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. 2024 Nov 12;25(22):12123.
doi: 10.3390/ijms252212123.

Identification of Biochemical Determinants for Diagnosis and Prediction of Severity in 5q Spinal Muscular Atrophy Using 1H-Nuclear Magnetic Resonance Metabolic Profiling in Patient-Derived Biofluids

Afshin Saffari  1 , Moritz Niesert  2 , Claire Cannet  3 , Astrid Blaschek  4 , Andreas Hahn  5 , Jessika Johannsen  6 , Musa Kockaya  7 , Heike Kölbel  8 , Georg F Hoffmann  2 , Peter Claus  9   10   11 , Stefan Kölker  1 , Wolfgang Müller-Felber  4 , Andreas Roos  8 , Ulrike Schara-Schmidt  8 , Friedrich K Trefz  1 , Katharina Vill  4 , Wolfgang Wick  12 , Markus Weiler  12 , Jürgen G Okun  1 , Andreas Ziegler  1   13
Affiliations

Identification of Biochemical Determinants for Diagnosis and Prediction of Severity in 5q Spinal Muscular Atrophy Using 1H-Nuclear Magnetic Resonance Metabolic Profiling in Patient-Derived Biofluids

Afshin Saffari et al. Int J Mol Sci. .

Abstract

This study explores the potential of 1H-NMR spectroscopy-based metabolic profiling in various biofluids as a diagnostic and predictive modality to assess disease severity in individuals with 5q spinal muscular atrophy. A total of 213 biosamples (urine, plasma, and CSF) from 153 treatment-naïve patients with SMA across five German centers were analyzed using 1H-NMR spectroscopy. Prediction models were developed using machine learning algorithms which enabled the patients with SMA to be grouped according to disease severity. A quantitative enrichment analysis was employed to identify metabolic pathways associated with disease progression. The results demonstrate high sensitivity (84-91%) and specificity (91-94%) in distinguishing treatment-naïve patients with SMA from controls across all biofluids. The urinary and plasma profiles differentiated between early-onset (type I) and later-onset (type II/III) SMA with over 80% accuracy. Key metabolic differences involved alterations in energy and amino acid metabolism. This study suggests that 1H-NMR spectroscopy based metabolic profiling may be a promising, non-invasive tool to identify patients with SMA and for severity stratification, potentially complementing current diagnostic and prognostic strategies in SMA management.

Keywords: 1H-NMR spectroscopy; metabolic profiling; metabolomics; spinal muscular atrophy.

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Conflict of interest statement

A.H., A.R., G.F.H., F.T., J.O., M.F., M.K., M.N., M.S. and S.K. have no potential conflicts of interest to declare. A.B. received honoraria for speaking at scientific meetings and serving on scientific advisory boards from Roche and Novartis, manufacturers of SMA therapeutics. A.S. and A.Z. received research funding from Biogen through a sponsored research agreement. A.Z. received consult fees, lecture fees, and travel support from Biogen, Roche, Novartis, Pfizer, and ITF Pharma. C.C. and D.P. are employees of Bruker Biospin GmbH & Co KG, Germany, a manufacturer of NMR-Spectrometers. H.K. received travel support from Biogen, Novartis, and Roche, manufacturers of therapeutics for SMA. U.S.-S. received consulting fees, lecture honoraria, and travel support from Biogen, Novartis, and Roche. H.K. received travel support from Sanofi, Novartis, Biogen, and Roche. J.J. received fees for advisory board attendance from Biogen, Novartis, and Roche. K.V. received travel support and lecture honoraria from Biogen, Novartis, and Sanofi. P.C. received lecture honoraria from Roche and Novartis and is the CEO of SMATheria Gmbh, a private non-profit research institute focusing on SMA. U.S.-S. and W.M.F. received consult fees, lecture honoraria, and travel support from Roche, Biogen, and Novartis. M.W. received consult fees and attended the advisory boards of Biogen and Roche and received travel support from Biogen. W.W. received consulting fees from Roche. A.Z. received consulting fees, honoraria, and travel support from Biogen, Novartis, and Roche.

Figures

Figure A1
Figure A1
Visualization of metabolite levels in G12.0, G12.1, and healthy controls in different biomaterials, represented by heatmaps showing group average. Red indicates high levels compared to mean value; blue indicates low levels compared to mean value. Gray indicates missing values. (A) Urine, (B) plasma, and (C) CSF.
Figure 1
Figure 1
1H-NMR metabolic profiles in urine (A), plasma (B), and CSF (C) from patients with SMA vs. healthy controls. PCA/CA classification and MCCV showed clear discrimination between SMA group and healthy control group.
Figure 2
Figure 2
1H-NMR metabolic profiles of G12.0 (severely affected) and G12.1 (mildly to moderately affected) vs. healthy controls in urine (A), plasma (B), and CSF (C) samples. PCA/CA classification and MCCV showed clear discrimination between severity subgroups and the healthy control group.
Figure 3
Figure 3
A univariate analysis of the full 1H-NMR urinary spectrum for G12.0, G12.1, and the healthy control group. The discriminating region between the groups revealed by the Kruskal–Wallis test (at p < 0.05 significance level) is highlighted in light pink. The median of each group is represented by a line (G12.0 is red, G12.1 is blue, and the healthy control is purple), and the 5–95% percentile of each group is represented by the corresponding light color area. (A) full 1H-NMR urinary spectrum (0.8–9.0 ppm). (B) A zoomed in figure of the aliphatic region (0.8–2.5 ppm). (C) A zoomed in figure of a specific region illustrating the discrimination between all 3 groups (2.6–2.8 ppm).
Figure 4
Figure 4
A summary plot for the quantitative enrichment analysis of G12.0 vs. G12.1 in patients’ urine (A), plasma (B), and CSF (C) samples. The quantitative enrichment analysis was performed by mapping metabolites on the KEGG human database using Metaboanalyst 6.0. The top 25 enriched metabolite sets are plotted. The X-axis represents the −log(p-value) indicating the significance of pathway enrichment, while the Y-axis lists the affected metabolic pathways. The dot size reflects the enrichment ratio. (D) Additionally, a Venn diagram is included to visualize the overlap between enriched pathways. The metabolite sets that overlap between all three sample qualities are the citrate cycle (TCA cycle); alanine, aspartate, and glutamate metabolism; glyoxylate and dicarboxylate metabolism; starch and sucrose metabolism; neomycin, kanamycin, and gentamicin biosynthesis (D-Glucose); primary bile acid biosynthesis; galactose metabolism; cysteine and methionine metabolism; valine, leucine, and isoleucine biosynthesis; lipoic acid metabolism; glutathione metabolism; porphyrin metabolism; glycine, serine, and threonine metabolism; pyruvate metabolism; glycolysis/gluconeogenesis; pantothenate and CoA biosynthesis; phenylalanine, tyrosine, and tryptophan biosynthesis; phenylalanine metabolism; arginine and proline metabolism; butanoate metabolism; tyrosine metabolism; valine, leucine, and isoleucine degradation; and glycerophospholipid metabolism.
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