Maryam Moshref, Jerry Hung-Hao Lo, Andrew McKay, Julien Camperi, Joseph Schroer, Norikiyo Ueno, Shu Wang, Saurabh Gulati, Somayeh Tarighat, Steffen Durinck, Ho Young Lee, Dayue Chen
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引用次数: 0
Abstract
Genetically engineered CD8+ T cells are being explored for the treatment of various cancers. Analytical characterization represents a major challenge in the development of genetically engineered cell therapies, especially assessing the potential off-target editing and product heterogeneity. As conventional sequencing techniques only provide information at the bulk level, they are unable to detect off-target CRISPR translocation or editing events occurring in minor cell subpopulations. In this study, we report the analytical development of a single-cell multi-omics DNA and protein assay to characterize genetically engineered cell products for safety and genotoxicity assessment. We were able to quantify on-target edits, off-target events, and potential translocations at the targeting loci with per-cell granularity, providing important characterization data of the final cell product. Conclusion: A single-cell multi-omics approach provides the resolution required to understand the composition of cellular products and identify critical quality attributes (CQAs).
基因工程 CD8+ T 细胞正被用于治疗各种癌症。分析表征是基因工程细胞疗法开发过程中的一大挑战，尤其是评估潜在的脱靶编辑和产品异质性。由于传统测序技术只能提供大体水平的信息，因此无法检测发生在次要细胞亚群中的脱靶 CRISPR 易位或编辑事件。在本研究中，我们报告了一种单细胞多组学 DNA 和蛋白质检测方法的分析开发情况，该方法用于鉴定基因工程细胞产品的安全性和遗传毒性评估。我们能够量化靶上编辑、脱靶事件，以及每个细胞粒度的靶向基因座的潜在易位，为最终细胞产品提供重要的表征数据。结论单细胞多组学方法提供了了解细胞产品组成和确定关键质量属性 (CQAs) 所需的分辨率。
期刊介绍：
The translation of new discoveries in medicine to clinical routine has never been easy. During the second half of the last century, thanks to the progress in chemistry, biochemistry and pharmacology, we have seen the development and the application of a large number of drugs and devices aimed at the treatment of symptoms, blocking unwanted pathways and, in the case of infectious diseases, fighting the micro-organisms responsible. However, we are facing, today, a dramatic change in the therapeutic approach to pathologies and diseases. Indeed, the challenge of the present and the next decade is to fully restore the physiological status of the diseased organism and to completely regenerate tissue and organs when they are so seriously affected that treatments cannot be limited to the repression of symptoms or to the repair of damage. This is being made possible thanks to the major developments made in basic cell and molecular biology, including stem cell science, growth factor delivery, gene isolation and transfection, the advances in bioengineering and nanotechnology, including development of new biomaterials, biofabrication technologies and use of bioreactors, and the big improvements in diagnostic tools and imaging of cells, tissues and organs.
In today`s world, an enhancement of communication between multidisciplinary experts, together with the promotion of joint projects and close collaborations among scientists, engineers, industry people, regulatory agencies and physicians are absolute requirements for the success of any attempt to develop and clinically apply a new biological therapy or an innovative device involving the collective use of biomaterials, cells and/or bioactive molecules. “Frontiers in Bioengineering and Biotechnology” aspires to be a forum for all people involved in the process by bridging the gap too often existing between a discovery in the basic sciences and its clinical application.
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