bims-novged Biomed News
on Non-viral vectors for gene delivery
Issue of 2022‒11‒13
fourteen papers selected by
the Merkel lab
Ludwig-Maximilians University
  1. On the Influence of Nucleic Acid Backbone Modifications on Lipid Nanoparticle Morphology.
  2. Sodium Alginate-Doping Cationic Nanoparticle As Dual Gene Delivery System for Genetically Bimodal Therapy.
  3. Hyperbranched Poly(β-amino ester)s (HPAEs) Structure Optimisation for Enhanced Gene Delivery: Non-Ideal Termination Elimination.
  4. Spermidine/Spermine N1-Acetyltransferase 1 (SAT1)-A Potential Gene Target for Selective Sensitization of Glioblastoma Cells Using an Ionizable Lipid Nanoparticle to Deliver siRNA.
  5. Gelatin-Based Electrospun Nanofibers Cross-Linked Using Horseradish Peroxidase for Plasmid DNA Delivery.
  6. A Comparative Study of Mesoporous Silica and Mesoporous Bioactive Glass Nanoparticles as Non-Viral MicroRNA Vectors for Osteogenesis.
  7. Plasmid-DNA Delivery by Covalently Functionalized PEI-SPIONs as a Potential 'Magnetofection' Agent.
  8. Antibody-mediated delivery of CRISPR-Cas9 ribonucleoproteins in human cells.
  9. Non-lysosomal Route of mRNA delivery and Combining with Epigenetic Regulation Optimized Antitumor Immunoprophylactic Efficacy.
  10. Stability Study of mRNA-Lipid Nanoparticles Exposed to Various Conditions Based on the Evaluation between Physicochemical Properties and Their Relation with Protein Expression Ability.
  11. Silencing of proinflammatory NF-κB and inhibition of herpes simplex virus (HSV) replication by ultrasmall gold nanoparticles (2 nm) conjugated with small-interfering RNA.
  12. Nebulization of extracellular vesicles: A promising small RNA delivery approach for lung diseases.
  13. Single-helical formyl β-glucan effectively deliver CpG DNA with poly(dA) to macrophages for enhanced vaccine effects.
  14. An Innovative PTD-IVT-mRNA Delivery Platform for CAR Immunotherapy of ErbB(+) Solid Tumor Neoplastic Cells.
  1. Langmuir. 2022 Nov 11.
    On the Influence of Nucleic Acid Backbone Modifications on Lipid Nanoparticle Morphology.
    Kevin An, Daniel Kurek, Mark Mahadeo, Yao Zhang, Jenifer L Thewalt, Pieter R Cullis, Jayesh A Kulkarni.
      Nucleic acid therapeutics represent a major advance toward treating diseases at their root cause. However, nucleic acids are prone to degradation by serum endonucleases, clearance through the immune system, and rapid degradation in complex medium. To overcome these barriers, nucleic acids frequently include chemical modifications to improve stability or decrease immune responses. Lipid nanoparticles (LNPs) have enabled a dramatic reduction in the dose required to achieve a therapeutic effect by protecting these nucleic acids and improving their intracellular delivery. It has been assumed thus far that nonspecific ionic interactions drive LNP formation and chemical modifications to the nucleic acid backbone to confer improved stability do not impact LNP delivery in any way. Here, we demonstrate that these chemical modifications do impact LNP morphology substantially, and phosphorothioate modifications produce stronger interactions with ionizable amino lipids, resulting in enhanced entrapment. This work represents a major first step toward greater understanding of the interaction between the lipid components and nucleic acids within an LNP.
    DOI:  https://doi.org/10.1021/acs.langmuir.2c01492
  2. Biomacromolecules. 2022 Nov 08.
    Sodium Alginate-Doping Cationic Nanoparticle As Dual Gene Delivery System for Genetically Bimodal Therapy.
    Yu-Rong Zhan, Ping Chen, Xi He, Meng-Wei Hei, Ji Zhang, Xiao-Qi Yu.
      Photodynamic therapy occupies an important position in cancer therapy because of its minimal invasiveness and high spatiotemporal precision, and photodynamic/gene combined therapy is a promising strategy for additive therapeutic effects. However, the asynchronism and heterogeneity between traditional chemical photosensitizers and nucleic acid would restrict the feasibility of this strategy. KillerRed protein, as an endogenous photosensitizer, could be directly expressed and take effect in situ by transfecting KillerRed reporter genes into cells. Herein, a simple and easily prepared sodium alginate (SA)-doping cationic nanoparticle SA@GP/DNA was developed for dual gene delivery. The nanoparticles could be formed through electrostatic interaction among sodium alginate, polycation, and plasmid DNA. The title complex SA@GP/DNA showed good biocompatibility and gene transfection efficiency. Mechanism studies revealed that SA doping could facilitate the cellular uptake and DNA release. Furthermore, SA@GP/DNA was applied to the codelivery of p53 and KillerRed reporter genes for the synergistic effect combining p53-mediated apoptosis therapy and KillerRed-mediated photodynamic therapy. The ROS generation, tumor cell growth inhibition, and apoptosis assays proved that the dual-gene transfection could mediate the better effect compared with single therapy. This rationally designed dual gene codelivery nanoparticle provides an effective and promising platform for genetically bimodal therapy.
    DOI:  https://doi.org/10.1021/acs.biomac.2c01119
  3. Nanomaterials (Basel). 2022 Nov 04. pii: 3892. [Epub ahead of print]12(21):
    Hyperbranched Poly(β-amino ester)s (HPAEs) Structure Optimisation for Enhanced Gene Delivery: Non-Ideal Termination Elimination.
    Yinghao Li, Zhonglei He, Jing Lyu, Xianqing Wang, Bei Qiu, Irene Lara-Sáez, Jing Zhang, Ming Zeng, Qian Xu, Sigen A, James F Curtin, Wenxin Wang.
      Many polymeric gene delivery nano-vectors with hyperbranched structures have been demonstrated to be superior to their linear counterparts. The higher delivery efficacy is commonly attributed to the abundant terminal groups of branched polymers, which play critical roles in cargo entrapment, material-cell interaction, and endosome escape. Hyperbranched poly(β-amino ester)s (HPAEs) have developed as a class of safe and efficient gene delivery vectors. Although numerous research has been conducted to optimise the HPAE structure for gene delivery, the effect of the secondary amine residue on its backbone monomer, which is considered the non-ideal termination, has never been optimised. In this work, the effect of the non-ideal termination was carefully evaluated. Moreover, a series of HPAEs with only ideal terminations were synthesised by adjusting the backbone synthesis strategy to further explore the merits of hyperbranched structures. The HPAE obtained from modified synthesis methods exhibited more than twice the amounts of the ideal terminal groups compared to the conventional ones, determined by NMR. Their transfection performance enhanced significantly, where the optimal HPAE candidates developed in this study outperformed leading commercial benchmarks for DNA delivery, including Lipofectamine 3000, jetPEI, and jetOPTIMUS.
    Keywords:  DNA delivery; gene therapy; hyperbranched polymers; poly(β-amino ester)s; polymeric nano vectors; transfection
    DOI:  https://doi.org/10.3390/nano12213892
  4. Cancers (Basel). 2022 Oct 22. pii: 5179. [Epub ahead of print]14(21):
    Spermidine/Spermine N1-Acetyltransferase 1 (SAT1)-A Potential Gene Target for Selective Sensitization of Glioblastoma Cells Using an Ionizable Lipid Nanoparticle to Deliver siRNA.
    Vinith Yathindranath, Nura Safa, Babu V Sajesh, Kelly Schwinghamer, Magimairajan Issai Vanan, Rashid Bux, Daniel S Sitar, Marshall Pitz, Teruna J Siahaan, Donald W Miller.
      Spermidine/spermine N1-acetyltransferase 1 (SAT1) responsible for cell polyamine catabolism is overexpressed in glioblastoma multiforme (GB). Its role in tumor survival and promoting resistance towards radiation therapy has made it an interesting target for therapy. In this study, we prepared a lipid nanoparticle-based siRNA delivery system (LNP-siSAT1) to selectively knockdown (KD) SAT1 enzyme in a human glioblastoma cell line. The LNP-siSAT1 containing ionizable DODAP lipid was prepared following a microfluidics mixing method and the resulting nanoparticles had a hydrodynamic size of around 80 nm and a neutral surface charge. The LNP-siSAT1 effectively knocked down the SAT1 expression in U251, LN229, and 42MGBA GB cells, and other brain-relevant endothelial (hCMEC/D3), astrocyte (HA) and macrophage (ANA-1) cells at the mRNA and protein levels. SAT1 KD in U251 cells resulted in a 40% loss in cell viability. Furthermore, SAT1 KD in U251, LN229 and 42MGBA cells sensitized them towards radiation and chemotherapy treatments. In contrast, despite similar SAT1 KD in other brain-relevant cells no significant effect on cytotoxic response, either alone or in combination, was observed. A major roadblock for brain therapeutics is their ability to cross the highly restrictive blood-brain barrier (BBB) presented by the brain microcapillary endothelial cells. Here, we used the BBB circumventing approach to enhance the delivery of LNP-siSAT1 across a BBB cell culture model. A cadherin binding peptide (ADTC5) was used to transiently open the BBB tight junctions to promote paracellular diffusion of LNP-siSAT1. These results suggest LNP-siSAT1 may provide a safe and effective method for reducing SAT1 and sensitizing GB cells to radiation and chemotherapeutic agents.
    Keywords:  Spermidine/spermine N1-acetyltransferase 1 (SAT1); blood–brain barrier (BBB); brain drug delivery; cadherin peptide; gene therapy; glioblastoma (GB); lipid nanoparticles; microfluidic mixing; siRNA; transient modulation; tumor sensitization
    DOI:  https://doi.org/10.3390/cancers14215179
  5. Biomolecules. 2022 Nov 04. pii: 1638. [Epub ahead of print]12(11):
    Gelatin-Based Electrospun Nanofibers Cross-Linked Using Horseradish Peroxidase for Plasmid DNA Delivery.
    Kotoko Furuno, Keiichiro Suzuki, Shinji Sakai.
      The delivery of nucleic acids is indispensable for tissue engineering and gene therapy. However, the current approaches involving DNA/RNA delivery by systemic and local injections face issues such as clearance, off-target distribution, and tissue damage. In this study, we report plasmid DNA (pDNA) delivery using gelatin electrospun nanofibers obtained through horseradish peroxidase (HRP)-mediated insolubilization. The nanofibers were obtained through the electrospinning of an aqueous solution containing gelatin possessing phenolic hydroxyl (Ph) moieties (Gelatin-Ph) and HRP with subsequent HRP-mediated cross-linking of the Ph moieties by exposure to air containing 16 ppm H2O2 for 30 min. Then, Lipofectamine/pDNA complexes were immobilized on the nanofibers through immersion in the solution containing the pDNA complexes, resulting in transfection and sustained delivery of pDNA. Cells cultured on the resultant nanofibers expressed genome-editing molecules including Cas9 protein and guide RNA (gRNA), resulting in targeted gene knock-in and knock-out. These results demonstrated the potential of Gelatin-Ph nanofibers obtained through electrospinning and subsequent HRP-mediated cross-linking for gene therapy and tissue regeneration by genome editing.
    Keywords:  electrospinning; gelatin; gene delivery system; horseradish peroxidase; nanofiber
    DOI:  https://doi.org/10.3390/biom12111638
  6. Pharmaceutics. 2022 Oct 26. pii: 2302. [Epub ahead of print]14(11):
    A Comparative Study of Mesoporous Silica and Mesoporous Bioactive Glass Nanoparticles as Non-Viral MicroRNA Vectors for Osteogenesis.
    Sepanta Hosseinpour, Maria Natividad Gomez-Cerezo, Yuxue Cao, Chang Lei, Huan Dai, Laurence J Walsh, Saso Ivanovski, Chun Xu.
      Micro-ribonucleic acid (miRNA)-based therapies show advantages for bone regeneration but need efficient intracellular delivery methods. Inorganic nanoparticles such as mesoporous bioactive glass nanoparticles (MBGN) and mesoporous silica nanoparticles (MSN) have received growing interest in the intracellular delivery of nucleic acids. This study explores the capacity of MBGN and MSN for delivering miRNA to bone marrow mesenchymal stem cells (BMSC) for bone regenerative purposes, with a focus on comparing the two in terms of cell viability, transfection efficiency, and osteogenic actions. Spherical MBGN and MSN with a particle size of ~200 nm and small-sized mesopores were prepared using the sol-gel method, and then the surface was modified with polyethyleneimine for miRNA loading and delivery. The results showed miRNA can be loaded into both nanoparticles within 2 h and was released sustainedly for up to 3 days. Confocal laser scanning microscopy and flow cytometry analysis indicated a high transfection efficiency (>64%) of both nanoparticles without statistical difference. Compared with MSN, MBGN showed stronger activation of alkaline phosphatase and activation of osteocalcin genes. This translated to a greater osteogenic effect of MBGN on BMSC, with Alizarin red staining showing greater mineralization compared with the MSN group. These findings show the potential for MBGN to be used in bone tissue engineering.
    Keywords:  bone regeneration; gene delivery; mesoporous bioactive glass nanoparticle; mesoporous silica nanoparticle; microRNA therapy
    DOI:  https://doi.org/10.3390/pharmaceutics14112302
  7. Molecules. 2022 Nov 01. pii: 7416. [Epub ahead of print]27(21):
    Plasmid-DNA Delivery by Covalently Functionalized PEI-SPIONs as a Potential 'Magnetofection' Agent.
    René Stein, Felix Pfister, Bernhard Friedrich, Pascal-Raphael Blersch, Harald Unterweger, Anton Arkhypov, Andriy Mokhir, Mikhail Kolot, Christoph Alexiou, Rainer Tietze.
      Nanoformulations for delivering nucleotides into cells as vaccinations as well as treatment of various diseases have recently gained great attention. Applying such formulations for a local treatment strategy, e.g., for cancer therapy, is still a challenge, for which improved delivery concepts are needed. Hence, this work focuses on the synthesis of superparamagnetic iron oxide nanoparticles (SPIONs) for a prospective "magnetofection" application. By functionalizing SPIONs with an active catechol ester (CafPFP), polyethyleneimine (PEI) was covalently bound to their surface while preserving the desired nanosized particle properties with a hydrodynamic size of 86 nm. When complexed with plasmid-DNA (pDNA) up to a weight ratio of 2.5% pDNA/Fe, no significant changes in particle properties were observed, while 95% of the added pDNA was strongly bound to the SPION surface. The transfection in A375-M cells for 48 h with low amounts (10 ng) of pDNA, which carried a green fluorescent protein (GFP) sequence, resulted in a transfection efficiency of 3.5%. This value was found to be almost 3× higher compared to Lipofectamine (1.2%) for such low pDNA amounts. The pDNA-SPION system did not show cytotoxic effects on cells for the tested particle concentrations and incubation times. Through the possibility of additional covalent functionalization of the SPION surface as well as the PEI layer, Caf-PEI-SPIONs might be a promising candidate as a magnetofection agent in future.
    Keywords:  cytotoxicity; ligand exchange; magnetofection; pentafluorophenyl ester; plasmid-DNA; superparamagnetic iron oxide nanoparticles (SPIONs); surface functionalization; transfection
    DOI:  https://doi.org/10.3390/molecules27217416
  8. Protein Eng Des Sel. 2022 Nov 07. pii: gzac011. [Epub ahead of print]
    Antibody-mediated delivery of CRISPR-Cas9 ribonucleoproteins in human cells.
    Stephanie Ubiparipovic, Daniel Christ, Romain Rouet.
      The CRISPR genome editing technology holds great clinical potential for the treatment of monogenetic disorders such as sickle cell disease or muscular dystrophy. The therapeutic in vivo application of the technology relies on targeted delivery methods of the Cas9 and gRNA complex to specific cells and/or tissues. However, such methods are currently limited to direct organ delivery, preventing wide clinical application. Here, we show that monoclonal antibodies can be employed to deliver the Cas9/gRNA complex directly into human cells via cell-surface receptors. Using the SpyCatcher/SpyTag system, we conjugated the Fab fragment of the therapeutic antibodies Trastuzumab and Pertuzumab directly to the Cas9 enzyme and observed HER2-specific uptake of the ribonucleoprotein in a human HER2 expressing cell line. Following cellular uptake in the presence of an endosomolytic peptide, modest gene editing was also observed. This finding provides a blueprint for the targeted delivery of the CRISPR technology into specific cells using monoclonal antibodies.
    DOI:  https://doi.org/10.1093/protein/gzac011
  9. Adv Healthc Mater. 2022 Nov 11. e2202460
    Non-lysosomal Route of mRNA delivery and Combining with Epigenetic Regulation Optimized Antitumor Immunoprophylactic Efficacy.
    Yu Liu, Xu Liu, Jiaxin Huang, Yingying Shi, Zhenyu Luo, Junlei Zhang, Xuemeng Guo, Mengshi Jiang, Xiang Li, Hang Yin, Bing Qin, Guannan Guan, Lihua Luo, Yun Zhou, Jian You.
      Currently, mRNA-based tumor therapies are in full flow because in vitro-transcribed (IVT) mRNA has the potential to express tumor antigens to initiate the adaptive immune responses. However, the efficacy of such therapy relies heavily on the delivery system. Here, a Pardaxin-modified liposome loaded with tumor antigen-encoding mRNA and adjuvant (2',3'-cGAMP, (cyclic [G(2',5')pA(3',5')p])), termed P-Lipoplex-CDN is reported. Due to an non-lysosomal delivery route, the transfection efficiency on DCs is improved by reducing the lysosome disruption of cargos. The mRNA modified DCs efficiently induced tumor antigen-specific immune responses both in vitro and in vivo. As prophylactic vaccines, mRNA transfected DCs significantly delay the occurrence and development of tumors, and several immunized mice are even completely resistant to tumors. Interestingly, the efficacy depends on the major histocompatibility complex class I (MHC-I) expression level on tumor cells. Furthermore, epigenetic modification (decitabine, DAC) is applied as a combination strategy to deal with malignant tumor progression caused by deficient tumor MHC-I expression. This study highlighted the close relationship between mRNA-DCs vaccine efficacy and the expression level of tumor cell MHC-I molecules. Moreover, a feasible strategy for tumor MHC-I expression deficiency is proposed, which may provide clinical guidance for the design and application of mRNA-based tumor therapies. This article is protected by copyright. All rights reserved.
    Keywords:  MHC-I-restricted immunity; cancer therapy; cationic liposomes; epigenetic modification; mRNA-DCs
    DOI:  https://doi.org/10.1002/adhm.202202460
  10. Pharmaceutics. 2022 Oct 31. pii: 2357. [Epub ahead of print]14(11):
    Stability Study of mRNA-Lipid Nanoparticles Exposed to Various Conditions Based on the Evaluation between Physicochemical Properties and Their Relation with Protein Expression Ability.
    Mariko Kamiya, Makoto Matsumoto, Kazuma Yamashita, Tatsunori Izumi, Maho Kawaguchi, Shusaku Mizukami, Masako Tsurumaru, Hidefumi Mukai, Shigeru Kawakami.
      Lipid nanoparticles (LNPs) are currently in the spotlight as delivery systems for mRNA therapeutics and have been used in the Pfizer/BioNTech and Moderna COVID-19 vaccines. mRNA-LNP formulations have been indicated to require strict control, including maintenance at fairly low temperatures during their transport and storage. Since it is a new pharmaceutical modality, there is a lack of information on the systematic investigation of how storage and handling conditions affect the physicochemical properties of mRNA-LNPs and their protein expression ability. In this study, using the mRNA-LNPs with standard composition, we evaluated the effects of temperature, cryoprotectants, vibration, light exposure, and syringe aspiration from the vials on the physicochemical properties of nanoparticles in relation to their in vitro/in vivo protein expression ability. Among these factors, storage at -80 °C without a cryoprotectant caused a decrease in protein expression, which may be attributed to particle aggregation. Exposure to vibration and light also caused similar changes under certain conditions. Exposure to these factors can occur during laboratory and hospital handling. It is essential to have sufficient knowledge of the stability of mRNA-LNPs in terms of their physical properties and protein expression ability at an early stage to ensure reproducible research and development and medical care.
    Keywords:  mRNA-LNP; stability; storage condition
    DOI:  https://doi.org/10.3390/pharmaceutics14112357
  11. Nanoscale Adv. 2022 Oct 25. 4(21): 4502-4516
    Silencing of proinflammatory NF-κB and inhibition of herpes simplex virus (HSV) replication by ultrasmall gold nanoparticles (2 nm) conjugated with small-interfering RNA.
    Natalie Wolff, Sebastian Kollenda, Kai Klein, Kateryna Loza, Marc Heggen, Leonie Brochhagen, Oliver Witzke, Adalbert Krawczyk, Ingrid Hilger, Matthias Epple.
      Azide-terminated ultrasmall gold nanoparticles (2 nm gold core) were covalently functionalized with alkyne-terminated small-interfering siRNA duplexes by copper-catalyzed azide-alkyne cycloaddition (CuAAC; click chemistry). The nanoparticle core was visualized by transmission electron microscopy. The number of attached siRNA molecules per nanoparticle was determined by a combination of atomic absorption spectroscopy (AAS; for gold) and UV-Vis spectroscopy (for siRNA). Each nanoparticle carried between 6 and 10 siRNA duplex molecules which corresponds to a weight ratio of siRNA to gold of about 2.2 : 1. Different kinds of siRNA were conjugated to the nanoparticles, depending on the gene to be silenced. In general, the nanoparticles were readily taken up by cells and highly efficient in gene silencing, in contrast to free siRNA. This was demonstrated in HeLa-eGFP cells (silencing of eGFP) and in LPS-stimulated macrophages (silencing of NF-κB). Furthermore, we demonstrated that nanoparticles carrying antiviral siRNA potently inhibited the replication of Herpes simplex virus 2 (HSV-2) in vitro. This highlights the strong potential of siRNA-functionalized ultrasmall gold nanoparticles in a broad spectrum of applications, including gene silencing and treatment of viral infections, combined with a minimal dose of gold.
    DOI:  https://doi.org/10.1039/d2na00250g
  12. J Control Release. 2022 Nov 03. pii: S0168-3659(22)00730-1. [Epub ahead of print]352 556-569
    Nebulization of extracellular vesicles: A promising small RNA delivery approach for lung diseases.
    Yohan Han, Yin Zhu, Hannah A Youngblood, Sultan Almuntashiri, Timothy W Jones, Xiaoyun Wang, Yutao Liu, Payaningal R Somanath, Duo Zhang.
      Small extracellular vesicles (sEVs) are a group of cell-secreted nanovesicles with a diameter up to 200 nm. A growing number of studies have indicated that sEVs can reflect the pathogenesis of human diseases and mediate intercellular communications. Recently, sEV research has drastically increased due to their drug delivery property. However, a comprehensive method of delivering exogenous small RNAs-loaded sEVs through nebulization has not been reported. The methodology is complicated by uncertainty regarding the integrity of sEVs after nebulization, the delivery efficiency of aerosolized sEVs, their deposition in the lungs/cells, etc. This study demonstrates that sEVs can be delivered to murine lungs through a vibrating mesh nebulizer (VMN). In vivo sEV tracking indicated that inhaled sEVs were distributed exclusively in the lung and localized primarily in lung macrophages and airway epithelial cells. Additionally, sEVs loaded with small RNAs were successfully delivered into the lungs. The administration of siMyd88-loaded sEVs through inhalation reduced lipopolysaccharide (LPS)-induced lung injury in mice, supporting an application of this nebulization methodology to deliver functional small RNAs. Collectively, our study proposes a novel method of sEVs-mediated small RNA delivery into the murine lung through nebulization and presents a potential sEV-based therapeutic strategy for human lung diseases.
    Keywords:  Acute lung injury; Exosome; Inflammation; Nebulizer; miRNA; siRNA
    DOI:  https://doi.org/10.1016/j.jconrel.2022.10.052
  13. Int J Biol Macromol. 2022 Nov 03. pii: S0141-8130(22)02513-2. [Epub ahead of print]223(Pt A): 67-76
    Single-helical formyl β-glucan effectively deliver CpG DNA with poly(dA) to macrophages for enhanced vaccine effects.
    Yuying Xu, Minting Liang, Jintao Huang, Yapei Fan, Haiyue Long, Qunjie Chen, Zhe Ren, Chaoxi Wu, Yifei Wang.
      Single helical β-glucan is a one-dimensional host that can form a hybrid helix with DNAs/RNAs as delivery systems. However, unmodified β-glucan has a gelling tendency and a single helical conformation is challenging to obtain. Therefore, in this study, we developed a β-glucan formyl derivative with stable single helical conformation and no gelling tendency. Circular dichroism studies found that the formyl-β-glucan could form a hybrid helix with DNA CpG-poly(dA). The hybrid helix delivery system showed improved activation on antigen-presenting cells, thereby upregulating the mRNA and protein levels of inflammatory factors, and had an immune-enhancing effect on ovalbumin (OVA) immunized mice. These results indicate that formyl-β-glucan can be developed as a non-cationic supramolecular DNA delivery platform with low toxicity and high efficiency.
    Keywords:  CpG; Delivery; Immunomodulatory; helix; β-Glucan
    DOI:  https://doi.org/10.1016/j.ijbiomac.2022.10.258
  14. Biomedicines. 2022 Nov 10. pii: 2885. [Epub ahead of print]10(11):
    An Innovative PTD-IVT-mRNA Delivery Platform for CAR Immunotherapy of ErbB(+) Solid Tumor Neoplastic Cells.
    Sofia K Georgiou-Siafis, Androulla N Miliotou, Charikleia Ntenti, Ioannis S Pappas, Lefkothea C Papadopoulou.
      Chimeric antigen receptor (CAR) immunotherapy includes the genetic modification of immune cells to carry such a receptor and, thus, recognize cancer cell surface antigens. Viral transfection is currently the preferred method, but it carries the risk of off-target mutagenicity. Other transfection platforms have thus been proposed, such the in vitro transcribed (IVT)-mRNAs. In this study, we exploited our innovative, patented delivery platform to produce protein transduction domain (PTD)-IVT-mRNAs for the expression of CAR on NK-92 cells. CAR T1E-engineered NK-92 cells, harboring the sequence of T1E single-chain fragment variant (scFv) to recognize the ErbB receptor, bearing either CD28 or 4-1BB as co-stimulatory signaling domains, were prepared and assessed for their effectiveness in two different ErbB(+) cancer cell lines. Our results showed that the PTD-IVT-mRNA of CAR was safely transduced and expressed into NK-92 cells. CAR T1E-engineered NK-92 cells provoked high levels of cell death (25-33%) as effector cells against both HSC-3 (oral squamous carcinoma) and MCF-7 (breast metastatic adenocarcinoma) human cells in the co-incubation assays. In conclusion, the application of our novel PTD-IVT-mRNA delivery platform to NK-92 cells gave promising results towards future CAR immunotherapy approaches.
    Keywords:  ErbB; HSC-3 cells; IVT-mRNAs; MCF-7 cells; NK-92 cells; PTD-IVT-mRNAs; T1E scFV; chimeric antigen receptor (CAR) immunotherapy; delivery; protein transduction domain (PTD)
    DOI:  https://doi.org/10.3390/biomedicines10112885
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