Garvan Institute of Medical Research, Sydney, NSW 2010, Australia.
UNSW Sydney, Faculty of Medicine, St Vincent's Clinical School, Sydney, NSW, Australia.
Protein Eng Des Sel. 2022 Feb 17;35. doi: 10.1093/protein/gzac011.
The CRISPR genome editing technology holds great clinical potential for the treatment of monogenetic disorders such as sickle cell disease. The therapeutic in vivo application of the technology relies on targeted delivery methods of the Cas9 and gRNA complex to specific cells or tissues. However, such methods are currently limited to direct organ delivery, preventing clinical application. Here, we show that monoclonal antibodies can be employed to deliver the Cas9/gRNA complex directly into human cells via cell-surface receptors. Using the SpyCatcher/SpyTag system, we conjugated the Fab fragment of the therapeutic antibodies Trastuzumab and Pertuzumab directly to the Cas9 enzyme and observed HER2-specific uptake of the ribonucleoprotein in a human HER2 expressing cell line. Following cellular uptake in the presence of an endosomolytic peptide, modest gene editing was also observed. This finding provides a blueprint for the targeted delivery of the CRISPR technology into specific cells using monoclonal antibodies.
CRISPR 基因组编辑技术在治疗镰状细胞病等单基因疾病方面具有巨大的临床潜力。该技术的治疗性体内应用依赖于 Cas9 和 gRNA 复合物靶向递送到特定细胞或组织的方法。然而，目前这些方法仅限于直接器官递送，阻碍了其临床应用。在这里，我们表明单克隆抗体可以通过细胞表面受体将 Cas9/gRNA 复合物直接递送至人类细胞。我们使用 SpyCatcher/SpyTag 系统将治疗性抗体曲妥珠单抗和帕妥珠单抗的 Fab 片段直接连接到 Cas9 酶上，并观察到在人 HER2 表达细胞系中，该核糖核蛋白具有 HER2 特异性摄取。在存在内溶酶体肽的情况下摄取细胞后，也观察到适度的基因编辑。这一发现为使用单克隆抗体将 CRISPR 技术靶向递送至特定细胞提供了蓝图。
