The Many Uses of CRISPR: Scientists Tell All

Smartphones, superglue, electric cars, video chat. When will the magic of a new technology wear off? When you are so used to its presence that you no longer think about it? When something newer and better comes out? When you forget what it was like before?

Whatever the answer, the gene-editing technology CRISPR hasn’t reached that point yet. Ten years after Jennifer Doudna and Emmanuelle Charpentier first introduced the discovery of CRISPR, it remains at the heart of ambitious scientific projects and complex ethical discussions. It continues to create new avenues for discovery and reviving old studies. Biochemists use it, and so do other scientists: entomologists, cardiologists, oncologists, zoologists, botanists.

For these researchers, some magic is still there. But the excitement of complete novelty has been replaced by open possibilities and ongoing projects. Here are a few of them.

Cathie Martin, a botanist at the John Innes Center in Norwich, England, and Charles Xavier, founder of the superhero team X-Men: Both love mutants.

But while Professor X has an affinity for human superpowered mutants, Dr. Martin prefers the red and succulent kind. “We’ve always been hungry for mutants, because that allows us to understand function,” says Dr Martin, focusing on plant genomes in the hope of figuring out how to make plants. Products – especially tomatoes – are healthier, stronger and longer. lasting.

When CRISPR-Cas9 came out, one of Dr. Martin’s colleagues offered to make her a mutant tomato as a gift. She was a little skeptical, but she told him, “I’d rather a tomato that doesn’t produce chlorogenic acid,” a substance believed to have health benefits; tomatoes without it have not been found before. Dr. Martin wants to remove what she believes is the key gene sequence and see what happened. Soon after, a tomato without chlorogenic acid was introduced into her lab.

Instead of looking for mutants, it was now possible to create them. “Capturing those mutants was so effective and amazing, because it gave us confirmation of all the hypotheses we had,” said Dr Martin.

Most recently, researchers at Dr. Martin’s lab used CRISPR to create a tomato plant that can accumulate vitamin D when exposed to sunlight. Just one gram of leaves contains 60 times the recommended daily value for adults.

Dr. Martin explains that CRISPR can be used in a wide range of food modifications. It has the ability to remove allergens from nuts and create plants that use water more efficiently.

“I don’t claim that what we did with vitamin D will solve any food insecurity problem,” says Dr. Martin, “but that is just one example. People like to have something they can cling to, and this is there. It’s not a promise.”

Infectious Diseases

Bringing testing to remote parts of Africa

Christian Happi, a biologist who directs the African Center of Excellence for the Genomics of Infectious Diseases in Nigeria, has dedicated his career to developing methods to detect and prevent the spread of infectious diseases. infectious diseases transmitted to humans from animals. Many existing ways to do so are expensive and imprecise.

For example, to do a polymerase chain reaction, or PCR, test, you need to “go get an RNA extraction, have a $60,000 machine, and hire someone with special training,” says Dr. Happi. Carrying this type of experiment to most remote villages is both logistically expensive and not feasible.

Recently, Dr. Happi and his colleagues used CRISPR-Cas13a technology (a close relative of CRISPR-Cas9) to detect diseases in the body by targeting pathogen-associated gene sequences. . They were able to decode the SARS-CoV-2 virus within weeks of the pandemic’s arrival in Nigeria and developed an test that required no on-site equipment or trained technicians – just a spittoon.

Dr Happi said: “If you’re talking about the future of pandemic preparedness, that’s what you’re talking about. “I want my grandmother to use this in her village.”

The CRISPR-based diagnostic test performs well under thermal conditions, is fairly easy to use, and costs one-tenth the cost of a standard PCR test. However, Dr Happi’s lab is constantly evaluating the technology’s accuracy and trying to convince leaders in the public health system in Africa to accept it.

He calls their proposal “cheaper, faster, equipment-free and can be pushed into the furthest corners of the continent. This will allow Africa to occupy what I call its natural space. “

Genetic diseases

Searching for a cure for sickle cell disease

Originally there was zinc finger nuclease.

It’s the gene editing tool that Gang Bao, a biochemical engineer at Rice University, first used to treat sickle cell disease, a genetic disorder marked by red blood cells. malformation. Bao’s lab took more than two years of development, and then the zinc finger nuclease was able to successfully cut sickle cell chains only about 10% of the time.

Another technique takes two more years and is only slightly more effective. And then, in 2013, shortly after CRISPR was used to successfully edit genes in living cells, Dr. Bao’s team changed the method again.

“Since we started getting some initial results, CRISPR has taken us a whole month,” said Dr. Bao. This method successfully cut the target sequence about 60 percent of the time. It’s easier to do and more efficient. “It was wonderful,” he said.

The next challenge is to identify the side effects of this process. That is, how did CRISPR affect genes that were not intentionally targeted? After a series of animal tests, Dr. Bao believes the method will work in humans. In 2020, the Food and Drug Administration approved an ongoing clinical trial, led by Dr Matthew Porteus and his lab at Stanford University. And there’s also hope that with CRISPR’s versatility, it could be used to treat other genetic diseases. At the same time, other treatments that do not rely on gene editing have brought success for sickle cell disease.

Dr. Bao and his lab are still trying to identify all the side effects and grade three of using CRISPR. But Dr. Bao is optimistic that there will soon be an effective and safe gene-editing treatment for sickle cell disease. How soon? “I think three to five years from now,” he said, smiling.

Heart

Looking into the secret of the heart

It’s hard to change someone’s heart. And it’s not just because we’re often stubborn and stuck in our path. The heart makes new cells at a much slower rate than many other organs. Treatments that work for other parts of the human body are often much harder for the heart.

It’s also hard to know what’s in someone’s heart. Even if you sequence the entire genome, there are often some segments that remain a mystery to scientists and doctors (known as variants of uncertain significance). A patient may have heart disease, but there is no way to definitively link it back to their genes. “You’re stuck,” said Dr. Joseph Wu, director of the Stanford Heart Institute. “So traditionally we would just wait and tell the patient that we don’t know what’s going on.”

But over the past few years, Dr. Wu has been using CRISPR to see what effect the presence and absence of these surprising sequences have on heart cells, simulated in his lab with stem cells. Induced pluripotent stem cells are generated from blood. By cutting out specific genes and observing the effects, Dr. Wu and his colleagues were able to draw a link between individual patients’ DNA and heart disease.

It will be a long time before these diseases can be treated with CRISPR, but diagnosis is the first step. “I think this will have a big impact in terms of personalized medicine,” said Dr. Wu, who mentioned that he found at least three variants of uncertain significance when sequencing the genome. own genes. “What do these variations mean to me?”

Sorghum is used in bread, wine and cereals around the world. But it’s not commercially processed to the same extent as wheat or corn, and when it’s made, it’s often not as tasty.

Karen Massel, a biotechnologist at the University of Queensland in Australia, saw plenty of room for improvement when she first started studying the plant in 2015. And that’s because millions of people eat it. sorghum around the world, “if you make one small change, you can have a huge impact,” she said.

She and her colleagues used CRISPR to try to make sago plants frost-resistant, make them more heat-tolerant, prolong growth, change root structure — “we used gene editing. on a large scale,” she said.

Not only could this lead to tastier and healthier grains, she said, but it could also make crops more resilient to a changing climate. But precisely editing the genome of plants with CRISPR is still no small task.

“Half of the genes that we remove, we don’t know what they do,” says Dr. Massel. “The second we tried to get in there and play God, we realized we were a little lost.” However, using CRISPR in conjunction with more traditional breeding techniques, Dr. Massel remains optimistic, despite describing himself as a pessimist. And she hopes that further advances will lead to the commercialization of genetically modified foods, making them more accessible and acceptable.

In 2012, a 6-year-old girl developed acute lymphoblastic leukemia. Chemotherapy was unsuccessful and the case was too severe for a bone marrow transplant. There didn’t seem to be any other option, and the girl’s doctors told her parents to return home.

Instead, they went to Children’s Hospital of Philadelphia, where doctors used an experimental treatment called chimeric antigen receptor (CAR) T-cell therapy to turn the girl’s white blood cells against cancer again. Ten years later, the girl recovered from cancer.

Since then, Dr. Carl June, a professor of medicine at the University of Pennsylvania who helped develop CAR T-cell therapy, and his collaborators, including Dr. Ed Stadtmauer, a hematologist -oncology at Penn Medicine, has been working to improve it. That includes using CRISPR, the simplest and most precise tool for editing T cells outside the body. Dr. Stadtmauer, who specializes in the treatment of cancers of the lymphatic and blood systems, said, “Over the past decade or so, we have seen a revolution in the treatment of these diseases; it’s very useful and interesting. “

Over the past few years, Dr. Stadtmauer has helped conduct a clinical trial in which T cells that had undergone significant CRISPR editing were introduced into treatment-resistant cancer patients. Promising results.

“Patients with a very poor prognosis are now doing much better and some are being cured,” said Dr. Stadtmauer. He continued to monitor the patients and found that the edited T cells remained in the blood, ready to attack tumor cells in the event of a relapse.

The real benefit is that scientists now know that CRISPR-assisted treatments are possible.

“Although it really falls into the category of science fiction-biochemistry and science, the reality is that the field has changed a lot,” said Dr. Stadtmauer. He added that he’s less excited about science than how useful CRISPR has become. “Every day I can see 15 patients who need me,” he says. “That’s what motivates me.”