Role of Lipid-Based and Polymer-Based Non-Viral Vectors in Nucleic Acid Delivery for Next-Generation Gene Therapy

Purpose: Nucleic acid analogs and genome-editing proteins have broad therapeutic potential, but clinical translation is limited by poor cell- and organ-specific delivery, endosomal entrapment, enzymatic degradation, and short bioavailability. The review examines lipid- and polymer-based non-viral vectors as strategies to overcome these barriers.

Hypothesis: As a review, no formal experimental hypothesis is tested. The central premise is that cationic lipid- and polymer-based non-viral vectors—especially ionizable lipids, biodegradable poly(amine-co-ester)s, and conjugate systems—can safely and effectively deliver nucleic acids and genome-editing cargo by improving encapsulation, cellular uptake, endosomal escape, and target-site delivery.

Aims: Summarize new-generation cationic lipids, cationic polymers, and their conjugates for nucleic acid delivery. - Review lipid-based vectors for mRNA, siRNA, and antisense oligonucleotide (ASO) delivery. - Review cationic polymeric vectors for mRNA, siRNA, and ASO delivery. - Discuss conjugate delivery systems, genome-editing delivery, clinical trials, and advantages/disadvantages of lipid- versus polymer-based vectors.

Delivery system: Lipid-based systems: liposomes, lipid nanoparticles (LNPs), stabilized plasmid lipid particles (SPLPs), ionizable amino lipids (DLin-MC3-DMA, C12-200, Lipid 5, DMAP-BLP, L101), helper lipids (DOPE, DSPC, cholesterol), PEGylated lipids. Polymer-based systems: poly-L-lysine (PLL), polyethylenimine (PEI), pDMAEMA, PAMAM dendrimers, poly(β-amino ester)s (PBAEs), hyperbranched PBAEs, poly(amino-co-ester)s (PACE, aPACE), PLGA, chitosan. Conjugates/targeting: antibody-conjugated LNPs, PBAE-coated gold nanoparticles, GalNAc-siRNA/ASO conjugates, dynamic polyconjugates (DPCs), cholesterol-siRNA. Payloads: plasmid DNA, mRNA, siRNA, ASO, miRNA inhibitors, ZFN mRNA, CRISPR/Cas9 mRNA/sgRNA/RNP. Disease targets: cancer, hereditary transthyretin amyloidosis, hypercholesterolemia, Zika virus, Friedreich’s ataxia, hepatitis B, glioblastoma, tissue transplantation, lung disease.

Approach: Review article; no original experimental groups, n, doses, or controls. It synthesizes preclinical studies in cell lines (HEK293, HeLa, A549, HUVEC, U87, RG2, MDA-MB-231, etc.) and animal models (mice, rats, cynomolgus monkeys) plus clinical trial data for non-viral vectors.

Key methods: Headline data summarized from cited studies include: luciferase/EPO reporter expression, target mRNA/protein knockdown (TTR, PCSK9, BCL-2, Nogo-B, miR-21, PLK-1), tumor growth/survival, biodistribution, serum neutralizing titers, clinical pharmacokinetics/pharmacodynamics, and endosomal escape/trafficking analyses (e.g., STORM imaging).

Key results: DOTMA liposomes: ~100% plasmid DNA entrapment and 5–100-fold higher transfection than calcium phosphate or DEAE-dextran. - SPLPs: ~70 nm particles with ~70% encapsulation; 100–1,000-fold gene expression in distal tumor tissue after IV administration; minimal toxicity at 175 µg plasmid, whereas liposome complexes showed significant toxicity above 20 µg. - Patisiran: first FDA-approved LNP-siRNA for hTTR amyloidosis; IV infusion in 29 patients reduced TTR by 20–30% at 24 h after first dose, 85% after second dose, and up to 96% knockdown after third dose at 0.3 mg/kg. - Biodegradable L101 LNP-siRNA targeting PCSK9: >90% protein silencing in non-human primates with higher hepatic clearance. - aPACE mRNA delivery: up to 10⁶-fold increase in vitro vs regular PACE; EPO mRNA produced high blood levels for up to 48 h without systemic toxicity. - PACE 70 siRNA targeting Nogo-B: >60% knockdown in liver after splenic injection in mice. - PBAE-coated gold nanoparticles + anti-luciferase siRNA: >90% luciferase gene silencing in HeLa cells. - DPC co-injection of Chol-siRNA and PBAVE polymer: 500-fold improvement in efficacy vs Chol-siRNA alone in mice. - ZFN mRNA LNPs targeting TTR and PCSK9: 80% and 90% protein reduction, respectively, at 10-fold lower mRNA doses. - CRISPR/Cas9 DNA nanoclaves + PEI coating targeting EGFP: 25% reduction in EGFP expression in U2OS.EGFP tumors after 10 days.

Interpretation: Non-viral vectors have enormous potential for next-generation gene therapy, with lipid-based vectors showing better clinical translation and manufacturing, and polymeric vectors offering tunability, sustained release, and functional conjugation. However, ligand-targeted delivery for cationic lipids and polymers still needs further exploration to improve clinical applicability. With advances in personalized medicine, these platforms may deliver therapeutic cargo to selected organs and treat a wide range of diseases.

Limitations: Review only; no new primary data. Cationic carriers can cause cellular toxicity and nonspecific serum-protein binding. Polymeric vectors face scale-up/manufacturing challenges and poor clinical translation; LNPs often have poor drug loading and require extensive formulation optimization. Many studies remain preclinical, and long-term safety, targeting specificity, and broad clinical efficacy are not yet established.