CRISPR/Cas9 for Human Genome Engineering and Disease Research

Abstract

The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system, a versatile RNA-guided DNA targeting platform, has been revolutionizing our ability to modify, manipulate, and visualize the human genome, which greatly advances both biological research and therapeutics development. Here, we review the current development of CRISPR/Cas9 technologies for gene editing, transcription regulation, genome imaging, and epigenetic modification. We discuss the broad application of this system to the study of functional genomics, especially genome-wide genetic screening, and to therapeutics development, including establishing disease models, correcting defective genetic mutations, and treating diseases.

The 4D nucleome project

“…First, using CRISPR/Cas9 technologies DNA elements involved in specific chromatin structures, e.g. domain boundaries or chromatin loops, can be altered, re-located or deleted 57,58 . Second, defined chromatin structures such as chromatin loops will be engineered de novo by targeting proteins that can (inducibly) dimerize with their partner looping proteins (e.g.…”

Lessons from Enzyme Kinetics Reveal Specificity Principles for RNA-Guided Nucleases in RNA Interference and CRISPR-Based Genome Editing

“…RGN systems hold tremendous technological and therapeutic promise, and recent advances in increasing their delivery, stability, and specificity have markedly lowered the barriers to wide application of these potent and versatile tools (Haussecker and Kay, 2015; Hendel et al, 2015; Hu et al, 2016; Sullenger and Nair, 2016; Wittrup and Lieberman, 2015; Xiong et al, 2016). Here we approached the widespread problem of off-target effects.…”

“…RGNs have long been recognized as promising therapeutics because of their potential to specifically target mutated genes associated with disease, especially autosomal dominant disorders, and several RNAi-based therapies are currently in clinical trials (Sullenger and Nair, 2016; Wittrup and Lieberman, 2015; Xiong et al, 2016). Among potential applications, targeting single-nucleotide polymorphisms (SNPs), which are linked to hereditary diseases, such as Huntington’s disease, hypertrophic cardiomyopathy, and amyotrophic lateral sclerosis, has proven to be a particularly challenging problem because the mutated target loci or associated SNPs differ from the wild-type allele by only a single nucleotide.…”

CRISPR–Cas9 Structures and Mechanisms

“…Using single-guide RNA (sgRNA) libraries, CRISPRbased genome-wide screens can be leveraged to identify drug-target or disease-resistance genes, such as novel tumor suppressors or oncogenes, and to quickly assess drug targets (19,89). As such, CRISPR-Cas9-mediated genome engineering holds immense promise to treat or even cure genetic disorders, including many forms of cancer and neurodegeneration, as well as sickle cell anemia, cystic fibrosis, Duchenne muscular dystrophy, viral infections, immunological disorders, and cardiovascular diseases (4,30,62,95,107). Despite its advantages and great promise, there are some obstacles between CRISPR-Cas9 and its full therapeutic potential (16,21).…”