DNA 介导的生物响应编辑递送 CRISPR-Cas12a 用于胆固醇调节。
CRISPR-Cas12a delivery by DNA-mediated bioresponsive editing for cholesterol regulation.
机构信息
Department of Bioengineering, University of California, Los Angeles, Los Angeles, CA 90095, USA.
California NanoSystems Institute, University of California, Los Angeles, Los Angeles, CA 90095, USA.
出版信息
Sci Adv. 2020 May 20;6(21):eaba2983. doi: 10.1126/sciadv.aba2983. eCollection 2020 May.
CRISPR-Cas12a represents an efficient tool for genome editing in addition to the extensively investigated CRISPR-Cas9. However, development of efficient nonviral delivery system for CRISPR-Cas12a remains challenging. Here, we demonstrate a DNA nanoclew (NC)-based carrier for delivery of Cas12a/CRISPR RNA (crRNA) ribonucleoprotein (RNP) toward regulating serum cholesterol levels. The DNA NC could efficiently load the Cas12a/crRNA RNP through complementation between the DNA NC and the crRNA. Addition of a cationic polymer layer condensed the DNA-templated core and allowed further coating of a charge reversal polymer layer, which makes the assembly negatively charged under a physiological pH but reverts to positive charge under an acidic environment. When was selected as the target gene because of its important role in regulating the level of serum cholesterol, efficient disruption was observed in vivo (48%), significantly reducing the expression of PCSK9 and gaining the therapeutic benefit of cholesterol control (45% of cholesterol reduction).
CRISPR-Cas12a 除了广泛研究的 CRISPR-Cas9 之外，还是一种有效的基因组编辑工具。然而，开发高效的非病毒递送系统来递送 Cas12a/CRISPR RNA（crRNA）核糖核蛋白（RNP）仍然具有挑战性。在这里，我们展示了一种基于 DNA 纳米笼（NC）的载体，用于递送 Cas12a/crRNA RNP，以调节血清胆固醇水平。DNA NC 可以通过 DNA NC 和 crRNA 之间的互补作用有效地负载 Cas12a/crRNA RNP。添加阳离子聚合物层可以使 DNA 模板化核心凝聚，并允许进一步涂覆电荷反转聚合物层，这使得组装在生理 pH 下带负电荷，但在酸性环境下恢复为正电荷。当 被选为靶基因，因为它在调节血清胆固醇水平方面起着重要作用，观察到体内 的有效破坏 (48%)，显著降低了 PCSK9 的表达，并获得了胆固醇控制的治疗益处 (45%的胆固醇降低)。
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