Gene induction for the treatment of methylmalonic aciduria
| Autor: | Jean Francois Benoist, Ruimei Hu, Joseph P. Sarsero, Jim Vadolas, Leonie R. Wood, Nicole E. Buck, Mahmoud Shekari Khaniani, Heidi Peters, Panos A. Ioannou, Hady Wardan, Lingli Li |
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| Rok vydání: | 2009 |
| Předmět: |
Green Fluorescent Proteins
Drug Evaluation Preclinical Organophosphonates Biology Green fluorescent protein Exon Mutase Drug Discovery Genetics Humans RNA Messenger Molecular Biology Gene Genetics (clinical) Regulation of gene expression Reporter gene Adenine Methylmalonyl-CoA mutase Methylmalonyl-CoA Mutase Molecular biology Gene Expression Regulation Methylmalonic aciduria Biochemistry Molecular Medicine Cisplatin Zidovudine Metabolism Inborn Errors HeLa Cells |
| Zdroj: | The Journal of Gene Medicine. 11:361-369 |
| ISSN: | 1521-2254 1099-498X |
| DOI: | 10.1002/jgm.1297 |
| Popis: | Background Methylmalonic aciduria is an autosomal recessive inborn error of the propionate metabolic pathway. One form of this disorder is caused by mutations in methylmalonyl-coenzyme A mutase (MCM), resulting in reduced levels of enzyme activity. The pharmacological up-regulation of residual mutase activity is one approach to advance treatment strategies for individuals affected by this disorder. We describe the construction, characterization and use of a cellular genomic reporter assay for MCM expression that will potentially identify therapeutic pharmacological agents for methylmalonic aciduria treatment. Methods Homologous recombination was used to insert an enhanced green fluorescent protein (EGFP) cassette inframe before the last codon of exon 13 of the MCM gene (MUT) in a BAC clone. The construct was used to generate stable HeLa cell lines. EGFP expression was measured by flow cytometry and the real-time reverse transcriptase-polymerase chain reaction was used to quantify changes in MUT gene mRNA levels. Results The genomic reporter assay used to screen a selection of compounds. Cisplatin, zidovudine and adefovir were found to increase the levels of MCM mRNA and EGFP expression, providing support for the possible efficacy of these pharmacological compounds in treating methylmalonic aciduria. Conclusions This assay has the potential of being used in high-throughput screening of chemical libraries for the identification of novel compounds that specifically modulate the expression of MCM. Copyright © 2009 John Wiley & Sons, Ltd. |
| Databáze: | OpenAIRE |
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