What is CRISPR-Cas9?

CRISPR-Cas9 is a genome editing tool thats able to cut DNA in a targeted fashion, allowing scientists to accurately edit the building blocks of life.

It was actually first observed in the 1980s as part of single-celled bacterias defence mechanisms, which ensure that the cells are able to remove unwanted intruders. Scientists have found that, by adapting the technology, they are able to target genome sequences with unprecedented speed, precision and accuracy.

Picture CRISPR-Cas9 as like a find and replace search in a computer document, only instead of words, youre editing genetic sequences.

Accurately modifying DNA is a scientific holy grail, and the potential is enormous. It could be used to eradicate diseases even hereditary ones such as cystic fibrosis, sickle-cell anemia and Huntington's could become a thing of the past.

The name CRISPR is an acronym for the less catchy clustered regularly interspaced short palindromic repeats. The Cas part refers to CRISPR associated.

CRISPR is part of certain bacterias naturally occurring defences. When a bacteria detects an invading virus, it is able to copy and blend segments of the foreign DNA into its own genome around CRISPR.

The next time the virus is spotted, CRISPR has an exact copy of the genome sequence to look out for, which is where the Cas protein comes in: it can cut the DNA up, and disable unwanted genes with incredible accuracy.

Or, as Carl Zimmer explains: As the CRISPR region fills with virus DNA, it becomes a molecular most-wanted gallery, representing the enemies the microbe has encountered. The microbe can then use this viral DNA to turn Cas enzymes into precision-guided weapons. The microbe copies the genetic material in each spacer into an RNA molecule. Cas enzymes then take up one of the RNA molecules and cradle it. Together, the viral RNA and the Cas enzymes drift through the cell. If they encounter genetic material from a virus that matches the CRISPR RNA, the RNA latches on tightly. The Cas enzymes then chop the DNA in two, preventing the virus from replicating.

In 2012, scientists from the University of California, Berkeley, published a groundbreaking paper showing they were able to reprogramme the CRISPR-Cas immune system to edit genes at will. CRISPR-Cas9 uses a specific Cas protein and a hybrid RNA that can identify and edit any gene sequence. The possibilities are huge.

In short, CRISPR lists the DNA sequences to target, and then Cas9 does the cutting. Scientists just need to programme CRISPR with the right code, and Cas9 does the rest.

This could also apply to faulty genes sections currently causing problems could be removed with CRISPR-Cas9, and then replaced with healthy genetic code, theoretically solving the problem.

CRISPR is cutting edge technology, but while its true that its use has massively accelerated in recent years thanks to the above discovery, scientists have actually been aware of it in bacteria since the 1980s. Pubmed lists 5,775 papers discussing CRISPR but 5,575 of those have been in the three years since the UC Berkeley paper, and the number has jumped from 2,071 when I first wrote this article back in October 2015.

CRISPR-Cas9 isnt the first genomic editor, but it has a number of upsides that make it both simpler and far more efficient.

Firstly, CRISPR-Cas9 can edit multiple genes at once, whereas other genome editors such as zinc finger nuclease (ZFN) or transcription activator-like effector nucleases (TALENs) require painstaking modification of a single gene at a time. Its also quicker and cheaper, as you might expect.

Although ZFN and TALENs can recognise longer gene sequences than CRISPR-Cas9, custom proteins have to be created each time and its an inexact science, involving the creation of several variants before the winning combination is found.

On top of that, scientists tend to use ZFN and TALENs with organisms scientists know extremely well such as mice, rats and fruit flies. CRISPR-Cas9 should work with every organism ever evolved. Yes, including humans.

Yes, in China. Using human embryos sourced from a fertility clinic, scientists tried to use CRISPR-Cas9 to edit a gene that causes beta thalassemia in every cell. It should be noted that the donor embryos used were non-viable, and could not have resulted in a live birth.

In any case, it failed, and failed quite badly: 86 embryos were injected, and after 48 hours and around eight cells grown, 71 survived, and 54 of those were genetically tested. Just 28 had been successfully spliced, and very few contained the genetic material the researchers intended. If you want to do it in normal embryos, you need to be close to 100%, lead researcher Jungiu Huang told Nature. Thats why we stopped. We still think its too immature.

On top of that, its extremely likely more undocumented damage was done. As the New York Times explains: The Chinese researchers point out that in their experiment gene editing almost certainly caused more extensive damage than they documented; they did not examine the entire genomes of the embryo cells.

As you might imagine, it caused a huge amount of controversy in the scientific community.

In November 2016, another grouip of Chinese scientists became the first to use CRISPR-Cas9 on an adult human, injecting a lung cancer sufferer with the patient's immune cells modified by CRISPR to disable the PD-1 protein, theoretically making the patient's body fight back against the cancer. Results are still yet to be reported. The first American trial of CRISPR in humans is due to take place at the University of Pennsylvania later this year again with cancer.

Even though the Chinese scientists used embryos that were not going to develop into life, there are real ethical concerns about experimenting on human embryos indeed, just a month before the Chinese research was published, a group of American scientists urged the world not to do so.

Part of this comes down to how immature the technology is remember that its only been in active use since 2012, and it would be astonishing if it was fully matured at this point. Scientists warned that it was too misunderstood and dangerous to use on humans at this point, and the Chinese research certainly vindicates this concern. Even if it worked flawlessly, there are concerns that unforeseen consequences could occur over generations.

But, even if it were 100% safe and successful, there are other ethical concerns: while nobody argues that we should hold back the potential of wiping out killer genetic diseases such as Huntingtons and cystic fibrosis, CRISPR-Cas9 potentially offers the opportunity to change anything about a person. As long as the genetic sequence is identified, in theory, it can be edited.

Its one thing to remove life-impacting diseases before birth its quite another for parents to be able to design their babies to be stronger, faster or better looking. Even if you accept that this is something people should be allowed to do, the chances are this would be heavily commercialised, ensuring only the rich could afford all the extra life advantages this would afford, massively affecting inequality.

Of course, these ethical questions are a million miles away when the only recorded embryonic human experiment was such a high-profile set-back. However, CRISPR-Cas9 is now showing extremely promising results in smaller tests.

Examples include HIV infection prevention in human cells, curing genetic mouse diseases and a pair of monkeys born with targeted mutations. As Wired says, it "kills HIV and eats Zika like Pac-man," with hopes that cancer could be the next disease in its sights.

Yes. Stem cell researchers in the UK sought permission to modify human embryos in an attempt to understand early human development, and reduce the likelihood of miscarriage. In February 2016, theHuman Fertilisation and Embryology Authority (HFEA) granted permission.

As mentioned previously, Cas9 can only recognise genetic sequences of around 20 bases long, meaning that longer sequences cannot be targeted.

More significantly, the enzyme still sometimes cuts in the wrong place. Figuring out why this is will be a significant breakthrough in itself fixing it will be even bigger.

Then, of course, theres the issue that CRISPR didnt work terribly well in human embryos. Scientists need to discover what went wrong there, and what the difference is between the success in single cells and the more patchy results with embryos.

That isnt a simple question to answer. Its subject to an ongoing patent battle surprisingly, given CRISPR is naturally occurring in certain bacteria.

Technology Review explains that, although CRISPR-Cas9 was first described in Science in 2012 by Jennifer Doudna from UC Berkeley, Feng Zhang from the Broad Institute won a patent on the technique by submitting lab notebooks proving hed invented it first.

First to file patent rights means that this should be granted to Doudna, but the decision could have been decided based on first to invent rules, which would have favoured Zhang. In the end, the case was resolved in February 2017, when the US Patent Trial and Appeal Board resolved that UC Berkeley would be granted the patent for the use of CRISPR-Cas9 in any living cell, while Broad would get it in any eukaryotic cell which is to say cells in plants and animals.

Images: Petra B Fritz, VeeDunn, NIH Image Gallery, and Steve Jurvetson used under Creative Commons

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What is CRISPR-Cas9, and will it change the world? | Alphr - Alphr

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