Drug Screening Identifies Sigma-1-Receptor as a Target for the Therapy of VWM Leukodystrophy

Abstract

Vanishing white matter (VWM) disease is an autosomal genetic leukodystrophy caused by mutations in subunits of eukaryotic translation initiation factor 2B (eIF2B). The clinical symptoms exhibit progressive loss of white matter in both hemispheres of the brain, accompanied by motor functions deterioration, neurological deficits, and early death. To date there is no treatment for VWM disease. The aim of this work was to expedite rational development of a therapeutic opportunity. Our approach was to design a computer-aided strategy for an efficient and reliable screening of drug-like molecules; and to use primary cultures of fibroblasts isolated from the Eif2b5R132H/R132H VWM mouse model for screening. The abnormal mitochondria content phenotype of the mutant cells was chosen as a read-out for a simple cell-based fluorescent assay to assess the effect of the tested compounds. We obtained a hit rate of 0.04% (20 hits out of 50,000 compounds from the selected library). All primary hits decreased mitochondria content and brought it closer to WT levels. Structural similarities between our primary hits and other compounds with known targets allowed the identification of three putative cellular pathways/targets: 11β-hydroxysteroid dehydrogenase type 1, Sonic hedgehog (Shh), and Sigma-1-Receptor (S1R). In addition to initial experimental indication of Shh pathway impairment in VWM mouse brains, the current study provides evidence that S1R is a relevant target for pharmaceutical intervention for potential treatment of the disease. Specifically, we found lower expression level of S1R protein in fibroblasts, astrocytes, and whole brains isolated from Eif2b5R132H/R132H compared to WT mice, and confirmed that one of the hits is a direct binder of S1R, acting as agonist. Furthermore, we provide evidence that treatment of mutant mouse fibroblasts and astrocytes with various S1R agonists corrects the functional impairments of their mitochondria and prevents their need to increase their mitochondria content for compensation purposes. Moreover, S1R activation enhances the survival rate of mutant cells under ER stress conditions, bringing it to WT levels. This study marks S1R as a target for drug development toward treatment of VWM disease. Moreover, it further establishes the important connection between white matter well-being and S1R-mediated proper mitochondria/ER function.

Astrocyte Subtype Vulnerability in Stem Cell Models of Vanishing White Matter

“…In mouse cultures, only 13 DEGs between VWM and control were found, of which some genes have been related to VWM disease mechanisms previously. 28 Interestingly, VWM mouse cultures also showed increased Ets1 expression. Previous studies showed an increase in CD44 expression in VWM patient white matter astrocytes in postmortem tissue.…”

“…A recent study showed an impaired SHH pathway in primary astrocytes from the Eif2b5 R132H/R132H mouse, confirming the involvement of the SHH pathway in VWM. 28 Interestingly, VWM mouse cultures also showed increased Ets1 expression. Ets1 is a transcription factor that is involved in the T-cell immune response and was shown to be upregulated in astrocytes surrounding white matter lesions in a mouse model for multiple sclerosis.…”

“…This may further lead to astrocytic glutamate release, thereby potentiating inhibitory synaptic transmission, and to astrocytic ATP/adenosine-mediated heterosynaptic suppression affecting excitatory transmission. 28 Thus, these pathways deserve further study to determine whether these can target for therapeutic strategies. 33 Our human transcriptome data furthermore showed a number of DEGs involved in mitochondrial functioning, including CTGF, CYP1B1, NTRK2, CNR1, TBX2, and MT2A.…”

“…39 Mitochondrial mechanisms involved in VWM pathophysiology corroborate recent findings in primary astrocytes of Eif2b5 R132H/R132H mice indicating that an impaired oxidative phosphorylation in these cells is compensated by increased mitochondrial content and glycolysis. 28 Thus, these pathways deserve further study to determine whether these can target for therapeutic strategies.…”

Human and mouse iPSC-derived astrocyte subtypes reveal vulnerability in Vanishing White Matter

“…While not significant, SHH and SMO levels were also decreased (2-2.5 fold) in VWM white matter-like astrocytes. A recent study showed an impaired SHH pathway in primary astrocytes from the Eif2b5 R132H/R132H mouse, confirming the involvement of the SHH pathway in VWM (Atzmon et al, 2018). Interestingly, VWM mouse cultures also showed increased Ets1 expression.…”