The Many Uses of CRISPR: Scientists Tell All

Smartphones, superglue, electric cars, video chat. When will the wonders of new technology disappear? What if you get used to its existence and can’t think of it anymore? When newer and better things come? What if I forget the previous situation?

Whatever the answer, the gene editing technique CRISPR has not yet reached that point. Ten years after Jennifer Doudna and Emmanuelle Charpentier first introduced the discovery of CRISPR, it continues to be the center of ambitious scientific projects and complex ethical debates. It continues to create new paths for quest and rejuvenate old research. Biochemists use it, as do other scientists: entomologists, cardiologists, oncologists, zoologists, botanists.

For these researchers, some of the wonders are still there. But the excitement of something entirely new has been superseded by open possibilities and ongoing projects. Here are some of them.

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

However, Professor X has an affinity for super-strong human mutants, while Dr. Martin is partly in the red and juicy type. “We were always anxious for mutants because we could understand the functionality,” said Dr. Martin, who made foods, especially tomatoes in her case, healthier, stronger, and longer. Sustained she mentioned her research focusing on the plant genome in the hope of finding a way to do it.

When CRISPR-Cas9 appeared, one of Dr. Martin’s colleagues offered her to make a mutant tomato as a gift. She was a little skeptical, but she “wants a tomato that doesn’t produce chlorogenic acid,” she told him. This is a substance that is considered good for your health. Tomatoes without it have never been found. Dr. Martin wanted to get rid of what she believed was an important gene sequence and see what happened. Soon there was a chlorogenic acid-free tomato in her lab.

Instead of looking for mutants, it is now possible to create them. “Obtaining these mutants was very efficient and very nice, as it gave us confirmation of all these hypotheses we had,” said Dr. Martin. ..

Recently, researchers in Dr. Martin’s lab have used CRISPR to create tomato plants. Can accumulate vitamin D When exposed to sunlight. Only 1 gram of leaves contained 60 times the recommended daily intake for adults.

Dr. Martin explained that CRISPR can be used for a wide range of food improvements. It has the potential to remove allergens from nuts and create plants that use water more efficiently.

“I’m not saying that what we did with vitamin D solves the problem of food insecurity,” said Dr. Martin. “But that’s just a good example. People like to have something they can cling to, and this is there. It’s not a promise.”

Infection

Bringing tests to remote parts of Africa

Christian Happi, a biologist who heads the Africa Center of Excellence for Genomics of Infectious Diseases in Nigeria, has spent his career developing ways to detect and contain the spread of infectious diseases from animals to humans. Many of the existing methods for doing so are costly and inaccurate.

For example, to perform a polymerase chain reaction (PCR) test, you need to “extract RNA, have a $ 60,000 machine, and hire a specially trained person.” Bringing this kind of test to most remote villages is costly and logistically unbelievable.

Recently, Dr. Happi and his collaborators used CRISPR-Cas13a technology, a closely related species of CRISPR-Cas9, to detect diseases in the body by targeting gene sequences associated with pathogens. .. They sequenced the SARS-CoV-2 virus within a few weeks of the pandemic in Nigeria, did not require on-site equipment or trained technicians, only tubes for spitting. I was able to develop the tests I needed.

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

CRISPR-based diagnostic tests work well in the heat, are extremely easy to use, and cost one-tenth the cost of standard PCR tests. Still, Dr. Happi’s lab is continually assessing the accuracy of the technology and trying to convince leaders of the African public health system to adopt it.

He called their proposal “cheaper, faster, no equipment required, and can be pushed into the farthest corners of the continent.” This allows Africa to occupy what I call its natural space. “

Hereditary disease

Find a cure for sickle cell disease

Initially, there was a zinc finger nuclease.

It was the first genetic editing tool used by Gangbao, a biochemical engineer at Rice University, to treat sickle cell disease, a hereditary disease characterized by malformed red blood cells. Development in Dr. Bao’s laboratory took more than two years, but zinc finger nucleases were able to cleave sickle cell sequences for only about 10% of the time.

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

“CRISPR took about a month from the beginning to the first results,” said Dr. Bao. This method successfully cut the target sequence in about 60% of the time. It was easy to create and more effective. “It was just great,” he said.

The next challenge was to identify the side effects of the process. So how did CRISPR affect genes that weren’t intentionally targeted? After a series of experiments on animals, Dr. Bao was convinced that this method would also work for humans. In 2020 Food and Drug Administration Approves Clinical Trials, Leaded by Dr. Matthew Porteus and his laboratory at Stanford University, it is underway. Also, due to the versatility of CRISPR, it may be used to treat other hereditary disorders. At the same time, other therapies that do not rely on gene editing have been successful for sickle cells.

Dr. Bao and his lab are still trying to determine all the secondary and tertiary effects of using CRISPR. However, Dr. Bao is optimistic that safe and effective gene-editing therapies for sickle cells will soon be available. How fast? “I think it’s three to five years left,” he said with a laugh.

Cardiology

Look into the secret of your heart

It’s hard to change someone’s mind. And that’s not just because we’re often stubborn and stuck. The heart produces new cells at a much slower rate than many other organs. Effective treatments for other parts of the human body are much more difficult for the heart.

It’s also difficult to know what’s in someone’s heart. Even when sequencing the entire genome, there are often many segments that remain mysterious to scientists and physicians (called variants of uncertain importance). Patients may have heart disease, but there is no clear way to link it to their genes. “You are stuck,” said Dr. Joseph Wu, director of the Stanford Cardiovascular Institute. “Traditionally, we just waited and told the patient that we didn’t know what was going on.”

But for the past two years, Dr. Wu has used CRISPR to see how the presence or absence of these disrupted sequences affects heart cells. In his laboratory, we simulated using induced pluripotent stem cells generated from blood.By cutting out specific genes and observing their effects, Dr. Wu and his collaborators Draw a link Between individual patient DNA and heart disease.

It will take a long time before these diseases can be treated with CRISPR, but diagnosis is the first step. “I think this will have a huge impact from a personalized medicine perspective,” said Dr. Wu, who discovered at least three mutants of uncertain importance when determining his own genomic sequence. Said. “What do these variants mean to me?”

Sorgham is used in bread, alcohol and cereals around the world. However, it is not as commercially designed as wheat or corn, and when processed, it is often not very tasty.

Karen Massell, a bioengineer at the University of Queensland, Australia, thought there was considerable room for improvement when she began her research on plants in 2015. “Millions of people around the world eat sorghum, so making small changes can have a big impact,” she said.

She and her colleagues used CRISPR to attempt to change the frost resistance, heat resistance, growth period, and root structure of sorghum. “We use gene editing entirely,” she said.

This not only leads to more delicious and healthy grains, but also has the potential to increase the number of plants Resistant to changing climate, She said. However, accurately editing the crop genome with CRISPR is not yet a small task.

“Half of the genes we knocked out have no idea what they’re doing,” Dr. Massell said. “When we go in and try to play God, we find that we are a little out of our depth.” But when we use CRISPR in combination with more traditional breeding techniques , Dr. Massell is optimistic despite being a self-proclaimed 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 suffered from acute lymphoblastic leukemia. She had unsuccessful chemotherapy and the case was too advanced for her bone marrow transplant. There seemed to be no other choice, and the girl’s doctor told her parents to go home.

Instead, they went to the Philadelphia Children’s Hospital, where doctors used an experimental treatment called chimeric antigen receptor (CAR) T cell therapy to counteract the white blood cells of the girl against cancer. Ten years later Girl has no cancer..

Since then, his collaborators, including Dr. Carl June, a professor of medicine at the University of Pennsylvania who helped develop CAR T cell therapy, and Dr. Ed Stadtmauer, a hematologist-oncologist at Penn Medicine, have been working to improve it. I did. This includes the use of CRISPR, the simplest and most accurate tool for editing in vitro T cells. Dr. Stadtmauer, who specializes in the treatment of cancers of various blood types and lymphatic systems, said: It’s rewarding and exciting. “

For the past few years, Dr. Stadtmauer has been Clinical trials T cells that underwent significant CRISPR editing were inserted into patients with refractory cancer. The result was promising.

“Patients with a very dire prognosis are now in much better condition and some have healed,” said Dr. Stadtmauer. He continued to monitor the patient and found that the edited T cells were still present in the blood and were ready to attack the tumor cells in case of recurrence.

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

“It’s really a kind of science fiction novel-like biochemistry and science, but in reality the field is tremendously moving,” said Dr. Stadtmauer. He added that he was less excited about science than how useful CRISPR was. “I see 15 patients in need of me every day,” he said. “That’s what motivates me.”