Recent advancements in lipid–mRNA nanoparticles as a treatment option for cancer immunotherapy

Purpose: mRNA is an attractive platform for cancer immunotherapy, but its instability, susceptibility to RNase degradation, and inefficient endosomal escape limit therapeutic applications. Lipid-based nanocarriers are non-viral vectors that can protect mRNA, improve transfection, and deliver it to intracellular compartments suitable for translation.

Hypothesis: If mRNA is formulated into lipid-based nanoparticles—lipoplexes, cationic or ionizable LNPs, lipopolypexes, or lipid calcium phosphate nanoparticles—with optimized lipid composition, charge, pKa, and targeting, then mRNA can be stabilized, delivered to target cells, translated into antigens or immunomodulatory proteins, and enhance antitumor immunity for cancer immunotherapy.

Aims: Review lipid-based mRNA nanoparticle systems for cancer immunotherapy. - Describe structures and mechanisms of lipoplexes, cationic/ionizable LNPs, lipopolypexes, and lipid calcium phosphate nanoparticles. - Discuss cellular internalization and endosomal escape. - Highlight applications in cancer vaccines, CAR T-cell therapy, and monoclonal antibody delivery. - Summarize clinical trials and future perspectives.

Delivery system: Platform: lipid-based mRNA nanoparticles, including lipoplexes, cationic LNPs, ionizable LNPs, lipid–polymer hybrid nanoparticles (lipopolypexes), and lipid calcium phosphate (LCP) nanoparticles. - Key lipids: DOTMA, DOTAP, DOPE, ePC; ionizable lipids DODAP, DODMA, DLin-DMA, DLin-KC2-DMA, DLin-MC3-DMA, L5, L101, ATX100, ssPalmE, OF-02, A18-Iso5-2DC18; helper lipids cholesterol, DSPC, PEG-lipids. - Payloads: mRNA encoding tumor antigens, neoantigens, CAR, monoclonal antibodies, and immunomodulatory proteins; also siRNA co-delivery. - Targeting/functionalization: mannose for dendritic cells, lipid-to-mRNA ratio for organ tropism (anionic → spleen; cationic → lungs), and STING-activating zwitterionic lipids. - Preparation: microfluidic mixing, electrostatic complexation, and combinatorial lipidoid screening.

Approach: Review and synthesis of preclinical and clinical literature. In vitro models include murine bone marrow-derived DCs, human monocyte-derived DCs, T cells, and various cancer cell lines. In vivo models include murine tumor models. Clinical trials cited include NCT03480152, NCT03164772, NCT01446731, NCT03948763, NCT00890032, NCT03468244, and NCT00204516. As a review, it reports no primary experimental groups, n values, doses, or controls.

Key methods: No primary methods. The review discusses data generated by cited studies using: - mRNA encapsulation and transfection assays. - Protein expression and antigen presentation assays. - Flow cytometry for immune cell activation and CAR expression. - Cytotoxicity and hemocompatibility assays. - In vivo tumor growth, survival, and immune response measurements. - Clinical trial safety, immunogenicity, and efficacy readouts.

Key results: Optimal pKa for ionizable lipids: 6.2–6.5 for intravenous mRNA delivery; 6.6–6.9 for intramuscular mRNA delivery. - Modifying lipid-to-mRNA ratio can shift lipoplex charge: anionic lipoplexes target spleen, cationic formulations target lungs. - DOTAP + cholesterol efficiently transfects primary bone marrow-derived DCs. - C14-4 LNPs for CAR T-cell engineering produced comparable CAR surface expression to electroporation with reduced T cell toxicity. - Pam3-LNPs induced increased antigen-specific CD8+ T cells. - PD-L1/PD-L2 siRNA delivery via DLin-KC2-DMA LNPs efficiently reduced PD-L expression in human monocyte-derived DCs without affecting DC phenotype or migration. - L5 lipid showed efficient mRNA delivery in rodent and primate models with optimal pharmacokinetics and non-toxic side effects. - A18-Iso5-2DC18 LNPs target T cells in vivo, strongly bind STING, and induce potent cytolytic T lymphocyte responses and antitumor immunity. - Clinical trials listed include mRNA-4650 (NCT03480152), BI 136,849 + durvalumab + tremelimumab (NCT03164772), mRNA + docetaxel DCs (NCT01446731), mRNA-5671/V941 + pembrolizumab (NCT03948763), BTSC mRNA-loaded DCs (NCT00890032), personalized mRNA tumor vaccine (NCT03468244), and melanoma-associated antigen mRNA + GM-CSF (NCT00204516).

Interpretation: The authors conclude that lipid-mRNA nanoparticles are promising for cancer immunotherapy because they protect mRNA, improve transfection, and enable endosomal escape. They highlight applications in cancer vaccines, CAR T-cell therapy, and monoclonal antibody delivery, with several formulations in clinical trials. They predict that novel ionizable lipids and optimized formulations will improve efficacy, and that combining mRNA-LNP systems with existing immunotherapies can enhance antitumor responses.

Limitations: Review article; no primary data, effect sizes, n values, doses, or controls. - mRNA is unstable and degraded by RNases; endosomal escape remains a major barrier. - Low transfection efficiency in lymphocytes and monocytes remains unresolved. - Cationic lipids can induce toxic pro-apoptotic and pro-inflammatory responses and liver damage. - Endosomal escape mechanisms are not fully understood. - Clinical translation is still limited; most data are preclinical. - Key variables affecting transfection efficiency—lipid composition, mRNA/lipid ratio, linker biodegradability, hydrophobic tail unsaturation, fusogenicity—require further study. - Personalized vaccines and combination therapies need further optimization and validation.