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Molecular Mechanisms of intracellular delivery of Nanoparticles Monitored by an Enzyme‑Induced Proximity Labeling

Nano-Micro Letters 2024年第6期16卷 14-37页

作者： Junji Ren Zibin Zhang Shuo Geng Yuxi Cheng Huize Han Zhipu Fan Wenbing Dai Hua Zhang Xueqing Wang Qiang Zhang Bing He Department of Pharmaceutics School of Pharmaceutical Sciences Peking University38 Xueyuan RdHaidian DistrictBeijing 100191People’s Republic of China

Achieving increasingly finely targeted drug delivery to organs,tissues,cells,and even to intracellular biomacromolecules is one of the core goals of *** the delivery destination is refined to cellular and subcellular targets,it is essential to explore the delivery of nanomedicines at the molecular ***,due to the lack of technical methods,the molecular mechanism of the intracellular delivery of nanomedicines remains unclear to ***,we develop an enzyme-induced proximity labeling technology in nanoparticles(nano-EPL)for the real-time monitoring of proteins that interact with intracellular ***(lactic-co-glycolic acid)nanoparticles coupled with horseradish peroxidase(HRP)were fabricated as a model(HRP(+)-PNPs)to evaluate the molecular mechanism of nano delivery in *** adding the labeling probe biotin-phenol and the catalytic substrate H_(2)O_(2)at different time points in cellular delivery,nano-EPL technology was validated for the real-time in situ labeling of proteins interacting with ***-EPL achieves the dynamic molecular profiling of 740 proteins to map the intracellular delivery of HRP(+)-PNPs in macrophages over *** on dynamic clustering analysis of these proteins,we further discovered that different organelles,including endosomes,lysosomes,the endoplasmic reticulum,and the Golgi apparatus,are involved in delivery with distinct participation *** importantly,the engagement of these organelles differentially affects the drug delivery efficiency,reflecting the spatial–temporal heterogeneity of nano delivery in *** summary,these findings highlight a significant methodological advance toward understanding the molecular mechanisms involved in the intracellular delivery of nanomedicines.

关键词： Enzyme-induced proximity labeling intracellular delivery Nano-protein interaction Dynamic molecule profiling Macrophages

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Micro and Nanotechnology for intracellular delivery Therapy Protein

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Nano-Micro Letters 2012年第2期4卷 118-123页

作者： Dan Xu Zhenhua Hu Jing Su Fei Wu Weien Yuan School of Pharmacy Shanghai Jiao Tong University

Proteins therapy is of great importance in the treatment of protein deficiency disease. Most human diseases are related to the malfunctioning of one or more proteins. The most effective and direct approach is protein therapy, which delivers the proteins into the target cell to replace the dysfunction protein and maintain the balance of organism. However, clinical application is frequently hampered by various biological barriers to their successful delivery. This review aims to discuss the recent advances about microparticles and nanoparticles fabricated using micro and nanotechnology for intracellular delivery therapy protein and give some suggestions about the promising delivery system.

关键词： intracellular delivery Protein Nanoparticle

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Protein@PP-Zn nanocomplex assembled by coordination of zinc ions used for intracellular protein delivery

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Science China Chemistry 2023年第8期66卷 2354-2362页

作者： Liyun Cui Sainan Liu Fan Wu Hua Chen Yuanfeng Li Linqi Shi Yong Liu Rujiang Ma Key Laboratory of Functional Polymer Materials of Ministry of Education Institute of Polymer ChemistryCollege of ChemistryNankai UniversityTianjin300071China Wenzhou Institute University of Chinese Academy of SciencesWenzhou Key Laboratory of Interdiscipline and Translational MedicineWenzhou325001China Translational Medicine Laboratory the First Affiliated Hospital of Wenzhou Medical UniversityWenzhou325035China

In recent years,intracellular delivery of protein drugs has attracted great attention,and polymer-based systems have been extensively exploited to develop efficient and safe ***,efficient intracellular delivery of protein drugs remains a challenge because of the cell membrane barrier and endosome ***,we report a protein@PP-Zn nanocomplex,which consists of an imidazole-containing block polymer poly(ethylene glycol)-block-poly(β-amino ester)(PEG-b-PAE(Im),PP),zinc ions,and protein drugs,for efficient intracellular protein ***-b-PAE(Im)could conjugate proteins via the bridging effect of zinc ions which simultaneously coordinate with imidazole groups on polymer and electron donor groups,such as imidazole and primary amine groups,on protein to improve the loading stability of *** a slightly acidic environment near cancer cells,the protonation of PAE(Im)backbone increases the positive charge density of the nanocomplex and promotes *** under a more acidic environment in endosomes,further protonation of imidazole groups leads to the disintegration of the nanocomplex and the breakdown of endosomes because of the proton sponge ***,protein is released into the *** the assistance of the nanocomplex,proteins with different sizes and isoelectric points are effectively delivered into *** work provides a stable,efficient and universal strategy for intracellular protein delivery.

关键词： protein zinc ions intracellular delivery coordination polymer

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Combinatorial synthesis of redox-responsive cationic polypeptoids for intracellular protein delivery application

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Science China Chemistry 2020年第11期63卷 1619-1625页

作者： Zhicheng Le Tong Xiao Zhijia Liu Xingliang Liu Hong Liu Lixin Liu Yongming Chen School of Materials Science and Engineering Key Laboratory of Polymeric Composite and Functional Materials of Ministry of EducationGD Research Center for Functional Biomaterials Engineering and TechnologySun Yat-sen UniversityGuangzhou 510275China

Biologics play an essential role in treating various indications from cancers to the metabolic diseases,while the current development of new classes of intracellular-acting protein drugs is still hindered because of high molecular mass and overall hydrophilicity of proteins creating extremely poor permeability across cell ***,there remains an unmet need to develop safe,potent approaches to augment intracellular protein delivery ***,we described a facile multicomponent reaction system for generating a small library of redox-responsive cationic polypeptoids with high *** co-assembly of optimized polymer with protein leads to the formation of compacted nanocomplexes with smaller size and high encapsulation efficiency,thus improving cellular internalization via the macropinocytosis and/or caveolae-mediated endocytosis *** endo-lysosomal escape,the nanocomplexes can be disassociated to efficiently release cargo proteins into the cytosol,owing to the intracellular glutathione(GSH)-triggered rapid cleavage of disulfide bonds in polymers *** a result,we screened a promising platform reagent for efficient cytosolic protein delivery application.

关键词： multicomponent reaction cationic polypeptoid redox-responsive intracellular delivery protein

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Recent electroporation-based systems for intracellular molecule delivery

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Nanotechnology and Precision Engineering 2021年第4期4卷 45-52页

作者： Zaizai Dong Lingqian Chang Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education Beijing Advanced Innovation Center for Biomedical EngineeringSchool of Biological Science and Medical EngineeringBeihang UniversityBeijing 100191China

intracellular delivery of functional molecules,such as DNA probes and plasmids,is an important method for investigating cellular mechanisms and changing cell fates in *** various delivery methods,recent years have seen the emergence of electroporation-based techniques that provide versatile platforms for molecule delivery,with high efficiency and controlled *** this Review,we describe recent electroporation-based systems for intracellular molecule *** principles of electroporation for cell membrane perforation and cargo delivery are briefly *** on various scenarios for the application of electroporation,we review electroporation devices that variously employ structures based on nanochannels,nanostraws,and flow-through microfluidic channels for in vitro intracellular molecule *** also consider in vivo targeted therapies based on delivery of active molecules by electroporation according to the lesion ***,we discuss the current challenges facing electroporation-based techniques,as well as opportunities for their future development,which may lead to innovations in intracellular molecule delivery both for cellular analysis in the laboratory and treatment in the clinic.

关键词： Electroporation intracellular delivery Micro/nanodevice Targeted therapy

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A novel colon cancer targeting mAb FMU198 for intracellular negative gold nanoparticles delivery,cancer diagnosis and treatment

A novel colon cancer targeting mAb FMU198 for intracellular ...

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第九届全国免疫学学术大会

作者： Chaojun Song Li Fan Yongzhan Nie Xiaofeng Huang Boquan Jin Department of Immunology the Fourth Military Medical University

Backgrounds:FMU198 recognizes 79%of colon cancers while it is negative when tested on 24 different healthy human tissues analyzed by immunohistochemistry(IHC).Moreover,FMU198 is rapidly internalized when it binds to colon cancer *** drug carriers have been used widely for drug *** gold(Au)particles can loaddrug with high efficency but are hard to cross the cell membrane due to electrostatic repulsion against negative charge protein on the membrane ***,in our study,FMU198 was prepared for intracellular delivery of negative gold ***:Dynamic laser light scattering(DLS)and transmission electron microscopy(TEM)were employed to determine the characteristics of *** cytometry and confocal fluorescence microscopy were used to demonstrate FMU198 mediated cell-selective intracellular delivery of the Au *** order to demonstrate the targeting efficiency of FMU198 in vivo,bio-imaging study was conducted using in vivo fluorescence imaging system in Colo205tumor bearing nude ***:The size of FMU198 coated Au nanoparticles with spherical morphology is about 20 nm according to the DLS results with negative zeta potential and the result also indicated that the system could be stable both in PBS and medium with *** TEM results revealed that in case of Au-FMU198,the intracellular gold amount was significantly increased in cells compared to Au-isotype which showed very low *** cytometry and confocal fluorescence microscopy showed that the intracellular uptake of negatively charged particle is enhanced to a greater extent using Au-FMU198 as delivery *** vivo bio-imaging study demonstrated that the fluorescence intensity of Au-(Cy5)-FMU198 treated group is8.4-fold higher than that of Au-(Cy5)-isotype treated ones upon 24 h post ***:Therefore,we demonstrated the use of novel tumor targeting molecular FMU198 for intracellular negative gold nanoparticles delivery and t

关键词： colon cancer negative gold nanoparticles intracellular delivery

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A green light-enhanced cytosolic protein delivery platform based on BODIPY-protein interactions

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Nano Research 2023年第1期16卷 1042-1051页

作者： Yang Zhou Yifan Gao Li Pang Weirong Kang Kwan Man Weiping Wang State Key Laboratory of Pharmaceutical Biotechnology The University of Hong KongHong KongChina Department of Pharmacology and Pharmacy Li Ka Shing Faculty of MedicineThe University of Hong KongHong KongChina Dr.Li Dak-Sum Research Centre The University of Hong KongHong KongChina Department of Surgery School of Clinical MedicineHKU-SZH&LKS Faculty of MedicineThe University of Hong KongHong KongChina

Development of cytosolic protein delivery platforms brings new possibilities for various incurable *** based on polymer/protein self-assembly have shown their potential in protein ***,versatile photocontrolled platforms based on self-assembly for protein delivery are seldom ***,we report a boron-dipyrromethene(BODIPY)-modified polyamidoamine(PAMAM)with excellent photo-controllability and efficiency for the cytosolic delivery of various *** serum stability was achieved by coating hyaluronic acid and human serum albumin on the surface of BODIPY-modified PAMAM/protein *** nanoparticles under green light irradiation allowed efficient intracellular delivery of multiple cargo proteins with different charges and molecular weights and promoted endosome *** study provides valuable guidance for the development of BODIPY derivative-based protein delivery systems and advances the research in intracellular protein delivery.

关键词： photoresponsive nanoparticles intracellular delivery protein delivery polyamidoamine(PAMAM) boron-dipyrromethene(BODIPY)

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Long-term live-cell microscopy with labeled nanobodies delivered by laser-induced photoporation

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Nano Research 2020年第2期13卷 485-495页

作者： Jing Liu Tim Hebbrecht Toon Brans Eef Parthoens Saskia Lippens Chengnan Li Herlinde De Keersmaecker Winnok H.De Vos Stefaan C.De Smedt Rabah Boukherroub Jan Gettemans Ranhua Xiong Kevin Braeckmans Laboratory of General Biochemistry and Physical Pharmacy Faculty of PharmacyGhent UniversityGhent B-9000Belgium Department of Biomolecular medicine Faculty of Medicine and Health SciencesGhent UniversityGhent B-9000Belgium VIB-UGent Center for Inflammation Research VIBGhent B-9000Belgium VIB Bioimaging Core Ghent VIBGhent B-9000Belgium Department of Biomedical Molecular Biology Ghent UniversityGhent B-9000Belgium Univ.Lille CNRSCentrale LilleISENUniv.ValenciennesUMR 8520-IEMNLille F-59000Franc Laboratory of Cell Biology and Histology Department of Veterinary SciencesUniversity of Antwerp2020 AntwerpBelgium Centre for Advanced Light Microscopy Ghent UniversityGhent B-9000Belgium

Fluorescence microscopy is the method of choice for studying intracellular ***,its success depends on *** of specific and stable markers.A prominent example of markers that are rapidly gaining interest are nanobodies(Nbs.-15 kDa),which can be functionalized with bright and photostable organic *** to their relatively small size and high specificity,Nbs offer great potential for high-quality long-term subcellular imaging,but suffer from the fact that they cannot spontaneously cross the plasma membrane of live *** have recently discovered that laser-induced photoporation is well suited to deliver extrinsic labels to living cells without compromising their *** a laser-based technology,it is readily compatible with light microscopy and the typical cell recipients used for *** by these promising initial results,we demonstrate here for the first time successful long-term imaging of specific subcellular structures with labeled nanobodies in living *** illustrate this using Nbs that target GFP/YFP-protein constructs accessible in the cytoplasm,actin-bundling protein Fascin,and the histone H2A/H2B *** an efficiency of more than 80%labeled cells and minimal toxicity(-2%),photoporation proved to be an excellent intracellular delivery method for ***-lapse microscopy revealed that cell division rate and migration remained unaffected,confirming excellent cell viability and *** conclude that laser-induced photoporation labeled Nbs can be easily delivered into living cells,laying the foundation for further development of a broad range of Nbs with intracellular targets as a toolbox for long-term live-cell microscopy.

关键词： laser-induced photoporation vapor nanobubble long-term microscopy imaging nanobody intracellular delivery living cell labeling

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Synthesis of Protein Nano-Conjugates for Cancer Therapy

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Nano Research 2011年第5期4卷 425-433页

作者： Jianjun Zhang Juanjuan Du Ming Yan Anandika Dhaliwal Jing Wen Fengqi Liu Tatiana Segura Yunfeng Lu Department of Chemical and Biomolecular Engineering University of California at Los Angeles Los Angeles CA 90095 USA Biomedical Engineering Interdepartmental Program University of California at Los Angeles Los Angeles CA 90095 USA College of Chemistry Jilin University Changchun 130012 China

A eukaryotic cell contains thousands of proteins that regulate its cellular function; delivering functional proteins into cells to rectify cellular functions holds great promise for treatment of various diseases, especially cancers. In this context, ribonuclease （RNase）, an enzyme that breaks down ribonucleic acid （RNA）, has great potential for cancer therapy. However, its therapeutic application is hampered by poor intracellular delivery efficiency and inhibition by ubiquitous intracellular RNase inhibitors. In this work, by designing and synthesizing RNase nano-conjugates by in situ atom transfer radical polymerization （ATRP）, we demonstrate a simple solution to address both challenges. Compared with native RNase, nano-conjugates exhibit significantly enhanced intracellular delivery efficiency, inhibitor resistance, and a near five-fold increase in cytotoxicity. This work provides a novel platform for delivery of therapeutic proteins for cancer therapy and other applications.

关键词： Atom transfer radical polymerization (ATRP) ribonuclease (RNase) RNase inhibitor ribonucleic acid (RNA) cancer therapy intracellular delivery

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Photothermal scaffolds/surfaces for regulation of cell behaviors

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Bioactive Materials 2022年第2期7卷 449-477页

作者： Yangcui Qu Kunyan Lu Yanjun Zheng Chaobo Huang Guannan Wang Yanxia Zhang Qian Yu College of Biomedical Engineering&the Key Laboratory for Medical Functional Nanomaterials Jining Medical UniversityJining272067PR China Joint Laboratory of Advanced Biomedical Materials(NFU-UGent) College of Chemical EngineeringNanjing Forestry UniversityNanjing210037PR China Department of Cardiovascular Surgery of the First Affiliated Hospital&Institute for Cardiovascular Science Soochow UniversitySuzhou215006PR China State and Local Joint Engineering Laboratory for Novel Functional Polymeric Materials College of ChemistryChemical Engineering and Materials ScienceSoochow UniversitySuzhou215123PR China

Regulation of cell behaviors and even cell fates is of great significance in diverse biomedical applications such as cancer treatment,cell-based therapy,and tissue *** the past decades,diverse methods have been developed to regulate cell behaviors such as applying external stimuli,delivering exogenous molecules into cell interior and changing the physicochemical properties of the substrates where cells *** scaffolds/surfaces refer to a kind of materials embedded or coated with photothermal agents that can absorb light with proper wavelength(usually in near infrared region)and convert light energy to heat;the generated heat shows great potential for regulation of cell behaviors in different *** the current review,we summarize the recent research progress,especially over the past decade,of using photothermal scaffolds/surfaces to regulate cell behaviors,which could be further categorized into three types:(i)killing the tumor cells via hyperthermia or thermal ablation,(ii)engineering cells by intracellular delivery of exogenous molecules via photothermal poration of cell membranes,and(iii)releasing a single cell or an intact cell sheet via modulation of surface physicochemical properties in response to *** the end,challenges and perspectives in these areas are commented.

关键词： Photothermal scaffolds Photothermal surfaces Tumor ablation intracellular delivery Cell detachment

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