Dystrophin/mini-dystrophin expression analysis by immunoaffinity liquid chromatography-tandem mass spectrometry after gene therapy for DMD.

IF 4.5 3区 医学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY
Jason Walsh, Joe Palandra, Nicole Duriga, David Beidler, Avery McIntosh, Michael Binks, Hendrik Neubert
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引用次数: 0

Abstract

Adeno-associated virus (AAV)-based gene replacement therapies in Duchenne muscular dystrophy (DMD) aim to restore dystrophin function via the introduction of micro- or mini-dystrophins. We report dystrophin and mini-dystrophin concentrations generated by immunoaffinity liquid chromatography-tandem mass spectrometry (IA-LC-MS/MS) in skeletal muscle biopsies from ambulatory participants with DMD in a phase 1b study of fordadistrogene movaparvovec, an AAV9-based gene replacement construct. The assay performed robustly for 26 months, as demonstrated by limited variability in calibration standards for peptides LLQV (dystrophin and mini-dystrophin) and LEMP (mini-dystrophin only), quality control samples consisting of spiked mini-dystrophin in DMD skeletal muscle lysate, as well as unspiked, pooled, non-dystrophic skeletal muscle lysate (normal pool). Average values for LLQV in the normal pool tested as part of clinical sample and long-term stability runs were similar to validated values. Biopsy samples showed minor or absent LLQV and absent LEMP signals pre-treatment with fordadistrogene movaparvovec infusion, but signals substantially increased at Days 60 and 360, on average. There was strong concordance in LEMP and LLQV expression change between Days 60 and 360 (R2 = 0.91; p < 0.001). IA-LC-MS/MS enables reproducible, stable, and reliable quantification of dystrophin/mini-dystrophin following fordadistrogene movaparvovec infusion. ClinicalTrials.gov identifier: NCT03362502.

免疫亲和液相色谱-串联质谱法分析DMD基因治疗后肌营养不良蛋白/微肌营养不良蛋白的表达。
基于腺相关病毒(AAV)的杜氏肌营养不良症(DMD)基因替代疗法旨在通过引入微或迷你肌营养不良蛋白来恢复肌营养不良蛋白的功能。我们报告了一项基于aav9基因替代构建物fordadistrogene movaparvovec的1b期研究中,通过免疫亲和液相色谱-串联质谱(IA-LC-MS/MS)在DMD患者的骨骼肌活检中产生的肌营养不良蛋白和微肌营养不良蛋白浓度。该实验稳定运行了26个月,证明了肽LLQV(肌营养不良蛋白和迷你肌营养不良蛋白)和LEMP(仅迷你肌营养不良蛋白)校准标准的有限可变性,质量控制样品包括DMD骨骼肌溶解液中加标的迷你肌营养不良蛋白,以及未加标的,池化的,非营养不良骨骼肌溶解液(正常池)。作为临床样本和长期稳定运行的一部分,正常池中LLQV的平均值与验证值相似。活检样本显示轻微或不存在LLQV和不存在LEMP信号,但平均在第60天和360天信号显著增加。60 ~ 360天,LEMP和LLQV表达变化具有较强的一致性(R2 = 0.91;p
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来源期刊
Gene Therapy
Gene Therapy 医学-生化与分子生物学
CiteScore
9.70
自引率
2.00%
发文量
67
审稿时长
4-8 weeks
期刊介绍: Gene Therapy covers both the research and clinical applications of novel therapeutic techniques based on a genetic component. Over the last few decades, significant advances in technologies ranging from identifying novel genetic targets that cause disease through to clinical studies, which show therapeutic benefit, have elevated this multidisciplinary field to the forefront of modern medicine.
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