Somatic Cell Genome Editing
Evolving High Potency AAV Vectors for Neuromuscular Genome Editing
Recombinant adeno-associated viruses (AAV) have emerged as safe and effective vectors for clinical gene therapy applications including systemic treatment of neuromuscular diseases such as Spinal Muscular Atrophy (SMA), Duchenne Muscular Dystrophy (DMD), and Giant Axonal Neuropathy (GAN) amongst others. However, genome editing in neuromuscular tissue, in particular, is challenging. We are investigating a comprehensive and innovative approach to evolve high potency AAV variants for systemic neuromuscular genome editing. Funding provided by NIH.
Epigenome Editing Technologies for Treating Diverse Disease
A recent revolution in DNA-targeting technologies has created new opportunities for treating human disease. While most effort has focused on using genome editing to alter DNA sequences to treat rare, inherited diseases, there is a tremendous opportunity to use these same tools to change the regulation of genes, rather than changing DNA sequence. This has the potential to extend the impact of these technologies to a greater diversity and number of diseases, and our work is addressing several key challenges to realizing this promise. Funding provided by NIH.
Development of Cell and Tissue Platforms to Detect Adverse Biological Consequences of Somatic Cell Genome Editing
Genome editing is a transformative technology that has the potential to treat diverse human diseases, but systems do not currently exist to monitor possible adverse consequences in human tissues. To evaluate potential undesirable consequences unique to genome editing tools, such as genome integrity, immune responses, and loss of therapeutic efficacy due to cell turn over, this project combines advanced tissue engineering approaches to systematically and quantitatively assess the effects of various genome editing methods. Effects are being assessed through measures of tissue physiology, function, and regeneration, as well as genome integrity and immune system responses in skeletal and cardiac muscle – tissues that are severely affected by many genetic diseases with significant unmet clinical need. Funding provided by the NIH Common Fund.
Regulation of functional plasticity in inflammation
Pro-inflammatory lymphocytes have tremendous plasticity and can undergo functional conversion into either pathogenic or regulatory/anti-inflammatory effector subclasses. Harnessing this attribute holds enormous potential to control the trajectory of inflammatory disease. Using a combination of genetic fate-mapping approaches in mice to track cellular transitions, and integrative regulatory network analyses, we have begun to identify the regulators that mediate such plastic switches (Carr T, et al, Nature Communications, 2017; Zuberbuehler MK et al, Nature Immunology, 2019). Funding provided by NIH/NIGMS.
Contribution of non-coding Genetic variation to autoimmune disease
Genome wide association (GWA) studies have identified numerous single nucleotide polymorphisms (SNP) in the non-coding genome that are associated with enhanced risk of disease. Major challenges include identifying the causal SNP in disease, its bone fide gene target, and the mechanism of dysregulation. We are addressing these challenges using a combination of global chromatin confirmation capture (Hi-C, Hi-ChIP), enhancer landscape profiling (ATAC-seq, ChIP-seq), and expression profiling of pathogenic T cells in patients with multiple sclerosis. Funding provided by the National Multiple Sclerosis Society.
Contribution of non-coding Genetic variation to schizophrenia
Similar to the autoimmune disease (see above) and other common disorders, GWA and whole exome sequencing studies for schizophrenia points to noncoding variation as contributing to disease risk. We have performed extensive chromatin accessibility mapping using ATAC-seq on 100's on postmortem brain samples from schizophrenia cases and controls (Bryois et al., Nature Neuroscience, 2018). While we do not detect chromatin accessibility differences between cases and controls, we do find that heritability enrichment for the disorder is much higher in the regulatory elements we have identified. To followup these studies, we are performing high-throughput STARR-seq reporter and high-throughput CRISPR genome editing screens to narrow down causal risk elements and variants. Funding provided by the National Institute of Mental Health, and Open Philanthropy.
Contribution of non-coding Genetic variation to rare recessive disorders
The majority of rare recessive disorders are due to pathogenic coding variants. However, there are many patients that display a clear phenotypic and biochemical diagnosis, but the genetic testing is inconclusive where only one pathogenic variant is detected on one allele. We have identified a number of patients with these types of inconclusive genetic testing, and will search for novel splicing, structural, and noncoding pathogenic variants. Funding provided by the National Human Genome Research Institute.
Engineering TEchnologies to DeterminE Causal Relationships between Chromatin Structure and Gene REgulation
The structure of the human genome inside of a living cell is precisely and dynamically controlled to determine the level of each gene in different cell types, in response to environmental stimuli, and in various disease conditions. The chemical modification and three-dimensional folding of our DNA plays an even greater role than our inherited genetics in human development, disease progression, and drug response. There have been tremendous advances in mapping genome sequence and structure but our understanding of the relationship between genome structure and function is relatively poor. The objective of this proposal is to cross interdisciplinary boundaries to develop the necessary technologies to accurately predict, quantitatively monitor, and deterministically program genome structure to generate improved disease models that will catalyze transformative drug development. The team will also develop educational programs to inspire the next generation of scientists in this emerging discipline. Funding provided by the National Science Foundation