Brief Description of mRNA Delivery Systems
As a therapeutic use, mRNA cancer vaccines have the advantages of high safety, strong immune activation ability, and short production cycle. Moreover, mRNA cancer vaccines have shown potential for clinical application in multiple preclinical studies or clinical trials. These effective mRNA vaccines usually include three basic components, namely antigen, adjuvant and delivery system. The antigen can induce specific cellular immunity or humoral immunity, and achieve the desired immune effect with the help of specific adjuvants. Vaccine adjuvants are immune enhancers that increase the intensity of immune responses by binding to pattern recognition receptors (PRRs) of innate immune cells, ultimately enhancing the effectiveness of vaccines. In addition, naked mRNA is easily degraded, so it is necessary to match a suitable delivery system to maintain the stability of the mRNA and deliver it to the target cells.
Types of Delivery Systems of mRNA Cancer Vaccines
Lipid Nanoparticle (LNP)
Lipid nanoparticles (LNP) are the most cutting-edge and most concerned mRNA delivery carriers in clinical practice. LNP is an amphiphilic molecule that usually consists of four components: ionizable lipids, helper phospholipids, cholesterol, and PEGylated lipids. Among them, ionizable lipids combine with RNA (negative charge) to assist phospholipids to simulate cell membrane lipid bilayers, cholesterol can adjust the fluidity of lipid bilayers, and PEGylated lipids can improve the stability of nanoparticles. Two LNP-based mRNA COVID-19 vaccines developed by Moderna and Pfizer/BioNTech have been approved for marketing.
Several LNP-mRNA cancer vaccines are currently in clinical research stages. Among them, mRNA-4157 (also known as V940) has started Phase 3 clinical research. mRNA-4157 is a personalized mRNA cancer vaccine encapsulated by LNP and encoding 34 patient-specific tumor neoantigens. An early open-label phase 2 trial (KEYNOTE-942) enrolled 157 participants. The study data showed that compared with PD-1 monoclonal antibody alone, patients with mRNA-4157 combined with PD-1 monoclonal antibody had a higher risk of postoperative recurrence. The risk of death or death was reduced by 44%. On July 26, 2023, Moderna and Merck announced the launch of a Phase 3 trial, planning to enroll 1,089 patients, to evaluate the combination of personalized cancer vaccine mRNA-4157 and the PD-1 antibody pembrolizumab (Keytruda) For use in patients at high risk for melanoma who have undergone surgery. Final results are scheduled to be announced in 2029.
The transfection efficiency of LNP carrier is significantly better than other non-viral vectors, such as cationic lipids. However, the ratio of each lipid component will affect the delivery efficiency of LNP, so it is necessary to explore suitable LNP formulation to improve transfection or delivery efficiency when selecting LNP enveloped mRNA.
| BNT162b2 | mRNA-1273 | |
| Ionizable cationic Lipid | ALC-0315 | SM-102 |
| Sterol Lipids | 1,2-DSPC | 1,2-DSPC |
| PEGylated Lipids | Cholesterol | Cholesterol |
| Lipid molar ratio | 46.3:9.4:42.7:1.6 | 50:10:38.5:1.5 |
LNP components of BNT162b2 and mRNA-1273
Polymer Carrier
Common mRNA polymer carriers include PEI (Polyethyleneimine), polyester, polyamino acids and sugars (such as chitosan). Polyethylenimine is a cationic polymer material. The “proton sponge” effect of PEI can promote the escape of endolysosomes, protect mRNA from degradation, and safely release it into the cytoplasm. PEI can also exert an immune-stimulating effect. It is reported that PEI with higher molecular weight is cytotoxic. Combining PEI with biocompatible molecules polyethylene glycol or cyclodextrin can reduce the positive surface charge, or introduce hydrophobic molecules, which can reduce the cytotoxicity of PEI. Poly(beta-aminoester) was used as a DNA delivery carrier in its early days and is currently also used to deliver mRNA. PBAEs can bind to negatively charged mRNA to form complexes that promote cellular uptake and endolysosomal escape.
Overall, LNP is more widely used in mRNA cancer vaccines, and research on polymer carriers is slower.
Peptide Carrier
Protamine and cell-penetrating peptides are important cationic peptides for mRNA delivery, condensing and protecting mRNA through electrostatic interactions. CureVac, one of the three giants of mRNA, has established a protamine delivery vector platform (RNAactive technology platform). The pipeline that has entered the clinical stage includes CV9103, CV9104, CV9201, and CV9202, which are in the phase 1, phase 1/2 or phase 2 pre-research stage. Indications include a variety of solid tumors.
Cell penetrating peptide (CPP) is another peptide-based mRNA delivery carrier, typically containing 8 to 30 amino acids, that can penetrate biofilms and efficiently deliver mRNA to target cells.
Cell-based Delivery System
Cells are also an effective biological delivery carrier, capable of delivering molecules such as enzymes and mRNA to target sites. Dendritic cells (DC) are currently the most studied mRNA cell carriers, also known as mRNA-loaded DC cell vaccines. The mRNA encoding the antigen can be directly taken up by DCs through endocytosis. In fact, the first mRNA vaccine to enter clinical trials used DC cells as a carrier. However, DC cell vaccines require infusion of autologous DCs transfected with antigen mRNA in vivo, which is expensive and requires strict processes. Therefore, although DC cancer vaccines have demonstrated safety in clinical trials, clinical responses in patients have been sporadic or limited.
Extracellular Vesicle Carrier
Extracellular vesicles (EVs) are another type of mRNA delivery carrier. EVs in mammals can be divided into three types based on size and function: exosomes, microvesicles and apoptotic bodies ( apoptotic bodies). Among them, exosomes, with a size of 50-150 nm, are the most studied and characterized EVs. Exosomes are important messengers in the exchange of information between cells and can deliver and release molecules into the cytoplasm of the recipient, including mRNA, non-coding RNA, proteins, and lipids. The difficulty in developing exosome vectors lies in the complexity of their large-scale purification processes.
CDK-002 (exoSTING) is an mRNA cancer vaccine based on exosome delivery carrier. In an open-label Phase 1/2 clinical trial (NCT04592484), researchers evaluated the efficacy of exoSTING monotherapy in the treatment of advanced refractory solid tumors. Results showed that repeated dosing of exoSTING was well tolerated across all dose cohorts, and tumor shrinkage was observed in a subset of patients in both injected and uninjected lesions.
Mixed Carrier
Delivery of mRNA cancer vaccines can also use carriers made from a mixture of components, such as lipids, polymers, peptides, exosomes, and cell membranes. Mixed carriers can add the properties developers desire while maintaining the performance of each component. Lipid-polymer complexes in mixed carriers have been extensively studied and are generally composed of ionizable lipids, hydrophobic polymers, and polyethylene glycol lipids. High transfection efficiency can be achieved through component and ratio optimization. Cationic nanoemulsion (CNE) is an oil-in-water delivery vehicle composed of lipids and polymers. The lipid component includes DOTAP, and the nanoemulsion includes MF59 and AS03. It is also an approved vaccine adjuvant. Cationic nanoemulsions are also promising mRNA delivery vehicles.