Polymeric Nucleic Acid Delivery for Immunoengineering

Purpose: Nucleic acids can be used to engineer the immune system, but their clinical use requires better specificity and intracellular delivery efficiency. Viral vectors have manufacturing, cargo-capacity, safety, and immunological drawbacks; therefore, polymers are being developed as non-viral delivery vehicles whose chemical and physical properties can be tuned to improve efficacy.

Hypothesis: As a review, this paper does not test a single formal hypothesis. Its central premise is: if polymers are formulated with nucleic acids — DNA, mRNA, siRNA, shRNA, or immunostimulatory nucleic acids — then they can enhance delivery to immune cells and enable prophylactic vaccines, therapeutic immunomodulation, gene overexpression/knockdown, and innate immune activation.

Aims: Describe polymers as a non-viral method for delivering nucleic acids for immunoengineering. - Review major nucleic acid cargo types and the intracellular barriers they face. - Discuss polymer classes and design features, including cationic complexation, degradable linkages, and targeting modifications. - Summarise immune cell targets and recent applications in prophylactic vaccines and therapeutic immunoengineering.

Delivery system: Platform: Polymer-based nucleic acid delivery vehicles. - Cationic polymers: Poly(L-lysine) (PLL), polyethylenimine (PEI), poly(beta-amino ester)s (PBAEs), PAMAM dendrimers, chitosan. These electrostatically complex negatively charged nucleic acids into nanoparticles, often approximately 50–200 nm. - Hydrophobic/encapsulating polymers: PLGA, PLA, poly(ortho ester)s (POEs); nucleic acids are physically entrapped, sometimes with polyamines to improve encapsulation and delivery. - Modifications: Degradable linkages (e.g., disulfide-containing PAMAM, bioreducible PBAEs), T-cell-specific ligands, mannose, folate. - Payloads: Plasmid DNA, mRNA, siRNA, shRNA, CpG oligodeoxynucleotides, poly(I:C), cyclic dinucleotides (CDNs), immunostimulatory RNA. - Formulations: Nanocomplexes, micro/nanoparticles, microneedle patches, hydrogels, intertwining DNA-RNA nanocapsules (iDR-NCs).

Approach: Narrative literature review covering preclinical in vitro, ex vivo, and in vivo studies. - Immune targets: Antigen-presenting cells (APCs), especially dendritic cells, macrophages, B cells, T cells, myocytes, and tumour cells. - Disease contexts: Infectious disease/HIV, cancer/melanoma, ovarian cancer, breast tumour, murine adenocarcinoma. - Model systems summarised: Mouse skin immunisation, tumour challenge models, immunocompetent mouse melanoma model, APC maturation and T-cell activation assays. - Design rigor: No systematic review, meta-analysis, or primary group structure is provided; evidence is synthesised from cited primary studies.

Key methods: The review summarises methods used in the cited primary studies rather than presenting new methods: - Nanocomplex/nanoparticle formation and characterisation. - Cellular uptake and APC internalisation. - APC maturation markers and cytokine secretion. - Antigen presentation and antibody responses. - CD4⁺ and CD8⁺ T-cell activation. - RNAi-mediated knockdown of immunosuppressive genes (e.g., PD-L1, STAT3, SOCS1). - In vivo tumour growth, survival, and vaccination challenge outcomes.

Key results: No primary quantitative results are reported; the review summarises qualitative outcomes from cited studies: - CDN/PBAE nanoparticles promoted APC uptake and significantly reduced tumour growth after intratumoral injection in an immunocompetent mouse melanoma model. - CDN-loaded hydrogels extended CDN release, slowed tumour growth, and drastically improved survival. - iDR-NCs co-delivering CpG ODN, STAT3 shRNA, and antigen trafficked to draining lymph nodes, increased APC maturation, and reduced tumour burden after vaccination. - PD-L1 siRNA/folate-PEI knockdown sensitised ovarian cancer cells to T-cell-mediated killing. - SOCS1 siRNA + ovalbumin in PLGA nanoparticles improved dendritic cell internalisation and ovalbumin-specific T-cell activation. - DNA/mannose polymer microneedles elicited a strong antibody response; DNA/poly(I:C)/PBAE microneedles generated strong CD4 and CD8 responses against an HIV antigen. - TNF-α plasmid nanocomplexes and IL-12 plasmid polyphosphazene slowed tumour growth in breast and adenocarcinoma models, respectively.

Interpretation: The authors conclude that polymeric nucleic acid delivery can overexpress or knock down specific genes, act as adjuvants or danger signals, and genetically modify immune cells. Cationic polymers facilitate cellular entry via electrostatic nanocomplexes, while hydrophobic polymers physically entrap nucleic acids. APCs are common targets, and these technologies may lead to new, effective immunoengineering strategies.

Limitations: This is a narrative review, not a systematic review or meta-analysis; no quantitative synthesis or risk-of-bias assessment is provided. - Most cited strategies remain preclinical; clinical translation is limited. - Non-viral transfection efficiency in APCs is generally much lower than viral methods. - Ex vivo dendritic cell manipulation is labour-intensive, costly, and challenging for broad patient application. - DNA vaccines tend to have poor immunogenicity; RNA is chemically unstable; PLGA encapsulation can suffer from poor nucleic acid stability and low encapsulation efficiency without added materials. - Chitosan has poor water solubility at neutral pH and moderate transfection efficacy; PEI is toxic; PAMAM dendrimers face a trade-off between low-molecular-weight efficacy and high-molecular-weight toxicity. - The review is from 2018, so it predates later advances in mRNA vaccines, CRISPR delivery, and newer polymeric systems.