Method for treating Usher syndrome and composition thereof

Abstract

The invention relates to a method for targeted editing of target RNA containing G-to-A mutation in a USH2A gene transcript based on a LEAPER technology, comprising the following steps that adenosine deaminase recruitment RNA (arRNA) for editing the target RNA or a construct encoding the arRNA is introduced into cells, the arRNA comprises a complementary RNA sequence hybridized with the target RNA, the arRNA can recruit adenosine deaminaseacting on RNA (ADAR) so as to deaminize target adenosine in the target RNA, in-vivo editing from A to I bases on RNA is safely and effectively carried out, pathogenic mutation sites are repaired, and the purpose of treating diseases such as Usher syndrome is achieved.

Description

technical field [0001] The present application belongs to the field of gene editing therapy, and specifically relates to a method for targeted editing of gene mutations related to Usher Syndrome Type II (Usher Syndrome Type II) based on LEAPER (Leveraging Endogenous ADAR for Programmable Editing on RNA) technology. Background technique [0002] Usher syndrome, also known as hereditary deafness-retinitis pigmentosa syndrome, was described and named by Charles Howard Usher in 1914 12 , is an autosomal recessive rare genetic disease caused by genetic mutations that lead to congenital or gradual loss of vision and hearing. The incidence of Usher syndrome is about 1 / 12500 in Germany 8 , 1 / 28000 in Norway 4 , 1 / 23,000 in the United States, it is estimated that more than half of the 16,000 blind and deaf people in the United States suffer from Usher syndrome 1 . There are tens or even hundreds of thousands of patients around the world who are in dire need of treatment. [0003...

AI Technical Summary

Problems solved by technology

First, the nucleases that need to be expressed exogenously usually have a large molecular weight, which makes their delivery into the body by viral vectors less efficient

Second, due to the exogenous expression of nucleases, there is a potential off-target possibility of nucleases in this method, which will lead to a potential carcinogenic risk in its application

Finally, exogenously expressed nucleases such as Cas9 are found in bacteria rather than naturally occurring in humans or mammals, which makes it possible to elicit an immune response in the patient, which may cause damage to the patient itself, On the other hand, it may also neutralize the exogenously expressed nuclease, thereby losing its proper activity and affecting the therapeutic effect

[0011] In 2017, Professor Zhang Feng and his research group from the Massachusetts Institute of Technology reported an RNA editing technology called REPAIR (RNAEditing for Programmable A to I Replacement), through the exogenous expression of Cas13-ADAR fusion protein and single guide RNA (single guide RNA, sgRNA) can realize the editing from A to I targeting the target RNA, but this method, like CRISPR technology, still requires the expression of foreign proteins

Among these chemical modifications, some are non-natural modifications, which may cause toxicity or immunogenicity of the oligonucleotide; and some modifications may lead to different conformations of the same base chain, so that for the same RNA sequence, there may be Dozens of different conformational combinations, which increase the difficulty of its delivery into cells

[0013] Due to the defects of the above three techniques, and in view of the particularity of the USH2A gene itself, it is difficult for the above three techniques to be used in the treatment of USHER syndrome type Ⅱ

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