In vitro characterization and in vivo expression of human very-long chain acyl-CoA dehydrogenase

J. Lawrence Merritt, Dietrich Matern, Jerry Vockley, Jan Daniels, Tien V. Nguyen, David B. Schowalter

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency is a disorder of fatty acid beta-oxidation that can present at any age with cardiomyopathy, rhabdomyolysis, hepatic dysfunction, and/or nonketotic hypoglycemia. Through the expansion of newborn screening programs an increasing number of individuals with VLCAD deficiency are being identified prior to the onset of symptoms allowing early initiation of therapy. The development of a safe, durable, and effective VLCAD gene delivery system for use at the time of diagnosis could result in a significant improvement in the quality and duration of life for patients with VLCAD deficiency. To this end, we developed a construct containing the human VLCAD cDNA under the control of the strong CMV promoter (pCMV-hVLCAD). A novel rabbit polyclonal anti-VLCAD antibody was prepared using a 24 amino-acid peptide unique to the human VLCAD protein to study human VLCAD expression in immune competent mice. Antibody specificity was demonstrated in Western blots of human VLCAD deficient fibroblasts and in pCMV-hVLCAD transiently transfected VLCAD deficient fibroblasts. Transfected fibroblasts showed correction of the metabolic block as demonstrated by normalization of C14- and C16-acylcarnitine species in cell culture media and restoration of VLCAD activity in cells. Following tail vein injection of pCMV-hVLCAD into mice, we demonstrated expression of hVLCAD in liver. Altogether, these steps are important in the development of a durable gene therapy for VLCAD deficiency.

Original languageEnglish (US)
Pages (from-to)351-358
Number of pages8
JournalMolecular Genetics and Metabolism
Volume88
Issue number4
DOIs
StatePublished - Aug 2006

Fingerprint

human ACADVL protein
Long-Chain Acyl-CoA Dehydrogenase
Fibroblasts
Gene Transfer Techniques
Rhabdomyolysis
Antibody Specificity
Liver
Secondary Prevention
Cardiomyopathies
Hypoglycemia
Genetic Therapy
Culture Media
Tail
Veins
Fatty Acids
Complementary DNA
Cell Culture Techniques
Western Blotting
Quality of Life
Newborn Infant

Keywords

  • Gene therapy
  • Hydrodynamic plasmid injection
  • In vitro correction
  • Very-long chain acyl-CoA dehydrogenase
  • VLCAD antibody

ASJC Scopus subject areas

  • Biochemistry
  • Genetics
  • Endocrinology, Diabetes and Metabolism

Cite this

Merritt, J. L., Matern, D., Vockley, J., Daniels, J., Nguyen, T. V., & Schowalter, D. B. (2006). In vitro characterization and in vivo expression of human very-long chain acyl-CoA dehydrogenase. Molecular Genetics and Metabolism, 88(4), 351-358. https://doi.org/10.1016/j.ymgme.2006.02.010

In vitro characterization and in vivo expression of human very-long chain acyl-CoA dehydrogenase. / Merritt, J. Lawrence; Matern, Dietrich; Vockley, Jerry; Daniels, Jan; Nguyen, Tien V.; Schowalter, David B.

In: Molecular Genetics and Metabolism, Vol. 88, No. 4, 08.2006, p. 351-358.

Research output: Contribution to journalArticle

Merritt, JL, Matern, D, Vockley, J, Daniels, J, Nguyen, TV & Schowalter, DB 2006, 'In vitro characterization and in vivo expression of human very-long chain acyl-CoA dehydrogenase', Molecular Genetics and Metabolism, vol. 88, no. 4, pp. 351-358. https://doi.org/10.1016/j.ymgme.2006.02.010
Merritt, J. Lawrence ; Matern, Dietrich ; Vockley, Jerry ; Daniels, Jan ; Nguyen, Tien V. ; Schowalter, David B. / In vitro characterization and in vivo expression of human very-long chain acyl-CoA dehydrogenase. In: Molecular Genetics and Metabolism. 2006 ; Vol. 88, No. 4. pp. 351-358.
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