Lipid Nanoparticles (LNP) are a sophisticated technology used for drug development. With so many conditions stemming from aberrant cellular activity, the idea of delivering a biological “message in a bottle” seems almost obvious. Over the past few decades, many efforts were invested in finding the right technology to encapsulate biomolecules like mRNA, so that they are safely delivered to target cells. Indeed, the Nobel Prize in Physiology or Medicine was awarded in 2023 to Katalin Karikó and Drew Weissman who paved the path to the development of the mRNA-based Covid-19 vaccines.
One challenge still standing is the targeting of LNPs to specific cells and tissues in the body. A possible solution for this challenge is to load the LNPs with a ligand, a biomolecule that directs them to specific cells. Broadly speaking, these can be classified into two groups - modified lipids-based targeting and protein-based targeting.
Targeting based on modified lipids is more widely known, with several drugs at clinical trials utilizing this approach. One example is the development of inhaled RNA-based treatments for cystic fibrosis, using different lipid formulations to improve the uptake by lung cells1,2. Based on similar concepts, the approved chemotherapy drugs Myocet Liposomal and Doxil® both contain the same active compound, doxorubicin, but with different composition of lipids, that affect their tissue distribution3. Another interesting example can be found in the work of Kathryn A Whitehead from Carnegie Mellon University4, where the team assessed early-development mRNA-LNP-based drugs targeting pancreas cells using different ionized lipids. Interestingly, their work demonstrates that some LNPs are not taken directly by pancreas cells, but instead, first taken up by macrophages and only then transferred laterally into beta cells in the pancreas.
An honorary mention in this group has to go to the chemical modification of the RNA molecules themselves, mainly RNAi. This is pioneered by Alnylam5, which conjugates the sugar N-acetylgalactosamine (GalNAc) to RNA molecules and utilizes them as drugs. A few of their approved drugs include Givlaari®, Oxlumo® and more.
Protein-based targeting poses a greater challenge, and equally, a greater benefit when successful. To the best of our knowledge today, the specificity of protein-protein interactions, e.g., antibody-epitope, surpasses the specificity of lipid-protein or lipid-lipid interactions. Consequently, in cases of systemic administration, LNPs with protein-based targeting are more likely to travel throughout the body and be taken up by specific cells. A prime example for this is the approved drug Onpattro® (also by Alnylam) to treat the rare genetic disease Hereditary Variant Transthyretin Amyloidosis (ATTRv): A therapy based on RNAi, encapsulated by LNPs that bind the protein ApoE while in the bloodstream. Together they travel to the liver, where ApoE binds to cell-surface receptors, facilitating endocytosis into liver cells, where the RNAi inhibits the production of transthyretin, the protein that is responsible for the fibril deposits in the tissues6.
Another organ often targeted by LNPs is the bone marrow, including the hematopoietic stem cells in it: modifying bone marrow cells can be used to treat hereditary hematological conditions such as sickle-cell anaemia and thalassemia, as well as different hematological malignancies. Examples of such ligands include the cell surface receptors CD117, CD38 and CD477-9.
Intriguingly, antibody engineering also plays a role in LNP targeting, where attempts are made to load LNP with modified antibodies, including bispecific targeting antibodies10 and antibody fragments, such as Fab, which are smaller, therefore allowing loading of more units on each LNP11.
Loading LNPs with antibodies or antibody fragments raises several questions, among them, how do these proteins affect the overall stability of the particles and how does the antibody payload correlate with effective delivery and off-target effects. For example, will “antibody overload” lead to increased detection and opsonization by circulating immune cells? Interestingly, this could be either beneficial or harmful depending on the context and the desired target.
Considering these possibilities, correctly identifying, quantifying and tracking ligand introduction is immensely valuable. Analyzing ligand abundance at the early stages of development can be extremely cost-effective compared to relying on pre-clinical models only for these studies. Our LNP Profiler can detect antibody-based ligands and provide quantitative information on their abundance, as well as the fraction of LNPs that contain them, with or without nucleic acid cargo.