Tailoring Gene Transfer Efficacy through the Arrangement of Cationic and Anionic Blocks in Triblock Copolymer Micelles

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This article references 44 other publications.

1. 1

   Hou, X.; Zaks, T.; Langer, R.; Dong, Y. Lipid nanoparticles for mRNA delivery. Nat. Rev. Mater. 2021, 6 (12), 1078– 1094,  DOI: 10.1038/s41578-021-00358-0

   Google Scholar

   1

   Lipid nanoparticles for mRNA delivery

   Hou, Xucheng; Zaks, Tal; Langer, Robert; Dong, Yizhou

   Nature Reviews Materials (2021), 6 (12), 1078-1094CODEN: NRMADL; ISSN:2058-8437. (Nature Portfolio)

   A review. MRNA (mRNA) has emerged as a new category of therapeutic agent to prevent and treat various diseases. To function in vivo, mRNA requires safe, effective and stable delivery systems that protect the nucleic acid from degrdn. and that allow cellular uptake and mRNA release. Lipid nanoparticles have successfully entered the clinic for the delivery of mRNA; in particular, lipid nanoparticle-mRNA vaccines are now in clin. use against coronavirus disease 2019 (COVID-19), which marks a milestone for mRNA therapeutics. In this Review, we discuss the design of lipid nanoparticles for mRNA delivery and examine physiol. barriers and possible administration routes for lipid nanoparticle-mRNA systems. We then consider key points for the clin. translation of lipid nanoparticle-mRNA formulations, including good manufg. practice, stability, storage and safety, and highlight preclin. and clin. studies of lipid nanoparticle-mRNA therapeutics for infectious diseases, cancer and genetic disorders. Finally, we give an outlook to future possibilities and remaining challenges for this promising technol.
2. 2

   Tenchov, R.; Bird, R.; Curtze, A. E.; Zhou, Q. Lipid Nanoparticles horizontal line From Liposomes to mRNA Vaccine Delivery, a Landscape of Research Diversity and Advancement. ACS Nano 2021, 15 (11), 16982– 17015,  DOI: 10.1021/acsnano.1c04996

   Google Scholar

   2

   Lipid Nanoparticles-From Liposomes to mRNA Vaccine Delivery, a Landscape of Research Diversity and Advancement

   Tenchov, Rumiana; Bird, Robert; Curtze, Allison E.; Zhou, Qiongqiong

   ACS Nano (2021), 15 (11), 16982-17015CODEN: ANCAC3; ISSN:1936-0851. (American Chemical Society)

   A review. Lipid nanoparticles (LNPs) have emerged across the pharmaceutical industry as promising vehicles to deliver a variety of therapeutics. Currently in the spotlight as vital components of the COVID-19 mRNA vaccines, LNPs play a key role in effectively protecting and transporting mRNA to cells. Liposomes, an early version of LNPs, are a versatile nanomedicine delivery platform. A no. of liposomal drugs have been approved and applied to medical practice. Subsequent generations of lipid nanocarriers, such as solid lipid nanoparticles, nanostructured lipid carriers, and cationic lipid-nucleic acid complexes, exhibit more complex architectures and enhanced phys. stabilities. With their ability to encapsulate and deliver therapeutics to specific locations within the body and to release their contents at a desired time, LNPs provide a valuable platform for treatment of a variety of diseases. We present a landscape of LNP-related scientific publications, including patents and journal articles, based on anal. of the CAS Content Collection, the largest human-curated collection of published scientific knowledge. Rising trends are identified, such as nanostructured lipid carriers and solid lipid nanoparticles becoming the preferred platforms for numerous formulations. Recent advancements in LNP formulations as drug delivery platforms, such as antitumor and nucleic acid therapeutics and vaccine delivery systems, are discussed. Challenges and growth opportunities are also evaluated in other areas, such as medical imaging, cosmetics, nutrition, and agrochems. This report is intended to serve as a useful resource for those interested in LNP nanotechnologies, their applications, and the global research effort for their development.
3. 3

   Zu, H.; Gao, D. Non-viral Vectors in Gene Therapy: Recent Development, Challenges, and Prospects. AAPS J. 2021, 23 (4), 78,  DOI: 10.1208/s12248-021-00608-7

   Google Scholar

   3

   Non-viral Vectors in Gene Therapy: Recent Development, Challenges, and Prospects

   Zu Hui; Gao Danchen

   The AAPS journal (2021), 23 (4), 78 ISSN:.

   Gene therapy has been experiencing a breakthrough in recent years, targeting various specific cell groups in numerous therapeutic areas. However, most recent clinical studies maintain the use of traditional viral vector systems, which are challenging to manufacture cost-effectively at a commercial scale. Non-viral vectors have been a fast-paced research topic in gene delivery, such as polymers, lipids, inorganic particles, and combinations of different types. Although non-viral vectors are low in their cytotoxicity, immunogenicity, and mutagenesis, attracting more and more researchers to explore the promising delivery system, they do not carry ideal characteristics and have faced critical challenges, including gene transfer efficiency, specificity, gene expression duration, and safety. This review covers the recent advancement in non-viral vectors research and formulation aspects, the challenges, and future perspectives.
4. 4

   Kumar, R.; Santa Chalarca, C. F.; Bockman, M. R.; Bruggen, C. V.; Grimme, C. J.; Dalal, R. J.; Hanson, M. G.; Hexum, J. K.; Reineke, T. M. Polymeric Delivery of Therapeutic Nucleic Acids. Chem. Rev. 2021, 121 (18), 11527– 11652,  DOI: 10.1021/acs.chemrev.0c00997

   Google Scholar

   4

   Polymeric Delivery of Therapeutic Nucleic Acids

   Kumar, Ramya; Santa Chalarca, Cristiam F.; Bockman, Matthew R.; Bruggen, Craig Van; Grimme, Christian J.; Dalal, Rishad J.; Hanson, Mckenna G.; Hexum, Joseph K.; Reineke, Theresa M.

   Chemical Reviews (Washington, DC, United States) (2021), 121 (18), 11527-11652CODEN: CHREAY; ISSN:0009-2665. (American Chemical Society)

   A review. The advent of genome editing has transformed the therapeutic landscape for several debilitating diseases, and the clin. outlook for gene therapeutics has never been more promising. The therapeutic potential of nucleic acids has been limited by a reliance on engineered viral vectors for delivery. Chem. defined polymers can remediate technol., regulatory, and clin. challenges assocd. with viral modes of gene delivery. Because of their scalability, versatility, and exquisite tunability, polymers are ideal biomaterial platforms for delivering nucleic acid payloads efficiently while minimizing immune response and cellular toxicity. While polymeric gene delivery has progressed significantly in the past four decades, clin. translation of polymeric vehicles faces several formidable challenges. The aim of our Account is to illustrate diverse concepts in designing polymeric vectors towards meeting therapeutic goals of in vivo and ex vivo gene therapy. Here, we highlight several classes of polymers employed in gene delivery and summarize the recent work on understanding the contributions of chem. and architectural design parameters. We touch upon characterization methods used to visualize and understand events transpiring at the interfaces between polymer, nucleic acids, and the physiol. environment. We conclude that interdisciplinary approaches and methodologies motivated by fundamental questions are key to designing high-performing polymeric vehicles for gene therapy.
5. 5

   Rinkenauer, A. C.; Schubert, S.; Traeger, A.; Schubert, U. S. The influence of polymer architecture on in vitro pDNA transfection. J. Mater. Chem. B 2015, 3 (38), 7477– 7493,  DOI: 10.1039/C5TB00782H

   Google Scholar

   5

   The influence of polymer architecture on in vitro pDNA transfection

   Rinkenauer, Alexandra C.; Schubert, Stephanie; Traeger, Anja; Schubert, Ulrich S.

   Journal of Materials Chemistry B: Materials for Biology and Medicine (2015), 3 (38), 7477-7493CODEN: JMCBDV; ISSN:2050-7518. (Royal Society of Chemistry)

   A review. In 2012, the first gene therapy agent was approved by the Europe Medicines Agency leading to increased interest in this research field. Beside viruses, non-viral agents based on lipids or polymers represent aspiring alternatives to deliver the genetic material. Different hurdles have to be overcome depending on the kind of nucleic acid used, where plasmid DNA (pDNA) and small interfering RNA represent the common ones. The main challenge for transfection agents, in particular for pDNA delivery, is the transfer to the cell and into the cell nuclei. Within the group of transfection vesicles, cationic polymers show promising features and variability, as they can be synthesized with tailor-made phys. and chem. properties (architectures and functionalization). In the field of polymer-based gene delivery, the tuning potential of polymers by using different architectures like graft and star-shaped polymers as well as self-assembled block copolymers is immense. In particular, in the last few years numerous new polymer designs showed enhanced transfection properties in combination with good biocompatibility. Furthermore, new insights into the transfection mechanism demonstrated the continuous progress in this field. Polymer architecture influences the polyplex characteristics and the latter has an impact on the transfection mechanism, e.g. the interaction with the cellular membrane depends on the polyplex shape. Moreover, polyplex dissocn. can be easily influenced by the polymer chem., thus biodegradable linkers lead to well suited polymers with reduced toxicity and high delivery potential, and are also promising for in vivo applications. This review focuses on the influence of polymer architectures for pDNA transfection in vitro, showing recent developments and insights. The theor. background concerning the biol. challenges for cationic polymers and the impact of graft- or star-shaped architectures as well as self-assembled structures will be presented in detail.
6. 6

   Ahmed, M.; Narain, R. Progress of RAFT based polymers in gene delivery. Prog. Polym. Sci. 2013, 38 (5), 767– 790,  DOI: 10.1016/j.progpolymsci.2012.09.008

   Google Scholar

   6

   Progress of RAFT based polymers in gene delivery

   Ahmed, Marya; Narain, Ravin

   Progress in Polymer Science (2013), 38 (5), 767-790CODEN: PRPSB8; ISSN:0079-6700. (Elsevier Ltd.)

   The review provides a comprehensive account on the development of gene delivery vectors via reversible addn.-fragmentation chain transfer polymn. (RAFT) approach. Since the development and use of the first non-viral vector for gene delivery applications, a wide range of polymers were synthesized and studied for their gene delivery efficacies. With the advent of living radical polymn. (LRP), well-defined polymers with varying mol. wts., compns. and architectures were synthesized to evaluate their potency as gene delivery vectors. Atom transfer radical polymn. (ATRP) and RAFT are 2 widely used LRP approaches for gene delivery applications. This review focuses primarily on the synthesis and use of cationic polymers via RAFT for DNA and siRNA delivery in vitro and in vivo. RAFT polymn. has allowed the facile synthesis of tailor-made cationic polymers with pendent biocompatible moieties which are promising non-viral gene delivery vectors. Advanced structure-activity relationship studies between the polymers and gene expression were possible due to the remarkable control in the design of these polymers via the RAFT process. As a result, biocompatible and non-toxic polymers are produced under aq. conditions and are identified as potent gene delivery vectors in vitro and in vivo. Polymeric functionalized nanomaterials were produced and studied for gene delivery applications. The consistency of results obtained from different research groups and identification of improved gene expression efficacies of RAFT based gene delivery vectors indicate the scope of this polymn. approach for future studies.
7. 7

   Gonçalves, S. d. Á.; Vieira, R. P. Current status of ATRP-based materials for gene therapy. React. Funct. Polym. 2020, 147, 104453,  DOI: 10.1016/j.reactfunctpolym.2019.104453

   Google Scholar

   7

   Current status of ATRP-based materials for gene therapy

   Goncalves, Sayeny de Avila; Vieira, Ronierik Pioli

   Reactive & Functional Polymers (2020), 147 (), 104453CODEN: RFPOF6; ISSN:1381-5148. (Elsevier Ltd.)

   A review. Gene therapy is an auspicious alternative to treat diseases. However, the design of efficient vectors remains as a challenge due to the innumerous intracellular and extracellular barriers that should be faced during the gene delivery process. Among some types of carries, polymeric gene vectors have gained increasingly attention. Aiming to improve the polymeric vectors' performance, several strategies have been applied such as diversification of the monomers, synthesis routes, polymers architecture, addn. of specific targeting units, shielding domains, and inorg. nanoparticles. Besides, the use of controlled polymn. in the synthesis of these carries have led to improvements, esp. ATRP, a very robust and versatile technique. Therefore, the aims of this review are summarizing the recent advances in gene vectors produced through ATRP; propose a division according to the main gene carries characteristics and strategies used to improve their performance; and also provide a crit. anal. of the current and future perspectives on the use of ATRP in the synthesis of gene vectors.
8. 8

   Yin, H.; Kanasty, R. L.; Eltoukhy, A. A.; Vegas, A. J.; Dorkin, J. R.; Anderson, D. G. Non-viral vectors for gene-based therapy. Nat. Rev. Genet. 2014, 15 (8), 541– 555,  DOI: 10.1038/nrg3763

   Google Scholar

   8

   Non-viral vectors for gene-based therapy

   Yin, Hao; Kanasty, Rosemary L.; Eltoukhy, Ahmed A.; Vegas, Arturo J.; Dorkin, J. Robert; Anderson, Daniel G.

   Nature Reviews Genetics (2014), 15 (8), 541-555CODEN: NRGAAM; ISSN:1471-0056. (Nature Publishing Group)

   Gene-based therapy is the intentional modulation of gene expression in specific cells to treat pathol. conditions. This modulation is accomplished by introducing exogenous nucleic acids such as DNA, mRNA, small interfering RNA (siRNA), microRNA (miRNA) or antisense oligonucleotides. Given the large size and the neg. charge of these macromols., their delivery is typically mediated by carriers or vectors. In this Review, we introduce the biol. barriers to gene delivery in vivo and discuss recent advances in material sciences, nanotechnol. and nucleic acid chem. that have yielded promising non-viral delivery systems, some of which are currently undergoing testing in clin. trials. The diversity of these systems highlights the recent progress of gene-based therapy using non-viral approaches.
9. 9

   Jarak, I.; Pereira-Silva, M.; Santos, A. C.; Veiga, F.; Cabral, H.; Figueiras, A. Multifunctional polymeric micelle-based nucleic acid delivery: Current advances and future perspectives. Appl. Mater. Today 2021, 25, 101217,  DOI: 10.1016/j.apmt.2021.101217

   Google Scholar

   There is no corresponding record for this reference.
10. 10

    Yousefpour Marzbali, M.; Yari Khosroushahi, A. Polymeric micelles as mighty nanocarriers for cancer gene therapy: a review. Cancer Chemother. Pharmacol. 2017, 79 (4), 637– 649,  DOI: 10.1007/s00280-017-3273-1

    Google Scholar

    10

    Polymeric micelles as mighty nanocarriers for cancer gene therapy: a review

    Yousefpour Marzbali, Mahsa; Yari Khosroushahi, Ahmad

    Cancer Chemotherapy and Pharmacology (2017), 79 (4), 637-649CODEN: CCPHDZ; ISSN:0344-5704. (Springer)

    A review. Purpose: Gene therapy has shown extensive potential to treat human diseases occurring from the defection of genes like various types of cancers. The cationic polymers, as non-viral gene carriers, offer the ability to engineer carrier systems having customized features that can be adapted to suit any system. Upon polymeric micelle systems' core-shell structure, micelles can create the capacity to load genes/gene-drugs into the different micelle compartments, resp. Methods: The search will be managed in Pubmed, Medline, Cochrane library, Embase and Proquest for articles related to polymeric micelle-based gene delivery in order to cancer gene therapy using the accommodative search terms. A database of the first search of all search engines results will be made and repeated articles will be removed. After that, the related articles will be selected, and also the refs. of selected articles will be searched in order to find any other articles to complete the search database. Results: This study reviews kinds of polymeric nanomicelles, which have been used in gene therapy, crit. parameters for micelle-based gene delivery, challenges and advantages/disadvantages as well as biosafety of nanomicelles in gene delivery systems. Furthermore, the discussion has focused on stimuli-responsive polymers and strategy and mechanisms regarding tumor-selective gene delivery. Conclusions: This study provides an overview of the advantages/disadvantages of polymeric-based nanocarriers for cancer gene therapy.
11. 11

    Epps, T. H., III; O'Reilly, R. K. Block copolymers: controlling nanostructure to generate functional materials - synthesis, characterization, and engineering. Chem. Sci. 2016, 7 (3), 1674– 1689,  DOI: 10.1039/C5SC03505H

    Google Scholar

    11

    Block copolymers: controlling nanostructure to generate functional materials - synthesis, characterization, and engineering

    Epps, Thomas H., III; O'Reilly, Rachel K.

    Chemical Science (2016), 7 (3), 1674-1689CODEN: CSHCCN; ISSN:2041-6520. (Royal Society of Chemistry)

    A review. In this perspective, we survey recent advances in the synthesis and characterization of block copolymers, discuss several key materials opportunities enabled by block copolymers, and highlight some of the challenges that currently limit further realization of block copolymers in promising nanoscale applications. One significant challenge, esp. as the complexity and functionality of designer macromols. increases, is the requirement of multiple complementary techniques to fully characterize the resultant polymers and nanoscale materials. Thus, we highlight select characterization and theor. methods and discuss how future advances can improve understanding of block copolymer systems. In particular, we consider the application of theor./simulation methods to the rationalization, and prediction, of obsd. exptl. self-assembly phenomena. Finally, we explore several next steps for the field and emphasize some general areas of emerging research that could unlock addnl. opportunities for nanostructure-forming block copolymers in functional materials.
12. 12

    Mai, Y.; Eisenberg, A. Self-assembly of block copolymers. Chem. Soc. Rev. 2012, 41 (18), 5969– 5985,  DOI: 10.1039/c2cs35115c

    Google Scholar

    12

    Self-assembly of block copolymers

    Mai, Yiyong; Eisenberg, Adi

    Chemical Society Reviews (2012), 41 (18), 5969-5985CODEN: CSRVBR; ISSN:0306-0012. (Royal Society of Chemistry)

    A review. Block copolymer (BCP) self-assembly has attracted considerable attention for many decades. The present tutorial review introduces the primary principles of BCP self-assembly in bulk and in soln., by describing expts., theories, accessible morphologies, etc.
13. 13

    Pereira-Silva, M.; Jarak, I.; Santos, A. C.; Veiga, F.; Figueiras, A. Micelleplex-based nucleic acid therapeutics: from targeted stimuli-responsiveness to nanotoxicity and regulation. Eur. J. Pharm. Sci. 2020, 153, 105461,  DOI: 10.1016/j.ejps.2020.105461

    Google Scholar

    13

    Micelleplex-based nucleic acid therapeutics: From targeted stimuli-responsiveness to nanotoxicity and regulation

    Pereira-Silva, Miguel; Jarak, Ivana; Santos, Ana Claudia; Veiga, Francisco; Figueiras, Ana

    European Journal of Pharmaceutical Sciences (2020), 153 (), 105461CODEN: EPSCED; ISSN:0928-0987. (Elsevier B.V.)

    A review. Cell-targeted nucleic acid (NA) therapeutics, either DNA- or RNA-based, have experienced considerable attention regarding their potential applications in gene expression modulation aiming at disease management. However, the therapeutic potential of NAs as selective, safe and multispecific biomols. is principally hindered by their instability in biol. fluids and deficient cellular uptake, urgently calling for intelligent design strategies such as NA loading in effective nanosystems. In this regard, NA delivery nanosystems which bypass biol. hurdles and are capable of safeguarding the NA payload have been extensively explored so far. Micelleplexes consist of tailored and multifunctional micelle-like nanoassemblies of neg.-charged NAs complexed with cationic blocks, generally of polymeric nature, this way ensuring efficient NA protection and transportation, as well as enhanced cellular transfection and boosted intracellular trafficking. Herein, we review the biomedical applications of stable micelleplexes as robust and smart NA delivery nanosystems focusing on the fine-tuning of their properties toward stimuli-responsiveness and the nanosystem's versatility to accommodate distinct ligands for selective tissue-targeting purposes. Addnl., the nanosafety and regulatory considerations of micelleplexes will also be discussed toward the future clin. translation of micelleplexes for efficient NA delivery to cells, paving the way to next-generation micelleplex-based NA therapeutics.
14. 14

    Tan, Z.; Jiang, Y.; Zhang, W.; Karls, L.; Lodge, T. P.; Reineke, T. M. Polycation Architecture and Assembly Direct Successful Gene Delivery: Micelleplexes Outperform Polyplexes via Optimal DNA Packaging. J. Am. Chem. Soc. 2019, 141 (40), 15804– 15817,  DOI: 10.1021/jacs.9b06218

    Google Scholar

    14

    Polycation Architecture and Assembly Direct Successful Gene Delivery: Micelleplexes Outperform Polyplexes via Optimal DNA Packaging

    Tan, Zhe; Jiang, Yaming; Zhang, Wenjia; Karls, Logan; Lodge, Timothy P.; Reineke, Theresa M.

    Journal of the American Chemical Society (2019), 141 (40), 15804-15817CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)

    Cellular delivery of biomacromols. is vital to medical research and therapeutic development. Cationic polymers are promising and affordable candidate vehicles for these precious payloads. However, the impact of polycation architecture and soln. assembly on the biol. mechanisms and efficacy of these vehicles has not been clearly defined. In this study, four polymers contg. the same cationic poly(2-(dimethylamino)ethyl methacrylate) (D) block but placed in different architectures have been synthesized, characterized, and compared for cargo binding and biol. performance. The D homopolymer and its diblock copolymer poly(ethylene glycol)-block-poly(2-(dimethylamino) Et methacrylate) (OD) readily encapsulate pDNA to form polyplexes. Two amphiphilic block polymer variants, poly(2-(dimethylamino)ethyl methacrylate)-block-poly(Bu methacrylate) (DB) and poly(ethylene glycol)-block-poly(2-(dimethylamino)ethyl methacrylate)-block-poly(Bu methacrylate) (ODB), self-assemble into micelles, which template pDNA winding around the cationic corona to form micelleplexes. Micelleplexes were found to have superior delivery efficiency compared to polyplexes and detailed physicochem. and biol. characterizations were performed to pinpoint the mechanisms by testing hypotheses related to cellular internalization, intracellular trafficking, and pDNA unpackaging. For the first time, we find that the higher concn. of amines housed in micelleplexes stimulates both cellular internalization and potential endosomal escape, and the phys. motif of pDNA winding into micelleplexes, reminiscent of DNA compaction by histones in chromatin, preserves the pDNA secondary structure in its native B form. This likely allows greater payload accessibility for protein expression with micelleplexes compared to polyplexes, which tightly condense pDNA and significantly distort its helicity. This work provides important guidance for the design of successful biomol. delivery systems via optimizing the physicochem. properties.
15. 15

    Jiang, Y.; Reineke, T. M.; Lodge, T. P. Complexation of DNA with Cationic Copolymer Micelles: Effects of DNA Length and Topology. Macromolecules 2018, 51 (3), 1150– 1160,  DOI: 10.1021/acs.macromol.7b02201

    Google Scholar

    15

    Complexation of DNA with Cationic Copolymer Micelles: Effects of DNA Length and Topology

    Jiang, Yaming; Reineke, Theresa M.; Lodge, Timothy P.

    Macromolecules (Washington, DC, United States) (2018), 51 (3), 1150-1160CODEN: MAMOBX; ISSN:0024-9297. (American Chemical Society)

    Micelleplexes are promising gene delivery vehicles that form when DNA complexes with polycationic micelles. In this study, the influence of DNA length and topol. on the structure and colloidal stability of micelleplexes was explored using a model system. The cationic micelles were composed of poly(2-(dimethylamino)ethyl methacrylate)-block-poly(Bu methacrylate) and were complexed with linear DNA and circular plasmids of 2442, 4708, 7537 base pairs. The cationic micelles had a mean core radius of 8 ± 1 nm and a mean hydrodynamic radius of 34 ± 1 nm in buffer at pH 5 and 100 mM ionic strength. The formation of micelleplexes was monitored by turbidimetric titrn. as a function of N/P ratio (amine in micelle corona/phosphate on DNA) in acetate buffers of various ionic strengths. The structure and size evolution of micelleplexes were studied by dynamic light scattering and cryo-TEM, while the compn. of micelleplexes were estd. using static light scattering. The combination of these techniques revealed that increasing DNA length resulted in increased micelleplex size at N/P > 1; this was attributed to an increased propensity for longer DNA to bridge between micelles. At N/P < 1, however, longer DNA enhances the stability of micelleplexes against aggregation by providing addnl. steric repulsions between micelleplexes. At high ionic strength, increasing DNA length also shifts titrn. curves to higher N/P ratios as the structure of micelleplexes changes with DNA length. On the other hand, DNA topol. showed minimal influence on the titrn. curves, structure and long-term stability of micelleplexes. Overall, this work illustrates how polycationic micelles may serve as compaction agents for long chain DNA, and how factors such as DNA length can be used to tune the structure and colloidal stability of micelleplexes.
16. 16

    Wang, H.; Ding, S.; Zhang, Z.; Wang, L.; You, Y. Cationic micelle: A promising nanocarrier for gene delivery with high transfection efficiency. J. Gene Med. 2019, 21 (7), e3101  DOI: 10.1002/jgm.3101

    Google Scholar

    There is no corresponding record for this reference.
17. 17

    Lv, H.; Zhang, S.; Wang, B.; Cui, S.; Yan, J. Toxicity of cationic lipids and cationic polymers in gene delivery. J. Controlled Release 2006, 114 (1), 100– 109,  DOI: 10.1016/j.jconrel.2006.04.014

    Google Scholar

    17

    Toxicity of cationic lipids and cationic polymers in gene delivery

    Lv, Hongtao; Zhang, Shubiao; Wang, Bing; Cui, Shaohui; Yan, Jie

    Journal of Controlled Release (2006), 114 (1), 100-109CODEN: JCREEC; ISSN:0168-3659. (Elsevier B.V.)

    A review. Gene therapy, as a promising therapeutics to treat genetic or acquired diseases, has achieved exciting development in the past 2 decades. Appropriate gene vectors can be crucial for gene transfer. Cationic lipids and polymers, the most important non-viral vectors, have many advantages over viral ones as non-immunogenic, easy to produce and not oncogenic. They hold the promise to replace viral vectors to be used in clinic. However, the toxicity is still an obstacle to the application of non-viral vectors to gene therapy. For overcoming the problem, many new cationic compds. were developed. This article provides a review with respect to toxicity of cationic lipids and polymers in gene delivery. The authors evaluate the structural features of cationic compds. and summarize the relationship of toxicity and structure and hope to provide available suggestions on the development of these cationic compds.
18. 18

    Fang, Y.; Xue, J.; Gao, S.; Lu, A.; Yang, D.; Jiang, H.; He, Y.; Shi, K. Cleavable PEGylation: a strategy for overcoming the ″PEG dilemma″ in efficient drug delivery. Drug Delivery 2017, 24 (2), 22– 32,  DOI: 10.1080/10717544.2017.1388451

    Google Scholar

    18

    Cleavable PEGylation: a strategy for overcoming the "PEG dilemma" in efficient drug delivery

    Fang, Yan; Xue, Jianxiu; Gao, Shan; Lu, Anqi; Yang, Dongjuan; Jiang, Hong; He, Yang; Shi, Kai

    Drug Delivery (2017), 24 (2), 22-32CODEN: DDELEB; ISSN:1071-7544. (Taylor & Francis Ltd.)

    To prolong the circulation time of drug, PEGylation has been widely used via the enhanced permeability and retention (EPR) effect, thereby providing new hope for better treatment. However, PEGylation also brings the "PEG dilemma", which is difficult for the cellular absorption of drugs and subsequent endosomal escape. As a result, the activity of drugs is inevitably lost after PEG modification. To achieve successful drug delivery for effective treatment, the crucial issue assocd. with the use of PEG-lipids, i.e., "PEG dilemma" must be addressed. In this paper, we introduced the development and application of nanocarriers with cleavable PEGylation, and discussed various strategies for overcoming the PEG dilemma. Compared to the traditional ones, the vehicle systems with different environmental-sensitive PEG-lipids were discussed, which cleavage can be achieved in response to the intracellular as well as the tumor microenvironment. This smart cleavable PEGylation provides us an efficient strategy to overcome "PEG dilemma", thereby may be a good candidate for the cancer treatment in future.
19. 19

    Kierstead, P. H.; Okochi, H.; Venditto, V. J.; Chuong, T. C.; Kivimae, S.; Frechet, J. M. J.; Szoka, F. C. The effect of polymer backbone chemistry on the induction of the accelerated blood clearance in polymer modified liposomes. J. Controlled Release 2015, 213, 1– 9,  DOI: 10.1016/j.jconrel.2015.06.023

    Google Scholar

    19

    The effect of polymer backbone chemistry on the induction of the accelerated blood clearance in polymer modified liposomes

    Kierstead, Paul H.; Okochi, Hideaki; Venditto, Vincent J.; Chuong, Tracy C.; Kivimae, Saul; Frechet, Jean M. J.; Szoka, Francis C.

    Journal of Controlled Release (2015), 213 (), 1-9CODEN: JCREEC; ISSN:0168-3659. (Elsevier B.V.)

    A variety of water-sol. polymers, when attached to a liposome, substantially increase liposome circulation half-life in animals. However, in certain conditions, liposomes modified with the most widely used polymer, polyethylene glycol (PEG), induce an IgM response resulting in an accelerated blood clearance (ABC) of the liposome upon the second injection. Modification of liposomes with other water-sol. polymers: HPMA (poly[N-(2-hydroxypropyl) methacrylamide]), PVP (poly(vinylpyrrolidone)), PMOX (poly(2-methyl-2-oxazoline)), PDMA (poly(N,N-di-Me acrylamide)), and PAcM (poly(N-acryloyl morpholine)), increases circulation times of liposomes; but a precise comparison of their ability to promote long circulation or induce the ABC effect has not been reported. To obtain a more nuanced understanding of the role of polymer structure/MW to promote long circulation, we synthesized a library of polymer diacyl chain lipids with low polydispersity (1.04-1.09), similar polymer mol. wts. (2.1-2.5 kDa) and incorporated them into 100 nm liposomes of a narrow polydispersity (0.25-1.3) composed of polymer-lipid/hydrogenated soy phosphatidylcholine/cholesterol/diD: 5.0/54.5/40/0.5. We confirm that HPMA, PVP, PMOX, PDMA and PAcM modified liposome have increased circulation times in rodents and that PVP, PDMA, and PAcM do not induce the ABC effect. We demonstrate for the first time, that HPMA does not cause an ABC effect whereas PMOX induces a pronounced ABC effect in rats. We find that a single dose of liposomes coated with PEG and PMOX generates an IgM response in rats towards the resp. polymer. Finally, in this homologous polymer series, we observe a pos. correlation (R = 0.84 in rats, R = 0.92 in mice) between the circulation time of polymer-modified liposomes and polymer viscosity; PEG and PMOX, the polymers that can initiate an ABC response were the two most viscous polymers. Our findings suggest that polymers that do not cause an ABC effect such as, HPMA or PVP, deserve further consideration as polymer coatings to improve the circulation of liposomes and other nanoparticles.
20. 20

    Bludau, H.; Czapar, A. E.; Pitek, A. S.; Shukla, S.; Jordan, R.; Steinmetz, N. F. POxylation as an alternative stealth coating for biomedical applications. Eur. Polym. J. 2017, 88, 679– 688,  DOI: 10.1016/j.eurpolymj.2016.10.041

    Google Scholar

    20

    Polyoxazolination as an alternative stealth coating for biomedical applications

    Bludau, Herdis; Czapar, Anna E.; Pitek, Andrzej S.; Shukla, Sourabh; Jordan, Rainer; Steinmetz, Nicole F.

    European Polymer Journal (2017), 88 (), 679-688CODEN: EUPJAG; ISSN:0014-3057. (Elsevier Ltd.)

    Polyethylene glycol (PEG) polymers are currently used in a variety of medical formulations to reduce toxicity, minimize immune interactions and improve pharmacokinetics. Despite its widespread use however, the presence of anti-PEG antibodies indicates that this polymer has the potential to be immunogenic and antigenic. Here we present an alternative polymer, poly(2-oxazoline) (POx) for stealth applications, specifically shielding of a proteinaceous nanoparticle from recognition by the immune system. Tobacco mosaic virus (TMV) was used as our testbed due to its potential for use as a nanocarrier for drug delivery and mol. imaging applications.
21. 21

    Hu, Y.; Hou, Y.; Wang, H.; Lu, H. Polysarcosine as an Alternative to PEG for Therapeutic Protein Conjugation. Bioconjugate Chem. 2018, 29 (7), 2232– 2238,  DOI: 10.1021/acs.bioconjchem.8b00237

    Google Scholar

    21

    Polysarcosine as an Alternative to PEG for Therapeutic Protein Conjugation

    Hu, Yali; Hou, Yingqin; Wang, Hao; Lu, Hua

    Bioconjugate Chemistry (2018), 29 (7), 2232-2238CODEN: BCCHES; ISSN:1043-1802. (American Chemical Society)

    The performance of many therapeutic proteins, including human interferon-α2b (IFN), is often impeded by their intrinsic instability to protease, poor pharmacokinetics, and strong immunity. Although PEGylation has been an effective approach to improve the pharmacokinetics of many proteins, a few noticeable limitations have aroused vast research efforts in seeking alternatives to PEG for bioconjugation. Herein, we report our investigation on the use of polysarcosine (PSar), a nonionic and hydrophilic polypeptoid, for IFN modification. The site-specific conjugate PSar-IFN, generated by native chem. ligation in high yield, is systematically compared with a similarly produced PEG-interferon conjugate (PEG-IFN) to evaluate the in vitro and in vivo behaviors. PSar is found to show comparable ability in stabilizing IFN from protease digestion in vitro and prolonging the circulation half-life in vivo. Interestingly, PSar-IFN retains more activity in vitro and accumulates more in the tumor sites upon systemic administration than PEG-IFN. Most importantly, PSar-IFN is significantly more potent in inhibiting tumor growth and elicits considerably less anti-IFN antibodies in mouse than PEG-IFN. Together, our results demonstrate for the first time that PSar is an outstanding candidate for therapeutic protein conjugation. Considering the low toxicity, biodegradability, and excellent stealth effect of PSar, this study suggests that such polypeptoids hold enormous potential for many biomedical applications including protein delivery, colloidal stabilization, and nanomedicine.
22. 22

    Hu, C. M.; Fang, R. H.; Luk, B. T.; Zhang, L. Polymeric nanotherapeutics: clinical development and advances in stealth functionalization strategies. Nanoscale 2014, 6 (1), 65– 75,  DOI: 10.1039/C3NR05444F

    Google Scholar

    22

    Polymeric nanotherapeutics: clinical development and advances in stealth functionalization strategies

    Hu, Che-Ming J.; Fang, Ronnie H.; Luk, Brian T.; Zhang, Liangfang

    Nanoscale (2014), 6 (1), 65-75CODEN: NANOHL; ISSN:2040-3372. (Royal Society of Chemistry)

    A review. Long-circulating polymeric nanotherapeutics have garnered increasing interest in research and in the clinic owing to their ability to improve the soly. and pharmacokinetics of therapeutic cargoes. Modulation of carrier properties promises more effective drug localization at the disease sites and can lead to enhanced drug safety and efficacy. In the present review, we highlight the current development of polymeric nanotherapeutics in the clinic. In light of the importance of stealth properties in therapeutic nanoparticles, we also review the advances in stealth functionalization strategies and examine the performance of different stealth polymers in the literature. In addn., we discuss the recent development of biol. inspired "self" nanoparticles, which present a differing stealth concept from conventional approaches.
23. 23

    Zhang, Y.; Satterlee, A.; Huang, L. In vivo gene delivery by nonviral vectors: overcoming hurdles?. Mol. Ther. 2012, 20 (7), 1298– 1304,  DOI: 10.1038/mt.2012.79

    Google Scholar

    23

    In Vivo Gene Delivery by Nonviral Vectors: Overcoming Hurdles?

    Zhang, Yuan; Satterlee, Andrew; Huang, Leaf

    Molecular Therapy (2012), 20 (7), 1298-1304CODEN: MTOHCK; ISSN:1525-0016. (Nature Publishing Group)

    A review. The promise of cancer gene therapeutics is hampered by difficulties in the in vivo delivery to the targeted tumor cells, and systemic delivery remains to be the biggest challenge to be overcome. Here, the authors conc. on systemic in vivo gene delivery for cancer therapy using nonviral vectors. In this review, the authors summarize the existing delivery barriers together with the requirements and strategies to overcome these problems. The authors will also introduce the current progress in the design of nonviral vectors, and briefly discuss their safety issues.
24. 24

    Hattori, Y. Progress in the development of lipoplex and polyplex modified with anionic polymer for efficient gene delivery. J. Genet. Med. Gene Ther. 2017, 1 (1), 3– 18,  DOI: 10.29328/journal.jgmgt.1001002

    Google Scholar

    There is no corresponding record for this reference.
25. 25

    Bernkop-Schnurch, A. Strategies to overcome the polycation dilemma in drug delivery. Adv. Drug Delivery Rev. 2018, 136–137, 62– 72,  DOI: 10.1016/j.addr.2018.07.017

    Google Scholar

    25

    Strategies to overcome the polycation dilemma in drug delivery

    Bernkop-Schnurch Andreas

    Advanced drug delivery reviews (2018), 136-137 (), 62-72 ISSN:.

    Because of polycationic auxiliary agents such as chitosan, polyethyleneimine and cell penetrating peptides as well as cationic lipids assembling to polycationic systems, drug carriers can tightly interact with cell membranes exhibiting a high-density anionic charge. Because of these interactions the cell membrane is depolarized and becomes vulnerable to various uptake mechanisms. On their way to the target site, however, the polycationic character of all these drug carriers is eliminated by polyanionic macromolecules such as mucus glycoproteins, serum proteins, proteoglycans of the extracellular matrix (ECM) and polyanionic surface substructures of non-target cells such as red blood cells. Strategies to overcome this polycation dilemma are focusing on a pH-, redox- or enzyme-triggered charge conversion at the target site. The pH-triggered systems are making use of a slight acidic environment at the target site such as in case of solid tumors, inflammatory tissue and ischemic tissue. Due to a pH shift from 7.2 to slightly acidic mainly amino substructures of polymeric excipients are protonated or shielding groups such as 2,3 dimethylmaleic acid are cleaved off unleashing the underlying cationic character. Redox-triggered systems are utilizing disulfide linkages to bulky side chains such as PEGs masking the polycationic character. Under mild reducing conditions such as in the tumor microenvironment these disulfide bonds are cleaved. Enzyme-triggered systems are targeting enzymes such as alkaline phosphatase, matrix metalloproteinases or hyaluronidase in order to eliminate anionic moieties via enzymatic cleavage resulting in a charge conversion from negative to positive. Within this review an overview about the pros and cons of these systems is provided.
26. 26

    Richter, F.; Leer, K.; Martin, L.; Mapfumo, P.; Solomun, J. I.; Kuchenbrod, M. T.; Hoeppener, S.; Brendel, J. C.; Traeger, A. The impact of anionic polymers on gene delivery: how composition and assembly help evading the toxicity-efficiency dilemma. J. Nanobiotechnol. 2021, 19 (1), 292,  DOI: 10.1186/s12951-021-00994-2

    Google Scholar

    26

    The impact of anionic polymers on gene delivery: how composition and assembly help evading the toxicity-efficiency dilemma

    Richter, Friederike; Leer, Katharina; Martin, Liam; Mapfumo, Prosper; Solomun, Jana I.; Kuchenbrod, Maren T.; Hoeppener, Stephanie; Brendel, Johannes C.; Traeger, Anja

    Journal of Nanobiotechnology (2021), 19 (1), 292CODEN: JNOAAO; ISSN:1477-3155. (BioMed Central Ltd.)

    Cationic polymers have been widely studied for non-viral gene delivery due to their ability to bind genetic material and to interact with cellular membranes. However, their charged nature carries the risk of increased cytotoxicity and interaction with serum proteins, limiting their potential in vivo application. Therefore, hydrophilic or anionic shielding polymers are applied to counteract these effects. Herein, a series of micelle-forming and micelle-shielding polymers were synthesized via RAFT polymn. The copolymer poly[(Bu acrylate)-b-(2-(di-Me amino)ethyl acrylamide)] (P(nBA-b-DMAEAm)) was assembled into cationic micelles and different shielding polymers were applied, i.e., poly(acrylic acid) (PAA), poly(4-acryloyl morpholine) (PNAM) or P(NAM-b-AA) block copolymer. These systems were compared to a triblock terpolymer micelle comprising PAA as the middle block. The assemblies were investigated regarding their morphol., interaction with pDNA, cytotoxicity, transfection efficiency, polyplex uptake and endosomal escape. The naked cationic micelle exhibited superior transfection efficiency, but increased cytotoxicity. The addn. of shielding polymers led to reduced toxicity. In particular, the triblock terpolymer micelle convinced with high cell viability and no significant loss in efficiency. The highest shielding effect was achieved by layering micelles with P(NAM-b-AA) supporting the colloidal stability at neutral zeta potential and completely restoring cell viability while maintaining moderate transfection efficiencies. The high potential of this micelle-layer-combination for gene delivery was illustrated for the first time.
27. 27

    Dutta, K.; Das, R.; Medeiros, J.; Kanjilal, P.; Thayumanavan, S. Charge-Conversion Strategies for Nucleic Acid Delivery. Adv. Funct. Mater. 2021, 31 (24), 2011103,  DOI: 10.1002/adfm.202011103

    Google Scholar

    27

    Charge-Conversion Strategies for Nucleic Acid Delivery

    Dutta, Kingshuk; Das, Ritam; Medeiros, Jewel; Kanjilal, Pintu; Thayumanavan, S.

    Advanced Functional Materials (2021), 31 (24), 2011103CODEN: AFMDC6; ISSN:1616-301X. (Wiley-VCH Verlag GmbH & Co. KGaA)

    A review. Nucleic acids are considered as one of the most potent therapeutic modalities, as their roles go beyond storing genetic information and chem. energy or as signal transducers. Attenuation or expression of desired genes through nucleic acids have profound implications in gene therapy, gene editing, and even in vaccine development. Although nucleic acid therapeutics bring in overwhelming possibilities toward the development of mol. medicines, there are significant loopholes in their effective clin. translation. One of the major pitfalls lies in the traditional design concepts of nucleic acid drug carriers, namely, cationic charge induced cytotoxicity. Targeting this bottleneck, several innovative carrier designs have been proposed accommodating charge-conversion approaches, whereby built-in functionalities convert from cationic to neutral or anionic, or even from anionic to cationic enabling the carrier to overcome several crit. barriers for therapeutics delivery, such as serum deactivation, instability in circulation, low transfection, and poor endosomal escape. This review will critically analyze various mol. designs of charge-converting nanocarriers in a classified approach for the successful delivery of nucleic acids. Accompanied by the narrative on recent clin. nucleic acid candidates, the review concludes with a discussion on the pitfalls and scope of these emerging approaches.
28. 28

    Hu, Y.; Gong, X.; Zhang, J.; Chen, F.; Fu, C.; Li, P.; Zou, L.; Zhao, G. Polymers 2016, 8 (4), 99,  DOI: 10.3390/polym8040099

    Google Scholar

    28

    Activated charge-reversal polymeric nano-system: the promising strategy in drug delivery for cancer therapy

    Hu, Yichen; Gong, Xiao; Zhang, Jinming; Chen, Fengqian; Fu, Chaomei; Li, Peng; Zou, Liang; Zhao, Gang

    Polymers (Basel, Switzerland) (2016), 8 (4), 99/1-99/21CODEN: POLYCK; ISSN:2073-4360. (MDPI AG)

    Various polymeric nanoparticles (NPs) with optimal size, tumor-targeting functionalization, or microenvironment sensitive characteristics have been designed to solve several limitations of conventional chemotherapy. Nano-sized polymeric drug carrier systems have remarkably great advantages in drug delivery and cancer therapy, which are still plagued with severe deficiencies, esp. insufficient cellular uptake. Recently, surface charge of medical NPs has been demonstrated to play an important role in cellular uptake. NPs with pos. charge show higher affinity to anionic cell membranes such that with more efficient cellular internalization, but otherwise cause severe aggregation and fast clearance in circulation. Thus, surface charge-reversal NPs, specifically activated at the tumor site, have shown to elegantly resolve the enhanced cellular uptake in cancer cells vs. non-specific protein adsorption dilemma. Herein, this review mainly focuses on the effect of tumor-site activated surface charge reversal NPs on tumor treatment, including the activated mechanisms and various applications in suppressing cancer cells, killing cancer stem cell and overcoming multidrug resistance, with the emphasis on recent research in these fields. With the comprehensive and in-depth understanding of the activated surface charge reversal NPs, this approach might arouse great interest of scientific research on enhanced efficient polymeric nano-carriers in cancer therapy.
29. 29

    Sun, Y.; Liu, H.; Yang, T.; Lang, L.; Cheng, L.; Xing, H.; Yang, L.; Ding, P. Amphoteric poly(amido amine)s with adjustable balance between transfection efficiency and cytotoxicity for gene delivery. Colloids Surf., B 2019, 175, 10– 17,  DOI: 10.1016/j.colsurfb.2018.11.045

    Google Scholar

    29

    Amphoteric poly(amido amine)s with adjustable balance between transfection efficiency and cytotoxicity for gene delivery

    Sun, Yanping; Liu, Hui; Yang, Tianzhi; Lang, Lang; Cheng, Lin; Xing, Haonan; Yang, Li; Ding, Pingtian

    Colloids and Surfaces, B: Biointerfaces (2019), 175 (), 10-17CODEN: CSBBEQ; ISSN:0927-7765. (Elsevier B.V.)

    In order to balance transfection efficiency and cytotoxicity as well as screen the optimal polymers for gene delivery, a series of amphoteric copolymers (poly(CBA-AGM/GABA)s) composed of different ratios between agmatine (AGM) and γ-aminobutyric acid (GABA) monomers were synthesized. The AGM contg. pos. charged guanidinium groups was used to improve transfection efficiency, while the GABA contg. neg. charged carboxyl groups was used to decrease cytotoxicity. It is hypothesized that the amphoteric poly(CBA-AGM/GABA)s synthesized at the optimal ratio of both components would well balance transfection efficiency and cytotoxicity. By comparing these polymers' essential features in gene delivery, the ideal ratio between AGM and GABA was optimized. AGM80, which contained 80% AGM and 20% GABA, showed favorable properties for gene delivery, including moderate DNA condensation capacity, high cellular uptake, strong nuclear localization ability, high transfection efficiency, and low cytotoxicity, indicating that this polymer is very promising as a potent and nontoxic gene carrier.
30. 30

    Ma, J.; Zhang, J.; Chi, L.; Liu, C.; Li, Y.; Tian, H. Preparation of poly (glutamic acid) shielding micelles self-assembled from polylysine-b-polyphenylalanine for gene and drug codelivery. Chin. Chem. Lett. 2020, 31 (6), 1427– 1431,  DOI: 10.1016/j.cclet.2020.02.034

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    30

    Preparation of poly(glutamic acid) shielding micelles self-assembled from polylysine-b-polyphenylalanine for gene and drug codelivery

    Ma, Jing; Zhang, Jingpeng; Chi, Lin; Liu, Chong; Li, Yanhui; Tian, Huayu

    Chinese Chemical Letters (2020), 31 (6), 1427-1431CODEN: CCLEE7; ISSN:1001-8417. (Elsevier B.V.)

    A novel amphiphilic cationic block copolymer polylysine-b-polyphenylalanine (PLL-b-PPhe) was synthesized and self-assembled into micelles in aq. soln., then shielded with poly(glutamic acid) (marked as PG/PLL-b-PPhe) to codeliver gene and drug for combination cancer therapy. Here, doxorubicin (DOX) was selected to be loaded into PLL-b-PPhe micelles and the drug loading efficiency was 8.0%. The drug release studies revealed that the PLL-b-PPhe micelles were pH sensitive and the released DOX could reach to 53.0%, 65.0%, 72.0% at pH 7.4, 6.8 and 5.0, resp. In order to reduce pos. charge and cytotoxicity of PLL-b-PPhe micelles, PG was used as shelding, simultaneously condensed with Bcl2 siRNA to form gene carrier system. Compared with PEI, PG/PLL-b-PPhe had excellent gene transfection efficiency, esp. when the molar ratio of PLL to PPhe was 30:60 and the mixed mass ratio of PLL-b-PPhe to gene was 5:1. More importantly, DOX and Bcl2 siRNA gene codelivery system displayed remarkable cytotoxicity against B16F10 cells. Confocal laser scanning microscopy (CLSM) and flow cytometry were used to characterize endocytosis of the codelivery system, and confirmed that both DOX and Bcl2 siRNA had been endocytosed into B16F10 cells. The above results indicated that gene and drug codelivery was a promising strategy in future cancer therapy.
31. 31

    Solomun, J. I.; Martin, L.; Mapfumo, P.; Moek, E.; Amro, E.; Becker, F.; Tuempel, S.; Hoeppener, S.; Rudolph, K. L.; Traeger, A. pH-sensitive packaging of cationic particles by an anionic block copolymer shell. J. Nanobiotechnol. 2022, 20 (1), 336,  DOI: 10.1186/s12951-022-01528-0

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    31

    pH-sensitive packaging of cationic particles by an anionic block copolymer shell

    Solomun, Jana I.; Martin, Liam; Mapfumo, Prosper; Moek, Elisabeth; Amro, Elias; Becker, Friedrich; Tuempel, Stefan; Hoeppener, Stephanie; Rudolph, K. Lenhard; Traeger, Anja

    Journal of Nanobiotechnology (2022), 20 (1), 336CODEN: JNOAAO; ISSN:1477-3155. (BioMed Central Ltd.)

    Cationic non-viral vectors show great potential to introduce genetic material into cells, due to their ability to transport large amts. of genetic material and their high synthetic versatility. However, designing materials that are effective without showing toxic effects or undergoing non-specific interactions when applied systemically remains a challenge. The introduction of shielding polymers such as polyethylene glycol (PEG) can enhance biocompatibility and circulation time, however, often impairs transfection efficiency. Herein, a multicomponent polymer system is introduced, based on cationic and hydrophobic particles (P(nBMA46-co-MMA47-co-DMAEMA90), (PBMD)) with high delivery performance and a pH-responsive block copolymer (poly((N-acryloylmorpholine)-b-(2-(carboxy)ethyl acrylamide)) (P(NAM72-b-CEAm74), PNC)) as shielding system, with PNAM as alternative to PEG. The pH-sensitive polymer design promotes biocompatibility and excellent stability at extracellular conditions (pH 7.4) and also allows endosomal escape and thus high transfection efficiency under acidic conditions. PNC shielded particles are below 200 nm in diam. and showed stable pDNA complexation. Further, interaction with human erythrocytes at extracellular conditions (pH 7.4) was prevented, while acidic conditions (pH 6) enabled membrane leakage. The particles demonstrate transfection in adherent (HEK293T) as well as difficult-to-transfect suspension cells (K-562), with comparable or superior efficiency compared to com. linear poly(ethylenimine) (LPEI). Besides, the toxicity of PNC-shielded particles was significantly minimized, in particular in K-562 cells and erythrocytes. In addn., a pilot in vivo expt. on bone marrow blood cells of mice that were injected with PNC-shielded particles, revealed slightly enhanced cell transfection in comparison to naked pDNA. This study demonstrates the applicability of cationic hydrophobic polymers for transfection of adherent and suspension cells in culture as well as in vivo by co-formulation with pH-responsive shielding polymers, without substantially compromising transfection performance.
32. 32

    Liang, K.; Bae, K. H.; Lee, F.; Xu, K.; Chung, J. E.; Gao, S. J.; Kurisawa, M. Self-assembled ternary complexes stabilized with hyaluronic acid-green tea catechin conjugates for targeted gene delivery. J. Controlled Release 2016, 226, 205– 216,  DOI: 10.1016/j.jconrel.2016.02.004

    Google Scholar

    32

    Self-assembled ternary complexes stabilized with hyaluronic acid-green tea catechin conjugates for targeted gene delivery

    Liang, Kun; Bae, Ki Hyun; Lee, Fan; Xu, Keming; Chung, Joo Eun; Gao, Shu Jun; Kurisawa, Motoichi

    Journal of Controlled Release (2016), 226 (), 205-216CODEN: JCREEC; ISSN:0168-3659. (Elsevier B.V.)

    Nanosized polyelectrolyte complexes are attractive delivery vehicles for the transfer of therapeutic genes to diseased cells. Here we report the application of self-assembled ternary complexes constructed with plasmid DNA, branched polyethylenimine and hyaluronic acid-green tea catechin conjugates for targeted gene delivery. These conjugates not only stabilize plasmid DNA/polyethylenimine complexes via the strong DNA-binding affinity of green tea catechin, but also facilitate their transport into CD44-overexpressing cells via receptor-mediated endocytosis. The hydrodynamic size, surface charge and phys. stability of the complexes are characterized. We demonstrate that the stabilized ternary complexes display enhanced resistance to nuclease attack and polyanion-induced dissocn. Moreover, the ternary complexes can efficiently transfect the difficult-to-transfect HCT-116 colon cancer cell line even in serum-supplemented media due to their enhanced stability and CD44-targeting ability. Confocal microscopic anal. demonstrates that the stabilized ternary complexes are able to promote the nuclear transport of plasmid DNA more effectively than binary complexes and hyaluronic acid-coated ternary complexes. The present study suggests that the ternary complexes stabilized with hyaluronic acid-green tea catechin conjugates can be widely utilized for CD44-targeted delivery of nucleic acid-based therapeutics.
33. 33

    Ito, T.; Iida-Tanaka, N.; Koyama, Y. Efficient in vivo gene transfection by stable DNA/PEI complexes coated by hyaluronic acid. J. Drug Target. 2008, 16 (4), 276– 81,  DOI: 10.1080/10611860801900728

    Google Scholar

    33

    Efficient in vivo gene transfection by stable DNA/PEI complexes coated by hyaluronic acid

    Ito, Tomoko; Iida-Tanaka, Naoko; Koyama, Yoshiyuki

    Journal of Drug Targeting (2008), 16 (4), 276-281CODEN: JDTAEH; ISSN:1061-186X. (Informa Healthcare)

    Plasmid DNA was mixed with polyethyleneimine (PEI) and hyaluronic acid (HA) to afford ternary complexes with neg. surface charge regardless of the mixing order. They showed reduced non-specific interactions with blood components. When DNA and PEI were mixed at a high concn. such as that used in in vivo expts., they soon aggregated, and large particles were formed. On the other hand, pre-addn. of HA to DNA prior to PEI effectively diminished the aggregation, and 10% (in vol.) of the complexes remained as small particles with a diam. below 80 nm. Those neg. charged small ternary complexes induced a much stronger extra-gene expression in tumor than binary DNA/PEI complex after intratumoral or i.v. injection into the mice bearing B16 cells.
34. 34

    Rinkenauer, A. C.; Schallon, A.; Günther, U.; Wagner, M.; Betthausen, E.; Schubert, U. S.; Schacher, F. H. A Paradigm Change: Efficient Transfection of Human Leukemia Cells by Stimuli-Responsive Multicompartment Micelles. ACS Nano 2013, 7 (11), 9621– 9631,  DOI: 10.1021/nn402072d

    Google Scholar

    34

    A Paradigm Change: Efficient Transfection of Human Leukemia Cells by Stimuli-Responsive Multicompartment Micelles

    Rinkenauer, Alexandra C.; Schallon, Anja; Guenther, Ulrike; Wagner, Michael; Betthausen, Eva; Schubert, Ulrich S.; Schacher, Felix H.

    ACS Nano (2013), 7 (11), 9621-9631CODEN: ANCAC3; ISSN:1936-0851. (American Chemical Society)

    The controlled nonviral delivery of genetic material using cationic polymers into cells has been of interest during the past three decades, yet the ideal delivery agent featuring utmost transfection efficiency and low cytotoxicity still has to be developed. Here, we demonstrate that multicompartment micelles from stimuli-responsive triblock terpolymers, polybutadiene-block-poly(methacrylic acid)-block-poly(2-(dimethylamino)ethyl methacrylate) (BMAAD), are promising candidates. The structures exhibit a patchy shell, consisting of amphiphilic (interpolyelectrolyte complexes, MAA and D) and cationic patches (excess D), generating a surface reminiscent to those of certain viruses and capable of undergoing pH-dependent changes in charge stoichiometry. After polyplex formation with plasmid DNA, superior transfection efficiencies can be reached for both adherent cells and human leukemia cells. Compared to the gold std. PEI, remarkable improvements and a no. of advantages were identified for this system, including increased cellular uptake and an improved release of the genetic material, accompanied by fast and efficient endosomal escape. Furthermore, high sedimentation rates might be beneficial regarding in vitro applications.
35. 35

    Convertine, A. J.; Benoit, D. S.; Duvall, C. L.; Hoffman, A. S.; Stayton, P. S. Development of a novel endosomolytic diblock copolymer for siRNA delivery. J. Controlled Release 2009, 133 (3), 221– 9,  DOI: 10.1016/j.jconrel.2008.10.004

    Google Scholar

    35

    Development of a novel endosomolytic diblock copolymer for siRNA delivery

    Convertine, Anthony J.; Benoit, Danielle S. W.; Duvall, Craig L.; Hoffman, Allan S.; Stayton, Patrick S.

    Journal of Controlled Release (2009), 133 (3), 221-229CODEN: JCREEC; ISSN:0168-3659. (Elsevier B.V.)

    The gene knockdown activity of small interfering RNA (siRNA) has led to their use as target validation tools and as potential therapeutics for a variety of diseases. The delivery of these double-stranded RNA macromols. has proven to be challenging, however, and in many cases, is a barrier to their deployment. Here we report the development of a new diblock copolymer family that was designed to enhance the systemic and intracellular delivery of siRNA. These diblock copolymers were synthesized using the controlled reversible addn. fragmentation chain transfer polymn. (RAFT) method and are composed of a pos.-charged block of dimethylaminoethyl methacrylate (DMAEMA) to mediate siRNA condensation, and a second endosomal-releasing block composed of DMAEMA and propylacrylic acid (PAA) in roughly equimolar ratios, together with Bu methacrylate (BMA). A related series of diblock compns. were characterized, with the cationic block kept const., and with the ratio of DMAEMA and PAA to BMA varied. These carriers became sharply hemolytic at endosomal pH regimes, with increasing hemolytic activity seen as the percentage of BMA in the second block was systematically increased. The diblock copolymers condensed siRNA into 80-250 nm particles with slightly pos. Zeta potentials. SiRNA-mediated knockdown of a model protein, namely glyceraldehyde 3-phosphate dehydrogenase (GAPDH), in HeLa cells generally followed the hemolytic activity trends, with the most hydrophobic second block (highest BMA content) exhibiting the best knockdown. This pH-responsive carrier designed to mediate endosomal release shows significant promise for the intracellular delivery of siRNA.
36. 36

    Gody, G.; Maschmeyer, T.; Zetterlund, P. B.; Perrier, S. Exploitation of the Degenerative Transfer Mechanism in RAFT Polymerization for Synthesis of Polymer of High Livingness at Full Monomer Conversion. Macromolecules 2014, 47 (2), 639– 649,  DOI: 10.1021/ma402286e

    Google Scholar

    36

    Exploitation of the Degenerative Transfer Mechanism in RAFT Polymerization for Synthesis of Polymer of High Livingness at Full Monomer Conversion

    Gody, Guillaume; Maschmeyer, Thomas; Zetterlund, Per B.; Perrier, Sebastien

    Macromolecules (Washington, DC, United States) (2014), 47 (2), 639-649CODEN: MAMOBX; ISSN:0024-9297. (American Chemical Society)

    We report the synthesis by the reversible addn.-fragmentation chain transfer process of well-defined decablock polymers with a final dispersity as low as 1.15 and a fraction of living chain as high as 97% after 10 successful block extensions, each taken to >99% monomer conversion. By using model decablock homopolymers of poly(N,N-dimethylacrylamide) and poly(4-acryloylmorpholine) of relatively low DP (10 units per block in av.), we describe the theor. and exptl. considerations required to access high-order multiblock copolymers with excellent control over mol. wt. distributions and high livingness.
37. 37

    Moad, G. Mechanism and Kinetics of Dithiobenzoate-Mediated RAFT Polymerization - Status of the Dilemma. Macromol. Chem. Phys. 2014, 215 (1), 9– 26,  DOI: 10.1002/macp.201300562

    Google Scholar

    37

    Mechanism and Kinetics of Dithiobenzoate-Mediated RAFT Polymerization - Status of the Dilemma

    Moad, Graeme

    Macromolecular Chemistry and Physics (2014), 215 (1), 9-26CODEN: MCHPES; ISSN:1022-1352. (Wiley-VCH Verlag GmbH & Co. KGaA)

    A review. Dithiobenzoates are among the most popular agents for reversible addn.-fragmentation chain transfer (RAFT) polymn. This is attributed to the better control over mol. wt. and end-group fidelity found in RAFT polymn. of methacrylates and methacrylamides. However, in polymn. of styrenes, acrylates, and acrylamides, their use has diminished, mainly in favor of trithiocarbonates, because of issues with retardation, as well as hydrolytic and thermal instability. This paper critically assesses developments in understanding the mechanism and kinetics of dithiobenzoate-mediated RAFT polymn. from 2006 to 2013, with specific ref. to the choice of reagents, polymn. conditions, side reactions, and factors leading to retardation.
38. 38

    Keddie, D. J. A guide to the synthesis of block copolymers using reversible-addition fragmentation chain transfer (RAFT) polymerization. Chem. Soc. Rev. 2014, 43 (2), 496– 505,  DOI: 10.1039/C3CS60290G

    Google Scholar

    38

    A guide to the synthesis of block copolymers using reversible-addition fragmentation chain transfer (RAFT) polymerization

    Keddie, Daniel J.

    Chemical Society Reviews (2014), 43 (2), 496-505CODEN: CSRVBR; ISSN:0306-0012. (Royal Society of Chemistry)

    A review. The discovery of reversible-deactivation radical polymn. (RDRP) has provided an avenue for the synthesis of a vast array of polymers with a rich variety of functionality and architecture. The prepn. of block copolymers has received significant focus in this burgeoning research field, due to their diverse properties and potential in a wide range of research environments. This tutorial review will address the important concepts behind the design and synthesis of block copolymers using reversible addn.-fragmentation chain transfer (RAFT) polymn. RAFT polymn. is arguably the most versatile of the RDRP methods due to its compatibility with a wide range of functional monomers and reaction media along with its relative ease of use. With an ever increasing array of researchers that possess a variety of backgrounds now turning to RDRP, and RAFT in particular, to prep. their required polymeric materials, it is pertinent to discuss the important points which enable the prepn. of high purity functional block copolymers with targeted molar mass and narrow molar mass distribution using RAFT polymn. The key principles of appropriate RAFT agent selection, the order of monomer addn. in block synthesis and potential issues with maintaining high end-group fidelity are addressed. Addnl., techniques which allow block copolymers to be accessed using a combination of RAFT polymn. and complementary techniques are touched upon.
39. 39

    Gody, G.; Maschmeyer, T.; Zetterlund, P. B.; Perrier, S. Pushing the Limit of the RAFT Process: Multiblock Copolymers by One-Pot Rapid Multiple Chain Extensions at Full Monomer Conversion. Macromolecules 2014, 47 (10), 3451– 3460,  DOI: 10.1021/ma402435n

    Google Scholar

    39

    Pushing the Limit of the RAFT Process: Multiblock Copolymers by One-Pot Rapid Multiple Chain Extensions at Full Monomer Conversion

    Gody, Guillaume; Maschmeyer, Thomas; Zetterlund, Per B.; Perrier, Sebastien

    Macromolecules (Washington, DC, United States) (2014), 47 (10), 3451-3460CODEN: MAMOBX; ISSN:0024-9297. (American Chemical Society)

    We describe an optimized method to prep. multiblock copolymers. The approach is based on our previously reported use of reversible addn.-fragmentation chain transfer (RAFT) polymn., which here has been optimized into a fast, versatile, efficient, and scalable process. The one-pot, multistep sequential polymn. proceeds in water, to quant. yields (>99%) for each monomer addn., thus circumventing requirements for intermediate purifn., in 2 h of polymn. per block. The optimization of the process is initially demonstrated via the synthesis of a model decablock homopolymer (10 blocks) of 4-acryloylmorpholine with an av. d.p. of 10 for each block (D = 1.15 and livingness >93% for the final polymer). Both the potential and the limitations of this approach are illustrated by the synthesis of more complex high-order multiblock copolymers: a dodecablock copolymer (12 blocks with 4 different acrylamide monomers) with an av. d.p. of 10 for each block and two higher mol. wt. pentablock copolymers (5 blocks with 3 different acrylamide monomers) with an av. d.p. of 100 per block.
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    Richter, F.; Martin, L.; Leer, K.; Moek, E.; Hausig, F.; Brendel, J. C.; Traeger, A. Tuning of endosomal escape and gene expression by functional groups, molecular weight and transfection medium: a structure-activity relationship study. J. Mater. Chem. B 2020, 8 (23), 5026– 5041,  DOI: 10.1039/D0TB00340A

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    Tuning of endosomal escape and gene expression by functional groups, molecular weight and transfection medium: a structure-activity relationship study

    Richter, Friederike; Martin, Liam; Leer, Katharina; Moek, Elisabeth; Hausig, Franziska; Brendel, Johannes C.; Traeger, Anja

    Journal of Materials Chemistry B: Materials for Biology and Medicine (2020), 8 (23), 5026-5041CODEN: JMCBDV; ISSN:2050-7518. (Royal Society of Chemistry)

    The use of genetic material by non-viral transfer systems is still in its initial stages, but there are high expectations for the development of targeted therapies. However, nucleic acids cannot enter cells without help, they must be well protected to prevent degrdn. and overcome a variety of biol. barriers, the endosomal barrier being one of the greatest cellular challenges. Herein, the structure-property-relation was studied in detail, using well-defined polymers. Polyacrylamides were synthesized via RAFT polymn. resulting in a polymer library of (1) different cationic groups as aminoethyl acrylamide (AEAm), dimethylaminoethyl acrylamide (DMAEAm), dimethylaminopropyl acrylamide (DMAPAm) and guanidinopropyl acrylamide (GPAm); (2) different d.p.; and studied (3) in different cell culture settings. The influence of molar mass and cationic moiety on complex formation with pDNA, cytotoxicity and transfection efficiency of the polymers were studied. The systematic approach identified a pH-independent guanidinium-contg. homopolymer (PGPAm89) as the polymer with the highest transfection efficiency and superior endosomal release under optimal conditions. Since PGPAm89 is not further protonated inside endosomes, common escape theories appear unsuitable. Therefore, the interaction with bis(monoacryloylglycerol)phosphate, a lipid specific for endosomal vesicles, was studied. The authors' research suggests that the interactions between amines and lipids may be more relevant than anticipated.
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    Leer, K.; Reichel, L. S.; Kimmig, J.; Richter, F.; Hoeppener, S.; Brendel, J. C.; Zechel, S.; Schubert, U. S.; Traeger, A. Optimization of Mixed Micelles Based on Oppositely Charged Block Copolymers by Machine Learning for Application in Gene Delivery. Small 2023, e2306116  DOI: 10.1002/smll.202306116

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    Martin, L.; Gody, G.; Perrier, S. Preparation of complex multiblock copolymers via aqueous RAFT polymerization at room temperature. Polym. Chem. 2015, 6 (27), 4875– 4886,  DOI: 10.1039/C5PY00478K

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    Preparation of complex multiblock copolymers via aqueous RAFT polymerization at room temperature

    Martin, Liam; Gody, Guillaume; Perrier, Sebastien

    Polymer Chemistry (2015), 6 (27), 4875-4886CODEN: PCOHC2; ISSN:1759-9962. (Royal Society of Chemistry)

    We describe a simple new approach towards complex multiblock copolymer architectures at ambient temps. Using the redox pair TBHP/AsAc for initiation, aq. reversible addn.-fragmentation chain transfer (RAFT) polymn. is used to develop a one-pot sequential monomer addn. process that yields high order multiblock copolymers at 25 °C. Working at ambient temp. permits the polymn. of monomers yielding polymers with lower crit. soln. temp. (LCST) and reduces the risk of side reactions by chain transfer. Our approach is initially demonstrated with the prepn. of well-defined low and high d.p. (DP) poly(4-acryloylmorpholine) (PNAM) and poly(2-hydroxyethyl acrylate) (PHEA) multiblock homopolymers with D under 1.35. To highlight the potential of our approach, more challenging multiblock copolymers were prepd.: a pentablock copolymer of high DP (an av. of 100 per block) including low LCST blocks of poly(N-isopropylacrylamide) (PNIPAM) and poly(N,N-diethylacrylamide) (PDEA), two polyacrylate multiblock copolymers (DP of 10 per block) using a range of different functional acrylate monomers, and a heptablock copolymer (DP 10 per block) consisting of both polyacrylate and polyacrylamido blocks, all with a final dispersity of around 1.3.
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    Pergushov, D. V.; Müller, A. H. E.; Schacher, F. H. Micellar interpolyelectrolyte complexes. Chem. Soc. Rev. 2012, 41 (21), 6888– 6901,  DOI: 10.1039/c2cs35135h

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    Micellar interpolyelectrolyte complexes

    Pergushov, Dmitry V.; Mueller, Axel H. E.; Schacher, Felix H.

    Chemical Society Reviews (2012), 41 (21), 6888-6901CODEN: CSRVBR; ISSN:0306-0012. (Royal Society of Chemistry)

    A review. Interpolyelectrolyte complexes (IPECs) are typically formed when 2 polyelectrolytes of opposite charge are mixed together in soln. The authors present an overview of different strategies for the prepn. of micellar IPECs, i.e., structures where such IPEC domains form the core or the shell of micelles. In addn., vesicular architectures are considered, where the IPEC domain forms a membrane layer. One intriguing feature of IPECs is that their formation can be directed, their stability towards changes in pH or ionic strength can (to a certain extent) be predicted, and their size can be controlled. Esp. the use of ionic/non-ionic block copolymers offers unique potential for the prepn. of well-defined and sophisticated nanostructured materials. The authors also discuss possible applications, esp. in the field of life sciences, including biocompatibility, the controlled uptake/release of guest substances, the immobilization of enzymes, or the controlled formation of inorg./org. hybrid materials.
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    Solomun, J. I.; Cinar, G.; Mapfumo, P.; Richter, F.; Moek, E.; Hausig, F.; Martin, L.; Hoeppener, S.; Nischang, I.; Traeger, A. Solely aqueous formulation of hydrophobic cationic polymers for efficient gene delivery. Int. J. Pharm. 2021, 593, 120080,  DOI: 10.1016/j.ijpharm.2020.120080

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    Solely aqueous formulation of hydrophobic cationic polymers for efficient gene delivery

    Solomun, Jana I.; Cinar, Gizem; Mapfumo, Prosper; Richter, Friederike; Moek, Elisabeth; Hausig, Franziska; Martin, Liam; Hoeppener, Stephanie; Nischang, Ivo; Traeger, Anja

    International Journal of Pharmaceutics (Amsterdam, Netherlands) (2021), 593 (), 120080CODEN: IJPHDE; ISSN:0378-5173. (Elsevier B.V.)

    Cationic polymers are promising gene delivery vectors due to their ability to bind and protect genetic material. The introduction of hydrophobic moieties into cationic polymers can further improve the vector efficiency, but common formulations of hydrophobic polymers involve harsh conditions such as org. solvents, impairing intactness and loading efficiency of the genetic material. In this study, a mild, aq. formulation method for the encapsulation of high amts. of genetic material is presented. A well-defined pH-responsive hydrophobic copolymer, i.e. poly((n-butylmethacrylate)-co-(methylmethacrylate)-co-(2-(dimethylamino) ethylmethacrylate)), (PBMD) was synthesized by reversible addn. fragmentation chain transfer (RAFT) polymn. Exploiting the pH-dependent soly. behavior of the polymer, stable pDNA loaded nanoparticles were prepd. and characterized using anal. ultracentrifugation (AUC), cryo-transmission electron microscopy (cryo-TEM) and dynamic light scattering (DLS). This novel formulation approach showed high transfection efficiencies in HEK293T cells, while requiring 5- to 10-fold less pDNA compared to linear polyethylenimine (LPEI), in particular at short incubation times and in serum-contg. media. Furthermore, the formulation was successfully adopted for siRNA and mRNA encapsulation and the com. approved polymer Eudragit E(PO/100). Overall, the aq. formulation approach, accompanied by a tailor-made hydrophobic polymer and detailed physicochem. and application studies, led to improved gene delivery vectors with high potential for further applications.