X-linked myotubular myopathy (XLMTM) is a severe congenital disease characterized by profound muscle weakness, respiratory failure, and early death. No approved therapy for XLMTM is currently available. Adeno-associated virus (AAV)-mediated gene replacement therapy has shown promise as an investigational therapeutic strategy. We aimed to characterize the transcriptomic changes in muscle biopsies of individuals with XLMTM who received resamirigene bilparvovec (AT132; rAAV8-Des-hMTM1) in the ASPIRO clinical trial and to identify potential biomarkers that correlate with therapeutic outcome. We leveraged RNA-sequencing data from the muscle biopsies of 15 study participants and applied differential expression analysis, gene co-expression analysis, and machine learning to characterize the transcriptomic changes at baseline (pre-dose) and at 24 and 48 weeks after resamirigene bilparvovec dosing. As expected, MTM1 expression levels were significantly increased after dosing (p < 0.0001). Differential expression analysis identified upregulated genes after dosing that were enriched in several pathways, including lipid metabolism and inflammatory response pathways, and downregulated genes were enriched in cell-cell adhesion and muscle development pathways. Genes involved in inflammatory and immune pathways were differentially expressed between participants exhibiting ventilator support reduction of either greater or less than 6 h/day after gene therapy compared to pre-dosing. Co-expression analysis identified similarly regulated genes, which were grouped into modules. Finally, the machine learning model identified five genes, including MTM1, as potential RNA biomarkers to monitor the progress of AAV gene replacement therapy. These findings further extend our understanding of AAV-mediated gene therapy in individuals with XLMTM at the transcriptomic level.

X连锁肌管肌病（XLMTM）是一种严重的先天性疾病，其特征是严重的肌肉无力、呼吸衰竭和过早死亡。目前尚无批准的 XLMTM 疗法。腺相关病毒（AAV）介导的基因替代疗法已显示出作为一种研究性治疗策略的前景。我们的目的是描述在 ASPIRO 临床试验中接受 resamirigene bilparvovec（AT132；rAAV8-Des-hMTM1）的 XLMTM 个体肌肉活检的转录组学变化，并确定与治疗结果相关的潜在生物标志物。我们利用来自 15 名研究参与者肌肉活检的 RNA 测序数据，并应用差异表达分析、基因共表达分析和机器学习来表征基线（给药前）以及 resamirigene bilparvovec 后 24 和 48 周时的转录组变化剂量。正如预期的那样，给药后MTM1表达水平显着增加 (p < 0.0001)。差异表达分析确定了给药后上调的基因在多种途径中富集，包括脂质代谢和炎症反应途径，而下调的基因在细胞间粘附和肌肉发育途径中富集。与给药前相比，基因治疗后每天呼吸机支持减少大于或小于 6 小时的参与者之间参与炎症和免疫途径的基因表达存在差异。共表达分析识别出类似调控的基因，并将其分组为模块。最后，机器学习模型确定了包括MTM1在内的 5 个基因作为潜在的 RNA 生物标志物，用于监测 AAV 基因替代疗法的进展。这些发现进一步扩展了我们在转录组水平上对 AAV 介导的 XLMTM 个体基因治疗的理解。