Gene editing, particularly the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 system, is now being extensively used in drug discovery for disease modeling, functional screening, target identification and more. Cambridge Healthtech Institute’s conference on CRISPR for Disease Modeling and Target Discovery will bring together experts, from target discovery to functional screening, to talk about how CRISPR is being used to unravel cellular pathways in disease and identify potential targets for drug intervention. This is a unique opportunity to hear from experts in pharma/biotech, academic and government labs specifically about their experiences leveraging the utility of CRISPR-based gene editing to create relevant cell lines, knock-outs and in vivo tools for modeling diseases, identifying and validating targets, functional screening, epigenome engineering, and more. It’s also an opportunity that brings together users and solution providers to talk about ways to overcome some of the inherent challenges in specificity, efficiency, delivery and off-target effects, associated with CRISPR/Cas9.

Final Agenda

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Tuesday, September 26

7:00 am Registration Open and Morning Coffee

CRISPR-Based Functional & Phenotypic Screens

8:00 Welcome Remarks

Tanuja Koppal, Ph.D., Conference Director, Cambridge Healthtech Institute

8:05 Chairperson’s Opening Remarks

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

8:10 Pooled Screening with CRISPR-Cas9: Where Are We Now?

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.

8:40 Large-scale CRISPR screening: Mining for the Deep

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

Large-scale CRISPR-Cas9 screening across many different cell lines provides the opportunity to identify gene-gene correlations based on similar phenotypic profiles. Such analyses identify known interactions and biological networks but also discover novel associations. In particular these interactions can be explored for novel therapeutic strategies for cancer.  The results of such efforts will be discussed.  

9:10 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 used parallel shRNA and CRISPR screening to explore the biology of essential and non-essential genes, and have identified the target and mechanism of action of a novel host-targeting antiviral drug. More recently, we have used pairwise expression of sgRNAs to identify synergistic combinations of drug targets, and adapted our screening systems for new applications in mutagenesis and directed evolution.

9:40 Grand Opening Coffee Break in the Exhibit Hall with Poster Viewing

10:25 Efficient CRISPR Screen Design and Analysis

Xiaole Shirley Liu, Ph.D., Professor, Department of Biostatistics and Computational Biology, Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute and Harvard School of Public Health

Genome-wide CRISPR-Cas9 screen has been widely used to interrogate gene functions. However, the analysis remains challenging and rules to design better libraries beg further refinement. I will discuss methods we have developed for the efficient design and analysis of CRISPR screens. I will also discuss our analysis of CRISPR screens comparing cells with and without drug treatment to understand the drug mechanism of action.

10:55 TECHNOLOGY PANEL DISCUSSION: Trends in CRISPR Technology and Applications

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

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

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

Xiaole Shirley Liu, Ph.D., Professor, Department of Biostatistics and Computational Biology, Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute and Harvard School of Public Health

David Piper, Ph.D., Research & Development Leader, Cell Biology, Thermo Fisher Scientific

Shawn Shafer, Ph.D., Director of Advanced Genomics, MilliporeSigma

Paul Diehl, Ph.D., COO, Cellecta

Abhishek Saharia, Ph.D., Director, Product Management, Synthego

This panel will bring together speakers and 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.

11:55 Functional Genomics Screening with Invitrogen™ LentiArray™ CRISPR Libraries

David Piper, Ph.D., Research & Development Leader, Cell Biology, Thermo Fisher Scientific

Investigating pathways and validating gene functions and molecular targets that underlie disease relevant biological processes remain to be a challenge in drug discovery. The CRISPR-Cas9 system provides an efficient method for specific, complete, and permanent gene knockout and is a potent tool for making new discoveries and identifying gene function. The Invitrogen™ LentiArray™ CRISPR library collections harnesses this capability and expands it into high throughput applications to create the next generation of tools for functional genomics screening thereby accelerating therapeutic research and biomarker discoveries. Described here are the various ready to use predefined CRISPR library collections and custom service options available through Thermo Fisher Scientific along with example application data to demonstrate gene targeting efficacy and screening workflows. We have demonstrated here a knock-out screening approach that utilizes the Invitrogen™ LentiArray™ CRISPR library to interrogate the impact of individual gene knock-outs on the NFκB pathway as measured by a functional cell-based reporter assay using our CellSensor™ NF-κB-bla ME180 cell line. We describe the library design concepts, assay development, initial screening results and validation of specific identified hits. Together with our CRISPR library and service offering we expect these approaches to be scalable to the entire human genome and portable to multiple cell types and end-point assays including both high-throughput plate-based assays and high-content imaging based assays.

12:25 pm Session Break

 12:35 Luncheon Presentation: Going the Extra Mile: Advancing Screening with Lentiviral CRISPR and RNAi Libraries

Suzanne Hibbs, MBA, Study Coordination Manager, Cell Design Studio, MilliporeSigma

CRISPR Cas9 nucleases have revolutionized the field of gene editing and high-throughput screens for target identification. MilliporeSigma seeks to share best approaches utilized and methods learned from our years of genome editing experience. In collaboration with the Wellcome Trust Sanger Institute, we have developed the first commercially available, genome-wide, truly arrayed guide RNA CRISPR-Cas9 lentiviral screening library. We will compare complementary screening technologies, such as RNAi, from small gene pathways to entire genomes.

1:15 Refreshment Break in the Exhibit Hall with Poster Viewing

Unraveling Disease Pathways Using CRISPR/Cas9

1:50 Chairperson’s Remarks

James Inglese, Ph.D., Head, Assay Development & Screening Technologies, National Center for Advancing Translational Sciences, NIH

1:55 Pooled CRISPR-Cas9 Screens for Host Factors Modulating AAV and HSV Infection

Patrick Collins, Ph.D., Senior Scientist, Genome Analysis Unit, Amgen

Adeno-associated virus and herpes simplex virus are vectors for two approved, virus-mediated therapies. To identify host factors modulating infection by these two vectors, we performed genome-wide screens using a pooled CRISPR library in U-2 OS Cas9 stable cells. We then infected with AAV or HSV and selected for cells with altered GFP transgene expression or cells resistant to HSV-mediated lysis. We will detail our approach and compare the results of our screens to other efforts aimed at identifying host factors for these vectors.

2:25 CRISPR-Cas9 Editing of Herpes Simplex Virus Genomes during Lytic and Latent Infection

Hyung Suk Oh, Ph.D., Research Associate, Laboratory of David Knipe, Ph.D., Department of Microbiology and Immunobiology, Harvard Medical School

There is a great medical need to cure latent viral infections, and CRISPR-Cas9 is a potential approach to edit and inactivate latent viral DNA genomes. We have used CRISPR-Cas9 technology to edit lytic herpes simplex virus genomes and reduce lytic infection. In addition, we have employed CRISPR-Cas9 to edit quiescent viral genomes in cell culture. Studies are underway to target latent infection in murine systems.

2:55 CRISPR/Cas9 Genome-Wide sgRNA Libraries for More Effective Genetic Screens

Paul Diehl, Ph.D., COO, Cellecta

Genome-wide loss-of-function screening is a fundamental method to identify genes responsible for driving biological processes. Complex pooled lentiviral-based libraries expressing large numbers of genetic disruptors, such as shRNAs (RNAi) or sgRNAs (CRISPR), make large-scale cell screening practical. CRISPR-based technologies offer not only an effective alternative, but distinct advantage.

 3:10 Conducting Functional Genomic CRISPR Screens with Arrayed Chemically Modified Synthetic sgRNA Libraries

Abhishek Saharia, Ph.D., Director, Product Management, Synthego

We have developed an arrayed library of chemically modified synthetic sgRNA that enables rapid screening of complex phenotypes in almost all human cells. Using this arrayed and multiplexed library, we demonstrate highly effective gene editing and knockout rates (up to 98%), highlighting the value of this arrayed CRISPR screening tool.

3:25 Refreshment Break in the Exhibit Hall with Poster Viewing and Poster Competition Winner Announced

4:05 Identification of Genetic Modifiers of Somatic CAG Instability in Huntington’s Disease by in vivo CRISPR/Cas9 Genome Editing

 Vanessa Wheeler, Ph.D., Associate Professor of Neurology, Center for Genomic Medicine, Massachusetts General Hospital and Harvard Medical School

A recent genome-wide association study identified DNA repair genes as modifiers of HD age at onset. These studies implicate the CAG repeat expansion process as a disease driver and highlight the  potential therapeutic impact of targeting these genes. To this end, we are using the CRISPR-Cas9 toolbox in HD mouse models, to dissect underlying disease mechanisms by determining if these genes modify CAG repeat expansion and early disease phenotypes in the mouse.

4:35 Application of Genome Editing to Develop HTS Assays for Rare and Neglected Disease Drug Discovery

James Inglese, Ph.D., Head, Assay Development & Screening Technologies, National Center for Advancing Translational Sciences, NIH

Dravet Syndrome is a convulsive disorder caused by heterozygous loss-of-function mutations in the voltage-gated sodium channel alpha-subunit encoded by the SCN1A gene. Here we describe a quantitative high-throughput screening (qHTS) assay engineered from the Neuro2a cell line in which one of the two SCN1A alleles has been replaced by a bioluminescence coincidence reporter using CRISPR/Cas9-medieated homologous recombination. Characterization of the assay and preliminary small molecule screening results will be presented.

5:05 Interactive Breakout Discussion Groups

Join a breakout discussion group. These are informal, moderated discussions with brainstorming and interactive problem solving, allowing participants from diverse backgrounds to exchange ideas and experiences and develop future collaborations around a focused topic. Details on the topics and moderators are below. Please click here for full details on all breakouts.

CRISPR/Cas9 for Drug Discovery Applications

Moderators: James Inglese, Ph.D., Head, Assay Development & Screening Technologies, National Center for Advancing Translational Sciences, NIH

Patrick Collins, Ph.D., Senior Scientist, Genome Analysis Unit, Amgen

• Impact of CRISPR/Cas9 for drug discovery in pharma and academia
• Applications for functional screens, creating cell lines and disease models
• Technical limitations
• CRISPR delivery

Applications for Pooled CRISPR/Cas9 Screening: What Works and What Doesn’t

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

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

• Best practices on setting up high-throughput CRISPR screens
• Data analysis and validation; use of complementary methods
• Insights on inherent challenges and ways to overcome it

Setting Up Pooled and Arrayed CRISPR Screens: What You Need to Know Before and After

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

• Challenges and solutions for CRISPR gRNA design
• Methods for detecting engineered changes
• Evaluating and testing the reagents and libraries
• Data analysis

6:05 Welcome Reception in the Exhibit Hall (Sponsorship Opportunity Available)

7:10 Close of Day

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Wednesday, September 27

7:30 am Registration Open and Morning Coffee

CRISPR for Target Identification

8:00 Chairperson’s Remarks

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

8:05 Validating and Invalidating Cancer Genetic Dependencies with CRISPR/Cas9

Jason Sheltzer, Ph.D., Principal Investigator, Cold Spring Harbor Laboratory

We have investigated putative genetic dependencies in triple-negative breast cancer using CRISPR/Cas9-mediated genome editing. In certain cases, on-target mutagenesis cast doubt on previously-reported cancer cell addictions. We focus on MELK and OTS167, a MELK inhibitor currently in clinical trials, and report that breast cancer cells tolerate the complete loss of MELK with no apparent fitness defect. OTS167 remains active against MELK-knockout clones, demonstrating that it necessarily kills cells through an off-target effect.

8:35 CRISPR/Cas9: A Functional Screening of the Kinome Reveals a New Potential Treatment for an Aggressive Pediatric Brain Tumor

Simone Treiger Sredni, M.D., Ph.D., Associate Professor of Pediatric Neurosurgery, Ann and Robert H. Lurie Children’s Hospital of Chicago and Northwestern University, Feinberg School of Medicine

We have used a lentiviral CRISPR library to perform a systematic functional screening of the kinome by editing 160 kinases in a highly malignant pediatric brain tumor called atypical teratoid/rhabdoid tumor (AT/RT). We found that the polo-like kinase 4 (PLK-4) is essential for tumor survival, growth and migration. More importantly, we found that cells respond to PLK-4 small molecule inhibitors. This is the first time PLK-4 has been described in brain or pediatric tumors.

9:05 An IL-17 Signaling Phenotypic Screen Using the Orthogonal Methods of CRISPR and Small Molecule Libraries to Identify New Targets

Peter Slivka, Ph.D., Senior Scientist, Foundational Immunology, AbbVie Bioresearch Center

IL17 plays a critical role in the development and maintenance of autoimmune diseases including psoriasis and psoriatic arthritis. To identify novel targets in the IL17 signaling cascade, we conducted two phenotypic screens in primary keratinocytes using IL17 as a stimulus. The first screen utilized a pooled CRISPR library and the second utilized an annotated small molecule library. This talk will compare the results from the two screens and highlight a novel target of interest identified by both screens.

9:35 Coffee Break in the Exhibit Hall with Poster Viewing

10:20 Nucleic Acid Detection with RNA-Guided RNA Targeting CRISPR Cas13

Jonathan Gootenberg, Ph.D. Student, Laboratory of Dr. Feng Zhang, Broad Institute of MIT and Harvard

Rapid, inexpensive, and sensitive nucleic acid detection may aid point-of-care pathogen detection, genotyping, and disease monitoring. We combine the RNA-targeting CRISPR effector Cas13a with isothermal amplification to establish a CRISPR-based diagnostic (CRISPR-Dx), providing rapid DNA or RNA detection with attomolar sensitivity and single-base mismatch specificity. We use this Cas13a-based molecular detection platform, termed SHERLOCK, to detect specific strains of Zika and Dengue virus, distinguish pathogenic bacteria, genotype human DNA, and identify cell-free tumor DNA mutations.

10:50 Harnessing RNA Targeting CRISPR Systems for Transcriptome Engineering and Human Health

Omar Abudayyeh, M.D., Ph.D. Student, Harvard-MIT Health Sciences and Technology, Laboratory of Dr. Feng Zhang, Broad Institute of MIT and Harvard

RNA plays important and diverse roles in biology, but molecular tools to manipulate and measure RNA are limited. Here, we demonstrate that RNA-targeting CRISPR effector Cas13a can be engineered for mammalian cell RNA knockdown and binding. LwCas13a can be heterologously expressed in mammalian and plant cells for targeted knockdown of either reporter or endogenous transcripts and targeted RNA binding for transcript imaging. Our results establish CRISPR-Cas13a as a flexible platform for RNA targeting with wide applicability for studying RNA in mammalian cells.

11:20 Enjoy Lunch on Your Own

12:35 pm Plenary Keynote Program

(click here for details)

2:00 Refreshment Break in the Exhibit Hall with Poster Viewing

2:45 Close of Conference

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