Balke-Want H, Keerthi V, Cadinanos-Garai A, Fowler C, Gkitsas N, Brown A K, Tunuguntla R, Abou-El-Enein M, Feldman S A
Stanford Center for Cancer Cell Therapy, Stanford Cancer Institute, Stanford University, Stanford, USA.
USC/CHLA Cell Therapy Program, University of Southern California, and Children's Hospital Los Angeles, Los Angeles, USA.
Immunooncol Technol. 2023 Mar 9;18:100375. doi: 10.1016/j.iotech.2023.100375. eCollection 2023 Jun.
Chimeric antigen receptor (CAR) T cell therapy has made significant strides in the treatment of B-cell malignancies, but its application in treating solid tumors still poses significant challenges. Particularly, the widespread use of viral vectors to deliver CAR transgenes into T cells comes with limitations, including high costs and regulatory restrictions, which hinder the translation of novel genetic engineering concepts into clinical applications. Non-viral methods, such as transposon/transposase and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems, offer promising alternatives for stable transgene insertion in CAR-T cells. These methods offer the potential to increase accessibility and efficiency in the development and delivery of CAR-T cell therapies. The main challenge in using non-viral methods, however, is their low knock-in efficiency, which leads to low transgene expression levels. In this review, we discuss recent developments in non-viral approaches for CAR-T cell production, the manufacturing requirements for clinical-grade production of non-viral CAR-T cells, and the adjustments needed in quality control for proper characterization of genomic features and evaluation of potential genotoxicity.
嵌合抗原受体（CAR）T细胞疗法在B细胞恶性肿瘤的治疗方面取得了重大进展，但其在实体瘤治疗中的应用仍然面临重大挑战。特别是，广泛使用病毒载体将CAR转基因导入T细胞存在局限性，包括成本高昂和监管限制，这阻碍了新型基因工程概念转化为临床应用。非病毒方法，如转座子/转座酶和规律成簇间隔短回文重复序列（CRISPR）/Cas系统，为在CAR-T细胞中稳定插入转基因提供了有前景的替代方案。这些方法有可能提高CAR-T细胞疗法开发和递送的可及性和效率。然而，使用非病毒方法的主要挑战是其低敲入效率，这导致转基因表达水平较低。在这篇综述中，我们讨论了CAR-T细胞生产的非病毒方法的最新进展、非病毒CAR-T细胞临床级生产的制造要求，以及在质量控制中为正确表征基因组特征和评估潜在基因毒性所需的调整。
