Princeton Neuroscience Institute, Princeton University, Princeton, NJ 08544, USA.
Princeton Neuroscience Institute, Princeton University, Princeton, NJ 08544, USA.
J Virol Methods. 2023 Apr;314:114688. doi: 10.1016/j.jviromet.2023.114688. Epub 2023 Feb 1.
Adeno-associated virus (AAV) has great potential as a source of treatments for conditions that might respond to potent and ubiquitous transgene expression. However, among its drawbacks, the genetic "payload" of AAV vectors is limited to <4.9 kb and some commonly used gene promoters are sizeable and susceptible to transcriptional silencing. We recently described a short (404 bp), potent, and persistent promoter obtained from the genome of pseudorabies virus (PrV) called alphaherpesvirus latency-associated promoter 2 (LAP2). Here, we evaluated the biodistribution and potency of transgene expression in mouse peripheral tissues in response to local and systemic administration of AAV8-LAP2 and AAV9-LAP2. We found that administration of these vectors resulted in levels of transgene expression that were similar to the larger EF1α promoter. LAP2 drives potent transgene expression in mouse liver and kidney when administered systemically and in skeletal muscle in response to intramuscular delivery. Notably, in skeletal muscle, administration of vectors with LAP2 and EF1α promoters resulted in preferential transduction of myofibers type 2. A direct side-by-side comparison between LAP2 and the EF1α promoter revealed that, despite its smaller size, LAP2 was equally potent to the EF1α promoter and resulted in widespread gene expression after IV and IM administration of AAV8 or AAV9 vectors. Collectively, these findings suggest that constructs that include LAP2 may have the capacity to deliver large therapeutically effective payloads in support of future gene therapy protocols.
腺相关病毒 (AAV) 作为治疗某些疾病的来源具有巨大潜力,这些疾病可能对强效且普遍的转基因表达有反应。然而,AAV 载体的遗传“有效载荷”受到限制,只能达到<4.9 kb,并且一些常用的基因启动子较大,容易受到转录沉默的影响。我们最近描述了一种来自伪狂犬病病毒 (PrV) 的短(404 bp)、有效且持久的启动子,称为α疱疹病毒潜伏相关启动子 2(LAP2)。在这里,我们评估了 AAV8-LAP2 和 AAV9-LAP2 局部和全身给药后在小鼠外周组织中转基因表达的生物分布和效力。我们发现,这些载体的给药导致转基因表达水平与更大的 EF1α 启动子相似。LAP2 在全身给药时可在小鼠肝脏和肾脏中驱动强效的转基因表达,在肌肉内给药时可在骨骼肌中表达。值得注意的是,在骨骼肌中,用 LAP2 和 EF1α 启动子的载体给药导致肌纤维类型 2 的优先转导。LAP2 与 EF1α 启动子的直接并排比较表明,尽管其体积较小,但 LAP2 与 EF1α 启动子同样有效,并且在 IV 和 IM 给予 AAV8 或 AAV9 载体后可导致广泛的基因表达。总之,这些发现表明,包含 LAP2 的构建体可能有能力在未来的基因治疗方案中提供大量有效的治疗有效载荷。