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Research Article

In-silico Molecular Interaction of Short Synthetic Lipopeptide/Importin-alpha and In-vitro Evaluation of Transgene Expression Mediated by Liposome- Based Gene Carrier

Author(s): Tarwadi Tarwadi, Jalal A. Jazayeri*, Sabar Pambudi, Alfan D. Arbianto, Heni Rachmawati, Rahmana E. Kartasasmita and Sukmadjaja Asyarie

Volume 20, Issue 5, 2020

Page: [383 - 394] Pages: 12

DOI: 10.2174/1566523220666201005104224

Price: $65

Abstract

Background: Lipopeptide-based gene carriers have shown low cytotoxicity, are capable of cell membrane penetration, are easy to manufacture and therefore are great potential candidates for gene delivery applications.

Objectives: This study aims to explore a range of short synthetic lipopeptides, (Lau: Lauryl; Pal: Palmitoyl) consisting of an alkyl chain, one cysteine (C), 1 to 2 histidine (H), and lysine (K) residues by performing in-silico molecular interaction and in-vitro evaluation.

Methods: The molecular interactions between the lipopeptides and Importin-α receptor were performed using AutoDock Vina and Amber14. The lipopeptide/DNA complexes were evaluated in- -vitro for their interactions, particle size, zeta potential and transgene expression. Transfection efficiency of the lipopeptides and Pal-CKKHH-derived liposome was carried out based on luciferase transgene expression.

Results: The in-silico interaction showed that Lau-CKKH and Pal-CKKHH hypothetically expedited nuclear uptake. Both lipopeptides had lower binding energy (-6.3 kcal/mol and -6.2 kcal/mol, respectively), compared to the native ligand, viz, nuclear localization sequence (-5.4 kcal/mol). The short lipopeptides were able to condense DNA molecules and efficiently form compacted nanoparticles. Based on the in-vitro evaluation on COS-7, Pal-CKKHH was found to be the best transfection agent amongst the lipopeptides. Its transfection efficiency (ng Luc/mg total protein) increased up to ~3-fold higher (1163 + 55) as it was formulated with helper lipid DOPE (1:2). The lipopeptide- based liposome (Pal-CKKHH: DOPE=1:2) also facilitated luciferase transgene expression on human embryonic kidney cells (293T) and human cervical adenocarcinoma cells (HeLa) with transfection efficiency 1779 +52 and 260 + 22, respectively.

Conclusion: Our study for the first time has shown that the fully synthesized short lipopeptide Pal- CKKHH is able to interact firmly with the Importin-α. The lipopeptide is able to condense DNA molecules efficiently, facilitate transgene expression, expedite the nuclear uptake process, and hence has the characteristics of a potential transfection agent.

Keywords: Lipopeptide, importin- α, in-silico, liposome, particle size, transfection.

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