The cell membrane is notoriously selective about what it lets in. Charged molecules? Mostly rejected at the door. That’s a problem, because some of the most promising drug targets sit behind that barrier, and reaching them requires chemistry the membrane won’t tolerate.
Continue reading “Charged Molecular Glues: The Molecular Trojan Horse That Slipped Past Biology’s Toughest Bouncer “Lipid nanoparticles (LNPs) have transformed mRNA delivery. From COVID-19 vaccines to the first approved RNAi therapeutic, ONPATTRO (Patisiran), LNPs have proven their ability to ferry nucleic acid cargo into cells with speed and efficiency (Huang, 2019). Despite this transformation, most clinically deployed LNP formulations share a significant constraint: following intravenous administration, roughly 90% of the injected dose clears to the liver within an hour. If your target is a hepatocyte, that is hardly a hindrance. It’s a serious limitation if you need to reach the spleen, lymph nodes, pancreas or other extrahepatic tissues, all of which are organs of major interest in immunotherapy, vaccine development and metabolic disease research.
A new paper from Pieter Cullis’s laboratory at the University of British Columbia (UBC) offers a structural solution to that problem. Their design, termed a ‘liposomal LNP’, reengineers the architecture of the particle itself to achieve dramatically longer circulation lifetimes and improved transfection in tissues that standard formulations have largely missed.
LNPs enable fast transfection of a wide variety of cells, facilitating the transport of mRNA, DNA and siRNA into cells to induce transient expression in a short period of time (mere hours for mRNA and one to two days for DNA). They are a powerful tool that rose to broad public awareness through their use in COVID-19 vaccines, which delivered spike protein mRNA as cargo. Beyond vaccines, LNPs have been applied therapeutically as the delivery vehicle for ONPATTRO, which treats polyneuropathy of hereditary transthyretin-mediated amyloidosis (Huang, 2019).
The most widely studied LNP formulation, such as the ONPATTRO-like composition, consists of four components: an ionizable lipid, a helper lipid, cholesterol and a PEG-lipid. At physiological pH, the ionizable lipid is neutral and resides in a hydrophobic oil-like core surrounded by a lipid monolayer. This structure is highly effective at transfecting hepatocytes, but its rapid hepatic clearance limits its utility for reaching other tissues.
Standard ONPATTRO-like formulations have a lipid monolayer surrounding an oil droplet core. The UBC team’s publication reasoned that dramatically increasing the proportion of bilayer-forming lipids, specifically equimolar egg sphingomyelin (ESM) and cholesterol, could fundamentally change what the LNP looks like (Cheng, 2025).
The authors explored various bilayer-to-ionizable lipid molar ratios (RB/I) to see how they modified the structure of the particle. They found that an RB/I of 4 resulted in particles that transition to a true liposomal architecture consisting of a lipid bilayer enclosing an aqueous interior with a solid core suspended inside. Cryo-electron microscopy confirmed that approximately 84% of particles at this ratio adopt this bilayer structure, with the solid core occupying roughly 30% of the interior. LNP sizes across all tested ratios remained in the 40–60 nm range, confirming that the structural shift happens without meaningful changes in particle size.
The liposomal LNP exploits pH-driven structural transitions at both the assembly and delivery stages, explaining how a particle dominated by bilayer lipids can remain transfection-competent.
Assembly: When an ethanol-lipid mixture meets an acidic aqueous buffer (pH 4) containing mRNA, the positively charged ionizable lipid binds the negatively charged mRNA, forming a core complex. This complex acts as a nucleation point for the deposition of ESM/cholesterol bilayer lipid. As pH rises to 7.4 during formulation, ionizable lipids in the outer monolayer shift to a neutral form and migrate inward, expanding an oil droplet core. The mRNA dissociates from the oil core and resides in the aqueous interior where it is protected within the bilayer.
Delivery: After endocytosis, the acidic endosomal environment reverses this process. The ionizable lipids become positively charged again and migrate to the outer surface of the LNP, causing the solid mRNA-containing core to extrude outward from the liposomal bilayer. This positively charged protrusion interacts with the negatively charged endosomal membrane, triggering fusion and releasing the mRNA into the cytoplasm for translation. The authors describe this as a localized “warhead” mechanism—a structural consequence of the bilayer architecture, rather than a simple membrane-disruption event.
The authors utilized NanoLuc® mRNA as a reporter payload throughout the study. From in vitro transfection efficiency to whole-animal imaging, it allowed the authors to detect differential expression that would have been difficult to detect with less sensitive reporters.
The performance of the Liposomal LNP tells a compelling story. In vitro, the RB/I = 4 formulation matched or exceeded the transfection potency of the ONPATTRO-like composition in Huh7 cells across a wide dose range, while also proving to be the most stable on the shelf. After 63 weeks at 4°C, it maintained greater than 80% mRNA encapsulation with less than 20% size increase, and produced the highest mRNA integrity and translatability of any tested ratio.
SPECT/CT imaging of whole animals with the Liposomal LNP showed a circulation half-life approximately 15-fold longer than the standard ONPATTRO-like formulation, a direct consequence of the bilayer exterior adsorbing roughly half the plasma protein load. This reduced exterior plasma protein load means less macrophage recognition, less clearance and more time in circulation to reach tissues beyond the liver. That improved lifetime in circulation translated into improved tissue access. Ex vivo organ analysis showed 50-fold greater luminescence in the spleen and 150-fold greater in the inguinal lymph node compared to the standard formulation. Meaningful signal was also detected in the pancreas, a tissue rarely reached through conventional LNP formulations. Immunofluorescence confirmed delivery was localized to macrophages at the marginal zone of the spleen and subcapsular sinus of the lymph node.
It is also worth noting that the liposomal morphology held up when tested with the ionizable lipids used in the BNT162b2 and the mRNA-1273 COVID vaccines, suggesting this is a generalizable design.
Throughout this study, NanoLuc® mRNA served as the reporter payload. In vitro, NanoLuc® luminescence normalized to total protein provided a sensitive, linear measure of transfection efficiency across a wide dose range. In vivo, it enabled whole-animal IVIS imaging using the Nano-Glo® Fluorofurimazine substrate, with quantification extended to ex vivo organ homogenates using the Nano-Glo® Luciferase Assay System. NanoLuc® Luciferase sensitivity enabled detecting differential expression in tissues as small as inguinal lymph nodes and the pancreas. Detecting meaningful signal from a lymph node or pancreas can be challenging and thus successful detection demonstrates the exceptional performance of NanoLuc® Luciferase.
The authors synthesized their NanoLuc® mRNA in-house, a capability not universally available to research groups. We have partnered with the University of British Columbia RNA and Formulation Core to close that gap, enabling distribution of NanoLuc® mRNA across the core’s academic and industry network. Researchers who want to investigate LNP delivery, optimize formulations or validate mRNA constructs can now work with the RNA and Formulation Core to acquire NanoLuc® mRNA without the overhead of in-house synthesis. Work from UBC has contributed foundational understanding for LNP formulations, and now through their core they enable NanoLuc® mRNA development for any interested scientist.
The work from UBC demonstrates what becomes possible when mRNA delivery can reach beyond the liver, but the findings are only useful if researchers can access the tools to replicate and build on them. That’s where the UBC RNA and Formulation Core comes in. By partnering with Promega to distribute NanoLuc® mRNA, the Core gives researchers direct access to the same reporter used in this study, without the overhead of in-house synthesis. Whether you’re optimizing an LNP formulation, validating extrahepatic delivery or exploring mRNA constructs for a new application, you can now work with the Core to get started.
Interested in learning more about the UBC RNA and Formulation Core? Explore their website.
Learn more about the full NanoLuc® portfolio.
Cheng, M.H.Y. et al. (2025) Liposomal lipid nanoparticles for extrahepatic delivery of mRNA. Nature Communications 16, 4135.
Huang, Y.Y. (2019) Approval of the first-ever RNAi therapeutics and its technological development history. Prog. Biochem. Biophys. 46, 313–322.
Despite its many mysteries, a chrysalis is one of the most familiar transformations in nature. We know what goes in. We know what comes out. For a long time, what happened in between was essentially invisible to us. Not because we weren’t curious, but because the mechanism was sealed inside something the size of a thumbnail, and we had no way in.
This same invisibility exists on a much older and much larger scale.
Sometime around two billion years ago, a cell swallowed a bacterium and, instead of digesting it, kept it alive inside itself. This process, called endosymbiosis, is arguably the single most consequential event in the history of complex life. The bacterium became a permanent resident, and over billions of years of co-evolution, it became something else entirely: the mitochondria that power every complex cell on earth. Without it, the living world as we know it doesn’t exist.
Scientists have known for decades that this kind of cellular acquisition had to have occurred. What has proved harder to explain is not that it happened, but how it started. What did the earliest molecular steps actually look like from the inside?
In the ocean, there is a microscopic single-celled organism called Rapaza viridis. It hunts algae by propelling itself through the water on whip-like appendages called flagella. That hunt may be showing us the beginning of a modern endosymbiosis: the same process that gave every complex cell its mitochondria and every plant its chloroplasts.
Continue reading “Stolen Chloroplasts and the Chrysalis of Complex Life “
Most cancer research relies on young, healthy mice. Most cancer patients are not young. Could this disconnect between model organism and patient, which ignores the impact of the physiological realities of aging, explain why some therapies perform well in the lab but not in clinical trials? Focusing on lung cancer, a study published in Nature investigated what happens when these two realities, young and old, finally meet in the lab (1). The answers could reshape how we think about cancer research and the impact of aging on metastasis and disease progression.
Continue reading “New Study Suggests Cancer Research Has an Age Problem”
Kierkegaard observed that one of humanity’s enduring tensions is that while life can only be understood backwards, it must be lived forwards. It’s a truth medicine knows intimately: in the treatment that worked until it didn’t, the resistance that arrived without warning, the moment a doctor has to tell a patient that the drug that was helping has stopped. Not because anyone made a mistake, but because the critical knowledge that would have mattered arrived too late, if at all.
A recent paper from the National Cancer Institute is, in a small but meaningful way, science’s pursuit of that elusive foresight: an understanding that emerges early enough, for once, to change what happens next.
For decades, chemotherapy has worked by brute force, flooding the body with toxins designed to kill rapidly dividing cells. The problem is that rapid division isn’t unique to cancer. Hair follicle cells, gut lining cells and immune cells also divide rapidly, which is why patients lose hair, lose energy and become susceptible to infection. Chemotherapy targets a behavior, but the drug has no way to tell a healthy cell from a cancerous one.
Antibody-drug conjugates (ADCs) change that. Instead of targeting what cancer cells do, they target what cancer cells are. Cancer cells tend to display certain proteins on their surface in far greater numbers than healthy cells do. The antibody is engineered to seek out those proteins specifically. It navigates to its target, binds and waits for the cell to do what cells routinely do: pull it inside. Once there, the cell’s own digestive machinery (the lysosome) breaks down the chemical tether holding the toxin to the antibody, releasing the toxin to kill the cell from within. More than a dozen ADCs have received FDA approval in recent years, and the field is evolving fast.
But cancer cells don’t simply accept their fate. Even when an ADC delivers its payload perfectly—the antibody finds its target, the cell pulls it inside, the lysosome cuts the tether—a pump embedded in the cell membrane can grab the released toxin and throw it back out before it causes damage.
The delivery worked. The package got ejected anyway.
These pumps—ATP-binding cassette transporters, or more plainly, efflux pumps—are a normal feature of cell biology. Their job is cellular housekeeping, clearing out unwanted or toxic substances before they cause damage. Under the pressure of drug treatment, cancer cells do what life has always done under pressure: the ones best equipped to survive do. The same mechanism that has shaped living things for billions of years now works against the treatment. Not all cancer cells are identical, and the ones that happen to produce more pumps survive while others don’t, gradually shifting the tumor toward resistance.
Continue reading “Before the First Dose”
The brain is one of the most complex and fascinating parts of biology. Thankfully, it’s also remarkably good at protecting itself. When exposed to a pathogen, an injury or even misfolded proteins, microglia and astrocytes function as the central nervous system’s (CNS) primary immune defenders. They mount an inflammatory response by releasing cytokines and working to contain the damage. Yet this same system can malfunction or not resolve, which manifests as devastating consequences.
Chronic neuroinflammation is now recognized as a shared characteristic across some of the most common and difficult-to-treat neurological conditions. A 2023 review in Signal Transduction and Targeted Therapy highlighted the dualistic nature of neuroinflammation: while acute responses serve a protective role, chronic or dysregulated inflammatory signaling can initiate and accelerate neurodegeneration, identifying these pathways as priority targets for therapeutic intervention (Zhang et al., 2023). A 2025 review in Science reinforced this view, noting that within Multiple Sclerosis, disease-modifying therapies targeting neuroinflammation have seen the most clinical success (Shi & Yong, 2025). This could suggest applications within neurological conditions where the same inflammatory mechanisms are at work.
Understanding how and where these inflammatory signals originate in the CNS is an active area of preclinical research. One cytokine being actively studied is IL-6. IL-6 is produced by several cell types, including astrocytes and microglia in the CNS. As a key mediator of inflammatory responses, it mediates pro-inflammatory effects through its trans-signaling, which occurs via soluble IL-6 receptors. Dysregulation of this mechanism may contribute to the chronic neuroinflammation seen in several neurological conditions. Characterizing how and when IL-6 is secreted from CNS cells is an important step toward understanding the neuroinflammatory processes underlying these disorders.
Continue reading “Detecting Neuroinflammation in Microglia and Astrocytes”
Drug discovery researchers face a fundamental constraint in their work to develop safe, effective therapeutics: the vast majority of the human proteome remains inaccessible to conventional small molecule approaches. Proteins without defined binding pockets, those lacking known chemical probes, and protein targets that fail to translate from biochemical assays into cellular models have long been considered out of reach of standard drug discovery screening tools. As Dixit et al. describe, developing biochemical or cellular assays for all genome-encoded targets “is not scalable and likely impossible as most proteins have ill-defined or unknown activity” — these are what the authors call “the dark undruggable expanses” of the proteome [1].
That gap is now narrowing. Promega Corporation recently launched the TarSeer™ BRETSA™ Target Engagement System, a live-cell target engagement platform designed to bring previously challenging targets within reach of early-stage drug discovery.
Drug discovery teams regularly encounter a frustrating disconnect. A compound may show strong binding activity in a biochemical assay, only to fail when tested in a cellular environment. Without target-specific cellular assays, which generally aren’t available for poorly characterized proteins, researchers face difficult choices when deciding which compounds to advance through the drug development pipeline.
Continue reading “Why BRETSA™ Target Engagement Matters for Drug Discovery”
Adoptive T-cell therapies rely on generating metabolically fit, functional cells during ex vivo expansion—but this process often pushes T cells toward highly glycolytic, terminally differentiated states that limit their persistence and therapeutic potential. These metabolic programs begin shifting within hours of activation, therefore understanding early metabolic remodeling is essential for designing culture conditions that support durable, cytotoxic, and memory-enriched T-cell populations.
Researchers at Promega set out to address this challenge by systematically mapping how media composition and activation strength shape T-cell metabolism during the first 72 hours after stimulation. Using a suite of bioluminescent assays, they profiled intracellular energy cofactors, redox balance, and extracellular metabolites across several conditions. This approach revealed distinct, media-driven metabolic states that not only emerged early but also predicted downstream expansion, proliferation, and cytotoxic function.
Their work demonstrates how integrating metabolic profiling into in vitro expansion workflows can provide a more informed framework for optimizing T-cell manufacturing strategies.
Continue reading “Your Media Choice Might Be Designing Your T-Cell Fate”This blog was written by guest contributor Tian Yang, Associate Product Manager, Promega, in collaboration with Kristin Huwiler, Manager, Small Molecule Drug Discovery, Promega.
During the development of chemical probes or small-molecule drugs, compound affinity (Kd) or potency (IC50) is used to characterize compound-target interactions, to guide structure-activity relationship analysis and lead optimization and to assess compound selectivity.
However, neither parameter provides information on how quickly a compound engages with and dissociates from the target. The dissociation constant Kd reflects the relative concentrations of unbound and bound state of the compound at thermodynamic equilibrium, and while IC50 is an empirical metric that measures the concentration at which a biochemical or cellular process is reduced to half of the maximum value, IC50 values are typically determined when the process is assumed to be at equilibrium or steady-state. For a closed system, like cells in a culture dish, these thermodynamic parameters are quite informative. In an open system like the human body, where compound-target interactions often do not reach equilibrium, the kinetic parameters, in addition to the thermodynamic parameters, are needed to better understand and characterize compound target engagement over time (1,2).
Continue reading “Residence Time: The Impact of Binding Kinetics on Compound-Target Interactions”Platelets are best known for their role in blood clotting, but they also participate in other biological processes that influence how cells communicate and behave. In research models, scientists have observed that tumor cells can interact with platelets in ways that affect how they move and attach to new environments. A recent study by Morris et al., published in Scientific Reports, explored the molecular details behind these platelet–cell interactions and the role of calcium in regulating them.
The study focused on integrins, which are surface proteins that help cells anchor to their surroundings and communicate with the extracellular matrix. Two integrins, αIIbβ3 and αvβ3, are particularly important because they mediate platelet–platelet and platelet–cancer cell binding. Their structure and function depend on divalent cations such as calcium, which stabilize receptor conformation and support ligand binding.
When extracellular calcium levels were manipulated, platelet behavior changed in distinct ways.
Continue reading “How Calcium Shapes Cell Communication and Invasion”