Comparison of anti-EGFR-Fab’ conjugated immunoliposomes modified with two different conjugation linkers for siRNA delivery in SMMC-7721 cells

Li Deng,1,* Yingying Zhang,1,* Lulu Ma,1,5,* Xiaolong Jing,1,3 Xingfa Ke,1,3 Jianhao Lian,1,3 Qiang Zhao,1,3 Bo Yan,1,3 Jinfeng Zhang,4 Jianzhong Yao,2 Jianming Chen1 1Department of Pharmaceutical Science, 2Department of Medicinal Chemistry, School of Pharmacy, Second Military Medical University, Shanghai, People’s Republic of China; 3Department of Pharmacy, Fujian University of Traditional Chinese Medicine, Fujian, People’s Republic of China; 4Shanghai TCM Integrated Hospital, Shanghai, People’s Republic of China; 5Department of Pharmacy, First Affiliated Hospital of Bengbu Medical College, Bengbu, People’s Republic of China *These authors contributed equally to this paper Background: Targeted liposome-polycation-DNA complex (LPD), mainly conjugated with antibodies using functionalized PEG derivatives, is an effective nanovector for systemic delivery of small interference RNA (siRNA). However, there are few studies reporting the effect of different conjugation linkers on LPD for gene silencing. To clarify the influence of antibody conjugation linkers on LPD, we prepared two different immunoliposomes to deliver siRNA in which DSPE-PEG-COOH and DSPE-PEG-MAL, the commonly used PEG derivative linkers, were used to conjugate anti-EGFR Fab’ with the liposome. Methods: First, 600 µg of anti-EGFR Fab’ was conjugated with 28.35 µL of a micelle solution containing DSPE-PEG-MAL or DSPE-PEG-COOH, and then post inserted into the prepared LPD. Various liposome parameters, including particle size, zeta potential, stability, and encapsulation efficiency were evaluated, and the targeting ability and gene silencing activity of TLPD-FPC (DSPE-PEG-COOH conjugated with Fab’) was compared with that of TLPD-FPM (DSPE-PEG-MAL conjugated with Fab’) in SMMC-7721 hepatocellular carcinoma cells. Results: There was no significant difference in particle size between the two TLPDs, but the zeta po