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Hydrophobe-substituted bPEI derivatives: boosting transfection on primary vascular cells

疏水基取代的分枝状聚乙酰亚胺衍生物:促进原发性血管细胞中的转染

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Abstract

Gene therapy targeted to vascular cells represents a promising approach for prevention and treatment of pathological conditions such as intimal hyperplasia, in-stent and post-angioplasty restenosis. In this context, polymeric non-viral gene delivery systems are a safe alternative to viral vectors but a further improvement in efficiency and cytocompatibility is needed to improve their clinical success. Herein, a library of 24 branched polyethylenimine (bPEI) derivatives modified with hydrophobic moieties was synthesised, characterised and tested in vitro on primary vascular cells, aiming to identify delivery agents with superior transfection efficiency and low cytotoxicity. Low molecular weight PEIs (0.6, 1.2 and 2 kDa) were grafted with long (C18) and short (C3) aliphatic chains, featuring different unsaturation degrees and degrees of substitution. 0.6 kDa bPEI-based derivatives were generally ineffective in transfection on vascular smooth muscle cells (VSMCs), while among the other derivatives some promising vectors were identified. Forcing polyplexes on the cell surface by means of centrifugation invariably boosted transfection levels but increased cytotoxicity as well. Of note, a propionyl-substituted derivative (PEI2-PrA1, C3:0) was the most effective on both VSMCs and endothelial cells (ECs), with higher and more sustained gene expression in combination with markedly lower cytotoxicity with respect to the gold standard 25 kDa bPEI. In addition, a linoleoyl-substituted derivative (PEI1.2-LA6, C18:2) owing to its high efficiency in VSMCs and relative inefficacy in ECs, combined with tolerable cytotoxicity was proposed as a vector for specific VSMCs targeting.

摘要

针对血管细胞的基因治疗代表了一种有望用于预防和治疗内膜增生、血管支架狭窄和血管成形术后狭窄等病理状态的方法. 聚合 物非病毒载体的基因传递系统可以安全替代病毒载体, 但是为了提高临床效果, 它们的治疗效率及细胞相容性还需要进一步改善. 本文合 成了一系列24种被疏水基团修饰的分枝状聚乙酰亚胺衍生物(bPEI), 并进行了表征及在体外原发性血管细胞内的测试, 旨在筛选出具有优 异的转染效率和低细胞毒性的传递剂. 低分子量的聚乙酰亚胺(0.6, 1.2 and 2 kDa)以不同取代程度接枝上了不同饱和度的长(C18)和短(C3) 的不饱和脂肪链. 丙酰取代衍生物(PEI2-PrA1, C3:0)在血管平滑肌细胞和内皮细胞转染中是最有效的, 与著名的黄金标准25 kDa bPEI相比, 具有更优异、更持久的基因表达, 且毒性更低. 此外, 亚油酰基取代衍生物(PEI1.2-LA6, C18:2) 由于在血管平滑肌细胞转染过程中效率高, 而在内皮细胞中相对无效, 且其具有可容忍的细胞毒性, 可作为特定靶向于血管平滑肌细胞的载体.

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Acknowledgments

Pezzoli D and Tsekoura EK were awarded a postdoctoral and doctoral scholarship, respectively, from the NSERC CREATE Programin Regenerative Medicine, www.ncprm.ulaval.ca. The studies were financially supported by the Natural Science and Engineering Research Council of Canada, (Discovery Grant to Uludağ H and Mantovani D), the Canadian Institute for Health Research (Operating grant to Uludağ H), and the Fonds de Recherche du Quebec sur les Natures et Technologies (Bilateral Grant to Mantovani D). We thank Dr. Vishwa Somayaji for 1H-NMR analysis of the polymer samples

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Authors and Affiliations

  1. Laboratory for Biomaterials and Bioengineering, CRC- I, Department of Mining, Metallurgical and Materials Engineering and CHU de Québec Research Centre, Laval University, Quebec City, QC, G1V 0A6, Canada

    Daniele Pezzoli & Diego Mantovani

  2. Department of Chemical & Material Engineering, Faculty of Engineering, University of Alberta, Edmonton, AB, T6G 1H9, Canada

    Eleni K. Tsekoura, K.C. Remant Bahadur & Hasan Uludağ

  3. Department of Chemistry, Materials and Chemical Engineering “Giulio Natta”, Politecnico di Milano, Milan, 20131, Italy

    Gabriele Candiani

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  1. Daniele Pezzoli
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  2. Eleni K. Tsekoura
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  3. K.C. Remant Bahadur
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  4. Gabriele Candiani
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  5. Diego Mantovani
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Correspondence to Daniele Pezzoli or Hasan Uludağ.

Additional information

Author contributions Pezzoli D, Uludağ H, Mantovani D and Candiani G conceived the idea and designed the experiments; Pezzoli D, Tsekoura EK and Bahadur KC R performed the experiments; Pezzoli D and Uludağ H analysed the data and wrote the manuscript with support from Candiani G and Mantovani D. All authors contributed to the general discussion.

Conflict of interest Bahadur KCR and Uludağ H hold ownership position in RJH Biosciences Inc. intended to commercialise the described polymers.

Supplementary information Supplementary data are available in the online version of the paper.

Daniele Pezzoli received his PhD degree in Materials Engineering at Politecnico di Milano (Milan, Italy) in 2011, under the supervision of Prof. Gabriele Candiani. He is currently a postdoctoral fellow in the Laboratory for Biomaterials and Bioengineering of Université Laval (Quebec City, Canada), led by Prof. Diego Mantovani. His current research is focused on vascular tissue engineering using collagen gel-based scaffolds and on the development of polymeric non-viral gene delivery systems for tissue engineering applications.

Hasan Uludağ has been with the University of Alberta since 1997, designing functional biomaterials to realize the therapeutic potential of nucleic acids. He obtained dual BSc degrees in Biomedical Engineering and Biology from Brown University (USA) in 1989. He then completed his PhD degree in 1993 at the Department of Chemical Engineering at the University of Toronto. He spent four years at Genetics Institute Inc. (USA), where he contributed to development of a bone-inducing BMP device.

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Pezzoli, D., Tsekoura, E.K., Remant Bahadur, K. et al. Hydrophobe-substituted bPEI derivatives: boosting transfection on primary vascular cells. Sci. China Mater. 60, 529–542 (2017). https://doi.org/10.1007/s40843-017-9030-7

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  • DOI: https://doi.org/10.1007/s40843-017-9030-7

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