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Tuesday, September 19 • 11:05am - 1:30pm
Session 1: Technology
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Microscopy is fast evolving field with development in resolution, imaging modalities, flexibility of acquisition and much more. This session will show how some of these exciting advances are being applied in automated screening setups to allow screening more complex cellular systems for higher clinical relevance.

11:05 - 11:30:
Droplet-Microarray – a miniaturized platform for high throughput screenings
Anna Popova, Institute for Toxicology and Genetics, Karlsruhe Institute of Technology

Phenotypic high throughput screening (HTS) of live cells is a methodology that lies at the basis of fundamental research, drug discovery and toxicology. The main problematic of screenings in our days lies in relatively large volumes per experiment and at the same time a need for higher throughout. Miniaturization of high throughput screenings has been a major trend in past years. Since further miniaturization of microtiter plates is not possible, alternative technologies are emerging to address this problem. Here we present a Droplet-Microarray (DMA) platform - a novel miniaturized platform for HTS of live cells. DMA consists of an array of superhydrophilic spots on superhydrophobic background. Due to extreme difference in wettability between superhydrophilic and superhydrophobic areas cell suspension applied on such surface spontaneously splits in array of homogeneous separated droplets of nL volumes, enabling pipetting-free seeding of cells. We have developed protocols for parallel addition of compounds and reagents simultaneously to the whole array without a need for pipetting of each individual compartment. The DMA platform is transparent and compatible with all kinds of microscopy. In addition, the DMA has dimensions of microscope glass slide or microtiter plate and, therefore, is compatible with all existing microscopes. All the standard protocols including staining, fixation and immunofluorescence can be performed on DMA platform, making it easy to incorporate it into existing screening workflows. In the past years we have evaluated DMA platform for cell-based screening applications. We successfully cultured over 10 different adherent and suspension cell lines, stem cells and primary patient-derived cells in miniaturized format. We have established protocols for transfection and compound treatment of cells on the DMA platform. We optimized procedures for staining, fixation and microscopy of cells of suspension and adherent nature. We tested DMA platform in a proof-of-concept screen on primary patient-derived tumor cells with antineoplastic compounds. We evaluated DMA platform for screening of cells in 3D environments by means of hydrogels, spheroids and embryonic bodies. Moreover, we further extended the portfolio of applications of DMA platform to whole – organism screenings. Using DMA platform we created single-embryo-array where single zebrafish embryo is trapped in an individual droplet, and evaluated this array for toxicity screening applications. Single-embryo-array based on the DMA principle enables detailed microscopic analysis of embryos, which are fixed in particular position in array format, significantly simplifying imaging process and image analysis of live animals. We believe that DMA technology carries a great potential to be adopted for various high content screening applications. On one hand, it is flexible and universal due to its compatibility with various cell types, different functional assays, standard protocols and all types of microscopy. On the other hand, it is highly miniaturized (3-80 nL) and pipetting-free. Thus, the DMA platform is a chip technology for cell-based assays that enables miniaturized and parallelized high throughput screenings of live cells in 2D and 3D, as well as whole-organisms.

11:30 - 12:00:
Applications of non-invasive phenotypic screens using label free imaging by digital holography; three selected examples
Gerardo Turcatti, EPFL 

In the fast evolving field of screening by imaging, a main challenge is to conciliate complex models or assays with more in vivo relevance and throughput. In this context, we are developing assays with representative cells or models amenable to screening using holographic microscopy (DHM) for imaging cells. DHM is a label-free interferometric microscopy technique, which provides a quantitative measurement of the optical path length. It is a two-step process where a hologram consisting of an interference pattern is first recorded on a digital camera and the quantitative phase images are reconstructed numerically using a specific algorithm. We illustrate here three phenotypic screening applications using DHM as imaging read-out: a) a drug screening method for adipocytic differentiation through lipid quantification b) a screen for quantification of cardiomyocytes beating dynamics c) a screening assay for measuring trans-epithelial fluxes in CFTR mutations using gastrointestinal-derived organoids.

12:00 - 12:30:
Cancer organoids for screening of chemotherapeutics and personalized medicine
Francesco Pampaloni, Buchmann Institute for Molecular Life Sciences (BMLS)

Three-dimensional (3D) cell cultures allows establishing physiologically relevant in vitro tissue models, which are of great interest for both industry and clinical research.  In my talk, I describe two 3D cell culture platforms for drug screening based on live fluorescent microscopy recently developed in our group. The first platform is based on U343 glioma 3D multicellular spheroids, which represent a realistic brain cancer model. With the U343 spheroids we established a drug screening pipeline for the identification of autophagy modulators. Since the autophagic flux is often dysregulated in cancer, the discovery of novel autophagy modulating drugs is of great clinical relevance. This screening assay with 3D tumor spheroids is robust, reproducible and scalable and represents a valuable tool for both basic research and pharmaceutical industry [1].The second drug screening platform uses hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC) stem cell organoids. The tumor stem cell organoids are obtained from liver biopsies and can be virtually limitless expanded in culture [2]. I illustrate the pros and contra of this new technology and our pipeline for the testing of chemotherapeutics with high-throughput microscopy, discuss realistic approaches for a personalized cancer therapy, and show first data recorded with Light Sheet Fluorescence Microscopy (LSFM). 

12:30 - 13:00:
Combining high-content imaging and pathway profiling technologies to advance in vitro pharmacogenomics and discovery of novel drug combinations
Neil Carragher, Institute of Genetics and Molecular MedicineUniversity of Edinburgh

Recent advances in high-content screening, CRISPR/Cas-based genome editing and induced pluripotent stem cell technologies are converging to stimulate the new discipline of Phenomics Drug Discovery. I will describe how Phenomics Drug Discovery incorporates state-of-the-art imaging, genomics, proteomics and informatics tools to explore drug mechanism-of-action (MOA) studies across genetically defined in vitro cell models. We demonstrate how multiparametric high content screening assays combined with novel image-informatics and machine learning methods advance in vitro pharmacogenomic studies across a broader variety of phenotypes, disease models and therapeutic classes. We also demonstrate how ultrasensitive Reverse Phase Protein Microarray (RPPA) technologies can combine with imaging to characterize drug MOA at the post-translational pathway network level. Finally, I present recent examples of how our Phenomics Drug Discovery platform enabled the rapid discovery of a novel orally-available, ATP-competitive, kinase inhibitor and novel drug combinations which display potent anti-tumour activity across 2D, 3D and in vivo models.

13:00 - 13:30:
Multiscale cytometry and regulation of 3D cell cultures on a chip
Charles Baroud, Ecole Polytechnique

Three-dimensional cell culture is emerging as a more relevant alternative to the traditional two-dimensional format. Yet the ability to perform cytometry at the single cell level on intact three-dimensional spheroids or together with temporal regulation of the cell microenvironment remains limited. I will describe a microfluidic platform to perform high-density three-dimensional culture, controlled stimulation, and observation in a single chip. The method extends the capabilities of droplet microfluidics for performing long-term culture of adherent cells. Using arrays of 500 spheroids per chip, in-situ immunocytochemistry and image analysis provide multiscale cytometry that yields information at the population scale, at the scale of 10,000 or more single spheroids, and nearly 1,000,000 single cells. This allows us to correlate functionality with cellular location within the spheroids and the local structure. Also, an individual spheroid can be extracted for further analysis or culturing. Then by combining this approach with a more advanced droplet handling approach, I will demonstrate how a complete drug response curve can be obtained in a single sweep on a chip. This approach will enable a shift towards quantitative studies on three-dimensional cultures, under dynamic conditions, with implications for stem cells, organs-on-chips, or cancer...

avatar for Charles Baroud

Charles Baroud

Ecole Polytechnique
Charles Baroud was trained as a physicist and engineer at MIT, then received a PhD at the University of Texas at Austin. Since then he has founded and now leads the microfluidics group at Ecole Polytechnique in Paris. The group has performed fundamental fluid dynamics research on... Read More →

Neil Carragher

Professor Neil Carragher graduated from the University of Aberdeen, Scotland UK in 1992 with a B.Sc Honours degree in “Cell and Immunobiology”.  He then took up a position within industry at the Yamanouchi Research Institute (now Astellas Pharma), Oxford, England UK where he also gained his PhD. He then held consecutive postdoctoral positions within the Department of Pathology, University of Washington, Seattle, USA and at the Beatson Institute for Cancer Research, Glasgow, Scotland UK. In 2004 Neil returned to the pharmaceutical industry as Principal Scientist with AstraZeneca where he pioneered early multiparametric high-content phenotypic screening approaches. In 2010 he once again made the career switch from industry to academia and took up the post of Principal Investigator of Drug Discovery at the University of Edinburgh. Primary research interests include advancing high-content analysis, phenotypic screening, Reverse Phase Protein Array technology, drug combinations and cancer drug... Read More →
avatar for Francesco Pampaloni

Francesco Pampaloni

Francesco Pampaloni is staff scientist with permanent position at the Buchmann Institute for Molecular Life Sciences (BMLS) in Frankfurt am Main (Germany). He leads the research on three-dimensional cell cultures and he is coordinator and scientific manager of the EU Horizon 2020 project LSFM4LIFE. He graduated in Physical Chemistry at the University of Florence (Italy). Awarded with an EU Marie-Curie Fellowship programme, he moved to the University of Regensburg (Germany), where he obtained his Ph.D. with excellence with a work on Optical Force Microscopy. From 2003 to 2009 post-doc and staff scientist at EMBL Heidelberg (Germany). Since 2010 at the Buchmann Institute for Molecular Life Sciences (BMLS) of the Goethe University Frankfurt. He authored 40+ publications and four patents. He is main inventor and driving force of the High-Throughput LSFM (HT-LSFM) and the Tissue Culture LSFM... Read More →
avatar for Anna Popova

Anna Popova

Institute for Toxicology and Genetics, Karlsruhe Institute of Technology
avatar for Gerardo Turcatti

Gerardo Turcatti

Director of the Biomolecular Screening Facility, EPFL
Dr. Gerardo Turcatti directs the Biomolecular Screening Facility (BSF) he created at the EPFL in 2006. In the frame of the National program NCCR-Chemical Biology, he leads the project ‘ACCESS’ ‘An Academic Chemical Screening Platform for Switzerland’ since 2011. Previously he co-founded and acted as CTO of Manteia S.A., a Swiss-based company that developed revolutionary high throughput DNA sequencing technologies currently commercialized by Illumina Inc. Prior to this experience, Dr Turcatti had a long multidisciplinary career in... Read More →

Tuesday September 19, 2017 11:05am - 1:30pm
Rosales I Courtyard by Marriot Madrid Princesa Hotel