Orthogonal therapy that combines CRISPR-based gene editing and prodrug-based chemotherapy is a promising approach to combat multidrug-resistant cancer. However, its potency to precisely regulate different therapeutic modalities in vivo is limited due to the lack of an integrated platform with high spatiotemporal resolution. Taking advantage of CRISPR technology, a Pt(IV)-based prodrug and orthogonal emissive upconversion nanoparticles (UCNPs), we herein rationally designed the first logic-gated CRISPR-Cas13d-based nanoprodrug for orthogonal photomodulation of gene editing and prodrug release for enhanced cancer therapy. The nanoprodrug (URL) was constructed by encapsulating a green light-activatable Pt(IV) prodrug and UV light-activatable Cas13d gene editing tool into UCNPs. We demonstrated that URL maintained excellent orthogonal emission behaviors under 808 and 980 nm excitations, allowing wavelength-selective photoactivation of Cas13d and the prodrug for downregulation of the resistance-related gene and induction of chemo-photodynamic therapy, respectively. Moreover, the photomodulation superiority of URL for overcoming drug resistance was highlighted by integrating it with a Boolean logic gate for programmable modulation of multiple cell behaviors. Importantly, in vivo studies demonstrated that URL can promote Pt(IV) prodrug activation and ROS generation and massively induce on-target drug accumulation by Cas13d-mediated drug resistance attenuation, delivering an ultimate chemo-photodynamic therapeutic performance in efficiently eradicating primary tumors and preventing further liver metastasis. Collectively, our results suggest that URL expands the Cas13d-based genome editing toolbox into prodrug nanomedicine and accelerates the discovery of new orthogonal therapeutic approaches.

中文翻译：

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一种可正交激活的 CRISPR-Cas13d 纳米前药，可逆转化学抗性以增强化学光动力疗法

结合基于 CRISPR 的基因编辑和基于前体药物的化学疗法的正交疗法是对抗多药耐药癌症的一种很有前途的方法。然而，由于缺乏具有高时空分辨率的集成平台，其在体内精确调节不同治疗方式的能力受到限制。利用 CRISPR 技术，一种基于 Pt( IV ) 的前药和正交发射上转换纳米颗粒 (UCNP)，我们在此合理设计了第一个基于逻辑门控 CRISPR-Cas13d 的纳米前药，用于基因编辑的正交光调制和增强癌症的前药释放治疗。纳米前药 (URL) 是通过封装绿光可激活的 Pt( IV) 前药和 UV 光可激活的 Cas13d 基因编辑工具进入 UCNPs。我们证明了 URL 在 808 和 980 nm 激发下保持出色的正交发射行为，允许 Cas13d 的波长选择性光激活和前药分别用于下调耐药相关基因和诱导化学光动力疗法。此外，通过将 URL 与用于对多细胞行为进行可编程调制的布尔逻辑门集成，突出了 URL 在克服耐药性方面的光调制优势。重要的是，体内研究表明 URL 可以促进 Pt( IV) 前体药物激活和 ROS 生成，并通过 Cas13d 介导的耐药性减弱大量诱导靶向药物积累，在有效根除原发性肿瘤和防止进一步肝转移方面提供最终的化学光动力治疗性能。总的来说，我们的结果表明 URL 将基于 Cas13d 的基因组编辑工具箱扩展为前药纳米药物，并加速了新的正交治疗方法的发现。