteries in rats as well as the exact mechanism and cell types involved in the initial development of these lesions have not been fully elucidated. We hypothesized that by using a mixed culture of cells from rat mesenteric tissue, we would be able to identify an RNA expression signature that could predict the invivo development of DIVI. Five compounds designed to inhibit Phosphodiesterase 4 activity (PDE4i) were chosen as positive controls. PDE4i's are well known to induce DIVI in the mesenteric vasculature of rats and there is microscopic evidence that this is associated, at least in part, with a proinflammatory mechanism. We surveyed, by qRT-PCR, the expression of 96 genes known to be involved in inflammation and using a Random-Forest model, identified 12 genes predictive of invivo DIVI outcomes in rats. Using these genes, we were able to cross-validate the ability of the Random-Forest modeling to predict the concentration at which PDE4i caused DIVI invivo.

Common pitfalls in preclinical cancer target validation.

An alarming number of papers from laboratories nominating new cancer drug targets contain findings that cannot be reproduced by others or are simply not robust enough to justify drug discovery efforts. This problem probably has many causes, including an underappreciation of the danger of being misled by off-target effects when using pharmacological or genetic perturbants in complex biological assays. This danger is particularly acute when, as is often the case in cancer pharmacology, the biological phenotype being measured is a 'down' readout (such as decreased proliferation, decreased viability or decreased tumour growth) that could simply reflect a nonspecific loss of cellular fitness. These problems are compounded by multiple hypothesis testing, such as when candidate targets emerge from high-throughput screens that interrogate multiple targets in parallel, and by a publication and promotion system that preferentially rewards positive findings. In this Perspective, I outline some of the common pitfalls in preclinical cancer target identification and some potential approaches to mitigate them.

Insights from animal models of bladder cancer: recent advances, challenges, and opportunities.

Bladder cancer (urothelial cancer of the bladder) is the most common malignancy affecting the urinary system with increasing incidence and mortality. Treatment of bladder cancer has not advanced in the past 30 years. Therefore, there is a crucial unmet need for novel therapies, especially for high grade/stage disease that can only be achieved by preclinical model systems that faithfully recapitulate the human disease. Animal models are essential elements in bladder cancer research to comprehensively study the multistep cascades of carcinogenesis, progression and metastasis. They allow for the investigation of premalignant phases of the disease that are not clinically encountered. They can be useful for identification of diagnostic and prognostic biomarkers for disease progression and for preclinical identification and validation of therapeutic targets/candidates, advancing translation of basic research to clinic. This review summarizes the latest advances in the currently available bladder cancer animal models, their translational potential, merits and demerits, and the prevalent tumor evaluation modalities. Thereby, findings from these model systems would provide valuable information that can help researchers and clinicians utilize the model that best answers their research questions.

Nuevas técnicas de imagen cardiaca en la detección precoz de cardiotoxicidad secundaria a tratamientos oncológicos

The undeniable advances in the field of oncology have finally led to a decrease in overall cancer-related mortality. However, this population of long-term cancer survivors is now facing a shift toward a substantial increase in cardiovascular morbidity and mortality. Because the development of overt cardiotoxicity can be associated with poor outcomes, preclinical identification of cardiac toxicity is important. This will promote early instauration of treatments to prevent overt heart dysfunction and allow oncologists to continue cancer therapy in an uninterrupted manner. Surveillance strategies for the early detection of cardiac injury include cardiac imaging and biomarkers during treatment. In this review, we outline existing cardiac imaging modalities to detect myocardial changes in patients undergoing cancer treatment and in survivors, and their strengths and limitations.

Identifying fast-onset antidepressants using rodent models.

Depression is a leading cause of disability worldwide and a major contributor to the burden of suicide. A major limitation of classical antidepressants is that 2-4 weeks of continuous treatment is required to elicit therapeutic effects, prolonging the period of depression, disability and suicide risk. Therefore, the development of fast-onset antidepressants is crucial. Preclinical identification of fast-onset antidepressants requires animal models that can accurately predict the delay to therapeutic onset. Although several well-validated assay models exist that predict antidepressant potential, few thoroughly tested animal models exist that can detect therapeutic onset. In this review, we discuss and assess the validity of seven rodent models currently used to assess antidepressant onset: olfactory bulbectomy, chronic mild stress, chronic forced swim test, novelty-induced hypophagia (NIH), novelty-suppressed feeding (NSF), social defeat stress, and learned helplessness. We review the effects of classical antidepressants in these models, as well as six treatments that possess fast-onset antidepressant effects in the clinic: electroconvulsive shock therapy, sleep deprivation, ketamine, scopolamine, GLYX-13 and pindolol used in conjunction with classical antidepressants. We also discuss the effects of several compounds that have yet to be tested in humans but have fast-onset antidepressant-like effects in one or more of these antidepressant onset sensitive models. These compounds include selective serotonin (5-HT)2C receptor antagonists, a 5-HT4 receptor agonist, a 5-HT7 receptor antagonist, NMDA receptor antagonists, a TREK-1 receptor antagonist, mGluR antagonists and (2R,6R)-HNK. Finally, we provide recommendations for identifying fast-onset antidepressants using rodent behavioral models and molecular approaches.

Emerging Cardiac Imaging Modalities for the Early Detection of Cardiotoxicity Due to Anticancer Therapies

The undeniable advances in the field of oncology have finally led to a decrease in overall cancer-related mortality. However, this population of long-term cancer survivors is now facing a shift toward a substantial increase in cardiovascular morbidity and mortality. Because the development of overt cardiotoxicity can be associated with poor outcomes, preclinical identification of cardiac toxicity is important. This will promote early instauration of treatments to prevent overt heart dysfunction and allow oncologists to continue cancer therapy in an uninterrupted manner. Surveillance strategies for the early detection of cardiac injury include cardiac imaging and biomarkers during treatment. In this review, we outline existing cardiac imaging modalities to detect myocardial changes in patients undergoing cancer treatment and in survivors, and their strengths and limitations.

SC21.04 Patient-Derived Xenografts for Guiding Therapy of Lung Cancer

Preclinical drug screening and biomarker discovery in the NCI-60 cancer cell line panel as well as the xenograft developed by growing these cell lines subcutaneously in immunodeficient mice have repeatedly failed to predict clinical responses. In an attempt to circumvent the limited predictive values of conventional preclinical models, there has been increasing attention in the development and characterization of Patient-derived tumor xenograft (PDX) models. The PDX models, which were created by direct implantation of patient’s tumor in immunodeficient mice, have shown to reflect principal histologic and genetic characteristics of original patient tumors and retain tumor heterogeneity better than any other preclinical model. These models have been shown to be predictive of clinical outcomes and are being used for translational research, preclinical drug screening and biomarker identification and validation. The PDX model may also be used in the application of ‘co-clinical trial’ approach, in which it is developed from a patient enrolled in a clinical trial and treated with the same experimental agents to emulate clinical response. This strategy permits the assessment of drug response simultaneously in the patient and mouse model, providing an interesting platform to investigate resistance mechanism, predictive biomarkers and novel combination strategies in a real-time manner. I will present the utility of PDX models, which faithfully replicated the histologic, genomic and pharmacologic features observed in the original patients, and ‘co-clinical trial’ that mirror a phase II trial of agents targeting fibroblast growth factor receptor (FGFR) in non-small cell lung cancer. Predictive biomarker, patient-derived tumor xenograft, co-clinical trial

Ultrasensitive Detection of Porcine Epidemic Diarrhea Virus from Fecal Samples Using Functionalized Nanoparticles.

Porcine epidemic diarrhea virus (PEDV) is the main causative agent of porcine diarrhea, which has resulted in devastating damage to swine industry and become a perplexed global problem. PEDV infection causes lesions and clinical symptoms, and infected pigs often succumb to severe dehydration. If there is not a timely and effective method to control its infection, PEDV will spread rapidly across the whole swine farm. Therefore, preclinical identification of PEDV is of great significance for preventing the outbreak and spread of this disease. In this study, a functionalized nanoparticles-based PCR method (UNDP-PCR) specific for PEDV was developed through systematic optimization of functionalized magnetic beads and gold nanoparticles which were further used to specifically enrich viral RNA from the lysate of PEDV stool samples, forming a MMPs-RNA-AuNPs complex. Then, oligonucleotides specific for PEDV coated on AuNPs were eluted from the complex and were further amplified and characterized by PCR. The detection limitation of the established UNDP-PCR method for PEDV was 25 copies in per gram PEDV stool samples, which is 400-fold more sensitive than conventional RT-PCR for stool samples. The UNDP-PCR for PEDV exhibited reliable reproducibility and high specificity, no cross-reaction was observed with other porcine viruses. In 153 preclinical fecal samples, the positive detection rate of UNDP-PCR specific for PEDV (30.72%) was much higher than that of conventional RT-PCR (5.88%) and SYBR Green real-time RT-PCR. In a word, this study provided a RNA extraction and transcription free, rapid and economical method for preclinical PEDV infection, which showed higher sensitivity, specificity and reproducibility, and exhibited application potency for evaluating viral loads of preclinical samples.

Development of a Biochemical Diagnosis of Parkinson Disease by Detection of α-Synuclein Misfolded Aggregates in Cerebrospinal Fluid

Parkinson disease (PD) is a highly prevalent and incurable neurodegenerative disease associated with the accumulation of misfolded α-synuclein (αSyn) aggregates. An important problem in this disease is the lack of a sensitive, specific, and noninvasive biochemical diagnosis to help in clinical evaluation, monitoring of disease progression, and early differential diagnosis from related neurodegenerative diseases. To develop a novel assay with high sensitivity and specificity to detect small quantities of αSyn aggregates circulating in cerebrospinal fluid (CSF) of patients affected by PD and related synucleinopathies. The strategy evaluated in this proof-of-concept study uses the protein misfolding cyclic amplification (PMCA) technology that detects minute amounts of misfolded oligomers by taking advantage of their ability to nucleate further aggregation, enabling a very high amplification of the signal. The technology was first adapted with synthetic αSyn oligomers prepared in vitro and used to screen in 2 blinded cohorts of CSF samples from German and Japanese patients with PD (n = 76) and individuals serving as controls affected by other neurologic disorders (n = 65), neurodegenerative diseases (n = 18), and Alzheimer disease (n = 14). The kinetics of αSyn aggregation were measured by αSyn-PMCA in the presence of CSF samples from the participants to detect αSyn oligomeric seeds present in this biological fluid. The assays were conducted from November 15, 2013, to August 28, 2015. Kinetic parameters correlated with disease severity at the time of sample collection, measured by the Hoehn and Yahr scale, with the lowest grade indicating unilateral involvement with minimal or no functional impairment, and the highest grade defining patients with complete confinement to wheelchair or bed. Studies with synthetic αSyn aggregates showed that αSyn-PMCA enabled to detect as little as 0.1 pg/mL of αSyn oligomers. The αSyn-PMCA signal was directly proportional to the amount of αSyn oligomers added to the reaction. A blinded study of CSF samples correctly identified patients affected by PD with an overall sensitivity of 88.5% (95% CI, 79.2%-94.6%) and specificity of 96.9% (95% CI, 89.3%-99.6%). The αSyn-PMCA results for different patients correlated with the severity of the clinical symptoms of PD (Japanese cohort: rs = -0.54, P = .006; German cohort: rs = -0.36, P = .02). The findings suggest that detection of αSyn oligomers by αSyn-PMCA in the CSF of patients affected by PD may offer a good opportunity for a sensitive and specific biochemical diagnosis of the disease. Further studies are needed to investigate the usefulness of αSyn-PMCA to monitor disease progression and for preclinical identification of patients who may develop PD.

Reference Values for Cardiac and Aortic Magnetic Resonance Imaging in Healthy, Young Caucasian Adults.

BackgroundReference values for morphological and functional parameters of the cardiovascular system in early life are relevant since they may help to identify young adults who fall outside the physiological range of arterial and cardiac ageing. This study provides age and sex specific reference values for aortic wall characteristics, cardiac function parameters and aortic pulse wave velocity (PWV) in a population-based sample of healthy, young adults using magnetic resonance (MR) imaging.Materials and MethodsIn 131 randomly selected healthy, young adults aged between 25 and 35 years (mean age 31.8 years, 63 men) of the general-population based Atherosclerosis-Monitoring-and-Biomarker-measurements-In-The-YOuNg (AMBITYON) study, descending thoracic aortic dimensions and wall thickness, thoracic aortic PWV and cardiac function parameters were measured using a 3.0T MR-system. Age and sex specific reference values were generated using dedicated software. Differences in reference values between two age groups (25–30 and 30–35 years) and both sexes were tested.ResultsAortic diameters and areas were higher in the older age group (all p<0.007). Moreover, aortic dimensions, left ventricular mass, left and right ventricular volumes and cardiac output were lower in women than in men (all p<0.001). For mean and maximum aortic wall thickness, left and right ejection fraction and aortic PWV we did not observe a significant age or sex effect.ConclusionThis study provides age and sex specific reference values for cardiovascular MR parameters in healthy, young Caucasian adults. These may aid in MR guided pre-clinical identification of young adults who fall outside the physiological range of arterial and cardiac ageing.

Preclinical identification of vaccine induced protective correlates in human leukocyte antigen expressing transgenic mice infected with Coccidioides posadasii

There is an emerging interest to develop human vaccines against medically-important fungal pathogens and a need for a preclinical animal model to assess vaccine efficacies and protective correlates. HLA-DR4 (DRB1∗0401 allele) transgenic mice express a human major histocompatibility complex class II (MHC II) receptor in such a way that CD4+ T-cell response is solely restricted by this human molecule. In this study HLA-DR4 transgenic mice were immunized with a live-attenuated vaccine (ΔT) and challenged by the intranasal route with 50–70 Coccidioides posadasii spores, a potentially lethal dose. The same vaccination regimen offers 100% survival for C57BL/6 mice. Conversely, ΔT-vaccinated HLA-DR4 mice displayed 3 distinct manifestations of Coccidioides infection including 40% fatal acute (FAD), 30% disseminated (DD) and 30% pulmonary disease (PD). The latter 2 groups of mice had reduced loss of body weight and survived to at least 50days postchallenge (dpc). These results suggest that ΔT vaccinated HLA-DR4 mice activated heterogeneous immunity against pulmonary Coccidioides infection. Vaccinated HLA-DR4 mice displayed early expansion of Th1 and Th17 cells and recruitment of inflammatory innate cells into Coccidioides-infected lungs during the first 9dpc. While contraction rates of Th cells and the inflammatory response during 14–35dpc significantly differed among the 3 groups of vaccinated HLA-DR4 mice. The FAD group displayed a sharply reduced Th1 and Th17 response, while overwhelmingly recruiting neutrophils into lungs during 9–14days. The FAD group approached moribund by 14dpc. In contrast, vaccinated HLA-DR4 survivors gradually contracted Th cells and inflammatory response with the greatest rate in the PD group. While vaccinated HLA-DR4 mice are susceptible to Coccidioides infection, they are useful for evaluation of vaccine efficacy and identification of immunological correlates against this mycosis.

Human iPSC-Derived Neuronal Model of Tau-A152T Frontotemporal Dementia Reveals Tau-Mediated Mechanisms of Neuronal Vulnerability.

SummaryFrontotemporal dementia (FTD) and other tauopathies characterized by focal brain neurodegeneration and pathological accumulation of proteins are commonly associated with tau mutations. However, the mechanism of neuronal loss is not fully understood. To identify molecular events associated with tauopathy, we studied induced pluripotent stem cell (iPSC)-derived neurons from individuals carrying the tau-A152T variant. We highlight the potential of in-depth phenotyping of human neuronal cell models for pre-clinical studies and identification of modulators of endogenous tau toxicity. Through a panel of biochemical and cellular assays, A152T neurons showed accumulation, redistribution, and decreased solubility of tau. Upregulation of tau was coupled to enhanced stress-inducible markers and cell vulnerability to proteotoxic, excitotoxic, and mitochondrial stressors, which was rescued upon CRISPR/Cas9-mediated targeting of tau or by pharmacological activation of autophagy. Our findings unmask tau-mediated perturbations of specific pathways associated with neuronal vulnerability, revealing potential early disease biomarkers and therapeutic targets for FTD and other tauopathies.

Whole Blood Gene Expression Profiling in Preclinical and Clinical Cattle Infected with Atypical Bovine Spongiform Encephalopathy.

Prion diseases, such as bovine spongiform encephalopathies (BSE), are transmissible neurodegenerative disorders affecting humans and a wide variety of mammals. Variant Creutzfeldt-Jakob disease (vCJD), a prion disease in humans, has been linked to exposure to BSE prions. This classical BSE (cBSE) is now rapidly disappearing as a result of appropriate measures to control animal feeding. Besides cBSE, two atypical forms (named H- and L-type BSE) have recently been described in Europe, Japan, and North America. Here we describe the first wide-spectrum microarray analysis in whole blood of atypical BSE-infected cattle. Transcriptome changes in infected animals were analyzed prior to and after the onset of clinical signs. The microarray analysis revealed gene expression changes in blood prior to the appearance of the clinical signs and during the progression of the disease. A set of 32 differentially expressed genes was found to be in common between clinical and preclinical stages and showed a very similar expression pattern in the two phases. A 22-gene signature showed an oscillating pattern of expression, being differentially expressed in the preclinical stage and then going back to control levels in the symptomatic phase. One gene, SEL1L3, was downregulated during the progression of the disease. Most of the studies performed up to date utilized various tissues, which are not suitable for a rapid analysis of infected animals and patients. Our findings suggest the intriguing possibility to take advantage of whole blood RNA transcriptional profiling for the preclinical identification of prion infection. Further, this study highlighted several pathways, such as immune response and metabolism that may play an important role in peripheral prion pathogenesis. Finally, the gene expression changes identified in the present study may be further investigated as a fingerprint for monitoring the progression of disease and for developing targeted therapeutic interventions.

"Braking" the Cycle of Resistance in Endocrine Therapy for Breast Cancer.

Endocrine resistance leads to recurrence and death from breast cancer. Animal models of endocrine resistance enable preclinical identification of efficacious therapeutic combinations and further our understanding of resistance. This strategy provides new insights into optimally targeting interactions between estrogen receptor (ESR-1) activity and the cell cycle by CDK4/6 inhibitors. See related article by Wardell et al., p. 5121.

Abstract 2639: Preclinical evaluation and biomarker identification for the anti-LGR5 mAb BNC101 in K-Ras mutant CRC and other solid tumor indications

Abstract BNC101 is a monoclonal antibody (mAb) targeting LGR5 that has successfully completed IND-enabling studies in preparation for Phase I clinical studies in 2015. LGR5 is a validated cancer stem cell (CSC) receptor overexpressed in colorectal cancer (CRC), pancreatic cancer and most other solid tumors. Loss and gain-of-function studies indicate that LGR5 is a functional cancer receptor involved in tumor growth and survival. Sorted LGR5+ primary CRC are highly tumorigenic compared to LGR5- cells in limiting dilution in vivo xenograft studies. BNC101 was selected as the lead LGR5 candidate for clinical development using Bionomics’ CSC Rx Discovery™ platform. Key lead selection criteria for BNC101 were anti-tumor and anti-CSC activity in vitro and in vivo, in patient-derived xenograft (PDX) tumor models from multiple tumor types. CSC activity was evaluated by functional in vivo limiting dilution assays (LDA) using tumor cells treated with BNC101 +/- standard-of-care chemotherapy (SOC) combinations. In LDA studies with the K-Ras mutant CRC PDX tumors CT1 and CT3, 5/8 (63%) and 6/8 (75%) mice implanted with serially diluted cells from BNC101 treated tumors remained tumor free, compared to 1/8 (13%) and 2/8 (25%) in the control groups. Combination with SOC further improved BNC101 activity against CT3 tumors, where 8/8 mice re-implanted for LDA had 6 months progression-free-survival (vs 3/8 in chemo group). In the pancreatic PDX model JH109, BNC101 combined with SOC flat-lined established tumors in 3/7 mice, compared to 0/7 tumors eradicated with control SOC alone. BNC101 is also active in multiple pancreatic, triple-negative breast (TNBC) and small-cell lung (SCLC) cancer xenograft models. Ongoing translational studies have identified a panel of Wnt genes modulated by BNC101 treatment that could potentially be used to identify BNC101 responsive versus resistant patients in the clinic. Furthermore, BNC101 treatment significantly reduced LGR5+ HLA+ circulating-tumor-cells (CTCs) in the peripheral blood of tumor-bearing mice, providing a pharmacodynamic (PD) biomarker of target engagement to further evaluate in clinical studies. Altogether, the preclinical data package supports our therapeutic hypothesis that BNC101 targeting of LGR5+ CSCs will significantly improve PFS and OS in solid tumor indications. Additional translational biomarker data to support the clinical development of BNC101 will also be presented. Citation Format: Peter Chu, Kristen Smith, Farbod Shojaei, Colin Walsh, John Norton, Jose Iglesias, Christopher Reyes. Preclinical evaluation and biomarker identification for the anti-LGR5 mAb BNC101 in K-Ras mutant CRC and other solid tumor indications. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2639. doi:10.1158/1538-7445.AM2015-2639

Variation in the Williams syndrome GTF2I gene and anxiety proneness interactively affect prefrontal cortical response to aversive stimuli.

Characterizing the molecular mechanisms underlying the heritability of complex behavioral traits such as human anxiety remains a challenging endeavor for behavioral neuroscience. Copy-number variation (CNV) in the general transcription factor gene, GTF2I, located in the 7q11.23 chromosomal region that is hemideleted in Williams syndrome and duplicated in the 7q11.23 duplication syndrome (Dup7), is associated with gene-dose-dependent anxiety in mouse models and in both Williams syndrome and Dup7. Because of this recent preclinical and clinical identification of a genetic influence on anxiety, we examined whether sequence variation in GTF2I, specifically the single-nucleotide polymorphism rs2527367, interacts with trait and state anxiety to collectively impact neural response to anxiety-laden social stimuli. Two hundred and sixty healthy adults completed the Tridimensional Personality Questionnaire Harm Avoidance (HA) subscale, a trait measure of anxiety proneness, and underwent functional magnetic resonance imaging (fMRI) while matching aversive (fearful or angry) facial identity. We found an interaction between GTF2I allelic variations and HA that affects brain response: in individuals homozygous for the major allele, there was no correlation between HA and whole-brain response to aversive cues, whereas in heterozygotes and individuals homozygous for the minor allele, there was a positive correlation between HA sub-scores and a selective dorsolateral prefrontal cortex (DLPFC) responsivity during the processing of aversive stimuli. These results demonstrate that sequence variation in the GTF2I gene influences the relationship between trait anxiety and brain response to aversive social cues in healthy individuals, supporting a role for this neurogenetic mechanism in anxiety.

Value of monitoring Nrf2 activity for the detection of chemical and oxidative stress.

Beyond specific limits of exposure, chemical entities can provoke deleterious effects in mammalian cells via direct interaction with critical macromolecules or by stimulating the accumulation of reactive oxygen species (ROS). In particular, these chemical and oxidative stresses can underpin adverse reactions to therapeutic drugs, which pose an unnecessary burden in the clinic and pharmaceutical industry. Novel pre-clinical testing strategies are required to identify, at an earlier stage in the development pathway, chemicals and drugs that are likely to provoke toxicity in humans. Mammalian cells can adapt to chemical and oxidative stress via the action of the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), which up-regulates the expression of numerous cell defence genes and has been shown to protect against a variety of chemical toxicities. Here, we provide a brief overview of the Nrf2 pathway and summarize novel experimental models that can be used to monitor changes in Nrf2 pathway activity and thus understand the functional consequences of such perturbations in the context of chemical and drug toxicity. We also provide an outlook on the potential value of monitoring Nrf2 activity for improving the pre-clinical identification of chemicals and drugs with toxic liability in humans.

SRC inhibition represents a potential therapeutic strategy in liposarcoma

Liposarcomas (LS) are the most common malignant mesenchymal tumors, with an overall long-term mortality rate of 60%. LS comprise three major subtypes, i.e., well-differentiated/dedifferentiated liposarcoma (WDLS/DDLS), myxoid/round cell liposarcoma (MLS) and pleomorphic liposarcoma (PLS). Aiming at the preclinical identification of novel therapeutic options, we here investigate the functional significance of SRC in primary human LS and in LS-derived cell lines. Immunohistochemical and Western blot analyses reveal relevant levels of activated p-(Tyr416)-SRC in LS of the different subtypes with particular activation in MLS and PLS. Dysregulation of the SRC modifiers CSK and PTP1B was excluded as major reason for the activation of the kinase. Consistent siRNA-mediated knockdown of SRC or inhibition by the SRC inhibitor Dasatinib led to decreased proliferation of LS cell lines of the different subtypes, with MLS cells reacting particularly sensitive in MTT assays. Flow cytometric analyses revealed that this effect was due to a significant decrease in mitotic activity and an induction of apoptosis. SRC inhibition by Dasatinib resulted in dephosphorylation of SRC itself, its interacting partners FAK and IGF-IR as well as its downstream target AKT. Consistent with a particular role of SRC in cell motility, Dasatinib reduced the migratory and invasive potential of MLS cells in Boyden chamber and Matrigel chamber assays. In summary, we provide evidence that SRC activation plays an important role in LS biology and therefore represents a potential therapeutic target, particularly in MLS and PLS.