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- Sliz, E., et al.
(författare)
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Evidence of a causal effect of genetic tendency to gain muscle mass on uterine leiomyomata
- 2023
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Uterine leiomyomata (UL) are the most common tumours of the female genital tract and the primary cause of surgical removal of the uterus. Genetic factors contribute to UL susceptibility. To add understanding to the heritable genetic risk factors, we conduct a genome-wide association study (GWAS) of UL in up to 426,558 European women from FinnGen and a previous UL meta-GWAS. In addition to the 50 known UL loci, we identify 22 loci that have not been associated with UL in prior studies. UL-associated loci harbour genes enriched for development, growth, and cellular senescence. Of particular interest are the smooth muscle cell differentiation and proliferation-regulating genes functioning on the myocardin-cyclin dependent kinase inhibitor 1A pathway. Our results further suggest that genetic predisposition to increased fat-free mass may be causally related to higher UL risk, underscoring the involvement of altered muscle tissue biology in UL pathophysiology. Overall, our findings add to the understanding of the genetic pathways underlying UL, which may aid in developing novel therapeutics.
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| 7. |
- Aoki, Y, et al.
(författare)
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Fine Tuning of Phosphorothioate Inclusion in 2'-O-Methyl Oligonucleotides Contributes to Specific Cell Targeting for Splice-Switching Modulation
- 2021
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Ingår i: Frontiers in physiology. - : Frontiers Media SA. - 1664-042X. ; 12, s. 689179-
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Tidskriftsartikel (refereegranskat)abstract
- Splice-switching antisense oligonucleotide- (SSO-) mediated correction of framedisrupting mutation-containing premessenger RNA (mRNA) transcripts using exon skipping is a highly promising treatment method for muscular diseases such as Duchenne muscular dystrophy (DMD). Phosphorothioate (PS) chemistry, a commonly used oligonucleotide modification, has been shown to increase the stability of and improve the pharmacokinetics of SSOs. However, the effect of PS inclusion in 2′-O-methyl SSOs (2OMe) on cellular uptake and splice switching is less well-understood. At present, we demonstrate that the modification of PS facilitates the uptake of 2OMe in H2k-mdx myoblasts. Furthermore, we found a dependency of SSO nuclear accumulation and high splice-switching activity on PS inclusion in 2OMe (2OMePS), as tested in various reporter cell lines carrying pLuc/705. Increased exon-inclusion activity was observed in muscle, neuronal, liver, and bone cell lineages via both the gymnotic uptake and lipofection of 2OMePS. Using the photoactivatable ribonucleoside-enhanced crosslinking and a subsequent proteomic approach, we identified several 2OMePS-binding proteins, which are likely to play a role in the trafficking of 2OMePS to the nucleus. Ablation of one of them, Ncl by small-interfering RNA (siRNA) enhanced 2OMePS uptake in C2C12 myoblasts and upregulated luciferase RNA splicing in the HeLa Luc/705 reporter cell line. Overall, we demonstrate that PS inclusion increases nuclear delivery and splice switching in muscle, neuronal, liver, and bone cell lineages and that the modulation of 2OMePS-binding partners may improve SSO delivery.
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| 8. |
- Bazaz, S, et al.
(författare)
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Novel Orthogonally Hydrocarbon-Modified Cell-Penetrating Peptide Nanoparticles Mediate Efficient Delivery of Splice-Switching Antisense Oligonucleotides In Vitro and In Vivo
- 2021
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Ingår i: Biomedicines. - : MDPI AG. - 2227-9059. ; 9:8
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Tidskriftsartikel (refereegranskat)abstract
- Splice-switching therapy with splice-switching oligonucleotides (SSOs) has recently proven to be a clinically applicable strategy for the treatment of several mis-splice disorders. Despite this, wider application of SSOs is severely limited by the inherently poor bioavailability of SSO-based therapeutic compounds. Cell-penetrating peptides (CPPs) are a class of drug delivery systems (DDSs) that have recently gained considerable attention for improving the uptake of various oligonucleotide (ON)-based compounds, including SSOs. One strategy that has been successfully applied to develop effective CPP vectors is the introduction of various lipid modifications into the peptide. Here, we repurpose hydrocarbon-modified amino acids used in peptide stapling for the orthogonal introduction of hydrophobic modifications into the CPP structure during peptide synthesis. Our data show that α,α-disubstituted alkenyl-alanines can be successfully utilized to introduce hydrophobic modifications into CPPs to improve their ability to formulate SSOs into nanoparticles (NPs), and to mediate high delivery efficacy and tolerability both in vitro and in vivo. Conclusively, our results offer a new flexible approach for the sequence-specific introduction of hydrophobicity into the structure of CPPs and for improving their delivery properties.
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| 9. |
- Gustafsson, O, et al.
(författare)
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Efficient Peptide-Mediated In Vitro Delivery of Cas9 RNP
- 2021
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Ingår i: Pharmaceutics. - : MDPI AG. - 1999-4923. ; 13:6
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Tidskriftsartikel (refereegranskat)abstract
- The toolbox for genetic engineering has quickly evolved from CRISPR/Cas9 to a myriad of different gene editors, each with promising properties and enormous clinical potential. However, a major challenge remains: delivering the CRISPR machinery to the nucleus of recipient cells in a nontoxic and efficient manner. In this article, we repurpose an RNA-delivering cell-penetrating peptide, PepFect14 (PF14), to deliver Cas9 ribonucleoprotein (RNP). The RNP-CPP complex achieved high editing rates, e.g., up to 80% in HEK293T cells, while being active at low nanomolar ranges without any apparent signs of toxicity. The editing efficiency was similar to or better compared to the commercially available reagents RNAiMAX and CRISPRMax. The efficiency was thoroughly evaluated in reporter cells and wild-type cells by restriction enzyme digest and next-generation sequencing. Furthermore, the CPP-Cas9-RNP complexes were demonstrated to withstand storage at different conditions, including freeze-thaw cycles and freeze-drying, without a loss in editing efficiency. This CPP-based delivery strategy complements existing technologies and further opens up new opportunities for Cas9 RNP delivery, which can likely be extended to other gene editors in the future.
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