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Aberrant proteostasis is thought to be involved in the pathogenesis of neurodegenerative diseases. Some proteostasis abnormalities are ameliorated by chaperones. Chaperones are divided into three groups: molecular, pharmacological and chemical. Chemical chaperones intended to alleviate stress in organelles, such as the endoplasmic reticulum (ER), are now being administered clinically. Of the chemical chaperones, 4-phenylbutyrate (4-PBA) has been used as a research reagent, and its mechanism of action includes chaperone effects and the inhibition of histone deacetylase. Moreover, it also binds to the B-site of SEC24 and regulates COPII-mediated transport from the ER. Although its therapeutic effect may not be strong, elucidating the mechanism of action of 4-PBA may contribute to the identification of novel therapeutic targets for neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Alport syndrome is a hereditary kidney disease caused by collagen IV mutations that interfere with the formation and deposition of the α3α4α5 protomer into the glomerular basement membrane. In this issue, Yu et al. show that the chemical chaperone tauroursodeoxycholic acid prevented kidney structural changes and function decline in mice with a pathogenic missense Col4a3 mutation by increasing mutant α3α4α5 protomer glomerular basement membrane deposition and preventing podocyte apoptosis induced by endoplasmic reticulum stress., (Copyright © 2024 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)

Proteins form native structures through folding processes, many of which proceed through intramolecular hydrophobic effect, hydrogen bond and disulfide-bond formation. In vivo , protein aggregation is prevented even in the highly condensed milieu of a cell through folding mediated by molecular chaperones and oxidative enzymes. Chemical approaches to date have not replicated such exquisite mediation. Oxidoreductases efficiently promote folding by the cooperative effects of oxidative reactivity for disulfide-bond formation in the client unfolded protein and chaperone activity to mitigate aggregation. Conventional synthetic folding promotors mimic the redox-reactivity of thiol/disulfide units but do not address client-recognition units for inhibiting aggregation. Herein, we report thiol/disulfide compounds containing client-recognition units, which act as synthetic oxidoreductase-mimics. For example, compound βCD W SH/SS bears a thiol/disulfide unit at the wide rim of β-cyclodextrin as a client recognition unit. βCD W SH/SS shows promiscuous binding to client proteins, mitigates protein aggregation, and accelerates disulfide-bond formation. In contrast, positioning a thiol/disulfide unit at the narrow rim of β-cyclodextrin promotes folding less effectively through preferential interactions at specific residues, resulting in aggregation. The combination of promiscuous client-binding and redox reactivity is effective for the design of synthetic folding promoters. βCD W SH/SS accelerates oxidative protein folding at highly condensed sub-millimolar protein concentrations., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Jae-Ho Jeon,1,* Somyoung Im,1,* Hyo Shin Kim,1 Dongyun Lee,1 Kwiwan Jeong,2 Jin-Mo Ku,2 Tae-Gyu Nam1 1Department of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University ERICA campus, Ansan, Gyeonggi-do, 15588, Republic of Korea; 2Gyeonggi Bio-Center, Gyeonggido Business and Science Accelerator, Suwon, Gyeonggi-do, 16229, Republic of Korea*These authors contributed equally to this workCorrespondence: Tae-Gyu Nam, Tel +82-31-400-5807, Fax +82-31-400-5958, Email tnam@hanyang.ac.krAbstract: The endoplasmic reticulum (ER) is responsible for structural transformation or folding of de novo proteins for transport to the Golgi. When the folding capacity of the ER is exceeded or excessive accumulation of misfolded proteins occurs, the ER enters a stressed condition (ER stress) and unfolded protein responses (UPR) are triggered in order to rescue cells from the stress. Recovery of ER proceeds toward either survival or cell apoptosis. ER stress is implicated in many pathologies, such as diabetes, cardiovascular diseases, inflammatory diseases, neurodegeneration, and lysosomal storage diseases. As a survival or adaptation mechanism, chaperone molecules are upregulated to manage ER stress. Chemical versions of chaperone have been developed in search of drug candidates for ER stress-related diseases. In this review, synthetic or semi-synthetic chemical chaperones are categorized according to potential therapeutic area and listed along with their chemical structure and activity. Although only a few chemical chaperones have been approved as pharmaceutical drugs, a dramatic increase in literatures over the recent decades indicates enormous amount of efforts paid by many researchers. The efforts warrant clearer understanding of ER stress and the related diseases and consequently will offer a promising drug discovery platform with chaperone activity.Keywords: endoplasmic reticulum stress, unfolded protein response, chemical chaperone, drug discovery, diabetes, cardiovascular disease, neurodegeneration, lysosomal storage disease

Abstract There are no available therapies targeting the underlying molecular mechanisms of neurodegenerative diseases. Although chaperone therapies that alleviate endoplasmic reticulum (ER) stress recently showed promise in the treatment of neurodegenerative diseases, the detailed mechanisms remain unclear. We previously reported that mice with central nervous system-specific deletion of Derlin-1, which encodes an essential component for ER quality control, are useful as models of neurodegenerative diseases such as spinocerebellar degeneration. Cholesterol biosynthesis is essential for brain development, and its disruption inhibits neurite outgrowth, causing brain atrophy. In this study, we report a novel mechanism by which chemical chaperones ameliorate brain atrophy and motor dysfunction. ER stress was induced in the cerebella of Derlin-1 deficiency mice, whereas the administration of a chemical chaperone did not alleviate ER stress. However, chemical chaperone treatment ameliorated cholesterol biosynthesis impairment through SREBP-2 activation and simultaneously relieved brain atrophy and motor dysfunction. Altogether, these findings demonstrate that ER stress may not be the target of action of chaperone therapies and that chemical chaperone-mediated improvement of brain cholesterol biosynthesis is a promising novel therapeutic strategy for neurodegenerative diseases.

Amyotrophic lateral sclerosis (ALS) is a progressive neurological disorder with currently no cure. Central to the cellular dysfunction associated with this fatal proteinopathy is the accumulation of unfolded/misfolded superoxide dismutase 1 (SOD1) in various subcellular locations. The molecular mechanism driving the formation of SOD1 aggregates is not fully understood but numerous studies suggest that aberrant aggregation escalates with folding instability of mutant apoSOD1. Recent advances on combining organelle-targeting therapies with the anti-aggregation capacity of chemical chaperones have successfully reduce the subcellular load of misfolded/aggregated SOD1 as well as their downstream anomalous cellular processes at low concentrations (micromolar range). Nevertheless, if such local aggregate reduction directly correlates with increased folding stability remains to be explored. To fill this gap, we synthesized and tested here the effect of 9 ER-, mitochondria- and lysosome-targeted chemical chaperones on the folding stability of truncated monomeric SOD1 (SOD1bar) mutants directed to those organelles. We found that compound ER-15 specifically increased the native state stability of ER-SOD1bar-A4V, while scaffold compound FDA-approved 4-phenylbutyric acid (PBA) decreased it. Furthermore, our results suggested that ER15 mechanism of action is distinct from that of PBA, opening new therapeutic perspectives of this novel chemical chaperone on ALS treatment. [ABSTRACT FROM AUTHOR]

Osmolytes are well known to protect the protein structure against different chemical and physical denaturants. Since their actions with protein surfaces are mechanistically complicated and context dependent, the underlying molecular mechanism is not fully understood. Here, we combined single-molecule magnetic tweezers and molecular dynamics (MD) simulation to explore the mechanical role of osmolytes from two different classes, trimethylamine N -oxide (TMAO) and trehalose, as mechanical stabilizers of protein structure. We observed that these osmolytes increase the protein L mechanical stability by decreasing unfolding kinetics while accelerating the refolding kinetics under force, eventually shifting the energy landscape toward the folded state. These osmolytes mechanically stabilize the protein L and plausibly guide them to more thermodynamically robust states. Finally, we observed that osmolyte-modulated protein folding increases mechanical work output up to twofold, allowing the protein to fold under a higher force regime and providing a significant implication for folding-induced structural stability in proteins.

The importance of studying the structural stability of proteins is determined by the structure–function relationship. Protein stability is influenced by many factors among which are freeze–thaw and thermal stresses. The effect of trehalose, betaine, sorbitol and 2-hydroxypropyl-β-cyclodextrin (HPCD) on the stability and aggregation of bovine liver glutamate dehydrogenase (GDH) upon heating at 50 °C or freeze–thawing was studied by dynamic light scattering, differential scanning calorimetry, analytical ultracentrifugation and circular dichroism spectroscopy. A freeze–thaw cycle resulted in the complete loss of the secondary and tertiary structure, and aggregation of GDH. All the cosolutes suppressed freeze–thaw- and heat-induced aggregation of GDH and increased the protein thermal stability. The effective concentrations of the cosolutes during freeze–thawing were lower than during heating. Sorbitol exhibited the highest anti-aggregation activity under freeze–thaw stress, whereas the most effective agents stabilizing the tertiary structure of GDH were HPCD and betaine. HPCD and trehalose were the most effective agents suppressing GDH thermal aggregation. All the chemical chaperones stabilized various soluble oligomeric forms of GDH against both types of stress. The data on GDH were compared with the effects of the same cosolutes on glycogen phosphorylase b during thermal and freeze–thaw-induced aggregation. This research can find further application in biotechnology and pharmaceutics. [ABSTRACT FROM AUTHOR]

(1) Background: Congenital factor (F) VII deficiency is caused by mutations in the F7 gene. Patients with modest differences in FVII levels may display large differences in clinical severity. The variant p.A354V-p.P464Hfs is associated with reduced FVII antigen and activity. The aim of the study was to investigate the clinical manifestation of this variant and the underlying molecular mechanisms. (2) Methods: Analyses were conducted in 37 homozygous patients. The recombinant variant was produced in mammalian cells. (3) Results: We report a large variation in clinical phenotypes, which points out genetic and acquired components beyond F7 mutations as a source of variability. In contrast, patients displayed similarly reduced FVII plasma levels with antigen higher than its activity. Comparative analysis of the recombinant variant and of plasma samples from a subset of patients indicated the presence of an elongated variant with indistinguishable migration. Treatment of cells with the chemical chaperone 4-phenylbutyrate (4-PBA) improved the intracellular trafficking of the variant and increased its secretion to the conditioned medium up to 2-fold. However, the effect of 4-PBA on biological activity was marginal. (4) Conclusions: Chemical chaperones can be used as biochemical tools to study the intracellular fate of a trafficking-defective FVII variant.

Abstract Background Imbalance between protein synthesis and endoplasmic reticulum (ER) capacity to modify and fold proteins lead to the accumulation of unfolded proteins resulting in ER stress and apoptosis. Chaperones are major defense molecules assisting in protein folding, transport, and cellular signaling. ER stress plays a major role in the pathogenesis of diabetes mellitus (DM) and its complications, e.g., diabetic cataract. In the present investigation, the chemical chaperones 4-phenylbutyric acid (4-PBA), tauroursodeoxycholic acid (TUDCA), and trimethylamine N-oxide (TMAO) are used as potential therapeutic agents for alleviation of DM-induced ER stress and diabetic cataract in rats. Animals are subjected to biochemical analysis of blood and lenses for ER stress and apoptosis markers. Moreover, ophthalmologic examination and histopathologic examination of the lenses were done to confirm the results. Results Both ophthalmic and lens histopathologic examination revealed that treatment with 4-PBA and TUDCA retarded the occurrence of cataract markedly. Whereas, treatment with TMAO caused a partial improvement of cataract. Moreover, biochemical tests showed that both 4-PBA and TUDCA produced a remarkable improvement in the ER marker levels (VEGF and caspase-12), GSH, MDA, TAC levels in lens tissues. On the other hand, TMAO had no significant effect on these parameters. However, Western blot analysis of lens homogenates showed a suppressed expression of GRP78 and CHOP after treatment with 4-PBA, TUDCA, and TMAO. Moreover, all treated groups showed a significant improvement of lens soluble proteins and their UV spectra absorption. A significant improvement in fasting blood sugar, GSH, serum MDA, and TAC were noted in all treated groups. 4-PBA produced a significant decrease in insulin resistance, whereas TUDCA and TMAO showed insignificant change. Conclusion The present research found that the tested chaperones could be used as a therapeutic approach for clinically relevant disorders featuring ER dysfunction such as DM and for reducing its complications in the eye mainly cataract. However, TUDCA and 4-PBA were found to have a more potential efficacy in reducing most of the tested parameters as compared to TMAO.

The Bradyrhizobium

Munc18-1 is an evolutionary conserved gene whose mutations are linked to various neurological diseases in human. In order to better understand the exact nature of the mutations, the authors here utilize several model systems to show mutant Munc18-1 can aggregate and deplete functional pool of Wt protein, and that chemical chaperones can reverse the cellular deficits.

Accumulation of misfolded and mistrafficked rhodopsin on the endoplasmic reticulum of photoreceptor cells has a pivotal role in the pathogenesis of retinitis pigmentosa and a subset of Leber’s congenital amaurosis. One potential strategy to reduce rhodopsin misfolding and aggregation in these conditions is to use opsin-binding compounds as chemical chaperones for opsin. Such molecules have previously shown the ability to aid rhodopsin folding and proper trafficking to the outer cell membranes of photoreceptors. As means to identify novel chemical chaperones for rhodopsin, a structure-based virtual screening of commercially available drug-like compounds (300,000) was performed on the main binding site of the visual pigment chromophore, the 11-cis-retinal. The best 24 virtual hits were examined for their ability to compete for the chromophore-binding site of opsin. Among these, four small molecules demonstrated the ability to reduce the rate constant for the formation of the 9-cis-retinal-rhodopsin complex, while five molecules surprisingly enhanced the formation of this complex. Compound 7, 13, 20 and 23 showed a weak but detectable increase in the trafficking of the P23H mutant, widely used as a model for both retinitis pigmentosa and Leber’s congenital amaurosis, from the ER to the cell membrane. The compounds did not show any relevant cytotoxicity in two different human cell lines, with the only exception of 13. Based on the structures of these active compounds, a series of in silico studies gave important insights on the potential structural features required for a molecule to act either as chemical chaperone or as stabiliser of the 11-cis-retinal-rhodopsin complex. Thus, this study revealed a series of small molecules that represent a solid foundation for the future development of novel therapeutics against these severe inherited blinding diseases.

Endoplasmic reticulum (ER) stress and oxidative stress lead to protein misfolding, and the resulting accumulation of protein aggregates is often associated with the pathogenesis of neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and prion disease. Small molecules preventing these pathogenic processes may be effective interventions for such neurodegenerative disorders. In this paper, we identify several novel oxindole compounds that can prevent ER stress- and oxidative stress-induced cell death. Among them, derivatives of the lead compound GIF-0726-r in which a hydrogen atom at the oxindole ring 5 position is substituted with a methyl (GIF-0852-r), bromine (GIF-0854-r), or nitro (GIF-0856-r) group potently suppressed global ER stress. Furthermore, GIF-0854-r and -0856-r prevented protein aggregate accumulation in vitro and in cultured hippocampal HT22 neuronal cells, indicating that these two compounds function effectively as chemical chaperones. In addition, GIF-0852-r, -0854-r, and -0856-r prevented glutamate-induced oxytosis and erastin-induced ferroptosis. Collectively, these results suggest that the novel oxindole compounds GIF-0854-r and -0856-r may be useful therapeutics against protein-misfolding diseases as well as valuable research tools for studying the molecular mechanisms of ER and oxidative stress.