Fallah Ali, Estiri Hajar, Parrish Elizabeth, Soleimani Mansoureh, Zeinali Sirous, Zadeh-Vakili Azita
Department of Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
RNAx Ltd., London, UK.
Cell J. 2020 Jan;21(4):433-443. doi: 10.22074/cellj.2020.6309. Epub 2019 Jul 29.
Tumor necrosis factor-alpha (TNF-α), checkpoint inhibitors, and interleukin-17 (IL-17) are critical targets in inflammation and autoimmune diseases. Monoclonal antibodies (mAbs) have a successful portfolio in the treatment of chronic diseases. With the current progress in stem cells and gene therapy technologies, there is the promise of replacing costly mAbs production in bioreactors with a more direct and cost-effective production method inside the patient's cells. In this paper we examine the results of an investigational assessment of secukinumab gene therapy.
In this experimental study, the DNA sequence of the heavy and light chains of secukinumab antibodies were cloned in a lentiviral vector. Human chorionic villous mesenchymal stem cells (CMSCs) were isolated and characterized. After lentiviral packaging and titration, part of the recombinant viruses was used for transduction of the CMSCs and the other part were applied for systemic gene therapy. The engineered stem cells and recombinant viruses were applied for ex vivo and gene therapy, respectively, in different groups of rat models. and secukinumab expression was confirmed with quantitative real-time polymerase chain reaction (qRT-PCR), western blot, and ELISA by considering the approved secukinumab as the standard reference.
Cell differentiation assays and flow cytometry of standard biomarkers confirmed the multipotency of the CMSCs. Western blot and qRT-PCR confirmed gene expression of secukinumab at both the mRNA and protein level. ELISA testing of serum from treated rat models confirmed mAb overexpression for both and gene therapies.
In this study, a lentiviral-mediated and gene therapy was developed to provide a moderate dose of secukinumab in rat models. Biosimilar gene therapy is an attractive approach for the treatment of autoimmune disorders, cancers and other chronic diseases.
肿瘤坏死因子-α（TNF-α）、检查点抑制剂和白细胞介素-17（IL-17）是炎症和自身免疫性疾病的关键靶点。单克隆抗体（mAb）在慢性病治疗方面成果丰硕。随着干细胞和基因治疗技术的不断进步，有望用一种更直接且具成本效益的患者细胞内生产方法取代生物反应器中昂贵的单克隆抗体制备。本文我们研究了司库奇尤单抗基因治疗的一项研究评估结果。
在本实验研究中，司库奇尤单抗抗体重链和轻链的DNA序列被克隆到慢病毒载体中。分离并鉴定人绒毛膜间充质干细胞（CMSCs）。慢病毒包装和滴定后，一部分重组病毒用于转导CMSCs，另一部分用于全身基因治疗。工程干细胞和重组病毒分别应用于不同组大鼠模型的体外和基因治疗。以获批的司库奇尤单抗为标准参考，通过定量实时聚合酶链反应（qRT-PCR）、蛋白质印迹法和酶联免疫吸附测定（ELISA）确认司库奇尤单抗的表达。
标准生物标志物的细胞分化测定和流式细胞术证实了CMSCs的多能性。蛋白质印迹法和qRT-PCR在mRNA和蛋白质水平均证实了司库奇尤单抗的基因表达。对治疗后大鼠模型血清的ELISA检测证实了两种基因治疗均有单克隆抗体过表达。
在本研究中，开发了一种慢病毒介导的司库奇尤单抗基因治疗方法，以在大鼠模型中提供中等剂量的司库奇尤单抗。生物类似物基因治疗是治疗自身免疫性疾病、癌症和其他慢性病的一种有吸引力的方法。
