What if an incurable disease such as AIDS, Cancer, and thousand others could be cured by simply fixing the genome or DNA sequences? Researchers are betting they can with CRISPR, a powerful technology that allows scientists to quickly target, delete, and repair any mutated sequence of DNA in any gene. Other gene-editing tools have emerged in recent years, but none seems to match the precision, low cost, and usability of CRISPR, which is rapidly transforming genetic research and has entered testing as a medical treatment.
CRISPR enables scientists to add, delete, or alter specific parts of the genome, or DNA sequence, in living cells. It consists of two key molecules: Cas9, an enzyme that cuts the two strands of DNA at a certain location; and a small piece of RNA known as guide RNA, which is a pre-designed RNA sequence that ensures that Cas9 cuts where researchers want it to.
It’s no exaggeration to say that CRISPR has been revolutionary. Today, barely a week goes by without news of another CRISPR “breakthrough.” With CRISPR, we can do genetic experiments that would have been unimaginable just a few years ago, not just on inherited disorders but also on genes that contribute to acquired diseases, including AIDS, cancer, and heart diseases.
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The List of Deadly Sickness That Can Be Treated Using CRISPR Technology
With CRISPR, scientists may have the ability to remove or correct disease-causing genes or insert new ones that could theoretically cure disease, including cancer. It has the potential to revolutionize cancer therapy, chiefly in the realm of immunotherapy. Until now, more than 86 people with different forms of cancer have been treated with CRISPR in China, and the results of its success should be available soon. Also, in the US, the first CRISPR trial is planned to target cancer using CRISPR to remove PD-1 as well as a T cell receptor from T cells engineered to express a cancer-targeting receptor.
In China, Hangzhou Cancer Hospital is testing the potential of the gene-editing tool in patients with advanced cancer of the esophagus. This is one of the first and most advanced CRISPR clinical trials. And cancer-disease could be where the first applications of CRISPR could happen. The tests at the Chinese hospital Hospital started with the extraction of T cells from the patient. By using CRISPR, the cells are modified to remove the gene that encodes for a receptor called PD-1 that some tumors are able to bind to and instruct the immune system not to attack. The cells are then reinfused into the patient with a higher capacity to attack tumor cells.
Scientists in Japan have used CRISPR-Cas9 technology to stop human immunodeficiency virus type 1 (HIV-1) replication in latently infected T cells that can’t be controlled using existing drug treatments. The team at Kobe University used CRISPR/Cas9 to remove two regulatory genes of HIV-1 genes, tat, and rev, which causes 95 percent of HIV infections – within infected human cell lines, and so stop further production of the virus. This was repeated in each of the six major subtypes of HIV-1. The team said there were also no off-target effects seen from the genome editing, and it did not affect the survival of the cultured cells.
Although advances have meant that HIV has gone from being a deadly disease to a chronic condition manageable with drug treatment, there is no cure as the virus can sit in dormant reservoirs within cells. It does this by incorporating itself within the chromosomes of a patient’s cells. As soon as treatment is stopped, the virus rebounds, and so drug therapy must be lifelong.
‘These results show that the CRISPR/Cas9 system is a promising method for treating HIV infection,’ said Professor Masanori Kameoka, one of the study authors. ‘We now need to investigate how we can selectively introduce a CRISPR/Cas9 system that targets HIV-1 genes into the infected cells of patients. In order to safely and effectively introduce the CRISPR/Cas9 system, the vectors must be improved,’ said Professor Kameoka. ‘We hope this research will provide us with useful information in developing a treatment method that can completely cure the HIV-1 infection.’
Experimental gene therapy has cured mice of diabetes, and although work is at a very early stage, scientists hope the technique can free people from its effects. American scientists adapted the gene-editing technology known as CRISPR (clustered, regularly interspaced, short palindromic repeat) to treat mouse models of type 1 diabetes successfully.
The CRISPR system is loaded into a harmless virus called an adeno-associated virus (AAV), which carries the tool to the target. But the entire protein, consisting of dCas9, the switches, and the guide RNAs, is too big to fit inside one of these AAVs. To solve that issue, the researchers split the protein into two, loading dCas9 into one virus and the switches and guide RNAs into another. The guide RNAs were tweaked to make sure both parts still ended up at the target together, and to make sure the gene was strongly activated.
To test how well the new technique worked, the researchers experimented with mice that had three different diseases – kidney damage, type 1 diabetes, and muscular dystrophy. In each case, the mice were treated with specialized CRISPR systems to increase the expression of certain genes, which would hopefully reverse the symptoms. In the kidney-damaged mice, the team targeted two genes that play a role in kidney function. Sure enough, there was an increase in the levels of a protein linked to those genes, and kidney function improved. In the diabetic mice, the targeted genes were those that promote the growth of insulin-producing cells, and after treatment, the mice were found to have lower blood glucose levels. And finally, the treatment also worked to reverse the symptoms of muscular dystrophy.
After that promising start, further work is underway on the system. The researchers plan to try to apply the technique to other cell types to help treat other diseases and conduct more safety tests before human trials can begin..
4. Parkinson’s Disease (PD)
Parkinson’s disease (PD) is the second most common neurodegenerative disease and is estimated to afflict up to 5.8 million people worldwide, with the increased mean age of the population in the Western world, the prevalence of the disease is projected to rise. The current mainstay treatment of Parkinson’s disease (PD) consists of dopamine replacement therapy which does not delay disease progression. The field of gene editing system offers a potential means to improve current therapy.
Researchers from MIT have developed ‘REPAIR’, a new version of the CRISPR/Cas9 gene-editing system that can edit RNA instead of DNA — so as not to alter a person’s genome — and may one day be used to treat Parkinson’s and a variety of other diseases. REPAIR — the RNA Editing for Programmable A to I Replacement — can change a single RNA nucleotide, potentially reversing some of the disease-causing mutations at the RNA level. The classic CRISPR/Cas9 system involves a guide RNA (gRNA) and a protein called the Cas9 nuclease. The gRNA guides the Cas9 nuclease to a precise location in the genome, where Cas9 can cause a double-stranded break. This prompts the cell’s repair machinery to fix the break, leading to mutations in the gene so as to cause it not to be expressed.
The researchers have engineered a new system for mammalian gene editing, which targets the RNA sequence instead of the DNA sequence. DNA codes for RNA molecules, which then code for proteins. In this way, targeting RNA can still change the gene product (a protein), but without making a change in the entire genome. This new ability to edit RNA opens up more potential opportunities to recover that function and treat many diseases, in almost any kind of cell. “The ability to correct disease-causing mutations is one of the primary goals of genome editing,” Dr. Feng Zhang, the study’s senior author and an associate professor in Brain and Cognitive Sciences and Biological Engineering departments at MIT, said in a press release. The team plans to continue to improve REPAIRv2 by manipulating the delivery system to improve its effectiveness when introduced into human cells.
5. Huntington’s Disease
People having Huntington’s disease, the nerve cells of the brain start to break down over time. The disease is fatal, often within 10 to 30 years, and as of now, there is no cure. Known to be caused by a single genetic mutation— it’s triggered by an inherited gene, making it something researchers call a Mendelian disorder. Because of that, it’s a prime target for scientists working with technologies that edit specific parts of the genetic code. In a recent study, scientists took a step toward using what’s often referred to as the most revolutionary genetic technology in existence, CRISPR, to tweak the genes that cause Huntington’s.
The key is a Cas9 “nickase”—an enzyme that can cut one strand of DNA instead of both strands. The researchers paired it with CRISPR and tested it in cells taken from a Huntington’s patient. The gene-editing technique inactivated the huntingtin gene and cut off production of the neuron-destroying protein, they reported in the journal Frontiers in Neuroscience. However, as exciting as CRISPR is, it’s a long way from reality because of the potential for off-target gene editing.
6. Duchenne Muscular Dystrophy (DMD)
Known to be caused by mutations in the DMD gene, Duchenne Muscular Dystrophy encodes for a protein necessary for the contraction of muscles. Thus, children born with this disease suffer progressive muscular dystrophy, and there is currently no treatment available beyond palliative care. Today, scientists report they’ve halted the progression of the disease in some of those doggy descendants using the gene-editing tool known as CRISPR. Researchers have used CRISPR to treat Duchenne muscular dystrophy in four dogs, according to a study published yesterday (August 30, 2018) in Science. By editing cells in one-month-old beagles serving as models of the disease, the team boosted the expression of the gene coding for dystrophin—a protein whose dysfunction underlies Duchenne muscular dystrophy (DMD)—to up to 92 percent of normal levels in some tissues.
In the current study, Olson and his colleagues used viral vectors to deliver CRISPR directly to the muscles of two dogs that can not make functional dystrophin. They found that after six weeks of treatment, the animals produced the protein at around 60 percent of normal levels in some muscle fibers, and microscopic examination showed that muscle integrity had improved. When the team next administered the vectors into the bloodstream of another two dogs, the animal receiving the highest dose produced dystrophin at up to 70 percent of normal levels in skeletal muscle after eight weeks, and 92 percent in heart muscle. Anecdotally, the dogs “showed obvious signs of behavioral improvement—running, jumping—it was quite dramatic,” Olson tells Wired.
The work is still a long way from being applied to humans. The researchers are first planning to run longer-term canine trials, Olson tells Wired. “We just have to be really, really, really careful with this,” he says. “We don’t want to have any slip-ups from trying to move too quickly.”
7. Cystic Fibrosis (CF)
A common lethal, genetic disease, cystic fibrosis causes persistent lung infections and breathing difficulty. It is caused by mutations in the CF transmembrane conductance regulator (CFTR). Although there are treatments available to deal with the symptoms, the life expectancy for a person with this disease is only around 40 years. CRISPR-Cas9 is a promising innovation whose approach may represent a valid tool to repair the CFTR mutation, and hopeful results were obtained in tissue and animal models of CF disease. Trials in humans are just getting started, but hopes are high.
A leading biotech company in CRISPR technology, Editas Medicine’s approach relies on gene editing, in which the wrong DNA sequence of the defective CFTR gene is replaced by the correct one using the CRISP/Cas-9technology. CRISPR/Cas9 uses a protein-RNA complex composed of an enzyme (or protein) — Cas9 — bound to a guide RNA (gRNA) molecule that recognizes the wrong DNA sequence and cuts it out. The cell can then fill the excised portion with the correct gene sequence. The delivery mode is expected to be by adeno-associated virus (AAV) or lipid nanoparticle (LNP).
Thought Editas’ research in this CF treatment is still in the early stages. However, according to research, the use of CRISPR/Cas9 technology in a model of intestinal stem cell organoids from CF patients. The defective CFTR gene was replaced, and CFTR function restored, demonstrating the potential use of this technology to correct mutated CFTR genes.
8. Genetic Blindness
Exhibiting the potential to improve conditions for patients with rare diseases, it’s not unlikely that CRISPR could also restore retinal function such as hereditary forms of blindness or the degenerative disease retinitis pigmentosa. By using mouse models, it was reported that the researchers have successfully applied CRISPR technology to a dominantly inherited disease for the first time. For many hereditary forms of blindness, the specific mutations causing the disease are known to make it easy to instruct CRISPR-Cas9 to target and modify that gene.
Historically, autosomal dominant retinitis pigmentosa and other similar diseases have presented a unique challenge to researchers because only one copy of a mutated gene is inherited from an individual’s parents, and the individual has one normal gene on a pair of autosomal chromosomes. The challenge lies in the desire to edit only the mutant copy of a gene without affecting the healthy one. A strategy, however, designed by Dr. Tsang and colleagues could potentially cut out the old gene and replace it with a good gene, without affecting its normal function. The “ablate-and-replace” strategy can be used to develop CRISPR toolsets for all types of mutations that reside in the same gene and is not exclusive for a type of mutation.
Editas Medicine is working on a CRISPR therapy for Leber Congenital Amaurosis, the most common cause of inherited childhood blindness, for which there is no treatment. The company aims to target the most common mutation behind the disease, using CRISPR to restore the function of the photoreceptor cells before the children lose sight completely.
9. Blood disorders
The first CRISPR trial in Europe will seek to treat Beta-thalassemia. A second trial will test the same therapy in the treatment of sickle cell diseases, another blood disorder affecting oxygen transportation. However, the FDA has put the US trial on hold to clear some safety questions before going ahead. Hemophilia is another blood disorder that CRISPR technology could tackle. CRISPR Therapeutics is working with Casebia on an in vivo CRISPR therapy where the gene-editing tool is delivered directly to the liver.
Beta-thalassemia is a blood disorder that affects oxygen transport in the blood. A therapy developed by CRISPR Therapeutics and Vertex Pharmaceuticals consists in harvesting hematopoietic stem cells from the patient and using CRISPR technology to make them produce fetal hemoglobin, a natural form of the oxygen-carrying protein that binds oxygen much better than the adult form.