Barcoded nanoparticles deliver nucleic acids to treat cancer, viral infections, and neurodegenerative diseases.
Lipid nanoparticles containing unique DNA barcodes could speed the optimization of targeted nucleic acid drugs. The barcodes allow researchers to inject hundreds of different particles into a mouse and determine the tissues and cells where each nanoparticle collects.
This nano-barcoding system could eventually be used to study thousands of nanoparticles in a living organism and greatly increase the discovery and understanding of nanoparticle drug delivery.
Drugs acting through RNA interference, or RNAi, have been tested in clinical trials to treat cancer, viral infections, and neurodegenerative diseases. These RNA-based drugs prevent mRNA, a nucleic acid that contains instructions for protein synthesis, from being produced or read. Researchers tailor the sequence of a RNA drug to bind to genetic information linked to a specific protein involved in disease.
To protect a RNA drug as it travels to its target cell, researchers package it inside nanoparticles commonly made from lipids. Some lipids can help cells swallow the nanoparticles, while others help direct nanoparticles to particular tissues. Researchers determine the best nanoparticle packaging by testing how different formulations deliver nucleic acids to cultured cells. Then they test the best performing nanoparticles in animals to see if the nanoparticles still reach their target tissues.
However, cell-based tests cannot capture all the ways nanoparticles are destroyed, blocked, or misdirected in the body. Also, cells used for the tests are often different from those intended to receive the nanoparticles in the body.
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James Dahlman, a bioengineer at Georgia Institute of Technology, realized that in vitro cell culture tests may not be the best way to predict how the nanoparticles will behave in vivo, under physiological conditions. He began thinking of alternative tests and quickly ran into a conundrum: Testing either a few thousand particles in many thousands of mice or figuring out how how to test many particles in one mouse.
Dahlman’s solution to that problem – labeling particles with a unique DNA barcode so he can test many at once – could change the field of nucleic acid drug delivery. He was recently named as one of MIT Technology Review’s 35 Innovators under 35.
Last year, a team led by Dahlman and Eric Wang at the University of Florida, first showed that the multi-particle test worked: The researchers tracked a dose of 30 different lipid nanoparticles to eight different tissues in a mouse. They sequenced the DNA barcodes in each tissue to determine where each nanoparticle delivered its contents.
The researchers make the labeled nanoparticles using a microfluidic mixing system to package short strands of DNA barcodes inside lipid nanoparticles. This process is similar to the one used clinically to package RNAi therapeutics. Next, the researchers identify stable particles 20-200 nm in diameter using high throughput dynamic light scattering. Finally, they inject the barcoded particles into mice.
To determine where the nanoparticles delivered their DNA contents, the researchers isolate tissue from the lung, liver, heart, or other organs, then separate the tissue into different cell types using fluorescence-activated cell sorting (FACS) and sequence the DNA barcodes in each cell type.
Earlier this year, Dahlman and his colleagues used this technique to see if nanoparticles that delivered their barcodes to cultured cells also targeted the same cells in mice. The researchers measured how well 281 different lipid nanoparticles delivered DNA barcodes to cultured macrophages or endothelial cells, cell types found in many tissues and implicated in many diseases. They also injected the particles into mice and identified barcodes in macrophages and endothelial cells isolated from heart, lung, or bone marrow. In all the tissues they’ve tested so far, barcode delivery to cells in a dish did not predict delivery to those cells in an animal, as Dahlman suspected when he started this project.
Dahlman and his team have also explored how the composition of the lipid nanoparticles helps direct them to specific cell types. The researchers found that including a specific type of cholesterol in the lipid nanoparticles directs more particles to liver endothelial cells than hepatocytes that make up most of the liver.
Studying how the chemical structure of nanoparticles affects their delivery would be too tedious to examine carefully in traditional cell culture screens. “It’s exciting to ask questions in ways we couldn’t ask them before,” he says. This barcoding technique could be used for other types of nanoparticles too, as long as they are nontoxic and stable in solution before injection.
“All data in our hands, with appropriate controls, shows that this is real,” Dahlman says. “If others validate it in their hands, this could make an actual impact in the field. I really hope that’s the case.”
Melissea Fellet is an independent technology writer.
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