Abstract：
Conventional gene therapy involving supplementation only treats loss-of-function diseases and is limited by viral packaging sizes, precluding therapy of large genes. The discovery of CRISPR/Cas has led to a paradigm shift in the field of genetic therapy, with the promise of precise gene editing, thus broadening the range of diseases that can be treated. The initial uses of CRISPR/Cas have focused mainly on gene editing or silencing of abnormal variants via utilising Cas endonuclease to trigger the target cell endogenous non-homologous end joining. Subsequently, the technology has evolved to modify the Cas enzyme and even its guide RNA, leading to more efficient editing tools in the form of base and prime editing. Further advancements of this CRISPR/Cas technology itself have expanded its functional repertoire from targeted editing to programmable transactivation, shifting the therapeutic focus to precise endogenous gene activation or upregulation with the potential for epigenetic modifications. In vivo experiments using this platform have demonstrated the potential of CRISPR-activators (CRISPRa) to treat various loss-of-function diseases, as well as in regenerative medicine, highlighting their versatility to overcome limitations associated with conventional strategies. This review summarises the molecular mechanisms of CRISPRa platforms, the current applications of this technology in vivo, and discusses potential solutions to translational hurdles for this therapy, with a focus on ophthalmic diseases.
摘要：
涉及补充的常规基因治疗仅治疗功能丧失的疾病，并且受到病毒包装大小的限制。排除大基因的治疗。CRISPR/Cas的发现导致了基因治疗领域的范式转变，有了精确基因编辑的承诺，从而扩大了可以治疗的疾病范围。CRISPR/Cas的最初用途主要集中在通过利用Cas内切核酸酶来触发靶细胞内源性非同源末端连接的异常变体的基因编辑或沉默。随后，该技术已经进化到修饰Cas酶，甚至它的指导RNA，导致更有效的编辑工具的形式的基础和主要编辑。这种CRISPR/Cas技术本身的进一步发展已经将其功能库从有针对性的编辑扩展到可编程的反式激活，将治疗重点转移到具有表观遗传修饰潜力的精确内源性基因激活或上调。使用该平台的体内实验已经证明了CRISPR激活剂（CRISPRa）治疗各种功能丧失疾病的潜力，以及再生医学，强调它们的多功能性，以克服与传统策略相关的限制。这篇综述总结了CRISPRa平台的分子机制，这项技术目前在体内的应用，并讨论了这种疗法的转化障碍的潜在解决方案，专注于眼科疾病。
