Gene editing, particularly using the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 system, is now being extensively used as a research and functional screening tool in drug discovery. Cambridge Healthtech Institute’s third annual conference on CRISPR for Target Discovery will bring together experts from early target discovery to high-throughput functional screening, to talk about how CRISPR is being applied to unravel cellular pathways and identify potential targets for drug intervention. Learn about ways in which CRISPR/Cas9 is being used to create relevant cell lines and in vivo disease models, its use in functional screening, epigenome engineering, and for identifying and validating targets. What can you do to overcome some of the inherent challenges in specificity, efficiency, delivery and off-target effects, associated with CRISPR/Cas9? Hear from experts in pharma/biotech, academic and government labs who will share their experiences leveraging the utility of CRISPR-based gene editing for diverse applications.

Who should attend: Students, Post-Docs, Lab Technicians, Managers, Scientists, Clinicians, Team Leads, Directors and Executives from Pharma, Biotech, Academia, Government, Contract Research Labs and Technology Providers involved in Target Discovery, Disease Modeling, Functional Screening, High-Throughput Screening, Assay Development and Translational Research.

Final Agenda

Tuesday, June 13

7:00 am Registration Open and Morning Coffee

CRISPR-DERIVED CELL LINES AND KNOCK-OUTS FOR DISEASE MODELING

8:25 Chairperson’s Opening Remarks

Madhu Lal-Nag, Ph.D., Group Leader, Trans-NIH RNAi Facility, National Center for Advancing Translational Sciences, National Institutes of Health

8:35 Functional Genomics in Dissecting Skin Malignancy

Yejing Ge, Ph.D., Postdoctoral Fellow, Laboratory of Dr. Elaine Fuchs, Department of Mammalian Cell Biology and Development, Rockefeller University

Tissue stem cells govern tissue regeneration and wound-repair. Tumors often hijack normal cellular programs and exploit them for malignancy. Here I use CRIPSR combined with in utero lentiviral injection to knockout candidate genes specifically in the skin epidermis, and interrogate their functions during wounding and tumorigenesis. In doing so, I unravel stem cell plasticity as a molecular mechanism underlying “tumors are wounds that do not heal.”

9:05 Using CRISPR/Cas9 to Create a Collection of GFP-tagged Human iPSC Lines to Model Cell Organization and Dynamics

Ru Gunawardane, Ph.D., Director, Stem Cells and Gene Editing, Allen Institute for Cell Science

The Allen Institute for Cell Science is creating a dynamic visual model of hiPSC organization by utilizing the CRISPR/Cas9 system to endogenously label major cellular organelles with a fluorescent protein. We will present the methodologies used for endogenous gene tagging, screening for precise editing, and cell biological and genomic QC. We will also discuss the potential applications of these lines for basic science and disease modeling.

9:35 Modeling Human Pain Using CRISPR/Cas9 Genome Editing

Yung-Chih Cheng, Ph.D., Postdoctoral Research Fellow, F.M. Kirby Neurobiology Center, Boston Children’s Hospital, Department of Neurobiology, Harvard Medical School

Pain is a critical sensation allowing people to escape from damage. Scientists have discovered mutations in Nav1.7 channel leads to pathologies associated with exaggerated pain or insensitivity to pain. Here, we use CRISPR technology to generate pathological pain model in mouse and in hiPSC derived sensory neuron based on Nav1.7 channel. These engineered pain models enabled us to mimic the clinical aspects of the genetic forms of pain conditions and provide novel therapeutics for pain disorders in humans.

10:05 Grand Opening Coffee Break in the Exhibit Hall with Poster Viewing

10:50 Permanent Correction of Diverse Dilated Cardiomyopathy Mutations by Genome Editing

Chengzu Long, Ph.D., Assistant Professor, Division of Cardiology, New York University School of Medicine

Dilated cardiomyopathy (DCM) is one of the most common causes of heart failure, which affects over 38 million patients worldwide. Gene mutations are major causes of idiopathic DCM. To address several challenges for clinical applications of gene editing of DCM in vivo, we performed CRISPR/Cas9-mediated gene editing on representative iPSC (induced pluripotent stem cells)-derived cardiomyocytes from multiple DCM patients and restored their function.

11:20 Modeling Endothelial Dysfunction in LMNA-Related Dilated Cardiomyopathy

Nazish Sayed, M.D., Ph.D., Instructor, Cardiovascular Institute, Stanford University School of Medicine

The mechanisms that underlie “cardiolaminopathy” remain elusive. Although LMNA mutations are known to induce endothelial dysfunction, little is known about the EC-specific phenotype. Our data shows that iPSC-ECs derived from LMNA-mutated patients exhibit decreased EC functionality. Genome editing of iPSCs enabled us to recapitulate the EC-disease phenotype to dissect the effects of LMNA mutations. This study is a first step towards understanding cardiolaminopathy by modeling endothelial dysfunction.

11:50 Driver-Map™ Genome-Wide Expression Profiling Solution for Biomarker Discovery

Paul Diehl, Ph.D., COO, Cellecta, Inc.

Cellecta’s Driver-Map Genome-Wide Expression Profiling assay combines the sensitivity of multiplex PCR with the dynamic range of NGS. The approach achieves 100-fold more sensitivity than RNAseq over a greater dynamic range. Just 10 pg of total RNA shows over 5 orders of magnitude variation in gene expression levels. Applications include analysis of immune cell infiltration, identification of active pathways in tumor and xenograft samples, and profiling of biomarkers from blood.

12:20 pm Luncheon Presentation: High Throughput Screening: Best Technology and Practices

Andrew Ravanelli, Ph.D., Senior Research & Development Scientist, Genome & Epigenome Editing, MilliporeSigma

CRISPR Cas9 nucleases have revolutionized the field of gene editing and high-throughput lentiviral screens continue to hold ever-increasing promise for both basic research and development of future therapies to benefit human health. Even with such powerful technologies at hand, researchers new to the field may find screening of multiple targets to be challenging and time-consuming. MilliporeSigma seeks to share best approaches learned and methods applied over our years of genome editing experience. Here, we will detail the planning steps and workflow overview essential to a successful lentiviral screening experiment. We also present experimental data from the Sanger Wellcome Trust, first of its kind, genome-wide, truly arrayed guide RNA screening libraries for CRISPR-Cas9. Finally, we will compare screening technologies and describe multiple, flexible options for screening gene targets, from small gene panels to entire genomes.

12:50 Session Break

CRISPR FOR FUNCTIONAL AND PHENOTYPIC SCREENS

1:40 Chairperson’s Remarks

John Doench, Ph.D., Associate Director, Genetic Perturbation Platform, Broad Institute of Harvard and MIT

1:50 Pooled Screens with CRISPR Technology - Don’t Miss Out

John Doench, Ph.D., Associate Director, Genetic Perturbation Platform, Broad Institute of Harvard and MIT

The ease of programming Cas9 with an sgRNA presents an abundance of potential target sites, but the on-target activity and off-target effects of individual sgRNAs can vary. We will discuss improved models that allow for increased on-target efficacy, metrics for understanding potential off-target sites, and how the combination of these findings can be used to design optimal libraries for genetic screens.

2:20 Development of New CRISPR/Cas9-based Tools to Study Drug Interactions Through Knockout and Directed Evolution

Michael Bassik, Ph.D., Assistant Professor, Department of Genetics, Stanford University

We have compared genome-wide shRNA and CRISPR/Cas9 screens to identify novel drug targets, with highly complementary results. By systematic pairwise expression of sgRNAs directed against known drug targets, we identify rare synthetic lethal drug combinations for leukemia in ultra-high-throughput. Finally, we have developed a strategy to use dCas9 to recruit hyperactive AID*D, mutagenizing the endogenous target of a drug to map its target binding.

2:50 Arrayed High-Throughput Screening with Synthetic crRNA Libraries

Melissa Kelley, Ph.D., Senior Research & Development Leader, Dharmacon, part of GE Healthcare

In large-scale screening studies, one-well-per-gene arrayed synthetic crRNA libraries have an advantage over pooled CRISPR screens for endpoint assays and high-content imaging. We will describe an arrayed synthetic crRNA library screen in a Cas9-expressing cell cycle reporter cell line where we utilized high-content, multiparametric analysis. The experimental design, assay optimization and analysis employed for hit identification, stratification, and validation to successfully identify cell cycle regulation genes will be described.

3:20 Refreshment Break in the Exhibit Hall with Poster Viewing

4:05 Identification of Drug Response Modifiers Using Pooled CRISPR Screening

Roderick Beijersbergen, Ph.D., Group Leader, Netherlands Cancer Institute and Head, NKI Robotics and Screening Center

The CRISPR-Cas9 system has shown to be very efficient in disruption of genes allowing for the comprehensive identification of genes required for cell survival and proliferation. Extending this platform with tools to abrogate the expression of two or more genes simultaneously allows for the identification of genetic interactions associated with synthetic sick of synthetic lethal phenotypes. The identification of genetic interactions in the context of cancer specific genomic alterations will be presented.

4:35 High Efficiency Synthetic sgRNA for CRISPR

Kevin Holden, Ph.D., Head, Synthetic Biology, Synthego

Synthego demonstrates, through a collaborative effort with key researchers utilizing CRISPR, that synthetic 100-mer sgRNA produces consistent and superior genome editing in a variety of cell types including adherent mammalian cell lines, primary T-cells, iPSCs and model organism embryos. This technology will help to enable successful and consistent genome editing for both basic research models and for sensitive primary human cells to be used for therapeutic applications.

4:50 Technology Panel: Trends in CRISPR Technology and Applications

This panel will bring together 3-5 technical experts from leading technology and service companies to discuss trends and improvements in library design, assay reagents and platforms, and data analysis tools that users can expect to see soon to explore new applications.

Moderator: John Doench, Ph.D., Associate Director, Genetic Perturbation Platform, Broad Institute of Harvard and MIT

Participants: Paul Diehl, Ph.D., COO, Cellecta, Inc.

Kevin Holden, Ph.D., Head, Synthetic Biology, Synthego

Melissa Kelley, Ph.D., Senior Research & Development Leader, Dharmacon, part of GE Healthcare

Andrew Ravanelli, Ph.D., Senior Research & Development Scientist, Genome & Epigenome Editing, MilliporeSigma

Adrienne Watson, Ph.D., Senior Research Scientist, Surrogen, A Recombinetics Company

5:35 Welcome Reception in the Exhibit Hall with Poster Viewing

6:45 Close of Day

Wednesday, June 14

7:00 am Registration Open

7:30 Interactive Breakout Discussion Groups with Continental Breakfast

This session features various discussion groups that are led by a moderator/s who ensures focused conversations around the key issues listed. Attendees choose to join a specific group and the small, informal setting facilitates sharing of ideas and active networking. Continental breakfast is available for all participants.

Using CRISPR Edited iPS Cells for Drug Discovery
Moderators:
Chengzu Long, Ph.D., Assistant Professor, Division of Cardiology, New York University School of Medicine
Ru Gunawardane, Ph.D., Director, Stem Cells and Gene Editing, Allen Institute for Cell Science

• How to screen iPSC clones after genome editing (PCR-based, qPCR-based, drug-based etc.)
• How to characterize gene edited iPSC lines for high quality before using them for drug discovery/screening
• CRISPR diversity for genome editing - how many crRNAs to try and what criteria to use?

Using CRISPR to Create Good In Vitro and In Vivo Tumor Models: Challenges and Opportunities
Moderators:
Danilo Maddalo, Ph.D., Lab Head, ONC Pharmacology, Novartis Institutes for BioMedical Research, Novartis Pharma AG
Madhu Lal-Nag, Ph.D., Group Leader, Trans-NIH RNAi Facility, National Center for Advancing Translational Sciences, National Institutes of Health

• Limitation of CRISPR-based mouse models
• How can we improve CRISPR-based mouse models
• Future perspectives/integration of these models into the drug development pipeline

CRISPR FOR ONCOLOGY DRUG DISCOVERY

8:35 Chairperson’s Remarks

Danilo Maddalo, Ph.D., Lab Head, ONC Pharmacology, Novartis Institutes for BioMedical Research, Novartis Pharma AG

8:45 In vivo Generation of Oncogenic Signatures with the CRISPR/Cas9 System

Danilo Maddalo, Ph.D., Lab Head, ONC Pharmacology, Novartis Institutes for BioMedical Research, Novartis Pharma AG

Precise genomic editing leading to cancer formation represents a powerful tool in preclinical research. Generation of signatures resulting in cancer development/resistance can be investigated and identified by in vivo delivery of the CRISPR/Cas9 system. In this talk I will discuss the methods for generating preclinical animal models, the impact the ‘genome editing revolution’ has had, and the future applications for drug discovery and target identification.

9:15 Genome Editing and Gene Silencing in 3D Tumor Models: Opportunities for Overlap

Madhu Lal-Nag, Ph.D., Group Leader, Trans-NIH RNAi Facility, National Center for Advancing Translational Sciences, National Institutes of Health

The development of our 3D oncology RNAi and CRISPR/Cas9 screening platform is a step towards understanding the contribution of the tumor microenvironment to cancer cell viability. Our work aims to study and understand the role of tumor heterogeneity, clonal evolution, dormancy and cell death and in doing so, to bring forth some novel mechanisms of action that will help uncover novel druggable targets.

9:45 Engineered Swine Models of Cancer

Adrienne Watson, Ph.D., Senior Research Scientist, Surrogen, A Recombinetics Company

Huge advancements in technology to engineer genetically modified swine, who share immense genetic and physiological similarity to humans, have enabled the development of swine models of human cancer. We describe the latest innovations in cancer modeling in swine, including Recombinetics’ model of Neurofibromatosis Type 1, to show the benefits of using swine as a large animal model in research and the vast applications and opportunities of swine models of cancer.

10:15 Coffee Break in the Exhibit Hall with Poster Viewing

11:00 In vivo and ex vivo Cancer Engineering via Somatic Genome Editing

Peter Cook, Ph.D., Postdoctoral Researcher, Laboratory of Andrea Ventura, M.D., Ph.D., Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center

Using CRISPR-Cas9 genome editing, we have developed a strategy for modeling cancer-associated genomic rearrangements. An induced chromosomal deletion generating an uncharacterized receptor tyrosine kinase fusion transforms mouse adult neural stem cells, generating brain tumors that are highly similar to human high grade gliomas and respond to a specific kinase inhibitor. This technique holds promise for more accurate modeling of human tumors and targeted therapy development.

11:30 Knocking Out Specific miRNAs Using a Double CRISPR Approach Identifies Their Function During Development: Embryogenesis Recapitulating Tumorigenesis

 G. Ian Gallicano Ph.D., Associate Professor, Department of Biochemistry and Molecular & Cellular Biology, and Director, Transgenic Core Facility, Georgetown University Medical Center 

My laboratory investigates miRNA mechanisms during development. Knocking out individual miRNAs to study their function has been difficult using homologous recombination technology. However, CRISPR technology has solved this problem. We used a two CRISPR approach to remove individual miRNAs resulting in new embryonic stem cells lines. During development oncogenes are activated and subsequently deactivated, a process that goes awry in many cancers.  As a result, CRISPR technology used during development could shed light on understanding tumorigenesis.   

12:00 pm Bridging Luncheon Presentation (Sponsorship Opportunity Available) or Enjoy Lunch on Your Own

12:30 Session Break

1:00 Coffee and Dessert in the Exhibit Hall with Poster Viewing

1:30 PLENARY KEYNOTE SESSION

3:30 Refreshment Break in the Exhibit Hall with Poster Viewing

4:15 Close of Conference