siRNA Knocking Down in HepG2 Cells Identifies PFKFB4 and HNF4α as Key Genes Important for Cancer Cell Survival.

IF 3.8 4区医学 Q2 GENETICS & HEREDITY Current gene therapy Pub Date : 2025-01-17 DOI:10.2174/0115665232337735250101121115

Amer Imraish, Walhan Alshaer, Tuqa Abu-Thiab, Fatima Enaya, Hamzeh J Al-Ameer, Afnan Al-Hunaiti, Mohammad Abumurad, Malek Zihlif

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Abstract

Introduction: Liposomes are versatile delivery systems for encapsulating small interfering RNAs (siRNAs) because they enhance cellular uptake and gene silencing. This study compares the new liposome formula to commercial lipofectamine in delivering siRNAs targeting hepatic carcinoma genes, focusing on HNF4-α and PFKFB4.

Methods: Flow cytometry and confocal microscopy revealed efficient internalization of PE-Rhod- B labeled lipoplexes in HepG2 cells, while cytotoxicity assays demonstrated significant reductions in cell viability, particularly with siHNF4-α and siPFKFB4.

Results: The newly formulated liposomes showed superior efficacy, achieving nearly 93% cytotoxicity at 100 nM, compared to just 50% with lipofectamine at the same concentration. Furthermore, real-time PCR confirmed that the liposome-encapsulated siHNF4-α reduced HNF4-α mRNA expression by tenfold at 100 nM, compared to a twofold reduction with lipofectamine at 200 nM. Similarly, siPFKFB4 delivered via liposomes showed a dose-dependent 35-fold reduction in PFKFB4 mRNA expression at 100 nM, outperforming the maximum reduction achieved by lipofectamine. The IC50 values for all siRNA treatment groups were significantly lower when using the liposome formula, reflecting improved delivery efficiency.

Conclusion: These results demonstrate the potential of liposome formulations for therapeutic siRNA delivery. The encapsulation enhances cellular uptake and gene silencing efficiency, making the liposome formula a promising candidate for targeted gene therapy in hepatic carcinoma. Further research should explore it's in vivo biodistribution and potential combination therapies.

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HepG2细胞siRNA敲低鉴定PFKFB4和HNF4α是影响癌细胞存活的关键基因

脂质体是封装小干扰rna （sirna）的通用递送系统，因为它们增强细胞摄取和基因沉默。本研究比较了新脂质体配方与商用脂质体在递送靶向肝癌基因的sirna方面的差异，重点是HNF4-α和PFKFB4。方法：流式细胞术和共聚焦显微镜显示PE-Rhod- B标记的脂质体在HepG2细胞中有效内化，而细胞毒性实验显示细胞活力显著降低，特别是siHNF4-α和siPFKFB4。结果：新配制的脂质体表现出优越的功效，在100 nM下达到近93%的细胞毒性，而相同浓度的脂质体仅为50%。此外，实时PCR证实脂质体包裹的siHNF4-α在100 nM时使HNF4-α mRNA的表达减少了10倍，而脂质体在200 nM时使HNF4-α mRNA的表达减少了2倍。同样，通过脂质体递送的siPFKFB4在100 nM时显示PFKFB4 mRNA表达的剂量依赖性降低了35倍，优于脂质体所达到的最大降幅。使用脂质体配方时，所有siRNA处理组的IC50值均显著降低，反映了递送效率的提高。结论：这些结果证明了脂质体制剂治疗siRNA递送的潜力。脂质体的包封增强了细胞摄取和基因沉默效率，使其成为肝癌靶向基因治疗的有希望的候选药物。进一步的研究应探讨其在体内的生物分布和潜在的联合治疗。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Current gene therapy 医学-遗传学

CiteScore

6.70

自引率

2.80%

发文量

期刊介绍： Current Gene Therapy is a bi-monthly peer-reviewed journal aimed at academic and industrial scientists with an interest in major topics concerning basic research and clinical applications of gene and cell therapy of diseases. Cell therapy manuscripts can also include application in diseases when cells have been genetically modified. Current Gene Therapy publishes full-length/mini reviews and original research on the latest developments in gene transfer and gene expression analysis, vector development, cellular genetic engineering, animal models and human clinical applications of gene and cell therapy for the treatment of diseases. Current Gene Therapy publishes reviews and original research containing experimental data on gene and cell therapy. The journal also includes manuscripts on technological advances, ethical and regulatory considerations of gene and cell therapy. Reviews should provide the reader with a comprehensive assessment of any area of experimental biology applied to molecular medicine that is not only of significance within a particular field of gene therapy and cell therapy but also of interest to investigators in other fields. Authors are encouraged to provide their own assessment and vision for future advances. Reviews are also welcome on late breaking discoveries on which substantial literature has not yet been amassed. Such reviews provide a forum for sharply focused topics of recent experimental investigations in gene therapy primarily to make these results accessible to both clinical and basic researchers. Manuscripts containing experimental data should be original data, not previously published.