how can crispr benefit society

^{Below are eight ways that CRISPR will likely impact the world:}

1. Remove malaria from mosquitos. Scientists have created mosquitoes that are resistant to malaria by deleting a segment of mosquito DNA. ...
2. Treating Alzheimer’s disease. CRISPR-based platforms have been developed to identify the genes controlling the cellular processes that lead to neurodegenerative diseases like Alzheimer's and Parkinson's, hopefully leading to new ...
3. Treating HIV. The HIV virus inserts its DNA into the cells of the human host. CRISPR has been successful in removing the virus’s DNA from the patient’s genome. ...
Develop new drugs. Pharmaceutical companies such as Bayer AG are investing hundreds of millions of dollars to develop CRISPR-based drugs to treat heart disease, blood disorders, and blindness.
Livestock. CRISPR/Cas9 has been utilized in China to delete genes in livestock that inhibit muscle and hair growth to grow larger stock for the country's commercial meat and wool ...
Agricultural crops. Researchers are using CRISPR to discover new ways to improve crop disease resistance and environmental stress tolerance in plants. ...
Develop new cancer treatments. CRISPR can modify immune cells to make them more effective at targeting and destroying cancer cells. ...
Reduce our need for plastic. CRISPR can be used to manipulate a type of yeast that transforms sugars into hydrocarbons, which can be used to make plastic—greatly reducing the ...

What are the pros and cons of CRISPR?

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What is CRISPR and how does it work?

The process of using a CRISPR system involves the following steps:

1. Use the endonuclease Cas9 to cut open a double-stranded DNA molecule and separate them into single strands, which remains attached to one end of each strand.
2. Edit the DNA sequence using CRISPR RNA and a CRISPR enzyme.
3. Create new strands from the edited DNA sequence with a DNA polymerase.

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How can CRISPR help humans?

Ways in Which CRISPR-Cas9 is Helping Humans

1. Ridding the World of Malaria. Scientists are determined that gene editing through Cas9 Can help them eliminate Malaria from mosquitoes.
2. Treating Many Viruses. Viruses can be very detrimental and sometimes even fatal to the human body. ...
3. Development Of New Drugs. Many pharmaceutical companies manufacture CRISPR-based drugs. ...
4. Eliminating Crop Diseases. ...

What diseases can CRISPR treat?

CRISPR is already showing promise for treating devastating blood disorders such as sickle cell disease and beta thalassemia. And doctors are trying to use it to treat cancer. And doctors are ...

How is CRISPR good for society?

CRISPR is having a major impact on diagnostics and therapeutics, where it allows medicine to become more personalized. Treatments for cancer and blood disorders are furthest along because of how CRISPR is performed, she said. “The most tested medical applications of CRISPR have been for cancer.

What are some benefits of using CRISPR?

Arguably, the most important advantages of CRISPR/Cas9 over other genome editing technologies is its simplicity and efficiency. Since it can be applied directly in embryo, CRISPR/Cas9 reduces the time required to modify target genes compared to gene targeting technologies based on the use of embryonic stem (ES) cells.

How does gene editing benefit society?

Potential benefits of human genome editing include faster and more accurate diagnosis, more targeted treatments and prevention of genetic disorders.

How can CRISPR change the world?

Thanks to its pinpoint accuracy and relatively low production costs, CRISPR could potentially change everything involving genes: from curing diseases and improving agriculture, to repairing genetic disorders like sickle cell anemia or hemophilia.

What are some pros and cons of CRISPR?

The ProsIt's Simple to Amend Your Target Region. OK, setting up the CRISPR-Cas9 genome-editing system for the first time is not simple. ... There Are Lots of Publications Using CRISPR-Cas9 Genome Editing. ... It's Cheap. ... Setting up from Scratch Is a Considerable Time Investment. ... It Is Not Always Efficient. ... Off-Target Effects.

What are the pros for Crispr gene editing?

What are the advantages of CRISPR over other genome editing tools? The CRISPR-Cas9 system can modify DNA with greater precision than existing technologies. An advantage the CRISPR-Cas9 system offers over other mutagenic techniques, like ZFN and TALEN, is its relative simplicity and versatility.

What are three of the positive effects pros of using gene editing technology on humans?

It could help eliminate hundreds of diseases. It could eliminate many forms of pain and anxiety . It could increase intelligence and longevity. It could change the scale of human happiness and productivity by many orders of magnitude.

Is Gene Therapy important to society?

Gene therapy holds promise for treating a wide range of diseases, such as cancer, cystic fibrosis, heart disease, diabetes, hemophilia and AIDS. Researchers are still studying how and when to use gene therapy. Currently, in the United States, gene therapy is available only as part of a clinical trial.

What are the diseases that CRISPR can help?

More likely, however, a CRISPR-modified world will be a world where genome-tinkering may be employed to provide solutions in agriculture and pest resistance, sustainable farming, cancer and other genetic diseases such as Muscular Dystrophy, Diabetes, Thalassemia’s and Huntington’s Disease. Even as exciting as making human-pig chimaera embryos to grow human organs to relieve the worldwide shortage of organs needed for transplants.

Where are we with CRISPR?

A future of CRISPR-based genome editing is unavoidable at this stage. In fact, decades from now we may look back through genetically altered eyes, free of diseases, free of cancer and struggling to remember a world where our DNA was an inherited constant. But what does this mean? Theoretically, the applications for CRISPR-Cas9 are endless; ranging from revolutionary advances in biomedical therapeutics to some of the science fiction reminiscent of 1997’s cult movie, “Gattaca”. Earlier this year, scientists from Temple University published a study in Scientific Reports demonstrating for the first time the CRISPR-mediated elimination of a HIV genome from human immune cells. Using a non-pathogenic strain of virus as a transmission vector, scientists were able to use CRISPR-Cas9 to effectively excise the viral HIV-1 DNA from infected human T-cells. These CRISPR-modified cells failed to support viral replication, thus being protected from future re-infection.

What is CRISPR-Cas9?

In its essence, CRISPR is a multifaceted molecular scissor with the ability to precisely snip out any gene of interest that can be found in our DNA. CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats. When paired with the endonuclease enzyme, Cas9 (CRISPR-associated protein 9), CRISPR-Cas9 is the biggest biomedical breakthrough since the early days of recombinant DNA (rDNA) in the 1970’s. The basis of this technology is an adaptation of a bacterial defence system to locate and destroy the genomes of invading viruses. Like bacteria, we are able to recognise stretches of DNA amidst entire genomes that we can accurately target and latch on to using RNA guides. By adding a nuclease such as the Cas9, we can use this guide system to seek and snip out genes of interest. CRISPR-Cas9 genome editing is now being used by laboratories all over the world to cheaply and accurately modulate specific genes in lab cultures, merely at a whim.

What is CRISPR gene editing?

Using a method known as “Gene Drive”, CRISPR gene editing can be used to bypass (or selectively enhance) the restraints of mendelian inheritance to speed up the propagation of genetic modifications rapidly through future generations.

Is genomic editing a step in the future?

However, with the news that the UK Human Fertilisation and Embryology Authority (HFEA) has officially endorsed genomic editing of human embryos for research , this first step may become a leap in the not-so-distant future. Beyond the biomedical space, researchers are proposing to use CRISPR to change the world around us.

How does CRISPR help mosquitoes?

Gene-editing techniques like CRISPR could directly combat infectious diseases, but some researchers have decided to slow the spread of disease by eliminating its means of transmission. Scientists at the University of California, Riverside developed a kind of mosquito that is uniquely susceptible to changes made with CRISPR, giving scientists unprecedented control over the traits that the organism passes to its offspring. The result: yellow, three-eyed, wingless mosquitoes, created by altering genes responsible for eye, wing, and cuticle development.

What is CRISPR gene editing?

CRISPR gene editing has proven to be promising in the field of agricultural research. Scientists from Cold Spring Harbor Laboratory in New York used the tool to increase the yield of tomato plants. The lab developed a method to edit the genes that determine tomato size, branching architecture and, ultimately, shape of the plant for a greater harvest.

How many embryos did CRISPR remove?

The results were promising: Of the 54 embryos that were injected with the CRISPR-Cas9 machinery 18 hours after fertilization, 36 did not show any mutations in the gene (practically no chance of developing the disease) and 13 were partially free of mutations (with a 50 percent chance of inheriting HCM).

What is the technology used to edit genes?

Compared to other tools used for genetic engineering, CRISPR (also known by its more technical name, CRISPR-Cas9) is precise, cheap, easy to use, and remarkably powerful.

When will CRISPR be used in China?

Another gene-editing trial in China is due to begin in July 2018 and will attempt to use CRISPR to disrupt the genes of the human papillomavirus (HPV) — a virus that has been shown to cause cervical cancer tumor growth — effectively destroying it.

When was CRISPR first used?

Discovered in the early 1990s, and first used in biochemical experiments seven years later, CRISPR has rapidly become the most popular gene editing tool among researchers in fields such as human biology, agriculture, and microbiology.

How did Chinese researchers increase resistance to HIV in mice?

In 2017, a team of Chinese researchers successfully increased resistance to HIV in mice by replicating a mutation of a gene that effectively prevents the virus from entering cells. For now, scientists are only conducting these experiments in animals, but there’s reason to think the same methods could work in humans.

What are the benefits of CRISPR?

Some of the benefits are discussed below. Cancer Therapeutics: New immunotherapies can be developed using CRISPR to treat cancer. Scientists can genetically modify T-cells using CRISPR to locate and kill cancer cells.

What are the pros and cons of CRISPR?

While the benefits of CRISPR range from curing genetic conditions to organ transplants, ethicists fear its use in promoting desired traits rather than life-saving traits such as intelligence that could have long-term implications.

What are the ethical concerns of CRISPR?

Ethical Concerns of CRISPR [Cons] Changes to the Germ-line Cells: Genetically modifications to human embryos and reproductive cells such as eggs and sperms are called germline editing. Changes to the germline can be passed to the next generation.

How much did Darpa invest in 2017?

DARPA, US’s secretive Defense Advanced Research Projects Agency, announced to invest US$65 million in 2017 over four years in seven teams that will investigate ways to make gene editing technologies safer and targeted. The program relates to both intentional and unintended consequences of gene editing technologies.

Is CRISPR a good tool for terrorists?

Compared to other genetic engineering tools, CRISPR technology is relatively inexpensive and simple, which could make it attractive to terrorist organization s. The technology can be used to genetically modify bacteria or viruses to wage biological attacks against humans.

Can CRISPR kill cancer cells?

Scientists can genetically modify T-cells using CRISPR to locate and kill cancer cells. Curing Genetic Diseases: CRISPR technology can eliminate the genes that cause genetic diseases such as diabetes, cystic fibrosis.

Is CRISPR a good gene editing tool?

CRISPR has become one of the most powerful gene-editing tools today. Unlike other genetic engineering tools, CRISPR is cheap, relatively easy to use and precise. Undoubtedly, its popularity has surged amongst scientists in the biotechnology industry.

How does CRISPR help humans?

There are many ways in which CRISPR can help humans by eliminating a lot of diseases: 1. Ridding the World of Malaria. Scientists are determined that gene editing through Cas9 Can help them eliminate Malaria from mosquitoes. They said that it is possible to change the genetic DNA of the mosquitoes having Malaria.

Why is CRISPR a great innovation?

It is a great innovation for such people who have a weakened immune system, especially because this treatment has the potential to eliminate many viruses from the Earth (Crawford, 2017). 3. Development Of New Drugs. Many pharmaceutical companies manufacture CRISPR-based drugs.

What is CRISPR used for?

These drugs have proven to be beneficial in treating heart diseases, different types of blood disorders, and congenital diseases (Hesman Saey, 2019).

What is the purpose of CAS9?

The protein CAS9 Acts as a molecular scissor that can cut the defected DNA strands (Crawford, 2017). Gene editing has been made simple and easy by the CRISPR treatment. Many people are getting various health benefits by using this significant treatment.

Is CRISPR a good gene editing tool?

CRISPR Treatment is a very effective method for gene editing, with multiple factors making it beneficial for human beings. This treatment has brought a revolution in the field of science and medicine.

Can mosquitoes change their DNA?

They said that it is possible to change the genetic DNA of the mosquitoes having Malaria . Every year, a large number of people suffer from Malaria because it is so prevalent in mosquitoes. If scientists are successful in eliminating Malaria from the mosquitoes through CRISPR treatment, it can be very beneficial for humans.

What is CRISPR used for?

Berkeley researchers are hoping to use CRISPR to sterilize female mosquitoes to control the spread of malaria, dengue, and other mosquito-borne illnesses. Known as a gene drive, the technology propagates a particular set of genes throughout entire populations, offering a cost-effective and, advocates contend, environmentally friendly solution to insect-borne disease.

Who discovered CRISPR?

In October, UC Berkeley professor Jennifer Doudna shared the Nobel Prize in chemistry for the discovery of CRISPR/Cas9, a gene-slicing tool that can be programmed to make precise edits to DNA. Since its discovery, CRISPR has captured the imaginations of everyone from pig farmers to infectious disease researchers. Here are just some of the ways it is already being put to work.

What is the greatest advantage of CRISPR?

The single greatest advantage of CRISPR is that it has taken gene editing out of the hands of molecular biologists and made it accessible to a broader array of researchers. Since this discovery, new companies have been founded to leverage CRISPRs ability to reliably alter genes in almost any organism.

What is CRISPR?

Clustered Regularly Interspaced Short Palindromic Repeats are a naturally-occurring genetic sequence native to the immune systems of certain bacteria. When a virus invades the bacteria for the first time, the CRISPR sequence allow the bacteria to "remember" the virus. 3 If the virus attacks again, the CRISPR array produces RNA segments to target the virus as well as an enzyme known as cas9 to chop up the viral DNA. 3

What was the CRISPR technology used for?

This act was widely condemned in professional circles as reckless and unethical.2The news also brought widespread public attention to CRISPR, the cutting-edge technology that was used to produce the genetically-altered embryos. In this article we want to look past the sensational news out of China, and focus more on the perfectly legitimate ways that CRISPR could revolutionize the drug world and the corresponding implications for plan sponsors.

How does CRISPR help in drug development?

CRISPR can help test drug candidates. One of the most crucial elements of drug development process is testing candidate drugs for efficacy and toxicity. For ethical reasons, this testing is rarely done in living people. Instead, researchers rely on cell and animal models of the human diseases they seek to cure. 11.

Can CRISPR be used to edit DNA?

In 2012, scientists realized that CRISPR could be re-purposed as a quicker, easier, more precise way to edit DNA.4This discovery unlocked a vast new realm of possibilities.

Is CRISPR a good candidate for drug development?

For decades, many scientists considered the parts of the genome that does not encode for proteins to be “junk DNA” and therefore not a good candidate for drug development. 9.

Is CRISPR a rare disease?

CRISPR-derived therapies are being developed for a wide range of diseases but it is likely that the first drugs to make it to market will be for rare diseases. Judging by existing costs for orphan drugs and gene therapies, one can reasonably expect CRISPR-derived drugs to be very expensive.

What is the role of CRISPR in cancer?

With CRISPR, we have the chance to challenge various types of cancer at the molecular level, address the environmental damage we’ve caused on the planet, slow the spread of disease and disability, and improve the quality of life for everyone.

Who is the pioneer of CRISPR?

Harvard geneticist and CRISPR pioneer George Church believes he can use the tool to genetically modify endangered Indian elephants into “wooly mammoths” capable of surviving in the freezing wilderness of Siberia.

What is CRISPR Cas9?

CRISPR-Cas9 is a unique gene editing tool that allows scientists to cut out segments of DNA from the genome of any organism, move them around, or replace them entirely with remarkable precision . Think of it as the cut-and-paste tool in Microsoft Word except with ...

What is the impact of CRISPR Cas9 on DNA?

Once unsubstantiated fear and paranoia take hold, scientists will have a much tougher time implementing the research needed to save countless lives. CRISPR-Cas9 editing a strand of DNA. The research shows that more knowledge leads to more understanding and acceptance.

What is CRISPR in Microsoft Word?

As CRISPR co-discoverer Jennifer Doudna, a professor of biochemistry at the University of California, Berkley, describes it, CRISPR is essentially a “molecular scalpel for genomes.”.

Why did scientists genetically modify mosquitoes?

Scientists recently genetically modified the genome of mosquitoes to make them resistant to Plasmodium falciparum, the parasite responsible for causing malaria. With CRISPR’s precision and accuracy, they were able to insert the necessary genes into the mosquitoes’ DNA.

What percentage of people know little about gene editing?

But most people have no idea what they are concerned about—about 90 percent knew little or nothing about gene editing. Many respondents expressed doubts about using gene editing on human babies to reduce the risk of serious diseases—designer baby, Gattaca-type fears. “It’s messing with nature.

Why is CRISPR used?

“CRISPR could be used to create an affordable way to detect disease at the point of care, in the field or at home ,” she said.

How does CRISPR help with cancer?

CRISPR is having a major impact on diagnostics and therapeutics, where it allows medicine to become more personalized. Treatments for cancer and blood disorders are furthest along because of how CRISPR is performed, she said.

What is CRISPR gene editing?

CRISPR gene editing is revolutionizing genetics, cell biology, and now medicine. Every month it seems that scientists and clinicians are devising new applications for CRISPR, which stands for “clustered regularly interspaced short palindromic repeats.”.

What is the green target of CRISPR?

Artistic rendering of the CRISPR-Cas9 system in action. The Cas9 (green) is shown finding the DNA (blue) target that matches its programmed guide RNA (gold) and making a double-strand break, the first step in genome editing. Credit: Janet Iwasa/Innovative Genomics Institute

What is Megan Hochstrasser's job?

Megan Hochstrasser’s job is to keep up with the latest in the world of CRISPR. Hochstrasser is science communications manager for the Innovative Genomics Institute (IGI), a joint research entity that operates as a collaboration between the University of California, Berkeley, and the University of California, San Francisco.

What is the concern with CRISPR?

A major concern for implementing CRISPR/Cas9 for gene therapy is the relatively high frequency of off-target effects (OTEs), which have been observed at a frequency of ≥50% ( 31 ). Current attempts at addressing this concern include engineered Cas9 variants that exhibit reduced OTE and optimizing guide designs. One strategy that minimizes OTEs utilizes Cas9 nickase (Cas9n), a variant that induces single-stranded breaks (SSBs), in combination with an sgRNA pair targeting both strands of the DNA at the intended location to produce the DSB ( 32 ). Researchers have also developed Cas9 variants that are specifically engineered to reduce OTEs while maintaining editing efficacy ( Table 1 ). SpCas9-HF1 is one of these high-fidelity variants that exploits the “excess-energy” model which proposes that there is an excess affinity between Cas9 and target DNA which may be enabling OTEs. By introducing mutations to 4 residues involved in direct hydrogen bonding between Cas9 and the phosphate backbone of the target DNA, SpCas9-HF1 has been shown to possess no detectable off-target activity in comparison to wildtype SpCas9 ( 35 ). Other Cas9 variants that have been developed include evoCas9 and HiFiCas9, both of which contain altered amino acid residues in the Rec3 domain which is involved in nucleotide recognition. Desensitizing the Rec3 domain increases the dependence on specificity for the DNA:RNA heteroduplex to induce DSBs, thereby reducing OTEs while maintaining editing efficacy ( 38, 39 ). One of the more recent developments is the Cas9_R63A/Q768A variant, in which the R63A mutation destabilizes R-loop formation in the presence of mismatches and Q768A mutation increases sensitivity to PAM-distal mismatches ( 49 ). Despite the different strategies, the rational for generating many Cas9 variants with reduced OTEs has been to ultimately reduce general Cas9 and DNA interactions and give a stronger role for the DNA:RNA heteroduplex in facilitating the edits.

What is CRISPR in gene therapy?

The discovery of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated (Cas) proteins has expanded the applications of genetic research in thousands of laboratories across the globe and is redefining our approach to gene therapy. Traditional gene therapy has raised some concerns, as its reliance on viral vector delivery of therapeutic transgenes can cause both insertional oncogenesis and immunogenic toxicity. While viral vectors remain a key delivery vehicle, CRISPR technology provides a relatively simple and efficient alternative for site-specific gene editing, obliviating some concerns raised by traditional gene therapy. Although it has apparent advantages, CRISPR/Cas9 brings its own set of limitations which must be addressed for safe and efficient clinical translation. This review focuses on the evolution of gene therapy and the role of CRISPR in shifting the gene therapy paradigm. We review the emerging data of recent gene therapy trials and consider the best strategy to move forward with this powerful but still relatively new technology.

What is Cas9 in DNA repair?

CRISPR/Cas9 mediated gene editing. Cas9 in complex with the sgRNA targets the respective gene and creates DSBs near the PAM region. DNA damage repair proceeds either through the NHEJ pathway or HDR. In the NHEJ pathway, random insertions and deletions (indels) are introduced at the cut side and ligated resulting in error-prone repair. In the HDR pathway, the homologous chromosomal DNA serves as a template for the damaged DNA during repair, resulting in error-free repair.

What is the purpose of CRISPR/CAS9?

CRISPR/Cas9 is a simple two-component system used for effective targeted gene editing. The first component is the single-effector Cas9 protein, which contains the endonuclease domains RuvC and HNH. RuvC cleaves the DNA strand non-complementary to the spacer sequence and HNH cleaves the complementary strand. Together, these domains generate double-stranded breaks (DSBs) in the target DNA. The second component of effective targeted gene editing is a single guide RNA (sgRNA) carrying a scaffold sequence which enables its anchoring to Cas9 and a 20 base pair spacer sequence complementary to the target gene and adjacent to the PAM sequence. This sgRNA guides the CRISPR/Cas9 complex to its intended genomic location. The editing system then relies on either of two endogenous DNA repair pathways: non-homologous end-joining (NHEJ) or homology-directed repair (HDR) ( Figure 2 ). NHEJ occurs much more frequently in most cell types and involves random insertion and deletion of base pairs, or indels, at the cut site. This error-prone mechanism usually results in frameshift mutations, often creating a premature stop codon and/or a non-functional polypeptide. This pathway has been particularly useful in genetic knock-out experiments and functional genomic CRISPR screens, but it can also be useful in the clinic in the context where gene disruption provides a therapeutic opportunity. The other pathway, which is especially appealing to exploit for clinical purposes, is the error-free HDR pathway. This pathway involves using the homologous region of the unedited DNA strand as a template to correct the damaged DNA, resulting in error-free repair. Experimentally, this pathway can be exploited by providing an exogenous donor template with the CRISPR/Cas9 machinery to facilitate the desired edit into the genome ( 30 ).

How does HDR improve gene editing?

Enhancement of HDR efficiency has been achieved via suppression of the NHEJ pathway through chemical inhibition of key NHEJ modulating enzymes such as Ku ( 77 ), DNA ligase IV ( 78 ), and DNA-dependent protein kinases (DNA-PKcs) ( 79 ). Other strategies that improve HDR efficiency include using single-stranded oligodeoxynucleotide (ssODN) template, which contains the homology arms to facilitate recombination and the desired edit sequence, instead of double-stranded DNA (dsDNA). Rationally designed ssODN templates with optimized length complementarity have been shown to increase HDR rates up to 60% in human cells for single nucleotide substitution ( 80 ). Furthermore, cell cycle stage plays a key role in determining the DNA-damage repair pathway a cell may take. HDR events are generally restricted to late S and G2 phases of the cell cycle, given the availability of the sister chromatid to serve as a template at these stages, whereas NHEJ predominates the G1, S, and G2 phases ( 81 ). Pharmacological arrest at the S phase with aphidicolin increased HDR frequency in HEK293T with Cas9-guide ribonucleoprotein (RNP) delivery. Interestingly, cell arrest in the M phase using nocodazole with low concentrations of the Cas9-guide RNP complex yielded higher frequencies of HDR events in these cells, reaching a maximum frequency of up to 31% ( 82 ). Although HDR is considered to be restricted to mitotic cells, a recent study revealed that the CRISPR/Cas9 editing can achieve HDR in mature postmitotic neurons. Nishiyama et al. successfully edited the CaMKIIα locus through HDR in postmitotic hippocampal neurons of adult mice in vitro using an AAV delivered Cas9, guide RNA, and donor template in the CaMKIIα locus, which achieved successful HDR-mediated edits in ~30% of infected cells. Although HDR efficiency was dose-dependent on AAV delivered HDR machinery and off-target activity was not monitored, this study demonstrated CRISPR's potential utility for translational neuroscience after further developments ( 83 ). To further exploit cell-cycle stage control as a means to favor templated repair, Cas9 conjugation to a part of Geminin, a substrate for G1 proteosome degradation, can limit Cas9 expression to S, G2, and M stages. This strategy was shown to facilitate HDR events while mitigating undesired NHEJ edits in human immortalized and stem cells ( 84, 85 ). A more recent strategy combined a chemically modified Cas9 to the ssODN donor or a DNA adaptor that recruits the donor template, either of which improved HDR efficiency by localizing the donor template near the cleavage site ( 86 ). Despite these advancements, HDR is still achieved at a relatively low efficiency in eukaryotic cells and use of relatively harmful agents in cells such as NHEJ chemical inhibitors may not be ideal in a clinical setting.

What is the CRISPR locus?

The bacterial CRISPR locus was first described by Francisco Mojica ( 23) and later identified as a key element in the adaptive immune system in prokaryotes ( 24 ). The locus consists of snippets of viral or plasmid DNA that previously infected the microbe (later termed “spacers”), which were found between an array of short palindromic repeat sequences. Later, Alexander Bolotin discovered the Cas9 protein in Streptococcus thermophilus, which unlike other known Cas genes, Cas9 was a large gene that encoded for a single-effector protein with nuclease activity ( 25 ). They further noted a common sequence in the target DNA adjacent to the spacer, later known as the protospacer adjacent motif (PAM)—the sequence needed for Cas9 to recognize and bind its target DNA ( 25 ). Later studies reported that spacers were transcribed to CRISPR RNAs (crRNAs) that guide the Cas proteins to the target site of DNA ( 26 ). Following studies discovered the trans-activating CRISPR RNA (tracrRNA), which forms a duplex with crRNA that together guide Cas9 to its target DNA ( 27 ). The potential use of this system was simplified by introducing a synthetic combined crRNA and tracrRNA construct called a single-guide RNA (sgRNA) ( 28 ). This was followed by studies demonstrating successful genome editing by CRISPR/Cas9 in mammalian cells, thereby opening the possibility of implementing CRISPR/Cas9 in gene therapy ( 29) ( Figure 1 ).

How is CRISPR delivered?

Delivery of CRISPR Therapy. Nucleic acids encoding CRISPR/Cas9 or its RNP complex can be packaged into delivery vehicles. Once packaged, edits can be facilitated either ex vivo or in vivo. Ex vivo editing involves extraction of target cells from the patient, cell culture, and expansion in vitro, delivery of the CRISPR components to yield the desired edits, selection, and expansion of edited cells, and finally reintroduction of therapeutic edited cells into the patient. In vivo editing can be systemically delivered via intravenous infusions to the patient, where the CRISPR cargo travels through the bloodstream via arteries leading to the target tissue, or locally delivered with injections directly to target tissue. Once delivered, the edits are facilitated in vivo to provide therapeutic benefit.