bims-novged 2022-12-18 papers

bims-novged

Biomed News

on Non-viral vectors for gene delivery

Issue of 2022–12–18
nine papers selected by
the Merkel lab, Ludwig-Maximilians University

Engineering poly- and micelleplexes for nucleic acid delivery - A reflection on their endosomal escape.
Interaction Kinetics of Individual mRNA-Containing Lipid Nanoparticles with an Endosomal Membrane Mimic: Dependence on pH, Protein Corona Formation, and Lipoprotein Depletion.
A comprehensive update of siRNA delivery design strategies for targeted and effective gene silencing in gene therapy and other applications.
Elucidating the nanostructure of small interfering RNA-loaded lipidoid-polymer hybrid nanoparticles.
Engineered Ionizable Lipid Nanoparticles mediated Efficient siRNA Delivery to Macrophages for Anti-inflammatory Treatment of Acute Liver Injury.
Excipient-Free Ionizable Polyester Nanoparticles for Lung-Selective and Innate Immune Cell Plasmid DNA and mRNA Transfection.
Inhalable exosomes outperform liposomes as mRNA and protein drug carriers to the lung.
Down-regulation of MDM2 by small interference RNA induces apoptosis and sensitizes MCF-7 breast cells to resveratrol.
The Impact of Nanobody Density on the Targeting Efficiency of PEGylated Liposomes.

J Control Release. 2022 Dec 09. pii: S0168-3659(22)00822-7. [Epub ahead of print]353 518-534

Engineering poly- and micelleplexes for nucleic acid delivery - A reflection on their endosomal escape.

Benjamin Winkeljann, David C Keul, Olivia M Merkel.

  For the longest time, the field of nucleic acid delivery has remained skeptical whether or not polycationic drug carrier systems would ever make it into clinical practice. Yet, with the disclosure of patents on polyethyleneimine-based RNA carriers through leading companies in the field of nucleic acid therapeutics such as BioNTech SE and the progress in clinical studies beyond phase I trials, this aloofness seems to regress. As one of the most striking characteristics of polymer-based vectors, the extraordinary tunability can be both a blessing and a curse. Yet, knowing about the adjustment screws and how they impact the performance of the drug carrier provides the formulation scientist committed to its development with a head start. Here, we equip the reader with a toolbox - a toolbox that should advise and support the developer to conceptualize a cutting-edge poly- or micelleplex system for the delivery of therapeutic nucleic acids; to be specific, to engineer the vector towards maximum endosomal escape performance at minimum toxicity. Therefore, after briefly sketching the boundary conditions of polymeric vector design, we will dive into the topic of endosomal trafficking. We will not only discuss the most recent knowledge of the endo-lysosomal compartment but further depict different hypotheses and mechanisms that facilitate the endosomal escape of polyplex systems. Finally, we will combine the different facets introduced in the previous chapters with the fundamental building blocks of polymer vector design and evaluate the advantages and drawbacks. Throughout the article, a particular focus will be placed on cellular peculiarities, not only as an additional barrier, but also to give inspiration to how such cell-specific traits might be capitalized on.

Keywords:  Nonviral vector; Polyethyleneimine; Proton sponge hypothesis; RNA; Vaccine

DOI:  https://doi.org/10.1016/j.jconrel.2022.12.008
ACS Nano. 2022 Dec 13.

Interaction Kinetics of Individual mRNA-Containing Lipid Nanoparticles with an Endosomal Membrane Mimic: Dependence on pH, Protein Corona Formation, and Lipoprotein Depletion.

Nima Aliakbarinodehi, Audrey Gallud, Mokhtar Mapar, Emelie Wesén, Sahar Heydari, Yujia Jing, Gustav Emilsson, Kai Liu, Alan Sabirsh, Vladimir P Zhdanov, Lennart Lindfors, Elin K Esbjörner, Fredrik Höök.

  Lipid nanoparticles (LNPs) have emerged as potent carriers for mRNA delivery, but several challenges remain before this approach can offer broad clinical translation of mRNA therapeutics. To improve their efficacy, a better understanding is required regarding how LNPs are trapped and processed at the anionic endosomal membrane prior to mRNA release. We used surface-sensitive fluorescence microscopy with single LNP resolution to investigate the pH dependency of the binding kinetics of ionizable lipid-containing LNPs to a supported endosomal model membrane. A sharp increase of LNP binding was observed when the pH was lowered from 6 to 5, accompanied by stepwise large-scale LNP disintegration. For LNPs preincubated in serum, protein corona formation shifted the onset of LNP binding and subsequent disintegration to lower pH, an effect that was less pronounced for lipoprotein-depleted serum. The LNP binding to the endosomal membrane mimic was observed to eventually become severely limited by suppression of the driving force for the formation of multivalent bonds during LNP attachment or, more specifically, by charge neutralization of anionic lipids in the model membrane due to their association with cationic lipids from earlier attached LNPs upon their disintegration. Cell uptake experiments demonstrated marginal differences in LNP uptake in untreated and lipoprotein-depleted serum, whereas lipoprotein-depleted serum increased mRNA-controlled protein (eGFP) production substantially. This complies with model membrane data and suggests that protein corona formation on the surface of the LNPs influences the nature of the interaction between LNPs and endosomal membranes.

Keywords:  endosomal membrane; ionizable lipid nanoparticle; lipoprotein; mRNA delivery; protein corona

DOI:  https://doi.org/10.1021/acsnano.2c04829
Expert Opin Drug Discov. 2022 Dec 17. 1-13

A comprehensive update of siRNA delivery design strategies for targeted and effective gene silencing in gene therapy and other applications.

Ahmed Khaled Abosalha, Waqar Ahmad, Jacqueline Boyajian, Paromita Islam, Merry Ghebretatios, Sabrina Schaly, Rahul Thareja, Karan Arora, Satya Prakash.

   INTRODUCTION: RNA interference (RNAi) using small interfering RNA (siRNA) is a promising strategy to control many genetic disorders by targeting the mRNA of underlying genes and degrade it. However, the delivery of siRNA to targeted organs is highly restricted by several intracellular and extracellular barriers.
AREAS COVERED: This review discusses various design strategies developed to overcome siRNA delivery obstacles. The applied techniques involve chemical modification, bioconjugation to specific ligands, and carrier-mediated strategies. Nanotechnology-based systems like liposomes, niosomes, solid lipid nanoparticles (SLNs), dendrimers, and polymeric nanoparticles (PNs) are also discussed.
EXPERT OPINION: Although the mechanism of siRNA as a gene silencer is well-established, only a few products are available as therapeutics. There is a great need to develop and establish siRNA delivery systems that protects siRNAs and delivers them efficiently to the desired sitesare efficient and capable of targeted delivery. Several diseases are reported to be controlled by siRNA at their early stages. However, their targeted delivery is a daunting challenge.

Keywords:  Small interfering RNA (siRNA); bioconjugation; chemical modification; drug delivery; gene delivery; gene silencing; gene therapy; nanotechnology; targeted gene delivery

DOI:  https://doi.org/10.1080/17460441.2022.2155630
J Colloid Interface Sci. 2022 Nov 28. pii: S0021-9797(22)02125-7. [Epub ahead of print]633 907-922

Elucidating the nanostructure of small interfering RNA-loaded lipidoid-polymer hybrid nanoparticles.

Anas Aljabbari, Abhijeet Girish Lokras, Jacob Judas Kain Kirkensgaard, Thomas Rades, Henrik Franzyk, Aneesh Thakur, Yibang Zhang, Camilla Foged.

  We analyzed the structural and material properties of small interfering RNA (siRNA)-loaded lipid-polymer hybrid nanoparticles (LPNs) containing ionizable lipidoid and poly(dl-lactic-co-glycolic acid) (PLGA) using small-angle X-ray scattering, cryogenic transmission electron microscopy, polarized light microscopy, the Langmuir monolayer methodology, differential scanning calorimetry, and attenuated total reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy. Scattering analyses showed that bulk lipidoid self-assemble into lamellar structures with a d-spacing of 38 Å, whereas lipidoid-siRNA lipoplexes display an in-plane lateral organization of siRNA in between lipidoid bilayers with a repeat distance of approximately 55 Å. The siRNA-loaded LPNs adopted a core-shell structure with an interaxial alignment of siRNA between lipidoid shell bilayers. Langmuir monolayer experiments showed a distinct interaction between the lipidoid headgroups and siRNA, which was dependent on buffer subphase pH. Thermal analyses suggested that PLGA and lipidoid interact, which was evident from a shift in the phase transition temperature of lipidoid, and the thermotropic phase behavior of lipidoid was affected by inclusion of siRNA. ATR-FTIR data confirmed the shift or disappearance of characteristic absorption bands of siRNA after lipidoid binding. In conclusion, siRNA-loaded LPNs display a core-shell structure, wherein the polymeric core functions as a colloid matrix support for siRNA-loaded lipidoid shell layers.

Keywords:  Drug delivery; Lipidoid-polymer hybrid nanoparticles; Lipoplex, nanostructure; Nanomedicine; siRNA

DOI:  https://doi.org/10.1016/j.jcis.2022.11.141
Int J Pharm. 2022 Dec 12. pii: S0378-5173(22)01044-4. [Epub ahead of print] 122489

Engineered Ionizable Lipid Nanoparticles mediated Efficient siRNA Delivery to Macrophages for Anti-inflammatory Treatment of Acute Liver Injury.

Hongqian Zhang, Feng Ding, Zongwei Zhu, Qian Sun, Chuanxu Yang.

  Small interfering RNA (siRNA) mediating specific gene silencing provides a promising strategy for anti-inflammatory therapy. However, the development of potent carriers for anti-inflammatory siRNA to macrophages remains challenging. With the aim of realizing potent delivery of siRNA to macrophages, we engineered ionizable lipid nanoparticles (LNP) with the key component of synthetic lipid-like materials. By varying the amine molecules in the structure of synthetic lipid-like materials, a potent LNP (1O14-LNP) was identified, which exhibited efficient transfection of macrophages by facilitating efficient internalization and endosomal escape. The 1O14-LNP successfully delivered anti-inflammatory siRNA against interleukin-1β (siIL-1β) with more than 90% downregulation of IL-1β expression in LPS-activated macrophages. From in vivo studies, systemic administrated 1O14-LNP/siRNA mainly distributed in liver and efficiently captured by hepatic macrophages without notable sign of toxicity. Furthermore, LPS/D-GalN-induced acute liver injury model treated with 1O14-LNP/siIL-1β resulted in significant suppression of IL-1β expression and amelioration of liver tissue damage. These results demonstrate that the engineered ionizable LNP provides a powerful tool for siRNA delivery to macrophages and that the strategy of silencing of pro-inflammatory cytokines holds great potential for treating inflammatory diseases.

Keywords:  endosomal escape; inflammation; lipid nanoparticle; macrophages; siRNA delivery

DOI:  https://doi.org/10.1016/j.ijpharm.2022.122489
ACS Appl Mater Interfaces. 2022 Dec 16.

Excipient-Free Ionizable Polyester Nanoparticles for Lung-Selective and Innate Immune Cell Plasmid DNA and mRNA Transfection.

Atanu Chakraborty, Shruti Dharmaraj, Nhu Truong, Ryan M Pearson.

  Extrahepatic nucleic acid delivery using polymers typically requires the synthesis and purification of custom monomers, post-synthetic modifications, and incorporation of additional excipients to augment their stability, endosomal escape, and in vivo effectiveness. Here, we report the development of a single-component and excipient-free, polyester-based nucleic acid delivery nanoparticle platform comprising ionizable N-methyldiethanolamine (MDET) and various hydrophobic alkyl diols (Cp) that achieves lung-selective nucleic acid transfection in vivo. PolyMDET and polyMDET-Cp polyplexes displayed high serum and enzymatic stability, while delivering pDNA or mRNA to "hard-to-transfect" innate immune cells. PolyMDET-C4 and polyMDET-C6 mediated high protein expression in lung alveolar macrophages and dendritic cells without inducing tissue damage or systemic inflammatory responses. Improved strategies using readily available starting materials to produce a simple, excipient-free, non-viral nucleic acid delivery platform with lung-selective and innate immune cell tropism has the potential to expedite clinical deployment of polymer-based genetic medicines.

Keywords:  excipient-free; extrahepatic; innate immune cells; ionizable polyesters; transfection

DOI:  https://doi.org/10.1021/acsami.2c14424
Extracell Vesicle. 2022 Dec;1 100002

Inhalable exosomes outperform liposomes as mRNA and protein drug carriers to the lung.

Kristen D Popowski, Blanca López de Juan Abad, Arianna George, Dylan Silkstone, Elizabeth Belcher, Jaewook Chung, Asma Ghodsi, Halle Lutz, Jada Davenport, Mallory Flanagan, Jorge Piedrahita, Phuong-Uyen C Dinh, Ke Cheng.

  Respiratory diseases are among the leading causes of morbidity and mortality worldwide, coupled with the ongoing coronavirus disease 2019 (COVID-19) pandemic. mRNA lipid nanoparticle (LNP) vaccines have been developed, but their intramuscular delivery limits pulmonary bioavailability. Inhalation of nanoparticle therapeutics offers localized drug delivery that minimizes off targeted adverse effects and has greater patient compliance. However, LNP platforms require extensive reformulation for inhaled delivery. Lung-derived extracellular vesicles (Lung-Exo) offer a biological nanoparticle alternative that is naturally optimized for mRNA translation and delivery to pulmonary cells. We compared the biodistribution of Lung-Exo against commercially standard biological extracellular vesicles (HEK-Exo) and LNPs (Lipo), where Lung-Exo exhibited superior mRNA and protein cargo distribution to and retention in the bronchioles and parenchyma following nebulization administration. This suggests that inhaled Lung-Exo can deliver mRNA and protein drugs with enhanced pulmonary bioavailability and therapeutic efficacy.

Keywords:  Exosomes; Extracellular vesicles; Lung; Nebulizers

DOI:  https://doi.org/10.1016/j.vesic.2022.100002
Chem Biol Drug Des. 2022 Dec 14.

Down-regulation of MDM2 by small interference RNA induces apoptosis and sensitizes MCF-7 breast cells to resveratrol.

Ning-Xin Zhang, Jun-Feng Ma, Si-Qi Li, Zhe Yin, Li Li.

  Recent studies have demonstrated the mouse double minute gene (MDM2), a main oncogene, as a novel and interesting therapeutic target for cancer therapy. The aim of this study is investigating the involvement of MDM2 in anti-proliferative and anti-metastatic effects of resveratrol in breast cancer cells. MCF-7 cells were transfected with siRNA against MDM2 and resveratrol. Proliferation and apoptosis were evaluated by MTT assay and cell death ELISA assay, respectively. MDM2, p53, Bax, Bcl-2, caspase-3, MMP-2, MMP9 expression were determined by qRT-PCR and western blotting. Transfection with si-MDM2 significantly suppressed the expression of MDM2 expression, resulting in MCF-7 cell growth inhibition and spontaneous apoptosis. Pretreatment with Si-MDM2 synergically increased anti-proliferation and anti-metatstatic effects of resveratrol. No significant anti-cancer effects were detected with negative control siRNA treatment. Our findings suggest that silencing of MDM2 by specific siRNA effectively induce apoptosis and also enhanced anti-cancer effects of resveratrol. Therefore, siMDM2 may be a potent combination in breast therapy.

Keywords:  MDM2; Polyphenols; Resveratrol; breast cancer

DOI:  https://doi.org/10.1111/cbdd.14190
Int J Mol Sci. 2022 Nov 29. pii: 14974. [Epub ahead of print]23(23):

The Impact of Nanobody Density on the Targeting Efficiency of PEGylated Liposomes.

Bárbara S Mesquita, Marcel H A M Fens, Alessia Di Maggio, Esmeralda D C Bosman, Wim E Hennink, Michal Heger, Sabrina Oliveira.

  Nanoparticles (NPs) are commonly modified with tumor-targeting moieties that recognize proteins overexpressed on the extracellular membrane to increase their specific interaction with target cells. Nanobodies (Nbs), the variable domain of heavy chain-only antibodies, are a robust targeting ligand due to their small size, superior stability, and strong binding affinity. For the clinical translation of targeted Nb-NPs, it is essential to understand how the number of Nbs per NP impacts the receptor recognition on cells. To study this, Nbs targeting the hepatocyte growth factor receptor (MET-Nbs) were conjugated to PEGylated liposomes at a density from 20 to 800 per liposome and their targeting efficiency was evaluated in vitro. MET-targeted liposomes (MET-TLs) associated more profoundly with MET-expressing cells than non-targeted liposomes (NTLs). MET-TLs with approximately 150-300 Nbs per liposome exhibited the highest association and specificity towards MET-expressing cells and retained their targeting capacity when pre-incubated with proteins from different sources. Furthermore, a MET-Nb density above 300 Nbs per liposome increased the interaction of MET-TLs with phagocytic cells by 2-fold in ex vivo human blood compared to NTLs. Overall, this study demonstrates that adjusting the MET-Nb density can increase the specificity of NPs towards their intended cellular target and reduce NP interaction with phagocytic cells.

Keywords:  biologicals; cancer treatment; immune cell interactions; nanobodies; nanoparticulate drug delivery systems; protein corona

DOI:  https://doi.org/10.3390/ijms232314974