CRISPR on Gold
Gold helps to change human genes
Alexey Aleksenko, Forbes, 28.05.2019
The CRISPR gene editing system can be delivered to human cells using gold nanoparticles. Thus, geneticists managed to correct a hereditary defect in the hemoglobin gene and protect white blood cells from HIV.
Researchers from the Fred Hutchinson Cancer Research Center (Seattle) have managed to develop a technique that greatly simplifies the procedure for editing genes in human cells. The method is based on gold nanoparticles, thanks to which the components of the "molecular machine" of CRISPR editing are delivered directly to the cell nucleus. The results of the work are published in Nature Materials (Shahbazi et al., Targeted homology-directed repair in blood stem and progenitor cells with CRISPR nanoformulations).
CRISPR technology allows you to directly change the information recorded in DNA. To do this, two main components need to be delivered to the target DNA: a "guide" molecule that will find the exact place of "surgical intervention" among tens of thousands of genes, and an enzyme that cuts the DNA molecule at this point to begin the process of "repair". The difficulty is the delivery of the components of the complex into the cell. Inactivated virus particles are often used for this purpose. A more straightforward approach is an electric current pulse. It stimulates foreign DNA to enter the nucleus, but it can damage or even kill the cell.
"I wanted to find something simpler- some way to passively deliver the editing device to blood stem cells," says study leader Dr. Jennifer Ader.
Gold nanoparticles – gold balls with a diameter of about one billionth of a grain of salt – have been used in molecular biology and genetic engineering for a quarter of a century. With their help, biologists introduced foreign genes into plant cells: many genetically modified varieties were created in this way. The advantage of gold nanoparticles is that nucleic acid molecules easily adhere to them, and the particles themselves relatively easily enter the nucleus, overcoming various cellular barriers.
Dr. Ader and her collaborator Reza Shahbazi have been using gold particles to deliver various substances to the cell for many years. In this case, the researchers obtained gold nanoparticles from a laboratory colloidal solution. Then they were sorted by size: the ideal diameter was 19 nm. At this size, the surface of the particles is sufficient to accommodate all the necessary molecules, but at the same time the particle is still small enough to freely penetrate into the cell nucleus. From above, the particles were covered with polyethylenimine: this created a small positive charge on the surface, thanks to which the cell "took the medicine" more willingly.
To demonstrate the capabilities of the method, biologists edited the genes of stem cell cultures that are able to transform into various human blood cells. In one cell line, the CCR5 gene was edited: an altered version of this gene makes cells immune to HIV infection. It was this gene that was edited in the sensational experiments of the Chinese geneticist He Jiankui. Unlike He, who edited human germ cells (and thereby created people capable of transmitting a new gene to their descendants), researchers from the Fred Hutchinson Center worked with a cell culture that, in principle, is not capable of developing into a human body: changes in any case would affect only the patient's blood cells.
In another cell line, the gamma-hemoglobin gene was changed: such procedures provide the key to the treatment of blood diseases such as sickle cell anemia and thalassemia.
The researchers found that about 6 hours after the addition of the gold particles, the cells assimilated them, and over the next day or two, gene editing took place in them. The edited cells were injected into mice used as a model of human diseases. The effect peaked 8 weeks after administration. Cells with altered genes were found in mice five months later. They were present in the bone marrow, spleen and thymus gland. Among the treated cell culture, 10-20% were edited. This is a high result, but biologists intend to increase the effectiveness of the method to 50%. In the future, this will give doctors a method of treating serious hereditary diseases.
Konstantin Severinov, a professor at Skoltech, commented on this scientific work: "The delivery of gene editors by electroporation – in fact, by discharge of a high–voltage current - is toxic: many cells die from such a procedure. Viruses can be used for delivery in various ways, but the interaction of viral particles with a cell can also be toxic, and besides, not all cells are able to interact with viruses. In general, there is a common problem of delivering – no matter what exactly – to cells. Nanoparticles can be used for this purpose. According to the authors, this method is non-invasive, non-toxic and effective."
Professor Severinov, however, believes that the results should be treated with caution: "In general, it is necessary to solve not just the problem of delivery – cells tend to capture all sorts of rubbish inside, if it is the right size – but targeted delivery. How to introduce the substance we are interested in into a certain type of cell? This is a very big unsolved problem that limits not only the use of genomic editing methods, but also various methods of cancer treatment, and much more. There are no general solutions yet."
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