Thoracic aortic aneurysm (TAA) is an asymptomatic and progressive dilatation of the thoracic aorta. Ascending aortic dissection (AAD) is an acute intraparietal tear, occurring or not on a pre-existing dilatation. AAD is a condition associated with a poor prognosis and a high mortality rate. TAA and AAD share common etiology as monogenic diseases linked to transforming growth factor β signaling pathway, extracellular matrix defect, or smooth muscle cell protein mutations. They feature a complex pathogenesis including loss of smooth muscle cells, altered phenotype, and extracellular matrix degradation in aortic media layer. A better knowledge of the mechanisms responsible for TAA progression and AAD occurrence is needed to improve healthcare, nowadays mainly consisting of aortic open surgery or endovascular replacement. Recent breakthrough discoveries allowed a deeper characterization of the mechanisms of gene regulation. Since alteration in gene expression has been linked to TAA and AAD, it is conceivable that a better knowledge of the causes of this alteration may lead to novel theranostic approaches. In this review article, the authors will focus on epigenetic regulation of gene expression, including the role of histone methylation and acetylation, deoxyribonucleic acid methylation, and noncoding ribonucleic acids in the pathogenesis of TAA and AAD. They will provide a translational perspective, presenting recent data that motivate the evaluation of the potential of epigenetics to diagnose TAA and prevent AAD. Aortic progressive dilation (aneurysm) is a potentially life-threatening condition that puts patients at risk of aortic dissection or rupture, a devastating clinical event with high mortality. Most aneurysms are painless and cause no symptomatology to alert the patient or clinician to the associated risk. Unfortunately, the hidden nature of progressive extracellular matrix (ECM) defect (with or without dilation) means that often the diagnosis is only made at the time of an aortic dissection. Thousands of deaths annually can be attributed to aortic aneurysms and dissections.Epigenetics can be defined as stable heritable traits independent of changes in deoxyribonucleic acid (DNA) sequence. In many cardiovascular disorders, significant epigenetic modifications have been shown to affect disease development or progression. Work has progressed most rapidly in cardiomyocytic lineages where intensive effort has gone into understanding transcriptional networks at the epigenetic level.Dilating remodeling of arterial tissues is defined as the structural consequence of dysregulated biological activities controlling the ECM and cell homeostasis within the wall. Aneurysms, regardless of their localization, are characterized by breakdown of the ECM, and aortic smooth muscle cell (aSMC) loss, leading to progressive dilation of the arteries, thinning of the medial layer, and, eventually leading to rupture of the arterial wall. Risk factors for aortic aneurysm are dependent on the regional propensity of the affected segment. Descending thoracic aortic aneurysm (DTAA) demonstrates a pathology characterized by atheroma, aSMC disarray and apoptosis, intense phagocytosis, and intraluminal thrombus usually observed in abdominal aortic aneurysms (AAA). Enzymatic degradation of the aortic ECM leads to structural weakening, dilation, and eventually rupture if left untreated.Thoracic aortic aneurysm, a progressive dilation of the thoracic ascending aorta, is a chronic pathology involving an imbalance between proteolytic degradation of the ECM and wall compensating resistance to proteolytic injury,The identity of genes mediating TAA often represents ECM components or regulators and genes required for the contractile function of aSMC. Increased expression of matrix-degrading enzymes, mucoid accumulation, and aSMC apoptosis are commonly noted in most forms of aneurysmal disease. Disruption of structural ECM components such as collagens and elastin is a commonly accepted mechanism of aneurysm pathogenesis. It comes as no surprise, therefore, that mutations that directly influence the ECM components can cause aneurysm. Examples include mutations in ELN,In addition to genetic changes that directly alter ECM components, human mutations have been discovered that interfere with vascular smooth muscle-mediated aortic homeostasis in genes encoding members of the smooth muscle contraction apparatus, such as MYH11,Aortic dissection is an acute intramural rupture, which can occur in a normal or moderately dilated aorta,TAA and AAD share common etiologies, including monogenic diseases in young patients.As displayed inHistones are DNA-associated proteins responsible for chromatin compaction. They are prone to modifications such as acetylation, methylation, phosphorylation, adenosine diphosphate-ribosylation, and sumoylation that generally occur on specific amino acid residues (arginine and lysine) of their N-extremity.Performed by DNA methyltransferases,Noncoding RNAs are defined as RNA molecules lacking protein-coding potential. They are generally classified according to their size: small noncoding RNAs contain less than 200 nucleotides, while long noncoding RNAs (lncRNAs) contain at least 200 base pairs. Different types of small noncoding RNAs have been characterized, among which 20-nucleotides-long microRNAs (miRNAs) predominate.The mechanisms of gene regulation by lncRNAs are more complex and diverse than miRNAs. LncRNAs can act as signal, decoy, guide, scaffold, or enhancer.The presence of miRNAs in the circulation was revealed almost a decade agoExperimental studies reported an activation of TGF-β in aneurysms related to Marfan syndrome.In a second study performed on human aortic samples, including TAAs of different etiologies and normal aortas from which tissue extracts and aSMCs and fibroblasts were obtained in primary cultures, it was observed that all types of TAA share a complex dysregulation of Smad2 signaling, independent of TGF-β1 in TAA-derived aSMCs.In a third study, the histone-modifying enzymes, transcription factors, and cofactors responsible for Smad2 promoter activation in aSMC from TAA patients were explored to understand the mechanisms regulating Smad2 overexpression. It was shown that Smad2 promoter activation is driven by the recruitment of a multipartner complex, including the transcription factor p53 and histone acetyltransferases.In a fourth study, protease nexin-1 (PN-1) and plasminogen activator inhibitor-1 (PAI-1), both inhibitors of serine-protease were overexpressed in medial tissue extracts and primary aSMCs from TAA compared with acute dissections of ascending aortic and healthy aorta.A genome-wide DNA methylation study comparing aortic tissues from patients with two different etiologies of TAA, bicuspid and tricuspid aortic valves, revealed that several genes involved in cardiovascular development were differentially methylated,Although the functional contribution of miRNAs to the development of TAA and progression toward dissection is still poorly characterized, some associations between miRNAs expression levels, TAA and AAD, have been reported .Technological developments of high-throughput approaches such as microarray facilitated the characterization of disease-associated miRNA profiles.Using microarray and quantitative polymerase chain reaction (PCR), a panel of four up-regulated and three down-regulated miRNAs was identified in six aortic tissues from AAD patients compared with six control subjectsInvestigation of the functional role of miRNAs in TAA and AAD remains difficult in human aortic tissues in situ. Gender-associated differences need to be considered. The use of primary human aSMC and experimental models in mice allowed a better characterization of the mechanisms of action of miRNAs, as detailed hereafter.(C1039G/ + )mice and its inhibition with LNA-modified antisense oligonucleotides prevented aneurysm development, reduced apoptosis in aortic wall, and increased ECM components synthesis.MiR-29 family contains three members, miR-29a/-29b/-29c, all involved in fibrosis.C1039G/+mice but not from wild-type mice.C1039G/+mice, the authors proposed that up-regulation of miR-29b expression by TGF-β1 may involve the NFκB pathway.In vitro, miR-29b induced aSMC migration by inhibiting DNA methyltransferase 3 that negatively regulates matrix metalloproteinases 2 and 9 expression.These studies support a therapeutic potential for miR-29b for the prevention of TAA expansion, acting through modulation of aSMC synthetic phenotypic switch and ECM composition. The cross-talk between miR-29b and TGF-β is consistent with a role for miR-29b in the pathogenesis of conditions due to TGF-β pathway alteration, not limited to TAA and AAD. The observation that TGF-β impacts miR-29b expression suggests that TAA and AAD pathogenesis is controlled by complex epigenetic mechanisms that remain to be fully deciphered.C1039G/+mice, losartan administration reduced aneurysm formation and miR-29b expression.Intriguingly, in aorta from young FbnMicroRNA-143 and miR-145 are expressed from the same bicistronic precursor.In mice, miR-143 and miR-145 were enriched in aorta compared with other organs.In vitro, miR-145 has been proposed as a phenotypic marker of vascular smooth muscle cells (vSMC).Despite some heterogeneity observed in human studies, miR-143/-145 family is apparently deeply involved in the regulation of aSMC biology and ECM composition. Since aSMC phenotypic switch and ECM alterations are critical in aortic media dysfunction, miR-143 and miR-145 may play important roles in TAA and AAD pathogenesis that could be interesting for therapeutic purposes.Due to the importance of aSMC phenotypic switch in the pathogenesis of TAA and AAD, the following chapter focuses on miRNAs  and theMicroRNAs promoting aSMC differentiation. MiR-1 expression was induced by myocardin in aSMC,MicroRNAs promoting aSMC dedifferentiation. MiR-221 and miR-222 are two close homologous miRNAs sharing the same seed sequence. They are highly expressed in aSMC, inducing not only their proliferation but also their migration, and inhibiting their apoptosis, presumably through modulation of their target genes p27 (Kip1), p57 (Kip2), and c-Kit.MicroRNAs promoting phenotype switch. The role of miR-21 in aSMC phenotype switch is less clear since it appears to be able to promote aSMC differentiationOverall, the differential expression of multiple miRNAs in aneurysmal aortic tissues, as well as their functional involvement in aSMC biology and ECM composition, strongly supports their role in the pathogenesis of TAA and AAD. The challenge resides in the differentiation between simple bystanders and miRNAs that actively contribute to the disease, the latter category containing the most interesting therapeutic candidates. MiR-29b appears to be such a good candidate.To our knowledge, the biomarker potential of circulating miRNAs has not been studied in the context of TAA. The potential use of circulating miRNAs as markers of AAA and aneurysm growth has only recently been reported.Contrarily to miRNAs, a few studies have addressed the association between lncRNAs and TAA. The lncRNA AK056155 was up-regulated in serum and aortic tissue samples from patients with Loeys–Dietz syndrome compared with controls.Mice lacking the lncRNA GAS5 display a thickening of the aortic media.In vitro experiments revealed that, like miRNAs, lncRNAs affect aSMC proliferation and migration . The lnThe role of lncRNAs in TAA is only emerging. The observation that some lncRNAs are present in blood supports a potential use as biomarkers. A deeper characterization of the functional involvement and biomarker potential of lncRNAs in TAA and AAD is warranted.Genetic alterations critically account for the development of TAA and the progression toward dissection. However, other etiologies of TAA or AAD such as degenerative TAA or AAD occurring on medium-sized aorta are unrelated to any known DNA polymorphism. In this context, epigenetic modifications, which do not change DNA sequence, may also trigger, or at least contribute to, TAA and AAD. Several lines of evidence strongly support a causative role of epigenetic modifications and noncoding RNAs in the pathogenesis of TAA and AAD.r) observed in TAA is associated with a decreased media thickness (h), leading to a dramatic increase in wall tension (T) at a constant blood pressure (P), according to the Laplace's law (T = P.r/2h). The nuclear envelope is mechanically coupled to mechanotransduction through aSMC adhesion to matrix, coupling of integrins to actin, intermediate filaments (tensegritySince epigenetic modifications may occur under the influence of environmental changes such as hemodynamic modifications, it would be interesting to address whether chromatin remodeling mediated by histone modifications is related to the hemodynamic modifications featured in TAA. Indeed, the dilation (increased radius,The impact of noncoding RNAs on TAA and AAD pathogenesis is far from being well characterized. Increasing evidences show that both short and lncRNAs interact to regulate chromatin remodeling, aSMC phenotype, and ECM composition, sometimes through mediation of cell–cell communication.Since miRNAs and lncRNAs are present in the blood, it is conceivable that they may emanate, actively or passively, from the diseased vascular wall and inform about its stability or propensity to rupture. The possibility that noncoding RNAs could be used as biomarkers of TAA and dissection is attractive and remains to be tested.Additional research is warranted to reach a better knowledge of the epigenetic mechanisms gearing TAA development and aneurysm rupture. Several studies reported here showed the involvement of noncoding RNAs in TAA and dissection. However, their role in the pathogenesis of these diseases is only partly understood. Increasing interest is observed toward DNA methylation and histone modification. New insights into their role in TAA and dissection pathogenesis are expected to emerge in the upcoming years. Since epigenetic changes can be modulated, this may lead to novel tools for better diagnosis and treatment of these severe conditions.


The purpose of this study was to identify factors predicting behavioral and psychological symptoms of dementia(BPSD) in older adults with Dementia. This is a cross-sectional study, recruiting 157 participants from neurology general hospital as study subjects. Data collection was performed from June 2018 to May 2019. BPSD were classified using a modified version of the Cohen‐Mansfield agitation inventory(CMAI), which are physically non-aggressive behaviors(PNAB), physically aggressive behaviors(PGAB), verbally non-aggressive behaviors(VNAB), verbally aggressive behaviors(VAGB). The Cornell scale for depression in dementia(CSDD), korea activity of daily living scale(K-ADL), korean mini-mental state examination(K-MMSE), activity and sleep time through using actigraphy for 2weeks, salivary melatonin and cortisol level at 4 times a day done after waking up, after breakfast, before and after dinner, and medication were measured as influencing factors. The generalized linear mixed model analyses indicated that VNAB and VAGB were associated with severe depression, low melatonin level at the after waking up , and high melatonin level before dinner. On the PNAB and PGAB, severe depression and high activity showed great influences. These findings suggest that developing of intervention of BPSD must be started with detecting depression, ADL. Considering the factors of each type of symptom, tailoring an individual approach is recommended. In addition, this study identified that the activity through actigraphy and salivary melatonin measurement are useful tools to examine BPSD. It can be helpful in the objective evaluation of BPSD.


Drosophila as providing an unparalleled opportunity to combine dietary manipulation with models of human metabolic disease and cancer. Thus, combining diet and cancer models in Drosophila can rapidly and significantly advance our understanding of the conserved molecular mechanisms that link diet and diet-related metabolic disorders to poor cancer patient prognosis.Many of the links between diet and cancer are controversial and over simplified. To date, human epidemiological studies consistently reveal that patients who suffer diet-related obesity and/or type II diabetes have an increased risk of cancer, suffer more aggressive cancers, and respond poorly to current therapies. However, the underlying molecular mechanisms that increase cancer risk and decrease the response to cancer therapies in these patients remain largely unknown. Here, we review studies in mouse cancer models in which either dietary or genetic manipulation has been used to model obesity and/or type II diabetes. These studies demonstrate an emerging role for the conserved insulin and insulin-like growth factor signaling pathways as links between diet and cancer progression. However, these models are time consuming to develop and expensive to maintain. As the world faces an epidemic of obesity and type II diabetes we argue that the development of novel animal models is urgently required. We make the case for  The effect of nutrition on cancer has received much attention, spawning an industry of books and websites purporting a vast array of controversial dietary information and supplements for cancer patients. A plethora of ‘recommended’ diets exist, ranging from those that claim to prevent cancer to those suggesting food types and supplements that enhance the effectiveness of therapy and prevent relapse ,2. OftenThe earliest suggestion of a link between diet and cancer arose from laboratory mouse studies conducted in the 1940s . Later sThere have since been numerous epidemiological studies assessing the effect of various specific dietary factors on the incidence of cancer . Taken tNevertheless, in the last decade one clear theme has emerged: obesity and obesity-related diseases, in particular type II diabetes, are associated with increased risk of colon, uterine, liver, pancreatic, breast , and prostate cancers amongst others . In 2012Drosophila melanogaster genome in order to alter gene function, either throughout the whole animal or within specific cell types, allows the molecular mechanisms of gene function to be characterized at a level of resolution not possible in current vertebrate models. Additionally, the low cost and rapid generation of multiple genetic and cell biology tools in Drosophila results in efficient in vivo investigation of the metabolic and growth pathways that underlie tumorigenesis. The conservation of fundamental biology and physiology between flies and humans, has allowed Drosophila researchers to deliver pivotal breakthroughs in our understanding of tumor biology [Drosophila melanogaster, may increase our capacity to make a timely and relevant contribution to understanding the relationships between cancer, diet, and dietary-induced metabolic disorders.Despite the potential impact of diet on cancer, and its associated economic burden, the mechanistic links between diet, dietary related metabolic disorders, and cancer remain poorly understood. Such mechanistic studies cannot be conducted using in vitro tissue-engineered tumor models because these fail to mimic the interactions between tumor cells, organ systems and diet that occur in whole animals. Thus, a significant barrier to current cancer research is the need to use whole animal models. Transgenic mouse models that manipulate nutrient status and cancer are in their infancy  and comp biology ,27,28. TTo gain knowledge of the links between diet and cancer, animal models in which the aetiology of human cancers can be recapitulated whilst altering dietary inputs and monitoring metabolic and tumor biology are required. The laboratory mouse has a long history of providing fundamental insights into the mechanisms of cancer initiation, progression and metastasis . Two majThe current gold standard of xenograft models is the ‘human-in-mouse’ xenografts  in which advanced tumor fragments or metastases are implanted into an immunodeficient mouse subcutaneously, or, with greater difficulty, orthotopically . The major advantage of this system is that initially tumors retain human-specific microenvironment features, which are more reflective of the histology, natural architecture and genetic heterogeneity seen in primary tumors . Unfortuin situ where inflammatory processes can interact with the developing tumor. Ideal GEMMs of human cancers feature tumors that carry the corresponding human mutation and that arise in a small subset of cells within normal tissue, where the steps in cancer progression  closely resemble human pathologies [Genetically-engineered mouse models circumvent several issues that arise using patient-derived xenograft models because tumors arise hologies . Howeverhologies . An advahologies . HoweverDespite the substantial human epidemiological evidence that reveals a clear association between cancer and diet, this area of human health is noticeably understudied. Yet, even with such limited study, increased tumor growth, and more aggressive metastatic disease has been observed in animals reared on high calorie diets for a variety of mouse cancer models, including pancreatic , prostatMin) has led to the development of several excellent mouse models to study the early stages of human colon cancer and the effects of dietary manipulation on cancer progression. Min mice have a mutation that results in truncation of the adenomatous polyposis coli (Apc) protein, similar to that found in patients with familial adenomatous polyposis and in 80% of patients with sporadic colon cancers [Min/+Apc mice fed an ‘obesogenic American-diet’ showed increased polyp size, but no change in total polyp number [Min/+Apc mice have also been used to demonstrate that introducing a source of high dietary fiber (rice bran) decreased tumor burden [The most well-studied models combining cancer and diet manipulation are colorectal cancer models using xenografts or GEMMs ,40. For  cancers . Heterozp number . This rep number , providir burden . These sIt is clear that mouse models can provide invaluable insights into the relationship between diet and cancer progression. However one problem with these studies is the lack of detail provided to describe the diets used in different laboratories. Currently, there are a number of high fat or high sugar diets commercially available such as the ‘Western’, ‘Sweet Stuff’, or ‘Strictly Vegan’ diets from the Jackson Laboratories , or the high and low fat diets from BioServ Inc.  and Research Diets . Furthermore, several research groups use ‘in house’ designer diets such as the ‘American-diet’ . CarefulAlthough the mechanisms that underlie increased susceptibility to cancer in individuals with type II diabetes or obesity remain largely unknown, several concepts are beginning to emerge through the study of mouse models that combine cancer with a model of obesity/ type II diabetes ) ,47. For A-FABP, ALDH1 expression levels and tumor growth were reduced in transplanted breast tumor cells. Furthermore, -/-A-FABP mutant mice bearing orthotopically injected breast cancer cells fed a high fat diet show reduced tumor burden compared to -/-A-FABP mice on a low fat diet. Finally, in a GEMM of post-menopausal breast cancer (MMTV-Transforming Growth Factor) loss of A-FABP also showed reduced ALDH1 positive cells and decreased tumor growth [Recent studies have begun to shed light on the potential molecular mechanisms that may link obesity with aggressive cancer progression and poor outcomes in breast cancer patients . Emerginr growth . These rr growth . Thus, ADrosophila melanogaster is an excellent model for addressing fundamental aspects of the interaction between diet and cancer. They are inexpensive to maintain, small, and have a simple diet, and therefore a vast number can be kept in the laboratory. Their short lifecycle (9–10 days) and exemption from many regulatory considerations ensures rapid and timely progress can be made, particularly with respect to cancer studies that can take several months or years to complete in mice. In addition, Drosophila researchers have developed powerful genetic techniques and dietary models that allow for the rapid identification and characterization of genes involved in multiple human diseases, which include cancer and metabolic disorders [Drosophila offers unparalleled opportunities for genetically manipulating multiple genes and pathways in a tissue-specific manner (needed for cancer studies) within a whole animal model (needed for dietary studies). Second, the similarity of cellular processes such as growth and metabolism and the emerging evidence of functional conservation of genes between Drosophila and mammals mean that studies in flies can directly contribute to the understanding of human disease [Despite progress using mice to model diet, obesity-type II diabetes, and cancer interactions, significant issues remain with these models. Predominantly, mouse models that manipulate nutrient status are in their infancy, and complex mouse models targeting multiple cancer pathways are technically challenging and expensive to maintain. We argue that the vinegar fly isorders ,58. In p disease ,60. Drosophila to investigate human cancers and metabolic disorders, there are also several limitations that should be considered. First, a clear physiological difference between flies and mammals is the optimal body temperature. Flies are ectotherms and can be successfully reared between 18 and 27 °C whilst humans are endothermic and closely maintain a body temperature of approximately 37 °C. As ectoderms do not have to maintain body temperature it has been proposed that these animals expend more energy on growth and reproduction than endoderms [Drosophila [Drosophila also lack a closed circulatory system rendering the modeling of tumor induced angiogenesis and associated changes in the tumor microenvironment difficult. Finally, Drosophila models of drug discovery may be limited due to differences in drug metabolism pathways between flies and humans [Notwithstanding the advantages of using ndoderms . Hence, osophila . Drosophd humans ,62.Drosophila [Although humans and flies differ greatly in terms of their gross morphology, many of the digestive and endocrine systems that control nutrient uptake, storage, and metabolism in humans are also present in osophila ,64. Foodosophila . Key orgosophila ,67  that is equivalent to glucagon, whilst iPCs located within the fly brain are analogous to pancreatic cells  to coordinate metabolic homeostasis . One tarDrosophila [Drosophila models can be used to investigate various aspects of vertebrate metabolic function including diet, nutrient uptake and the regulation of energy storage or expenditure.The biochemical pathways responsible for regulating appropriate circulating sugar levels and energy storage in mammals are also highly conserved in osophila ,72. Consosophila , cellulaosophila , mitochoosophila , the horosophila . In mammosophila . Thus, DDrosophila larvae reared on a high sugar diet  or a high fat diet  exhibit hallmarks of human type II diabetes [iLP2) mRNA, and protein expression, and elevated circulating iLP2 levels. Importantly, larvae raised on a HSD or HFD supplemented with insulin show reduced responses to insulin and diminished activation of the InR pathway target Protein Kinase B (Akt), indicating Drosophila larvae raised on a HSD or HFD display insulin resistance, which is a defining feature of type II diabetes. Finally, insulin resistant diabetes is often present in obese patients, and larvae raised on a HSD and HFD accumulate stored fat in the form of triacylglycerides and free fatty acids [Drosophila is an excellent model to explore the genetic and molecular mechanisms that link dietary fats and sugars to human diseases [Importantly, diabetes ,79. Thesty acids ,79,80. Tdiseases .Drosophila [Drosophila studies are shedding light on the relationship between cancer and diet. In flies, it is possible to generate groups of tumor cells in patches of epithelial tissue surrounded by a normal cellular and signaling environment, thus mimicking the sporadic nature of human tumorigenesis [PTEN, a negative regulator of PI3K signaling) was shown to be increased under conditions of nutrient restriction  [PTEN mutant cells increase tumor growth via cell multiplication under conditions of amino acid starvation and energy stress. The increased proliferation of PTEN mutant cells facing severe nutrient restriction comes at the expense of neighboring wildtype cells. PTEN mutant tissue is metabolically more active and outcompetes the surrounding wildtype tissue for nutrients via the InR pathway. In addition, under conditions of nutrient restriction PTEN mutant cells also sustain their energy needs by systemically reducing the growth of organs throughout the entire animal [Since the identification of the first tumor suppressor mutations in osophila , flies hosophila ,28, therigenesis . Furtherigenesis . In one  by 25%) . Thus, Pe animal . The mecDrosophila sugar-enhanced cancer model (SECM) [Drosophila to model human tumor development, and the ability of a high sugar diet (HSD) to induce type II diabetes in growing larvae. Drosophila SECM tumor cells contain the Q156STOPcsk mutation that results in activation of the potent oncogene Src. In addition, these tumor cells also overexpress an activated isoform of the small GTPase Ras (V12Ras). This mimics the elevated Src and Ras signaling observed in a number of human cancers including breast, colorectal and pancreatic cancers [V12Ras/Src co-activated cells develop small, slow-growing tumors within the epithelial tissue. By contrast, in larvae raised on a HSD the V12Ras/Src co-activated cells develop large primary tumors, as well as metastatic secondary tumors due to increased Wingless  signaling [The laboratory of Ross Cagan was the first to establish a l (SECM) . This mo cancers . In larvignaling . Thus, rDrosophila SECM is the ease with which secondary mutations that alter tumor growth can be then introduced into these flies, enabling mechanistic studies of links between diet and cancer signaling pathways to be performed. For example, the Cagan lab showed that members of the conserved AMP activated kinase family, Drosophila Salt Inducible Kinases 2 and 3 (SIK2 and 3), were required for tumor growth specifically on a high sugar diet [A major advantage of the gar diet . These sgar diet ,91,92,93Drosophila show that SIK3 improves animal survival when exposed to increased dietary sugar levels [Studies in r levels . SIK3 dir levels . The PPPr levels . In contr levels ,96. We dr levels . Drosophila and mouse cancer models that also recapitulate diet, obesity and/or type II diabetes interactions will be crucial to identify growth signaling and/or metabolic pathways that may provide new avenues for cancer diagnostics and therapies.In ovarian cancers, 85% of tumor samples show elevated hSIK3 levels . In addiDrosophila and mice has traditionally focused on caloric restriction, where energy intake is reduced by decreasing the amount of food (diluting food) to around 10–50% or increased by over-nutrition. However, diet is complex and there is debate as to whether the effects of caloric restriction and over-nutrition are due to altered calories or altered intake of specific macronutrient proportions; primarily protein, fat and sugar [Drosophila development and growth [Much of the cancer-related dietary research in both nd sugar ,103. Thend sugar ,107,108.d growth ,111,112.z-axis . The coordinates of the point representing an individual are defined by its nutritional history. Any quantitative phenotype of interest, e.g., animal growth, tumor growth or number of metastases, can be plotted as a surface in the z-axis . Statistthe diet . By combDrosophila genetics is the UAS-GAL4 system, which is used to manipulate gene expression in a tissue and timing-specific manner. This system is based upon the yeast GAL4 transcriptional activator and its DNA target, Upstream Activating Sequence (UAS) [GAL4 driver lines with defined patterns of expression have been generated by many researchers. Furthermore, a large number of UAS lines that express Drosophila or human genes, or induce RNA interference (RNAi) and therefore reduce gene expression, have been described. These tools underpin multiple experimental approaches that have enabled Drosophila researchers to stay at the forefront of biological research [LexA/LexAop transgene systems have also been developed in Drosophila [Drosophila equivalent of the liver (fat body) or gut. This builds capacity to undertake systemic approaches in Drosophila cancer models, allowing researchers to shed light on the interplay between tumor growth, metastasis, and organ systems throughout the body. By taking advantage of these dual transgene expression systems in Drosophila it will now be possible to generate tumors within the fly and manipulate the expression of circulating hormones, and metabolic genes and pathways in specific organs, thereby modeling the complex interactions between cancer progression and metabolic processes in distant organs. Drosophila researchers are thus poised to generate new insights into communication between tumor cells and peripheral organs, which is critical for distributing energy stores and maintaining cancer growth. The workhorse of ce (UAS) . A wide research . Alternaosophila ,117,118.Drosophila provides an excellent model system, as the conservation of fundamental physiology, and metabolic and growth pathways between flies and humans means that studies in Drosophila can deliver pivotal breakthroughs in our understanding of nutrition and cancer growth pathways that are applicable to human health.We are currently faced with an epidemic of patients with obesity and type II diabetes. Crucially, these individuals face an increased risk of cancer and decreased survival rates. Resolving the effects of dietary macronutrients on cancer risk and progression remains a fundamental challenge, with profound implications for human health. Animal models that allow precise manipulation of dietary input , combined with the capacity to assess nutrient levels, genetically manipulate metabolic and/or growth-signaling networks in specific tissues and/or tumor cells are needed.


Room for improvement exists regarding recommendations for screening, staging, therapy selection, and frequency of surveillance of gastrointestinal cancers. Screening is costly and invasive, improved staging demands increased sensitivity and specificity to better guide therapy selection. Surveillance requires increased sensitivity for earlier detection and precise management of recurrences. Peripherally collected blood-based liquid biopsies enrich and analyze circulating tumor cells and/or somatic genomic material, including circulating tumor DNA along with various subclasses of RNA. Such assays have the potential to impact clinical practice at multiple stages of management in gastrointestinal cancers. This review summarizes current basic and clinical evidence for the utilization of liquid biopsy in cancers of the esophagus, pancreas, stomach, colon, and rectum. Technical aspects of various liquid biopsy methodologies and targets are reviewed and evidence supporting current commercially available assays is examined. Finally, current clinical applicability, potential future uses, and pitfalls of applying liquid biopsy to the screening, staging and therapeutic management of these diseases are discussed. Effective management of gastrointestinal tract cancers depends on early detection, accurate staging, therapy selection, and timely surveillance. All components of management hinge on the presence or absence of systemic disease, which was historically detectable only after the development of bulky metastases. Such metastases are currently detected with a combination of serum markers, radiography, and invasive procedures such as endoscopy.Solid tumors have long been known to shed malignant cells and genomic material into the circulation antecedent to the development of macrometastases . In receUntil recently, liquid biopsy of peripheral blood lacked the sensitivity and specificity required for clinical utility in cancer management . AdvanceCTC-based assays exclude cells grossly unlike CTCs based on size and surface markers. This enrichment step is required due to the relatively miniscule amount of circulating tumor material present in a standard 2–4 mL collection of peripheral blood. Following enrichment, conclusive identification of CTCs requires either cytologic or genetic analysis. Cytologic detection with surface markers allows for the further assessment of cell viability and, in some cases, improved prognostication compared to genetic detection . NotablyGenetic material is released into circulation through cell death or active processes such as exosome release. ctDNA-based liquid biopsy excludes benign cell-free DNA to identify tumor-derived ctDNA, probing circulating oligonucleotides that harbor tumor-specific mutations. ctDNA most commonly occurs as fragments on the order of 100–200 base pairs, which are shorter on average than fragments of benign circulating DNA. Assays performed in plasma offer superior sensitivity to serum-based assays due to increased release of benign cell-free DNA encountered during the processing of serum [Intratumoral heterogeneity is a primary reason for treatment failure owing to drug resistance and therapeutic failure . TherefoCTCs and tumor microemboli (clusters of tumor-derived cells) have been detected in the peripheral blood of patients with both subtypes (adenocarcinoma and squamous cell carcinoma) of EC ,14. CTCsCurrent ctDNA-based liquid biopsy generally targets circulating oligonucleotides containing point mutations or alterations in methylation. Digital PCR-based detection methods offer high sensitivity for ctDNA containing mutations already identified in the primary tumor; however, assays that instead employ next-generation sequencing cast a broader net and are more likely to detect mutations discordant from the primary tumor genotype.Esophageal cancer screening is invasive and limited to high-risk patients with persistent symptoms of acid reflux . UnfortuLiquid biopsy may represent a sensitive and minimally invasive alternative to endoscopic screening. A small study of CTC-based liquid biopsy differentiated squamous cell EC patients from healthy controls with 86.3% sensitivity and 90.3% specificity . ctDNA iCurrent staging is radiographic, involving PET-CT and endoscopic ultrasound. Nearly 50% of patients without evidence of disseminated disease at the time of curative-intent therapy develop distant metastases within five years, mainly due to hematogenous dissemination . Thus, aClearly, room for improvement exists in staging, and liquid biopsy has the potential to improve on current modalities. Prospective studies of 74–106 patients with EC demonstrated that the presence of CTCs prior to treatment correlated positively with the degree of local invasion, presence of nodal disease , and prectDNA-based liquid biopsy has the potential to improve prognostication in both squamous and adenocarcinoma. While the total level of cell-free DNA (cfDNA) is not correlated with overall survival , ctDNA aThe above studies suggest that liquid biopsy can further stratify patients with clinically apparent localized EC based on endoscopy and radiography. Ideally, clinical trials to refine indications for neoadjuvant and adjuvant therapy could be enriched with “high-risk” individuals rather than all-comers with liquid biopsy.Management is stage-dependent, but in a curative setting involves a multimodality approach with chemo/radiation therapy and surgical resection of the primary tumor. One study demonstrates that persistent CTC positivity following radiation therapy may be a negative prognostic factor . AbsolutMET amplifications detected in ctDNA can serve as markers of resistance to trastuzumab and novel HER-kinase inhibitors in adenocarcinoma [Multiple studies revealed that somatic mutations detected in ctDNA were discordant with mutations in the tumor tissue in 13–39% of patients . The Perarcinoma .Response to therapy, determination of resistance to targeted treatment, and frequency of surveillance may be facilitated by liquid biopsy as well. Further prospective studies in which patients receive personalized therapy based on specific mutations identified in ctDNA are necessary to confirm the utility of ctDNA in clinical practice. Whether patients should undergo serial liquid biopsies throughout the course of treatment to provide clinically relevant trends are another area that needs to be explored in a prospective fashion.7 white blood cells collected [Both CTCs and ctDNA have been detected in the peripheral blood of patients with gastric cancer. One study showed that >2 CTCs per mL of peripheral blood had 85.3% sensitivity and 90.3% specificity for distinguishing 116 patients with known gastric cancer from 31 healthy controls. The false positive rate in this study was 6.5% (2/31) . The conollected . CTCs haollected . In addiollected .No average-risk screening for GC is currently employed in the United States. In Japan, where the incidence is higher, nationwide endoscopic screening has diminished GC-related mortality over the past decade . Based oNo current tests offer the sensitivity and specificity that would be required to supplant endoscopic screening in high incidence locations such as Japan, or to justify broad screening in the United States. The CancerSEEK platform based on ctDNA demonstrated nearly 70% sensitivity to all-comers, however, sensitivity for stage I disease was extremely low with currently available assays . More stDetection of CTCs and ctDNA is a promising adjunct to current established staging modalities in gastric cancer. A study of 44 patients demonstrated that the CTC count at diagnosis is predictive of advanced stage, peritoneal dissemination, presence of metastases, and poor survival . FurtherHER2 amplification status between primary tumors and CTCs in patients with metastatic GC [A well-established treatment for advanced esophagogastric cancer is anti-HER2 antibodies in tumors with HER2 amplification or overexpression. However, resistance to these antibodies poses a barrier to effective therapy. A prospective study of 88 patients demonstrated discordance in tatic GC . As targAlthough not broadly employed yet in clinical environments, it appears that the most actionable current use for liquid biopsy is likely in monitoring response to therapy and acquired resistance and guiding targeted therapy with the ability to detect sensitivity and resistance patterns. A small study demonstrated discordant genotypes acquired from tissue biopsy between primary and metastatic lesions, leading to clinically significant treatment reassignment . That saAnother GC therapy currently under clinical investigation is anti-EGFR agents in patients with EGFR amplification. While determining the utility of these drugs, it was discovered that ctDNA could be utilized to reveal pre-existing or acquired genomic events that ultimately lead to anti-EGFR resistance .A final role for liquid biopsy in predicting overall response to therapy in GC is one demonstrated throughout most surgically resectable gastrointestinal cancers. This is the correlation between elevations of CTC levels in post-treatment patients with overall poor prognosis. This, combined with other prognostic data, provides an even more accurate prognosis .Promising research has demonstrated a potential role for liquid biopsy in the diagnosis and management of PDAC. In patients with PDAC, both CTCs and ctDNA have been identified in peripheral blood. Enrichment is challenging in PDAC due to small numbers of CTC in peripheral blood and pilot studies demonstrated that 7.5 mL whole blood is required for adequate sensitivity; in addition, cell size-based isolation (ISET) was more sensitive than the Cellsearch system . FollowiA study of 34 patients demonstrated that PDAC patients released ctDNA into the circulation at a rate that was orders of magnitude higher than healthy controls and genomic and epigenetic mutations were detectable in 5 mL of whole blood . The optPancreatic cancer is currently the 4th leading cause of cancer death in the United States, primarily due to late presentation of illness, demonstrating an unmet need for effective PDAC screening. A recent study suggested that PDAC takes at least 15 years to develop from initial mutations to metastatic cancer , highligCTCs are present in patients across all stages of PDAC, more so in patients with advanced disease. However, they are limited for screening use at present due to high rates of false positives in non-malignant disease states, such as chronic pancreatitis (up to 63%) . One stuKRAS, CDKN2A, SMAD4, and TP53. A pilot study of exome sequencing demonstrated KRAS as the most commonly mutated gene in ctDNA (90% to 95% of cases) [KRAS mutations in up to 20% of chronic pancreatitis patients [KRAS mutations with CA 19-9 levels differentiated PDAC (47 patients) from chronic pancreatitis (31 patients) with a sensitivity of up to 98% and a specificity of 77% [Four principal mutations have been identified in ctDNA as most commonly found in PDAC: f cases) . As withpatients . Howevery of 77% . While pCurrent staging for PC is done radiographically, with patients’ resectability status determined by presence of local invasion, involvement of major peripancreatic vessels, and the presence of remote disease. The most evident potential benefit of liquid biopsy lies in prognostic indicators with the potential to guide management and determine resectability. A meta-analysis of 16 studies regarding the use of CTCs and disseminated tumor cells (CTCs in bone marrow) as prognostic indicators demonstrated that detection of CTCs in peripheral blood and CTCs in bone marrow is associated with both progression-free and overall survival .KRAS or ERBB2 mutations in peripheral blood significantly impacted overall and progression-free survival. Furthermore, specific mutations, hypermethylation, and a high absolute quantity of ctDNA were associated with inferior survival [KRAS) in the peripheral blood or portal blood. One study demonstrated that a KRAS mutation in the portal venous circulation was associated with increased risk of developing metastases [KRAS mutations were present in 31%. KRAS mutation-positive patients demonstrated an overall survival of 13.6 months compared to 27.6 months in those with wild-type cancers [KRAS mutations were a worse prognostic indicator when patients with all stages of disease were analyzed [As for the role of ctDNA, a meta-analysis of 18 studies including 1243 patients determined that the presence of ctDNA with survival . Potentitastases . Further cancers . Interesanalyzed .Even after curative intent resection, five-year survival in PDAC is low. Contributing to this is recurrence, with a majority of resected patients experiencing recurrence within two years of surgery. Frequent physical exams, CT imaging, and CA 19-9 levels are currently utilized, however, no clear guidelines for surveillance are currently in place.KRAS or TP53 mutations) metastatic PDAC who underwent chemotherapy, detectable mutations significantly decreased during treatment and the resulting levels correlated with progression-free survival [Liquid biopsy may predict overall survival and response to therapy in PDAC. An example of this is CTC positivity, which dropped from 80.5% of patients to 29.3% after a cycle of fluorouracil-based chemotherapy in 41 patients with advanced PDAC . Berger survival . The samFurther research is warranted to demonstrate the utility of liquid biopsy in monitoring PDAC recurrence. One study demonstrated that specific CTC phenotypes  are predictive of recurrence . Higher The role of liquid biopsy in colorectal cancer (CRC) is evolving rapidly. CTCs, ctDNA, and exosomal DNA have been isolated from the blood of patients with CRC. Location of the blood-draw impacts the sensitivity of liquid biopsy in CRC, as blood from the portal system is enriched with tumor material compared to peripheral venous blood . A smallDetection limits reach as low as 3 CTCs per 2 mL whole blood . CTCs inTotal cell-free DNA and ctDNA were elevated in the peripheral blood of CRC patients compared to both healthy controls and patients with inflammatory bowel disease . The halCRC is unique among gastrointestinal malignancies in that a relatively mature screening program exists for average-risk patients. Colonoscopic screening is effective yet invasive, associated with poor compliance from patients, and is labor- and resource-intensive. Less invasive means of screening, such as fecal immunochemical tests and DNA tests , have been developed and deployed clinically in recent years. It is important to note that any positive noninvasive screen for colon cancer is followed up with a diagnostic colonoscopy. If sufficient sensitivity and specificity can be achieved, liquid biopsy would save disease-free patients the cost, inconvenience, and morbidity of screening colonoscopy. No liquid biopsy-based assays were recommended for CRC screening in the 2018 Guideline Updates from the American Cancer Society, however, as trial data are collected and as technology improves, such tests may have a greater role in CRC screening in the future.Multianalyte detection systems (CancerSEEK) analyzed specific mutations in ctDNA along with other traditional peptide markers for CRC and other cancers in preclinical experiments. Data collected in this study were run through a supervised machine-learning algorithm that was able to identify CRC positivity in 100% of patients with known disease . CTC-basIndications for adjuvant chemotherapy are well-defined in patients with stage III disease, however, patients with stage II disease require further stratification to determine whether systemic therapy is indicated. The ability to better identify those patients that would benefit from systemic therapy is critical to improving survival and limiting toxicity and should be investigated further. A meta-analysis of 11 studies demonstrated that ctDNA positivity at the time of diagnosis is associated with inferior overall survival in colon cancer, however, the strength of this association varies with timing and technique . ExosomaCurrent staging and surveillance systems poorly identify patients who may benefit from invasive regional, and even distant, curative therapies, such as cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC). Indications for such therapy are currently based on imaging findings in patients who are already symptomatic. A pilot study of 14 patients suggested that postoperative CTC-positive patients with suspicion of peritoneal disease would benefit from CRS/HIPEC in terms of overall survival . The indKRAS that impact the efficacy of targeted therapy plays a major role in the staging of CRC. Digital PCR specific for KRAS and BRAF mutations was able to detect variant alleles in the ctDNA of 66% of patients with locally advanced rectal cancer that harbored these mutations in surgically resected specimens. On the other hand, 15% of preoperative patients had KRAS mutations detected via ctDNA that were not detected during endoscopic biopsy [RAS mutations when biopsy of the primary tumor is unfeasible [The presence of mutations such as c biopsy . Some exfeasible .KRAS, TP53, or APC mutation was sufficient evidence of residual disease after curative-intent therapy [Liquid biopsy has demonstrated promise in detecting minimal residual disease or recurrence during the surveillance of colon cancer. Standard post-treatment surveillance requires the use of non-specific serum markers (CEA), endoscopy, and an abdominopelvic CT scan. A well-designed study of 230 patients determined that ctDNA with either a  therapy . Support therapy .RAS wild-type. However, most patients go on to develop resistance to these agents. Thus, treatment decisions in metastatic CRC hinge on the presence of treatment resistance. One well-designed study demonstrated that ctDNA-based liquid biopsy was extremely sensitive (>85%) for primary resistance to HER2 inhibitors [Most patients with CRC are treated with 5-fluorouracil-based chemotherapy. Targeted agents such as panitumumab and cetuximab are routinely added to these regimens if the tumor is hibitors . Presumahibitors . In factnsitive >% for prihibitors . Notablyhibitors . Taken tAdvances in regional therapy for rectal cancer in particular have made distant recurrences the primary cause of mortality. A point of clinical equipoise is whether rectal cancer patients benefit from adjuvant systemic therapy after neoadjuvant chemoradiation and surgical resection. A well-designed cohort study with 159 patients demonstrated that the presence of ctDNA after resection is indicative of residual disease and a high probability of recurrence. In these patients, treatment with adjuvant systemic chemotherapy significantly improved OS . AnotherAs a screening tool, liquid biopsy has the potential to save the cost and morbidity of invasive endoscopic screening procedures in diseases such as CRC and improve early detection of diseases such as pancreatic adenocarcinoma. It appears ctDNA-based liquid biopsy is generally more sensitive for the early detection of gastrointestinal cancers than CTC-based assays. While surgical techniques and outcomes continue to improve, procedures such as esophagectomy, gastrectomy, and pancreaticoduodenectomy still come with a high risk of morbidity and mortality. The ability to accurately stage patients with micrometastatic disease could save patients, who would not benefit from upfront resection, the morbidity inherent to such operations. Presence of CTCs may be an indicator of unresectability in certain gastrointestinal tumors, whereas the presence of ctDNA in determining resectability is less clear. Both CTC- and ctDNA-based liquid biopsy offer broad advantages over currently employed surveillance of gastrointestinal tumors using radiography, endoscopy, and tumor markers. Regarding the selection of systemic therapy regimens, ctDNA is generally more predictive of response to therapy and more useful in identifying actionable mutations for targeted therapy. Improvements in surveillance would benefit thousands of patients each year with locally advanced disease who may be candidates for systemic or extensive regional therapy, including cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (HIPEC) .A catalog of actively enrolling clinical trials investigating liquid biopsy in gastrointestinal cancers is shown in Liquid biopsy technology has improved alongside basic understanding of cancer biology in recent years. The convergence of ideas from these two fields has led to the development of clinical-grade assays with the potential to impact all stages in the management of gastrointestinal cancers. Although still evolving, if applied correctly, existing and future technologies will hopefully improve access to care, management, and outcomes across all gastrointestinal cancers.


Preoperative localization of functionally eloquent cortex  is common clinical practice in order to avoid or reduce postoperative morbidity. This review aims at providing a general overview of magnetoencephalography (MEG) and high-density electroencephalography (hdEEG) based methods and their clinical role as compared to common alternatives for functional cortical mapping of (1) verbal language function, (2) sensorimotor cortex, (3) memory, (4) visual, and (5) auditory cortex. We highlight strengths, weaknesses and limitations of these functional cortical mapping modalities based on findings in the recent literature. We also compare their performance relative to other non-invasive functional cortical mapping methods, such as functional Magnetic Resonance Imaging (fMRI), Transcranial Magnetic Stimulation (TMS), and to invasive methods like the intracarotid Amobarbital Test (WADA-Test) or intracranial investigations. Functional cortical mapping (FCM) aims at localizing eloquent functional cortex using a range of invasive and non-invasive methods . Its maiThe advent of non-invasive FCM methods has substantially influenced the care of neurosurgical candidates. Indeed, the availability of FCM results based on non-invasive approaches before surgery allows for a better estimation of the risk-benefit ratio of the planned neurosurgical procedure with better patients' counsel, optimized neurosurgical strategies, as well as tailored resection extents. Overlap between functionally eloquent cortex as identified by non-invasive FCM and lesional or epileptogenic zones may even argue against surgery or for alternative therapeutic strategies.This paper reviews currently available FCM methods with a special emphasis on electromagnetic source imaging. Strengths, weaknesses and limitations of electromagnetic source imaging in relation to other modalities commonly used for mapping of verbal language, sensorimotor, memory, visual, and auditory functions are also presented.Identification of anatomical landmarks in structural cerebral imaging represents an easy and fast approach to localize functionally eloquent cortex. However, inter-rater reliability is significantly lower than with FCM results, even within the same subject and across successive analyses over several days . FurtherAmong the various non-invasive FCM techniques that can be used in humans, fMRI is by far the most commonly used . Active Magnetoencephalography (MEG) and EEG are non-invasive methods that record magnetic and electric fields, respectively. These are generated by excitatory or inhibitory postsynaptic potentials at the apical dendrites of neocortical pyramidal cells. Determination of the anatomical location of the sources generating the measured signal is known as magnetic and electric source imaging . This is achieved by combining MEG or EEG data with structural MRI. For the purpose of clarity, we will henceforth only use MEG and EEG to refer to MSI- or ESI-based FCM.In contrast to EEG, MEG is sensitive mainly to tangential neocortical source components. Consequently, it detects activity mainly from the sulcal walls, while small areas at the crown or in the sulcal depth barely contribute to detectable signals . SensitiTMS is a non-invasive form of neurostimulation. Magnetic fields applied focally are used to induce or inhibit electric activity of targeted neurons via electromagnetic induction. Neuronavigation using the patient's individual structural MRI  allows for better targeted stimulation (nTMS). In the context of FCM, it is used primarily for verbal language and primary motor (M1) cortex localization –17 in spWada and Rasmussen described in 1960 the intracarotid injection of amobarbital for the lateralization of cerebral speech dominance . Sodium Direct current stimulation (DCS) is used in awake craniotomy for language-, motor- and memory mapping. For a review, see . IntracrPresurgical and potentially intraoperative investigation of verbal language function is mandatory to avoid postoperative language deficits if surgery involves resection near (presumed) language eloquent cortex.Language related cortex is extensive and bilateral, although one hemisphere is usually dominant. Two broad processing streams can be distinguished: a ventral stream for speech comprehension  and a dorsal stream for sensory-motor integration . For comprehensive reviews, see, e.g., , 24. In Assessment of hemispheric dominance and intra-hemispheric cortical representation of speech processing and production are the main objectives of language FCM . IAT hasA frequently used approach for assessment of hemispheric receptive verbal language dominance was proposed by Papanicolaou et al. . It evalOther strategies apply beamforming  and evalFurthermore, distributed source models, such as e.g., MR-FOCUSS have been successfully used for verbal language lateralization, as demonstrated by Bowyer et al. . Their aActive participation for the assessment of verbal language dominance using MEG may not be needed, as passive (listening) paradigms have also been described , 39.IAT does not allow for an arterial injection of amobarbital that is selective enough to discriminate the sublobar structures involved in verbal language processing. Therefore, in clinical practice, intrahemispheric verbal language localization is primarily evaluated using fMRI or MEG preoperatively, or using direct current stimulation (DCS) intraoperatively.Various protocols for the assessment of areas involved in verbal language comprehension , 40–42 aMost studies suggest a high inter-subject variability of language-related activations, as well as high degrees of cortical plasticity in patients with brain disorders , 46, 47.Furthermore, nTMS, as well as nTMS-based DTI fiber tracking, have been applied successfully  but are,fMRI possibly provides better prediction of postoperative verbal language and memory deficits compared to IAT , 51, andMultiple approaches have been evaluated and validated for MEG verbal language lateralization. Stimulation paradigms usually test for language comprehension or language production. Agreement with IAT is about 86–92% for word recognition tasks , 32, andData on the comparison of localization accuracy of MEG mapping and DCS are sparse . Hirata Clinical MEG societies regard MEG as a validated tool for presurgical evaluation of patients in respect to the assessment of verbal language-dominant hemisphere and consider it as a potential replacement for IAT in most patients . HoweverIn patients with lesions or epileptogenic zones located at the central region, mapping of sensorimotor cortex is utilized to locate primary somatosensory (S1) and motor (M1) areas . While sClinical MEG/EEG mapping of S1 cortex typically utilizes either electrical or mechanical stimulation. The former follows principles of, e.g., median or tibial nerve electrical stimulation as frequently applied in neurological and neurosurgical practice , 73–79. The spectrum of FCM paradigms and analytical techniques to locate M1 cortex mainly rely on motor evoked fields/potentials (MEF/MEP) and on the suppression of rolandic  beta rhythm(s). Additional methods can also be used such as cortico-muscular (CMC) or cortico-kinematic (CKC) coherence.Motor evoked field/potential or readiness paradigms , 83–85 uMotor activity is accompanied by event-related desynchronization (ERD) or suppression of oscillatory activity in the alpha and beta frequency bands . This muCoherence approaches  evaluate the functional coupling of neuronal activity with either muscular activity [as measured by electromyography (EMG)] or movement kinematics. Statistical approaches localize cortical areas of significant coupling with these external reference signals. Stimulation paradigms typically utilize isometric contractions for CMC and recording of (active or passive) movement kinematics with, e.g., accelerometers for CKC , 91–93. Of note, MEG and fMRI activations may also be used to support identification of the corticospinal tract . Aoyama MEG-based localization of the SM1 cortex shows high agreement with DCS and fMRI—for a review, see, e.g., . ValidatIn comparison to fMRI, MEG shows comparable, and in some patients even superior results , 97–102.In patients with tumors or vascular lesions in the vicinity of the SM1 cortex, localization of SM1 cortex using fMRI may be difficult due to lesion-induced alterations of regional cerebral blood flow and susceptibility artifacts , 104. InMEG also presents an additional key strength over fMRI, which is the ability to investigate in one single MEG session different neurophysiological processes  that can be altered or affected differently by brain lesions or patients' clinical status. Thus, MEG provides the unique opportunity to acquire several MEG “functional localizers” of the SM1 cortex in a reasonable time for the patients . “FunctiNavigated transcranial magnetic stimulation (nTMS) is increasingly applied for presurgical mapping of SM1 cortex . A numbeStudies evaluating EEG for FCM of SM1 cortex are limited. Klamer et al.  comparedDue to the overlap in declarative memory function and lesions associated with mesial temporal lobe epilepsy (MTLE), memory impairment is common in this group of patients. Verbal memory decline can be observed in 30–85% of patients who undergo left temporal resection, whereas non-verbal memory deterioration after right (- or left) temporal resection affects 30–50% , 114–116IAT is considered the gold standard for the assessment of declarative memory function. Impaired memory performance is usually found in about 20–30 % of cases injected ipsilateral to the seizure onset zone and in 60–80% after contralateral injection –119. BasIn addition to the methodological drawbacks of IAT, there is no guarantee that amobarbital can be sufficiently delivered to the targeted hippocampal formation , 125.fMRI, MEG, PET, and TMS, as well as several combined/integrated methods are non-invasive alternatives evaluated for declarative memory functional mapping .In this context, fMRI is by far the most popular and widely used solution. For a comprehensive review, see e.g., .To the best of our knowledge, MEG (or EEG) are not in  clinical use for declarative memory FCM. The most important obstacle might be the detection of hippocampal activation due to the limited sensitivity of MEG to deep sources—a problem that has been addressed, leading to the implementation of research protocols including the evaluation of deep structures via MEG –130. CliEven though it is considered the gold standard, the role of IAT in the prediction of postoperative declarative memory outcome remains controversial. Rathore et al.  reportedfMRI results show good concordance with IAT, which is also the basis for most validation studies. Throughout the literature, there is a great heterogeneity regarding the experimental paradigms used, but verbal memory tasks show the most consistent and clinically useful results .To the best of our knowledge no validated method of declarative memory functional mapping relying on MEG or EEG exists.Damage to the visual cortex or optic radiations may result in partial or complete anopia, whereas congenital defects or lesions might result in functional reorganization , 134. PrEarly components of visual evoked activity in EEG and MEG  localize to V1 cortex. Paradigms apply pattern reversal stimuli, such as checkerboards, presented to a hemifield or a single quadrant –137. SouEEG-based visual evoked potentials are used for intraoperative neurophysiological monitoring, which evaluates response latencies and amplitudes rather than localization . StudiesDCS, e.g., during invasive Video-EEG monitoring for presurgical evaluation of refractory focal epilepsy, provides high accuracy and selectivity for different aspects of the visual system . A compafMRI has shown the ability to accurately localize V1 cortex and differentiate correlates of different stimulus aspects, such as visual field eccentricity and angular position by sophisticated paradigm design . LocalizAuditory mapping is clinically applied in patients with lesions or structural alterations within or near Heschl's gyrus . LesionaAuditory stimulation in EEG/MEG recordings typically use monaural presentation of brief sine or click tones with white noise masking of the contralateral ear . AverageUni- and bilateral auditory stimulation results in bilateral activation of auditory cortices, reflected by bilateral dipolar generators. In EEG, this leads to merging potentials with a maximum negativity over the vertex and positivity over the basal temporal lobes. Due to the different sensitivity and rotated orientation of magnetic fields, the two components are easier to separate in MEG. Furthermore, generators of auditory activity have a predominantly tangential orientation, which improves SNR in MEG .Early studies have shown the accuracy of MEG , 157 andThe evaluation of functional connectivity has gained considerable interest in the last years, partially due to methodological and technical advances. Going beyond the absolute activation and focusing on functional integration patterns between brain areas has opened new opportunities for FCM. These new approaches enable identification of functional networks, rather than task-based individual activated areas. The main hypothesis assumes that this provides a more complete and realistic view of human brain function. While most studies currently focus on physiological activity in healthy subjects, there are promising findings with potential for novel clinical application . For exaEvaluation of functional connectivity patterns in the resting brain  potentially also provides relevant information for localization of functionally eloquent cortices in fMRI  and MEG Machine learning algorithms are transforming multiple medical fields. Clinical applications are already in use in various domains—predominantly in image processing, classification and segmentation . Roland In the future, machine learning might find further application in FCM, e.g., based on methods and findings in the field of brain-computer interfaces . HoweverFinally, studies demonstrated that FCM based on the investigation of event related enhancement of high gamma activity as recorded by invasive recordings or electrocorticography during awake craniotomy might be of high interest for the mapping of verbal language and motor eloquent cortex –173. StiFor any validation of FCM methods, the choice of the reference standard used to validate the methods is crucial. DCS either intraoperatively or during invasive monitoring with subdural or depth electrodes is considered as one gold standard. However, data on current flow from the electrodes through the cortex is sparse in addition to how such currents interact with the neuronal architecture beyond single cell responses. In addition, different methods might be sensitive to different aspects of neural activity. Lascano et al.  describeIn addition, the predictive value of DCS itself may be limited, as illustrated by Ilmberger et al . The autIn a prospective clinical study by Hermann et al. , resectiTate et al.  identifiEspecially in patients with atypical hemispheric dominance, all non-invasive methods show limitations. Bauer et al.  publisheIn light of these limitations, non-invasive localization methods that seek to go beyond mere lateralization can provide only a conservative estimate of essential functional cortex. If such areas are spared by the neurosurgical procedure, functional deficit can thus probably be avoided in most cases. However, on the other hand, a portion of these regions could potentially be resected without any functional deterioration, e.g., to achieve gross total resection in tumor surgery or to completely remove the epileptogenic zone. In the context of epilepsy surgery, this “conservative” perspective on functionally eloquent cortex may be warranted, especially as quality of life is the central goal. However, tumor surgery requires more aggressive strategies, which would benefit from increased specificity. Focusing on patients undergoing tumor surgery might therefore help to better determine if this conservative approach is indeed mandatory in all situations.The current challenge is to identify markers, which provide a more accurate and robust estimation of this essential, necessary cortex.In general, studies on the direct comparison of different FCM methods in the same patient population are sparse. Most publications focus on the diagnostic accuracy of a single method vs. a mostly invasive reference standard, i.e., DCS or IAT, etc. When postsurgical outcomes are considered, follow-ups are frequently short. Only limited data is available relating presurgical FCM to long-term outcomes and functional reorganization. Results on clinical M/EEG FCM of the auditory or visual system are generally sparse, potentially due to the limited clinical relevance. A main issue for validation of not only M/EEG-based FCM, but also the prognostic value of fMRI, TMS, and even DCS is the very limited availability of prospective, randomized clinical studies.While EEG and MEG register similar activity with differences in sensitivity, EEG is rarely applied for FCM in a clinical context. This is in stark contrast to the wide utilization of EEG in clinical routine for diagnosis and in neurocognitive studies , althougBoth MEG and EEG are sparsely used for FCM in tumor patients in comparison to, e.g., fMRI, while most clinical MEG centers perform functional mapping in the context of epilepsy surgery , 179. ReIn conclusion, electromagnetic source imaging provides additional information for functional mapping with reasonable spatial resolution, exquisite temporal resolution and direct information about neural activity. Due to their non-invasive nature, these methods can be applied early in the presurgical workup and can be utilized to optimize the application of invasive means, such as DCS. Further evaluation is needed to investigate their respective clinical added-value.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


During the last decade, cancer research has significantly evolved into precision medicine and targeted therapies are now part of the many standard treatment regimens in various cancers. As recent cancer genome projects highlighted the importance of epigenetic alterations in cancer development, this area has become a growing focus of interest; thus several drug classes are now being studied that target various components of epigenetic network. One of these epigenetic targets is the bromodomain and extraterminal motif (BET) proteins, which play key roles in gene transcription and cell cycle regulation . The BETIn vitro experiments demonstrated high activity of these agents against USC cell lines with high c-Myc expression by causing a dose-dependent decrease in the phosphorylated levels of c-Myc and a dose-dependent increase in caspase activation. In vivo results were also similar, in which both GS-5829 and GS-626510 exhibited a significantly slower rate of tumor growth, compared with vehicle control as well as mice undergoing treatment with JQ1 (p < 0.05). Importantly, the drug activity was not affected by the presence or absence of gain of function mutations in the PIK3CA gene, which was previously shown to be related with resistance to trastuzumab treatment in USC [ClinicalTrials.gov identifier: NCT02392611, NCT02607228, NCT02983604), while GS-626510, due to its shorter half-life, has so far been assessed only in preclinical studies.The incidence and mortality rates of endometrial cancer (EC) are globally increasing, and, if current trends continue, the incidence of EC will double by 2030 in the United States . HoweverThe relationship between cMyc regulation and BET inhibition is complex due to the fact that BET proteins have many roles beyond cancer and the effects of Myc suppression are not uniform across tumor types. However, our results are promising and deserve further investigation. Myc gain-of-function may represent a novel target in USC and treatment with BET inhibitors used alone or in combination with other targeted agents might be a potential option in patients with advanced/recurrent USC unresponsive to chemotherapy.


Visuospatial neglect constitutes a supramodal cognitive deficit characterized by reduction or loss of spatial awareness for the contralesional space. It occurs in over 40% of right- and 20% of left-brain-lesioned stroke patients with lesions located mostly in parietal, frontal and subcortical brain areas. Visuospatial neglect is a multifaceted syndrome - symptoms can be divided into sensory, motor and representational neglect - and therefore requires an individually adapted diagnostic and therapeutic approach. Several models try to explain the origins of visuospatial neglect, of which the “interhemispheric rivalry model” is strongly supported by animal and human research. This model proposes that allocation of spatial attention is balanced by transcallosal inhibition and both hemispheres compete to direct attention to the contralateral hemi-space. Accordingly, a brain lesion causes an interhemispheric imbalance, which may be re-installed by activation of lesioned, or deactivation of unlesioned (over-activated) brain areas through noninvasive brain stimulation. Research in larger patient samples is needed to confirm whether noninvasive brain stimulation can improve long-term outcomes and whether these also affect activities of daily living and discharge destination. Stroke is one of the leading reasons for long-term disabilities in adults and is responsible for large and increasing health-costs in the United States, projecting to more than double from $36.7 billion in 2015 to $94.3 billion in 2035 motor, and representational  neglect. It can furthermore affect different ranges of space  and frames of references . Some authors combine said classifications and describe for example, extrapersonal neglect in terms of visual or auditory  deficits and personal neglect in terms of somatosensory deficits  stimuli , peripersonal (reaching space), and extrapersonal (far space) neglect. Those different radii around the subject can be effected selectively, though this is rather rare. A recent study found prevalence rates for single-domain neglect ranging from 11%  to 22% . Similarly, patients often omit or misread words or letters on the contralesional side of pages or words (often referred to as “neglect dyslexia”).extrapersonal neglect often overlook people and objects in the left hemi-space. They may bump into obstacles or doorframes. Serious difficulties can occur when they move outdoors, for example, crossing a street can become dangerous, as oncoming cars are not perceived on time. Patients therefore may not be able to move independently at all outside their home.Patients with Cortical representations of space are not limited to viewer-centered reference frames, for which reason visuospatial neglect symptoms typically occur with respect to spatial coding systems that are more complex than a simple “Cartesian” coordinate system dividing space along two or three axes around the viewer. In visuospatial neglect, at least two different coding systems can be impaired.egocentric or viewer-centered neglect, coding in relation to egocentric coordinates, with the own body as the center of the reference frame, is impaired. This can be described as an inability to orient to stimuli that are situated in the contralesional hemi-space with respect to the midsagittal plane of the body.In allocentric neglect, patients fail to allocate attention to the contralesional half of an object, regardless of object location. This deficit can be revealed when copying figures.In Evidence to date disagrees on whether egocentric and allocentric neglect are dissociated , which complicates diagnosis. Moreover, symptoms underlying other deficits can lead to an impression of visuospatial neglect and severity of single subsymptoms can vary greatly requiring careful diagnostic procedures.Visuospatial neglect is often associated with large lesions and commonly occurs concomitantly with other deficits The term homonymous hemianopia refers to visual field loss on one side of the vertical midline due to unilateral post-chiasmatic brain lesions. Visuospatial neglect and homonymous hemianopia co-occur often and symptoms can appear similar. In homonymous hemianopia, patients typically suffer from hemianopic alexia and problems in visual exploration, while spatial representations of both hemi-spaces are intact. Differentiation can be based on affected modalities , visual behaviour, covert attention, and performance in cancellation and drawing tests , or when two actions have to be performed with both hands simultaneously (motor extinction)”  seems to be a promising tool to further elucidate functional relations of anatomically relevant regions in visuospatial neglect , lack appropriate measures to detect visuospatial neglect reliably, and finally, medical staff are often not appropriately trained to assess visuospatial neglect. Systematic screening is crucial for planning rehabilitative efforts and can significantly improve rehabilitation outcome in visuospatial neglect patients  and transcranial direct current stimulation (tDCS), which are safe and can be applied in a wide range of patients with neurological and psychiatric disorders . Indeed, visuospatial neglect-like difficulties in symmetry judgement . Plow et al.  observedStudies in healthy subjects support the hypothesis of a rivalry of spatial attention between the two hemispheres and emphasize the role of the right parietal cortex in spatial processing. However, as these results have been obtained in young subjects, their translational value for the use of noninvasive brain stimulation in rehabilitation of visuospatial neglect is limited. Age affects the lateralization of task-related activity in healthy subjects Cabeza,  and furtNoninvasive brain stimulation is not only applied as a research tool to improve the pathophysiological understanding of visuospatial neglect, but also as a treatment device. Based on the interhemispheric rivalry model, several stimulation approaches can be deduced: excitation of the lesioned hemisphere, inhibition of the non-lesioned – but hyperactive – hemisphere, or a combination of both. Due to safety concerns regarding the potentially increased risk of seizures, most TMS-studies focussed on inhibiting the non-lesioned hemisphere. The majority of studies used TMS-protocols, but in recent years a few studies using tDCS-protocols were published. To evaluate the efficiacy of noninvasive brain stimulation in post-stroke visuospatial neglect, we conducted a systematic review adhering to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses , study designs and inclusion criteria  were applied. To our knowledge, no studies to date explored novel electrical stimulation methods such as transcranial random noise stimulation (tRNS) or transcranial alternating current stimulation (tACS) in patients with visuospatial neglect. Furthermore, more recently developed deeper-reaching coils like H-coils or double-cone-coils have not been applied in patients with visuospatial neglect.We searched MEDLINE, EMBASE and CENTRAL . Medical-Subject-Headings (MeSH)-Terms and direct entries were used. We additionally screened reference lists of relevant articles. The search strategy was as follows: ) AND ).We included peer-reviewed cohort studies and case reports of stroke patients with visuospatial neglect written in English language, in which noninvasive brain stimulation was used as the primary intervention to treat visuospatial neglect. Due to the heterogenity of visuospatial neglect and diagnostic difficulties , no restrictions regarding diagnostic criteria or outcome measures were applied.The database search resulted in 6277 hits. Four more publications were added afterwards. In total, 25 studies  published between September 1999 and April 2019 were included in the systematic review Fig. .Fig. 3Fln = 227) of these studies applied rTMS over the contralesional PPC. Five more recent studies applied cTBS over the contralesional PPC and only two studies applied stimulation over contralesional prefrontal areas, one using iTBS , as well as symptom heterogenitiy, pose a general problem in studies investigating visuospatial neglect. Additionally, the conducted studies are heterogeneous with regard to study designs, inclusion criteria, time since stroke, and outcome measures, hampering comparability and making conclusions difficult.When these results are considered together, the therapeutic value of noninvasive brain stimulation in visuospatial neglect seems very promising. Except for one study that investigated exclusively chronic stroke patients , compare different stimulation protocols , explore possible benefits of MRI-guided targeting, and further elucidate underlying processes of visuospatial neglect, for example by using single-subject designs.Visuospatial neglect is a frequent and disabling impairment after stroke. In light of its multifaceted nature with heterogenous manifestations, diagnosis and treatment pose challenges in clinical routine.Normal performance in bedside testing or a single paper-pencil test does not rule out visuospatial neglect. If visuospatial neglect is suspected, it is crucial that different tests covering a wide range of symptoms are administered. Neuropsychological diagnosis should therefore be based on patient observation and comprehensive testing. Often disregarded, concomitant cognitive impairment in non-spatial attention and executive functioning must be addressed carefully in neuropsychological assessment and therapy.A wide range of therapeutic approaches has been applied in visuospatial neglect, including cognitive, behavioural, and pharmacological therapy. However, there is no consensus as to which rehabilitation methods and combinations are most effective. In recent years, technological advances have led to a better understanding of underlying neurophysiological, and anatomical factors of visuospatial neglect. The role of large-scale networks in visuospatial attention and their disruption in visuospatial neglect is now widely recognized and, in the course of this, noninvasive brain stimulation has emerged as a promising approach to study visuospatial neglect pathophysiology and advance treatment. Noninvasive brain stimulation as a rehabilitation tool is based on the interhemispheric rivalry model, in which excitatory and inhibiting protocols are employed to reestablish interhemispheric balance. As interindividual variability in treatment response is an important issue in visuospatial neglect, future stimulation protocols should be based on individually designed, network-based protocols. For example, each patient should be stimulated over different stimulation targets in order to identify the protocol that is most likely to (a) lead to behavioural changes and (b) modify brain activity and connectivity toward “normalization”.In this review we have discussed the broad and complex nature of visuospatial neglect, presented currently used diagnostic and therapeutic options with a special emphasis on noninvasive brain stimulation, and offered recommendations for future studies. We hope that this review therefore contributes valuable information to advance the investigation of visuospatial neglect.