Dayananda Sagar University, College of Pharmaceutical Sciences, Bengaluru, Karnataka, India.
Gene editing (e.g., CRISPR-Cas systems) and mRNA therapeutics hold great promise for addressing genetic and acquired diseases, but their clinical translation is impeded by delivery challenges including cargo instability, immunogenicity, off-target effects, and inadequate cellular uptake. MXenes and other two-dimensional (2D) transition metal carbides/nitrides have emerged as versatile nanomaterials with tunable surface chemistry, high surface area, and potential for functional modification. This review critically examines recent advances (2018–2025) in MXene-based and related 2D nanocarriers for CRISPR RNP, sgRNA, and mRNA delivery, focusing on: (i) materials design (surface terminations, defect control, hybridization); (ii) cargo loading and protection strategies; (iii) cellular uptake, endosomal escape, intracellular release; (iv) in vitro and in vivo efficacy; (v) biosafety, biodegradation, and immunogenicity; (vi) manufacturing, characterization and regulatory challenges. We highlight comparative strengths and limitations relative to lipid nanoparticles (LNPs), polymeric nanocarriers, and gold nanoparticles. Finally, we propose a translational roadmap to guide future research toward clinical application. This review aims to fill a gap in the literature: there is no existing comprehensive summary focused explicitly on MXene/2D carbide/nitride nanocarriers for gene editing and mRNA therapies.
Gene-editing technologies, notably CRISPR-Cas9, Cas12a, and base editors, together with messenger RNA (mRNA) therapeutics, have revolutionized the possibility of correcting genetic disorders, cancer immunotherapy, vaccination, and treating infectious diseases [1,2]. However, delivery remains a major bottleneck: nucleic acids and RNPs are sensitive to nuclease degradation, have poor cellular uptake, risk off-target distribution, and elicit immune responses [3,4].
Lipid nanoparticles (LNPs) and polymeric systems are the most advanced non-viral carriers; yet they have limitations including stability, scalability, endosomal escape efficiency, and immunogenicity [5,6]. In this context, two-dimensional transition metal carbides/nitrides, known as MXenes, have gained attention because of their unique physicochemical properties: large surface area, tunable surface terminations (–OH, –F, –O), high electrical conductivity, and facile functionalization [7,8].
MXene has been used in various applications in biosensing, diagnostic devices, and therapy in general [9,10], and other non-viral carriers for gene editing or mRNA [5,11]. However, there is no review that systematically brings together MXene materials specifically as nanocarriers for CRISPR RNPs and mRNA, analyzing design principles, cargo protection, biosafety, and translation.
2. MXene Family: Physical Structure, Surface Chemistry, and Synthesis Relevant to Nucleic Acid Cargo
MXenes are produced by etching MAX phase precursors (e.g., Ti?AlC?) to remove the ‘A’ layer (e.g., Al), resulting in 2D sheets with formula M???X?T? where M = transition metal (e.g., Ti, Nb), X = C or N, T = surface terminations (–O, –OH, –F) [12]. The nature and density of T terminals determine hydrophilicity, surface charge, biocompatibility, and capacity for functionalization [7,13]. Sheet size, layer thickness, defect density, oxidation state (which degrades over time in aqueous/oxidative environments) affect both loading capacity and stability of cargo [14,15].
Synthesis methods vary: (i) chemical etching (e.g., HF, LiF + HCl) to produce multilayer MXenes; (ii) delamination using intercalants (e.g., dimethyl sulfoxide) to get single- or few-layer MXene sheets [16,17]. Post-synthesis functionalization (polymer grafting, lipid coating, covalent ligand attachment) helps reduce immunogenicity and improve dispersibility in biological media [8,18].
3. Cargo Loading & Protection Strategies
3.1 Adsorption and Electrostatic Complexation
Because many MXenes (depending on surface terminations) are negatively charged or have mixed charges, their surfaces can bind positively charged nucleic acids or RNPs via electrostatic interactions. For example, Ti?C?Tx modified with polyethylenimine (PEI) has been reported to load sgRNA or mRNA and protect from RNase degradation [19].
3.2 Hybrid Coatings: Polymers, Lipids, and MOF Hybrids
Hybrid systems where MXenes are coated with polymers like PEG, PEI, or by lipid bilayers provide improved stability and biocompatibility, reduce aggregation, and help in endosomal escape [20,21]. MOF (metal-organic framework)-MXene hybrids have been explored in other cargo scenarios and could be promising for nucleic acids [22].
3.3 Lyophilization and Cryoprotectants
To improve storage stability of mRNA or RNPs bound to MXene, lyophilization with cryoprotectants (e.g., trehalose, sucrose) is essential to prevent aggregation and preserve activity after rehydration. Although explicit studies with MXene and mRNA are few, related nanocarrier studies show significant loss of expression without such stabilization [23,24].
4. Cellular Uptake, Endosomal Escape, and Intracellular Release
MXene nanocarriers can be internalized via endocytosis (clathrin-, caveolin- or macropinocytosis pathways) depending on size, surface charge, and functional ligands [25]. Surface functionalization with cell-penetrating peptides or targeting ligands increases specificity and uptake [26,27].
Endosomal escape is a key challenge: some hybrid MXene-polymer composites are able to disrupt endosomal membranes via the “proton sponge” effect (e.g., PEI coatings) or via pH-sensitive linkers that degrade in acidic endosomes [20]. Photo or redox responsive terminations (disulfide bonds, glutathione-sensitive linkers) have also been proposed in preliminary works [28].
Release of cargo (mRNA, RNP) inside the cytosol requires disassembly of carrier, minimal binding to allow function, and avoidance of degradation.
5. In Vitro and In Vivo Efficacy: Evidence Summary
|
Study |
Material |
Cargo Type |
Model (cell / animal) |
Output |
Toxicity |
|
Chen et al., 2022 [29] |
Ti?C?Tx-PEI nanosheets |
sgRNA + Cas9 RNP |
HEK293 cells |
~45% indel rate at targeted locus |
Moderate cytotoxicity at high PEI dose |
|
Li et al., 2023 [30] |
Ti?C?Tx-lipid hybrid |
mRNA (luciferase) |
Mice (intramuscular) |
~10-fold higher luminescence vs mRNA-LNP control |
Rapid clearance; mild inflammatory cytokines |
|
Wang et al., 2024 [31] |
Nb?C MXene nanosheets + PEG |
mRNA vaccine model (antigen mRNA) |
Mice (intravenous) |
Strong antigen expression, good immune response |
Long-term clearance (30 days) not reported |
|
Zhao et al., 2024 [32] |
Ti?C?Tx grafted with redox-sensitive polymer |
Cas12a RNP |
Human hepatocytes |
~40% editing; minimal off-target effects |
Low toxicity; but scale-up not addressed |
6. Biosafety, Biodegradation, Immunogenicity
6.1 Metal Ion Leaching and Oxidative Degradation
MXenes often contain residual metal ions (e.g., Ti, Nb), and when exposed to air or aqueous environment, can oxidize (forming oxides) altering structure and possibly releasing metal ions [15,33]. These may contribute to cytotoxicity, reactive oxygen species (ROS) generation, and inflammatory responses. Few studies have systematically quantified ion leaching in physiologically relevant conditions [34].
6.2 Immunogenicity and In-Vivo Clearance
Immune activation (e.g., cytokine secretion) has been observed in animal studies after systemic administration of MXene hybrids [30]. Protein corona formation can affect circulation time and biodistribution [35]. Reticuloendothelial system (RES) uptake (liver, spleen) is often high due to particle size and opsonization [31].
6.3 Biodegradation
Data on long-term degradation are sparse. Some MXenes degrade under oxidative conditions producing oxide byproducts, but clearance via renal or hepatic pathways, long-term accumulation, and histopathological effects are under-explored [31,36].
7. Comparison with Established Carriers
|
Carrier Type |
Stability |
Delivery Efficiency |
Immunogenicity |
Manufacturing / Scalability |
Regulatory Readiness |
|
LNPs |
Good (especially with ionizable lipids) |
High for mRNA; less so for large RNPs |
Known immune activation; PEG concerns [37] |
Scalable; several in clinical use |
Several clinically approved (e.g., COVID-19 vaccines) |
|
Polymer Nanoparticles (PEI, polyplexes) |
Variable; often less stable |
Moderate; issues of endosomal escape |
PEI high cytotoxicity; immunogenicity concerns [38] |
Easier synthesis; but batch variation problematic |
Some in trials; regulatory concerns on toxicity |
|
Gold Nanoparticles |
Excellent stability; easy functionalization |
Decent for small nucleic acid; for large complexes lower efficiency |
Generally low immunogenicity; but accumulation risk [39] |
Scalable; but cost and metal accumulation worrying |
Few clinical uses for gene therapy delivery |
|
MXene / 2D carbides/ nitrides |
Promising surface tunability and high cargo loading; stability a concern due to oxidation |
Early studies show competitive editing / expression (~30-50%) [29-32] |
Early signs of immunogenicity; more data needed |
Synthesis of MXenes is improving; reproducibility and purity remain challenges |
No MXene-based nucleic acid therapeutic yet in clinic; regulatory parameters yet underdeveloped |
8. Manufacturing, Characterization, & Regulatory Challenges
Synthesis reproducibility & purity: Etching methods introduce residual acids/fluorides; batch-to-batch variability in size, oxidation state, surface terminations [16]. Purification and characterization (TEM, XRD, XPS) required.
Endotoxin, metal ion contamination, sterility: Regulatory bodies (FDA, EMA) require rigorous QC for gene therapy carriers; MXene nanomaterials must meet these for clinical translation [40].
Scalable functionalization: Coatings (polymer, lipid) or modifications must be scalable, stable, scalable cost, maintain reproducibility.
Regulatory toxicity testing: Chronic toxicity, accumulation, long-term biodistribution; clinical dosage scaling; need standardized animal model data.
Analytical standards for nucleic acid cargo delivery: Assays for quantifying RNP editing efficiency, off-target effects, immune activation, mRNA expression, in vivo and ex vivo, must be standardized.
9. Challenges & Future Directions
Figure 1. Translational roadmap for MXene-based CRISPR/mRNA nanocarriers
CONCLUSION
MXene and related 2D transition metal carbides/nitrides present an exciting and under-explored frontier in the delivery of gene-editing and mRNA therapeutics. Their unique physical and chemical characteristics, when properly engineered, could help overcome several limitations of existing carriers. However, significant challenges remain in ensuring biosafety, biodegradation, reproducibility, and regulatory compliance. With targeted future work especially in vivo safety, stimuli-responsive release, and head-to-head efficacy studies MXene-based platforms could become viable competitors to lipid nanoparticles and other established systems in clinical gene therapy and mRNA applications.
REFERENCES
Rakshitha T R, Rakshitha J S, Skandanaa , Greeshma R, MXene and 2D Transition Metal Carbide/Nitride Nanocarriers for Gene-Editing and mRNA Therapeutics: Design Principles, Biosafety Challenges, and Translational Roadmap, Int. J. of Pharm. Sci., 2025, Vol 3, Issue 9, 2433-2439. https://doi.org/10.5281/zenodo.17175096
10.5281/zenodo.17175096
