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Kuan Yan

Bio: Kuan Yan is an academic researcher from Leiden University. The author has contributed to research in topics: Medicine & Tumor microenvironment. The author has an hindex of 12, co-authored 21 publications receiving 387 citations.

Papers
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Journal ArticleDOI
15 May 2014-Oncogene
TL;DR: Evidence is provided for AnxA2 as a mediator of EGFR endocytosis and signaling in breast cancer via regulation of cofilin activation.
Abstract: Enhanced epidermal growth factor receptor (EGFR) activity has been strongly linked to breast cancer progression and mediators of EGFR endocytosis may well be involved. We developed a semi-automated high-content fluorescence microscopy-based EGFR endocytosis screen to identify proteins that mediate EGFR endocytosis in human HBL100 breast cancer cells. Knockdown of 172 individual endocytosis and actin-regulatory genes with small interfering RNAs led to the identification of 14 genes of which the contribution to EGFR endocytosis in breast cancer is until now poorly defined, including DNAJC6, GDI2, FGD6, HAX1, NECAP2 and AnxA2. We show that depletion of the actin and endocytosis regulatory protein annexin A2 (AnxA2) in a panel of four triple negative breast cancer (TNBC) cell lines affected EGFR endocytosis. Depletion of AnxA2 in the aggressive and highly metastatic MDA-MB-231 TNBC cell line resulted in the inhibition of EGFR transport beyond the early endosomes. This inhibition coincided with enhanced epidermal growth factor (EGF)-induced cell migration and downstream signaling via c-Jun N-terminal kinase (JNK) and Akt. Moreover, AnxA2 knockdown increased lung metastasis formation in mice. The effect of AnxA2 knockdown on EGFR endocytosis in MDA-MB-231 was related to dephosphorylation/activation of the actin-severing protein cofilin, as re-expression of an inactive S3E-cofilin mutant, but not an active S3A-cofilin mutant, re-established EGFR endocytosis to control levels. Together, our data provide evidence for AnxA2 as a mediator of EGFR endocytosis and signaling in breast cancer via regulation of cofilin activation.

53 citations

Journal ArticleDOI
TL;DR: Phenotypic profiling using complex 3-D cell cultures steers hit selection towards more relevant in vivo phenotypes, and may shed light on subtle mechanistic variations in drug candidates, enabling data-driven decisions for oncology target validation.
Abstract: Background Monolayer cultures of immortalised cell lines are a popular screening tool for novel anti-cancer therapeutics, but these methods can be a poor surrogate for disease states, and there is a need for drug screening platforms which are more predictive of clinical outcome. In this study, we describe a phenotypic antibody screen using three-dimensional cultures of primary cells, and image-based multi-parametric profiling in PC-3 cells, to identify anti-cancer biologics against new therapeutic targets.

51 citations

Journal ArticleDOI
TL;DR: This review provides an overview of recent advanced imaging strategies together with quantitative image analysis that can be implemented to understand the dynamics of matrix adhesions and its molecular components in relation to tumour cell migration.
Abstract: Cell migration is essential in a number of processes, including wound healing, angiogenesis and cancer metastasis. Especially, invasion of cancer cells in the surrounding tissue is a crucial step that requires increased cell motility. Cell migration is a well-orchestrated process that involves the continuous formation and disassembly of matrix adhesions. Those structural anchor points interact with the extra-cellular matrix and also participate in adhesion-dependent signalling. Although these processes are essential for cancer metastasis, little is known about the molecular mechanisms that regulate adhesion dynamics during tumour cell migration. In this review, we provide an overview of recent advanced imaging strategies together with quantitative image analysis that can be implemented to understand the dynamics of matrix adhesions and its molecular components in relation to tumour cell migration. This dynamic cell imaging together with multiparametric image analysis will help in understanding the molecular mechanisms that define cancer cell migration.

38 citations

Journal ArticleDOI
23 Jun 2017
TL;DR: It is shown that a pathophysiologically relevant 3D cyst culture model of PKD coupled to phenotypic profiling can be used to identify potentially therapeutic compounds and predict and validate molecular targets for PKD.
Abstract: Polycystic kidney disease (PKD) is a prevalent disorder characterized by renal cysts that lead to kidney failure. Various signaling pathways have been targeted to stop disease progression, but most interventions still focus on alleviating PKD-associated symptoms. The mechanistic complexity of the disease, as well as the lack of functional in vitro assays for compound testing, has made drug discovery for PKD challenging. To identify modulators of PKD, Pkd1–/– kidney tubule epithelial cells were applied to a scalable and automated 3D cyst culture model for compound screening, followed by phenotypic profiling to determine compound efficacy. We used this screening platform to screen a library of 273 kinase inhibitors to probe various signaling pathways involved in cyst growth. We show that inhibition of several targets, including aurora kinase, CDK, Chk, IGF-1R, Syk, and mTOR, but, surprisingly, not PI3K, prevented forskolin-induced cyst swelling. Additionally, we show that multiparametric phenotypic classification discriminated potentially undesirable (i.e., cytotoxic) compounds from molecules inducing the desired phenotypic change, greatly facilitating hit selection and validation. Our findings show that a pathophysiologically relevant 3D cyst culture model of PKD coupled to phenotypic profiling can be used to identify potentially therapeutic compounds and predict and validate molecular targets for PKD.

37 citations

Journal ArticleDOI
TL;DR: Photobleaching assays of whole cells revealed an opposite residence time of FAK and paxillin in maturing and disassembling FAs, which depends on the ventral and peripheral cellular position of the FAs.
Abstract: Focal adhesions (FAs) are specialized membrane-associated multi-protein complexes that link the cell to the extracellular matrix and enable cell proliferation, survival and motility. Despite the extensive description of the molecular composition of FAs, the complex regulation of FA dynamics is unclear. We have used photobleaching assays of whole cells to determine the protein dynamics in every single focal adhesion. We identified that the focal adhesion proteins FAK and paxillin exist in two different states: a diffuse cytoplasmic pool and a transiently immobile FA-bound fraction with variable residence times. Interestingly, the average residence time of both proteins increased with focal adhesion size. Moreover, increasing integrin clustering by modulating surface collagen density increased residence time of FAK but not paxillin. Finally, this approach was applied to measure FAK and paxillin dynamics using nocodazole treatment followed by washout. This revealed an opposite residence time of FAK and paxillin in maturing and disassembling FAs, which depends on the ventral and peripheral cellular position of the FAs.

35 citations


Cited by
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Book ChapterDOI
E.R. Davies1
01 Jan 1990
TL;DR: This chapter introduces the subject of statistical pattern recognition (SPR) by considering how features are defined and emphasizes that the nearest neighbor algorithm achieves error rates comparable with those of an ideal Bayes’ classifier.
Abstract: This chapter introduces the subject of statistical pattern recognition (SPR). It starts by considering how features are defined and emphasizes that the nearest neighbor algorithm achieves error rates comparable with those of an ideal Bayes’ classifier. The concepts of an optimal number of features, representativeness of the training data, and the need to avoid overfitting to the training data are stressed. The chapter shows that methods such as the support vector machine and artificial neural networks are subject to these same training limitations, although each has its advantages. For neural networks, the multilayer perceptron architecture and back-propagation algorithm are described. The chapter distinguishes between supervised and unsupervised learning, demonstrating the advantages of the latter and showing how methods such as clustering and principal components analysis fit into the SPR framework. The chapter also defines the receiver operating characteristic, which allows an optimum balance between false positives and false negatives to be achieved.

1,189 citations

Journal ArticleDOI
Ye Fang1, Richard M. Eglen1
18 May 2017
TL;DR: This review of leading 3D cell culture technologies and their impact on drug discovery, including spheroids, organoids, scaffolds, hydrogels, organs-on-chips, and 3D bioprinting, discusses the implementation of these technologies in compound identification, screening, and development.
Abstract: The past decades have witnessed significant efforts toward the development of three-dimensional (3D) cell cultures as systems that better mimic in vivo physiology. Today, 3D cell cultures are emerging, not only as a new tool in early drug discovery but also as potential therapeutics to treat disease. In this review, we assess leading 3D cell culture technologies and their impact on drug discovery, including spheroids, organoids, scaffolds, hydrogels, organs-on-chips, and 3D bioprinting. We also discuss the implementation of these technologies in compound identification, screening, and development, ranging from disease modeling to assessment of efficacy and safety profiles.

509 citations

Journal ArticleDOI
TL;DR: In this article, the authors propose a set of principles to facilitate the definition and development of disease-relevant assays, and discuss new opportunities for exploiting the latest advances in cell-based assay technologies in drug discovery.
Abstract: The common and persistent failures to translate promising preclinical drug candidates into clinical success highlight the limited effectiveness of disease models currently used in drug discovery. An apparent reluctance to explore and adopt alternative cell- and tissue-based model systems, coupled with a detachment from clinical practice during assay validation, contributes to ineffective translational research. To help address these issues and stimulate debate, here we propose a set of principles to facilitate the definition and development of disease-relevant assays, and we discuss new opportunities for exploiting the latest advances in cell-based assay technologies in drug discovery, including induced pluripotent stem cells, three-dimensional (3D) co-culture and organ-on-a-chip systems, complemented by advances in single-cell imaging and gene editing technologies. Funding to support precompetitive, multidisciplinary collaborations to develop novel preclinical models and cell-based screening technologies could have a key role in improving their clinical relevance, and ultimately increase clinical success rates.

384 citations

Journal ArticleDOI
TL;DR: Drug combinations of two or more compounds with different mechanisms of action are an alternative approach to increase the success rate of drug repositioning.

256 citations

Journal ArticleDOI
TL;DR: Insight into the cAMP dynamics executed by the Epac signalosome will help to optimize the pharmacological treatment of devastating diseases, such as diabetes, cognitive impairment, renal and heart failure, (pulmonary) hypertension, asthma, and chronic obstructive pulmonary disease.
Abstract: Since the discovery nearly 60 years ago, cAMP is envisioned as one of the most universal and versatile second messengers. The tremendous feature of cAMP to tightly control highly diverse physiologic processes, including calcium homeostasis, metabolism, secretion, muscle contraction, cell fate, and gene transcription, is reflected by the award of five Nobel prizes. The discovery of Epac (exchange protein directly activated by cAMP) has ignited a new surge of cAMP-related research and has depicted novel cAMP properties independent of protein kinase A and cyclic nucleotide-gated channels. The multidomain architecture of Epac determines its activity state and allows cell-type specific protein-protein and protein-lipid interactions that control fine-tuning of pivotal biologic responses through the "old" second messenger cAMP. Compartmentalization of cAMP in space and time, maintained by A-kinase anchoring proteins, phosphodiesterases, and β-arrestins, contributes to the Epac signalosome of small GTPases, phospholipases, mitogen- and lipid-activated kinases, and transcription factors. These novel cAMP sensors seem to implement certain unexpected signaling properties of cAMP and thereby to permit delicate adaptations of biologic responses. Agonists and antagonists selective for Epac are developed and will support further studies on the biologic net outcome of the activation of Epac. This will increase our current knowledge on the pathophysiology of devastating diseases, such as diabetes, cognitive impairment, renal and heart failure, (pulmonary) hypertension, asthma, and chronic obstructive pulmonary disease. Further insights into the cAMP dynamics executed by the Epac signalosome will help to optimize the pharmacological treatment of these diseases.

246 citations