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Poster Session [clear filter]
Monday, October 21
 

2:30pm BST

Poster Presentation #1- High-Throughput Drug Screening with Advanced Cell-Based Assays
High-Throughput drug screening with advanced cell-based assays

Booij T.H., Hirt C., Keller D., Taube D., Prummer M., Riegler E.M., Stirnimann C., Schwank G.

Lab automation has enabled miniaturization and high-throughput screening approaches in drug research, allowing increased numbers of potential drugs to be tested in in vitro assays. Screens have largely been performed using simple 2D cell culture assays to minimize assay variability. However, it is increasingly being recognized that simple 2D cell culture assays may not adequately represent human disease, causing poor translatability from in vitro results to in vivo experiments. As a result, more advanced assays have gained popularity, such as 3D cell culture assays with cultured cell lines, or the use of patient material as PDX- or organoid models. While the use of such assays has, initially, largely been limited to neoplastic disorders such as cancer and cystic diseases, these models are gaining in popularity for other indications, especially where in vitro experimentation has not been general practice.

A limitation of more advanced cell-based assays with PDX- or organoid model systems has been to automate the liquid-handling procedures to enable high-throughput drug screening in these models. The main factors holding back this development include reagent costs, difficulties with liquid handling of viscous reagents, and the increased assay variation that is inherent to the growth of ‘mini-organs’ in normal tissue culture.
We have established a statistically robust organoid screening platform for pancreatic cancer that integrates fully-automated liquid handling, plate transfer and readouts. This screening platform was applied to screen a drug library that can allow for rapid drug repurposing, possibly providing new lead compounds for this, so far, largely incurable disease.

Speakers
DK

David Keller

Lab Technician, ETH Zurich NEXUS Personalized Health Technologies


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #10- Phenotypic Screening in 3D Culture Including the Extracellular Matrix
Ninety percent of potential drug candidates fail within the clinical trials, mainly because of a lack of efficacy. Biologists and pharmacologists now largely acknowledge the lack of relevant in vitro models in the early drug development stages.

To mimic human tissues, several parameters need to be taken into account: the target cells but also the extra-cellular matrix (ECM). The ECM has diverse and crucial roles as anchorage of the cells, migration barrier and track, signal reservoir, low-affinity co-receptor, signal presenter and is a source of biochemical forces. All these roles of the matricial micro-environment are really important in maintaining organ’s homeostasis and proper intercellular communications. Depending on the organ, ECM composition and physicochemical properties are different.

Cellular assays in 3D culture have shown many advantages to better mimic in vivo situation. Nevertheless, 3D technology used for phenotypic screening is often really simple, as the use of ULA plates. But we can go further by using 3D technologies, which allow reproducing the microenvironment of healthy or pathological organ of interest, as BIOMIMESYS technology. During this talk, we will see how the cellular microenvironment impacts the proliferation and/or differentiation of cells. Few examples will be presented in oncology, CNS and metabolic diseases fields.

Speakers
NM

Nathalie Maubon, Ph.D.

President, HCS Pharma
With 22 years of experience in the pharmaceutical research (public research, pharmaceutical industry and biotechnology company), Nathalie Maubon is a specialist in the development and automation of robust models for high-throughput screening (HCS, HTS, ADMET). She has held several... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #2- Time-Lapse Imaging of Cell Behaviour on 2D Substrates and in 3D Aggregates: Watch it Live with DRAQ9
The importance of cell migration and proliferation in wound healing, tumour metastasis and tissue remodelling is under continued investigation while the ability to track single cells and cellular communities has become a potent tool to measure their biological functional capacity and a systems response to perturbation. Delivering a tracking capability and robust assay readouts is multi-faceted. It requires a cell label that is non-toxic, stable throughout, as cell identifier/locator. We describe a cell permeant, far-red fluorescent, cytoplasmic probe DRAQ9 (BioStatus) that is non-toxic, and present continuously as real-time tracker so fluorescence is undiminished over time e.g. during spheroid aggregation, formation and expansion; in 2D scratch-wounds; or a hybrid assay, as single cells emerge onto 2D substrate from a spheroid. In all cases, DRAQ9 labels cells for unique identification over time. This stable tracker provides a readout ripe for automated cell tracking in aggregates, migration in 2D or indeed through mitosis.

Additionally, in end-point analyses (e.g. high throughput toxicity assays) a fundamental first step to 3D object image analysis is correct boundary identification, where failure results in inaccurate downstream morphometric analysis. DRAQ9 functions as a whole 3D-object “paint”. This allows accurate reproducible boundary identification, using an inexpensive universal platform, simplifying image analysis, for rapid, effective screening.

A459 cells were cultured in wells using a 2 Well insert (IBIDI). After 24 h inserts were removed, cells washed and DMEM (with 10% FBS, 2 μM DRAQ9) added then returned to the incubator for 20 min. Time-series images were acquired every 20 min. using 20x objective lens, for 24 h. Cells were manually tracked and analysed using imageJ. Spheroids were generated in a U-bottomed low-adherence plate from A549 cells cultured in DMEM (with 2 μM DRAQ9, upon spheroid culture) for 7 days. Time-series images were acquired every 30 min. using 10x objective lens, for 4 days. After 7 days spheroids were re-plated to adhere for outgrowth and time-series images acquired every 20 min. using 10x objective lens, for 48 h. A widefield microscope was used throughout.

Manual cell tracking can be completed with confidence due to DRAQ9's cytoplasmic localisation and absence in nuclei. Distinct cell patterning enables automated cell tracking algorithms. Mitosis can be visualised from changes in DRAQ9 signal due to altered morphology. Spheroid painting allows confident morphometric analysis of 3D cultures over time due to non-toxic properties of DRAQ9 for time-series analysis of cultures. Emerging cell behaviour can be visualised from 3D cultures.

DRAQ9 labeling of live cell cultures enables real-time tracking of cell migration and cell division. Visualisation of cell-cell and cell-matrix interactions provides rich information on cell behaviour within a niche. Far-red fluorescence makes it compatible with imaging on sub-optimal imaging substrates e.g. tissue culture plastic. Changes in fluorescence intensity adds information on cellular division, allowing such events to be localised to a cell niche. DRAQ9-brightfield image overlays make manual tracking easier to perform, increasing confidence in cell identification, especially during cell division. Importantly, absence of DRAQ9 in nuclei provides a mask for automated image analysis, reducing image analysis times.

Speakers
RE

Roy Edward, BSc Hons

Sales and Marketing Director, BioStatus Limited
Trained in Biochemistry (St Andrews Univ.). 30+ years of international experience in biotools, in biomagnetic separation and proteomics.Beyond the UK, worked in the USA and Norway with roles in product and marketing management, international distributor management and business development.Written... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #3- Magnetic Electrospun Microfibre Scaffold Assemblies: Examples of there use for Cell-Based Screening Applications
In an effort to create a more physiological three-dimensional (3-D) environment for cell growth while maintaining the logistics of well plate screening for drug discovery purposes, we have develop a 3-D micro-scaffold platform from electrospun material used in conjunction with well plates for higher throughput screening. We have engineered electrospun material to form micro-scaffold islands of cells. Cells grow on, around and into the material, forming a micro-island of adherent cells that are effectively 3-D in solution. The incorporation of iron nano-particles into the fibres during manufacture results in micro-scaffold islands that can be physically manipulated using electro-magnetism. We can culture cells on micro-scaffolds in culture vessels in media within the incubator in addition to moving them within wells using arrays of neodymium magnets, attracting scaffolds to the edges of wells to prevent damage during media replacement or align scaffolds in the centre of wells for imaging purposes. Using sheathed electromagnetic tips we can move micro-scaffolds from well to well with no adverse effects to cells. In addition we have developed a liquid handling device capable of identifying the location of scaffolds in a petri dish then performing a ˜pick and place™ procedure putting them into user defined wells of a plates. Furthermore, we have incorporated fluorescent dyes or quantum dots into the fibres such that fibres can be visualised using fluorescence microscopy, while quantum dots can be used as a barcode to distinguish between two cell populations on different scaffolds within the same well. Cells can be transfected while cultured on micro-scaffolds using either lipofection or electroporation. Recombinant cells can be cryo-preserved on micro-scaffolds. We will show data of exemplar assays using recombinant, primary and differentiated stem cell assays on micro-scaffolds. We have used this approach to examine a number of luminescence and fluorescence readouts in recombinant cell lines and have differentiated iPSC™s to cortical neurones in 3-D on micro-scaffolds to characterise then use imaging as a readout. Our data suggests that this micro-scaffold approach may open up new opportunities for both recombinant cell based screening and differentiated stem cell assays using single and co-cultures in a well plate-based format more amenable to higher throughputs with very little manual intervention.

Speakers
GA

Gary Allenby, Ph.D.

Chief Scientific Officer, Aurelia Bioscience
PhD in reproductive tox, worked in pre-clinical pharma for 25 years in Hit ID, Hits to Lead, Target Biology sections developing cell based assays for compound screening. Founding entrepreneur of Aurelia Bioscience in 2012, delivering tailored assay development for compounds pharmacological... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #4- 3D Multicellular Intestine-on-a-chip Model for Disease Modeling and Drug Discovery
The development of efficient drugs and therapies for treatment of inflammatory bowel disease (IBD) is hampered because of the lack of reliable in vitro and in vivo models. Current models generally fail to recapitulate key aspects of the intestine, resulting in low translatability to the human situation. With the increasing IBD incidence worldwide, there is an unmet need for predictive in vitro models that recapitulate the complex structure of the human intestine. Here, we show the development and characterization of a 3D multicellular perfused intestine-on-a-chip model and its application for investigating and interfering with intestinal inflammation.

Forty independent polarized 3D perfused leak-tight epithelial tubular structures composed of Caco-2 and HT29-MTX-E12 cells were obtained in the microfluidic platform, the OrganoPlate 3-lane. Next, immune competent cells were added to the system and shown to be functional, as demonstrated by the release of cytokines upon activation.

To mimic intestinal inflammation, the model was exposed to tumor necrosis factor (TNF) α and/or interleukin (IL)-1β, whose effects were assessed by measuring the transepithelial electrical resistance (TEER) and secretion of proinflammatory cytokines at the apical and basal sides. The cytokines induced an inflammatory state in the culture, as demonstrated by the impaired barrier integrity and increased IL-8 secretion. We also assessed the applicability of the model in screening anti-inflammatory compounds by using the well-known anti-inflammatory drug TPCA-1. Exposure to the drug prevented an inflammatory state in the model.

Overall, these results suggest that this model provides the versatile modularity of mimicking key features of intestinal inflammation, which positions it at the forefront of high-throughput screening efforts for supporting drug discovery and providing a platform for personalized medicine.

Speakers
LG

Linda Gijzen

Scientist, Mimetas BV
Linda Gijzen, MsC, got her master’s degree in Toxicology and Pathobiology from the Radboud University, the Netherlands, in 2016. Since then she is working as a scientist at the Model Development department at Mimetas, the organ-on-a-chip company, in Leiden. There she is involved... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #5- Patient-Derived Explants (PDEs) as a Powerful Ex Xivo Platform to Predict Clinical Response to Novel Anti-Cancer Drugs
One of the leading goals of the Leicester Cancer Research Centre is to investigate novel and more relevant pre-clinical tumour models that are able to better predict patient outcomes.

Accordingly, we have established a rapidly expanding ‘Cancer Explant Facility’ whereby Patient-Derived Explants (PDEs) from freshly resected tumour specimens, are cultured ex vivo and tested with a wide panel of anti-cancer drugs including currently used therapies as well as novel drugs. This ex vivo model is a powerful platform with multiple advantages over animal models: it is patient-specific, it improves the predictivity of drug response in situ and, by accurately retaining the complexity of the tumour ex vivo, overcomes the differences between the human and animal immune systems that can lead to differential response to targeted immune-oncology therapies in animal models.

In our experimental method, drug responses are assessed using multiplexed immunofluorescence (mIF), which allows the ability to distinguish between drug effects in tumour areas versus the surrounding stroma as well as the potential contribution of the host immune system. So far, this fast and reproducible technique has been applied to different types of cancer (melanoma, lung, renal, breast and CRC liver metastasis) to analyse the effect of anti-cancer agents on cell viability, proliferation and immune cell mobility. Finally, by combining this platform with gene expression, genetic and epigenetic analysis, we also aim to identify novel biomarkers that correlate with, and predict, the response of cancers to anti-cancer drugs as well as their toxicity in preclinical settings, prior to clinical testing.

Speakers
GV

Giuditta Viticchie, PhD

Post Doctoral Research Associate, University of Leicester
2008: Master’s degree in Chemistry at the University of Rome “Tor Vergata”, Italy.2012: PhD in Biochemistry and Molecular Biology at the University of Rome “Tor Vergata”, Italy2012-2013: Post Doctoral Fellow at the MRC Toxicology Unit Leicester, UK, with Italian fellowship... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #6-Real-Time Live-Cell Analysis of 3D Tumour Spheroid Invasion
Cell invasion, a hallmark of malignant cancers, plays a key role in tumour dissemination and metastasis and is responsible for the higher incidence of cancer-related deaths. The ability of tumour cells to form a metastatic tumour primarily involves cell morphological reorganization and degradation of the surrounding extracellular matrix (ECM).Conventional in vitro invasion models, such as the filter based transwell invasion assay (2D) are widely used to assess tumour cell invasion.However, lack of comprehensive cell-ECM interactions may limit their physiological relevance. Here, we provide a simple 3D tumour spheroid invasion model to recapitulate key features of the metastatic phenotype and demonstrate use of live cell imaging to kinetically acquire and quantify 3D single spheroid invasion.



To exemplify the assay platform and assess the metastatic potential of tumour cells in 3D, U87-MG, A172 and HT1080 spheroids were formed for 72 h in Ultra-Low Attachment round bottom 96-well plates and subsequently embedded in Matrigel (4.5 mg/mL). Spheroid invasive properties were measured using novel brightfield image analysis software (IncuCyte® Live-Cell Analysis System) which tracks and quantifies changes in spheroid size (whole spheroid area or invading cell area) over time. To account for variation in size following spheroid formation, whole spheroid area was normalized to spheroid size at t = 0 h and automatically plotted. Cell type specific temporal invasive potential was observed. At 168 h, whole spheroid area of U87-MG spheroids (ratio: 32) increased to approximately 2x and 4x the size of HT1080 (ratio: 18) and A172 (ratio: 8) spheroids respectively. HT1080 spheroids embedded in varying concentrations of Matrigel (1.13 – 4.5 mg/mL) demonstrated that spheroid invasive capacity is Matrigel concentration-dependent. While cell motility was reduced at the highest Matrigel concentration, marked density and elongation of invadopodia was seen. Using available metrics and analysis approaches we could determine the effects of known anti-metastatic compounds on spheroid invasion and spheroid proliferation. Cytochalasin D (300 nM) and PP242 (30 µM) inhibitors caused comparable reduction of U87-MG spheroid size (90% and 80% respectively). However, marked inhibition of invadopodia was only evident with Cytochalasin D. Subtraction of the invading cell area from the whole spheroid area provided a measure of the spheroid body size and revealed the anti-proliferative rather than anti-invasive properties of PP242. A separate but identical study conducted in the absence of Matrigel further supported the anti-proliferative effects of PP242 but not Cytochalasin D.

These data demonstrate the capability to kinetically visualise and quantify 3D single spheroid invasion assays and illustrates the utility of this approach for high throughput real-time anti-metastatic compound profiling. This model can be further developed with the incorporation of stromal cells to address additional aspects of tumour cell invasion.

Speakers
MO

Miniver Oliver

Research Scientist, Sartorius
Research Scientist at Sartorius, working as part of the Applications group for the IncuCyte platform. Expertise indeveloping 3D cellular assays for quantitative live cell analysis.


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #7- High-Content Assay for Morphological Characterization of 3D Neuronal Networks in a Microfluidic Platform
Establishment of physiologically-relevant in vitro models is crucial to further understanding of the mechanisms of neurological diseases as well as targeted drug development. While iPSC-derived neurons show great promise for compound screening and disease modeling, use of three-dimensional (3D) cultures is emerging as a valid approach for neuronal cell based assay development. 3D cultures are recognized as more closely recapitulating aspects of the human tissues including the architecture, cell organization, as well as cell-cell and cell-matrix interactions.

This poster presents the development of a high-throughput 3D neurite outgrowth assay using iPSC-derived neurons developed in the microfluidic OrganoPlate® platform, with the goal of establishing 3D models for neurodegenerative diseases and neurotoxicology screens. 3D cultures were treated with known neurotoxic compounds and the morphology and viability of neurons were assessed. The OrganoPlate® from MIMETAS is a high-throughput platform that combines the most recent advances in 3D cell culture, Phaseguides™ and microfluidics. The OrganoPlate contains 96 tissue chips suitable for long term culture of live cells, is amenable for screening purposes, and is compatible with standard laboratory equipment or automated systems, like the ImageXpress® Micro Confocal High-Content Imaging System.

Speakers
AB

Andy Bashford

Application Scientist, Molecular Devices
Andy works as a Field Applications Scientist for Molecular Devices covering the whole portfolio of imaging solutions.Prior to this he worked as a development scientist for the European Collection of Authenticated Cell Cultures (ECACC) where he was responsible for developing assays... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #8- LaFOS: A Novel 4D Morphometric Phenotypic Profiling Method to Uncover Therapeutic Vulnerabilities in Patient-Derived 3D Tumor Models
3D-spheroid cultures are powerful tools for in vitro models for biology, but size and shape diversity within the culture is largely ignored and also their current end point data outputs (like metabolic measurements) are resource intensive and we cannot afford real-time kinetics at the primary drug screening level. We intent to develop robust Label free Oncology Score (LaFOS) using brightfield image of 3D tumor spheroids in different time intervals. We developed a computational image scoring methodology, that integrates morphometric profiling, segmentation, filtering, and analysis for brightfield images of 3D culture and provide us with cytotoxicity level without any metabolic end point readouts like cell viability assay. LaFOS is accurate and flexible like current existing platforms, and we illustrate its potential by studying 600 plus small molecules that are used in the clinic or in advanced phases of development. We tested this computational method in parallel with cell viability assay (cell titer Glo) in 15 primary patient derived models and 3 commercial cancer cell lines in comparison with traditional 2D monolayer models. We found Pearson correlation of 0.81 for LaFOS methodology and 0.95 for cell viability assay in finding the cellular cytotoxicity. Our results suggest that, there is differential drug response between 2D and 3D model for specific class of compounds. The compound connectivity map showed different targets specificity and network structure between 2D and 3D models. We observed pathway specific enrichment of (EGFR, PI3K and mTOR inhibitors) in 3D models and cell cycle/division related enrichment (HDAC, CDK, Topo and Tubulin) in 2D models. RNA-sequence analysis of 13 cell lines showed 151 pan-differentially expressed genes in 2D vs 3D. Immunofluorescence and western blot analysis showed HIF-1α was high in 3D models and Ki67 was found high in 2D models this data correlates to architecture dependent drug response and gene expression data. Also 3D models showed more activity of pAKT and pERK could be a reason for pathway specific inhibitor activity in 3D models. We did a validation of hits obtained from in vitro screen using in vivo PDX model for ZSTK474 a PI3K inhibitor. Overall we developed a computational platform (LaFOS) that can measure 3D spheroid cell viability in real-time fashion using morphometric signature obtained from label free brightfield phenotypes.

Speakers
GP

Giridharan Periyasamy, PhD

Platform Leader, Genome Institute of Singapore
Dr. Giridharan Periyasamy is a Platform Leader in Centre for High Throughput Phenomics (CHiP-GIS) at the Genome institute of Singapore. He is the domain expert in Assay automation, miniaturization, robotics, and High Content Imaging technology. His research focus is on the Cancer... Read More →


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer

2:30pm BST

Poster Presentation #9- Microfluidic Drug Efficacy Testing of 3D Tumour Models
In recent years, physiologically relevant 3D in vitro models of cancer have been continuously refined in order to better mimic the tumour microenvironment when testing chemotherapies and immunotherapies. An increasing interest in precision treatment of cancer patients has highlighted the need for technologies that are capable of maximising the testing of compounds in 3D tumour models obtained from cancer biopsies. Microfluidics and organ-on-a-chip technologies, via precise control of fluids and cells in solution, offer great opportunities for screening biopsy-derived spheroids, organoids, co-cultures and primary tumour fragments in a sophisticated and cost-effective manner.

We have developed microfluidic platforms for high quality and multiplexed drug screening assays on both spheroid, organoids and tissue fragments. Readouts, such as the spheroid volume, spheroid shape and viability are generated from both brightfield and epifluorescence microscopy. The technology has been validated using a variety of cell lines, primary and biopsy cancerous tissue. As proof of concept, human prostate biopsies were grown as a heterogeneous co-culture and used for the generation and screening of thousands of spheroids, showing sensitivity to docetaxel, but resistance to enzalutamide, despite the presence of intact androgen receptors. A further application of the platform lies in the co-culture of several cell types, in order to reconstruct a more realistic tumour microenvironment. Several breast cancer cell lines and cancer-associated fibroblasts (CAFs) were tested to allow the assessment of the impact of drugs and inhibitors on specific cell populations. Lastly, ovarian tumour biopsy tissue fragments were cultured for precision medicine purposes.

Speakers
TM

Theresa Mulholland, BsC

Senior Scientist, ScreenIn3D Limited
Senior scientist at ScreenIn3D, interested in the development of novel assays for drug development and screening, using complex microfluidic 3D in vitro models


Monday October 21, 2019 2:30pm - 3:00pm BST
Sherry Coutu Seminar Suite Foyer
 
Tuesday, October 22
 

1:30pm BST

Poster Presentation #11-Using 3D Neural Spheroids to Accelerate Drug Discovery with Kinetic and High Content Imaging
Three-dimensional platforms of human induced pluripotent stem cell (hiPSC)-derived neurons enable greater physiological relevance and promise to breathe new life into drug discovery programs. A fundamental requirement of this re-vitalization is high-throughput and informative interrogation methodologies.StemoniX has developed microBrain®3D, ready to use spheroids comprised of human iPSC-derived cortical neurons and astrocytes supplied in ready to use 96- and 384-well plates.Real-time, high-throughput kinetic imaging has demonstrated the value of this preparation in functional screening and disease model settings while advancements in tissue clearing and spheroid processing are enabling in-depth, granular structural examinations. In particular, our data demonstrate a dramatic benefit of tissue clearing for confocal analysis of neuronal spheroids stained with antibodies and Cell Painting dyes as well as enhanced throughput enabled by physical stabilization of spheroids on flat-bottom imaging plates. This presentation will highlight recent uses of microBrain 3D in functional 384-well based screens, patient-derived disease models, phenotypic rescue, and image-based interrogation. Together 3D human neural spheroids with functional and structural experimental methodologies hold the potential to overcome several bottlenecks in CNS-based drug discovery.

Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #12-Feasibility of Manufacturing-scale Bioproduction of Novel Next-generation 3D Organoid Cancer Models in Support of the Human Cancer Models Initiative
To meet the need for improved approaches to study cancer in vitro, there has been a surge in the development of novel research models utilizing advanced culture methods. These methods permit in vitro growth of cancer types previously not possible, and/or models with enhanced in vivo relevancy compared to traditional continuous cell lines. However, availability of these early-stage research models is currently limited and there is a lack of data on the ability to scale up production of these models to support the needs of the global research community. We sought to investigate the protocols, expansion capacity, cryopreservation ability, genetic stability, and feasibility of larger-scale bioproduction of a subset of the models generated by the Human Cancer Models Initiative (HCMI), an international collaborative effort between Cancer Research UK, the foundation Hubrecht Organoid Technology, the National Cancer Institute, and the Wellcome Sanger Institute. The HCMI’s initial goal is the development of 1,000 novel human cancer models, paired with bioinformatics and patient clinical data, particularly from rare or underrepresented cancer types. One advanced culture method being utilized, three-dimensional organoid “microtissue” culture, potentially poses challenges for traditional large-scale bioproduction processes. It requires growth embedded within an undefined extracellular matrix and complex media formulations containing multiple small molecules and recombinant proteins with unknown stability and shelf-life. Additionally, organoid growth media typically includes multiple sources of undefined conditioned media containing critical growth factors. We cultured organoid models derived from human colon, pancreas, esophagus, and mammary tissues developed by laboratories contributing to the HCMI. Multiple unique donors were available for all tissues and both cancer and non-cancer models were available for two tissue types. Most models were maintained in culture continuously for at least 60 days (7-27 population doublings, > 10 passages). Tissue and donor variability was evident in model characteristics, including morphology (assessed by microscopy and immunocytochemistry), growth rate, and genetic stability (measured by short tandem repeats analysis). All models were amenable to scale up beyond multiwell plates, and all models could recover from cryopreservation. While organoid culture represents a significant divergence from typical two-dimensional monolayer culture of continuous cell lines, our results show that these next-generation in vitro models are suitable for larger-scale bioproduction. This is vital to ensure the widespread availability of these models within the research community to facilitate applications like pre-clinical drug discovery and basic cancer research.

Speakers
NP

Nirmal Perera, Bachelor of Medicine, Bachelor of Surgery (M.B.B.S), Medicine. Masters of Research, Genetics and Gene The

ATCC Field Applications Scientist, LGC
Dr. Nirmal Perera is a Technical Sales and Field Application Specialist for ATCC with LGC. He's been in his current role for almost 2 years now, attending and presenting at conferences all over the world.


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #13-A Functional Study of the Endomembrane System in Mammalian Cells Growing in a 3D Environment
The human body is a complex assembly of tissues and organs, each of which is made of different types of cells. Specialisation of function is achieved via compartmentalisation within each cell, as well as defined communication networks that exist between endomembrane compartments. The endomembrane system is essential for the synthesis and distribution of proteins, lipids and carbohydrates, and as such needs to be tightly regulated. One protein family of master regulators of the endomembrane system is the Rab small GTPases. Rab proteins govern the communication between organelles, via the GDP-GTP cycle and by exhibiting specific membrane localisation patterns. To date, their distribution and function have mostly been investigated in conventional two-dimensional in vitro cell models, however three-dimensional cell models, so far applied to investigate cancer and drug development, offer new possibilities. In this work, a functional study is being used to understand the localisation and role of Rab proteins in the endomembrane system in a three-dimensional environment. For this, a three-dimensional cell model (spheroids), using HeLa Kyoto cells grown on micropatterned plates, has been developed. This approach allows highly uniform spheroids to be grown in a tightly controlled manner. In order to localise various Rab proteins, stably expressing cell lines expressing GFP-tagged Rab proteins have been produced and grown as spheroids. Automated high-content screening microscopy is now being used to quantitatively describe the localisation and function of Rab proteins in this new cell system, and in turn, compare the results to existing data coming from classical two-dimensional cell models.

Speakers
MM

Margaritha Mysior, B.Sc., M.Sc.

PhD candidate, University College Dublin
I absolved a Bachelor (2012-2015) and Master (2015-2017) degree program at the University of Applied Science Gelsenkirchen, Recklinghausen and Bocholt. During my Master thesis (October 2016 - August 2017) I characterised five unidentified regulators of the retrograde transport pathway... Read More →


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #14- GBM Infiltration of Cerebral Organoids Models Brain Tumour Invasion
Introduction
Glioblastoma (GBM) is the most common and most aggressive brain tumours in adults. No cure is available and complete surgical resection is not achievable due to GBM cell infiltration into the healthy brain parenchyma. Therefore, interventions that specifically target the invasive GBM phenotype are highly desirable. However, it is challenging to model the process of GBM infiltration in real time in the laboratory in relevant model systems encompassing both tumour and neural tissue compartments. Previously we demonstrated the ability of patient derived GBM spheroids to spontaneously infiltrate mouse derived early-stage cerebral organoids (CO). Here we present an enhanced culture system enabling tractable ex vivo investigation of human GBM into human-derived CO.

Methods
Adaptation of previously described methods in murine CO has allowed us to see spontaneous infiltration of GFP-expressing GBM patient derived cell model spheroids into early stage human COs and quantified using live cell imaging and histological analysis.

Results
Patient derived GBM infiltrated in all cases CO whereas neural progenitors (NP) were unable to infiltrate. The GBM spheroid and CO self-assembly and tumour cell infiltration were characterised quantitatively using several parameters including total GBM cell infiltration, migration distance, and cell compartment spread in 3D. This ‘assembloid’ assay was also able to differentiate ‘early’ and ‘late’ invasion capabilities of GBM cell models from different patients and GBM subtypes, as well as successful infiltration of a freshly resected tumour piece.

Conclusion
We demonstrate that GBM/CO assembloids can be adapted for scalable imaging applications (for a throughput of n>100 and >10 conditions per time lapse experiment). They can be used as an ex vivo experimental model system that aims, in real-time, to recapitulate the infiltrative characteristic of GBM that makes current therapy so ineffective.

Looking forward, in collaboration with the Stem Cell Hotel (King’s College London) and industry partner Cellesce (Cardiff), we plan to automate the quantification of multiple relevant imaging parameters that may enable predictive biological readouts in the future. Furthermore, we plan to use brain tumour assembloids to establish proof-of-principle work for laboratory testing of personalised treatments as a clinical decision system that may aid treatment stratification of brain tumour patients.

Speakers
BI

Bronwyn Irving, MSci

PhD Student, University of Leeds
Bronwyn is currently a PhD student at the University of Leeds following her passion for cancer research after previously working at UCL’s Institute of Child Health and Newcastle University’s Northern Institute for Cancer Research.


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #15- High Content Screening Platforms for the Evaluation of Steatosis Induction in 2D and 3D HepG2 cultures
Steatosis, or the increased accumulation of lipids in the liver, can be induced by several factors including exposure to steatogenic drugs and through pertubations in lipid metabolism indicative of non-alcoholic fatty liver disease (NAFLD). Hepatic lipid accumulation is a key driver of morphological and functional changes to liver architecture that leads to the development of acute and chronic liver disease and fibrosis. We have developed a number of high content screening platforms to screen for drug induced steatosis using 2D HepG2 cells and 3D HepG2 C3A spheroid cultures with detection of lipid accumulation by LipidTOX green and Nile red.Additionally, we have adapted our 3D spheroid platform to evaluate steatosis induced by dietary fatty acid uptake. We demonstrate our high throughput screening platforms utilising automated ECHO650 and Biomek NxP compound dispense combined with High Content Imaging of lipid accumulation using the ImageXpress Micro Confocal. Our assays are highly reproducible and have been validated using a panel of known steatogenic drugs. Development of such platforms will enable the early screening of candidates with steatogenic potential with a view to de-risking and de-selecting adverse compounds during the Discovery phase of drug development.

Speakers
KO

Katalin Orban, MSc, PharmD

Senior Scientist, Sygnature Discovery
Katalin Orban currently works in the DMPK and Physical Sciences Department at Sygnature Discovery where she contributes to the strategy, interpretation and overall DMPK assessment of NCEs in small molecule drug discovery.Katalin previously graduated from the Semmelweis University... Read More →


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #16- Genedata Imagence®: An Evaluation of Deep Learning for High Content Analysis
Machine learning has seen some revolutionary and remarkable developments over the last few years. Exciting breakthroughs have been achieved in artificial neural networks, in particular around deep network architectures (= deep learning), which work based on raw-pixel image information and provide a classification accuracy exceeding human expert judgement. Deep neural networks are now the state-of-the-art machine learning models across a variety of areas, including image processing and analysis and are widely implemented in academia and industry.

Genedata has developed the first commercially available solution for deep learning-based High Content Image analysis. Genedata Imagence® allows for the application of deep networks to the analysis of High Content Imaging, creating a workflow that cuts image analysis time, increases data quality, reproducibility of results and seamlessly integrates with Genedata Screener® for image data analysis. This deep learning approach outperforms conventional approaches for feature extraction and phenotype classification.

Here we provide the evaluation of a recent pilot of Genedata Imagence® for High Content Analysis. We compare the Genedata Imagence® workflow with our conventional in-house High Content Imaging and Analysis workflow through a series of assays that have been developed within the target validation biology group at LifeArc. These include, immunocytochemistry of shRNA knock-down clones, antibody internalisation assay, High Content cell health assay, growth cone collapse assay and osteoclast differentiation assay.The generation of training data and subsequent training of the network was carried out for each assay and results of the data analysis were compared to in-house conventional analysis. We discuss a few limitations but show overall an excellent data quality produced by this novel deep learning module for High Content Image analysis.

Speakers
ZI

Zaynab Isseljee

Scientist, LifeArc
Zaynab Isseljee is an enthusiastic research scientist with ten years’ experience in drug discovery. Graduated with a BSC (Hons) in Biomedical Science at King's College London with an industrial placement year at Novartis Horsham Research Centre. Since 2009, Zaynab has worked at... Read More →


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #17- Deep Learning-Derived Features Outperform Classical Computer Vision in Low Dimension-Embedding of High-Content Screening Data
When automating analysis of high-content screening images, a key step is reducing the object image (e. g. a cell) to a vector of features that describe that object for subsequent comparison with other objects. In classical computer vision, many of these features have an implicit biological meaning (e. g. cell size, intensity, aspect ratio, etc).

Depending on the research question, feature vectors then are subjected to univariate or multivariate analysis. Nonlinear dimensionality reduction techniques such as tSNE and its variants [1] are particularly useful when representing or embedding high-dimensional data in two or three dimensions.Ideally, such visualizations should clearly separate different phenotypes observed in the experiment. However, the embedding quality relies heavily on the quality of the input features derived from the HCS images.

In this poster, we use a high-content screening translocation assay to compare the quality of embedding produced by features extracted using classical versus deep learning-based feature approaches. We compare them based on ability to separate phenotypes and robustness against batch effects. We show that while classical and deep-learning-derived feature sets or a combination of both all produce excellent results, when using classical features, attaining this level of quality requires expert tuning of the feature extraction process to the assay of interest. In contrast, deep-learning-based feature extraction is fully automated and does not require expert knowledge.

Using deep learning-based features, we observe that the embedding quality depends on the network architecture. Standard CNN architectures perform poorly, while tailored architectures outperform classical methods. Finally, we show that for embedding and visualization, adding classical features to the deep-learning based features does not increase resolution and is thus unnecessary.

[1] L. J. P. van der Maaten and G. E. Hinton. Visualizing High-Dimensional Data Using t-SNE. Journal of Machine Learning Research 9(Nov):2579-2605, 2008

Speakers
MF

Matthias Fassler

Scientific Account Management, Genedata
Matthias Fassler is a Scientific Account Manager at Genedata, Basel.


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #18- Novel Organ-on-Chip Microfluidic Device based on Hydraulically Actuated Hydrogel Layers
Organs-on-chips (ooc) are small devices aiming to recapitulate the physiology and/or pathology of a living organism. Contrarily to commonly used flasks and dishes, ooc devices are designed to recapitulate the physico-chemical microenvironment of the cells. Thanks to these mimics of higher relevance, ooc devices are able to enhance cell differentiation compared to 2D culture techniques.
The field of ooc is still young, and many improvements remain to be done. For instance, one of the first organ-on-chip, produced by Huh et al. (2010), submitted epithelial and endothelial cells to mechanical stimuli by stretching a thin perforated PDMS membrane on which cells were attached. While extremely efficient, this setup presents several shortcomings, such as the high stiffness of PDMS compared to that of soft living tissues, and the poor relevance of a perforated membrane to study extravasation.
Here we present a novel ooc microfluidic device allowing the coculture of cells in and on a soft hydrogel. Thus, a greater tissue complexity can be mimicked, including notably a “stromal compartment” in addition to the usual epithelium-endothelium dual culture. Relying on a pressure differential between each side of the hydrogel and resistances to adjust the flow, the hydrogel can be stretched. Thus, in the final setup, most physico-chemical parameters (stiffness, flow, strain, functionalization, etc.) are under the control of the operator to best suit the desired organ and application.

Nicolas MINIER1, 2, Samy GOBAA1

Speakers
NM

Nicolas Minier

Institut Pasteur


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #19- Fully Automated and Scalable 3D-Baed Toxicity Testing in Human Neural Microtissues

Recent advances in stem cell technology have led to the development of three-dimensional (3D) culture systems, which have fueled hopes to bring about the next generation of more physiologically relevant high throughput screens (HTS). However, current protocols yield either complex but highly heterogeneous aggregates (“organoids”) or 3D structures with less physiological relevance (“spheroids”). Here, we present a fully scalable, HTS-compatible workflow for the automated generation, maintenance, and high content analysis of human midbrain-like automated microtissues (mANTs) in a standard SBS 96-well format. The resulting microtissues possess a highly homogeneous morphology, size, global gene expression, cellular composition, and internal organization. They present significant features of the developing human midbrain and display spontaneous aggregate-wide synchronized neural activity. By automating the entire workflow from generation to analysis, we provide essential intra- and inter-batch reproducibility as demonstrated via RNA sequencing and quantitative whole mount high content imaging. Dose response toxicity testing showcases our ability to quantify drug effects on single cells within a complex 3D environment using mANTs. In conclusion, our workflow has the potential to form the basis for 3D-tissue-based screening in a variety of applications including toxicology studies, disease modelling, and drug development.
 

Speakers
HR

Henrik Renner, Msc.

PhD Student, Max Planck Institute for molecular Biomedicine
- PhD student in the lab of Hans Schöler at the MPI in Münster (since 2016), Germany (creating 3D HTS tissue models for midbrain)- Msc. Karolinska Institute (2016)- Bsc. University of Göttingen (2014)


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #20-AI-driven Optimization of Multidrug Combinations in Heterotypic 3D Human Co-cultures
Heterotypic three-dimensional (3D) short-term human cellular models were developed based on the histopatological patient tissues to predict the efficacy of new (combined) treatment regimens for treatment of various stages of human colorectal carcinoma (CRC). These spheroidal callular co-cultures included human CRC cell lines of various genetics and stage co-cultured with human endothelial cells and fibroblasts.

The exposure of those spheroids with several low-dose three- or four-drug combinations, designed using artificial-intelligence-based method allowed efficient optimization of the most effective treatment strategy.

The optimized synergistic cell type-specific drug combinations induced cell metabolic activity inhibition, that indirectly corresponding to cell viability inhibition, by up to 90%. Moreover, the activity of those optimized drug combinations was often more potent in 3D models as compared to simple 2D cell cultures and facilitated further translation to more complex in vivo model.

Summarizing, our technology represents an innovative step to obtain enhanced accuracy of the activity of cell type-specific treatments. Our multicellular spheroids resemble in many aspects solid tumors and serve as a promising model to mimic the in vivo situation.

Speakers
PN

Patrycja NowakSliwinska, Ph.D., D.Sc.

Professor, Institute of Pharmaceutical Sciences of Western Switzerland
She performed her post-doctoral work in the Medical Photonics Group at the Swiss Federal Institute of Technology (EPFL) in Lausanne, Switzerland, and in the Angiogenesis Laboratory at the VU University Medical Center in Amsterdam, The Netherlands. She obtained the certificate of advanced... Read More →


Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer

1:30pm BST

Poster Presentation #21- Icy2.0: An open and free software for bioimage analysis
We present Icy2.0 (http://icy.bioimageanalysis.org), which is a major evolution of our previous free and open software for bioimage analysis Icy. This release has three important features: 1) the ability to handle very large images (> 4Gb); 2) the possibility to process long sequences through an optimized streaming procedure with adaptive caching; and 3) a totally redesigned web site for improved interactions within the Icy community. Icy2.0 has retained and consolidated the major features that made its success in the bioimaging community, namely to make it available to the bioimaging community an image analysis software suite that encompasses the large variety of biological applications (microscopy, particle tracking, HCS/HTS, digital pathology, animal behavior…) and users (biologist, bioimage analyst, physicist, developer) and to give access to advanced image analysis methods and solutions.

Since its launching, it has been the constant philosophy of the Icy team to promote sharing of source code and know-how and to facilitate the use of quantitative approaches and open new scientific perspectives in terms of exhaustiveness, reproducibility and robustness of the analysis of bioimaging data sets. For those reasons, Icy2.0 has a fully redesigned web-site, that includes new communication channels between the end-users and the developers, new tutorial material and improved maintenance cycles. Icy2.0 now provides more than 400 dedicated plugins covering a large variety of state-of-the-art image analysis methods ranging from active contours models to Machine Learning through statistical spatial analysis, which empower users with the most recent and adapted quantitative image analysis and visualization tools. Protocols, which are a graphical front-end that enable software development without any programming knowledge, have also been improved and now include the possibility to develop sophisticated image processing pipelines in a more readable and interactive manner.

Icy2.0 has a large community of developers and users (4000+ regular users, 700+ students trained, 1500+ visits per month) that are using Icy for distributing their algorithms and plugins. For its long-term sustainability and development, the Icy2.0 project benefits from the institutional support of the French national Infrastructure FranceBioImaging and of Institut Pasteur. Ongoing work aims at improving the interoperability and convergence of Icy2.0 with other open and free image analysis packages as well as facilitating the integration and use of AI packages.



Speakers

Tuesday October 22, 2019 1:30pm - 2:00pm BST
Sherry Coutu Seminar Suite Foyer