Superior COL7A1 and TGM1 gene expression in difficult-to-transfect skin cell mediated by highly branched poly(β-amino esters) through stepwise fractionation

• Published in: Journal of controlled release Vol. 370; pp. 82 - 94
• Main Authors: Pan, Chaolan, Wang, Chenfei, Zhao, Yitong, Bo, Tao, Han, Liping, Yao, Dingjin, Wang, Yumeng, Wang, Xiaoxiao, Shi, Linjing, Zhao, Anqi, Cao, Qiaoyu, Chen, Fuying, He, Wei, Ye, Ying, Zhang, Si, Li, Ming
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.06.2024
• Subjects: Branched poly(β-amino ester)s; COL7A1 and TGM1 gene expression; Gene therapy; Gene transfection; Stepwise fractionation; Gene transfection; Branched poly(β-amino ester)s; Gene therapy; COL7A1 and TGM1 gene expression; Stepwise fractionation
• ISSN: 0168-3659; 1873-4995
• DOI: 10.1016/j.jconrel.2024.04.030

Delivering functional gene into targeted skin cells or tissues to modulate the genes expression, has the potential to treat various hereditary cutaneous disorders. Nevertheless, the lack of safe and effective gene delivery vehicles greatly limits the clinical translation of gene therapy for inherited skin diseases. Herein, we developed a facile elution fractionation strategy to isolate eight HPAEs with Mw ranging from 7.6 to 131.8 kg/mol and Đ < 2.0 from the one crude HPAE23.7k, and investigated the expression efficiency for TGM1 and COL7A1 plasmids. Gene transfection results revealed that the intermediate MW HPAEs, HPAE20.6k, exhibited the highest gene transfection efficiency (46.4%) and the strongest mean fluorescence intensity (143,032 RLU), compared to other isolated components and the crude product. Importantly, best-performing isolated HPAE effectively delivered COL7A1 (15,974 bp) and TGM1 (7181 bp) plasmids, promoting the efficient expression of type VII collagen (C7) and transglutaminase-1 proteins in cutaneous cells. Our study establishes a straightforward step-by-step elution fractionation strategy for the development of HPAEs gene delivery vectors, expediting their clinical translation in inherited skin diseases. HPAEs crude polymer is fractionated to eight HPAEs with Mw ranging from 7.6 to 131.8 kg/mol and Đ < 2.0. The best-performing isolated HPAE effectively delivered COL7A1 (15,974 bp) and TGM1 (7181 bp) plasmids, promoting the efficient expression of type VII collagen (C7) and transglutaminase-1 proteins in cutaneous cells. [Display omitted] •We establish a straightforward step-by-step elution fractionation strategy to isolate HPAEs with different molecular weight.•Isolated intermediate molecular weight HPAEs exhibit high transfection efficiency and biocompatibility in diverse cell types.•The top-performing HPAEs efficiently deliver TGM and COL7A1 DNA to potentially rescue hereditary cutaneous disorders.