Nanocontainers for macromolecules
Researchers from Johns Hopkins University have reported the creation of a nanoscale container that can penetrate into cells and deliver protein drugs and gene therapy drugs of any size, including CRISPR gene editing tools. They consist of a self-assembling biodegradable polymer. If further tests confirm the effectiveness and safety of nanocontainers, they can be used for the efficient transport of medical compounds into specially selected target cells.
Almost all medications are distributed throughout the body and do not target specific cells. Some drugs contain antibodies that attach to certain receptors on the surface of target cells, but there are no systems for delivering biological drugs directly into the cell, where there would be a higher chance of effective action with fewer side effects.
Work on delivery systems is underway in many laboratories. To do this, special forms of viruses are used, known for their ability to "infect" cells directly, to deliver drugs, although weakened versions of such delivery systems can cause an undesirable response of the immune system. Other treatments are more complex, requiring, for example, that the patient's blood be removed and then passed through an electric field to increase the permeability of the cell membrane for drugs.
The nanoscale container developed by Jordan Green and his colleagues is based on the principle of viruses, many of which have an almost spherical shape and carry both negative and positive charges. With a more neutral overall charge, viruses can approach the cells. Many biological drugs consist of highly charged large proteins and nucleic acids, which tend to repel cells. Therefore, they cannot penetrate the cells on their own.
To overcome this obstacle, researchers have developed a new biodegradable polymer material. To do this, they combined four molecules that eventually disintegrate and dissolve in water. Molecules have both positive and negative charges. With such a balance, molecules both repel and attract, depending on the charge of each of them, and hydrogen atoms bind to biological preparations located nearby. As a result, a nanostructure containing the drug is assembled.
Scheme of self-assembly of a nanocontainer from poly-(β-aminoester) molecules. Source: article in Science Advances.
Positively charged molecules of the nanoscale container interact with the cell membrane, and the container is absorbed by the cell pocket – the endosome. Once inside, the nanoscale container ruptures the endosome, the polymers decompose and dissolve, leaving the drug inside the cell.
To test their invention, the researchers created nanocontainers with protein molecules and injected them into a Petri dish with mouse kidney cells. The protein "cargo" was labeled with a green fluorescent label. Soon, almost all cells turned bright green, which proved the successful delivery of the protein.
Then a larger protein molecule was packed into the nanocontainer – human immunoglobulin, usually used to correct the immune system. 90% of the embryonic human kidney cells in the cup turned green, which marked the immunoglobulin.
Confocal microscopy of human embryonic renal cells. Nanocontainers with human immunoglobulin were delivered inside the cells. Cell nuclei are colored blue, endosomes are colored pink, immunoglobulin proteins are labeled with a green fluorescent label. Source: Johns Hopkins Medicine.
The experiments carried out prove that protein molecules are evenly distributed inside cells, and do not get stuck or destroyed in endosomes.
To complicate the experiment, scientists have created a nanocontainer containing a complex of proteins and nucleic acids based on CRISPR, which is able to turn off the green fluorescence signal and make cells glow red if the CRISPR system removes part of the cell genome. The researchers noted that the deletion of genes occurred in 77% of cells. Considering that with other gene editing systems, the desired result is obtained in less than 10% of cases, such an effect can be considered quite high.
In the latest experiment, the group implanted cancer cells into the brains of mice. Nanocontainers with gene editing components were injected directly into the brains of mice, the presence of a red glow would indicate successful gene editing. The researchers found cancer cells that glowed red a few millimeters from the site of their introduction.
Currently, the research team is trying to make nanocontainers more stable so that they can be injected into the bloodstream and directed to cells with certain genetic traits.
Scientists have applied for patents.
Article Y. Rui et al. Carboxylated branched poly(β-amino ester) nanoparticles enable robust cytosolic protein delivery and CRISPR-Cas9 gene editing is published in the journal Science Advances.
Aminat Adzhieva, portal "Eternal Youth" http://vechnayamolodost.ru based on Johns Hopkins Medicine: Little Size Holds Big Impact: Johns Hopkins Scientists Develop Nanocontainer to Ship Titan-Size Gene Therapies and Drugs Into Cells.