Phenotypic screening continues to be a valuable tool for drug discovery. Although target-based screening is gaining popularity, a disproportionate number of first-in-class drugs with novel mechanisms of action are discovered via phenotypic screening. Cambridge Healthtech Institute’s Phenotypic Drug Discovery will provide case studies of phenotypic screening in various physiologically relevant 3D models, including models in immuno-oncology, infectious disease, inflammation, neurodegeneration and cancer.

Final Agenda

Arrive early and attend (Mon-Tues): High-Content Analysis

Tuesday, November 7

11:00 am Conference Registration

12:00 pm Luncheon Presentation: Testing Cancer Therapeutics in Cultured Human 3D Tissues

Leo Price, Ph.D., CSO, OcellO

PDX models have been used to test a variety of targeted therapies in vivo. To expand the potential of these models, 3D in vitro methods have been developed showing in vivo-like growth characteristics, invasion, and responses to therapeutics. Combined with advanced imaging analysis, a unique high throughput in vitro PDX screening platform allows efficient identification of active and selective molecules while enabling selection of the optimal PDX models for subsequent validation of candidates in vivo.

12:45 Dessert Break with Exhibit and Poster Viewing

Phenotypic Screening of 3D Models

1:25 Chairperson’s Opening Remarks

Sophie A. Lelièvre, D.V.M., Ph.D., LL.M. (public health), Professor, Department of Basic Medical Sciences, College of Veterinary Medicine; Associate Director, Collaborative Science, Purdue University Center for Cancer Research; Scientific Director, 3D Cell Culture Core (3D3C) Facility, Birck Nanotechnology Center, Discovery Park; Coleader, International Breast Cancer & Nutrition (IBCN); Professor of Cancer Pharmacology, West Lafayette-IU Medical School

1:30 Exploiting 3D Cell Cultures for Drug Discovery and Regenerative Medicine

Jonathan S. Dordick, Ph.D., Howard Isermann Professor of Chemical and Biological Engineering and Vice President for Research, Rensselaer Polytechnic Institute

Myriad factors (e.g., cytokines and other proteins, small molecules, and extracellular matrix components) and their combinations strongly influence cell function and fate, which has direct relevance in drug discovery, human toxicology and regenerative medicine. We have developed in vitro, 3D cellular environments that can be engineered with the capacity to emulate the in vivo cellular environment. To demonstrate the breadth of the 3D culture platforms, human neural stem cell lines are being used to elucidate factors that guide neuronal differentiation, and primary cells in vitro are being compared to identical cells engrafted into humanized mouse models. As a result, highly predictive human outcomes may be predicted.

2:00 FEATURED PRESENTATION: Translational Applications of Organ-on-a-Chip Technologies

Murat Cirit, Ph.D., Director of Translational Center of Tissue Chip Technologies, Massachusetts Institute of Technology

In vitro models have been developed and utilized in various stages for the preclinical development. Compared to animal models, in vitro models have advantages such as high-throughput capability, low cost, well-controlled experimental parameters and fewer ethical concerns etc. The simplicity of the conventional in vitro models makes them incapable of achieving adequate physiological relevance for mimicking the human body, however, which is a dynamic system that has complex three-dimensional microenvironment, intracellular communications and organ interactions. Hence, there is an urgent need to develop more physiologically relevant in vitro systems for better simulating the human body in response of drugs and providing more reliable in vitro in vivo translation (IVIVT) from preclinical results to clinical outcomes.

2:30 Gradient-on-a-Chip to Recreate Microenvironmental Heterogeneity and Identify Cellular Response Thresholds

Sophie A. Lelièvre, D.V.M., Ph.D., LL.M. (public health), Professor, Department of Basic Medical Sciences, College of Veterinary Medicine; Associate Director, Collaborative Science, Purdue University Center for Cancer Research; Scientific Director, 3D Cell Culture Core (3D3C) Facility, Birck Nanotechnology Center, Discovery Park; Coleader, International Breast Cancer & Nutrition (IBCN); Professor of Cancer Pharmacology, West Lafayette-IU Medical School

Cell exposure to chemicals traveling through the extracellular matrix depends on variable local concentrations. We designed a microfluidics-based gradient-on-a-chip for controlled delivery of a chemical gradient within the liquid bed of an open culture chamber. As proof-of-concept we used a gradient of reactive oxygen species involved in cancer progression. Preinvasive tumors displayed increasing oxidative stress and phenotypic responses, indicative of a more aggressive status, as the gradient concentration increased. The response level was influenced by matrix stiffness, a known factor in cancer progression, and tumor grade.

3:00 Microengineered 3D Hydrogels for Tissue Engineering and Surgical Applications

Nasim Annabi, Ph.D., Assistant Professor, Chemical Engineering, Northeastern University

Tissue engineering is an interdisciplinary field aimed at maintaining, restoring and enhancing the normal function of organs and tissues through the use of live cells, and by incorporating concepts from engineering, biological sciences and medicine. One of the central themes in the field of tissue engineering is the development of tissue constructs that mimic the three-dimensional (3D) architecture of native tissues. To date, tissue engineering has been successfully implemented in the engineering of several types of tissues including bone, cartilage, and vascular systems. Despite the significant progress in this field, many challenges still remain, which hinder the development of fully functional tissue construct. Micro- and nanoscale technologies have been shown to hold great potential to address the current challenges in tissue engineering. These technologies have immensely benefited the fields of experimental biology and medicine, and have allowed the design of complex biomaterials that can be used for cell-based studies. Our research is focused on merging micro/nanofabrication techniques with advanced protein-based biomaterials for tissue engineering applications. Our group has been actively involved in engineering novel cell-laden elastomeric biomaterials with unique physical properties by using recombinant protein technologies. We use these elastomers as 3D matrices for various soft tissue engineering applications. In addition, we utilize them as highly elastic and adhesive hydrogels for wound closure after different surgical procedures. Our work encompasses a wide range of scientific subjects, from materials science to biology. In this presentation, I will outline our work in the development of microscale hydrogels to modulate cell-microenvironment interactions for tissue engineering applications. I will also highlight some of the clinical applications of our engineered biomaterials as surgical sealants and tissue adhesives.

3:30 Refreshment Break with Exhibit and Poster Viewing

4:10 Nonlinear Optical Microscopic Imaging for Non-Invasive and Label-Free Screening of Viability and Function in 3D Engineered Tissues

Mary-Ann Mycek, Ph.D., Associate Professor & Associate Chair, Biomedical Engineering; Faculty Member, Applied Physics Program; Core Member, Comprehensive Cancer Center, University of Michigan

Label-free nonlinear optical microscopic imaging with quantitative image analysis non-invasively screened viability and function in engineered tissues in 3D. In situ optical measures of living tissue morphology and function could serve as quality control criteria for engineered tissues prior to implantation in patients, a critical regulatory requirement in regenerative medicine. Applications to engineered oral mucosa and skeletal muscle will be discussed.

4:40 A 3D ex vivo Orthotopic Xenograft Screening Platform to Identify Novel Drug and Drug Target Candidates for Brain Cancers

Shawn D Hingtgen, Ph.D., Assistant Professor, Clinical Assistant Professor, Molecular Pharmaceutics, University of North Carolina Chapel Hill

In preclinical development of brain cancer therapeutics, it has become increasingly apparent that conventional, 2D cell culture models (i.e., brain cancer cell lines propagated in plastic) diverge substantially from primary tumors in genomic as well as phenotypic characteristics. To address this, we are developing a novel 3D bona fide brain tissue-based, high-content and high-context assay platform for the identification and validation of new brain cancer drug and drug target candidates.

5:10 Engineered Assembly of Stem Cell-Derived Human Tissues for Discovery and Therapy

William L. Murphy, Ph.D., Harvey D. Spangler Professor, Biomedical Engineering and Orthopedics; Co-Director, Stem Cell and Regenerative Medicine Center; Director, Human MAPs Center, University of Wisconsin

The next generation of high throughput cell-based assay formats will require a broadly applicable set of tools for human tissue assembly and analysis. To address these needs, we have recently focused on: i) generating iPS-derived cells that properly represent the diverse phenotypic characteristics of developing or mature human somatic cells; ii) assembling engineered organoids that are robust and reproducible; iii) translating engineered organoids to microscale systems for high throughput screening; iv) combining genomic analyses with bioinformatics to gain insights into engineered organoid assembly and the pathways influenced by drugs and toxins; and v) assembling cellular constructs that can be efficiently delivered in therapeutic applications. This talk will emphasize recent studies on engineered assembly of 3-dimensional “cellular scaffolds” and engineered vascular, neural, and hepatic organoids. The talk will also introduce the use of our engineered organoids to develop models of developmental disorders, degenerative diseases, and infectious disease effects.

5:40 Close of Day

Wednesday, November 8

7:30 am Breakfast Breakout Roundtable Discussions

 

7:30 am Breakfast Breakout Roundtable Discussions

Concurrent breakout discussion groups are interactive, guided discussions hosted by a facilitator to discuss some of the key issues presented earlier in the day’s sessions. Delegates will join a table of interest and become an active part of the discussion at hand. To get the most out of this interactive session and format please come prepared to share examples from your work, vet some ideas with your peers, be a part of group interrogation and problem solving, and, most importantly, participate in active idea sharing.

Table 1: The Art of Choosing Appropriate 3D Cell Culture Models

Moderator: Sophie A. Lelièvre, D.V.M., Ph.D., LL.M. (public health), Professor, Department of Basic Medical Sciences, College of Veterinary Medicine; Associate Director, Collaborative Science, Purdue University Center for Cancer Research; Scientific Director, 3D Cell Culture Core (3D3C) Facility, Birck Nanotechnology Center, Discovery Park; Coleader, International Breast Cancer & Nutrition (IBCN); Professor of Cancer Pharmacology, West Lafayette-IU Medical School

  • When are organs-on-a-chip better suited than standard 3D cell cultures to address a biological question?
  • What is the importance of the cell model and of the culture medium for 3D culture compared to classical (2D) cell culture?
  • Is there a real advantage of using 3D bioprinting to design and build 3D culture models?

Table 2: Mimicking Tumor Immunosuppression in a Dish - Use of Cell Culture Models for Immuno-oncology

Moderator: Jakub Swiercz, Head of In Vitro Pharmacology & Screening, iTeos Therapeutics SA

  • What are current challenges to overcome?
  • Can one model mimic multiple immunosuppression mechanisms
  • 2D vs 3D models for compound screening in IO

Table 3: Multi-Organ Organ-on-a-Chip Models

Moderator: James J. Hickman, Ph.D., Professor, Nanoscience Technology, Chemistry, Biomolecular Science, and Electrical Engineering, University of Central Florida

Table 4: Toxicity Screening in 3D Models

Moderator: Matt Wagoner, Associate Director, Mechanistic and Investigative Toxicology, Takeda Pharmaceuticals

  • The devil’s in the details | Quantitative considerations for toxicity screening
  • The Valley-dation of Death | Building a bridge between academic innovation and the high-confidence assays needed by industry

Table 5: Screening Complex Biological Models

Moderator: Andreas Vogt, Ph.D., Associate Professor, Department of Computational and Systems Biology, University of Pittsburgh Drug Discovery Institute

  • Are complexity and heterogeneity obstacles or opportunities? Should we abandon a good model just because it is difficult to analyze and standardize? What influences that decision?
  • How much assay development is needed? How much is enough?
  • Are canonical measures of assay performance appropriate for complex systems? What are the alternatives?
 

Case Studies: High-Content Analysis & Phenotypic Screening

8:25 Chairperson’s Remarks

David Nolte, Ph.D., Professor, Physics, Purdue University; President, Animated Dynamics, Inc.

8:30 Breakout Roundtable Report-Outs

9:00 Development and Validation of a Phenotypic Screening Platform for the Identification of Novel Immuno-Oncology Targets

Jakub Swiercz, Head of In Vitro Pharmacology & Screening, iTeos Therapeutics SA

iTeos Therapeutics has developed a target discovery and drug repurposing platform based on phenotypic screening assays. We have established a co-culture assay combining tumor immune suppressive cells and T-cells. This assay is flexible to allow the screening of chemogenomics, shRNA and cDNA libraries. Multi-parameter readouts are combined to assess both T cell activation and proliferation.

9:30 Quantifying Stem Cell Colony Phenotypes: Live Cell Imaging and Visualization and Analysis of Very Large Datasets

Anne Plant, Ph.D., Chief of the Biosystems and Biomaterials Division, National Institute of Standards and Technology

Automated instruments easily allow many GB of image data to be collected in an experiment, which is more data than can be visualized on a desktop computer. We have developed tools that allow panning and zooming of 1 GB images composed of hundreds of fields of view, which permit visualization of colonies as they expand and merge over 5 days in culture. Our interest is in validation of the quantitative analysis of these data, which ultimately allows validation of putative biomarkers of pluripotency and differentiation.

10:00 Networking Coffee Break

10:30 Chairperson’s Remarks

David Nolte, Ph.D., Professor, Physics, Purdue University; President, Animated Dynamics, Inc.

10:35 Balancing Hypothesis Driven and Empirical Drug Discovery

Jonathan A. Lee, Ph.D., Senior Research Advisor, Quantitative Biology, Eli Lilly and Company

Target agnostic, empirical, drug discovery strategies have contributed a disproportionate number of first in class medicines. Accordingly, this target agnostic-empirical or “phenotypic” drug discovery (PDD) strategy has been the subject of active debate/discussion between academic, biotechnology, and pharmaceutical scientists. PDD is challenging and is not a magic bullet or a quick fix. Multiple considerations including scientific, process timeline, business strategy, organizational risk tolerance, and innovation/follower strategies form a complex decision landscape. Aspects of these tradeoffs are illustrated in the context of a phenotypic angiogenesis project which identified potent, non-kinase targeted molecules which were active in vivo and were differentiated from standard of care.

11:05 High Throughput Flow Cytometry (HyperCyt) for Infectious Disease Targets

Larry A. Sklar, Regents Professor of Pathology and Distinguished University Professor; Co-Leader Program in Cancer Therapeutics, UNM Comprehensive Cancer Center; Director, Drug Rescue, Repurposing, and Repositioning Network, UNM CTSC; Director, University of New Mexico Center for Molecular Discovery; Associate Director and Maralyn S. Budke/Robert E. Anderson Endowed Chair in Cancer Drug Discovery; PI, UNM Center of Excellence NCI Experimental Therapeutics Chemical Biology Consortium

High throughput flow cytometry enables the screening of small molecules toward infectious or pathogenic targets including bacteria, fungi, and virus in suspension, as well as host pathogen interactions. We have conducted collaborative screens for: 1) TOR pathway analysis in yeast; 2) quorum sensing pathway analysis in MRSA; 3) proteome expression in yeast; 4) yeast two; 5) efflux transporter expression in yeast; 6) efflux transporter inhibition in yeast and fungi; 7) virus and bacteria interactions with host cells; 8) analysis of intracellular pH; 9) molecular assemblies relevant to botulinum protease inhibition, and proteasome molecular assembly in yeast. Examples of high content or phenotypic screening projects leading to the identification of small molecules such as 2) above will be presented.

11:35 Phenotypic Profiling of Intracellular Motion to Stratify Patient Chemosensitivity

David Nolte, Ph.D., Professor, Physics, Purdue University; President, Animated Dynamics, Inc.

Biodynamic screening of drug-response phenotypes, using living ex vivo biopsies, measures changes of intracellular dynamics in response to applied therapeutics. This talk will present the current status of several ongoing pre-clinical and clinical trials of biodynamic phenotypic profiling. Biodynamic profiling could help preselect qualified patients for improved clinical trial outcomes, and could help select the most efficacious therapies for cancer patients.

12:05 pm Luncheon Presentation (Sponsorship Opportunity Available) or Enjoy Lunch on Your Own

12:50 Session Break

1:35 Snatching HTS Performance from the Claws of Biological Complexity: Integrating Heterogeneity Indices with Multivariate Analysis to Quantitatively Characterize Zebrafish Transgene Morphology

Andreas Vogt, Ph.D., Associate Professor, Department of Computational and Systems Biology, University of Pittsburgh Drug Discovery Institute

By using the pixel-level distribution of transgene intensity as a proxy to cell-level data, we applied population-based intensity, shape, and heterogeneity measurements to quantitatively describe and characterize transgene expression in each embryo. Subsequent linear discriminant analysis on multiple discriminating features then captured and condensed this information into a single numerical value, resulting in HTS compatible assay performance in multi-day variability studies. The method is phenotype-agnostic and should be applicable to other complex and heterogeneous biological assay systems.

2:05 An Automated Method for Analyzing 4D High Content Imaging Data to Profile the Gene Set Controlling Embryonic Development

Rebecca Green, Ph.D., Research Scientist, Ludwig Institute for Cancer Research

We film embryogenesis, following targeted depletion, using two custom-engineered C. elegans strains: the first strain was designed to monitor changes in cell fate specification, by dynamically tracking fluorescently labeled endoderm, mesoderm and ectoderm nuclei, and the second strain was designed to track morphogenic changes during epithelial and neuronal development by monitoring changes in their tissue position and tissue shape. We evaluate phenotypic similarity by comparing the complex parameter profiles for each condition in n-dimensional space and develop a measure to correct for the observed non-uniformity in Euclidean distance. This automated method is highly effective in identifying groups of genes that yield similar phenotypes, suggesting that they function together in specific developmental pathways. This work represents the first fully automated high-content screen of an intact developing organism.

R. Khaliullin, S. D. Ochoa, Z. Zhao, S. Wang, J. Hendel, R. Biggs, A. Gerson-Gurwitz, A. Desai, K. Oegema* and R.A. Green*

2:35 Close of Conference

 

Arrive early and attend (Mon-Tues): High-Content Analysis