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

Chaolan Pan; Chenfei Wang; Yitong Zhao; Tao Bo; Liping Han; Dingjin Yao; Yumeng Wang; Xiaoxiao Wang; Linjing Shi; Anqi Zhao; Qiaoyu Cao; Fuying Chen; Wei He; Ying Ye; Si Zhang; Ming Li

doi:10.1016/j.jconrel.2024.04.030

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

J Control Release. 2024 Apr 23:370:82-94. doi: 10.1016/j.jconrel.2024.04.030. Online ahead of print.

Authors

Chaolan Pan¹, Chenfei Wang², Yitong Zhao³, Tao Bo⁴, Liping Han⁵, Dingjin Yao⁶, Yumeng Wang¹, Xiaoxiao Wang⁷, Linjing Shi⁸, Anqi Zhao¹, Qiaoyu Cao¹, Fuying Chen¹, Wei He¹, Ying Ye¹, Si Zhang⁹, Ming Li¹⁰

Affiliations

¹ Department of Dermatology, Children's Hospital of Fudan University, National Children's Medical Center, Shanghai 201102, China.
² Department of Dermatology, Children's Hospital of Fudan University, National Children's Medical Center, Shanghai 201102, China. Electronic address: wangchenfei@fudan.edu.cn.
³ School of Medicine, Anhui University of Science and Technology, Huainan, Anhui 232000, China.
⁴ NHC Key Laboratory of Glycoconjugate Research, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China.
⁵ NHC Key Laboratory of Glycoconjugate Research, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; Department of Blood Transfusion, Huashan Hospital, Fudan University, Shanghai, China.
⁶ Shanghai EditorGene Technology Co., Ltd, Shanghai, 200000, China.
⁷ Department of Dermatology, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200092, China.
⁸ Department of Dermatology, The Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, China.
⁹ NHC Key Laboratory of Glycoconjugate Research, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China. Electronic address: zhangsi@fudan.edu.cn.
¹⁰ Department of Dermatology, Children's Hospital of Fudan University, National Children's Medical Center, Shanghai 201102, China. Electronic address: mingli@fudan.edu.cn.

PMID: 38643938
DOI: 10.1016/j.jconrel.2024.04.030

Abstract

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 M_w ranging from 7.6 to 131.8 kg/mol and Đ < 2.0 from the one crude HPAE_23.7k, and investigated the expression efficiency for TGM1 and COL7A1 plasmids. Gene transfection results revealed that the intermediate MW HPAEs, HPAE_20.6k, 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.

Keywords: Branched poly(β-amino ester)s; COL7A1 and TGM1 gene expression; Gene therapy; Gene transfection; Stepwise fractionation.