Isogenic GAA-KO Murine Muscle Cell Lines Mimicking Severe Pompe Mutations as Preclinical Models for the Screening of Potential Gene Therapy Strategies
Pompe disease (PD) is a rare disorder caused by mutations in the acid alpha-glucosidase (GAA) gene. Most gene therapies (GT) partially rely on the cross-correction of unmodified cells through the uptake of the GAA enzyme secreted by corrected cells. In the present study, we generated isogenic murine...
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Published in | International journal of molecular sciences Vol. 23; no. 11; p. 6298 |
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Main Authors | , , , , , , , , , , , , |
Format | Journal Article |
Language | English |
Published |
Switzerland
MDPI AG
04.06.2022
MDPI |
Subjects | |
Online Access | Get full text |
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Summary: | Pompe disease (PD) is a rare disorder caused by mutations in the acid alpha-glucosidase (GAA) gene. Most gene therapies (GT) partially rely on the cross-correction of unmodified cells through the uptake of the GAA enzyme secreted by corrected cells. In the present study, we generated isogenic murine GAA-KO cell lines resembling severe mutations from Pompe patients. All of the generated GAA-KO cells lacked GAA activity and presented an increased autophagy and increased glycogen content by means of myotube differentiation as well as the downregulation of mannose 6-phosphate receptors (CI-MPRs), validating them as models for PD. Additionally, different chimeric murine GAA proteins (IFG, IFLG and 2G) were designed with the aim to improve their therapeutic activity. Phenotypic rescue analyses using lentiviral vectors point to IFG chimera as the best candidate in restoring GAA activity, normalising the autophagic marker p62 and surface levels of CI-MPRs. Interestingly, in vivo administration of liver-directed AAVs expressing the chimeras further confirmed the good behaviour of IFG, achieving cross-correction in heart tissue. In summary, we generated different isogenic murine muscle cell lines mimicking the severe PD phenotype, as well as validating their applicability as preclinical models in order to reduce animal experimentation. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Current address: Hospital del Mar- Parc de Salut MAR, 08003 Barcelona, Spain. Current address: Department of Neurology, Columbia University Medical Center, New York, NY 10032, USA. These authors contributed equally to the work. Current address: Comecer, Via Maestri del Lavoro 90, 48014 Castel Bolognese, Italy. |
ISSN: | 1422-0067 1661-6596 1422-0067 |
DOI: | 10.3390/ijms23116298 |