gene editing, the ability to make highly specific changes in the DNA sequence of a living organism, essentially customizing its genetic makeup. Gene editing is performed using enzymes, particularly nucleases that have been engineered to target a specific DNA sequence, where they introduce cuts into the DNA strands, enabling the removal of existing DNA and the insertion of replacement DNA. Key among gene-editing technologies is a molecular tool known as CRISPR-Cas9, a powerful technology discovered in 2012 by American scientist Jennifer Doudna, French scientist Emmanuelle Charpentier, and colleagues and refined by American scientist Feng Zhang and colleagues. CRISPR-Cas9 functioned with precision, allowing researchers to remove and insert DNA in the desired locations.
The significant leap in gene-editing tools brought new urgency to long-standing discussions about the ethical and social implications surrounding the genetic engineering of humans. Many questions, such as whether genetic engineering should be used to treat human disease or to alter traits such as beauty or intelligence, had been asked in one form or another for decades. With the introduction of facile and efficient gene-editing technologies, particularly CRISPR-Cas9, however, those questions were no longer theoretical, and the answers to them stood to have very real impacts on medicine and society.
The idea of using gene editing to treat disease or alter traits dates to at least the 1950s and the discovery of the double-helix structure of DNA. In the mid-20th-century era of genetic discovery, researchers realized that the sequence of bases in DNA is passed (mostly) faithfully from parent to offspring and that small changes in the sequence can mean the difference between health and disease. Recognition of the latter led to the inescapable conjecture that with the identification of molecular mistakes that cause genetic diseases would come the means to fix those mistakes and thereby enable the prevention or reversal of disease. That notion was the fundamental idea behind gene therapy and from the 1980s was seen as a holy grail in molecular genetics.
The development of gene-editing technology for gene therapy, however, proved difficult. Much early progress focused not on correcting genetic mistakes in the DNA but rather on attempting to minimize their consequence by providing a functional copy of the mutated gene, either inserted into the genome or maintained as an extrachromosomal unit (outside the genome). While that approach was effective for some conditions, it was complicated and limited in scope.
In order to truly correct genetic mistakes, researchers needed to be able to create a double-stranded break in DNA at precisely the desired location in the more than three billion base pairs that constitute the human genome. Once created, the double-stranded break could be efficiently repaired by the cell using a template that directed replacement of the bad sequence with the good sequence. However, making the initial break at precisely the desired locationand nowhere elsewithin the genome was not easy.
Before the advent of CRISPR-Cas9, two approaches were used to make site-specific double-stranded breaks in DNA: one based on zinc finger nucleases (ZFNs) and the other based on transcription activator-like effector nucleases (TALENs). ZFNs are fusion proteins composed of DNA-binding domains that recognize and bind to specific three- to four-base-pair-long sequences. Conferring specificity to a nine-base-pair target sequence, for example, would require three ZFN domains fused in tandem. The desired arrangement of DNA-binding domains is also fused to a sequence that encodes one subunit of the bacterial nuclease Fok1. Facilitating a double-stranded cut at a specific site requires the engineering of two ZFN fusion proteinsone to bind on each side of the target site, on opposite DNA strands. When both ZFNs are bound, the Fok1 subunits, being in proximity, bind to each other to form an active dimer that cuts the target DNA on both strands.
TALEN fusion proteins are designed to bind to specific DNA sequences that flank a target site. But instead of using zinc finger domains, TALENs utilize DNA-binding domains derived from proteins from a group of plant pathogens. For technical reasons TALENs are easier to engineer than ZFNs, especially for longer recognition sites. Similar to ZFNs, TALENs encode a Fok1 domain fused to the engineered DNA-binding region, so, once the target site is bound on both sides, the dimerized Fok1 nuclease can introduce a double-stranded break at the desired DNA location.
Unlike ZFNs and TALENs, CRISPR-Cas9 uses RNA-DNA binding, rather than protein-DNA binding, to guide nuclease activity, which simplifies the design and enables application to a broad range of target sequences. CRISPR-Cas9 was derived from the adaptive immune systems of bacteria. The acronym CRISPR refers to clustered regularly interspaced short palindromic repeats, which are found in most bacterial genomes. Between the short palindromic repeats are stretches of sequence clearly derived from the genomes of bacterial pathogens. Older spacers are found at the distal end of the cluster, and newer spacers, representing more recently encountered pathogens, are found near the proximal end of the cluster.
Transcription of the CRISPR region results in the production of small guide RNAs that include hairpin formations from the palindromic repeats linked to sequences derived from the spacers, allowing each to attach to its corresponding target. The RNA-DNA heteroduplex formed then binds to a nuclease called Cas9 and directs it to catalyze the cleavage of double-stranded DNA at a position near the junction of the target-specific sequence and the palindromic repeat in the guide RNA. Because RNA-DNA heteroduplexes are stable and because designing an RNA sequence that binds specifically to a unique target DNA sequence requires only knowledge of the Watson-Crick base-pairing rules (adenine binds to thymine [or uracil in RNA], and cytosine binds to guanine), the CRISPR-Cas9 system was preferable to the fusion protein designs required for using ZFNs or TALENs.
A further technical advance came in 2015, when Zhang and colleagues reported the application of Cpf-1, rather than Cas9, as the nuclease paired with CRISPR to achieve gene editing. Cpf-1 is a microbial nuclease that offers potential advantages over Cas9, including requiring only one CRISPR guide RNA for specificity and making staggered (rather than blunt) double-stranded DNA cuts. The altered nuclease properties gave potentially greater control over the insertion of replacement DNA sequences than was possible with Cas9, at least in some circumstances. Researchers suspect that bacteria house other genome-editing proteins as well, the evolutionary diversity of which could prove valuable in further refining the precision and versatility of gene-editing technologies.
See more here:
Gene editing | Definition, History, & CRISPR-Cas9 | Britannica
- New gene therapy for sickle cell disease has been a long time coming, but is it a complete game-changer? - UCLA Health Connect - December 29th, 2023
- Sickle Cell Gene Therapy Reimbursement: Will Experience Matter? And Which Kind? - Pink Sheet - December 29th, 2023
- Cell and gene therapy will be top industry trend for pharma in 2024 - The Financial Express - December 29th, 2023
- Excision Bio Seeks to Suppress HIV Replication With CRISPR Gene Therapy - AJMC.com Managed Markets Network - December 29th, 2023
- Global Cell and Gene Therapy Manufacturing Industry is projected to surpass a valuation of US$ 240 B - PharmiWeb.com - December 29th, 2023
- Penn's gene therapy layoffs are the latest in biotech belt-tightening in Philly and beyond - The Philadelphia Inquirer - December 29th, 2023
- Adeno-associated Virus Gene Therapy Market is Projected to Grow at a CAGR of 43.4% from 2023-2033 - EIN News - December 29th, 2023
- How Does Gene Therapy Work? Types, Uses, Safety - Healthline - May 9th, 2023
- Gene therapy: Comprehensive overview and therapeutic applications - April 7th, 2023
- Thanks to collaboration between Stand Up Therapeutics and VectorBuilder, a paraplegic patient will get gene therapy for the first time - Business... - January 21st, 2023
- A short history of gene therapy - Boston Children's Answers - December 27th, 2022
- Gene Therapy Analytical Development Summit 2022 | Home - December 27th, 2022
- $3.5-Million Hemophilia Gene Therapy Is World's Most Expensive Drug - Scientific American - December 10th, 2022
- CAR T Global Consultant Inc. Announce their Collaboration with Titronbio - a company founded in Shanghai China by a renowned leader in the field of... - December 10th, 2022
- Cell and Gene Therapy Manufacturing Services Market Size In 2023 | Financial Performance, In-Depth Insight of Trends, Key Players (Thermo Fisher... - December 10th, 2022
- How Gene Therapy Can Cure or Treat Diseases | FDA - December 2nd, 2022
- FDA Approves First Gene Therapy to Treat Adults with Hemophilia B - FDA.gov - November 24th, 2022
- Fact Check-mRNA vaccines are distinct from gene therapy ... - Reuters - October 29th, 2022
- Gene therapy: The Potential for Treating Type 1 Diabetes - Healthline - October 13th, 2022
- The promised land of gene therapy: Commercialization of novel gene-editing technology in beta-thalassemia - PMLiVE - October 13th, 2022
- Gene Therapy Hits Its Stride in the Clinic - Genetic Engineering & Biotechnology News - October 13th, 2022
- FDA Expands Oversight of Cell and Gene Therapies - Pharmaceutical Technology Magazine - October 13th, 2022
- Rocket Pharmaceuticals Announces Presentations Highlighting Lentiviral Gene Therapies at the 29th Annual Congress of the European Society of Gene... - October 13th, 2022
- Health Alert for Parents: How one boy is thriving following treatment with a gene therapy after receiving an early diagnosis - PR Newswire - October 13th, 2022
- M6P Therapeutics Presents Novel AAV Gene Therapy Approach for the Treatment of Gaucher Disease at the ESGCT 29th Annual Congress - Business Wire - October 13th, 2022
- Two Cell and Gene Therapies Manufactured at Lonza Houston Reach FDA Approval - Contract Pharma - October 13th, 2022
- Ascidian starts up with $50M and a twist on RNA editing - BioPharma Dive - October 13th, 2022
- Viral Vector Manufacturing, Non-Viral Vector Manufacturing and Gene Therapy Manufacturing Market Report 2022 - ResearchAndMarkets.com - Business Wire - October 13th, 2022
- Exopharm identifies two new lead programs as exosomes gain prominence in gene therapy - Stockhead - October 13th, 2022
- What Do We Know About Gene Therapy and Cystic Fibrosis? - The Epoch Times - October 13th, 2022
- CSafe and BioLife Solutions, Inc. announce partnership to expand supply chain solutions for the cell and gene therapy market - PR Newswire APAC - PR... - October 13th, 2022
- Genomics in Cancer Care Market is estimated to be US$ 72.61 billion by 2032 with a CAGR of 16.3% during the forecast period 2032 - By PMI -... - October 13th, 2022
- After $100M deal, J&J links gene therapy to improved vision in early-phase trial - FierceBiotech - October 4th, 2022
- Bayer Head Admits COVID-19 Vaccine is Gene Therapy - October 4th, 2022
- New Gene Therapy Shows Promise for Treating Age Related Macular ... - October 4th, 2022
- BioMarin resubmits its hemophilia gene therapy to the FDA - BioPharma Dive - October 4th, 2022
- Why Philly ranks #2 among best cell and gene therapy hubs in the US - Technical.ly - October 4th, 2022
- AMPLIFYBIO ACQUIRES PACT PHARMA ASSETS TO ENHANCE CELL AND GENE THERAPY CHARACTERIZATION CAPABILITIES - PR Newswire - October 4th, 2022
- bluebird's (BLUE) Gene Therapy Approvals to Drive the Top Line - Zacks Investment Research - October 4th, 2022
- Researchers Develop Potential Gene Therapy to Treat Blindness - Managed Healthcare Executive - October 4th, 2022
- Voyager Therapeutics Announces Data Presentations at the 29th Annual Congress of the European Society of Gene & Cell Therapy - GlobeNewswire - October 4th, 2022
- Viral Vector Manufacturing, Non-Viral Vector Manufacturing and Gene Therapy Manufacturing Market by Scale of Operation, Type of Vector, Application... - October 4th, 2022
- Sanofi partners with Scribe to gain gene editing tools for cell therapy work - BioPharma Dive - October 4th, 2022
- Real Endpoints Marketplace announces collaboration with bluebird bio to help scale delivery of a first-of-its-kind value-based contract for one-time... - October 4th, 2022
- Vertex given green light to seek US approval of CRISPR-based therapy - BioPharma Dive - October 4th, 2022
- Atsena Therapeutics Announces Positive Results from Phase I/II Clinical Trial of ATSN-101 for the Treatment of GUCY2D-associated Leber Congenital... - October 4th, 2022
- Meet with the OrganaBio executives in-person at The Cell & Gene Meeting on the Mesa - Business Wire - October 4th, 2022
- Ocular Gene Therapy: A Literature Review With Focus on Current Clinical Trials - Cureus - September 25th, 2022
- Will experience support use of first-ever retinal gene therapy? - Ophthalmology Times - September 25th, 2022
- Rejuvenate Bio CEO highlights ambitious approach of using gene therapy to reverse aging - MedCity News - September 25th, 2022
- Gene therapy R&D market is projected to grow at a CAGR of 30.1% by 2032: Visiongain Reports Ltd - GlobeNewswire - September 25th, 2022
- Emulate Announces New Gene Therapy Application Enabling Accelerated Development of Treatments for Genetic Diseases with Organ-on-a-Chip Technology -... - September 25th, 2022
- CSL flexes gene therapy muscle with latest drug - Sydney Morning Herald - September 25th, 2022
- FDA Updates for the Week of Sept. 19, 2022 - Managed Healthcare Executive - September 25th, 2022
- AVROBIO Receives Rare Pediatric Disease Designation From FDA For First Gene Therapy In Development For Cy - Benzinga - September 25th, 2022
- Preparing Europe for a new generation of innovative therapies - Open Access Government - September 25th, 2022
- KSQ Therapeutics Announces Leadership and Board Additions - Business Wire - September 25th, 2022
- Hemophilia gene therapies from BioMarin, CSL pick up an early ICER endorsement - FiercePharma - September 16th, 2022
- Challenges In Gene Therapy - University of Utah - September 16th, 2022
- Hopkinton bioscience company forms advisory board for cell and gene therapy - Worcester Business Journal - September 16th, 2022
- Gene Therapy Restores Some Vision in Colorblind Children - Pharmacy Times - September 16th, 2022
- Coave Therapeutics partners with IMN on gene therapy - Labiotech.eu - September 16th, 2022
- MeiraGTx's Industry-Leading Gene Therapy Manufacturing Facility in Shannon, Ireland Formally Introduced by Head of Irish Government -... - September 16th, 2022
- Bluebird CFO resigns ahead of gene therapy launch - BioPharma Dive - September 16th, 2022
- Enrollment completed in STARLIGHT Phase 2 clinical trial of MCO-010 optogenetic gene therapy for Stargardt disease - Ophthalmology Times - September 16th, 2022
- Global Cancer Gene Therapy Market to Reach $11.35 Billion by 2030 at a CAGR of 23.3% - PR Newswire - September 16th, 2022
- How WhiteLab Genomics is using AI to aid gene and cell therapy development - TechCrunch - September 16th, 2022
- CAR-T Cell Therapy Market Segmentation and Forecast Analysis up to 2027 - BioSpace - September 16th, 2022
- Cluster-busters against Cancer and a Potential New Gene Therapy for LCA - BioSpace - September 16th, 2022
- Ori Biotech and CTMC team up on cell therapy delivery - BioPharma-Reporter.com - September 16th, 2022
- Adverum (ADVM) Doses First Patient in the Mid-Stage wAMD Study - Zacks Investment Research - September 16th, 2022
- Engensis Gene Therapy for ALS Found Safe in Small Phase 2a Trial |... - ALS News Today - September 8th, 2022
- A new gene therapy based on antibody cells is about to be tested in humans - MIT Technology Review - September 8th, 2022
- Myrtelles rAAV-Olig001-ASPA Gene Therapy Candidate for Canavan Disease Receives Advanced Therapy Medicinal Product Classification from the European... - September 8th, 2022
- Sangamo presses ahead with Fabry disease gene therapy - BioPharma Dive - September 8th, 2022
- The gene therapy that could transform the lives of millions - ABC News - September 8th, 2022
- As Philly becomes a hub for life sciences, a new program will train workers for jobs in the field - The Philadelphia Inquirer - September 8th, 2022
- Mayflower BioVentures to Announce New Cell & Gene Therapy Companies within Months - BioSpace - September 8th, 2022
- Carroucell Raises 1.5 Million to Introduce Breakthrough Microcarriers and Customizable Processes to Cell and Gene Therapy Market - Business Wire - September 8th, 2022
- AnGes : Announces Discontinuation of Development of HGF Gene Therapy Product for Additional Indication of Chronic Arterial Occlusive Disease with Rest... - September 8th, 2022