Transfection Capability Research Articles

IPSC-mediated genetic manipulation promotes natural killer cell-centered cancer immunotherapy

Natural killer (NK) cells demonstrate potent cytotoxic activities and the capacity to secrete cytokines. Their distinctive capability to trigger cell death, bypassing the need for major histocompatibility complex recognition, opens promising avenues for their use in clinical settings such as allogeneic transplantation and tumor immunotherapy. Although the ability of NK cells to kill hematological tumors has been widely recognized, their effectiveness in treating solid tumors is not as pronounced. The intricate interplay of NK cells with the tumor microenvironment, specifically in the context of solid malignancies, has been noted to attenuate the anti-cancer prowess of NK cells and foster the ability of malignant cells to elude immune surveillance. Successful NK cell-centered immunotherapy hinges on obtaining a substantial quantity of NK cells with potent tumor-killing capabilities. However, the current challenge lies in the limited ex vivo expansion of NK cells and the inefficiency of gene introduction methods. Induced pluripotent stem cells (iPSCs) are multipotent stem cells with relatively easier gene transfection capability and theoretically unlimited proliferation potential. NK cells derived from iPSCs circumvent the challenge of difficult genetic modification in NK cells, offering various potential strategies to counteract the immune suppression induced by the tumor microenvironment.

Efficient mRNA Delivery In Vitro and In Vivo Using a Polycharged Biodegradable Nanomaterial.

As RNA rises as one of the most significant modalities for clinical applications and life science research, efficient tools for delivering and integrating RNA molecules into biological systems become essential. Herein, we report a formulation using a polycharged biodegradable nano-carrier, N1-501, which demonstrates superior efficiency and versatility in mRNA encapsulation and delivery in both cell and animal models. N1-501 is a polymeric material designed to function through a facile one-step formulation process suitable for various research settings. Its capability for mRNA transfection is investigated across a wide range of mRNA doses and in different biological models, including 18 tested cell lines and mouse models. This study also comprehensively analyzes N1-501's application for mRNA transfection by examining factors such as buffer composition and pH, incubation condition, and media type. Additionally, N1-501's superior in vivo mRNA transfection capability ensures its potential as an efficient and consistent tool for advancing mRNA-based therapies and genetic research.

Self‐promoted Controlled Ring‐opening Polymerization via Side Chain‐mediated Proton Transfer for the Synthesis of Tertiary Amine‐pendant Polypeptoids

AbstractProton transfer is essential in virtually all biochemical processes, with enzymes facilitating this transfer by optimizing the proximity and orientation of reactants through site‐specific hydrogen bonds. Proton transfer is also crucial in the rate‐determining step for the ring‐opening polymerization of N‐carboxyanhydrides (NCAs), widely used to prepare various peptidomimetic materials. This study utilizes side chain‐assisted strategy to accelerate the rate of chain propagation by using NCAs with tertiary amine pendants. This moiety enables hydrogen bond formation between the incoming NCA and the polymer amino growing end. The tertiary amine side chain of the NCA forms a proton shuttle, via a less constrained transition state, to facilitate the proton transfer process. Moreover, the tertiary amine side chains enable the precipitation of NCA monomers through in situ protonation during the monomer synthesis. This greatly facilitates the synthesis of these unreported monomers, allowing the direct controlled synthesis of tertiary amine‐pendant polypeptoids. This side chain‐promoted polymerization has rarely been reported. Additionally, the tertiary amine side chains, as widely used functional groups, endow the polymers with unique properties including pH‐ and thermo‐responsiveness, tunable pKas, and siRNA transfection capability. The self‐promoted synthesis, facile monomer preparation, and attractive properties make tertiary amine‐pendant polypeptoids promising materials for various applications.

Self-promoted Controlled Ring-opening Polymerization via Side Chain-mediated Proton Transfer for the Synthesis of Tertiary Amine-pendant Polypeptoids.

Proton transfer is essential in virtually all biochemical processes, with enzymes facilitating this transfer by optimizing the proximity and orientation of reactants through site-specific hydrogen bonds. Proton transfer is also crucial in the rate-determining step for the ring-opening polymerization of N-carboxyanhydrides (NCAs), widely used to prepare various peptidomimetic materials. This study utilizes side chain-assisted strategy to accelerate the rate of chain propagation by using NCAs with tertiary amine pendants. This moiety enables hydrogen bond formation between the incoming NCA and the polymer amino growing end. The tertiary amine side chain of the NCA forms a proton shuttle, via a less constrained transition state, to facilitate the proton transfer process. Moreover, the tertiary amine side chains enable the precipitation of NCA monomers through in situ protonation during the monomer synthesis. This greatly facilitates the synthesis of these unreported monomers, allowing the direct controlled synthesis of tertiary amine-pendant polypeptoids. This side chain-promoted polymerization has rarely been reported. Additionally, the tertiary amine side chains, as widely used functional groups, endow the polymers with unique properties including pH- and thermo-responsiveness, tunable pKas, and siRNA transfection capability. The self-promoted synthesis, facile monomer preparation, and attractive properties make tertiary amine-pendant polypeptoids promising materials for various applications.

DOTAP Modified Formulations of Aminoacid Based Cationic Liposomes for Improved Gene Delivery and Cell Viability.

The liposomal systems proved remarkably useful for the delivery of genetic materials but enhancing their efficacy remains a significant challenge. While structural alterations could result in the discovery of more effective transfecting lipids, improving the efficacy of widely used lipid carriers is also crucial in order to compete with viral vectors for gene delivery. Herein, we developed formulations of commercially available lipid, 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) with synthetic amino acid based cationic lipids. Two cationic lipids were synthesized using amino acids, with either cystine (CTT) or arginine (AT) in the head group. These lipids were used to formulate co-liposomal structures with different lipid compositions. The liposomal formulations were broadly categorised into two types: amino acid-based liposomes without DOTAP (CTTD and ATD) and those with DOTAP (DtATD and DtCTTD). Optimized lipid-DNA complexes of DOTAP-incorporated formulations (DtATD and DtCTTD) exhibited enhanced efficacy in transfection compared to formulations lacking DOTAP as well as commercial formulations such as DOTAP:DOPE. Notably, DtCTTD displayed superior transfection capabilities in prostate cancer (PC3) and lung cancer (A549) cell lines when compared to the widely used commercial transfection reagent, Lipofectamine. Collectively, the findings from this study suggest that DOTAP-incorporated formulations derived from amino acid-based liposomes, hold promise as effective tools for improving transfection efficacy with reduced toxicity, offering potential advancements in gene delivery applications.

Biocompatible fluorinated poly(2-hydroxypropenimine)s for safe and efficient gene transfection

The development of non-viral gene delivery systems often requires balancing transfection efficiency and cytotoxicity, particularly when using cationic polymers. Poly(2-hydroxypropenimine) (PHP), with its hydroxyl groups, presents a more biocompatible alternative to traditional polymers such as polyethyleneimine (PEI). In this study, we explored the potential to enhance PHP’s transfection capabilities and reduce its cytotoxicity through a targeted fluorination strategy. Specifically, fluorinated alkyl groups were grafted onto PHP via an ethylene oxide ring-opening reaction, with the reaction conditions optimized for temperature, solvent, and time to ensure efficiency. Transfection experiments using HEK 293 cells demonstrated that fluorinated PHP variants, especially PHP-FH816, significantly improved transfection efficiency, achieving a 75.1 % gene expression rate at an N/P ratio of 2, compared to 44.0 % for unmodified PHP. The transfection experiments in Hela cells also showed substantial transfection efficiency. The gene expression rate was 53.2 % at an N/P ratio of 2 compared to 31.6 % for unmodified PHP. Notably, the fluorinated polymers exhibited lower cytotoxicity and increased serum stability, which suggests their potential as safer gene delivery systems. This study carefully evaluates the feasibility of using fluorinated cationic polymers as next-generation gene delivery vectors, balancing efficacy and safety. It underscores the strategic application of fluorination technology in developing modified polymer vectors that could lead to more efficient and safer non-viral gene therapy solutions.

Effect of Degree of Substitution and Molecular Weight on Transfection Efficacy of Starch-Based siRNA Delivery System

RNA interference (RNAi) is a promising approach for gene therapy in cancers, but it requires carriers to protect and deliver therapeutic small interfering RNA (siRNA) molecules to cancerous cells. Starch-based carriers, such as quaternized starch (Q-Starch), have been shown to be biocompatible and are able to form nanocomplexes with siRNA, but significant electrostatic interactions between the carrier and siRNA prevent its release at the target site. In this study, we aim to characterize the effects of the degree of substitution (DS) and molecular weight (Mw) of Q-Starch on the gene silencing capabilities of the Q-Starch/siRNA transfection system. We show that reducing the DS reduces the electrostatic interactions between Q-Starch and siRNA, which now decomplex at more physiologically relevant conditions, but also affects additional parameters such as complex size while mostly maintaining cellular uptake capabilities. Notably, reducing the DS renders Q-Starch more susceptible to enzymatic degradation by α-amylase during the initial Q-Starch pretreatment. Enzymatic cleavage leads to a reduction in the Mw of Q-Starch, resulting in a 25% enhancement in its transfection capabilities. This study provides a better understanding of the effects of the DS and Mw on the polysaccharide-based siRNA delivery system and indicates that the polysaccharide Mw may be the key factor in determining the transfection efficacy of this system.

High Charge Density in Peptide Dendrimers is Required to Destabilize Membranes: Insights into Endosome Evasion.

Peptide dendrimers are a type of branched, symmetric, and topologically well-defined molecule that have already been used as delivery systems for nucleic acid transfection. Several of the most promising sequences showed high efficiency in many key steps of transfection, namely, binding siRNA, entering cells, and evading the endosome. However, small changes to the peptide dendrimers, such as in the hydrophobic core, the amino acid chirality, or the total available charges, led to significantly different experimental results with unclear mechanistic insights. In this work, we built a computational model of several of those peptide dendrimers (MH18, MH13, and MH47) and some of their variants to study the molecular details of the structure and function of these molecules. We performed CpHMD simulations in the aqueous phase and in interaction with a lipid bilayer to assess how conformation and protonation are affected by pH in different environments. We found that while the different peptide dendrimer sequences lead to no substantial structural differences in the aqueous phase, the total charge and, more importantly, the total charge density are key for the capacity of the dendrimer to interact and destabilize the membrane. These dendrimers become highly charged when the pH changes from 7.5 to 4.5, and the presence of a high charge density, which is decreased for MH47 that has four fewer titratable lysines, is essential to trigger membrane destabilization. These findings are in excellent agreement with the experimental data and help us to understand the high efficiency of some dendrimers and why the dendrimer MH47 is unable to complete the transfection process. This evidence provides further understanding of the mode of action of these peptide dendrimers and will be pivotal for the future design of new sequences with improved transfection capabilities.

Antigen Presenting Cell Mimetic Lipid Nanoparticles for Rapid mRNA CAR T Cell Cancer Immunotherapy.

Chimeric antigen receptor (CAR) T cell therapy has achieved remarkable clinical success in the treatment of hematological malignancies. However, producing these bespoke cancer-killing cells is a complicated ex vivo process involving leukapheresis, artificial T cell activation, and CAR construct introduction. The activation step requires the engagement of CD3/TCR and CD28 and is vital for T cell transfection and differentiation. Though antigen-presenting cells (APCs) facilitate activation in vivo, ex vivo activation relies on antibodies against CD3 and CD28 conjugated to magnetic beads. While effective, this artificial activation adds to the complexity of CAR T cell production as the beads must be removed prior to clinical implementation. To overcome this challenge, this work develops activating lipid nanoparticles (aLNPs) that mimic APCs to combine the activation of magnetic beads and the transfection capabilities of LNPs. It is shown that aLNPs enable one-step activation and transfection of primary human T cells with the resulting mRNA CAR T cells reducing tumor burden in a murine xenograft model, validating aLNPs as a promising platform for the rapid production of mRNA CAR T cells.

Cyclization-enhanced poly(β-amino ester)s vectors for efficient CRISPR gene editing therapy

Among non-viral gene delivery vectors, poly(β-amino ester)s (PAEs) are one of the most versatile candidates because of their wide monomer availability, high polymer flexibility, and superior gene transfection performance both in vitro and in vivo. Over two decades, PAEs have evolved from linear to highly branched structures, significantly enhancing gene delivery efficacy. Building on the proven efficient sets of monomers in highly branched PAEs (HPAEs), this work introduced a new class of cyclic PAEs (CPAEs) constructed via an A2 + B4 + C2 cyclization synthesis strategy and identified their markedly improved gene transfection capabilities in gene delivery applications. Two sets of cyclic PAEs (CPAEs) with rings of different sizes and topologies were obtained. Their chemical structures were confirmed via two-dimensional nuclear magnetic resonance and the photoluminescence phenomena, and their DNA delivery behaviours were investigated and compared with the HPAE counterparts. In vitro assessments demonstrated that the CPAEs with a macrocyclic architecture (MCPAEs), significantly enhanced DNA intracellular uptake and facilitated efficient gene expression while maintaining perfect biocompatibility. The top-performance MCPAEs have been further employed to deliver a plasmid coding dual single guide RNA-guided CRISPR-Cas9 machinery to delete COL7A1 exon 80 containing the c.6527dupC mutation. In recessive dystrophic epidermolysis bullosa (RDEB) patient-derived epidermal keratinocytes, MCPAEs facilitated the CRISPR plasmid delivery and achieved efficient targeted gene editing in multiple colonies.

Improving gene transfection efficiency of highly branched poly(β-amino ester)s through the in-situ conversion of inactive terminal groups

Highly branched poly(β-amino ester)s (HPAEs) have emerged as a safe and efficient type of non-viral gene delivery vectors. However, the presence of inactive terminal secondary amine groups compromises their gene transfection capability. In this study, HPAEs with similar topological structures and chemical compositions but varying numbers of terminal secondary 4-amino-1-butanol (S4) and secondary/tertiary 3-morpholinopropylamine (MPA) groups were synthesized. The results demonstrate that an increased number of secondary/tertiary MPA groups in-situ significantly enhances the DNA binding capability of HPAEs, leading to the formation of smaller HPAE/DNA polyplexes with higher zeta potential, ultimately resulting in superior gene transfection efficiency in bladder epithelial cells. This study establishes a simple yet effective strategy to maximize the gene transfection potency of HPAEs by converting the inactive terminal groups in-situ without the need for complex modifications to their topological structure and chemical composition.

Precise control of microfluidic flow conditions is critical for harnessing the in vitro transfection capability of pDNA-loaded lipid-Eudragit nanoparticles.

Microfluidics is widely regarded as a leading technology for industrial-scale manufacture of multicomponent, gene-based nanomedicines in a reproducible manner. Yet, very few investigations detail the impact of flow conditions on the biological performance of the product, particularly biocompatibility and therapeutic efficiency. Herein, this study investigated the engineering of a novel lipid-Eudragit hybrid nanoparticle in a bifurcating microfluidics micromixer for plasmid DNA (pDNA) delivery. Nanoparticles of ~150 nm in size, with uniform polydispersity index (PDI = 0.2) and ξ-potential of 5-11 mV were formed across flow rate ratios (FRR, aqueous to organic phase) of 3:1 and 5:1, respectively. The hybrid nanoparticles maintained colloidal stability and structural integrity of loaded pDNA following recovery by ultracentrifugation. Importantly, in vitro testing in human embryonic kidney cell line (HEK293T) revealed significant differences in biocompatibility and transfection efficiency (TE). Lipid-Eudragit nanoparticles produced at FRR 3:1 displayed high cellular toxicity (0-30% viability), compared with nanoparticles prepared at FRR 5:1 (50-100% viability). Red fluorescent protein (RFP) expression was sustained for 24-72 h following exposure of cells to nanoparticles, indicating controlled release of pDNA and trafficking to the nucleus. Nanoparticles produced at FRR 5:1 resulted in markedly higher TE (12%) compared with those prepared at FRR 3:1 (2%). Notably, nanoparticles produced using the bench-scale nanoprecipitation method resulted in lower biocompatibility (30-90%) but higher RFP expression (25-38%). These findings emphasize the need for in-depth analysis of the effect of formulation and flow conditions on the physicochemical and biological performance of gene nanomedicines when transitioning from bench to clinic.

Alkylated Sulfonium Modification of Low Molecular Weight Polyethylenimine to Form Lipopolymers as Gene Vectors.

Hydrophobic modification of low molecular weight polyethylenimine (PEI) is an efficient method to form ideal gene-transfer carriers. Sulfonium-a combination of three different functional groups, was conjugated onto PEI 1.8k at a conjugation ratio of 1:0.1 to form a series of sulfonium PEI (SPs). These SPs were hydrophobically modified and characterized by Fourier transform infrared and HNMR. DNA-condensing abilities of SPs were tested with gel retardation experiment, and their cytotoxicity was evaluated via the MTT assay. The particle size and zeta potential of SP/DNA nanoparticles were measured and evaluated for cellular uptake and transfection ability on HepG2 cell line. The results showed that the sulfonium moiety was attached to PEI 1.8k with a high yield at a conjugation ratio of 1:0.1. SPs containing longer alkyl chains condensed DNA completely at an SP/DNA weight ratio of 2:1. The formed nanoparticle size was in the range of 168-265 nm, and the zeta potential was +16-45 mV. The IC50 values of SPs were 6.5-43.2 μg/mL. The cytotoxicity of SPs increased as the hydrophobic chain got longer. SP/DNA showed much stronger cellular uptakes than PEI 25k; however, pure SPs presented almost no gene transfection on cells. Heparin release experiment showed that SP's strong binding of DNA resulted in low release of DNA and thus hindered the gene transfection process. By mixing SP with PEI 1.8k, the mixture presented adjustable DNA binding and releasing. The mixture formed by 67% SP and 33% PEI 1.8k showed strong gene transfection. In conclusion, sulfonium is an effective linkage to carry hydrophobic groups to adjust cell compatibilities and gene transfection capabilities of PEI.

Design and synthesis of a shikimoyl-functionalized cationic di-block copolypeptide for cancer cell specific gene transfection.

Targeted and efficient gene delivery systems hold tremendous potential for the improvement of cancer therapy by enabling appropriate modification of biological processes. Herein, we report the design and synthesis of a novel cationic di-block copolypeptide, incorporating homoarginine (HAG) and shikimoyl (LSA) functionalities (HDA-b-PHAGm-b-PLSAn), tailored for enhanced gene transfection specifically in cancer cells. The di-block copolypeptide was synthesized via sequential N-carboxyanhydride (NCA) ring-opening polymerization (ROP) techniques and its physicochemical properties were characterized, including molecular weight, dispersity, secondary conformation, size, morphology, and surface charge. In contrast to the cationic poly-L-homoarginine, we observed a significantly reduced cytotoxic effect of this di-block copolypeptide due to the inclusion of the shikimoyl glyco-polypeptide block, which also added selectivity in internalizing particular cells. This di-block copolypeptide was internalized via mannose-receptor-mediated endocytosis, which was investigated by competitive receptor blocking with mannan. We evaluated the transfection efficiency of the copolypeptide in HEK 293T (noncancerous cells), MDA-MB-231 (breast cancer cells), and RAW 264.7 (dendritic cells) and compared it with commonly employed transfection agents (Lipofectamine). Our findings demonstrate that the homoarginine and shikimoyl-functionalized cationic di-block copolypeptide exhibits potent gene transfection capabilities with minimal cytotoxic effects, particularly in cancer cells, while it is ineffective for normal cells, indicative of its potential as a promising platform for cancer cell-specific gene delivery systems. To evaluate this, we delivered an artificially designed miRNA-plasmid against Hsp90 (amiR-Hsp90) which upon successful transfection depleted the Hsp90 (a chaperone protein responsible for tumour growth) level specifically in cancerous cells and enforced apoptosis. This innovative approach offers a new avenue for the development of targeted therapeutics with an improved efficacy and safety profile in cancer treatment.

Delivery of RNA to the Blood-Brain Barrier Endothelium Using Cationic Bicelles.

Blood-brain barrier (BBB) dysfunction is prevalent in Alzheimer's disease and other neurological disorders. Restoring normal BBB function through RNA therapy is a potential avenue for addressing cerebrovascular changes in these disorders that may lead to cognitive decline. Although lipid nanoparticles have been traditionally used as drug carriers for RNA, bicelles have been emerging as a better alternative because of their higher cellular uptake and superior transfection capabilities. Cationic bicelles composed of DPPC/DC7PC/DOTAP at molar ratios of 63.8/25.0/11.2 were evaluated for the delivery of RNA in polarized hCMEC/D3 monolayers, a widely used BBB cell culture model. RNA-bicelle complexes were formed at five N/P ratios (1:1 to 5:1) by a thin-film hydration method. The RNA-bicelle complexes at N/P ratios of 3:1 and 4:1 exhibited optimal particle characteristics for cellular delivery. The cellular uptake of cationic bicelles laced with 1 mol% DiI-C18 was confirmed by flow cytometry and confocal microscopy. The ability of cationic bicelles (N/P ratio 4:1) to transfect polarized hCMEC/D3 with FITC-labeled control siRNA was tested vis-a-vis commercially available Lipofectamine RNAiMAX. These studies demonstrated the higher transfection efficiency and greater potential of cationic bicelles for RNA delivery to the BBB endothelium.

Branch Unit Distribution Matters for Gene Delivery.

As a key nonviral gene therapy vector, poly(β-amino ester) (PAE) has demonstrated great potential for clinical application after two decades of development. However, even after extensive efforts in structural optimizations, including screening chemical composition, molecular weight (MW), terminal groups, and topology, their DNA delivery efficiency still lags behind that of viral vectors. To break through this bottleneck, in this work, a thorough investigation of highly branched PAEs (HPAEs) was conducted to correlate their fundamental internal structure with their gene transfection performance. We show that an essential structural factor, branch unit distribution (BUD), plays an important role for HPAE transfection capability and that HPAEs with a more uniform distribution of branch units display better transfection efficacy. By optimizing BUD, a high-efficiency HPAE that surpasses well-known commercial reagents (e.g., Lipofectamine 3000 (Lipo3000), jetPEI, and Xfect) can be generated. This work opens an avenue for the structural control and molecular design of high-performance PAE gene delivery vectors.

Evaluation of Poly(N-Ethyl Pyrrolidine Methacrylamide) (EPA) and Derivatives as Polymeric Vehicles for miRNA Delivery to Neural Cells.

MicroRNAs (miRNAs) are endogenous, short RNA oligonucleotides that regulate the expression of hundreds of proteins to control cells' function in physiological and pathological conditions. miRNA therapeutics are highly specific, reducing the toxicity associated with off-target effects, and require low doses to achieve therapeutic effects. Despite their potential, applying miRNA-based therapies is limited by difficulties in delivery due to their poor stability, fast clearance, poor efficiency, and off-target effects. To overcome these challenges, polymeric vehicles have attracted a lot of attention due to their ease of production with low costs, large payload, safety profiles, and minimal induction of the immune response. Poly(N-ethyl pyrrolidine methacrylamide) (EPA) copolymers have shown optimal DNA transfection efficiencies in fibroblasts. The present study aims to evaluate the potential of EPA polymers as miRNA carriers for neural cell lines and primary neuron cultures when they are copolymerized with different compounds. To achieve this aim, we synthesized and characterized different copolymers and evaluated their miRNA condensation ability, size, charge, cytotoxicity, cell binding and internalization ability, and endosomal escape capacity. Finally, we evaluated their miRNA transfection capability and efficacy in Neuro-2a cells and rat primary hippocampal neurons. The results indicate that EPA and its copolymers, incorporating β-cyclodextrins with or without polyethylene glycol acrylate derivatives, can be promising vehicles for miRNA administration to neural cells when all experiments on Neuro-2a cells and primary hippocampal neurons are considered together.

Synthesis and characterization of polyethyleneimine/silk fibroin/gold nanoparticle nanocomposites: Potential application as a gene carrier in breast cancer cell lines

There is a need in advancing the design of non-viral gene carriers, thus herein using the pulsed laser ablation in liquid (PLAL) technique, polyethyleneimine-capped gold nanoparticles (PEI-AuNPs) were synthesized and then combined with Bombyx mori silk fibroin (SF) to form the nanocomposites (PEI-SF-AuNPs). After the characterization analyses were carried out, PEI-AuNPs and PEI-SF-AuNPs were tested for their in vitro biocompatibility in MCF-7 and MDA-MB231 breast cancer cells. Finally, the transfection capability of the selected group was observed. PEI capping enabled the cationic modification of the AuNP surface and SF reduced the surface charge (from 35 to 4 mV). PEI-AuNPs were highly monodispersed in shape and size (8–9 nm). The nanoparticle group synthesized in 5% (w/v) PEI concentration (PEI-AuNP5) had increased cytotoxicity as compared to the group synthesized in 1% (w/v) PEI concentration (PEI-AuNP1), but the conjugation of SF with PEI-AuNPs alleviated this cytotoxic tendency significantly. The group (PEI-AuNP1) with the most negligible cytotoxicity proved to be successful in the transfection of siRNA analog, i.e., siGLO green transfection indicator. Overall, the PEI-AuNP1 nanocomposites could be suggested as potent siRNA carriers due to the above beneficial properties in gene-based breast cancer therapies.

Iron Oxide Nanoparticles Decorated with Functional Peptides for a Targeted siRNA Delivery to Glioma Cells.

Glioma is a deadly form of brain cancer, and the difficulty of treating glioma is exacerbated by the chemotherapeutic resistance developed in the tumor cells over the time of treatment. siRNA can be used to silence the gene responsible for the increased resistance, and sensitize the glioma cells to drugs. Here, iron oxide nanoparticles functionalized with peptides (NP-CTX-R10) were used to deliver siRNA to silence O6-methylguanine-DNA methyltransferase (MGMT) to sensitize tumor cells to alkylating drug, Temozolomide (TMZ). The NP-CTX-R10 could complex with siRNA through electrostatic interactions and was able to deliver the siRNA to different glioma cells. The targeting ligand chlorotoxin and cell penetrating peptide polyarginine (R10) enhanced the transfection capability of siRNA to a level comparable to commercially available Lipofectamine. The NP-siRNA was able to achieve up to 90% gene silencing. Glioma cells transfected with NP-siRNA targeting MGMT showed significantly elevated sensitivity to TMZ treatment. This nanoparticle formulation demonstrates the ability to protect siRNA from degradation and to efficiently deliver the siRNA to induce therapeutic gene knockdown.

Designing Synthetic Polymers for Nucleic Acid Complexation and Delivery: From Polyplexes to Micelleplexes to Triggered Degradation.

Gene delivery as a therapeutic tool continues to advance toward impacting human health, with several gene therapy products receiving FDA approval over the past 5 years. Despite this important progress, the safety and efficacy of gene therapy methodology requires further improvement to ensure that nucleic acid therapeutics reach the desired targets while minimizing adverse effects. Synthetic polymers offer several enticing features as nucleic acid delivery vectors due to their versatile functionalities and architectures and the ability of synthetic chemists to rapidly build large libraries of polymeric candidates equipped for DNA/RNA complexation and transport. Current synthetic designs are pursuing challenging objectives that seek to improve transfection efficiency and, at the same time, mitigate cytotoxicity. This Perspective will describe recent work in polymer-based gene complexation and delivery vectors in which cationic polyelectrolytes are modified synthetically by introduction of additional components─including hydrophobic, hydrophilic, and fluorinated units─as well as embedding of degradable linkages within the macromolecular structure. As will be seen, recent advances employing these emerging design strategies are promising with respect to their excellent biocompatibility and transfection capability, suggesting continued promise of synthetic polymer gene delivery vectors going forward.