Browsing by Author "Grinberg, Daniel"
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- Splicing therapeutics for patients affected by lysosomal storage disordersPublication . Matos, Liliana; Gonçalves, Vânia; Canals, Isaac; Jordan, Peter; Grinberg, Daniel; Pérez, Belén; Prata, Maria João; Alves, SandraIn this study, we have used a modified U1 snRNA that completely matches the splice donor site of HGSNAT gene exon 2, which corrected the effect of the common 5’ splice site mutation c.234+1G>A in Mucopolysaccharidosis IIIC. In another approach using an antisense oligonucleotide (AO) we have succeeded in the correction of the c.66G>A splicing mutation in CSTB gene (Unverricht–Lundborg disease). Besides that, we have performed the functional analysis of some IDS gene splicing mutations (Mucopolysaccharidosis II) and used AOs to exploit an alternative therapy for one of those mutations (c.1122C>T on exon 8).
- Therapeutic strategies based on modified U1 snRNAs and chaperones for Sanfilippo C splicing mutationsPublication . Matos, Liliana; Canals, Isaac; Dridi, Labri; Choi, Yoo; Prata, Maria Joâo; Jordan, Peter; Desviat, Lourdes R.; Perez, Belén; Pshezhetsky, A.V.; Grinberg, Daniel; Alves, Sandra; Vilageliu, LluisaMutations affecting RNA splicing represent more than 20% of the mutant alleles in Sanfilippo syndrome type C, a rare lysosomal storage disorder that causes severe neurodegeneration. Many of these mutations are localized in the conserved donor or acceptor splice sites, while few are found in the nearby nucleotides. METHODS: In this study we tested several therapeutic approaches specifically designed for different splicing mutations depending on how the mutations affect mRNA processing. For three mutations that affect the donor site (c.234 + 1G > A, c.633 + 1G > A and c.1542 + 4dupA), different modified U1 snRNAs recognizing the mutated donor sites, have been developed in an attempt to rescue the normal splicing process. For another mutation that affects an acceptor splice site (c.372-2A > G) and gives rise to a protein lacking four amino acids, a competitive inhibitor of the HGSNAT protein, glucosamine, was tested as a pharmacological chaperone to correct the aberrant folding and to restore the normal trafficking of the protein to the lysosome. RESULTS: Partial correction of c.234 + 1G > A mutation was achieved with a modified U1 snRNA that completely matches the splice donor site suggesting that these molecules may have a therapeutic potential for some splicing mutations. Furthermore, the importance of the splice site sequence context is highlighted as a key factor in the success of this type of therapy. Additionally, glucosamine treatment resulted in an increase in the enzymatic activity, indicating a partial recovery of the correct folding. CONCLUSIONS: We have assayed two therapeutic strategies for different splicing mutations with promising results for the future applications