To be honest I had hard time falling asleep yesterday. The news about winning a scholarship from Lithuanian Council for Culture brought me so much excitement. I already started to collect the data for my artist research "Chimeras of nanotechnologies: the end of the sci-fi genre". The project will be developed during summer 2021.
Chimera is a spooky word. Playing with DNA is spooky too. I would like to shed some light on the subject, first of all, for myself, and eventually for those who will dig into my research. So let's start.
What does the word "chimera" mean? A genetic chimerism or chimera is a single organism composed of cells with more than one distinct genotype. For example, two-colored rose chimera.
Genotype. A genotype is an organism’s complete set of genetic material. Often though, genotype is used to refer to a single gene or set of genes, such as the genotype for eye color.
In order to manipulate DNA, there are several methods. One being for example CRISPR, and other - DNA origami.
1. EDITING DNA
CRISPR is a technology that can be used to edit genes and, as such, will likely change the world. The essence of CRISPR is simple: it's a way of finding a specific bit of DNA inside a cell. After that, the next step in CRISPR gene editing is usually to alter that piece of DNA. CRISPR has made it cheap and easy.
EDITING DNA - CRISPR - ‘GENETIC SCISSORS’
What is Crispr and why did it win the Nobel prize?
BY JAMIE DURRANI
7 OCTOBER 2020
The science behind the prize-winning gene editing tool that could change our lives
Emmanuelle Charpentier and Jennifer Doudna have scooped the 2020 Nobel prize in chemistry ‘for the development of a method for genome editing’. Specifically, they’ve been awarded the prize for their discovery of the Crispr–Cas9 genome editing technique that allows scientists to make precise alterations to the genetic code of living organisms. Crispr–Cas9 is a powerful tool that could revolutionise many aspects of our lives, from medical treatments to the way we produce food. It’s also seen its fair share of controversy in recent years. Here, we take a deeper look at these ‘genetic scissors’ and why they’ve won the Nobel prize.
Why did it win?
Since Charpentier and Doudna began investigating the Crispr–Cas9 system in 2011, the field has exploded. Due to the relative simplicity and affordability of Crispr systems, researchers around the world have been able to apply the tools to all manner of different problems. Today there are entire journals, conferences and companies dedicated to the technique.
The ability to cut any DNA molecule at a chosen site has huge potential – from treating genetic illnesses to creating disease-resistant crops. Trials have even shown how Crispr-delivered genetic modifications can spread through populations of mosquitoes and stop malaria infections – such ‘gene-drives’ offer a way to eliminate the disease altogether. And in the face of the Covid-19 pandemic, researchers have found ways to use Crispr in rapid coronavirus diagnostic tests and have also proposed using it to attack the virus’s genome.
As Claes Gustafsson, chair of the Nobel committee for chemistry, said at the award announcement, ‘There is enormous power in this genetic tool, which affects us all.’
CLUSTERED REGULARLY INTERSPACED SHORT PALINDROMIC REPEATS
How does it work?
The whole Crispr gene editing tool has been adapted from the immune system of bacteria. The term Crispr comes from ‘clustered regularly interspaced short palindromic repeats’, which refers to distinct genetic sequences found in the genomes of bacteria. Each Crispr sequence is transcribed into RNA sequences that will target the DNA of a virus. These sequences also include cas (Crispr-associated) genes that code for DNA-cutting Cas enzymes. Together, the guide RNA and Cas enzyme form a complex that hunts out viral DNA and chops it up.
In Crispr gene editing, scientists repurpose this system by designing a guide RNA sequence of around 20 nucleobases that matches up to a DNA sequence they wish to target in a cell’s genome. This RNA sequence is paired with the Cas9 enzyme that will cut the DNA strand at the targeted site. The whole DNA sequence coding for both these components of the Crispr-Cas9 tool can be delivered to the target cell via a plasmid.
Source: © Jonathan Jarnstead/The Royal Swedish Academy of Sciences
The guide RNA sequence brings the DNA cleaving Cas9 enzyme to a specific spot in a cell’s genome, where the enzyme then makes a cut in the DNA. In this way, the system acts as a precision pair of gene-cutting scissors.
The tool can therefore be used to edit a cell’s genome with incredible precision – for example, it can cut out a dysfunctional gene associated with a hereditary illness. And if the healthy version of the gene is also delivered to the cell, the cell’s own repair system will then incorporate the healthy strands at the site where it has been cleaved.
Charpentier teamed up with Doudna to investigate the system further. Together they revealed how the Cas9 protein, CrisprRNA and tracrRNA worked together to snip DNA strands into two parts. They then simplified the system by combining the CrisprRNA and tracrRNA into a single molecule – guide RNA – making it easier to use, and showed how this could be used to cut any DNA strand at a site of their choosing, opening the door to using the tool in all manner of genome editing experiments.
While previous tools for genetic editing existed before Crispr-Cas9, the new tools are much simpler and cheaper. This has led to the huge expansion of the field by making gene editing accessible for scientists all around the globe.
<...> area of controversy surrounds the potential consequences of using genome editing tools at all. As the genome is so complex, we can’t always know what will happen when we edit genes. Some genes have multiple and often unknown functions – editing them to correct for one problem could end up creating new unforeseen ones. This is particularly important when it comes to editing germline cells (those that can be passed on to an organism’s children), because the modified genes can be inherited by future generations.
As a relatively new technique, we also know that Crispr itself isn’t perfect. Some studies have shown off-target cuts, where the tool has snipped DNA strands at additional locations to the desired site. This clearly can have harmful consequences, and so many researchers are looking into ways to improve the technique and make it more suitable for medical uses.
With these concerns in mind, scientists worldwide – including Doudna and Charpentier – have called for a moratorium on editing human germline cells, until we can know more about the consequences. Such calls intensified after the rogue Chinese scientist He Jiankui edited human embryos that were then brought to term in 2018. He is now serving a three year prison sentence for conducting the study.
GREAT GOOD AND GREAT HARM
CRISPR’s co-discoverer, microbiologist Jennifer Doudna had a dream about Hitler:
It’s happened. The first children genetically engineered with the powerful DNA-editing tool called CRISPR-Cas9 have been born to a woman in China. Their altered genes will be passed to their children, and their children’s children. It is not the first time human ingenuity has created something capable of doing us great good and great harm. Are we up to the challenge of guiding how CRISPR will shape the future?
He shocked the world’s scientists in November 2018 when he announced that his team at Southern University of Science and Technology in Shenzhen had used the CRISPR gene-editing system to edit DNA in human embryos to make them less susceptible to HIV. The edits were designed to disrupt a gene that codes for a protein that allows HIV to enter immune cells.
Interesting to see the original presentation of He.
Article in Nature.com: "On 30 December, the People’s Court of Nanshan District of Shenzhen announced that, in the pursuit of “fame and profit”, He and two colleagues had flouted regulations and research and medical ethics by altering genes in human embryos that were then implanted into two women, according to Xinhua News Agency. One woman gave birth to twin girls in late 2018; the court said a third baby has been born but did not say when, a revelation that fits with a claim made by He in November 2018 to have implanted a gene-edited embryo in a second woman.
he court fined He 3 million yuan (US$430,000). Collaborators Zhang Renli and Qin Jinzhou received lesser prison sentences and fines."
Chinese government didn't want bad publicity so was silencing discussions about He Jiankui on social media. Jiankui's paper seems to be never published, and also it looks like there are three chinese babies that saw the daylight, from two different mothers, walking this earth with edited genes now. Lulu, Nana and another - mysterious child. Some scientist posted articles demanding for explanation. He is in jail at the moment probably and his return to work in the lab is unlikely. Another two scientists who worked with he seems to be also punished, but some say, that more people knew about what He was doing. Anyway, this punishment show scare other scientists of doing things deliberately, without consulting and getting permission. At the same time, there are three babies now living (hopefully), who might have immunity to HIV virus. Best case scenario - they grow up, have their own babies and this immunity spreads through the world. But there could be other scenarios. Some scientist was pointing out that the results of this experiment could be awry, and that's why there's no information about Lulu and Nana. From the presentation sheets He showed at the conference some say they could see that the results were "mosaic", meaning that not all cells could have adopted the properties of being immune to the virus.
WE NEED TO KNOW WHAT HAPPENED TO CRISPR TWINS LULU AND NANA
Technologyreview.com: "Why must the information be public? It’s because He’s work reveals serious, unresolved safety concerns. It’s not clear that any effort to directly edit human embryos, even if done ethically and with full social approval, can reliably avoid these problems.
International committees convened by the World Health Organization, the US National Academies of Medicine and Sciences, and the Royal Society are currently working to propose regulatory frameworks for doing clinical germline gene editing safely, if it is to be done at all.
How can the committees properly do their work without fully understanding all the scientific problems with the single real-world application of clinical germline gene editing that’s been attempted to date?
Most worrying is that scientists like Denis Rebrikov in Russia aspire to follow in He’s footsteps. Rebrikov has said he’ll be able to edit the human germline safely. But how can Rebrikov credibly claim to be able to do better than He if the nature of the problems with He’s work aren’t widely known? How can the Russian authorities properly evaluate the safety of his proposals without being able to refer to He’s work for guidance?
It’s time for the scientific community to fully understand what happened with Lulu and Nana, and to avoid stumbling down a path toward further ill-starred experiments with clinical germline gene editing.
Kiran Musunuru is an associate professor of cardiovascular medicine and genetics at the Perelman School of Medicine at the University of Pennsylvania and the author of The CRISPR Generation, a book about the history of gene editing and the Chinese twins."
Article in Forbes: "He shocked the world’s scientists in November 2018 when he announced that his team at Southern University of Science and Technology in Shenzhen had used the CRISPR gene-editing system to edit DNA in human embryos to make them less susceptible to HIV. The edits were designed to disrupt a gene that codes for a protein that allows HIV to enter immune cells."
MUTATION & MOSAICISM
There are also concerns about adverse effect called off-target mutation in CRISPR/Cas9 editing and mosaicism, a condition in which many different cells develop in the same embryo. Off-target mutation may cause health hazards, while mosaicism may create HIV susceptible cells. Fyodor Urnov, Director at the Altius Institute for Biomedical Sciences at Washington, asserted that "This [off-target mutation] is a key problem for the entirety of the embryo-editing field, one that the authors sweep under the rug here," and continued, "They [He's team] should have worked and worked and worked until they reduced mosaicism to as close to zero as possible. This failed completely. They forged ahead anyway."
The first successful gene-editing experiment of CCR5 in humans was in 2014. Researchers at the University of Pennsylvania, Philadelphia, Albert Einstein College of Medicine, New York, and Sangamo BioSciences, California, reported that they modified CCR5 on the blood cells (CD4 T cells) using zinc-finger nuclease which they introduced (infused) into HIV patients. After complete treatment, the patients showed decreased viral load, and in one, HIV disappeared.
In January 2019, scientists in China reported the creation of five identical cloned gene-edited monkeys, using the same cloning technique that was used with Zhong Zhong and Hua Hua – the first ever cloned monkeys – and Dolly the sheep, and the same gene-editing Crispr-Cas9 technique allegedly used by He Jiankui in creating the first ever gene-modified human babies Lulu and Nana. The monkey clones were made in order to study several medical diseases.
The first clinical trial of CRISPR-Cas9 for the treatment of genetic blood disorders was started in August 2018. The study was jointly conducted by CRISPR Therapeutics, a Swiss-based company, and Vertex Pharmaceuticals, headquartered in Boston.Preliminary report announced on 19 November 2019 states that the first two patients, one with β-thalassemia and the other with sickle cell disease, were treated successfully.
In April 2019, use of the CRISPR technology to edit human genes to treat for the first time cancer patients, with whom standard treatments were not successful, has been reported.
In June 2019, Denis Rebrikov at the Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology in Moscow announced through Nature that he was planning to repeat He's experiment once he got official approval from the Russian Ministry of Health and other authorities. Rebrikov asserted that he would use safer and better method than that of He, saying, "I think I'm crazy enough to do it."In a subsequent report on 17 October, Rebrikov said that he was approached by a deaf couple for help. He already started in vitro experiment to repair a gene that causes deafness, GJB2, using CRISPR.
ARE WE PLAYING GOD?
HUMAN NATURE (2019)
“With an extraordinary new technology called CRISPR, we can now edit DNA—including human DNA. But how far should we go? Gene editing promises to eliminate certain genetic disorders like sickle cell disease. But the applications quickly raise ethical questions. Is it wrong to engineer soldiers to feel no pain, or to resurrect an extinct species? And is there harm in allowing parents to choose their child’s features, like eye color or height? The scientists who pioneered human genome studies and CRISPR grapple with these questions. (Premiered September 9, 2020)”
Is it wrong to engineer soldiers to feel no pain, or to resurrect an extinct species?
Here's another big topic, using DNA for building, as wood.
2. BUILDING DNA STRUCTURE
When it comes to creating nanotechnology, one cannot simply build it with their hands. Instead, researchers need something nano-sized that is able to self-assemble. DNA origami is a method of creating nano-sized shapes by folding strands of DNA. This can be used to manufacture nanomachines, sensors, and nanorobots for use in fields ranging from biophysics to physical computing.
"We use DNA the way a carpenter uses wood," says Paul Rothemund from the California Institute of Technology, who invented the technique, and gave it its name: DNA origami.
TO BE CONTINUED...