The Many Uses of CRISPR: Scientists Tell It All

Smartphones, superglue, electric cars, video chat. When does the wonder of a new technology disappear? When do you get so used to his presence that you no longer think about him? When will something new and better come out? When you forget how things used to be?

Whatever the answer, CRISPR gene-editing technology has yet to reach that level. ten years later Jennifer Doudna and Emmanuelle Charpentier first presented their discovery of CRISPR, it remains at the center of ambitious scientific projects and complex ethical discussions. It continues to create new research opportunities and revitalize old research. It is used by biochemists, as well as other scientists: entomologists, cardiologists, oncologists, zoologists, botanists.

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

But while Professor X has an affinity for superpowered human mutants, Dr. Martin is not indifferent to the red and juicy type. “We have always coveted mutants because it allowed us to understand functionality,” says the doctor. Martin spoke about her research, which focuses on plant genomes in the hope of finding ways to make foods, especially tomatoes, in her case, healthier, stronger and more durable.

When CRISPR-Cas9 appeared, one of Dr. Martin’s colleagues offered to give her a mutant tomato as a gift. She was somewhat skeptical, but told him, “I’d really like a tomato that doesn’t produce chlorogenic acid,” a substance considered to be beneficial to health; tomatoes have never been seen without it before. Dr. Martin wanted to delete what she thought was a key gene sequence and see what happens. Soon, a tomato without chlorogenic acid appeared in her laboratory.

Instead of looking for mutants, it was now possible to create them. “Getting these mutants was so effective and so wonderful because it gave us confirmation of all our hypotheses,” says the doctor. Martin said.

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

Dr. Martin explained that CRISPR can be used in a wide range of food modifications. This could potentially remove nut allergens and create plants that use water more efficiently.

“I am not saying that what we have done with vitamin D will solve any food safety problems,” says the doctor. Martin said, “But this is just a good example. People like to have something they can hold on to, and that’s it. This is not a promise.”

Infection

Christian Happi, a biologist who heads the African Center of Excellence for Infectious Disease Genomics in Nigeria, has dedicated his career to developing methods to detect and contain the spread of infectious diseases transmitted to humans from animals. Many of the existing ways to do this are costly and inaccurate.

For example, to do a polymerase chain reaction, or PCR, you need to “extract RNA, have a $60,000 machine, and hire someone who is specially trained,” says Dr. Wilson. Happy said. Conducting such tests in the most remote villages is both expensive and logistically impractical.

Recently dr. Happy and his collaborators used CRISPR-Cas13a technology (a close relative of CRISPR-Cas9) to detect diseases in the body by targeting genetic sequences associated with pathogens. They were able to sequence the SARS-CoV-2 virus within weeks of the pandemic arriving in Nigeria and develop a test that required no on-site equipment or trained technicians, just a saliva tube.

“If you talk about the future of pandemic preparedness, that’s what you mean,” the doctor said. Happy said. “I would like my grandmother to use this in her village.”

The CRISPR-based diagnostic test works well in hot weather, is fairly easy to use, and costs one-tenth the cost of a standard PCR test. However, dr. Happi Labs is constantly evaluating the accuracy of the technology and trying to convince leaders in African health systems to adopt it.

He called their proposal one that is “cheaper, faster, requires no equipment, and can be delivered to the most remote corners of the continent.” This would allow Africa to occupy what I call its natural space.”

hereditary disease

In the beginning, there was zinc finger nuclease.

It was a gene-editing tool that Gan Bao, a biochemical engineer at Rice University, first used to treat sickle cell anemia, an inherited disease characterized by misshapen red blood cells. It took dr. Bao’s lab worked on the development for more than two years, and then zinc finger nuclease successfully excised the sickle cell sequence only about 10 percent of the time.

The other method took another two years and was only slightly more effective. And then, in 2013, shortly after CRISPR was successfully used to edit genes in living cells, Dr. Bao’s team changed tactics again.

“From the very beginning to the first results, CRISPR took us about a month,” says the doctor. Bao said. The method successfully cuts the target sequence in about 60% of cases. It was easier to do and more efficient. “It was just amazing,” he said.

The next task was to determine the side effects of the process. That is, how did CRISPR affect genes that were not targeted? After a series of experiments on animals, D. Bao was convinced that this method would work for humans as well. In 2020 FDA approves clinical trial, led by Dr. Matthew Porteous and his lab at Stanford University, it’s going on. It is also hoped that, due to the versatility of CRISPR, it could be used to treat other hereditary diseases. At the same time, other therapies that are not based on gene editing has been successful for sickle cell.

Dr. Bao and his lab are still trying to determine all the secondary and tertiary effects of using CRISPR. But dr. Bao is optimistic that a safe and effective treatment for sickle cell anemia through gene editing will soon be available. How soon? “I think another three to five years,” he said, smiling.

Cardiology

It’s hard to change someone’s heart. And this is not only because we are often stubborn and get stuck in our path. The heart generates new cells much more slowly than many other organs. Treatments that are effective in other parts of the human anatomy are much more difficult for the heart.

It is also difficult to understand what is in a person’s heart. Even when you sequence an entire genome, there are often a number of segments that remain a mystery to scientists and doctors (so-called undetermined variants). The patient may have heart disease, but there is no way to definitively link it to his genes. “You are stuck,” said the doctor. Joseph Wu, director of the Stanford Institute for Cardiovascular Diseases. “So traditionally we just waited and told the patient we didn’t know what was going on.”

But over the past couple of years, Dr. Wu used CRISPR to see what effect the presence and absence of these confusing sequences had on heart cells modeled in his lab with blood-derived induced pluripotent stem cells. By cutting out certain genes and observing the effects, Dr. Wu and his collaborators were able to draw links between the DNA of individual patients 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 on personalized medicine,” the doctor said. Wu, who mentioned that he found at least three variants of undetermined significance when he sequenced his own genome. “What do these options mean to me?”

Sorghum is used in bread, alcohol and cereals throughout the world. But it hasn’t been industrially modified to the same extent as wheat or corn, and once processed, it often doesn’t taste as good.

Karen Mussel, a biotechnologist at the University of Queensland in Australia, saw plenty of room for improvement when she first began studying the plant in 2015. And since millions of people around the world eat sorghum, “if you make a small change, you can make a huge impact,” she said.

She and her colleagues have used CRISPR to try to make sorghum tolerant to frost, to make it more tolerant to heat, to extend its growth period, to change its root structure—“we use gene editing all over the place,” she said.

This can not only lead to tastier and healthier cereals, but also make plants healthier. climate resilient, she said. But accurately editing crop genomes with CRISPR is no easy task.

“Half of the genes that we turn off, we just don’t know what they do,” says the doctor. Massel said. “The second we try to get in there and play God, we realize we’re a little crazy.” But using CRISPR in combination with more traditional breeding methods, Dr. Massel is optimistic, despite the fact that he calls himself a pessimist. And she hopes that further advances will lead to the commercialization of gene-altered products, making them more accessible and acceptable.

In 2012, a 6-year-old girl suffered from acute lymphoblastic leukemia. Chemotherapy was unsuccessful and the case was too advanced for a bone marrow transplant. There seemed to be no other options, and the girl’s doctors advised her parents to return home.

Instead, they went to the 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. Ten years later girl got rid of 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-oncologist at Penn Medicine, are working to improve it. This includes the use of CRISPR, which is the simplest and most accurate tool for editing T cells outside the body. Dr. Stadtmauer, who specializes in treating various types of cancers of the blood and lymphatic system, said that “there has been a revolution in the treatment of these diseases in the last decade or so; it was helpful and interesting.”

Over the past couple of years Dr. Stadtmauer helped launch clinical trial in which T cells heavily edited by CRISPR were inserted into patients with treatment-resistant cancer. The results were promising.

Nine months into the trial, the edited T cells were not rejected by the patients’ immune systems and were still present in the blood. The real benefit is that scientists now know that CRISPR treatment is possible.

“Despite the fact that it’s actually kind of science fiction, biochemistry and science, the reality is that this field has advanced a lot,” says the doctor. Stadtmauer said. He added that he was not so much concerned with the science as with the usefulness of CRISPR. “Every day I see about 15 patients who need me,” he said. “That’s what motivates me.”