Article Dans Une Revue Gene Therapy Année : 2021

Dystrophin and mini-dystrophin quantification by mass spectrometry in skeletal muscle for gene therapy development in Duchenne muscular dystrophy

Vahid Farrokhi
Jason Walsh
Joe Palandra
  • Fonction : Auteur
Joanne Brodfuehrer
Teresa Caiazzo
Michael Binks
  • Fonction : Auteur
Srividya Neelakantan
Florence Yong
Pinky Dua
  • Fonction : Auteur
Hendrik Neubert

Résumé

Duchenne muscular dystrophy (DMD) is a lethal, degenerative muscle disorder caused by mutations in the DMD gene, leading to severe reduction or absence of the protein dystrophin. Gene therapy strategies that aim to increase expression of a functional dystrophin protein (mini-dystrophin) are under investigation. The ability to accurately quantify dystrophin/mini-dystrophin is essential in assessing the level of gene transduction. We demonstrated the validation and application of a novel peptide immunoaffinity liquid chromatography–tandem mass spectrometry (IA-LC-MS/MS) assay. Data showed that dystrophin expression in Becker muscular dystrophy and DMD tissues, normalized against the mean of non-dystrophic control tissues ( n = 20), was 4–84.5% (mean 32%, n = 20) and 0.4–24.1% (mean 5%, n = 20), respectively. In a DMD rat model, biceps femoris tissue from dystrophin-deficient rats treated with AAV9.hCK.Hopti-Dys3978.spA, an adeno-associated virus vector containing a mini-dystrophin transgene, showed a dose-dependent increase in mini-dystrophin expression at 6 months post-dose, exceeding wildtype dystrophin levels at high doses. Validation data showed that inter- and intra-assay precision were ≤20% (≤25% at the lower limit of quantification [LLOQ]) and inter- and intra-run relative error was within ±20% (±25% at LLOQ). IA-LC-MS/MS accurately quantifies dystrophin/mini-dystrophin in human and preclinical species with sufficient sensitivity for immediate application in preclinical/clinical trials.
Fichier principal
Vignette du fichier
41434_2021_Article_300.pdf (1) Télécharger le fichier
Origine Publication financée par une institution
licence

Dates et versions

hal-04811115 , version 1 (29-11-2024)

Licence

Identifiants

Citer

Vahid Farrokhi, Jason Walsh, Joe Palandra, Joanne Brodfuehrer, Teresa Caiazzo, et al.. Dystrophin and mini-dystrophin quantification by mass spectrometry in skeletal muscle for gene therapy development in Duchenne muscular dystrophy. Gene Therapy, 2021, 29 (10-11), pp.608-615. ⟨10.1038/s41434-021-00300-7⟩. ⟨hal-04811115⟩
17 Consultations
6 Téléchargements

Altmetric

Partager

More