Small promoters capable of driving potent neuron-restricted gene expression are required to support successful brain circuitry and clinical gene therapy studies. However, converting large promoters into functional MiniPromoters, which can be used in vectors with limited capacity, remains challenging. In this study, we describe the generation of a novel version of alphaherpesvirus latency-associated promoter 2 (LAP2), which facilitates precise transgene expression exclusively in the neurons of the mouse brain while minimizing undesired targeting in peripheral tissues. Additionally, we aimed to create a compact neural promoter to facilitate packaging of larger transgenes. Our results revealed that MiniLAP2 (278 bp) drives potent transgene expression in all neurons in the mouse brain, with little to no expression in glial cells. In contrast to the native promoter, MiniLAP2 reduced tropism in the spinal cord and liver. No expression was detected in the kidney or skeletal muscle. In summary, we developed a minimal pan-neuronal promoter that drives specific and robust transgene expression in the mouse brain when delivered intravenously via AAV-PHP.eB vector. The use of this novel MiniPromoter may broaden the range of deliverable therapeutics and improve their safety and efficacy by minimizing the potential for off-target effects.

需要能够驱动有效的神经元限制性基因表达的小型启动子来支持成功的脑回路和临床基因治疗研究。然而，将大型启动子转化为可用于容量有限的载体的功能性微型启动子仍然具有挑战性。在这项研究中，我们描述了一种新版本的α疱疹病毒潜伏相关启动子2（LAP2）的产生，它专门促进小鼠大脑神经元中转基因的精确表达，同时最大限度地减少外周组织中不需要的靶向。此外，我们的目标是创建一个紧凑的神经启动子，以促进更大转基因的包装。我们的结果表明，MiniLAP2 (278 bp) 在小鼠大脑的所有神经元中驱动有效的转基因表达，而在神经胶质细胞中几乎没有表达。与天然启动子相比，MiniLAP2 降低了脊髓和肝脏的向性。在肾脏或骨骼肌中未检测到表达。总之，我们开发了一种最小的泛神经元启动子，当通过 AAV-PHP.eB 载体静脉注射时，该启动子可在小鼠大脑中驱动特异性和稳健的转基因表达。这种新型 MiniPromoter 的使用可以扩大可提供治疗的范围，并通过最大限度地减少脱靶效应的可能性来提高其安全性和有效性。