Nanotechnology in medicine: successes and prospects
"In the world of science" No. 2-2009
Doctors have learned how to inject an active gene instead of a damaged one into a living cell, as well as observe the metabolism of various substances in real time. About this to the correspondent of "In the world of science" Academician of the Russian Academy of Sciences and the Russian Academy of Medical Sciences, Dean of the Faculty of Fundamental Medicine of the Lomonosov Moscow State University, Doctor of Biological Sciences, Professor Vsevolod Arsenyevich Tkachuk tells.
Recently, nanotechnology has been increasingly introduced into many areas of human activity. This trend has not bypassed medicine. Today, macromolecules and artificially prepared particles are used for the diagnosis, treatment of various diseases and restoration of damaged tissues. The new direction was called nanomedicine. Russian scientists have achieved considerable success in this field.
Doctors have been familiar with the nano area for a long time. Many of the biological objects they study are smaller than a micrometer. For example, peptides have a size of 1 nm, proteins – from 10 to 100 nm. The DNA of a human cell reaches 1.5 m in length, but in the "packed" state its diameter is only 100 nm. Antibodies, viruses, organelles have approximately the same size, and cells and bacteria already belong to the microcosm. For clarity, we can make such a comparison: if the cell is enlarged to the size of a lake, then proteins, antibodies, viruses will be like small fish swimming in it.
Magnetic nanoparticlesTo effectively treat the disease, you need to be able to diagnose it at the earliest stage.
For these purposes, magnetic nanoparticles containing a crystalline core of iron oxide are used. Such a diagnosis is good because there are no magnetic particles in the body, but iron is present, which is part of hemoglobin and is in the form of separate ions that have practically no magnetic properties.
When a suspension of magnetic nanoparticles is introduced into the body, each of them, being a foreign body, is captured by a macrophage (specialized protective cells that absorb bacteria and other foreign substances), which as a result becomes "labeled". Since he cannot digest an inorganic particle, he continues to move along with it further. If there is a tumor or an inflammatory process somewhere, macrophages rush there to fight infections, viruses, bacteria, and accumulate there for a certain time. Using magnetic resonance imaging, it is easy to detect areas of increased concentration of magnetic nanoparticles, and thus determine the foci of inflammation at the earliest stage of their occurrence.
Quantum dotsConsider a small piece of a semiconductor in which there is one "hole", i.e. a positively charged ion.
If its size is much larger than a micron, then it behaves like a normal macro object. A free electron in it with a small energy can connect with such an ion due to Coulomb forces, thereby neutralizing it. When a semiconductor crystal is reduced to a size of about 100 nm or less, a qualitative change in its physical properties occurs due to the appearance of quantum effects. An electron has wave properties, and therefore it cannot be localized in a volume of space smaller than its wavelength – it does not have enough energy for this. The result is a nanoobject called a "quantum dot". An electron with a small energy can neither fly away from the hole nor fall on it. It turns out a kind of potential pit, which has quantum properties due to its small size, in which the electron has a certain spectrum of energy levels. Accordingly, this whole "hole –electron" system also has a certain electromagnetic spectrum and resembles an ordinary atom, which also represents a potential well. However, only the properties of ordinary atoms always remain unchanged, and the radiation spectrum of a quantum dot can be adjusted at its discretion by changing its size. It is no coincidence that Nobel laureate J.I. Alferov called quantum dots artificial atoms whose properties can be controlled.
Today, we have already learned how to prepare a "suspension" of quantum dots of a certain size, having, for example, green or some other color convenient for conducting research. In addition, chemists can "sew" such molecules to quantum dots that are able to specifically bind to the necessary molecules or parts of small organic bodies located inside a living organism. With this binding, the size and color of the point change. An antibody can be "sewn" to a quantum dot, which will then bind a protein, or a substance that can chemically bind to a certain fragment of DNA - a gene. Biochemists have learned to attach characteristic molecules to nanoparticles-probes that bind to a certain piece of DNA, protein, vascular wall, or simply "hang out" in the blood or lymph.
All this is used to solve one of the main problems in diagnostic medicine – the background problem: it is usually very difficult to distinguish the signal coming from the studied place in the body from the various noises generated by the surrounding tissues. Here a special role is assigned to the use of quantum dots. As it turned out, when cancer cells are detected at the earliest stage of the disease, a molecule is first synthesized that binds only to a specific protein produced in the pathological cell. Then this molecule is sewn to a quantum dot having, for example, a red color. By observing the appearance of this color in the body, it is possible to determine where the malignant tumor is located with accuracy to individual cells. During experiments with mice, researchers injected red quantum dots into their tails that bind to thyroid cancer cells. There was an accumulation of dots in the area of this organ, and a characteristic red color was recorded.
A suspension of green quantum dots can also be injected into vessels. After they are distributed quickly and evenly enough, an extensive network of blood flow in the form of a characteristic emerald "coral tree" will be visible through the skin. And if a small vessel or capillary is damaged in some place, it will be noticeable by small breaks in the green grid of vessels. Such a signal is absolutely clear, and it cannot be confused with anything, because there is no green color in the tissues of the human body.
The image shows the visualization of microvessels using water–soluble quantum dots in the mouse skin (depth 100 microns).
The whole human genome in one dropScientists have already deciphered the structure of the human genome, consisting of 30 thousand genes.
This is about 3 billion nucleotides, which play the role of peculiar letters when recording hereditary information. It is also known in which part of the giant "DNA book" each gene is recorded. Most diseases are provoked by "failures" caused by mutations in genes, therefore, during a medical examination it is important to know whether the patient has an innate predisposition to any diseases caused by hereditary "errors" in DNA.
Currently, this is done using microchips. Approximately 100 quantum dots are applied to each square millimeter of such a device, each of which has its own probe sewn to it, capable of specifically binding to a certain DNA fragment and thus testing it.
The general principle of testing is as follows. Let's say you need to find out if a person has a predisposition to Alzheimer's disease or to heart failure. To do this, a blood test is taken from him and it is determined whether there is a gene there whose mutations cause this ailment. Then, in the laboratory, a small fragment of DNA is synthesized, for example, in the size of one hundred nucleotides. At the same time, only in this gene out of all 30 thousand available there is such a sequence. A fragment is sewn to a quantum dot and placed in a certain cell on the chip, another specific fragment is sewn to another quantum dot and inserted into the next cell, etc.
After a drop of blood is applied to the chip, the probes sewn to the quantum dots bind to certain DNA fragments. Then, with the help of a computer equipped with a microscope, all cells are "viewed" sequentially. If the color has changed in each quantum dot, it means that there has been a binding with all the genes that this microchip is testing, and everything is fine with them. If the color has not changed in some cell, it means that binding has not occurred there, and, consequently, there is a "breakdown" in the corresponding gene, i.e. a violation in the sequence of nucleotides. After that, it turns out exactly where the breakdown is located in the gene. If it is in the piece that is responsible for Alzheimer's disease, then there is a predisposition to this disease.
Microchips have already been created, in which the number of cells is measured in tens of thousands, and in each there is an indicator for a certain fragment of human DNA. They can belong to the same gene or to different ones, depending on the task at hand. Today, with the help of quantum dots, it is possible to diagnose not only hereditary diseases, but also various infections, etc. There is a "specialization" of chips. For example, one is made for all cardiovascular diseases, the second - for endocrine, the third – for oncological. A chip with a drop of the examined blood is placed under a microscope, a special program reads all the points, the information is processed on a computer, and the diagnosis for this disease is ready.
Unfortunately, this technology does not allow for a complete genetic diagnosis of a person. The hereditary information contained in DNA is so large that it will require millions of specialized microchips. But if you master the technology of manufacturing nanochips with a cell size of about 100 nanometers or even less, it will be possible to increase its informativeness millions of times. Nanochip technology will allow using only one square centimeter to diagnose a person by all genes and mutations. One drop of blood is enough – and you can learn everything about a person's genetic health. It is expected that in the coming years a nanochip will be constructed on which the entire human genome can be applied.
Delivery of genes strictly to the addressDNA has a spiral structure and consists of two complementary strands connected to each other, representing a sequence of four nucleotides, the location of which determines the structure of the synthesized protein.
And if there is a substitution of at least one of the nucleotides, then some "foreign" amino acid will appear in the protein molecule. As a result, the protein will not be able to pack properly and will perform its functions poorly. For example, with such a congenital disease as sickle cell anemia, the hemoglobin protein, due to a slight change in the amino acid sequence, loses its ability to take the desired shape and therefore cannot carry oxygen. This disease is incurable, and the person dies. Other protein disorders caused by hereditary errors in DNA are usually less dangerous, but, nevertheless, can also lead to various chronic diseases over time. And if earlier doctors tried to cure the disease itself, now they are trying to eliminate its cause – to help the body synthesize the right protein correctly. There is even a new direction in medicine – gene therapy. And many diseases caused by a malfunction in genes, both hereditary and acquired, are being cured today.
Studies have shown that only a certain number of genes are involved in the work of a cell, depending on its specialization. If the sequence of nucleotides is disrupted in any gene, then the protein synthesized on its basis cannot fully perform its functions, which leads to metabolic disorders with all the consequences that follow from this.
Recently, they have learned to introduce a full-fledged gene into a human cell to replace a damaged one, which makes it possible to synthesize the "right" protein. The main problem is the exact targeted delivery of the gene inside this cell. To solve it, the natural ability of the cell is used – the so-called endocytosis, i.e. the capture of various small particles and organic molecules by the outer membrane with their subsequent digestion. Viruses use this property: they get inside the cell and make it work for themselves.
The virus consists of DNA or RNA surrounded by a protein shell, and cannot reproduce independently. Such a parasite has a certain program: getting into a living cell, it disrupts its work, forcing it to copy itself in thousands and millions of copies. Viruses propagated in this way leave the destroyed cell to find a new victim.
And what if we use the "strategy" of viruses to introduce the right gene into the cell? It turned out that this is possible.
In the treatment of the disease, it is first found out whether it is caused by genetic errors. And if so, then it is determined which gene is malfunctioning. Then the required DNA fragment is synthesized, which is always a polyanion, since in it the positively charged nucleotide bases are connected to each other and hidden inside the twisted molecule, and the negatively charged phosphate groups are facing outward. To neutralize the external electric charge, a corresponding polycation is added to the gene. Under the action of interatomic forces, this whole complex structure collapses into a nanosheet. From above, such a ball is covered with another polyanion. This is done so that when the cell comes into contact with it, it swallows it, which it usually does with a virus. Inside the cell, the shell of the ball is destroyed by the action of digestive enzymes. The released gene penetrates into the nucleus, where it begins to work – it turns on the intracellular mechanisms of protein synthesis on its matrix. The gene introduced in this way manages to work for about two weeks, then the cell still recognizes the alien and destroys it.
It turned out that in patients with thrombophlebitis and diabetes, small blood vessels are destroyed in the legs, the supply of oxygen to cells is disrupted, they die off, forming so-called trophic ulcers. Traditional methods of treatment, as a rule, do not help, because proteins injected to restore blood vessels can "work" inside the body for no more than half an hour and during this time they do not have time to do anything. The pain of trophic ulcers is so severe that it is necessary to amputate the legs. Some of these patients, who are at the last, most severe stage of the disease, are periodically injected with genes responsible for the synthesis of missing proteins. After two months, the thinnest network of blood vessels is noticeably restored, and trophic ulcers are reduced.
Nanorobots and nanomotorsThe most promising direction of future research in nanomedicine is the creation of nanorobots that will play the role of a kind of nanodoctors.
Moving throughout the body inside the smallest vessels and inside the cells, they will eliminate various malfunctions and clean the vessels. One of the main problems in creating such devices is the manufacture of nanomotors, with the help of which nanorobots will be able to move inside tissues and inside a single cell. To achieve such goals, it is enough to learn how to use intracellular "transport".
Each cell can be compared to a huge metropolis, in which many factories for the production of protein and other organic compounds are connected to each other by a complex network of roads – actin (a type of protein) threads. Each molecule, once inside the cell, depending on its structure gets on a certain road and moves strictly along it to a certain place. Having understood the principle of operation of intracellular communication pathways, you can use them for accurate targeted drug delivery. To do this, it is enough to find out which path leads to the right place, as well as the types of molecules moving along it. By sewing a nanocontainer with a drug to a similar molecule, you can send it to the desired address. Moreover, such delivery occurs due to the universal energy of ATP, which is the most efficient "fuel" with an efficiency of 92%. In this reaction, the energy of the chemical bond immediately turns into mechanical, bypassing the thermal stage. Such a natural motor works with high efficiency in all living organisms without exception.
Scientists of the Moscow State University, the Russian Academy of Sciences and the Cardiocenter have developed a technology where the myosin protein, which has a natural ability to move along actin filaments, is sewn to a nanocontainer with a drug. As a result, a portion of the drug "crawls" along the intracellular rails to the right place without any energy expenditure.
Any pills, capsules that we swallow, in addition to the therapeutic effect, also have a side effect on the whole body. And the use of nanocontainers that deliver the drug "where it is needed" will reduce this side effect to almost zero.
BiosensorsIn diagnostics, it is important to be able to identify the intracellular processes that interest us at the moment and monitor them in real time.
Corresponding member of the Russian Academy of Sciences S.A. Lukyanov from the Institute of Bioorganic Chemistry proposed an original solution to the problem.
The scientist has developed a technology that changes the structure of the gene so that the fluorescent part of the protein is preserved, and the other acquires the ability to bind in a certain way to any pre-selected intracellular substance. At first glance, such a technology opens up wide possibilities for determining the path of any compound inside a living cell.
It should be noted that it is possible to create a fluorescent protein that binds to calcium. Moreover, when the ion binds, the fluorescence of the protein will increase. Next, a gene with an altered structure is introduced, for example, into a mouse germ cell. When the animal grows up, a change in the concentration of calcium in its cells will be accompanied by a change in the intensity of green light in various places. The cellular metabolism of a chemical element becomes visible in every detail. You can monitor how the concentration of calcium ions in cells changes in various diseases, how treatment affects their recovery, etc. In a similar way, you can monitor the metabolism of other substances. This method opens up wide opportunities for visualization of vital processes and allows you to directly check the effectiveness of a particular treatment.
In each cell there is a program of self-destruction, the so-called apoptosis, which is turned on after a certain period of time (see: Koroleva A., Skulachev V., Skulachev M. The choice between life and death // VMN, No. 2, 2008). Such an extremely important process for understanding the vital activity of the body is not yet fully clear. However, it is known that it is accompanied by an increase in the intracellular content of hydrogen peroxide. S.A. Lukyanov synthesized a gene producing a fluorescent protein that binds to hydroperite. This made it possible to observe the entire process of cellular apoptosis in real time. You can also track the path of a drug or any other substance injected into the body.
Do no harm!It is known that everything useful is useful only in certain quantities, taking too much, as a rule, causes harm.
Paracelsus also said that everything is poison, and only the right dose makes the substance safe. Accordingly, nanoparticles, despite their ultra-small size, can also be dangerous. In this regard, their testing procedures are being developed, similar to those used for conventional medicines. For example, it is possible that nanoparticles can cause protein aggregation, and this can subsequently lead to Alzheimer's disease, etc. Therefore, all nanopreparations undergo preclinical testing. Their influence on morphology, cell development and movement, metabolism, etc. is consistently checked. And only after that, the optimal nature, size and dose of nanoparticles are selected for each specific case.
The Great UnificationToday, biological sciences are developing rapidly.
Discoveries in these areas of human activity occur almost every day. Specialists study how viruses, bacteria, and various intracellular structures function, and then adopt their "work experience" to solve fundamental and applied problems. Physicists and engineers are diving deeper into the microcosm, mastering new nanotechnology and already know how to manipulate individual molecules. It can be expected that in the near future, the study of intracellular processes will combine quantum mechanics, molecular biology, genetic engineering, biochemistry, medicine and inorganic chemistry. As a result of such a "great unification", there will surely be a qualitative leap in understanding what life is, and medicine will be enriched with new methods for the diagnosis and treatment of a person.
Vasily Yanchilin talked
Portal "Eternal youth" www.vechnayamolodost.ru02.03.2009