概述聚类、间隔规则短回文重复序列（CRISPR）/相关蛋白9（CRISPR/Cas9）作为一种强大的基因组编辑工具正日益崭露头角，能够在活体细胞中对几乎任何基因组序列进行精确和有针对性的修改。这些技术在癌症、代谢性疾病和遗传性疾病等领域具有潜在的治疗应用。然而，一些重大挑战妨碍了其潜力的充分实现。具体而言，无论CRISPR-Cas9基因编辑者是以质粒DNA、mRNA/sgRNA还是核糖核蛋白（RNP）形式传递，其细胞膜渗透性均较差，限制了它们进入细胞内基因组进行编辑的能力。此外，这些编辑者缺乏组织或器官特异性，引发对组织层面上非特异性编辑引起的非预期基因毒性的担忧。尽管已开发出一系列传递载体来传递Cas9编辑者，但它们的有效性往往受到细胞外和细胞内层面的多项障碍的限制。此外，Cas9的长时间活性增加了基因组层面非特异性编辑的风险。因此，开发高效的传递载体以及分子开关以安全地调控Cas9活性至关重要。 在本文中，我们概述了我们最近在开发不同类型的材料方面取得的成就，这些材料可以有效地传递编码Cas9蛋白和单导RNA（sgRNA）的质粒DNA或Cas9 RNP进入细胞，以突显安全有效传递的载体设计考虑因素，并介绍了利用动态二硫化物化学设计可降解聚合物载体的方式，强调了它们在基因组编辑生物大分子的安全有效传递特性，并介绍了将细胞内基因组编辑生物大分子与基于微针的经皮传递相结合，以促进治疗性基因组编辑应用于炎症性皮肤疾病的方法。最后，我们回顾了如何利用光学、化学和遗传开关来控制Cas9活性，并与靶向传递相结合，以解决体内基因编辑的时空特异性，并以癌症和结肠炎治疗为概念验证示例来证明其精确治疗的能力。在最后一部分，我们将总结我们取得的进展，并根据我们自己的研究成果提出未来可能影响该领域的方向。

ConspectusClustered, regularly interspaced, short palindromic repeat (CRISPR)/associated protein 9 (CRISPR/Cas9) is emerging as a powerful genome-editing tool, enabling precise and targeted modifications of virtually any genomic sequence in living cells. These technologies have potential therapeutic applications for cancers, metabolic diseases, and genetic disorders. However, several major challenges hinder the full realization of their potential. Specifically, CRISPR-Cas9 gene editors, whether delivered as plasmid DNA, mRNA/sgRNA, or ribonucleoprotein (RNP), exhibit poor membrane permeability, restricting their access to the intracellular genome, where the editing occurs. Additionally, these editors lack tissue or organ specificity, raising concerns about off-target editing at the tissue level that causes unwanted genotoxicity. Though a range of delivery carriers has been developed to deliver Cas9 editors, their effectiveness is often limited by a number of barriers at both the extracellular and intracellular levels. Moreover, the prolonged activity of Cas9 increases the risk of off-target editing at the genomic level. Therefore, it is crucial to develop efficient delivery vectors, along with molecular switches to safely regulate Cas9 activity.In this Account, we summarize our recent achievements in developing different types of materials that can efficiently deliver the plasmid DNA encoding Cas9 protein and single-guide RNA (sgRNA), or Cas9 RNP into cells to highlight the design considerations of carriers for safe and efficient delivery in vitro and in vivo. After elucidating the chemical and physical factors that are responsible for encapsulating and delivering these biomacromolecules, we further elucidate how we design the biodegradable polymeric carriers using dynamic disulfide chemistry, emphasize their safe and efficient delivery features for genome-editing biomacromolecules, and also introduce the integration of the intracellular delivery of genome-editing biomacromolecules with microneedle-based transdermal delivery to promote therapeutic genome editing for inflammatory skin disorders. Finally, we review how we exploit optical, chemical, and genetic switches to control the Cas9 activity in conjunction with targeted delivery to address the spatiotemporal specificity of gene editing in vivo and demonstrate their precision therapy against cancer and colitis treatment as proof-of-concept examples. In the final part, we will summarize the progress we have made and propose the future directions that may impact the field based on our own research outcomes.