Amino Acid–Substituted Dextran‐Based Non‐Viral Vectors for Gene Delivery

To form bio‐inspired non‐viral vectors for DNA delivery, the polysaccharide dextran is allowed to react with Boc‐amino protected amino acids glycine, β‐alanine, and L‐lysine activated with 1,1’‐carbonyldiimidazole and subsequent dextran ester deprotection. A library of such dextran esters is made available to investigate the relationship between polymer structure, complex formation, stability, toxicity, and transfection. Only dextran esters of β‐alanine and L‐lysine are able to efficiently interact with DNA as shown by dye exclusion assays, to form nanosized complexes (70–110 nm) with positive zeta potential. With increasing substitution degree and complex charge ratios, the L‐lysine esters accomplish more effective binding and protection of DNA against enzymatic degradation than β‐alanine esters. However, luciferase reporter gene assays reveal higher transfection for β‐alanine than for L‐lysine esters due to a more effective DNA release and better suited buffing area of the amino groups triggering the endosomal release. Conclusively, β‐alanine‐substituted dextran derivatives may serve as promising non‐viral vectors.     

Novel Acid‐Labile,Thermoresponsive Poly(methacrylamide)s with Pendent Ortho Ester Moieties

Novel acid‐labile, thermoresponsive methacrylamide‐based (co)polymers with pendent ortho ester groups were prepared by free radical polymerization of N‐(2‐methoxy‐1,3‐dioxan‐5‐yl) methacrylamide (NMM) and N‐(2‐ethoxy‐1,3‐dioxan‐5‐yl)methacrylamide (NEM). These polymers are both thermoresponsive and acid‐sensitive in aqueous solution, which was proved by transmittance measurements, fluorescence, and ¹H NMR spectroscopy. The LCSTs of the (co)polymers were shifted to higher temperature by increasing the content of the more hydrophilic NMM units. All of these polymers can be hydrolyzed under acidic condition and the hydrolysis rate increased with the decrease in the pH value.

     

基于纳米材料的基因载体

基因疗法是治疗基因变异引起的先天性遗传疾病和后天获得性疾病以及癌症的新型有效方法。外源基因在细胞中安全、高效、稳定的表达是基因治疗成功的关键,这与基因治疗所使用的载体系统息息相关。基因载体主要分为病毒载体和非病毒载体两大类:病毒载体的转染效率较高,但副作用较大;非病毒载体作为一种新型的基因传递系统,可以弥补病毒载体的缺陷,尽管其转染效率稍逊于病毒载体,但在基因治疗领域具有不可替代的作用。随着纳米技术的出现和蓬勃发展,基于纳米材料的基因载体研究受到越来越多的关注。纳米基因载体具有如下潜在的优势:它制备相对简单,易于对其进行多功能的修饰;具有良好的生物相容性,一般不会引起强烈的机体免疫反应;粒径普遍很小,容易穿过人体的组织间隙而被细胞吸收,基因转运效率较高;可以较有效保护其所携带外源基因,利于基因更高效地表达。本文主要对基于金属、无机非金属、阳离子聚合物和脂质体纳米材料作为基因载体的研究进展进行综述和展望。     

Amphiphilic Block Copolymers Bearing Ortho Ester Side‐Chains: pH‐Dependent Hydrolysis and Self‐Assembly in Water

A new type of pH‐responsive block copolymer nanoparticle has been synthesized and characterized. The amphiphilic diblock copolymer, PEG‐b‐PMYM, contains acid‐labile ortho ester side‐chains in the hydrophobic block and can self‐assemble into micelle‐like nanoparticles in water at neutral pH. Hydrolysis of the ortho ester side‐chains follows a distinct exocyclic mechanism and shows pH‐dependent kinetics, which triggers changes in nanoparticle size and morphology. The nanoparticles have been found to be non‐toxic to cells in vitro. The ability to tune the size and morphology of biocompatible block copolymer nanoparticles by controlling the pH‐sensitive side‐chain hydrolysis represents a unique approach that may be exploited to improve the efficacy of nanometer‐scale drug delivery.

     

胆固醇修饰的低分子量聚乙烯亚胺接枝化聚[L-天冬酰胺-co-L-赖氨酸]作为基因载体的性能研究

通过将低分子量的聚乙烯亚胺(PEI600)及其胆固醇衍生物与聚(L-天冬酰胺-co-L-赖氨酸)(PSL)进行开环反应, 合成了一类新型的肿瘤靶向基因载体, 研究了这类载体与DNA形成复合物的性质以及介导绿色荧光蛋白质粒pEGFP-C1转染不同细胞的性能. 结果表明, 在复合质量比大于5∶1时, 各载体均能与DNA形成结构稳定的复合物. 同时转染实验结果证明, 通过在侧链引入一定数目的胆固醇, 可以明显提高载体对于癌细胞HepG2和Hela的转染效率. 这类新型的载体具有良好的细胞相容性、较高的转染效率以及易于进行靶向修饰等特点, 在基因治疗研究领域中将具有较好的潜在应用价值.     

Well‐Defined Poly(Ortho Ester Amides) for Potential Drug Carriers: Probing the Effect of Extra‐ and Intracellular Drug Release on Chemotherapeutic Efficacy

To compare the chemotherapeutic efficacy determined by extra‐ and intracellular drug release strategies, poly(ortho ester amide)‐based drug carriers (POEAd‐C) with well‐defined main‐chain lengths, are successfully constructed by a facile method. POEAd‐C3‐doxorubicin (DOX) can be rapidly dissolved to release drug at tumoral extracellular pH (6.5–7.2), while POEAd‐C6‐DOX can rapidly release drug following gradual swelling at intracellular pH (5.0–6.0). In vitro cytotoxicity shows that POEAd‐C3‐DOX exhibits more toxic effect on tumor cells than POEAd‐C6‐DOX at extracellular pH, but POEAd‐C6‐DOX has stronger tumor penetration and inhibition in vitro and in vivo tumor models. So, POEAd‐C6‐DOX with the intracellular drug release strategy has stronger overall chemotherapeutic efficacy than POEAd‐C3‐DOX with extracellular drug release strategy. It is envisioned that these poly(ortho ester amides) can have great potential as drug carriers for efficient chemotherapy with further optimization.

     

Poly(cyclic imino ether)s Beyond 2‐Substituted‐2‐oxazolines

2‐Oxazolines (2‐OZO) are 5‐membered cyclic imino ethers whose cationic ring‐opening polymerization (CROP) mechanism and resulting polymer properties are extensively studied. However, also 6‐ and 7‐membered cyclic imino ethers can be polymerized via CROP. Together with the much less studied 4‐ and 5‐substituted main‐chain chiral poly(2‐oxazoline)s (P‐2‐OZO), these compounds are interesting monomers to enhance the versatility of (co)poly(cyclic imino ether)s. To emphasize the potential of such alternative cyclic imino ether monomers, we provide an overview on the polymerizations of 2‐oxazine (2‐OZI) and chiral 4‐ and 5‐substituted 2‐OZO as well as of selected properties of the resulting polymers. In addition, the hydrolysis of these polymers into the corresponding poly(alkylene imine)s will be addressed.

     

Crosslinked Redox‐Responsive Micelles Based on Lipoic Acid‐Derived Amphiphiles for Enhanced siRNA Delivery

Successful application of gene silencing approaches critically depends on systems that are able to safely and efficiently deliver genetic material such as small interfering RNA (siRNA). Due to their beneficial well‐defined dendritic nanostructure, self‐assembling dendrimers are emerging as promising nanovectors for siRNA delivery. However, these kinds of vectors are plagued with stability issues, especially when considered for in vivo applications. Therefore, in the present study, disulfide‐based temporarily fixed micelles are developed that can degrade upon reductive conditions, and thus lead to efficient cargo release. In detail, lipoic acid‐derived crosslinked micelles are synthesized based on small polymerizable dendritic amphiphiles. Particularly, one candidate out of this series is able to efficiently release siRNA due to its redox‐responsive biodegradable profile when exposed to simulated intracellular environments. As a result, the reduction‐triggered disassembly leads to potent gene silencing. In contrast, noncrosslinkable, structurally related constructs fails under the tested assay conditions, thereby confirming the applied rational design approach and demonstrating its large potential for future in vivo applications.

     

Isomerization polymerization and copolymerization of glycidyl alkanoates catalyzed by methylaluminum bis(2,6‐di‐t‐butyl‐4‐methylphenoxide)

The isomerization polymerizations of glycidyl propionate (1b), octanoate (1c), and stearate (1d) with methylaluminum bis(2,6‐di‐tert‐butyl‐4‐methylphenoxide) (3) were investigated. The polymerizations selectively gave poly(2‐alkyl‐1,3‐dioxolane‐4,2‐diyloxymethylene)s (2), although the polymer yield as well as the polymer molecular weight significantly decreased as the acyl chain of 1 was lengthened. These polymers readily hydrolyzed to glycerin and the corresponding fatty acids under mild conditions. The copolymerizations of glycidyl acetate (1a) with these monomers were also examined. In any combination, the composition of the obtained copolymer was essentially identical with the feed ratio, while both copolymer yield and molecular weight decreased as the feed of 1a was decreased. © 1999 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 37: 435–444, 1999 (See graphics.)     

Biodegradable Coumaric Acid‐Based Poly(anhydride‐ester) Synthesis and Subsequent Controlled Release

To overcome drug delivery issues associated with its short half‐life in vivo, p‐coumaric acid (pCA), a naturally occurring bioactive, has been chemically incorporated into a poly(anhydride‐ester) backbone through solution polymerization. Nuclear magnetic resonance and Fourier transform infrared spectroscopies indicated that pCA was successfully incorporated without noticeable alterations in structural integrity. The polymer's weight‐average molecular weight and thermal properties were determined, exhibiting a molecular weight of over 26 000 Da and a glass transition temperature of 57 °C. In addition, in vitro hydrolytic release studies demonstrated pCA release over 30 d with maintained antioxidant activity, demonstrating the polymer's potential as a controlled release system.

     

Poly(ethylene glycol)‐block‐Poly(glycidyl methacrylate) with Oligoamine Side Chains as Efficient Gene Vectors

Well‐defined diblock copolymers, poly(ethylene glycol)‐block‐poly(glycidyl methacrylate)s (PEG‐b‐PGMAs), with different poly(glycidyl methacrylate) (PGMA) chains, were prepared via atom transfer radical polymerization (ATRP) from the same macromolecular initiator 2‐bromoisobutyryl‐terminated poly(ethylene glycol) (PEG). Ethyldiamine (EDA), diethylenetriamine (DETA), triethylenetetramine (TETA), and polyethyleneimine (PEI) with an of 400 (PEI₄₀₀) were used to decorate PEG‐b‐PGMAs to get the cationic polymers PEG‐b‐PGMA‐ oligoamines. These cationic polymers possessed high buffer capability and could condense plasmid DNA (pDNA) into nanoscaled complexes of 125–530 nm. These complexes showed the positive zeta potential of 20–35 mV at N/P ratios of 10–50. Most of them exhibited very low cytotoxicity and good transfection efficiency in 293T cells. The presence of the serum medium did not decrease the transfection efficiency due to the steric stabilization of the PEG chains.

     

Acid‐Labile Acyclic Cucurbit[n]uril Molecular Containers for Controlled Release

Stimuli‐responsive molecular containers are of great importance for controlled drug delivery and other biomedical applications. A new type of acid labile acyclic cucurbit[n ]uril (CB[n ]) molecular containers is presented that can degrade and release the encapsulated cargo at accelerated rates under mildly acidic conditions (pH 5.5–6.5). These containers retain the excellent recognition properties of CB[n ]‐type hosts. A cell culture study demonstrated that the cellular uptake of cargos could be fine‐tuned by complexation with different containers. The release and cell uptake of cargo dye was promoted by acidic pH.     

pH‐ and Sugar‐Induced Shape Memory Hydrogel Based on Reversible Phenylboronic Acid–Diol Ester Bonds

A simple strategy is provided to construct a novel pH‐ and sugar‐induced shape memory hydrogel based on dynamic phenylboronic acid (PBA)–diol interactions formed by PBA‐modified sodium alginate (Alg‐PBA) and poly(vinyl alcohol) (PVA). The dynamic PBA–diol ester bonds serve as temporary cross‐links and stabilize the deformed shape of the hydrogel. The disassociation of the PBA–diol ester bonds is explored in acidic conditions and aqueous solutions of glucose and fructose, which endow the hydrogel with shape memory performances.

     

Monodisperse Poly(lactide‐co‐glycolic acid)‐Based Nanocarriers for Gene Transfection

This contribution describes a simple, aerosol‐based method for fabricating monodisperse particles containing mixtures of poly(lactide‐co‐glycolic acid) [PLGA], protamine sulfate (Prot), and poly(l‐ lysine) [PLL] as nanocarriers for gene transfection. Aqueous solutions of PLGA, Prot, and PLL were collison‐atomized, and the resulting aerosolized droplets were dried “on the fly” to form solid particles, which then were electrostatically size‐classified into 50, 100, and 200 nm mobility diameter samples. Measurements of cell viability and transfection reveal that the fabricated nanocarriers have a lower cytotoxicity (>85% in cell viability) and a higher transfection efficiency [>8.7 × 10⁵ in relative light units (RLU) mg⁻¹] than does 25 kDa polyethyleneimine (≈50% and 6.8 × 10⁵ RLU mg⁻¹).     

Structure–Function Correlations of Poly(Amido Amine)s for Gene Delivery

Poly(amido amine)s' (PAAs) versatility are nearly unique among stepwise polymers. Different functional groups can be easily introduced into these polymers to add functionality such as cell internalization, charge‐shift, bioreducibility, “stealth” properties, and targeting moieties, while maintaining the bulk structural integrity of these polymers. The poly(amido amine)s are used as a unique research platform to elucidate their complex structure–function relationship. It is shown that guanidinium group, carboxyl group, disulfide bond, alkyl chain, branching, acetyl groups, benzoyl groups, and quaternary nicotinamide moieties can influence many steps of gene delivery, such as DNA condensation, cellular uptake, endosomal escape, nuclear entry, and finally gene expression. The authors systematically discuss the structure–function correlations of PAAs for gene delivery, and elaborate how the properties of polymers can be adjusted by changing the polymeric structure.

     

DOPC‐Detergent Conjugates: Fusogenic Carriers for Improved In Vitro and In Vivo Gene Delivery

Phospholipid‐detergent conjugates are proposed as fusogenic carriers for gene delivery. Eleven compounds are prepared and their properties are investigated. The ability of the conjugates to promote fusion with a negatively charged model membrane is determined. Their DNA delivery efficiency and cytotoxicity are assessed in vitro. Lipoplexes are administered in the mouse lung, and transgene expression Indeterminate inflammatory activity are measured. The results show that conjugation of 1,2‐dioleoyl‐sn‐glycero‐3‐phosphocholine (DOPC) with C₁₂E₄ produces a carrier that can efficiently deliver DNA to cells, with negligible ­associated toxicity. Fusogenicity of the conjugates shows good correlation with in vitro transfection efficiency and crucially depends on the length of the polyether moiety of the detergent. Finally, DOPC‐C₁₂E₄ reveals highly potent for in vivo DNA delivery and favorably compares to GL67A, the current golden standard for gene delivery to the airway, opening the way for further promising developments.