Novel Approach to Lipid Nanoparticle Engineering

To enhance mRNA distribution, Penn engineers have created an ideal “recipe” for ionizable lipids, essential lipid nanoparticles (LNPs) components, the molecules that underlie COVID-19 vaccines, and other cutting-edge treatments.

The technique, which was published in the journal Nature Biomedical Engineering, is similar to the iterative process of creating a recipe and could result in mRNA vaccines and treatments that are safer and more efficient.

To determine the right structure for the ionizable lipid, the researchers employed an iterative method, testing variations, much like how a chef perfects a meal by experimenting with flavors and textures. The structure of this lipid facilitates mRNA therapies for gene editing and vaccinations and affects how well LNPs carry their contents.

A Breakthrough in LNP Design

Nanoparticles have revolutionized the delivery of mRNA vaccines and treatments since they can pass through the body safely, reach the target cells, and effectively release their contents. When left alone, RNA is brittle and disintegrates before ever reaching its destination.

These nanoparticles' core constituents are ionizable lipids, unique molecules that can transition between charged and neutral states depending on their environment. This switch is necessary for the nanoparticle's travel: Ionizable lipids do not become harmful in the bloodstream because they remain neutral. However, once inside the target cell, they acquire a positive charge, which causes the mRNA payload to be released.

Under the direction of Associate Professor in Bioengineering Michael J. Mitchell, the researchers improved the structure of ionizable lipids to enhance this delivery method. The researchers went beyond current techniques that were constrained by accuracy and speed considerations and created a methodical, “directed chemical evolution” procedure.

Through five cycles, each further refining the lipids, they produced dozens of high-performing, biodegradable lipids, some exceeding industry requirements.

The Secret Sauce: Directed Chemical Evolution

The Penn Engineers took a novel approach to create safer, more potent ionizable lipids by combining two widely used techniques: combinatorial chemistry, which produces many different molecules rapidly through straightforward reactions, and medicinal chemistry, which involves carefully and slowly designing molecules one step at a time.

Low speed and low accuracy characterize the latter, while great accuracy and low speed characterize the former.

We thought it might be possible to achieve the best of both worlds. High speed and high accuracy, but we had to think outside the traditional confines of the field.”

Xuexiang Han, Study First Author and Postdoctoral Fellow, University of Pennsylvania

To create their perfect lipid “recipe,” the researchers combined accuracy and speed by utilizing the concept of directed evolution, a method that is applied in both biology and chemistry and simulates the process of natural selection.

First, many molecules are created, and they are vetted to see if they can carry mRNA. The top-performing lipids then serve as the basis for yet another round of chemical variations, and so on, until only the best-performing versions are left.

An Innovative Ingredient: A3 Coupling

An amine, an aldehyde, and an alkyne are the three chemical components of the A3 coupling reaction, which is a key component of the team's recipe for enhanced ionizable lipids.

The reaction, which has never been used to create ionizable lipids for LNPs, is a cost-effective and environmentally friendly option for quickly creating the vast quantities of ionizable lipid variants required as components for directed evolution because it only produces water as a byproduct and uses cheap, commercially available ingredients.

We found that the A3 reaction was not only efficient but also flexible enough to allow for precise control over the lipids’ molecular structure.”

Michael J. Mitchell, Associate Professor, University of Pennsylvania

This adaptability was essential for optimizing the ionizable lipid characteristics for secure and efficient mRNA delivery.

Why This Advance Matters

mRNA-based vaccines and treatments, which are positioned to cure a variety of ailments, from infectious diseases to genetic problems, are anticipated to be significantly impacted by this novel approach to creating ionizable lipids.

Editing genes that cause familial amyloidosis, a rare illness that causes aberrant protein deposits throughout the body, and enhancing the delivery of the COVID-19 mRNA vaccine are two high-priority applications for which the modified lipids in this work increased mRNA transport in preclinical models. In both instances, the modified lipids outperformed the industry-standard lipids already in use.

The novel strategy may hasten the development of mRNA therapeutics in general, independent of these particular uses. Traditional approaches can take years to generate an effective lipid, but the team's guided evolution process may shorten that time to a matter of weeks or months.

Our hope is that this method will accelerate the pipeline for mRNA therapeutics and vaccines, bringing new treatments to patients faster than ever before.”

Michael J. Mitchell, Associate Professor, University of Pennsylvania

A New Frontier for mRNA Delivery

The effectiveness of LNPs, a versatile and safe method of delivering genetic material, depends on the characteristics of their ionizable lipids. Researchers can now enhance these lipids with previously unheard-of speed and accuracy thanks to the Penn Engineers' iterative design methodology, which is advancing the development of the next generation of mRNA treatments.

Penn Engineers' novel LNP formula represents a significant advancement in mRNA technology, providing hope for a quicker and more effective route to medicines that could change people's lives.