‘Medicine of the Future’: Delivering mRNA Gene Editing Tools In Vivo

The research team used lipid nanoparticles to deliver the mRNA gene editing tools, allowing for the modification of diseased blood cells directly within the body.

By Nancy McCann

A scientific team of Children's Hospital of Philadelphia and University of Pennsylvania investigators developed a proof-of-concept model for delivering gene editing tools to the bone marrow to treat blood disorders, allowing for in vivo cellular reprogramming and providing a new and potentially safer way of controlling hematopoietic stem cells (HSC) fate by correcting genetic defects. If translated into the clinic, this approach could expand access and reduce the cost of gene therapies for blood disorders.

To treat sickle cell disease and beta thalassemia — devastating blood disorders that result in anemia, severe chronic pain, and premature death — with gene therapy, patients receive an expensive treatment, which is also risky as it requires myeloablation. Myeloablation involves toxic doses of chemotherapy to eliminate the mutated HSCs in the bone marrow and makes room for the functional cells.

The patient's HSCs are modified outside of the body and then transplanted back in the patient after myeloablation. It's a grueling experience for the patient, and it can only be done once.

But what if physicians could replace the diseased blood cells with corrected ones directly within the body, eliminating the need for the myeloablative conditioning treatment and streamlining the delivery of this potentially life-changing treatment?

Working to make this new treatment a reality are investigators Stefano Rivella, PhD; Laura Breda, PhD; and Michael Triebwasser, MD, PhD, from CHOP, together with Hamideh Parhiz, and Tyler Papp, PhD, at the University of Pennsylvania, and Drew Weissman, MD, PhD, director of the Penn Institute for RNA Innovation. The journal Science published their novel findings.

"The potential of this technology is transformative," Dr. Rivella said. "In this paper, the technology is meant for bone marrow and red blood cell diseases. But this technology can potentially treat other diseases. If we could deliver a cargo that corrects a mutation in the bone marrow, we can conceive alternative formulations to correct mutation in the brain, lung, or liver — this is the technology that will allow us to do that.

"This is a medicine of the future."

Delivering mRNA Gene Editing Tools

The research team used lipid nanoparticle (LNP) to deliver the mRNA gene editing tools. LNPs are highly effective at packaging and delivering mRNA to cells and, in 2020, this technology became widely used due to the LNP-mRNA platform for two leading COVID-19 vaccines.

But for COVID-19 vaccines, the LNP-mRNA construct did not target specific cells or organs within the body. This team of researchers wanted to target HSCs specifically, so they enhanced the surface of their experimental LNPs with antibodies that would recognize CD117, a receptor on the surface of HSCs. They then pursued three approaches to test the efficacy of their CD117/LNP formulation.

First, the researchers tested CD117/LNP encapsulating reporter mRNA to show successful in vivo mRNA expression and gene editing, working with a murine model that enabled the team to quantify the level of gene editing with a fluorescent marker.

Next, the researchers investigated whether this method could be used as a therapy for hematologic disease. They tested CD117/LNP encapsulating mRNA encoding a Cas9 base editor targeting the mutation that causes sickle cell disease. This type of gene editing converts the disease-causing hemoglobin mutation into a non-disease-causing variant.

Testing their construct on cells from donors with sickle cell disease, the researchers showed that CD117/LNP produced efficient base editing in vitro, leading to a corresponding increase in functional hemoglobin of up to approximately 92%. They also demonstrated a nearly complete absence of sickled cells, the crescent-shaped blood cells that cause the symptoms of the disease.

The researchers also explored whether LNPs could be used for in vivo conditioning, which would allow bone marrow to be depleted without chemotherapy or radiation. To do so, they used CD117/LNP encapsulating mRNA for PUMA, a protein that promotes cell death.

The researchers showed that in vivo targeting with CD117/LNP-PUMA effectively depleted HSC, allowing for successful infusion and uptake of healthy bone marrow cells, a process known as engraftment, without the need of chemotherapy or radiation. The engraftment rates observed in animal models were consistent with those reported to be sufficient for the cure of severe combined immunodeficiency using healthy donor bone marrow cells, suggesting this technique could be used for severe immunodeficiencies.

Advantages of New Delivery System

"We've created a sophisticated delivery system which doesn't require any intermediate steps like myeloablation," said Dr. Breda, scientific co-director of the Comprehensive Center for the Cure of Sickle Cell Disease and Other Red Blood Cell Disorders. "It's an intravenous infusion that goes directly to the bone marrow via the bloodstream."

Theoretically, this infusion can be done more than once, unlike a bone marrow transplant. If the optimal number of corrected cells wasn't reached with the first injection, more may be given, as long as it doesn't create an immune response to the drug. 

"While we would like this to be a one-and-done therapy," Dr. Breda said, "these formulations can be safely injected multiple times to achieve the optimal level of stem cell modification."

This IV treatment would likely be more cost effective than current therapy, as it does not require a hospital stay and is delivered in a vial, making worldwide access to the treatment possible.

"This is a big step forward in how we think about treating genetic diseases and could expand the access of gene therapies to patients who need them most," said Dr. Rivella, the Kwame Ohene-Frempong Endowed Chair in Pediatric Hematology.

For more information see the CHOP press release.