In lab experiments, the researchers have found that this device, a vector, is able to deliver DNA deeply enough into a cell to allow genetic material to be activated – a critical step in gene therapy. This vector is between 2 ½ and 10 times more effective than other experimental materials, according to the research.
In this case, Ohio State University scientists combined two ingredients – calcium phosphate and a lipid shell – to create a nanoparticle that protects DNA during its journey to the cell and then dissolves to allow for gene activation in the target cell. Nano refers to the tiny size of the particle in question – its general structure can be detected only by an atomic force microscope.
Calcium phosphate is a mineral found in bones and teeth. Lipids are fatty molecules that help maintain the structure of cell membranes. Alone, calcium phosphate is toxic and lipids get absorbed by cells. Together, they form a protective and inflexible structure that, thanks to complex chemical reactions, self-destructs once inside a cell.
“Our nanoparticle is a foreign body just like a viral vector is, but it has a self-destructive mechanism so it does not generate a strong response from the immune system,” said Chenguang Zhou, a graduate student in pharmaceutics at Ohio State and lead author of the study. “The material we use is also biocompatible. Calcium phosphate is in our bones and the lipids we use are synthetic, but can be biologically degraded. That’s why there is low toxicity.”
The research is published in a recent issue of the International Journal of Pharmaceutics.
Zhou noted that other researchers have tried to use liposomes – nanometer-sized bubbles made out of the same material as a cell membrane – to create nonviral vectors for gene delivery. While the material did a good job of protecting the DNA, it did not do a good job of releasing the gene into a cell.
“The liposome gets internalized into cells. It’s sort of like eating food that gets stuck in the stomach or intestines, but never gets to the rest of the body,” he said.
Similarly, calcium phosphate alone has been considered as a gene delivery vehicle. But because of its salty properties, it becomes unstable and expands in size, which makes it too big to penetrate some cell and vascular walls, and which can cause the immune system to reject it.
“So what we do is encapsulate a calcium phosphate core inside the liposome,” Zhou said. “And when this calcium phosphate gets inside a cell and that environment becomes acidic, it gets dissolved and then the gene can be very effectively released into the cytoplasm and transported to the nucleus. That is the theory.”
Zhou and colleagues have developed what they consider an easy method to manufacture this particle. They create a synthetic lipid and place it in a solution that contains calcium and phosphate, which becomes integrated with the lipid. As the acidic properties in the solution change, the calcium phosphate forms a core.
The scientists then mix a solution containing plasmid DNA with their newly formed particle, and all the materials become bundled together. Plasmid DNA is a circular DNA molecule that is able to turn on gene activity that starts a protein-building process without altering an entire genome, or the complete hereditary information of an organism.
Because this particular vector is intended for injection into the bloodstream as a cancer treatment, the particle is designed to protect the DNA from being digested by enzymes as it travels to target cells. A test that exposed this solution to serum, a component of blood, showed that the hybrid particle provided this protection, while unprotected DNA was digested by enzymes.
The researchers next applied this DNA-infused particle solution to mouse cells. The DNA contained the gene code for green fluorescent protein that would be turned on only after it entered the cell. They observed that the particle penetrated the cell membrane and, after a series of interactions occurred, the green fluorescent protein lit up inside the cell, indicating the DNA had reached its target.
For comparison, the group also monitored DNA movement on its own and in other types of vectors. Their hybrid vector was 24 times more effective at delivering genetic material to the cell than was DNA on its own, and 10 times more effective than calcium phosphate preparations.
“We know the particle gets to where it needs to go and what happens to the particle,” Zhou said. “Do we know that the DNA reaches the nucleus? That is something we still need to find out. But because we saw the green fluorescent protein expressed, we think it got to the nucleus or at least as far as the cytoplasm. What’s important is that the protein got inside the cell.”
The study also showed that this hybrid particle maintained its structure for at least 21 days and, when compared with a variety of other potential vector substances, did very little damage to cells, meaning it is not as toxic as most other materials.
With viral vectors, gene therapy is considered a one-time treatment because when the virus carrying new genes infects a cell, that interaction changes the recipient’s entire genome, effectively canceling the activity of the defective gene. Zhou said that with this nonviral vector, treatment would be designed as an intravenous injection on a regular basis until cells are “infected enough to make a change.”
The researchers next plan to test the particle’s ability to travel through the bloodstream and enter target cells in animals.
This work was supported by the National Science Foundation (NSF) and the National Institutes of Health. Zhou has a fellowship in Ohio State’s NSF Nanoscale Science and Engineering Center (NSEC) for Affordable Nanoengineering of Polymeric Biomedical Devices.
He completed the research with Ohio State co-authors Bo Yu and L. James Lee of the Department of Chemical and Biomolecular Engineering; Xiaojuan Yang and Robert Lee of the Division of Pharmaceutics; and Tianyao Huo and Rolf Barth of the Department of Pathology. Yu, Yang, L. James Lee and Robert Lee are also affiliated with the NSEC.Contact: Chenguang Zhou, (614) 292-5870; firstname.lastname@example.org
Chenguang Zhou | EurekAlert!
Complementing conventional antibiotics
24.05.2018 | Goethe-Universität Frankfurt am Main
Building a brain, cell by cell: Researchers make a mini neuron network (of two)
23.05.2018 | Institute of Industrial Science, The University of Tokyo
A research team led by physicists at the Technical University of Munich (TUM) has developed molecular nanoswitches that can be toggled between two structurally different states using an applied voltage. They can serve as the basis for a pioneering class of devices that could replace silicon-based components with organic molecules.
The development of new electronic technologies drives the incessant reduction of functional component sizes. In the context of an international collaborative...
At the LASYS 2018, from June 5th to 7th, the Laser Zentrum Hannover e.V. (LZH) will be showcasing processes for the laser material processing of tomorrow in hall 4 at stand 4E75. With blown bomb shells the LZH will present first results of a research project on civil security.
At this year's LASYS, the LZH will exhibit light-based processes such as cutting, welding, ablation and structuring as well as additive manufacturing for...
There are videos on the internet that can make one marvel at technology. For example, a smartphone is casually bent around the arm or a thin-film display is rolled in all directions and with almost every diameter. From the user's point of view, this looks fantastic. From a professional point of view, however, the question arises: Is that already possible?
At Display Week 2018, scientists from the Fraunhofer Institute for Applied Polymer Research IAP will be demonstrating today’s technological possibilities and...
So-called quantum many-body scars allow quantum systems to stay out of equilibrium much longer, explaining experiment | Study published in Nature Physics
Recently, researchers from Harvard and MIT succeeded in trapping a record 53 atoms and individually controlling their quantum state, realizing what is called a...
The historic first detection of gravitational waves from colliding black holes far outside our galaxy opened a new window to understanding the universe. A...
02.05.2018 | Event News
13.04.2018 | Event News
12.04.2018 | Event News
24.05.2018 | Ecology, The Environment and Conservation
24.05.2018 | Medical Engineering
24.05.2018 | Physics and Astronomy