| Chemical Engineering, Energy Engineering |
|
|
|
|
| Construction and evaluation of gene delivery system based on polyacylthiourea |
Yue WANG( ),Zhu-xian ZHOU,You-qing SHEN*() |
| College of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310027, China |
|
|
|
Abstract N, N-dimethylcysteamine modified PATU (PATU-DMCA) was synthesized through thiol-ene Michael addition reaction based on a novel dendrimer of polyacylthiourea (PATU). NMR results show that the dendrimer structure of PATU-DMCA is correct and integrated. Agarose gel electrophoresis assay and dynamic light scattering results demonstrate that PATU-DMCA of low generations with low molar ratios of N and P can package DNA to form nanoparticles with size of 60~100 nm and Zeta potential of 10~20 mV. In human cervical carcinoma cell HeLa and human lung cancer cell A549, the formed polyplexes with low molar ratios of N and P show higher transfection efficiency in serum-free medium than the classical dendrimer poly (amidoamine), while the cytotoxicity is equal to each other. The subcellular distribution study in HeLa indicate that the intracellular trafficking of the polyplexes include cell membrane adhesion, endocytosis into cell, entering into lysosome, and lysosomal escape via ‘proton sponge effect’ for transfection in cytoplasm and nucleus. Both PATU-DMCA and PATU itself have enormous potential for gene delivery.
|
|
Received: 24 July 2018
Published: 04 March 2019
|
|
|
|
Corresponding Authors:
You-qing SHEN
E-mail: wangyue_sunshine@163.com;shenyq@zju.edu.cn
|
聚酰基硫脲基因输送体系的构建与评价
基于新型树枝状大分子聚酰基硫脲(PATU)，通过巯基-烯Michael加成反应得到表面N, N-二甲基半胱胺修饰的聚酰基硫脲(PATU-DMCA). 核磁结果表明：PATU-DMCA树枝状结构正确且完整. 琼脂糖凝胶电泳及动态光散射粒径测定结果表明：PATU-DMCA在低代数、低氮磷摩尔比条件下可以有效包载DNA，形成粒径为60~100 nm、Zeta电位为10~20 mV的纳米复合物. 在人宫颈癌细胞HeLa和人肺癌细胞A549上，低氮磷摩尔比的PATU-DMCA的无血清转染效率优于经典的树枝状大分子基因载体聚酰胺胺，而两者细胞毒性相当. HeLa上的亚细胞分布结果表明：纳米复合物在细胞水平的转运过程基本包括黏附细胞膜、内吞入胞、进入溶酶体，通过“质子海绵”效应从溶酶体逃逸后进入细胞质及细胞核进行转染. PATU-DMCA以及PATU本身在基因输送中均具有很大的潜力.
关键词：
聚酰基硫脲,
基因输送,
高效合成,
转染效率
|
|
| [1] |
CORNU T I, MUSSOLINO C, CATHOMEN T Refining strategies to translate genome editing to the clinic[J]. Nature Medicine, 2017, 23 (4): 415- 423
doi: 10.1038/nm.4313
|
|
|
| [2] |
NALDINI L Gene therapy returns to centre stage[J]. Nature, 2015, 526 (7573): 351- 360
doi: 10.1038/nature15818
|
|
|
| [3] |
CHENG C J, BAHAL R, BABAR I A, et al MicroRNA silencing for cancer therapy targeted to the tumour microenvironment[J]. Nature, 2015, 518 (7537): 107- 110
doi: 10.1038/nature13905
|
|
|
| [4] |
ZHOU Z X, LIU X R, ZHU D C, et al Nonviral cancer gene therapy: delivery cascade and vector nanoproperty integration[J]. Advanced Drug Delivery Reviews, 2017, 115: 115- 154
doi: 10.1016/j.addr.2017.07.021
|
|
|
| [5] |
LACHELT U, WAGNER E Nucleic acid therapeutics using polyplexes: a journey of 50 years (and beyond)[J]. Chemical Reviews, 2015, 115 (19): 11043- 11078
doi: 10.1021/cr5006793
|
|
|
| [6] |
ZHANG Y, SATTERLEE A, HUANG L In vivo gene delivery by nonviral vectors: overcoming hurdles?[J]. Molecular Therapy, 2012, 20 (7): 1298- 1304
doi: 10.1038/mt.2012.79
|
|
|
| [7] |
KESHARWANI P, LYER A K Recent advances in dendrimer-based nanovectors for tumor-targeted drug and gene delivery[J]. Drug Discovery Today, 2015, 20 (5): 536- 547
doi: 10.1016/j.drudis.2014.12.012
|
|
|
| [8] |
TSCHICHE A, MALHOTRA S, HAAG R Nonviral gene delivery with dendritic self-assembling architectures[J]. Nanomedicine, 2014, 9 (5): 667- 693
doi: 10.2217/nnm.14.32
|
|
|
| [9] |
LUO K, HE B, WU Y, et al Functional and biodegradable dendritic macromolecules with controlled architectures as nontoxic and efficient nanoscale gene vectors[J]. Biotechnology Advances, 2014, 32 (4): 818- 830
doi: 10.1016/j.biotechadv.2013.12.008
|
|
|
| [10] |
HU J J, HU K, CHENG Y Y Tailoring the dendrimer core for efficient gene delivery[J]. Acta Biomaterialia, 2016, 35: 1- 11
doi: 10.1016/j.actbio.2016.02.031
|
|
|
| [11] |
SHCHARBIN D, SHAKHBAZAU A, BRYSZEWSKA M Poly(amidoamine) dendrimer complexes as a platform for gene delivery[J]. Expert Opinion on Drug Delivery, 2013, 10 (12): 1687- 1698
doi: 10.1517/17425247.2013.853661
|
|
|
| [12] |
SHAO S Q, ZHOU Q, SI J X, et al A non-cytotoxic dendrimer with innate and potent anticancer and anti-metastatic activities[J]. Nature Biomedical Engineering, 2017, 1 (9): 745- 757
doi: 10.1038/s41551-017-0130-9
|
|
|
| [13] |
SOWINSKA M, URBANCZYK-LIPKOWSKA Z Advances in the chemistry of dendrimers[J]. New Journal of Chemistry, 2014, 38 (6): 2168- 2203
doi: 10.1039/c3nj01239e
|
|
|
| [14] |
YAMAMOTO K Dendrimer complexes: fine control of metal assembly in macromolecules[J]. Journal of Polymer Science Part a-Polymer Chemistry, 2005, 43 (17): 3719- 3727
doi: 10.1002/(ISSN)1099-0518
|
|
|
| [15] |
DUFES C, UCHEGBU I F, SCHATZLEIN A G Dendrimers in gene delivery[J]. Advanced Drug Delivery Reviews, 2005, 57 (15): 2177- 2202
doi: 10.1016/j.addr.2005.09.017
|
|
|
| [16] |
YANG J P, ZHANG Q, CHANG H, et al Surface-engineered dendrimers in gene delivery[J]. Chemical Reviews, 2015, 115 (11): 5274- 5300
doi: 10.1021/cr500542t
|
|
|
| [17] |
ARIMA H, MOTOYAMA K, HIGASHI T Sugar-appended polyamidoamine dendrimer conjugates with cyclodextrins as cell-specific non-viral vectors[J]. Advanced Drug Delivery Reviews, 2013, 65 (9): 1204- 1214
doi: 10.1016/j.addr.2013.04.001
|
|
|
| [18] |
BARNER-KOWOLLIK C, DU PREZ F E, ESPEEL P, et al "Clicking" polymers or just efficient linking: what is the difference?[J]. Angewandte Chemie-International Edition, 2011, 50 (1): 60- 62
doi: 10.1002/anie.v50.1
|
|
|
| [19] |
KOLB H C, FINN M G, SHARPLESS K B Click chemistry: diverse chemical function from a few good reactions[J]. Angewandte Chemie-International Edition, 2001, 40 (11): 2004- 2021
doi: 10.1002/(ISSN)1521-3773
|
|
|
| [20] |
FELLA C, WALKER G F, OGRIS M, et al Amine-reactive pyridylhydrazone-based PEG reagents for pH-reversible PEI polyplex shielding[J]. European Journal of Pharmaceutical Sciences, 2008, 34 (4-5): 309- 320
doi: 10.1016/j.ejps.2008.05.004
|
|
|
| [21] |
ASAYAMA S, SUDO M, NAGAOKA S, et al Carboxymethyl poly(L-histidine) as a new pH-sensitive polypeptide to enhance polyplex gene delivery[J]. Molecular Pharmaceutics, 2008, 5 (5): 898- 901
doi: 10.1021/mp800094b
|
|
|
| [22] |
PACK D W, HOFFMAN A S, PUN S, et al Design and development of polymers for gene delivery[J]. Nature Reviews Drug Discovery, 2005, 4 (7): 581- 593
doi: 10.1038/nrd1775
|
|
|
| [23] |
ZHU D C, YAN H J, LIU X, et al Intracellularly disintegratable polysulfoniums for efficient gene delivery[J]. Advanced Functional Materials, 2017, 27 (16): 1606826
doi: 10.1002/adfm.201606826
|
|
|
| [24] |
QIU N S, LIU X R, ZHONG Y, et al Esterase-activated charge-reversal polymer for fibroblast-exempt cancer gene therapy[J]. Advanced Materials, 2016, 28 (48): 10613- 10622
doi: 10.1002/adma.201603095
|
|
|
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
| |
Shared |
|
|
|
|
| |
Discussed |
|
|
|
|
