Key takeaways
Nanoparticles could enable gene therapies for cancer and genetic disease that don't use modified viruses, which may cause new forms of cancer or dangerous immune system reactions in some patients.
Researchers from University of Michigan Engineering and Michigan Medicine used the nanoparticles to genetically modify human liver cancer, kidney and immune cells, which were grown in lab cultures.
The outer casing of the nanoparticles was made from protein, which could help prevent inflammation and liver and cell damage sometimes caused by fat-based nanoparticles used for gene therapies and mRNA vaccines.
In a demonstration that helps pave the way for gene therapies with fewer side effects, several human cell types were genetically modified with protein nanoparticles designed at University of Michigan Engineering and Michigan Medicine.
Gene therapy has been enormously successful for treating disorders of the blood, including sickle cell disease and leukemia. However, using a virus as a vector for treatment can create unwanted side effects, such as secondary cancers and immune system overreactions. With the nanoparticles, the research team aims to develop a safer method for delivering gene therapies.
In a proof-of-concept experiment, the researchers used nanoparticles to modify several types of human cells. They made human liver cancer cells, kidney cells and immune cells glow green by giving them genes for green fluorescent protein. The cells activated the new genes after they engulfed and digested the nanoparticles, releasing the DNA or messenger RNA packed inside.
"There are a lot of diseases where a protein is missing or dysfunctional due to a single mutation, and we can definitely correct for that by introducing a new gene," said Joerg Lahann , the Wolfgang Pauli Collegiate Professor of Chemical Engineering, director of the U-M Biointerfaces Institute and the corresponding author of the study, published in Advanced Materials.
"Typically, this is done with viruses, but the viruses can be toxic and activate the immune cells. So there has been a push in the field to replace virus-based gene editing strategies."
To help find alternatives, the National Institutes of Health funded the research.
An artificial virus goes rogue
Gene editing is already a central component of FDA-approved treatments for blood and bone marrow cancers , sickle cell and beta-thalassemia . These involve collecting and editing immune cells or blood stem cells with a genetically modified variant of HIV , which has evolved to insert itself into the genome of immune cells. In gene therapy, doctors change the virus so that it delivers beneficial genes that, for example, improve how the blood carries oxygen or help the immune system recognize and kill cancer cells. The edited cells are then given back to the patient.
Around 80% of adults and children who receive the cancer therapy, called CAR T-cell therapy, show complete cancer remission, and gene therapies are effective in 88% of sickle cell patients and 89% of beta-thalassemia patients . But some cancer patients develop a second cancer of the immune cells, and some sickle cell patients also develop blood cancer . Studies suggest the new cancers are caused by the virus, which breaks tumor-suppressing genes by inserting itself inside them.
The FDA has approved other viruses for direct injection into the body to fight skin cancer, congenital blindness and spinal muscular atrophy, but these viruses can sometimes cause infections and dangerous immune responses .
The researchers' nanoparticles could help avoid new cancers during gene therapy, because the ring-shaped (plasmid) DNA or RNA isn't inserted into a patient's genome, so no genes are broken up. The protein outer coating may also be safer than existing fat-based (lipid) nanoparticles, which can cause inflammation, fever and liver damage .
In the researchers' experiments, the protein of choice was serum albumin—a natural component of blood. Future recipes could use other proteins, such as neurotransmitters or signal proteins, to help the nanoparticles enter specific cell types, said co-author Michael Triebwasser , a clinical instructor at the U-M Medical School.
How the genes are delivered
The nanoparticles are made by a printing technique called electrohydrodynamic (EHD) jetting. First, protein is mixed with either DNA or RNA in water, which is placed into a syringe hanging above an aluminum plate. The researchers then apply an electric field that forces the charged protein, genetic material and water out of the syringe at high speeds. The acceleration quickly vaporizes the water and condenses the protein around the genetic material. The researchers then encase the particles in a synthetic, gel-like chemical called polyethylenimine to hold the particles together.
After cells engulf the particles, they are trapped in bubble-like compartments called endosomes, where cells digest food or kill pathogens. As the nanoparticles are digested, the positively charged polyethylenimine creates a charge imbalance. Water rushes into the endosome, popping it and releasing the genetic material.
Without integrating the released genetic material into the cell's DNA, the effects won't be permanent—mRNA lasts several days and plasmid DNA lasts several months, at most. As a workaround, patients could receive "booster" doses. Nanoparticles could also become "one-and-done" treatments when loaded with CRISPR-Cas9, a protein that inserts genes into a patient's genome with more precision than viruses.
"In future studies, we hope to test the nanoparticles' ability to modify human cells with therapeutic genes and identify potential side effects," said Fjorela Xhyliu , a doctoral graduate of chemical engineering and a co-first author of the study with Yao Yao, a doctoral graduate of health sciences from the U-M School of Dentistry, and Yeongun Ko, a former U-M postdoctoral research fellow in chemical engineering.
The nanoparticles were studied at the Michigan Center for Materials Characterization , the Biointerfaces Institute's Nanotechnicum Core and the Michigan Medicine Microscopy Core , all of which are operated and maintained with support from indirect cost allocations in federal grants.
Lahann is also a professor of materials science and engineering, biomedical engineering, and macromolecular science and engineering.
Study: Surface-Capped Protein Nanoparticles for Nonviral Gene Delivery (DOI: 10.1002/adma.202521796)
Written by Derek Smith
