Treatment with IFB-088 Improves Neuropathy in CMT1A and CMT1B Mice.

Charcot-Marie-Tooth disease type 1A (CMT1A), caused by duplication of the peripheral myelin protein 22 (PMP22) gene, and CMT1B, caused by mutations in myelin protein zero (MPZ) gene, are the two most common forms of demyelinating CMT (CMT1), and no treatments are available for either. Prior studies of the MpzSer63del mouse model of CMT1B have demonstrated that protein misfolding, endoplasmic reticulum (ER) retention and activation of the unfolded protein response (UPR) contributed to the neuropathy. Heterozygous patients with an arginine to cysteine mutation in MPZ (MPZR98C) develop a severe infantile form of CMT1B which is modelled by MpzR98C/ + mice that also show ER stress and an activated UPR. C3-PMP22 mice are considered to effectively model CMT1A. Altered proteostasis, ER stress and activation of the UPR have been demonstrated in mice carrying Pmp22 mutations. To determine whether enabling the ER stress/UPR and readjusting protein homeostasis would effectively treat these models of CMT1B and CMT1A, we administered Sephin1/IFB-088/icerguestat, a UPR modulator which showed efficacy in the MpzS63del model of CMT1B, to heterozygous MpzR98C and C3-PMP22 mice. Mice were analysed by behavioural, neurophysiological, morphological and biochemical measures. Both MpzR98C/ + and C3-PMP22 mice improved in motor function and neurophysiology. Myelination, as demonstrated by g-ratios and myelin thickness, improved in CMT1B and CMT1A mice and markers of UPR activation returned towards wild-type values. Taken together, our results demonstrate the capability of IFB-088 to treat a second mouse model of CMT1B and a mouse model of CMT1A, the most common form of CMT. Given the recent benefits of IFB-088 treatment in amyotrophic lateral sclerosis and multiple sclerosis animal models, these data demonstrate its potential in managing UPR and ER stress for multiple mutations in CMT1 as well as in other neurodegenerative diseases. (Left panel) the accumulation of overexpressed PMP22 or misfolded mutant P0 in the Schwann cell endoplasmic reticulum (ER) leads to overwhelming of the degradative capacity, activation of ER-stress mechanisms, and myelination impairment. (Right panel) by prolonging eIF2α phosphorylation, IFB-088 reduces the amount of newly synthesized proteins entering the ER, allowing the protein quality control systems to better cope with the unfolded/misfolded protein and allowing myelination to progress.

Enhanced axonal neuregulin-1 type-III signaling ameliorates neurophysiology and hypomyelination in a Charcot-Marie-Tooth type 1B mouse model.

Charcot-Marie-Tooth (CMT) neuropathies are a group of genetic disorders that affect the peripheral nervous system with heterogeneous pathogenesis and no available treatment. Axonal neuregulin 1 type III (Nrg1TIII) drives peripheral nerve myelination by activating downstream signaling pathways such as PI3K/Akt and MAPK/Erk that converge on master transcriptional regulators of myelin genes, such as Krox20. We reasoned that modulating Nrg1TIII activity may constitute a general therapeutic strategy to treat CMTs that are characterized by reduced levels of myelination. Here we show that genetic overexpression of Nrg1TIII ameliorates neurophysiological and morphological parameters in a mouse model of demyelinating CMT1B, without exacerbating the toxic gain-of-function that underlies the neuropathy. Intriguingly, the mechanism appears not to be related to Krox20 or myelin gene upregulation, but rather to a beneficial rebalancing in the stoichiometry of myelin lipids and proteins. Finally, we provide proof of principle that stimulating Nrg1TIII signaling, by pharmacological suppression of the Nrg1TIII inhibitor tumor necrosis factor-alpha-converting enzyme (TACE/ADAM17), also ameliorates the neuropathy. Thus, modulation of Nrg1TIII by TACE/ADAM17 inhibition may represent a general treatment for hypomyelinating neuropathies.

Laminin 211 inhibits protein kinase A in Schwann cells to modulate neuregulin 1 type III-driven myelination.

Myelin is required for proper nervous system function. Schwann cells in developing nerves depend on extrinsic signals from the axon and from the extracellular matrix to first sort and ensheathe a single axon and then myelinate it. Neuregulin 1 type III (Nrg1III) and laminin α2ß1γ1 (Lm211) are the key axonal and matrix signals, respectively, but how their signaling is integrated and if each molecule controls both axonal sorting and myelination is unclear. Here, we use a series of epistasis experiments to show that Lm211 modulates neuregulin signaling to ensure the correct timing and amount of myelination. Lm211 can inhibit Nrg1III by limiting protein kinase A (PKA) activation, which is required to initiate myelination. We provide evidence that excessive PKA activation amplifies promyelinating signals downstream of neuregulin, including direct activation of the neuregulin receptor ErbB2 and its effector Grb2-Associated Binder-1 (Gab1), thereby elevating the expression of the key transcription factors Oct6 and early growth response protein 2 (Egr2). The inhibitory effect of Lm211 is seen only in fibers of small caliber. These data may explain why hereditary neuropathies associated with decreased laminin function are characterized by focally thick and redundant myelin.

MpzR98C arrests Schwann cell development in a mouse model of early-onset Charcot-Marie-Tooth disease type 1B.

Mutations in myelin protein zero (MPZ) cause Charcot-Marie-Tooth disease type 1B. Many dominant MPZ mutations, including R98C, present as infantile onset dysmyelinating neuropathies. We have generated an R98C 'knock-in' mouse model of Charcot-Marie-Tooth type 1B, where a mutation encoding R98C was targeted to the mouse Mpz gene. Both heterozygous (R98C/+) and homozygous (R98C/R98C) mice develop weakness, abnormal nerve conduction velocities and morphologically abnormal myelin; R98C/R98C mice are more severely affected. MpzR98C is retained in the endoplasmic reticulum of Schwann cells and provokes a transitory, canonical unfolded protein response. Ablation of Chop, a mediator of the protein kinase RNA-like endoplasmic reticulum kinase unfolded protein response pathway restores compound muscle action potential amplitudes of R98C/+ mice but does not alter the reduced conduction velocities, reduced axonal diameters or clinical behaviour of these animals. R98C/R98C Schwann cells are developmentally arrested in the promyelinating stage, whereas development is delayed in R98C/+ mice. The proportion of cells expressing c-Jun, an inhibitor of myelination, is elevated in mutant nerves, whereas the proportion of cells expressing the promyelinating transcription factor Krox-20 is decreased, particularly in R98C/R98C mice. Our results provide a potential link between the accumulation of MpzR98C in the endoplasmic reticulum and a developmental delay in myelination. These mice provide a model by which we can begin to understand the early onset dysmyelination seen in patients with R98C and similar mutations.

P0S63del impedes the arrival of wild-type P0 glycoprotein to myelin in CMT1B mice.

More than 120 mutations in the Myelin Protein Zero gene (MPZ, P0) cause various forms of hereditary neuropathy. Two human mutations encoding either P0S63C or P0S63del have been shown to cause demyelination in mice through different gain of function pathomechanisms. P0S63del, for example, is retained in the endoplasmic reticulum (ER) and elicits a pathogenetic unfolded protein response (UPR). As P0 likely forms oligomers, another gain of abnormal function could include a dominant-negative interaction between P0S63del and normal P0 (P0wt). To test this idea, we generated a transgenic mouse that expressed a form of P0wt with a myc epitope tag at the C terminus (P0ct-myc). We show that P0ct-myc is trafficked and functions like P0wt, thus providing a new tool to study P0 in vivo. In mice that express both P0ct-myc and P0S63del, P0S63del specifically delays the transit of P0ct-myc through the ER and reduces the level of P0wt in the myelin sheath by half-a level previously shown to cause demyelination in mice and humans. Surprisingly, P0ct-myc does not co-immunoprecipitate with P0S63del, suggesting an indirect interaction. Thus, P0S63del causes not only a UPR-related toxic mechanism, but also a dominant-negative effect on P0wt that probably contributes to demyelinating neuropathy.

IL-1 genes in myasthenia gravis: IL-1A -889 polymorphism associated with sex and age of disease onset.

Myasthenia gravis (MG) is a multifactorial autoimmune disease of the neuromuscular junction. We investigated the relation between four polymorphisms of the interleukin (IL)-1 gene cluster on 2q12-22, and MG susceptibility and clinical features in a large cohort of individuals. No polymorphism was associated with MG susceptibility. However, the IL-1A -889 CC genotype was associated with early disease onset (p=0.0044) in the whole MG group and the subgroup of CC males developed MG about 18 years earlier than males carrying other IL-1A -889 genotypes (p=0.022). This finding suggests that IL-1A is a disease modifier in MG, or is in linkage disequilibrium with an unknown locus on chromosome 2.