bims-novged 2023-10-15 papers

bims-novged

Biomed News

on Non-viral vectors for gene delivery

Issue of 2023–10–15
fourteen papers selected by
the Merkel lab, Ludwig-Maximilians University

Lipid nanoparticles with PEG-variant surface modifications mediate genome editing in the mouse retina.
Effect of Polyplex Size on Penetration into Tumor Spheroids.
The alpha-adrenergic antagonist prazosin promotes cytosolic siRNA delivery from lysosomal compartments.
Effect of mRNA-LNP components of two globally-marketed COVID-19 vaccines on efficacy and stability.
mRNA-based vaccines and therapeutics: an in-depth survey of current and upcoming clinical applications.
Cancer/Testis Antigens as Targets for RNA-Based Anticancer Therapy.
An Ionizable Lipid Material with a Vitamin E Scaffold as an mRNA Vaccine Platform for Efficient Cytotoxic T Cell Responses.
siRNA Lipid-Polymer Nanoparticles Targeting E-Selectin and Cyclophilin A in Bone Marrow for Combination Multiple Myeloma Therapy.
Lyophilization process optimization and molecular dynamics simulation of mRNA-LNPs for SARS-CoV-2 vaccine.
Engineered mRNA Delivery Systems for Biomedical Applications.
Design of siRNA Bioconjugates for Efficient Control of Cancer-Associated Membrane Receptors.
Antimicrobial polymer-siRNA polyplexes as a dual-mode platform for the treatment of wound biofilm infections.
The Dual Role of the Innate Immune System in the Effectiveness of mRNA Therapeutics.
A potential paradigm in CRISPR/Cas systems delivery: at the crossroad of microalgal gene editing and algal-mediated nanoparticles.

Nat Commun. 2023 Oct 13. 14(1): 6468

Lipid nanoparticles with PEG-variant surface modifications mediate genome editing in the mouse retina.

Milan Gautam, Antony Jozic, Grace Li-Na Su, Marco Herrera-Barrera, Allison Curtis, Sebastian Arrizabalaga, Wayne Tschetter, Renee C Ryals, Gaurav Sahay.

Ocular delivery of lipid nanoparticle (LNPs) packaged mRNA can enable efficient gene delivery and editing. We generated LNP variants through the inclusion of positively charged-amine-modified polyethylene glycol (PEG)-lipids (LNPa), negatively charged-carboxyl-(LNPz) and carboxy-ester (LNPx) modified PEG-lipids, and neutral unmodified PEG-lipids (LNP). Subretinal injections of LNPa containing Cre mRNA in the mouse show tdTomato signal in the retinal pigmented epithelium (RPE) like conventional LNPs. Unexpectedly, LNPx and LNPz show 27% and 16% photoreceptor transfection, respectively, with striking localization extending from the photoreceptor synaptic pedicle to the outer segments, displaying pan-retinal distribution in the photoreceptors and RPE. LNPx containing Cas9 mRNA and sgAi9 leads to the formation of an oval elongated structure with a neutral charge resulting in 16.4% editing restricted to RPE. Surface modifications of LNPs with PEG variants can alter cellular tropism of mRNA. LNPs enable genome editing in the retina and in the future can be used to correct genetic mutations that lead to blindness.

DOI: https://doi.org/10.1038/s41467-023-42189-3
Mol Pharm. 2023 Oct 09.

Effect of Polyplex Size on Penetration into Tumor Spheroids.

Cristina Casadidio, Jet E M Hartman, Bárbara S Mesquita, Ragna Haegebaert, Katrien Remaut, Myriam Neumann, Jaimie Hak, Roberta Censi, Piera Di Martino, Wim E Hennink, Tina Vermonden.

  Ovarian cancer is one of the most lethal gynecological cancers in the world. In recent years, nucleic acid (NA)-based formulations have been shown to be promising treatments for ovarian cancer, including tumor nodules. However, gene therapy is not that far advanced in clinical reality due to unfavorable physicochemical properties of the NAs, such as high molecular weight, poor cellular uptake, rapid degradation by nucleases, etc. One of the strategies used to overcome these drawbacks is the complexation of anionic NAs via electrostatic interactions with cationic polymers, resulting in the formation of so-called polyplexes. In this work, the role of the size of pDNA and siRNA polyplexes on their penetration into ovarian-cancer-based tumor spheroids was investigated. For this, a methoxypoly(ethylene glycol) poly(2-(dimethylamino)ethyl methacrylate) (mPEG-pDMAEMA) diblock copolymer was synthesized as a polymeric carrier for NA binding and condensation with either plasmid DNA (pDNA) or short interfering RNA (siRNA). When prepared in HEPES buffer (10 mM, pH 7.4) at a nitrogen/phosphate (N/P) charge ratio of 5 and pDNA polyplexes were formed with a size of 162 ± 11 nm, while siRNA-based polyplexes displayed a size of 25 ± 2 nm. The polyplexes had a slightly positive zeta potential of +7-8 mV in the same buffer. SiRNA and pDNA polyplexes were tracked in vitro into tumor spheroids, resembling in vivo avascular ovarian tumor nodules. For this purpose, reproducible spheroids were obtained by coculturing ovarian carcinoma cells with primary mouse embryonic fibroblasts in different ratios (5:2, 1:1, and 2:5). Penetration studies revealed that after 24 h of incubation, siRNA polyplexes were able to penetrate deeper into the homospheroids (composed of only cancer cells) and heterospheroids (cancer cells cocultured with fibroblasts) compared to pDNA polyplexes which were mainly located in the rim. The penetration of the polyplexes was slowed when increasing the fraction of fibroblasts present in the spheroids. Furthermore, in the presence of serum siRNA polyplexes encoding for luciferase showed a high cellular uptake in 2D cells resulting in ∼50% silencing of luciferase expression. Taken together, these findings show that self-assembled small siRNA polyplexes have good potential as a platform to test ovarian tumor nodulus penetration..

Keywords:  3D in vitro model; gene delivery; tumor penetration; tumor stroma

DOI:  https://doi.org/10.1021/acs.molpharmaceut.3c00397
J Control Release. 2023 Oct 08. pii: S0168-3659(23)00664-8. [Epub ahead of print]

The alpha-adrenergic antagonist prazosin promotes cytosolic siRNA delivery from lysosomal compartments.

Thijs Van de Vyver, Cristina Muntean, Iuliia Efimova, Dmitri V Krysko, Lynn De Backer, Stefaan C De Smedt, Koen Raemdonck.

  The widespread use of small interfering RNA (siRNA) is limited by the multiple extra- and intracellular barriers upon in vivo administration. Hence, suitable delivery systems, based on siRNA encapsulation in nanoparticles or its conjugation to targeting ligands, have been developed. Nevertheless, at the intracellular level, these state-of-the-art delivery systems still suffer from a low endosomal escape efficiency. Consequently, the bulk of the endocytosed siRNA drug rapidly accumulates in the lysosomal compartment. We recently reported that a wide variety of cationic amphiphilic drugs (CADs) can promote small nucleic acid delivery from the endolysosomal compartment into the cytosol via transient induction of lysosomal membrane permeabilization. Here, we describe the identification of alternate siRNA delivery enhancers from the NIH Clinical Compound Collection that do not have the typical physicochemical properties of CADs. Additionally, we demonstrate improved endolysosomal escape of siRNA via a cholesterol conjugate and polymeric carriers with the α1-adrenergic antagonist prazosin, which was identified as the best performing delivery enhancer from the compound screen. A more detailed assessment of the mode-of-action of prazosin suggests that a different cellular phenotype compared to typical CAD adjuvants drives cytosolic siRNA delivery. As it has been described in the literature that prazosin also induces cancer cell apoptosis and promotes antigen cross-presentation in dendritic cells, the proof-of-concept data in this work provides opportunities for the repurposing of prazosin in an anti-cancer combination strategy with siRNA.

Keywords:  Cationic amphiphilic drugs; Combination therapy; Drug repurposing; Endosomal escape; Lysosomal membrane permeabilization; cancer immunotherapy; cancer therapy

DOI:  https://doi.org/10.1016/j.jconrel.2023.10.014
NPJ Vaccines. 2023 Oct 11. 8(1): 156

Effect of mRNA-LNP components of two globally-marketed COVID-19 vaccines on efficacy and stability.

Lizhou Zhang, Kunal R More, Amrita Ojha, Cody B Jackson, Brian D Quinlan, Hao Li, Wenhui He, Michael Farzan, Norbert Pardi, Hyeryun Choe.

During the COVID-19 pandemic, Pfizer-BioNTech and Moderna successfully developed nucleoside-modified mRNA lipid nanoparticle (LNP) vaccines. SARS-CoV-2 spike protein expressed by those vaccines are identical in amino acid sequence, but several key components are distinct. Here, we compared the effect of ionizable lipids, untranslated regions (UTRs), and nucleotide composition of the two vaccines, focusing on mRNA delivery, antibody generation, and long-term stability. We found that the ionizable lipid, SM-102, in Moderna's vaccine performs better than ALC-0315 in Pfizer-BioNTech's vaccine for intramuscular delivery of mRNA and antibody production in mice and long-term stability at 4 °C. Moreover, Pfizer-BioNTech's 5' UTR and Moderna's 3' UTR outperform their counterparts in their contribution to transgene expression in mice. We further found that varying N1-methylpseudouridine content at the wobble position of mRNA has little effect on vaccine efficacy. These findings may contribute to the further improvement of nucleoside-modified mRNA-LNP vaccines and therapeutics.

DOI: https://doi.org/10.1038/s41541-023-00751-6
J Biomed Sci. 2023 Oct 07. 30(1): 84

mRNA-based vaccines and therapeutics: an in-depth survey of current and upcoming clinical applications.

Yu-Shiuan Wang, Monika Kumari, Guan-Hong Chen, Ming-Hsiang Hong, Joyce Pei-Yi Yuan, Jui-Ling Tsai, Han-Chung Wu.

  mRNA-based drugs have tremendous potential as clinical treatments, however, a major challenge in realizing this drug class will promise to develop methods for safely delivering the bioactive agents with high efficiency and without activating the immune system. With regard to mRNA vaccines, researchers have modified the mRNA structure to enhance its stability and promote systemic tolerance of antigenic presentation in non-inflammatory contexts. Still, delivery of naked modified mRNAs is inefficient and results in low levels of antigen protein production. As such, lipid nanoparticles have been utilized to improve delivery and protect the mRNA cargo from extracellular degradation. This advance was a major milestone in the development of mRNA vaccines and dispelled skepticism about the potential of this technology to yield clinically approved medicines. Following the resounding success of mRNA vaccines for COVID-19, many other mRNA-based drugs have been proposed for the treatment of a variety of diseases. This review begins with a discussion of mRNA modifications and delivery vehicles, as well as the factors that influence administration routes. Then, we summarize the potential applications of mRNA-based drugs and discuss further key points pertaining to preclinical and clinical development of mRNA drugs targeting a wide range of diseases. Finally, we discuss the latest market trends and future applications of mRNA-based drugs.

Keywords:  Administration routes; Lipid nanoparticles; Targeting mRNA delivery system; mRNA therapeutics; mRNA vaccine

DOI:  https://doi.org/10.1186/s12929-023-00977-5
Int J Mol Sci. 2023 Sep 28. pii: 14679. [Epub ahead of print]24(19):

Cancer/Testis Antigens as Targets for RNA-Based Anticancer Therapy.

Kyeonghee Shim, Hyein Jo, Dooil Jeoung.

  In the last few decades, RNA-based drugs have emerged as a promising candidate in the treatment of various diseases. The introduction of messenger RNA (mRNA) as a vaccine or therapeutic agent enables the production of almost any functional protein/peptide. The key to applying RNA therapy in clinical trials is developing safe and effective delivery systems. Exosomes and lipid nanoparticles (LNPs) have been exploited as promising vehicles for drug delivery. This review discusses the feasibility of exosomes and LNPs as vehicles for mRNA delivery. Cancer/testis antigens (CTAs) show restricted expression in normal tissues and widespread expression in cancer tissues. Many of these CTAs show expression in the sera of patients with cancers. These characteristics of CTAs make them excellent targets for cancer immunotherapy. This review summarizes the roles of CTAs in various life processes and current studies on mRNAs encoding CTAs. Clinical studies present the beneficial effects of mRNAs encoding CTAs in patients with cancers. This review highlight clinical studies employing mRNA-LNPs encoding CTAs.

Keywords:  RNA delivery; RNA vaccines; cancer/testis antigens; clinical trials; lipid nanoparticles; tumor-associated antigens

DOI:  https://doi.org/10.3390/ijms241914679
ACS Nano. 2023 10 10. 17(19): 18758-18774

An Ionizable Lipid Material with a Vitamin E Scaffold as an mRNA Vaccine Platform for Efficient Cytotoxic T Cell Responses.

Ryotaro Oyama, Harumichi Ishigame, Hiroki Tanaka, Naho Tateshita, Moeko Itazawa, Ryosuke Imai, Naomasa Nishiumi, Jun-Ichi Kishikawa, Takayuki Kato, Jessica Anindita, Yoshifumi Nishikawa, Masatoshi Maeki, Manabu Tokeshi, Kota Tange, Yuta Nakai, Yu Sakurai, Takaharu Okada, Hidetaka Akita.

  RNA vaccines based on lipid nanoparticles (LNPs) with in vitro transcribed mRNA (IVT-mRNA) encapsulated are now a currently successful but still evolving modality of vaccines. One of the advantages of RNA vaccines is their ability to induce CD8+ T-cell-mediated cellular immunity that is indispensable for excluding pathogen-infected cells or cancer cells from the body. In this study, we report on the development of LNPs with an enhanced capability for inducing cellular immunity by using an ionizable lipid with a vitamin E scaffold. An RNA vaccine that contained this ionizable lipid and an IVT-mRNA encoding a model antigen ovalbumin (OVA) induced OVA-specific cytotoxic T cell responses and showed an antitumor effect against an E.G7-OVA tumor model. Vaccination with the LNPs conferred protection against lethal infection by Toxoplasma gondii using its antigen TgPF. The vitamin E scaffold-dependent type I interferon response was important for effector CD8+ T cell differentiation induced by the mRNA-LNPs. Our findings also revealed that conventional dendritic cells (cDCs) were essential for achieving CD8+ T cell responses induced by the mRNA-LNPs, while the XCR1-positive subset of cDCs, cDC1 specialized for antigen cross-presentation, was not required. Consistently, the mRNA-LNPs were found to selectively transfect another subset of cDCs, cDC2 that had migrated from the skin to lymph nodes, where they could make vaccine-antigen-dependent contacts with CD8+ T cells. The findings indicate that the activation of innate immune signaling by the adjuvant activity of the vitamin E scaffold and the expression of antigens in cDC2 are important for subsequent antigen presentation and the establishment of antigen-specific immune responses.

Keywords:  RNA vaccine; adjuvant; cellular immunity; innate immunity; lipid nanoparticle; vitamin E

DOI:  https://doi.org/10.1021/acsnano.3c02251
Cell Mol Bioeng. 2023 Aug;16(4): 383-392

siRNA Lipid-Polymer Nanoparticles Targeting E-Selectin and Cyclophilin A in Bone Marrow for Combination Multiple Myeloma Therapy.

Christian G Figueroa-Espada, Pedro P G Guimarães, Rachel S Riley, Lulu Xue, Karin Wang, Michael J Mitchell.

   Introduction: Multiple myeloma (MM) is a hematological blood cancer of the bone marrow that remains largely incurable, in part due to its physical interactions with the bone marrow microenvironment. Such interactions enhance the homing, proliferation, and drug resistance of MM cells. Specifically, adhesion receptors and homing factors, E-selectin (ES) and cyclophilin A (CyPA), respectively, expressed by bone marrow endothelial cells enhance MM colonization and dissemination. Thus, silencing of ES and CyPA presents a potential therapeutic strategy to evade MM spreading. However, small molecule inhibition of ES and CyPA expressed by bone marrow endothelial cells remains challenging, and blocking antibodies induce further MM propagation. Therefore, ES and CyPA are promising candidates for inhibition via RNA interference (RNAi).
Methods: Here, we utilized a previously developed lipid-polymer nanoparticle for RNAi therapy, that delivers siRNA to the bone marrow perivascular niche. We utilized our platform to co-deliver ES and CyPA siRNAs to prevent MM dissemination in vivo.
Results: Lipid-polymer nanoparticles effectively downregulated ES expression in vitro, which decreased MM cell adhesion and migration through endothelial monolayers. Additionally, in vivo delivery of lipid-polymer nanoparticles co-encapsulating ES and CyPA siRNA extended survival in a xenograft mouse model of MM, either alone or in combination with the proteasome inhibitor bortezomib.
Conclusions: Our combination siRNA lipid-polymer nanoparticle therapy presents a vascular microenvironment-targeting strategy as a potential paradigm shift for MM therapies, which could be extended to other cancers that colonize the bone marrow.
Supplementary Information: The online version contains supplementary material available at 10.1007/s12195-023-00774-y.

Keywords:  Blood cancer; Nanomedicine; RNAi

DOI:  https://doi.org/10.1007/s12195-023-00774-y
NPJ Vaccines. 2023 Oct 09. 8(1): 153

Lyophilization process optimization and molecular dynamics simulation of mRNA-LNPs for SARS-CoV-2 vaccine.

Mingyuan Li, Lin Jia, Yanbo Xie, Wenlin Ma, Zhihong Yan, Fufeng Liu, Jie Deng, Ali Zhu, Xue Siwei, Wen Su, Xiaofeng Liu, Shiqin Li, Haomeng Wang, Peng Yu, Tao Zhu.

Some studies have shown that lyophilization significantly improves the stability of mRNA-LNPs and enables long-term storage at 2-8 °C. However, there is little research on the lyophilization process of mRNA-lipid nanoparticles (LNPs). Most previous studies have used empirical lyophilization with only a single lyoprotectant, resulting in low lyophilization efficiency, often requiring 40-100 h. In the present study, an efficient lyophilization method suitable for mRNA-LNPs was designed and optimized, shortening the total length of the lyophilization process to 8-18 h, which significantly reduced energy consumption and production costs. When the mixed lyoprotectant composed of sucrose, trehalose, and mannitol was added to mRNA-LNPs, the eutectic point and collapse temperature of the system were increased. The lyophilized product had a ginger root-shaped rigid structure with large porosity, which tolerated rapid temperature increases and efficiently removed water. In addition, the lyophilized mRNA-LNPs rapidly rehydrated and had good particle size distribution, encapsulation rate, and mRNA integrity. The lyophilized mRNA-LNPs were stable at 2-8 °C, and they did not reduce immunogenicity in vivo or in vitro. Molecular dynamics simulation was used to compare the phospholipid molecular layer with the lyoprotectant in aqueous and anhydrous environments to elucidate the mechanism of lyophilization to improve the stability of mRNA-LNPs. This efficient lyophilization platform significantly improves the accessibility of mRNA-LNPs.

DOI: https://doi.org/10.1038/s41541-023-00732-9
Adv Mater. 2023 Oct 08. e2308029

Engineered mRNA Delivery Systems for Biomedical Applications.

Ji Wang, Haofang Zhu, Jingjing Gan, Gaofeng Liang, Ling Li, Yuanjin Zhao.

  Messenger RNA (mRNA)-based therapeutic strategies have shown remarkable promise in preventing and treating a staggering range of diseases. Optimizing the structure and delivery system of engineered mRNA has greatly improved its stability, immunogenicity, and protein expression levels, which has led to a wider range of uses for mRNA therapeutics. In this tutorial review, we first provide a thorough analysis of the optimization strategies used in the structure of mRNA and then describe delivery systems in great detail. Furthermore, we summarize the latest advancements in biomedical engineering for mRNA technology, including its applications in combatting infectious diseases, treating cancer, providing protein replacement therapy, conducting gene editing, and more. Lastly, we offer a perspective on forthcoming challenges and prospects concerning the advancement of mRNA therapeutics. Despite these challenges, mRNA-based therapeutics remain promising, with the potential to revolutionize disease treatment and contribute to significant advancements in the biomedical field. This article is protected by copyright. All rights reserved.

Keywords:  biomaterials; delivery systems; mRNA; particles; therapeutics

DOI:  https://doi.org/10.1002/adma.202308029
ACS Omega. 2023 Oct 03. 8(39): 36435-36448

Design of siRNA Bioconjugates for Efficient Control of Cancer-Associated Membrane Receptors.

Jong Won Lee, Jiwoong Choi, Eun Hye Kim, Jiwon Choi, Sun Hwa Kim, Yoosoo Yang.

Research on siRNA delivery has seen tremendous growth over the past few decades. As one of the major delivery strategies, siRNA bioconjugates offer the potential to enhance and extend the pharmacological properties of siRNAs while minimizing toxicity. In this paper, we suggest the development of a siRNA conjugate platform with peptides and proteins that are ligands of target receptors for cancer treatment. The siRNA bioconjugates target and block the receptor membrane proteins, enter the cells through receptor-mediated endocytosis, and inhibit the expression of that same target membrane receptor, thereby doubly controlling the function of the membrane proteins. The three kinds of bioconjugates targeting CD47, PD-L1, and EGFR were synthesized via two different copper-free click chemistry reactions. Results showed the cellular uptake of each conjugate, reduction of target gene expression, and efficient functional control of receptor proteins. This platform provides an effective approach for regulating membrane proteins in various diseases beyond cancer.

DOI: https://doi.org/10.1021/acsomega.3c05395
Mater Horiz. 2023 Oct 10.

Antimicrobial polymer-siRNA polyplexes as a dual-mode platform for the treatment of wound biofilm infections.

Taewon Jeon, Jessa Marie V Makabenta, Jungmi Park, Ahmed Nabawy, Yagiz Anil Cicek, Sarah S Mirza, Janelle Welton, Muhammad Aamir Hassan, Rui Huang, Jesse Mager, Vincent M Rotello.

Treatment of wound biofilm infections faces challenges from both pathogens and uncontrolled host immune response. Treating both issues through a single vector would provide enhanced wound healing. Here, we report the use of a potent cationic antimicrobial polymer to generate siRNA polyplexes for dual-mode treatment of wound biofilms in vivo. These polyplexes act both as an antibiofilm agent and a delivery vehicle for siRNA for the knockdown of biofilm-associated pro-inflammatory MMP9 in host macrophages. The resulting polyplexes were effective in vitro, eradicating MRSA biofilms and efficiently delivering siRNA to macrophages in vitro with concomitant knockdown of MMP9. These polyplexes were likewise effective in an in vivo murine wound biofilm model, significantly reducing bacterial load in the wound (∼99% bacterial clearance) and reducing MMP9 expression by 80% (qRT-PCR). This combination therapeutic strategy dramatically reduced wound purulence and significantly expedited wound healing. Taken together, these polyplexes provide an effective and translatable strategy for managing biofilm-infected wounds.

DOI: https://doi.org/10.1039/d3mh01108a
Int J Mol Sci. 2023 Oct 01. pii: 14820. [Epub ahead of print]24(19):

The Dual Role of the Innate Immune System in the Effectiveness of mRNA Therapeutics.

Albert Muslimov, Valeriy Tereshchenko, Daniil Shevyrev, Anna Rogova, Kirill Lepik, Vasiliy Reshetnikov, Roman Ivanov.

  Advances in molecular biology have revolutionized the use of messenger RNA (mRNA) as a therapeutic. The concept of nucleic acid therapy with mRNA originated in 1990 when Wolff et al. reported successful expression of proteins in target organs by direct injection of either plasmid DNA or mRNA. It took decades to bring the transfection efficiency of mRNA closer to that of DNA. The next few decades were dedicated to turning in vitro-transcribed (IVT) mRNA from a promising delivery tool for gene therapy into a full-blown therapeutic modality, which changed the biotech market rapidly. Hundreds of clinical trials are currently underway using mRNA for prophylaxis and therapy of infectious diseases and cancers, in regenerative medicine, and genome editing. The potential of IVT mRNA to induce an innate immune response favors its use for vaccination and immunotherapy. Nonetheless, in non-immunotherapy applications, the intrinsic immunostimulatory activity of mRNA directly hinders the desired therapeutic effect since it can seriously impair the target protein expression. Targeting the same innate immune factors can increase the effectiveness of mRNA therapeutics for some indications and decrease it for others, and vice versa. The review aims to present the innate immunity-related 'barriers' or 'springboards' that may affect the development of immunotherapies and non-immunotherapy applications of mRNA medicines.

Keywords:  DAMP; NF-κB; NLRP3; PAMP; adjuvants; cGAS/STING; innate immunity; mRNA

DOI:  https://doi.org/10.3390/ijms241914820
J Nanobiotechnology. 2023 Oct 10. 21(1): 370

A potential paradigm in CRISPR/Cas systems delivery: at the crossroad of microalgal gene editing and algal-mediated nanoparticles.

Shuying Feng, Xin Xie, Junjie Liu, Aifang Li, Qianqian Wang, Dandan Guo, Shuxuan Li, Yalan Li, Zilong Wang, Tao Guo, Jin Zhou, Doris Ying Ying Tang, Pau Loke Show.

  Microalgae as the photosynthetic organisms offer enormous promise in a variety of industries, such as the generation of high-value byproducts, biofuels, pharmaceuticals, environmental remediation, and others. With the rapid advancement of gene editing technology, CRISPR/Cas system has evolved into an effective tool that revolutionised the genetic engineering of microalgae due to its robustness, high target specificity, and programmability. However, due to the lack of robust delivery system, the efficacy of gene editing is significantly impaired, limiting its application in microalgae. Nanomaterials have become a potential delivery platform for CRISPR/Cas systems due to their advantages of precise targeting, high stability, safety, and improved immune system. Notably, algal-mediated nanoparticles (AMNPs), especially the microalgae-derived nanoparticles, are appealing as a sustainable delivery platform because of their biocompatibility and low toxicity in a homologous relationship. In addition, living microalgae demonstrated effective and regulated distribution into specified areas as the biohybrid microrobots. This review extensively summarised the uses of CRISPR/Cas systems in microalgae and the recent developments of nanoparticle-based CRISPR/Cas delivery systems. A systematic description of the properties and uses of AMNPs, microalgae-derived nanoparticles, and microalgae microrobots has also been discussed. Finally, this review highlights the challenges and future research directions for the development of gene-edited microalgae.

Keywords:  Algal-mediated nanoparticles; CRISPR/Cas system; Delivery system; Gene editing; Microalgae application

DOI:  https://doi.org/10.1186/s12951-023-02139-z