FIGURE SUMMARY
- Title
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Novel mRNA therapy restores GALT protein and enzyme activity in a zebrafish model of classical galactosemia
- Authors
- Delnoy, B., Haskovic, M., Vanoevelen, J., Steinbusch, L.K.M., Vos, E.N., Knoops, K., Zimmermann, L.J.I., Noga, M., Lefeber, D.J., Martini, P.G.V., Coelho, A.I., Rubio-Gozalbo, M.E.
- Source
- Full text @ J. Inherit. Metab. Dis.
Study design. Naked, LNP1 and LNP2-packaged hGALT mRNAs provided by Moderna Inc. were injected in CG zebrafish (galt KO) zebrafish at one-cell stage (0–1 h post fertilization, hpf) and intravenous (48–56 hpf). GALT activity, protein expression (merely one-cell stage injections) and Gal-1-P and galactonate (merely intravenous injections) were assessed at 5 days post fertilization (dpf). LNP, lipid nanoparticle; mRNA, messenger RNA; hpf, hours post fertilization; dpf, days post fertilization; LNP1, LNP1-packaged mRNA; LNP2, LNP2-packaged mRNA; Naked, naked mRNA; Gal-1-P, galactose-1-phosphate. Figure is created with BioRender.com
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hGALT expression and activity in 5 dpf CG zebrafish (galt KO) after single dose hGALT mRNA injection (100 ng/μl) at the one-cell stage. A: Western blot analysis: Non-injected WT and CG zebrafish (galt KO) controls present no band, which indicates that the antibody is specific for hGALT. Samples were loaded in duplicate (n = 150 per sample). B: GALT activity (nmol/mg protein/h), results are presented as mean ± SEM. All samples (n = 3 per experimental group, on average 30 zebrafish/sample), were measured in duplicate. MW, molecular weight ladder; WT, wildtype; NIC, non-injected CG zebrafish (galt KO) control; NKD, naked mRNA injected CG zebrafish (galt KO); LNP1, LNP1-packaged hGALT mRNA injected CG zebrafish (galt KO); LNP2, LNP2-packaged hGALT mRNA injected CG zebrafish (galt KO)
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hGALT activity in 5 dpf CG zebrafish (galt KO) after single dose intravenous hGALT mRNA injection (100 ng/μl). Results are presented as mean ± SEM. All samples (n = 6 for wildtype zebrafish and n = 4 for NIC and LNP2 injected zebrafish, ~90 zebrafish/sample), were measured in duplicate. WT, wild-type; NIC, non-injected CG zebrafish (galt KO) control; LNP2, LNP2-packaged hGALT mRNA injected CG zebrafish (galt KO)
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Gal-1-P and Galactonate in 5 dpf CG zebrafish (galt KO) after single dose intravenous hGALT mRNA injection (100 ng/μl). A. Gal-1-P (peak area) B. Galactonate (peak area)Results are presented as mean ± SEM. C, D, E. Gal-1-P levels and corresponding logarithm of GALT activity levels (nmol/mg protein/h) Sample size per condition (unexposed or exposed to galactose) was n = 6 for WT zebrafish and n = 4 for NIC and LNP2 injected zebrafish, ~90 zebrafish/sample see Table S1). WT, wild-type; NIC, non-injected CG zebrafish (galt KO) control; LNP2, LNP2-packaged hGALT mRNA injected CG zebrafish (galt KO)
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Galactose (100 mM) exposure: survival rates in 5 dpf CG zebrafish (galt KO) following single dose intravenous hGALT injection (100 ng/μl). Total sample size at the start of galactose exposure can be found in Table S2, results are presented as survival rate ± 95% CI. WT, wildtype; NIC, non-injected CG zebrafish (galt KO) control; LNP2, LNP2-packaged hGALT mRNA injected CG zebrafish (galt KO)
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Full text @ J. Inherit. Metab. Dis.