An mRNA cancer vaccine carries its own immune booster and lights up when it starts working, showing treatment success in mice within hours.
(Nanowerk Spotlight) Messenger RNA (mRNA) vaccines transformed infectious disease control, yet applying them to cancer remains a far tougher problem. Tumors originate from the body’s own tissue, making them difficult to distinguish from healthy cells. They also suppress immune responses that could destroy them.
To work, an mRNA cancer vaccine must deliver the genetic code for tumor antigens into the right immune cells and trigger a strong response, while providing early evidence that those cells are reacting effectively.
Lipid nanoparticles can protect fragile RNA and help it enter cells, but they often fail to generate enough immune activation for durable anti-tumor protection. Clinicians also lack a way to see whether the vaccine is taking effect until weeks later, when changes in antibodies or T cells become measurable.
Progress in RNA formulation and lipid chemistry has made targeted delivery possible, and advances in molecular imaging now allow scientists to monitor biological processes inside living tissue. But the two capabilities have rarely been integrated.
Design of the modular integrated platform for timely optimization of cancer immunotherapy. a) Schematic illustration of the interaction between vaccine delivery system optimization and vaccine efficacy assessment. b) Schematic illustration of the integrated modular platform combining self-adjuvanting PLNP mRNA delivery with early-stage, self-targeting vaccine efficacy monitoring for enhanced cancer immunotherapy. For the selfadjuvanting module: The modular self-adjuvanting peptide (C14-FK-Melittin) consists of melittin, a CTSB-responsive sequence (FK), and a C14 lipid tail. Together with ionizable lipids, phosphatidylcholine, cholesterol, and PEGylated lipids, this peptide forms the mRNA delivery vector. The resulting mRNA vaccine is administered intramuscularly. Upon endocytosis by DCs, C14-FK-Melittin releases melittin in response to CTSB, activating the STING pathway, while mRNA enters the cytoplasm to promote antigen expression. Mature DCs present these antigens to T cells, leading to T cell activation and subsequent tumor cell destruction. For the self-targeting imaging module: The VER probe tracks antigen presentation in real-time by harnessing the activity of ERAP1. Administered via footpad injection, VER is activated by ERAP1, emitting fluorescence to enable real-time monitoring of vaccine efficacy and antigen presentation. ER: Endoplasmic reticulum. MHC: Major histocompatibility complex. (Image: Reprinted with permission by Wiley-VCH Verlag) (click on image to enlarge)
At the core of this system is a peptide lipid nanoparticle, or PLNP, built from a clinically used lipid mixture containing an ionizable lipid, a helper phospholipid, cholesterol, and a polyethylene glycol lipid. These components form a stable structure that carries and protects the mRNA until it reaches immune cells.
The researchers added a modified form of melittin, a peptide from bee venom that can disrupt membranes and stimulate innate immunity. Because melittin is harmful if released too early, it was attached to the particle with a short linker recognized by cathepsin B, an enzyme common in the lysosomes of dendritic cells.
When dendritic cells internalize the nanoparticle, cathepsin B cleaves the linker, releasing melittin exactly where it can help mRNA escape into the cytoplasm and activate the STING pathway—a signaling system that induces interferon production and enhances antigen presentation.
The researchers tested several formulations, replacing between five and twenty percent of the main lipid with the melittin-based one. As melittin content increased, particle size grew slightly, but mRNA encapsulation remained high. In cultured human cells, the ten percent formulation produced the most protein from a test RNA, about four times higher than standard lipid particles. In dendritic cells derived from mouse bone marrow, this same formulation produced the greatest increase in maturation markers, indicating stronger immune activation.
In mice injected with luciferase mRNA, the melittin nanoparticles sustained gene expression and increased dendritic cell activation in nearby lymph nodes. The ten percent formulation produced nearly triple the number of mature dendritic cells compared with conventional particles. These nanoparticles also remained stable for at least a week when refrigerated and caused minimal damage to red blood cells or cultured immune cells.
To test antigen presentation, the researchers used mRNA encoding ovalbumin, a model antigen used in immunology. Dendritic cells transfected with melittin particles displayed the antigen on their surface ten times more often than untreated cells and roughly sixty percent more often than those treated with standard lipid nanoparticles. When cathepsin B cleavage was blocked or inhibited, activation dropped, confirming that melittin release inside lysosomes is essential for the effect.
Further analysis identified the STING pathway as the main driver of immune activation. Blocking STING reduced dendritic cell maturation about twofold, while inhibiting other innate immune sensors, such as toll-like receptors, had little effect. Microscopy showed STING clustering, a sign of activation. STING signaling increases production of type I interferons, which help cells present antigens to cytotoxic T cells.
This process raises levels of two key proteins: major histocompatibility complex class I (MHC I), which displays short antigen fragments on the cell surface, and ERAP1, an enzyme that trims those fragments to the correct length. Analysis of melanoma data sets revealed that tumors with higher STING expression also show higher MHC I and ERAP1 levels. Dendritic cells treated with the melittin nanoparticles exhibited similar increases, confirming the link between STING activation and antigen processing.
This insight led to the second element of the study: a fluorescent reporter that measures ERAP1 activity in real time. The reporter, called the vaccine efficacy reporter (VER), combines a near-infrared dye with a lipid polymer that directs it to lymph nodes. The dye remains non-fluorescent until ERAP1 removes a small leucine group, releasing light as a signal of enzyme activity.
In solution, the probe’s fluorescence increased fifteenfold after exposure to ERAP1. In dendritic cells, probe brightness rose when ERAP1 levels were boosted by interferon or when cells were treated with the melittin nanoparticles carrying antigen mRNA. The light signal matched the increase in MHC I expression, showing that the probe accurately reflected the antigen-processing step essential for T cell activation.
To evaluate the full system in animals, mice were immunized with different formulations and later injected with the fluorescent reporter. Within thirty minutes, lymph nodes from animals that received the melittin mRNA vaccine emitted about twice as much near-infrared light as those from control groups. The signal appeared in T cell–rich areas of the nodes, where immune priming occurs. Across mice, imaging intensity closely matched the number of mature dendritic cells and correlated with activated T cells in the spleen.
In tumor prevention experiments, vaccinated mice were challenged with melanoma cells expressing ovalbumin. Both RNA vaccines slowed tumor growth, but the melittin formulation produced the strongest effect, yielding tumors roughly thirteen times smaller than in untreated mice. Half of the vaccinated animals showed complete tumor disappearance and resisted a second tumor challenge without metastasis.
Their tumors and lymph nodes contained more activated dendritic cells, more helper and killer T cells, and fewer suppressive immune cells. The early imaging signal from the probe predicted these results: higher fluorescence corresponded to smaller tumors and stronger immune activation.
In a treatment model where mice already had tumors, the melittin mRNA vaccine again slowed tumor growth and extended survival. The lymph node signal correlated inversely with tumor size, serving as an early indicator of vaccine success. Tumor tissue from treated mice showed higher numbers of natural killer cells and fewer regulatory T cells, indicating a shift toward immune activity. Combining the vaccine with anti–PD-L1 therapy reduced tumor growth further, consistent with the idea that effective T cell priming enhances checkpoint blockade.
The study presents two linked advances. First, the delivery vehicle acts as both carrier and adjuvant, releasing melittin in a controlled way that enhances RNA translation and immune activation without excess toxicity. Second, the fluorescent reporter turns a hidden intracellular process—antigen trimming by ERAP1—into a visible signal that reveals vaccine activity in real time. Together these features create a platform that both delivers mRNA and reports whether the immune system is responding as intended.
The work remains preclinical and uses a model antigen, but its design points toward more adaptive vaccine strategies. By uniting delivery and monitoring, it shortens the time between vaccination and insight, allowing rapid optimization of dose and timing. The study illustrates how cancer vaccines could become not only more potent but also self-reporting, showing in real time how the immune system is being trained.
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