Antisense oligonucleotide-based therapy in human erythropoietic protoporphyria.

[erythropoietic protoporphyria]

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 .3 15 -48 T >C ) localized in intron 3 . The 315 -48 C 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-V 1 ), prevented usage of the cryptic splice site . In lymphoblastoid cell lines derived from symptomatic EPP subjects , transfection of ASO-V 1 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-V 1 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 .

Diseases
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Diseases presenting "falls below the critical threshold" symptom

erythropoietic protoporphyria

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