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. 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 , and discusses potential solutions to translational hurdles for this therapy, with a focus on ophthalmic diseases.

中文翻译：

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基因疗法和 CRISPR 激活剂在眼部的潜在治疗应用


涉及补充的传统基因治疗只能治疗功能丧失性疾病，并且受到病毒包装大小的限制，排除了大基因的治疗。 CRISPR/Cas的发现导致了基因治疗领域的范式转变，有望实现精确的基因编辑，从而扩大了可治疗的疾病范围。 CRISPR/Cas的最初用途主要集中在通过利用Cas核酸内切酶触发靶细胞内源性非同源末端连接来进行基因编辑或异常变异沉默。随后，该技术不断发展以修改 Cas 酶甚至其指导 RNA，从而以碱基和引物编辑的形式出现更高效的编辑工具。这种 CRISPR/Cas 技术本身的进一步进步已将其功能库从靶向编辑扩展到可编程反式激活，将治疗重点转向精确的内源基因激活或上调，并具有表观遗传修饰的潜力。使用该平台的实验已经证明了 CRISPR 激活剂 (CRISPRa) 治疗各种功能丧失性疾病以及再生医学的潜力，突显了它们克服传统策略相关局限性的多功能性。这篇综述总结了 CRISPRa 平台的分子机制、该技术的当前应用，并讨论了该疗法转化障碍的潜在解决方案，重点关注眼科疾病。