The field of genome editing is at the tip of the iceberg, where discovery of new nucleases, development of novel CRISPR applications in genome engineering, gene silencing, screening, disease modeling, therapeutics and epigenomics are occurring rapidly. Despite the immense potential of CRISPR, the technology still needs to overcome challenges in its precision and efficiency in delivery, as well as reducing off-target mutations.
Introduced at PEGS Boston for the first time, CRISPR for Genome Engineering invites experts to share their experiences on harnessing CRISPR for drug discovery and therapeutic applications, and to discuss strategies to overcome its technological challenges.
Final Agenda
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WEDNESDAY, MAY 2
7:30 am Registration(Commonwealth Hall) and Morning Coffee (Harbor Level)
8:30 Chairperson’s Remarks
John Feder, PhD, Associate Director, Genome Biology, Bristol-Myers Squibb
8:40 Cellular Engineering for Drug Discovery
Aaron T. Cheng, PhD, Head, PTS Discovery Genome Editing, Protein Cellular and Structural Sciences, R&D Platform Technology & Science, GlaxoSmithKline
This talk will highlight our application and learnings from use of CRISPR genome editing in building cellular systems for drug discovery.
9:10 Genome-Wide CRISPR Screens Identify New Genes and Pathways Driving Breast Cancer Development and Progression
Gus Frangou, PhD, Fellow, Molecular and Integrative Physiological Sciences, Harvard TH Chan School of Public Health
Identifying genetic drivers of metastatic breast cancer and the timing during which these lesions occur is critical to developing effective therapeutics. In this talk, novel modifications of CRISPR/Cas9 genome-editing technology we have developed, including high-efficiency in vivo phenotypic screens and inducible gene targeting, to interrogate the functions of cancer-driver mutations will be discussed. Significantly, these CRISPR/Cas9-based genetic screens provide a systematic phenotypic measurement of loss-of-function lesions in disease progression and provide novel insights into the molecular underpinnings of metastasis.
9:40 Dissecting Complex Biological Processes with Pooled CRISPR-Cas9 Screens
Patrick Collins, PhD, Senior Scientist, Genome Analysis Unit, Amgen
Pooled CRISPR screens have been shown by multiple groups to significantly outperform shRNA technology for identifying fitness genes. While every pooled screen inherently measures fitness over time, we can also apply selection(s) to study nearly any cell autonomous phenotype of interest. We will discuss the application of pooled CRISPR screens to study complex biological processes such as mechanisms of resistance and host factors modulating viral vector infection.
10:10 Coffee Break in the Exhibit Hall with Poster Viewing (Commonwealth Hall)
Tony Ho, MD, Executive Vice President, Head of R&D, CRISPR Therapeutics
I will be discussing the data that supports the first CRISPR-based cell therapy clinical trial for b-thalassemia, including selection of gRNA and evaluation of target events, efficacy of editing and pharmacodynamics effects. In addition, I will also discuss the use of CRISPR/Cas 9 in generating potent allogenic multi-edited CAR T therapeutics across variety of tumors.
11:25 Antibody-Display Libraries in Mammalian Cells Created Using CRISPR/Cas9 and TALE Nucleases
John McCafferty, PhD, CEO, Antibody Engineering, IONTAS, Ltd.
Using directed integration of antibody genes by CRISPR/Cas9 and TALE nucleases, we have constructed large libraries in mammalian cells containing a single antibody gene/cell. This has permitted construction of millions of monoclonal stable cell lines displaying IgG antibodies on their surface from which antibodies have been selected by flow cytometry for specificity, binding affinity, species cross-reactivity and expression level. Expression in production cell lines also enables high-throughput developability screening.
11:55 Genome-Scale Activation Screen Identifies a lncRNA Locus Regulating a Gene Neighborhood
Julia Joung, Graduate Student. Biological Engineering, MIT
Mammalian genomes contain thousands of loci that transcribe long non-coding RNAs (lncRNAs). Despite their potentially important roles, it remains challenging to identify functional lncRNA loci. Here, we developed a CRISPR–Cas9 activation screen targeting >10,000 lncRNA loci and found 11 that mediate melanoma drug resistance. Detailed analysis of one candidate revealed that its transcriptional activation resulted in dosage-dependent activation of four neighboring genes, one of which confers the resistance phenotype.
12:25 pm Poster Highlight: SMART Libraries and Phage-Induced Directed Evolution of Cas9 to Engineer Off-Target Activity
Barrett Steinberg, PhD, Scientist II, Protein Engineering, Editas Medicine
RNA-guided endonucleases such as Cas9 often show efficient editing in cells but can cleave at off-target loci in the genome. Engineered variants of S.pyogenes Cas9 (Sp.Cas9) have been developed to globally reduce off-target activity, but individual off-targets may remain or on-target activity may be compromised. In order to engineer decreased editing at specific off-targets and maintain on-target activity, we created a phage-based selection system in which we can negatively select for cleavage of specific sequence sin a competitive pool and positively select for cleavage at our desired on-target. Directed evolution using our system demonstrates a structure-independent methodology to effectively engineer nuclease activity.
12:55 Luncheon Presentation I: Build Better Biologics with Machine Learning and Synbio
Claes Gustafsson, Chief Commercial Officer, ATUM (formerly DNA2.0)
This presentation will showcase how ATUM combines recent developments in genome engineering, automation, big data and product analytics to increase efficiency of engineering and developability of biologics and cell lines. Cell lines generated using the LeapIn® transposase combined with optimized vector constructs, proprietary codon optimization and QSAR-based protein engineering allow for an information rich and efficient optimization of mAbs, bispecifics, CAR-T molecules, and the increasingly complex biologics approaching the market place.
1:25 Luncheon Presentation (Sponsorship Opportunity Available) or Enjoy Lunch on your Own
1:55 Session Break
2:10 Chairperson’s Remarks
Aaron T. Cheng, PhD, Head, PTS Discovery Genome Editing, Protein Cellular and Structural Sciences, R&D Platform Technology & Science, GlaxoSmithKline
2:15 Development and Optimization of CRISPR Gene Editing for Drug Discovery Applications
John Feder, PhD, Associate Director, Genome Biology, Bristol-Myers Squibb
2:45 Development of a Qualifiable NGS Assay for Precise and Accurate Quantitation of Small Insertions and Deletions (Indels) That Are Introduced in the Human Genome by Gene Editing
Marina Falaleeva, PhD, Scientist I, Development, Sangamo Therapeutics
Gene editing by targeted engineered nucleases in living cells is tracked by the detection of introduced indels (small insertions and deletions) at specific nucleotide sequences using Next-Generation Sequencing. Comparisons of nuclease efficiency and monitoring of results in patient samples requires the development of quantitative assays. The purpose of this talk is to discuss application of these established principles to the challenge of indel quantitation.
3:15 Panel Discussion:
Legal and Ethical Issues of Using Genome Editing:
Moderator: Colin A. Johnson, PhD, Professor of Medical & Molecular Genetics, Section of Ophthalmology and Neurosciences, Leeds Institute of Molecular Medicine
Panelists: Aaron T. Cheng, PhD, Head, PTS Discovery Genome Editing, Protein Cellular and Structural Sciences, R&D Platform Technology & Science, GlaxoSmithKline
- Modification of agricultural crops or livestock
- Germ-line editing of human embryos
- Somatic correction of oncogenic mutations in leukemia etc
- How does the technology interface with citizen science?
3:45 Refreshment Break in the Exhibit Hall with Poster Viewing (Commonwealth Hall)
4:45 Problem-Solving Breakout Discussions (Commonwealth Hall)
5:45 Networking Reception in the Exhibit Hall with Poster Viewing (Commonwealth Hall)
7:00 End of Day
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THURSDAY, MAY 3
8:00 am Registration(Commonwealth Hall) and Morning Coffee (Harbor Level)
8:30 Chairperson’s Remarks
Patrick Collins, PhD, Senior Scientist, Genome Analysis Unit, Amgen
8:35 Genome Editing to Model and Treat Cardiac and Neuromuscular Diseases
Chengzu Long, PhD, Asst Professor, Department of Medicine, New York University
Our projects focus on advancing the novel genome editing technology to model and treat cardiac and neuromuscular diseases. Duchenne muscular dystrophy (DMD) is a fatal muscle disease affecting 1 in 3,500 boys. Dilated cardiomyopathy (DCM) and heart failure are common and lethal consequences of DMD. We have advanced genome editing to cells from DMD patients by engineering the permanent skipping of mutant exons in the genomes of DMD patient-derived induced pluripotent stem cells (iPSCs).
9:05 Modelling the Pathogenic Effect of Mutations in Rare Mendelian Conditions
Colin A. Johnson, PhD, Professor of Medical & Molecular Genetics, Section of Ophthalmology and Neurosciences, Leeds Institute of Molecular Medicine
To gain novel unbiased insights into essential biological processes and to identify roles for unanticipated pathways in human genetic disease, we have developed our existing gene discovery research to include cellular disease modelling using CRISPR-Cas9 genome and base editing approaches. We will discuss recent advances in modelling ciliopathies, neuromuscular and neurodevelopmental conditions, and inherited retinal dystrophies.
9:35 Poster Highlight: Directed Evolution of CRISPR-Cas9 to Increase Its Specificity
Joonsun Lee, Researcher, Platform R&D, ToolGen, Inc.
We have developed a directed evolution approach to improve the specificity of SpCas9. By screening a library of random mutants of SpCas9 generated by error-prone PCR, a mutant named Sniper-Cas9 was identified, and it shows high specificities without sacrificing on-target activities in human cells. Sniper-Cas9 is fully compatible with extended or truncated sgRNAs and functions well in a preassembled ribonucleoprotein (RNP) form which facilitates DNA-free genome editing.
10:05 Coffee Break in the Exhibit Hall with Poster Viewing (Commonwealth Hall)
11:05 CRISPR-Cas9 Inactivation by Bacteriophage Proteins
Bettie Osuna, PhD, Graduate Student, Bondy-Denomy Lab, University of California, San Francisco
CRISPR-Cas adaptive immune systems defend prokaryotes from invasion by foreign genetic elements, including viruses (bacteriophages/phages). Phages have coevolved counter-attack strategies, including inactivation of CRISPR-Cas with “anti-CRISPR” proteins. We have discovered proteins encoded by Listeria monocytogenes phages, AcrIIA1-4, that suppress CRISPR-Cas9 activity. Our work aims to characterize anti-CRISPR-Cas9 mechanisms in this natural context to gain insight into CRISPR-Cas9 evolution and allow the use of anti-CRISPRs as “off-switches” in gene editing applications.
11:35 Structural Basis of CRISPR/Cas9 and Anti-CRISPR Mechanisms for Precise Genome Editing
Fuguo Jiang, PhD, Postdoc Fellow, Molecular and Cell Biology, University of California, Berkeley
CRISPR/Cas9 technology shows great promise in treating cancer diseases at a genetic level. The recently discovered natural Cas9-specific “anti-CRISPRs” present important tools that can be used to regulate CRISPR/Cas9-mediated genome editing specificity. I will talk about our structural studies on CRISPR/Cas9 and anti-CRISPR mechanisms, with the hope to support CRISPR/Cas9 site-specific genetic control and therapeutic applications.
12:05 pm Closing Q&A
12:35 End of CRISPR for Genome Engineering
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