Antisense Oligonucleotide-Based Therapy in Human Erythropoietic Protoporphyria
| Autor: | Katell Peoc'h, Joëlle Marie, Carole Beaumont, Bernard Grandchamp, Hervé Puy, Zoubida Karim, Caroline Schmitt, Jean-Charles Deybach, Laurent Gouya, Hubert de Verneuil, François Moreau-Gaudry, Arienne Mirmiran, Said Lyoumi, Rima Soaid, Véronique Guyonnet-Dupérat, Vincent Oustric, Sarah Ducamp, Hana Manceau |
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| Rok vydání: | 2014 |
| Předmět: |
Male
Protoporphyria Erythropoietic RNA Splicing Protoporphyrins Biology Cell Line Exon Report Genetics medicine Humans Genetics(clinical) RNA Messenger Allele Genetics (clinical) Messenger RNA Polymorphism Genetic Intron Transfection Oligonucleotides Antisense Ferrochelatase medicine.disease Molecular biology Pedigree RNA splicing biology.protein Female Erythropoietic protoporphyria |
| Zdroj: | The American Journal of Human Genetics. 94:611-617 |
| ISSN: | 0002-9297 |
| DOI: | 10.1016/j.ajhg.2014.02.010 |
| Popis: | In 90% of people with erythropoietic protoporphyria (EPP), the disease results from the inheritance of a common hypomorphic FECH allele, encoding ferrochelatase, in trans to a private deleterious FECH mutation. The activity of the resulting FECH enzyme falls below the critical threshold of 35%, leading to the accumulation of free protoporphyrin IX (PPIX) in bone marrow erythroblasts and in red cells. The mechanism of low expression involves a biallelic polymorphism (c.315−48T>C) localized in intron 3. The 315−48C allele increases usage of the 3′ cryptic splice site between exons 3 and 4, resulting in the transcription of an unstable mRNA with a premature stop codon, reducing the abundance of wild-type FECH mRNA, and finally reducing FECH activity. Through a candidate-sequence approach and an antisense-oligonucleotide-tiling method, we identified a sequence that, when targeted by an antisense oligonucleotide (ASO-V1), prevented usage of the cryptic splice site. In lymphoblastoid cell lines derived from symptomatic EPP subjects, transfection of ASO-V1 reduced the usage of the cryptic splice site and efficiently redirected the splicing of intron 3 toward the physiological acceptor site, thereby increasing the amount of functional FECH mRNA. Moreover, the administration of ASO-V1 into developing human erythroblasts from an overtly EPP subject markedly increased the production of WT FECH mRNA and reduced the accumulation of PPIX to a level similar to that measured in asymptomatic EPP subjects. Thus, EPP is a paradigmatic Mendelian disease in which the in vivo correction of a common single splicing defect would improve the condition of most affected individuals. |
| Databáze: | OpenAIRE |
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