Advances in Gene Editing Technology: New Gene Therapy Options in Personalized Medicine
Curators: Stephen J Williams, PhD and Aviva Lev-Ari, PhD, RN
Recent Advances in Gene Editing Technology Adds New Therapeutic Potential for the Genomic Era
Author and Curator: Stephen J Williams, PhD
2.1.3.2 Advances in Gene Editing Technology: New Gene Therapy Options in Personalized Medicine, Volume 2 (Volume Two: Latest in Genomics Methodologies for Therapeutics: Gene Editing, NGS and BioInformatics, Simulations and the Genome Ontology), Part 2: CRISPR for Gene Editing and DNA Repair
The fundamental shift presently occurring within the medical field as well as our understanding of underlying biology has been brought on by revolutionary advances in the disciplines referred to as ‘OMICS’ (genomics, metabolomics, transcriptomics, proteomics). This paradigm shift has brought a new, more “personalized” mindset in investigating, treating, detecting, and policy-decision making disease as well as the physician-patient relationship. This Volume One of Genomics explains this paradigm shift as our classical understanding of the gene has evolved with rapid development of molecular technologies and high-end computational methods to a vision beyond the classic model. This new model involves big data to focus of the “code of OMIC signature”, moving from our investigational focus of “one gene at a time” to analysis of the changes in the networks of protein and gene expression occurring during disease progression.
Moving toward this promise of genome-based therapeutics has required the concomitant development of methodologies unavailable to the researcher and drug developer for most of the 20th century. These new technologies have allowed for the sequencing of the whole genome (advanced and inexpensive pyrosequencing), analyze the proteome for changes in post-translational modifications (new mass spectroscopy techniques combined with automated high-throughput gel electrophoresis on robotic platforms), ability to track all the changes happening to a patient’s metabolic profile (LC-MS in combination with an array of biocurated database functions), and develop new therapeutics based on discrete disease-specific changes in protein, enzyme, and DNA/RNA (mutational analysis, and advanced molecular techniques to allow for manipulation of DNA/RNA such as gene editing and therapeutic vectors) all advancements being dependent on the massive advancements in computing power and software development.
Although this final chapter on a specific technology (Cas9-mediated gene editing) might seem out of place to the reader for the subject of this Genomics volume, as discussed above, the development of these omics-related technologies have spurred the advent of personalized therapies. For example, in the 1990’s (as highlighted in the earlier chapters of this book) Dr. Craig Venter founded Celera Genomics with the goals of 1) sequencing the human genome in a cost effective manner (using new DNA sequencing technology and workflow he and colleagues had developed, and 2) use the information from whole genome sequencing to develop a new line of genomic-based therapeutics. Other companies such as Human Genome Sciences, Myriad Genetics, Seattle Genetics and recently new ventures from 23andMe and Google Ventures were also founded based on the promise that high-end sequencing information could directly lead to this new era of genome-based therapeutics. And although many in the medical field have felt that the primary goal of these companies, in particular using genomic analysis to enhance drug development has been a bit disappointing, AS IN ALL SCIENTIFIC AND MEDICAL DISCOVERY, which involves both SERENDIPITY and INDIRECT HAPPENSTANCE, three important breakthroughs, directly related to the development of a post-genomics era personalized medicine approach, resulted from the aforementioned efforts. These were:
- The detection of disease-specific mutations in exomes resulting in “druggable” protein targets and ability to define the respective drug-responsive patient cohorts
Chronic myelogenous (or myeloid or myelocytic) leukemia (CML) was one of the first cancers attributed to a specific chromosomal aberration, namely the translocation event resulting in a fusion protein between part of the BCR (“breakpoint cluster region”) gene from chromosome 22 with the ABL gene on chromosome 9. Early drug development efforts were directed against the tyrosine kinase activity of the aberrant BCR/ABL protein. The first of this new class of drugs was imatinib mesylate (Gleevec™) showed early success but was later noticed that a subset of patients had significantly greater response rates. This led to more detailed investigation of Gleevec’s mechanism of action and was determined that Gleevec’s therapeutic action depended on the drug’s ability to bind to an ATP binding pocket within the BCR/ABL. Patients with a specific mutation in this ATP pocket (C944T and T1052C) were found resistant to Gleevec. This finding, that pateint’s DNA could be sequenced to stratify them in responder versus nonresponder groups became a cornerstone for tyrosine kinase inhibitor (TKI) development for various cancers. One example is the development of crizotanib, a TKI directed against a mutant version of the anaplastic lymphoma kinase (ALK) enzyme, namely in patients carrying the ALK-EML4 fusion gene. As with Gleevec, certain mutations in the ATP binding pocket confer resistance to the inhibitory effects of crizotanib. Therefore, the Whole Exome Sequencing (WES) has shown its utility not only in drug development against cancer-specific mutant targets but stratifies patient cohorts into eligible versus non-eligible for a specific personalized therapy.
- Ability to define at-risk populations based on genomic data and development of corresponding genetic risk assessment for disease
Tremendous advances have been made in the area of risk-assessment for a plethora of diseases, including various malignancies, heart disease, and metabolic diseases. These risk factors have been identified given our advances in whole genome sequencing and proteomic and metabolomics. And, although the aforementioned companies had not developed therapeutic agents using these technologies, their major contribution has been the development of the diagnostic tests which identify at-risk patients and susceptible populations for a given disease. For example, the development of tests for carriers of the BRCA1/BRAC2 breast/ovarian cancer susceptibility mutation or APC (for colon cancer) has led to the appearance of Family Risk Assessment Programs and radically changed the discourse between patient and physician. And although determining risk factors to a disease such as cardiac disease in a large population can be fraught with complexities, the advanced research tools together with gene-directed technologies discussed in this Volume and current chapter may give better clarity in this regard. In essence, the technology had been developed well before its use in the clinic had been identified.
- Supplying and verifying linkages of specific genetic alterations to heritable diseases and offering a framework for future advances in gene-replacement and mutation-correction therapy
Our abilities to phenotypically correct inheritable diseases thru a gene-therapy (either by gene replacement or correction of mutated genes) have been hampered by three main areas. First identifying the specific mutations for a given inheritable disease used to be an arduous time-consuming process (linkage analysis), especially in small affected populations. However as whole exome sequencing rapidly evolved this had no longer become a rate-limiting step toward the development of a gene-directed therapy. Second and more troubling was determining a process which could deliver therapeutic genes in a safe, reliable and persistent manner. The first attempts at gene-therapy, relying on DNA virus and retroviral based delivery met with disaster and set back the field of gene therapy for decades (this story is too long for an introduction but for reference see the link.) Recently there have been improvements in therapeutic gene-therapy delivery systems such as the use of conditionally replicative adenovirus (cRADs) and novel serotype AAV (Recombinant adeno-associated virus, a nonpathogenic single stranded DNA human parvovirus) which have greatly improved safety and therapeutic profiles). The third issue, directly related to this chapter on Cas9-mediatied DNA editing) is the ability to integrate therapeutic DNA into the genome in a safe manner or correct mutations in their proper place. It is well established that the random integration of pieces of DNA has spurious effects on gene expression or contribute to transformation by an insertional mutagenesis mechanism.
This chapter will discuss how CRISPR/Cas9-mediated gene editing is being used in ex vivo strategies, namely to insert T-cell specific genes, in definable and safe loci, for the development of the new CAR-T cancer immuno-based therapies. In addition CRISPR/Cas9-mediated gene editing has much hope and promise for correcting specific mutations related to inheritable diseases, although investigations are at an infantile yet rapidly expanding area. As discussed above, new technologies have preceded their clinical use, mostly in a serendipitous and advantageous manner. Therefore it is a natural progression, using the concepts and curations in previous chapters, to investigate how a new technology, such as CRISPR/Cas9 medicated gene editing will fit into the ‘OMICS era of medicine.
Introduction
Larry H Bernstein, MD, FCAP
This document is a review and of the brilliant accomplishment of the Doudna Laboratory at University of California, Berkeley. It also traces the developments leading up to this groundbreaking work. The principle investigator is a young woman of significant accomplishments with the astounding publication of 4 papers at this time in 2015 and 20 in 2014. She
is a member of the National Academy of Sciences, and recipient of the Breakthrough Prize and the Lurie Prize in Biomedical Sciences, R. B. Woodward Visiting Professor, Harvard University (2000-2001). She achieved the Henry Ford II Professor of Molecular Biophysics and Biochemistry, Center for Structural Biology, Department of Molecular Biophysics and Biochemistry, Yale University (1994-2002) nine years after completion of her B.A. at Pomona College, and her Ph.D. under Jack Stozak at Harvard in 1989, became a Searle Scholar in 1996, and a Howard Hughes Investigator in 1997.
Her work has encompassed the editing of genes using the CRISPR-Cas9 system, and her team replaced a gene in a human cell which was convincing replicated in the Broad Laboratory at Harvard. The laboratory is currently working on the just reported immunological implications for CRISPR-Cas9 with respect to editing prokaryotic CRISPR-Cas genomic loci that encode RNA-mediated adaptive immune systems that bear some functional similarities with eukaryotic RNA interference. This is because acquired and heritable immunity against bacteriophage and plasmids begins with integration of ∼30 base pair foreign DNA sequences into the host genome.
Of special note are the following applications:
21.4.2 CRISPR: Applications for Autoimmune Diseases @UCSF
Reporter: Aviva Lev-Ari, PhD, RN
21.4.3 In vivo validated mRNAs
http://www.appliedstemcell.com/products/knock-in-cell-lines/in-vivo-crispr/
Doudna’s Interview from the National Academy of Science in 2004
Doudna discusses her current work with signal recognition particles, a type of RNA that is found in virtually all cell types and is responsible for directing specific proteins to specific membranes. She also discusses how advances in genomic sequencing may help catalog the complete range of functional RNA molecules. (9 minutes)
SOURCE
http://www.nasonline.org/news-and-multimedia/podcasts/interviews/jennifer-doudna.html
The Doudna lab pursues mechanistic understanding of fundamental biological processes involving RNA molecules. Research in the lab is currently focused on three major areas:
- bacterial immunity via the CRISPR system,
- RNA interference in eukaryotes, and
- translational control logic.
http://rna.berkeley.edu/crispr.html
http://rna.berkeley.edu/rnai.html
Different subunits are colored with invader RNAs in the background. Art by Gerard W.M. Staals
http://rna.berkeley.edu/translation.html
Alu-element regulated miRNA interactions
The Voice of Aviva Lev-Ari, PhD, RN
Big Pharma, CRISPR and Cancer
In January, the pharmaceutical giant Novartis announced that it would be using Doudna’s CRISPR technology for its research into cancer treatments. It plans to edit the genes of immune cells so that they will attack tumors.
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaJdJh5t
- The biggest biotech discovery of the century is about to change medicine forever
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaIwAbQU
Evolution of the Discovery
- In 1987, Yoshizumi Ishino and colleagues at Osaka University in Japan published the sequence of a gene called iap belonging to the gut microbe E. coli.
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaL6md3C
- In 2007, Blake Wiedenheft joined Doudna’s lab as a postdoctoral researcher, eager to study the structure of Cas enzymes to understand how they worked.
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaNXI2R4 - In a new paper in Nature Reviews Genetics, Koonin and Mart Krupovic of the Pasteur Institute in Paris argue that the CRISPR-Cas system got its start when mutations transformed casposons from enemies into friends.
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaN6Ucnk
- In January 2013, the scientists went one step further: They cut out a particular piece of DNA in human cells and replaced it with another one.
- In the same month, separate teams of scientists at Harvard University and the Broad Institute reported similar success with the gene-editing tool.
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaOF9xi4
- At Editas, a company based in Cambridge, Massachusetts, scientists have been investigating the Cas9 enzyme made by another species of bacteria, Staphylococcus aureus and Streptococcus pyogenes
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaMPy9Yw
- Writing last year in the journal Reproductive Biology and Endocrinology, Motoko Araki and Tetsuya Ishii of Hokkaido University in Japan predicted that doctors will be able to use CRISPR to alter the genes of human embryos “in the immediate future.”
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/#ixzz3UaOg9jyD
International regulatory landscape and integration of corrective genome editing into in vitro fertilization, Motoko Araki and Tetsuya Ishii*
http://www.rbej.com/content/12/1/108
The Human Germ Line
Don’t edit the human germ line
Heritable human genetic modifications pose serious risks, and the therapeutic benefits are tenuous, warn Edward Lanphier, Fyodor Urnov and colleagues.
The CRISPR technique has dramatically expanded research on genome editing. But we cannot imagine a situation in which its use in human embryos would offer a therapeutic benefit over existing and developing methods. It would be difficult to control exactly how many cells are modified. Increasing the dose of nuclease used would increase the likelihood that the mutated gene will be corrected, but also raise the risk of cuts being made elsewhere in the genome.
http://www.nature.com/news/don-t-edit-the-human-germ-line-1.17111
Engineering the Perfect Baby
Industry Body Calls for Gene-Editing Moratorium
Doudna’s Interview from the National Academy of Science in 2004
Track 6: Future Directions (follow link to track 6; requires RealPlayer)
Doudna discusses her current work with signal recognition particles, a type of RNA that is found in virtually all cell types and is responsible for directing specific proteins to specific membranes. She also discusses how advances in genomic sequencing may help catalog the complete range of functional RNA molecules. (9 minutes)
SOURCE
http://www.nasonline.org/news-and-multimedia/podcasts/interviews/jennifer-doudna.html
VIEW VIDEOS on Gene Editing
https://www.physicsforums.com/threads/breakthrough-prize-genome-editing-with-crispr-cas9.798959/
https://www.quantamagazine.org/20150206-crispr-dna-editor-bacteria/
The Structural Biology of CRISPR-Cas Systems
Curr Opin Struct Biol. 2015 Feb 24;30C:100-111
Authors: Jiang F, Doudna JA
Abstract
Prokaryotic CRISPR-Cas genomic loci encode RNA-mediated adaptive immune systems that bear some functional similarities with eukaryotic RNA interference. Acquired and heritable immunity against bacteriophage and plasmids begins with integration of ∼30 base pair foreign DNA sequences into the host genome. CRISPR-derived transcripts assemble with CRISPR-associated (Cas) proteins to target complementary nucleic acids for degradation. Here we review recent advances in the structural biology of these targeting complexes, with a focus on structural studies of the multisubunit Type I CRISPR RNA-guided surveillance and the Cas9 DNA endonuclease found in Type II CRISPR-Cas systems. These complexes have distinct structures that are each capable of site-specific double-stranded DNA binding and local helix unwinding.
PMID: 25723899 [PubMed – as supplied by publisher]
SOURCE
About CRISPR
“This technology will revolutionize biology in the same way PCR did,” Rudolf Jaenisch introducing Jennifer Doudna, 6/13/2014 @KI Symposium @MIT.
Top CRISPR Related Publications
http://blog.appliedstemcell.com/top-crispr-related-publications/
What is CRISPR? Why are Cas9-CRISPR services so popular?
http://blog.appliedstemcell.com/what-is-crispr-why-are-cas9-crispr-services-so-popular/
Custom Rat Model Generation Service Using CRISPR/Cas9
http://www.appliedstemcell.com/services/animal-models/
Jennifer A. Doudna
Dr. Jennifer Doudna is a member of the departments of Molecular and Cell Biology and Chemistry atUC Berkeley, the Howard Hughes Medical Institute, and Lawrence Berkeley National Lab, along with the National Academy of Sciences, and the American Academy of Arts and Sciences.
http://rna.berkeley.edu/people.html
AWARDS for the Discovery
Jennifer Doudna, cosmology teams named 2015 Breakthrough Prize winners
Jennifer Doudna, The winner of the 2014 Lurie Prize in the Biomedical Sciences
Doudna was a Searle Scholar and received a 1996 Beckman Young Investigators Award, the 1999 NAS Award for Initiatives in Research and the 2000 Alan T. Waterman Award. She was elected to the National Academy of Sciences in 2002 and to the Institute of Medicine in 2010. In 2014, Doudna was awarded the Lurie Prize in Biomedical Sciences from the Foundation for the National Institutes of Health as well as the Dr. Paul Janssen Award for Biomedical Researchand Breakthrough Prize in Life Sciences, both shared with Emanuelle Charpentier.
SOURCE
http://en.wikipedia.org/wiki/Jennifer_Doudna
21.1 Introducing CRISPR/Cas9 Gene Editing Technology – Works by Jennifer A. Doudna
21.1.1 Ribozymes and RNA Machines – Work of Jennifer A. Doudna Reporter: Aviva Lev-Ari Ph.D. RN
21.1.2 Evaluate your Cas9 gene editing vectors: CRISPR/Cas Mediated Genome Engineering – Is your CRISPR gRNA optimized for your cell lines? Reporter: Aviva Lev-Ari Ph.D. RN
Reporter: Aviva Lev-Ari Ph.D. RN
Curator: Aviva Lev-Ari, PhD, RN
21.2 CRISPR in Other Labs
21.2.1 CRISPR @MIT – Genome Surgery
21.2.2 The CRISPR-Cas9 System: A Powerful Tool for Genome Engineering and Regulation
Yongmin Yan and Department of Gastroenterology, Hepatology & Nutrition, University of Texas M.D. Anderson Cancer, Houston, USADaoyan Wei*
Reporter: Aviva Lev-Ari Ph.D. RN
Reporter: Aviva Lev-Ari Ph.D. RN
21.2.5 CRISPR & MAGE @ George Church’s Lab @ Harvard
Genome Engineering: CRISPR & MAGE
Multiplex Automated Genome Engineering (MAGE), is an intentionally broad term. In practice, it has come to be associated with a very efficient oligonucleotide allele-replacment (lambda red beta), so far restricted mainly to E.coli. CRISPR, in contrast, works in nearly every organism tested.
Relevant companies: EnEvolv, Egenesis, Editas.
News:
Editas (NextBigFuture, 28-Nov-2013, Brian Wang)
A Call to Fight Malaria One Mosquito at a Time by Altering DNA (NY Times, 17-Jul-2014, Carl Zimmer)
Resources:
* Vectors: Addgene
* Computational: Center for Causal Consequences of Variation (CCV)
Relevant Lab Publications:
2013 Probing the limits of genetic recoding in essential genes. Science.
2013 Genomically Recoded Organisms Impart New Biological Functions. Science.
2013 CAS9 transcriptional activators for target specificity screening and paired nickases for cooperative genome engineering. Nature Biotech.
2009 Programming cells by multiplex genome engineering and accelerated evolution. Nature.
SOURCE
http://arep.med.harvard.edu/gmc/B2.html
21.3 Patents Awarded and Pending for CRISPR
21.3.1 Litigation on the Way: Broad Institute Gets Patent on Revolutionary Gene-Editing Method
Reporter: Aviva Lev-Ari, PhD, RN
21.3.2 The Patents for CRISPR, the DNA editing technology as the Biggest Biotech Discovery of the Century
Reporter: Aviva Lev-Ari, PhD, RN
2.4 CRISPR/Cas9 Applications
Kennedy EM1, Kornepati AV1, Goldstein M2, Bogerd HP1, Poling BC1, Whisnant AW1, Kastan MB2, Cullen BR3.
21.4.2 CRISPR: Applications for Autoimmune Diseases @UCSF
Reporter: Aviva Lev-Ari, PhD, RN
21.4.3 In vivo validated mRNAs
http://www.appliedstemcell.com/products/knock-in-cell-lines/in-vivo-crispr/
Summary
Larry H Bernstein, MD, FCAP
CRISPR-Cas is a prokaryotic defense system against invading genetic elements. In a collaboration with John van der Oost’s laboratory, we are studying the structure and function of the effector complex of the Type III-A CRISPR-Cas system of Thermus thermophilus: the Csm complex (TtCsm). Recently, we showed that multiple Cas proteins and a crRNA guide assemble to recognize and cleave invader RNAs at multiple sites . Our negative stain EM structure of the TtCsm complex exhibits the characteristic architecture of Type I and Type III CRISPR-associated ribonucleoprotein complexes, suggesting a model for cleavage of the target RNA at periodic intervals (in collaboration with Eva Nogales, UC Berkeley, HHMI).
Double-stranded RNA induces potent and specific gene silencing in a broad range of eukaryotic organisms through a pathway known as RNA interference (RNAi). RNAi begins with the processing of endogenous or introduced precursor RNA into micro-RNAs (miRNAs) and small interfering RNAs (siRNAs) 21-25 nucleotides in length by the enzyme Dicer. We previously determined the crystal structure of an intact Dicer enzyme, revealing how Dicer functions as a molecular ruler to measure and cleave duplex RNAs of a specific length. Current work focuses on the mechanism of a complex of proteins known as the RISC loading complex (RLC) which load miRNA into the endonuclease Argonaute. The RLC contains the enzyme Dicer as well as TRBP, an RNA-binding protein hypothesized to interact with miRNA and Dicer during RISC loading. We seek to determine the molecular underpinnings of these interactions, along with the role of TRBP in RISC loading.
MicroRNAs (miRNAs) regulate endogenous eukaryotic genes by repressing gene expression through direct base-pairing interactions with their target messenger RNAs (mRNAs). To date, the rules used to predict miRNA-mRNA interactions have been based on one-dimensional sequence analysis. A more complete picture of miRNA-mRNA interactions should take into account the ability of RNA to form two- and three-dimensional structures. We are investigating the role of mRNA structure in the efficiency and specificity of targeting by miRNAs. Specifically, we are investigating the structure of Alu elements found within the 3′ untranslated regions (UTRs) of many human mRNAs and whether these structured domains serve as targets of a subset of human miRNAs. We are using in vitro biochemical methods and cell-based assays to probe the relationship between miRNA binding and mRNA structure.
The 5’ UTR of mRNA is also the site of multiple regulatory mechanisms, including upstream open reading frames (uORFs), internal ribosome entry sites (IRESs), protein binding sites, and stable secondary structures. Genes that profoundly influence cellular state often are controlled by multiple of these regulatory mechanisms. We are attempting to further understand regulatory elements in the 5′ UTR of mammalian mRNA using a combination of in vitro, cell-based and high-throughput techniques.
What does this mean for the development of therapeutics in the near future?
New methods for programming cell phenotype have broadly enabled drug screening, disease modeling, and regenerative medicine. Current research explores genome engineering tools, such as CRISPR/Cas9-based gene regulation and epigenome editing, to more precisely reprogram gene networks and control cellular decision making.
Donald Zack, M.D., Ph.D., Associate Professor of Ophthalmology and Neuroscience, Johns Hopkins University School of Medicine, is using CRISPR/Cas9 technology to generate retinal cell type-specific reporter ES and iPS lines and to introduce retinal degeneration-associated mutations. These reporter lines can be used to follow retinal neuronal specification during differentiation, they allow the purification of specific cell types by sorting and immunopanning, and they also are useful for the development of drug screening assays.
Jacquin C. Niles, M.D., Ph.D., Associate Professor of Biological Engineering, Massachusetts Institute of Technology is using CRISPR-Cas9 technology to study functional genetics in the human malaria parasite, Plasmodium falciparum. The team has established strategies for achieving controllable gene expression, and has integrated these into an experimental framework that facilitates efficient interrogation of virtually any target parasite gene using CRISPR/Cas9 editing.
Sidi Chen, Ph.D., Postdoctoral Fellow, Laboratory of Dr. Feng Zhang, Broad Institute and the Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology is observing that cancer genomics has revealed hundreds to thousands of mutations associated with human cancer. To test the roles of these mutations, we applied CRISPR/Cas9-mediated genome editing platform to engineer specific mutations in oncogenes and tumor suppressor genes. This results in tumorigenesis in several internal organs in mice. Our method expedites modeling of multigenic cancer with virtually any combination of mutations.
Samuel Hasson, Ph.D., Principal Investigator, Neuroscience, Pfizer, Inc., observes that while RNAi-based functional genomics is a staple of gene pathway and drug target exploration, there is a need for tools to provide rapid orthogonal validation of gene candidates that emerge from RNAi campaigns. CRISPR, CRISPRi, and CRISPRa are not only developing into primary screening platforms, they are a promising method to compliment RNAi and enhance the quality of functional genomic datasets.
These are some of the developments that will be discussed in detail at an upcoming meeting in Boston, MA in June titled ‘Gene Editing for Drug Discovery’.
SOURCE
http://rna.berkeley.edu/crispr.html
http://rna.berkeley.edu/rnai.html
http://rna.berkeley.edu/translation.html
Doudna Lab Publications
http://rna.berkeley.edu/publications.html
The structural biology of CRISPR-Cas systems.
Jiang F, Doudna JA
Curr Opin Struct Biol 2015 Feb 24;30C:100-111
Rational design of a split-Cas9 enzyme complex.
Wright AV, Sternberg SH, Taylor DW, Staahl BT, Bardales JA, Kornfeld JE, Doudna JA
Proc Natl Acad Sci U S A 2015 Feb 23
Genomic Engineering and the Future of Medicine.
Doudna JA
JAMA 2015 Feb 24;313(8):791-792
Integrase-mediated spacer acquisition during CRISPR-Cas adaptive immunity.
Nuñez JK, Lee AS, Engelman A, Doudna JA
Nature 2015 Feb 18
Dicer-TRBP Complex Formation Ensures Accurate Mammalian MicroRNA Biogenesis.
Wilson RC, Tambe A, Kidwell MA, Noland CL, Schneider CP, Doudna JA
Mol Cell 2014 Dec 30
Enhanced homology-directed human genome engineering by controlled timing of CRISPR/Cas9 delivery.
Lin S, Staahl B, Alla RK, Doudna JA
Elife 2014 Dec 15;3
Cutting it close: CRISPR-associated endoribonuclease structure and function.
Hochstrasser ML, Doudna JA
Trends Biochem Sci 2014 Nov 18
RNA Targeting by the Type III-A CRISPR-Cas Csm Complex of Thermus thermophilus.
Staals RH, Zhu Y, Taylor DW, Kornfeld JE, Sharma K, Barendregt A, Koehorst JJ, Vlot M, Neupane N, Varossieau K, Sakamoto K, Suzuki T, Dohmae N, Yokoyama S, Schaap PJ, Urlaub H, Heck AJ, Nogales E, Doudna JA, Shinkai A, van der Oost J
Mol Cell 2014 Nov 20;56(4):518-530
Genome editing. The new frontier of genome engineering with CRISPR-Cas9. (Free Full Text)
Doudna JA, Charpentier E
Science 2014 Nov 28;346(6213):1258096
Doudna JA, Sontheimer EJ
Methods Enzymol 2014;546C:xix-xx
New tools provide a second look at HDV ribozyme structure, dynamics and cleavage.
Kapral GJ, Jain S, Noeske J, Doudna JA, Richardson DC, Richardson JS
Nucleic Acids Res 2014 Oct 17
Programmable RNA recognition and cleavage by CRISPR/Cas9.
O’Connell MR, Oakes BL, Sternberg SH, East-Seletsky A, Kaplan M, Doudna JA
Nature 2014 Sep 28
RNA-guided assembly of Rev-RRE nuclear export complexes.
Bai Y, Tambe A, Zhou K, Doudna JA
Elife 2014;3:e03656
Evolutionarily Conserved Roles of the Dicer Helicase Domain in Regulating RNAi Processing.
Kidwell MA, Chan JM, Doudna JA
J Biol Chem 2014 Aug 18
Structure-Guided Reprogramming of Human cGAS Dinucleotide Linkage Specificity.
Kranzusch PJ, Lee AS, Wilson SC, Solovykh MS, Vance RE, Berger JM, Doudna JA
Cell 2014 Aug 12
Insights into RNA structure and function from genome-wide studies.
Mortimer SA, Kidwell MA, Doudna JA
Nat Rev Genet 2014 May 13
Cas1-Cas2 complex formation mediates spacer acquisition during CRISPR-Cas adaptive immunity.
Nuñez JK, Kranzusch PJ, Noeske J, Wright AV, Davies CW, Doudna JA
Nat Struct Mol Biol 2014 May 4
CasA mediates Cas3-catalyzed target degradation during CRISPR RNA-guided interference.
Hochstrasser ML, Taylor DW, Bhat P, Guegler CK, Sternberg SH, Nogales E, Doudna JA
Proc Natl Acad Sci U S A 2014 Apr 18
Structures of Cas9 Endonucleases Reveal RNA-Mediated Conformational Activation.
Jinek M, Jiang F, Taylor DW, Sternberg SH, Kaya E, Ma E, Anders C, Hauer M, Zhou K, Lin S, Kaplan M, Iavarone AT, Charpentier E, Nogales E, Doudna JA
Science 2014 Feb 6
DNA interrogation by the CRISPR RNA-guided endonuclease Cas9.
Sternberg SH, Redding S, Jinek M, Greene EC, Doudna JA
Nature 2014 Jan 29
http://rna.berkeley.edu/fun/D-lab%202013/SMALL/crisprconf.jpg
http://www.nasonline.org/news-and-multimedia/podcasts/interviews/jennifer-doudna.html
The Doudna lab pursues mechanistic understanding of fundamental biological processes involving RNA molecules. Research in the lab is currently focused on three major areas:
- bacterial immunity via the CRISPR system,
- RNA interference in eukaryotes, and
- translational control logic.
http://rna.berkeley.edu/crispr.html
http://rna.berkeley.edu/rnai.html
http://rna.berkeley.edu/translation.html
Other related curation on Gene Editing published as Chapter 21 in
Genomics Orientations for Individualized Medicine
- Advances in Gene Editing Technology: New Gene Therapy Options in Personalized Medicine
Curators: Stephen J Williams, PhD and Aviva Lev-Ari, PhD, RN
- Recent Advances in Gene Editing Technology Adds New Therapeutic Potential for the Genomic Era
Author and Curator: Stephen J Williams, PhD
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