聚酰胺-胺的合成及在水处理中的应用

聚酰胺-胺(PAMAM)树枝状大分子作为一类三维的、分子尺寸和构型高度可控的高分子聚合物,因其具有无毒高效、易于修饰等优点,已在工业水处理领域引起了广泛的关注。本文介绍了PAMAM的主要合成路线及工艺优缺点,综述了近年来PAMAM及其改性衍生物在工业水处理中应用研究进展,分析其作用机理及影响因素,最后对未来的研究方向作以展望。     

How Far Are Non-Viral Vectors to Come of Age and Reach Clinical Translation in Gene Therapy?

Efficient delivery of genetic material into cells is a critical process to translate gene therapy into clinical practice. In this sense, the increased knowledge acquired during past years in the molecular biology and nanotechnology fields has contributed to the development of different kinds of non-viral vector systems as a promising alternative to virus-based gene delivery counterparts. Consequently, the development of non-viral vectors has gained attention, and nowadays, gene delivery mediated by these systems is considered as the cornerstone of modern gene therapy due to relevant advantages such as low toxicity, poor immunogenicity and high packing capacity. However, despite these relevant advantages, non-viral vectors have been poorly translated into clinical success. This review addresses some critical issues that need to be considered for clinical practice application of non-viral vectors in mainstream medicine, such as efficiency, biocompatibility, long-lasting effect, route of administration, design of experimental condition or commercialization process. In addition, potential strategies for overcoming main hurdles are also addressed. Overall, this review aims to raise awareness among the scientific community and help researchers gain knowledge in the design of safe and efficient non-viral gene delivery systems for clinical applications to progress in the gene therapy field.     

Dendritic polyamines: simple access to new materials with defined treelike structures for application in nonviral gene delivery

Polycationic dendrimers are interesting nonviral vectors for in vitro DNA delivery. We describe a simple approach to the synthesis of dendritic polyamines with different molecular weights and adjustable flexibility (degrees of branching; DB). Both parameters influence the transfection efficiency and the cell toxicity of the polymer. Functionalization of hyperbranched polyethylenimine (PEI) by a two-step procedure generated fully branched pseudodendrimers (analogues of polypropylenimine (PPI) and polyamidoamine (PAMAM) dendrimers). The DNA transfection efficiencies observed for these polymers depended on the cell line investigated. The highest efficiencies were observed for polymers whose unfunctionalized PEI cores had molecular weights in the range M(w)=6000-25 000 g mol(-1). The cytotoxicity of the dendrimers generally rises with increasing core size. The data collected for NIH/3T3 and COS-7 cells indicate a maximum transfection efficiency at around 60 % branching for the PPI analogues, and at a PEI-core molecular weight of M(w)=25 000 g mol(-1). PAMAM functionalization of PEI (M(w)=5000 and 21 000 g mol(-1)) leads to polymers with little or no cytotoxity in the cell lines investigated.     

Nanotechnology-Based Strategies to Overcome Current Barriers in Gene Delivery

Nanomaterials are currently being developed for the specific cell/tissue/organ delivery of genetic material. Nanomaterials are considered as non-viral vectors for gene therapy use. However, there are several requirements for developing a device small enough to become an efficient gene-delivery tool. Considering that the non-viral vectors tested so far show very low efficiency of gene delivery, there is a need to develop nanotechnology-based strategies to overcome current barriers in gene delivery. Selected nanostructures can incorporate several genetic materials, such as plasmid DNA, mRNA, and siRNA. In the field of nanotechnologies, there are still some limitations yet to be resolved for their use as gene delivery systems, such as potential toxicity and low transfection efficiency. Undeniably, novel properties at the nanoscale are essential to overcome these limitations. In this paper, we will explore the latest advances in nanotechnology in the gene delivery field.     

PAMAM树状大分子负载和释放阿霉素的耗散粒子动力学模拟

采用耗散粒子动力学模拟方法研究了药物输送载体聚酰胺-胺（PAMAM）树状大分子对抗癌药物阿霉素（DOX）的负载和释放行为。构建了PAMAM树状大分子的粗粒化模型,该模型能准确地重现树状大分子的构象性质。考察了PAMAM树状大分子代数（G）对DOX负载以及pH环境对DOX释放的影响。模拟结果表明,PAMAM树状大分子主要通过疏水作用将DOX包封于内部空腔,G6和G7 PAMAM树状大分子的负载能力较强,因为其孔隙率较高,内部有更多的疏水空腔。在低pH环境下,PAMAM树状大分子结构发生变化,DOX分子能快速地从其中释放,主要原因是PAMAM的伯胺、叔胺和DOX伯胺发生质子化,质子化基团间的静电排斥作用使得PAMAM树状大分子发生溶胀,导致其内部空腔暴露,促进了DOX的释放。本工作可以为基于树状大分子的药物输送体系的设计和优化提供参考。     

聚酰胺-胺型多聚物作为基因载体的研究现状

近年来,新型非病毒载体在基因治疗中应用广泛,特别是树枝状阳离子多聚物,以其低毒性和特殊的结构特性,成为人们研究的重点。其文对聚酰胺。胺树枝状阳离子多聚物（PAMAM dendrimers）的结构特点、性能、基因转移机制以及近年来在基因治疗中的应用进行论述,并对它作为基因治疗载体的应用前景进行了预测。     

Dendrimeric Guanidinoneomycin for Cellular Delivery of Bio‐macromolecules

We present the synthesis of polymeric amino‐ and guanidinoglycosides prepared by tethering neomycin and guanidinoneomycin to PAMAM dendrimers of generations 2 and 4. The ability of these conjugates to promote cellular uptake of high‐molecular‐weight cargo is discussed, together with their cytotoxicity and mechanisms of entry. We demonstrate that the presence of multiple guanidinoneomycin carriers on the PAMAM surface plays an important role in promoting cellular uptake of the dendrimers, maintaining the heparan sulfate specificity and negligible cytotoxicity typical of monomeric guanidinoglycoside molecular transporters.     

Gene Therapy in HIV‐Infected Cells to Decrease Viral Impact by Using an Alternative Delivery Method

The ability of dendrimer 2G‐[Si{O(CH₂)₂N(Me)₂⁺(CH₂)₂NMe₃⁺(I^?)₂}]₈ (NN16) to transfect a wide range of cell types, as well as the possible biomedical application in direct or indirect inhibition of HIV replication, was investigated. Cells implicated in HIV infection such as primary peripheral blood mononuclear cells (PBMC) and immortalized suspension cells (lymphocytes), primary macrophages and dendritic cells, and immortalized adherent cells (astrocytes and trophoblasts) were analyzed. Dendrimer toxicity was evaluated by mitochondrial activity, cell membrane rupture, release of lactate dehydrogenase, erythrocyte hemolysis, and the effect on global gene expression profiles using whole‐genome human microarrays. Cellular uptake of genetic material was determined using flow cytometry and confocal microscopy. Transfection efficiency and gene knockdown was investigated using dendrimer‐delivered antisense oligonucleotides and small interfering RNA (siRNA). Very little cytotoxicity was detected in a variety of cells relevant to HIV infection and erythrocytes after NN16 dendrimer treatment. Imaging of cellular uptake showed high transfection efficiency of genetic material in all cells tested. Interestingly, NN16 further enhanced the reduction of HIV protein 24 antigen release by antisense oligonucleotides due to improved transfection efficiency. Finally, the dendrimer complexed with siRNA exhibited therapeutic potential by specifically inhibiting cyclooxygenase‐2 gene expression in HIV‐infected nervous system cells. NN16 dendrimers demonstrated the ability to transfect genetic material into a vast array of cells relevant to HIV pathology, combining high efficacy with low toxicity. These results suggest that NN16 dendrimers have the potential to be used as a versatile non‐viral vector for gene therapy against HIV infection.     

Aging process of polyamidoamine dendrimers: Effect of pH and shaking in the fluorescence emission and aggregation-state

In the last years, it has been discovered and intensely studied the non-traditional intrinsic luminescence of polyamidoamine (PAMAM) dendrimers. Nevertheless, their aging process in aqueous suspension is scarcely studied, being unknown the causes of the changes in their luminescence properties. Hence, this work aims to characterize the amine-terminated (DG4.0) and carboxylic acid-terminated (DG4.5) PAMAM dendrimers of generations 4.0 and 4.5, respectively, through the aging process at three different pH conditions, stored with or without shaking. The UV–Vis absorption, the fluorescence emission, and the dendrimer-size distribution are studied for up to 16 days. In a different way than the already published works, this work demonstrates that there is no chemical change in dendrimers through the aging process, even though changes in fluorescence emission are observed. Besides, the changes in the agglomeration patterns of dendrimers are not related to the change in the fluorescence emission through aging. Moreover, large aggregates of DG4.5 are present in water and need to be disrupted by shaking before an in vivo administration.     

Development of Covalent Chitosan-Polyethylenimine Derivatives as Gene Delivery Vehicle: Synthesis,Characterization, and Evaluation

There is an increasing interest in cationic polymers as important constituents of non-viral gene delivery vectors. In the present study, we developed a versatile synthetic route for the production of covalent polymeric conjugates consisting of water-soluble depolymerized chitosan (dCS; M_W 6–9 kDa) and low molecular weight polyethylenimine (PEI; 2.5 kDa linear, 1.8 kDa branched). dCS-PEI derivatives were evaluated based on their physicochemical properties, including purity, covalent bonding, solubility in aqueous media, ability for DNA condensation, and colloidal stability of the resulting polyplexes. They were complexed with non-integrating DNA vectors coding for reporter genes by simple admixing and assessed in vitro using liver-derived HuH-7 cells for their transfection efficiency and cytotoxicity. Using a rational screening cascade, a lead compound was selected (dCS-Suc-LPEI-14) displaying the best balance of biocompatibility, cytotoxicity, and transfection efficiency. Scale-up and in vivo evaluation in wild-type mice allowed for a direct comparison with a commercially available non-viral delivery vector (in vivo-jetPEI). Hepatic expression of the reporter gene luciferase resulted in liver-specific bioluminescence, upon intrabiliary infusion of the chitosan-based polyplexes, which exceeded the signal of the in vivo jetPEI reference formulation by a factor of 10. We conclude that the novel chitosan-derivative dCS-Suc-LPEI-14 shows promise and potential as an efficient polymeric conjugate for non-viral in vivo gene therapy.     

Carbon nanohorns functionalized with polyamidoamine dendrimers as efficient biocarrier materials for gene therapy

Carbon nanohorns are suitable platforms for use in biological applications. Their high surface areas allow the incorporation of molecular entities, such as polyamidoamine dendrimers. In this work, we report the synthesis, structural characterization and biological data of new hybrid systems of carbon nanohorns that hold polyamidoamine dendrimers. One of these derivatives has been employed as an agent for gene delivery. The system is able to release interfering genetic material diminishing the levels of a house-keeping protein and a protein directly involved in prostate cancer development. Importantly, this hybrid material is also far less toxic than the corresponding free dendrimer. These results allow us to conclude that these nanomaterials can be exploited as useful non-viral agents for gene therapy.     

PAMAM Dendrimers Mediate siRNA Delivery to Target Hsp27 and Produce Potent Antiproliferative Effects on Prostate Cancer Cells

RNA interference (RNAi) holds great promise for the treatment of inherited and acquired diseases, provided that safe and efficient delivery systems are available. Herein we report that structurally flexible triethanolamine (TEA) core PAMAM dendrimers are able to deliver an Hsp27 siRNA effectively into prostate cancer (PC‐3) cells by forming stable nanoparticles with siRNA, protecting the siRNA nanoparticles from enzymatic degradation, and enhancing cellular uptake of siRNA. The Hsp27 siRNA resulted in potent and specific gene silencing of heat‐shock protein 27, an attractive therapeutic target in castrate‐resistant prostate cancer. Silencing of the hsp27 gene led to induction of caspase‐3/7‐dependent apoptosis and inhibition of PC‐3 cell growth in vitro. In addition, the siRNA–dendrimer complexes are non‐cytotoxic under the conditions used for siRNA delivery. Altogether, TEA core PAMAM dendrimer‐mediated siRNA delivery, in combination with RNAi that specifically targets Hsp27, may constitute a promising approach for combating castrate‐resistant prostate cancer, for which there is no efficacious treatment.     

多肽型阳离子脂质体的研究进展

多肽型阳离子脂质体作为一种重要的非病毒基因载体,克服了其他阳离子脂质体转染效率低、具有一定细胞毒性的缺点,在基因转运方面具有广阔的应用前景。在近年来多肽型阳离子脂质体研究的基础上,论述了多肽型阳离子脂质体在类脂的结构设计、脂质体制备及其在基因转运方面应用的研究现状,展望了多肽型阳离子脂质体的发展方向。     

聚甲基丙烯酸N,N-二甲氨基乙酯及其衍生物作为非病毒基因载体的研究进展

综述了聚甲基丙烯酸N,N-二甲氨基乙酯(PDMAEMA)及其衍生物作为非病毒基因载体的研究进展,包括PDMAEMA及其衍生物相对分子质量及结构与转染效率之间的关系、亲水性聚合物修饰的PDMAEMA和低相对分子质量的PDMAEMA接枝共聚物,并展望了PDMAEMA及其衍生物作为非病毒基因载体的发展方向。     

Perylenediimide-cored dendrimers and their bioimaging and gene delivery applications

In the past few years, perylenediimide-cored (PDI-cored) dendrimers have been successfully prepared and applied in various fields. In this review, we focus on the structural design and synthesis strategies of PDI-cored dendrimers that involve convergent and divergent synthesis approaches. When building the outer shell of PDI-cored dendrimers, first hydrophobic and then hydrophilic macromolecules are introduced. The advantages of water-soluble PDI-cored dendrimers include strong red fluorescence, excellent photostability, low cytotoxicity, high quantum yield, and versatile surface modification. The biological application of water-soluble PDI-cored dendrimers is broad, including fluorescence live-cell imaging, fluorescent labeling, and gene delivery, owing to the rapid development of these materials in the past decade. The challenges and outlooks in this field will also be discussed.     

树枝状大分子用于药物载体的研究进展

树枝状大分子是近年来出现的一类新型合成纳米高分子,具有可控的三维高度支化结构、表面分布着大量的功能基团和单分散等特点,使其在生物医学等领域中日益受到广泛关注。作为一种新型非生物载体,树枝状大分子内部空腔和表面功能基团均可与药物复合,在药物输送和基因转运等方面具有广阔的应用前景。     

Efficient Delivery of Plasmid DNA Using Cholesterol-Based Cationic Lipids Containing Polyamines and Ether Linkages

Cationic liposomes are broadly used as non-viral vectors to deliver genetic materials that can be used to treat various diseases including cancer. To circumvent problems associated with cationic liposome-mediated delivery systems such as low transfection efficiency and serum-induced inhibition, cholesterol-based cationic lipids have been synthesized that resist the effects of serum. The introduction of an ether-type linkage and extension of the aminopropyl head group on the cholesterol backbone increased the transfection efficiency and DNA binding affinity compared to a carbamoyl-type linkage and a mono aminopropyl head group, respectively. Under optimal conditions, each liposome formulation showed higher transfection efficiency in AGS and Huh-7 cells than commercially available cationic liposomes, particularly in the presence of serum. The following molecular structures were found to have a positive effect on transfection properties: (i) extended aminopropyl head groups for a strong binding affinity to plasmid DNA; (ii) an ether linkage that favors electrostatic binding to plasmid DNA; and (iii) a cholesterol backbone for serum resistance.     

Current Status of Gene Therapy Research in Polyglutamine Spinocerebellar Ataxias

Polyglutamine spinocerebellar ataxias (PolyQ SCAs) are a group of 6 rare autosomal dominant diseases, which arise from an abnormal CAG repeat expansion in the coding region of their causative gene. These neurodegenerative ataxic disorders are characterized by progressive cerebellar degeneration, which translates into progressive ataxia, the main clinical feature, often accompanied by oculomotor deficits and dysarthria. Currently, PolyQ SCAs treatment is limited only to symptomatic mitigation, and no therapy is available to stop or delay the disease progression, which culminates with death. Over the last years, many promising gene therapy approaches were investigated in preclinical studies and could lead to a future treatment to stop or delay the disease development. Here, we summed up the most promising of these therapies, categorizing them in gene augmentation therapy, gene silencing strategies, and gene edition approaches. While several of the reviewed strategies are promising, there is still a gap from the preclinical results obtained and their translation to clinical studies. However, there is an increase in the number of approved gene therapies, as well as a constant development in their safety and efficacy profiles. Thus, it is expected that in a near future some of the promising strategies reviewed here could be tested in a clinical setting and if successful provide hope for SCAs patients.     

A PEG-based biocompatible block catiomer with high buffering capacity for the construction of polyplex micelles showing efficient gene transfer toward primary cells

Nonviral gene vectors from synthetic catiomers (polyplexes) are a promising alternative to viral vectors. In particular, many recent efforts have been devoted to the construction of biocompatible polyplexes for in vivo nonviral gene therapy. A promising approach in this regard is the use of poly(ethylene glycol) (PEG)-based block catiomers, which form a nanoscaled core-shell polyplex with biocompatible PEG palisades. In this study, a series of PEG-based block catiomers with different amine functionalities were newly prepared by a simple and affordable synthetic procedure based on an aminolysis reaction, and their utility as gene carriers was investigated. This study revealed that the block catiomers carrying the ethylenediamine unit at the side chain are capable of efficient and less toxic transfection even toward primary cells, highlighting critical structural factors of the cationic units in the construction of polyplex-type gene vectors. Moreover, the availability of the polyplex micelle for transfection with primary osteoblasts will facilitate its use for bone regeneration in vivo mediated by nonviral gene transfection.     

A review of the developments of characteristics of PEI derivatives for gene delivery applications

Redox‐active stimuli have gained a great deal of interest as an indicating factor for designing bioresponsive matrices in gene delivery. Hence, a wide range of gene carriers has been designed incorporating the redox‐stimuli characteristics. The most important type of gene carriers is the class of redox responsive polymers. Among them, disulfide incorporated redox‐responsive polyethyleneimine (PEI) and its derivatives, as a result of their outstanding DNA entrapping characteristics and their intrinsic endosomolytic activity, have attracted considerable attention in recent studies. The review presents the main developments of the characteristics of PEI derivatives and their applications in gene delivery. It is found that despite the uniquely stated characteristics, the noncleavable structure of conventional PEI (high molecular weight PEI: 25k), which makes it a nondegradable material, as well as the frequent inclusion of positively charged amino groups, which reduces its blood circulation period, render conventional PEI a very toxic material for gene‐delivery applications. The extremely high cellular toxicity of conventional PEI has restricted its administration for real in‐vivo physiological media. Recent studies have shown that employing low molecular weight PEI cross‐linked by disulfide linkages (SS‐PEI) and assembling low molecular weight disulfide linkages PEI (LMW SS‐PEI) with bio‐detachable anionic groups were two successful approaches for increasing bioavailability of the PEI‐based gene carriers, while keeping outstanding cellular transfection. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42096.