bims-novged 2021-05-30 papers

on Non-viral vectors for gene delivery

Issue of 2021–05–30
ten papers selected by
the Merkel lab, Ludwig-Maximilians University and Benjamin Winkeljann, Ludwig-Maximilians University

Traceable Nano-Biohybrid Complexes by One-Step Synthesis as CRISPR-Chem Vectors for Neurodegenerative Diseases Synergistic Treatment.
Synthesis and validation of DOPY: a new gemini dioleylbispyridinium based amphiphile for nucleic acid transfection.
Integration of [12]aneN3 and Acenaphtho[1,2-b]quinoxaline as non-viral gene vectors with two-photon property for enhanced DNA/siRNA delivery and bioimaging.
cRGD peptide-conjugated polyethylenimine-based lipid nanoparticle for intracellular delivery of siRNA in hepatocarcinoma therapy.
Effects of lyoprotectants on long-term stability and transfection efficacy of lyophilized poly(lactide-co-glycolide)-graft-polyethylenimine/plasmid DNA polyplexes.
Promising PEGylated cationic dendrimers for delivery of miRNAs as a possible therapy against HIV-1 infection.
Biomineralized calcium carbonate nanohybrids for mild photothermal heating-enhanced gene therapy.
Nanomedicine-based delivery strategies for nucleic acid gene inhibitors in inflammatory diseases.
Targeted delivery system using silica nanoparticles coated with chitosan and AS1411 for combination therapy of doxorubicin and antimiR-21.
mRNA therapy restores euglycemia and prevents liver tumors in murine model of glycogen storage disease.

Adv Mater. 2021 May 28. e2101993

Traceable Nano-Biohybrid Complexes by One-Step Synthesis as CRISPR-Chem Vectors for Neurodegenerative Diseases Synergistic Treatment.

Jie Shen, Zhiguo Lu, Jianze Wang, Qiulian Hao, Weihong Ji, Yanyue Wu, Huan Peng, Ruichen Zhao, Jun Yang, Yan Li, Zhuyan Shi, Xin Zhang.

  Abnormal protein aggregations are essential pathological features of neurodegenerative diseases. Eliminating while inhibiting the regeneration of these protein aggregates is considered an effective treatment strategy. Herein, the CRISPR/Cas9 gene-editing tool is employed to inhibit the regeneration of disease-related proteins, while chemical drugs are applied to eliminate the proteins that are produced. To efficiently deliver CRISPR-chem drugs into brain lesions, traceable nano-biohybrid complexes (F-TBIO) are constructed by one-step synthesis and CRISPR/Cas9 plasmids (CF-TBIO) are loaded in a controllable manner. CF-TBIO can knock out the BACE1 gene and reduce the burden of amyloid-β, and thereby significantly improve the cognitive abilities of 2xTg-AD mice. In particular, by prolonging the dosing interval, the pathological damage and behavioral abilities of 2xTg-AD mice are still significantly improved. During the therapeutic process, CF-TBIO with a high relaxation rate provides accurate imaging signals in the complex brain physiological environment. The finding shows that CF-TBIO has great potential to serve as a CRISPR-chem drug-delivery platform for neurodegenerative diseases therapy.

Keywords:  CRISPR-chem vectors; CRISPR/Cas9; neurodegenerative diseases synergistic treatment; one-step synthesis; traceable nano-biohybrid complexes

DOI:  https://doi.org/10.1002/adma.202101993
Eur J Pharm Biopharm. 2021 May 22. pii: S0939-6411(21)00146-6. [Epub ahead of print]

Synthesis and validation of DOPY: a new gemini dioleylbispyridinium based amphiphile for nucleic acid transfection.

Eva Aubets, Rosa Griera, Alex J Felix, Gemma Rigol, Chiara Sikorski, David Limón, Chiara Mastrorosa, Maria Antònia Busquets, Lluïsa Pérez-García, Véronique Noé, Carlos J Ciudad.

  Nucleic acids therapeutics provide a selective and promising alternative to traditional treatments for multiple genetic diseases. A major obstacle is the development of safe and efficient delivery systems. Here, we report the synthesis of the new cationic gemini amphiphile 1,3-bis[(4-oleyl-1-pyridinio)methyl]benzene dibromide (DOPY). Its transfection efficiency was evaluated using PolyPurine Reverse Hoogsteen hairpins (PPRHs), a nucleic acid tool for gene silencing and gene repair developed in our laboratory. The interaction of DOPY with PPRHs was confirmed by gel retardation assays, and it forms complexes of 155 nm. We also demonstrated the prominent internalization of PPRHs using DOPY compared to other chemical vehicles in SH-SY5Y, PC-3 and DF42 cells. Regarding gene silencing, a specific PPRH against the survivin gene delivered with DOPY decreased survivin protein levels and cell viability more effectively than with N-[1-(2,3-Dioleoyloxy)propyl]-N,N,N-trimethylammonium methylsulfate (DOTAP) in both SH-SY5Y and PC-3 cells. We also validated the applicability of DOPY in gene repair approaches by correcting a point mutation in the endogenous locus of the dhfr gene in DF42 cells using repair-PPRHs. All these results indicate both an efficient entry and release of PPRHs at the intracellular level. Therefore, DOPY can be considered as a new lipid-based vehicle for the delivery of therapeutic oligonucleotides.

Keywords:  Cancer therapy; Cationic liposome; DOPY; Gene delivery; Gene repair; Gene silencing; Lipid-based vector; Nucleic acids; PPRHs; Transfection

DOI:  https://doi.org/10.1016/j.ejpb.2021.05.016
Bioorg Chem. 2021 May 11. pii: S0045-2068(21)00360-6. [Epub ahead of print]113 104983

Integration of [12]aneN3 and Acenaphtho[1,2-b]quinoxaline as non-viral gene vectors with two-photon property for enhanced DNA/siRNA delivery and bioimaging.

Jin-Yu Liu, Xu-Ying Liu, Rui Liu, Fang Tang, Jing-Bo Yang, Quan Tang, Zhong-Lin Lu, Hai-Jun Qiao, Lan He.

  Two-photon fluorescent Acenaphtho[1,2-b]quinoxaline (ANQ) and the hydrophilic di-(triazole-[12]aneN3) moieties were combined through an alkyl chain (ANQ-A-M) or a β-hairpin motif with two aromatic γ-amino acid residues (ANQ-H-M) to explore their capabilities for in vitro and in vivo gene delivery and tracing. ANQ-A-M and ANQ-H-M showed the same maximum absorption at 420 nm, and their fluorescent intensities around 650 nm were varied in different solvents and became poor in the protic solvents. Gel electrophoresis assays indicated that both compounds completely retarded the migration of pDNA at 20 μM in the presence of DOPE. However, the DNA condensation with ANQ-H-M was not reversible, and the particle size of the corresponding complexes were larger indicated from the SEM and DLS measurements. In vitro transfections indicated ANQ-A-M/DOPE achieved Luciferase and GFP expressions were to be 7.9- and 5.7-fold of those by Lipo2000 in A549 cells respectively. However, ANQ-H-M showed very poor transfection efficiency in Luciferase expression. With the help of single/two-photon fluorescence imaging it clearly demonstrated that the successful transfection of ANQ-A-M was attributed to its cellular uptake, apparent lysosomal escape, and reversible release of DNA; and the poor transfection of ANQ-H-M was resulted from the aggregation of the DNA complexes which prevented them from the cellular uptake, and also the strong binding ability which is not easy to release DNA. ANQ-A-M/DOPE also exhibited robust gene silencing (83% knockdown of Luciferase) and GFP expression (2.47-fold higher) efficiency compared with Lipo2000 in A549 and zebrafish, respectively. The work demonstrated that the linkage structure between fluorescent and di(triazole-[12]aneN3) played the important role for their gene delivery performance, and that ANQ-A-M represents a vector with the strong transfection efficiency in vitro and in vivo as well as the efficient real time bioimaging properties, which is potential for the development in biomedical research.

Keywords:  Biological tracking; Gene transfection; Non-viral gene vector; Two photon property; [12]aneN(3)

DOI:  https://doi.org/10.1016/j.bioorg.2021.104983
Drug Deliv. 2021 Dec;28(1): 995-1006

cRGD peptide-conjugated polyethylenimine-based lipid nanoparticle for intracellular delivery of siRNA in hepatocarcinoma therapy.

Shuang Yang, Dandan Wang, Xia Zhang, Yaojun Sun, Bin Zheng.

  The effective delivery system plays an important role in the application of siRNA in the antitumor study. However, until now, researches on the delivery systems targeting hepatocarcinoma cells are still being explored. Here we designed and prepared a novel siRNA delivery system, cRGD-PSH-NP, which was based on a modified polyethyleneimine (PSH) and DSPE-PEG2000-cRGD. cRGD-PSH-NP loaded with survivin siRNA (cRGD-PSH-NP/S) was composed of egg phosphatidylcholine, cationic PSH, PEGylated lipids, survivin siRNA, and cRGD peptide as a targeting ligand. The formulations of cRGD-PSH-NP/S were optimized and characterized. In vitro investigations showed excellent gene silencing and antitumor activity compared with the unmodified nanoparticles in HepG2 cells. In vivo antitumor efficacy of cRGD-PSH-NP/S exhibited potent tumor inhibition (74.71%) in HepG2-bearing nude mice without inducing toxicity. These data suggested further research of cRGD-PSH-NP/S in hepatocarcinoma therapy.

Keywords:  Targeting peptide; antitumor; nanoparticles; polyethyleneimine; siRNA

DOI:  https://doi.org/10.1080/10717544.2021.1928794
Nanomedicine (Lond). 2021 May 28.

Effects of lyoprotectants on long-term stability and transfection efficacy of lyophilized poly(lactide-co-glycolide)-graft-polyethylenimine/plasmid DNA polyplexes.

Joshua Woo, Jeoung Soo Lee.

  Aim: We investigated the effect of lyoprotectants on the long-term stability and transfection efficiency of lyophilized (Lyo.) polyplexes prepared from poly(lactide-co-glycolide)-graft-polyethylenimine (PgP) and plasmid DNA encoding green fluorescent protein (pGFP). Materials & methods: Lyo. PgP/pGFP polyplexes prepared with/without lyoprotectants were stored at -20°C over 6 months. Polyplex stability was analyzed by gel electrophoresis and heparin competition assay. Transfection efficiency and cytotoxicity were evaluated in rat glioma (C6) cells in medium containing 10% serum. Results: Lyo. PgP/pGFP polyplexes prepared with 5% sucrose as a lyoprotectant remained stable up to 6 months and retained transfection efficiency up to 4 months. Conclusion: Lyo. PgP-based polyplexes retain bioactivity during extended storage, potentially enabling transport to remote regions and less stable settings, increasing access to life-changing gene therapy.

Keywords:  cationic amphiphilic polymer; gene delivery; long-term shelf life; lyophilization; lyoprotectants; nanoparticles; polymeric micelle

DOI:  https://doi.org/10.2217/nnm-2021-0065
J Nanobiotechnology. 2021 May 28. 19(1): 158

Promising PEGylated cationic dendrimers for delivery of miRNAs as a possible therapy against HIV-1 infection.

E Royo-Rubio, I Rodríguez-Izquierdo, M Moreno-Domene, T Lozano-Cruz, F J de la Mata, R Gómez, M A Muñoz-Fernández, J L Jiménez.

   BACKGROUND: The appearance of resistance against new treatments and the fact that HIV-1 can infect various cell types and develop reservoirs and sanctuaries makes it necessary to develop new therapeutic approaches to overcome those failures.
RESULTS: Studies of cytotoxicity, genotoxicity, complexes formation, stability, resistance, release and particle size distribution confirmed that G2-SN15-PEG, G3-SN31-PEG, G2-SN15-PEG-FITC and G3-SN31-PEG-FITC dendrimers can form complexes with miRNAs being biocompatible, stable and conferring protection to these nucleic acids. Confocal microscopy and flow cytometry showed effective delivery of these four dendrimers into the target cells, confirming their applicability as delivery systems. Dendriplexes formed with the dendrimers and miRNAs significantly inhibited HIV-1 infection in PBMCs.
CONCLUSIONS: These dendrimers are efficient delivery systems for miRNAs and they specifically and significantly improved the anti-R5-HIV-1 activity of these RNA molecules.

Keywords:  Carbosilane dendrimers; Delivery; HIV-1 infection; Inhibition; microRNAs

DOI:  https://doi.org/10.1186/s12951-021-00899-0
Biomaterials. 2021 May 13. pii: S0142-9612(21)00241-6. [Epub ahead of print]274 120885

Biomineralized calcium carbonate nanohybrids for mild photothermal heating-enhanced gene therapy.

Yanjun Liu, Bingran Yu, Xiaoguang Dai, Nana Zhao, Fu-Jian Xu.

  It is of great significance to develop multifunctional gene carriers to achieve treatments with enhanced therapeutic effects in an inflammation-free manner. In this work, assembled micelles of polysaccharide were utilized for the biomineralization of calcium carbonate to produce one-dimensional Alg-CaCO3 nanoparticles. In order to introduce both functions of mild hyperthermia and gene transfection, polydopamine (PDA) coating was applied to conjugate cationic polymers on the surface of nanoparticles. The resultant ACDP nanohybrids exhibited enhanced performance as gene carriers under near infrared (NIR) light irradiation at a low power density. Meanwhile, the pH-responsive degradation of gene carriers could further promote gene release for better effectiveness. The enhanced gene therapy induces tumor cell apoptosis, which could prevent inflammatory responses. The feasibility of mild hyperthermia-enhanced gene therapy for tumor treatment was investigated in vitro and in vivo. In addition, dual-modal ultrasound (US) and photoacoustic (PA) imaging was also realized to monitor and guide the treatment processes. The current work provides a new avenue for the construction of multifunctional platform to realize cancer therapy with improved therapeutic effectiveness in an inflammation-free manner.

Keywords:  Biomineralization; Calcium carbonate; Gene therapy; Inflammation-free; Mild hyperthermia

DOI:  https://doi.org/10.1016/j.biomaterials.2021.120885
Adv Drug Deliv Rev. 2021 May 22. pii: S0169-409X(21)00183-6. [Epub ahead of print]

Nanomedicine-based delivery strategies for nucleic acid gene inhibitors in inflammatory diseases.

Elias Fattal, François Fay.

  Thanks to their abilities to modulate the expression of virtually any genes, RNA therapeutics have attracted considerable research efforts. Among the strategies focusing on nucleic acid gene inhibitors, antisense oligonucleotides and small interfering RNAs have reached advanced clinical trial phases with several of them having recently been marketed. These successes were obtained by overcoming stability and cellular delivery issues using either chemically modified nucleic acids or nanoparticles. As nucleic acid gene inhibitors are promising strategies to treat inflammatory diseases, this review focuses on the barriers, from manufacturing issues to cellular/subcellular delivery, that still need to be overcome to deliver them to sites of inflammation other than the liver. Furthermore, key examples of applications in rheumatoid arthritis, inflammatory bowel, and lung diseases are presented as case studies of systemic, oral, and lung nucleic acid delivery.

Keywords:  Gene inhibition; Inflammation; Nanoparticle; Nucleic acid

DOI:  https://doi.org/10.1016/j.addr.2021.05.019
Carbohydr Polym. 2021 Aug 15. pii: S0144-8617(21)00498-7. [Epub ahead of print]266 118111

Targeted delivery system using silica nanoparticles coated with chitosan and AS1411 for combination therapy of doxorubicin and antimiR-21.

Fatemeh Khatami, Maryam M Matin, Noor Mohammad Danesh, Ahmad Reza Bahrami, Khalil Abnous, Seyed Mohammad Taghdisi.

  Herein, a novel targeted delivery system was developed for intracellular co-delivery of doxorubicin (DOX) as a chemotherapeutic drug, antimiR-21 as an oncogenic antagomiR. In this system, DOX was loaded into mesoporous silica nanoparticles (MSNs) and chitosan was applied to cover the surface of MSNs. AS1411 aptamer as targeting nucleolin and antimiR-21 were electrostatically attached onto the surface of the chitosan-coated MSNs and formed the final nanocomplex (AACS nanocomplex). The study of drug release was based on DOX release under pH 7.4 and 5.5. Cellular toxicity and cellular uptake assessments of AACS nanocomplex were carried out in nucleolin positive (C26, MCF-7, and 4T1) and nucleolin negative (CHO) cell lines using MTT assay and flow cytometry analysis, respectively. Also, Anti-tumor efficacy of AACS nanocomplex was evaluated in C26 tumor-bearing mice. Overall, the results show that the combination therapy of DOX and antimiR-21, using AACS nanocomplex, could combat the cancer cell growth rate.

Keywords:  AS1411 aptamer; Chitosan; Colorectal cancer; Doxorubicin; Silica nanoparticles; Tumor; antimiR-21

DOI:  https://doi.org/10.1016/j.carbpol.2021.118111
Nat Commun. 2021 May 25. 12(1): 3090

mRNA therapy restores euglycemia and prevents liver tumors in murine model of glycogen storage disease.

Jingsong Cao, Minjung Choi, Eleonora Guadagnin, Maud Soty, Marine Silva, Vincent Verzieux, Edward Weisser, Arianna Markel, Jenny Zhuo, Shi Liang, Ling Yin, Andrea Frassetto, Anne-Renee Graham, Kristine Burke, Tatiana Ketova, Cosmin Mihai, Zach Zalinger, Becca Levy, Gilles Besin, Meredith Wolfrom, Barbara Tran, Christopher Tunkey, Erik Owen, Joe Sarkis, Athanasios Dousis, Vladimir Presnyak, Christopher Pepin, Wei Zheng, Lei Ci, Marjie Hard, Edward Miracco, Lisa Rice, Vi Nguyen, Mike Zimmer, Uma Rajarajacholan, Patrick F Finn, Gilles Mithieux, Fabienne Rajas, Paolo G V Martini, Paloma H Giangrande.

Glycogen Storage Disease 1a (GSD1a) is a rare, inherited metabolic disorder caused by deficiency of glucose 6-phosphatase (G6Pase-α). G6Pase-α is critical for maintaining interprandial euglycemia. GSD1a patients exhibit life-threatening hypoglycemia and long-term liver complications including hepatocellular adenomas (HCAs) and carcinomas (HCCs). There is no treatment for GSD1a and the current standard-of-care for managing hypoglycemia (Glycosade®/modified cornstarch) fails to prevent HCA/HCC risk. Therapeutic modalities such as enzyme replacement therapy and gene therapy are not ideal options for patients due to challenges in drug-delivery, efficacy, and safety. To develop a new treatment for GSD1a capable of addressing both the life-threatening hypoglycemia and HCA/HCC risk, we encapsulated engineered mRNAs encoding human G6Pase-α in lipid nanoparticles. We demonstrate the efficacy and safety of our approach in a preclinical murine model that phenotypically resembles the human condition, thus presenting a potential therapy that could have a significant therapeutic impact on the treatment of GSD1a.

DOI: https://doi.org/10.1038/s41467-021-23318-2