scholarly journals Pharmacological Chaperones: A Therapeutic Approach for Diseases Caused by Destabilizing Missense Mutations

2020 ◽  
Vol 21 (2) ◽  
pp. 489 ◽  
Author(s):  
Ludovica Liguori ◽  
Maria Monticelli ◽  
Mariateresa Allocca ◽  
Bruno Hay Mele ◽  
Jan Lukas ◽  
...  
Keyword(s):  
Small Molecules ◽  
Specific Binding ◽  
Total Activity ◽  
Missense Mutations ◽  
Lysosomal Storage ◽  
Pharmacological Chaperone ◽  
Mutant Proteins ◽  
Pharmacological Chaperones ◽  
Protein Mutants ◽  
Competitive Inhibitors

The term “pharmacological chaperone” was introduced 20 years ago. Since then the approach with this type of drug has been proposed for several diseases, lysosomal storage disorders representing the most popular targets. The hallmark of a pharmacological chaperone is its ability to bind a protein specifically and stabilize it. This property can be beneficial for curing diseases that are associated with protein mutants that are intrinsically active but unstable. The total activity of the affected proteins in the cell is lower than normal because they are cleared by the quality control system. Although most pharmacological chaperones are reversible competitive inhibitors or antagonists of their target proteins, the inhibitory activity is neither required nor desirable. This issue is well documented by specific examples among which those concerning Fabry disease. Direct specific binding is not the only mechanism by which small molecules can rescue mutant proteins in the cell. These drugs and the properly defined pharmacological chaperones can work together with different and possibly synergistic modes of action to revert a disease phenotype caused by an unstable protein.

A counterintuitive approach to treat enzyme deficiencies: use of enzyme inhibitors for restoring mutant enzyme activity

2008 ◽  
Vol 389 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Jian-Qiang Fan
Keyword(s):  
Enzyme Activity ◽  
Active Site ◽  
Protein Misfolding ◽  
Enzyme Inhibitors ◽  
Lysosomal Storage ◽  
New Paradigm ◽  
Inherited Disorders ◽  
Pharmacological Chaperone ◽  
Mutant Proteins ◽  
Enzyme Active Site

Abstract Pharmacological chaperone therapy is an emerging counterintuitive approach to treat protein deficiencies resulting from mutations causing misfolded protein conformations. Active-site-specific chaperones (ASSCs) are enzyme active-site directed small molecule pharmacological chaperones that act as a folding template to assist protein folding of mutant proteins in the endoplasmic reticulum (ER). As a result, excessive degradation of mutant proteins in the ER-associated degradation (ERAD) machinery can be prevented, thus restoring enzyme activity. Lysosomal storage disorders (LSDs) are suitable candidates for ASSC treatment, as the levels of enzyme activity needed to prevent substrate storage are relatively low. In addition, ASSCs are orally active small molecules and have potential to gain access to most cell types to treat neuronopathic LSDs. Competitive enzyme inhibitors are effective ASSCs when they are used at sub-inhibitory concentrations. This whole new paradigm provides excellent opportunity for identifying specific drugs to treat a broad range of inherited disorders. This review describes protein misfolding as a pathophysiological cause in LSDs and provides an overview of recent advances in the development of pharmacological chaperone therapy for the diseases. In addition, a generalized guidance for the design and screening of ASSCs is also presented.

scholarly journals Second-Generation Pharmacological Chaperones: Beyond Inhibitors

Molecules ◽  
2020 ◽  
Vol 25 (14) ◽  
pp. 3145 ◽  
Author(s):  
My Lan Tran ◽  
Yves Génisson ◽  
Stéphanie Ballereau ◽  
Cécile Dehoux
Keyword(s):  
Protein Misfolding ◽  
First Generation ◽  
Second Generation ◽  
Missense Mutations ◽  
Loss Of Function ◽  
Pharmacological Chaperone ◽  
Conformational Diseases ◽  
Pharmacological Chaperones ◽  
Drug Candidates ◽  
Binding Pockets

Protein misfolding induced by missense mutations is the source of hundreds of conformational diseases. The cell quality control may eliminate nascent misfolded proteins, such as enzymes, and a pathological loss-of-function may result from their early degradation. Since the proof of concept in the 2000s, the bioinspired pharmacological chaperone therapy became a relevant low-molecular-weight compound strategy against conformational diseases. The first-generation pharmacological chaperones were competitive inhibitors of mutant enzymes. Counterintuitively, in binding to the active site, these inhibitors stabilize the proper folding of the mutated protein and partially rescue its cellular function. The main limitation of the first-generation pharmacological chaperones lies in the balance between enzyme activity enhancement and inhibition. Recent research efforts were directed towards the development of promising second-generation pharmacological chaperones. These non-inhibitory ligands, targeting previously unknown binding pockets, limit the risk of adverse enzymatic inhibition. Their pharmacophore identification is however challenging and likely requires a massive screening-based approach. This review focuses on second-generation chaperones designed to restore the cellular activity of misfolded enzymes. It intends to highlight, for a selected set of rare inherited metabolic disorders, the strategies implemented to identify and develop these pharmacologically relevant small organic molecules as potential drug candidates.

Pharmacological chaperone corrects lysosomal storage in Fabry disease caused by trafficking-incompetent variants

2006 ◽  
Vol 290 (4) ◽  
pp. C1076-C1082 ◽  
Author(s):  
Gary Hin-Fai Yam ◽  
Nils Bosshard ◽  
Christian Zuber ◽  
Beat Steinmann ◽  
Jürgen Roth
Keyword(s):  
Enzyme Activity ◽  
Fabry Disease ◽  
Competitive Inhibitor ◽  
Apparent Difference ◽  
Residual Enzyme Activity ◽  
Lysosomal Storage ◽  
Electron Microscopic ◽  
Pharmacological Chaperone ◽  
Mutant Proteins

Fabry disease is a lysosomal storage disorder caused by deficiency of α-galactosidase A (α-Gal A) resulting in lysosomal accumulation of glycosphingolipid globotriosylceramide Gb3. Misfolded α-Gal A variants can have residual enzyme activity but are unstable. Their lysosomal trafficking is impaired because they are retained in the endoplasmic reticulum (ER) by quality control. Subinhibitory doses of the competitive inhibitor of α-Gal A, 1-deoxygalactonojirimycin (DGJ), stabilize mutant α-Gal A in vitro and correct the trafficking defect. We showed by immunolabeling that the chaperone-like action of DGJ significantly reduces the lysosomal Gb3 storage in human Fabry fibroblasts harboring the novel mutations T194I and V390fsX8. The specificity of the DGJ effect was proven by RNA interference. Electron microscopic morphometry demonstrated a reduction of large-size, disease-associated lysosomes and loss of characteristic multilamellar lysosomal inclusions on DGJ treatment. In addition, the pre-Golgi intermediates were decreased. However, the rough ER was not different between DGJ-treated and untreated cells. Pulse-chase experiments revealed that DGJ treatment resulted in maturation and stabilization of mutant α-Gal A. Genes involved in cell stress signaling, heat shock response, unfolded protein response, and ER-associated degradation show no apparent difference in expression between untreated and DGJ-treated fibroblasts. The DGJ treatment has no apparent cytotoxic effects. Thus our data show the usefulness of a pharmacological chaperone for correction of the lysosomal storage in Fabry fibroblasts harboring different mutations with residual enzyme activity. Pharmacological chaperones acting on misfolded, unstable mutant proteins that exhibit residual biological activity offer a convenient and cost-efficient therapeutic strategy.

scholarly journals In vitro pharmacological profiling of selected small molecules compound using recombinant human iduronate-2-sulphatase

2021 ◽  
Vol 5 (2) ◽  
pp. 26-30
Author(s):  
Affandi Omar ◽  
Dyg Pertiwi Abg Kamaludin ◽  
Salina Abdul Rahman ◽  
Rosnani Mohamed ◽  
Fatimah Diana Amin Nordin ◽  
...  
Keyword(s):  
Thermal Stability ◽  
Small Molecules ◽  
Heparan Sulphate ◽  
Stability Study ◽  
Lysosomal Storage ◽  
Pharmacological Chaperone ◽  
Enzyme Replacement ◽  
Mps Ii ◽  
System Manifestation

Background: Mucopolysaccharidoses type II (MPS II) is an X-linked lysosomal storage disease (LSD). It is due to mutation in IDS gene encoding iduronate-2-sulphatase (IDS) involved in the catabolism of dermatan sulphate and heparan sulphate. Currently, the treatments for MPS II patients are enzyme replacement therapy (ERT) and bone marrow transplantation (BMT). However, ERT is not effectively reducing the central nervous system manifestation and finding the suitable donor maybe quite challenging in BMT. Over the past decades, pharmacological chaperone has been an alternative approach for management of MPS II patient. Here, we described the in vitro profiling of small molecules in group of chondroitin/dermatan (CD) sulphate disaccharide, heparin oligosaccharides, unsaturated heparin disaccharides and 6-O-desulphated heparin oligosaccharide, using recombinant human iduronate-2-sulphatase (rhIDS). Twenty-one small molecule compounds with several concentrations were each screened by inhibition and thermal stability assays. Results: Our study revealed that condroitin dermatan trisulphate (CD3S), heparin tetrasaccharide (H4Sac), heparin octasaccharide (H8Sac) and heparin octadecasaccharide (H18Sac) showed high inhibition constant, Ki and low inhibition concentration, IC50 in comparison to others. In the thermal stability study, only rhIDS incubated with CD3S was found to preserve enzyme activity (20%) after incubated at 67oC. Conclusion: Overall, our experiments discovered that CD3S was able to bind, inhibit and chaperone rhIDS. These features suggest a potential pharmacological chaperone for MPS II.

scholarly journals A mutation-led search for novel functional domains in MeCP2

2018 ◽  
Author(s):  
Jacky Guy ◽  
Beatrice Alexander-Howden ◽  
Laura FitzPatrick ◽  
Dina DeSousa ◽  
Martha V. Koerner ◽  
...  
Keyword(s):  
Small Molecules ◽  
Rett Syndrome ◽  
Functional Domains ◽  
Cultured Neurons ◽  
Missense Mutations ◽  
Mutant Proteins ◽  
Methylated Dna ◽  
Therapeutic Value ◽  
Repressor Complex

AbstractMost missense mutations causing Rett syndrome affect domains of MeCP2 that have been shown to either bind methylated DNA or interact with a transcriptional co-repressor complex. Several mutations, however, including the C-terminal truncations that account for ~10% of cases, fall outside these characterised domains. We studied the molecular consequences of four of these “non-canonical” mutations in cultured neurons and mice to see if they reveal additional essential domains without affecting known properties of MeCP2. The results show that the mutations partially or strongly deplete the protein and also in some cases interfere with co-repressor recruitment. These mutations therefore impact the activity of known functional domains and do not invoke new molecular causes of Rett syndrome. The finding that a stable C-terminal truncation does not compromise MeCP2 function raises the possibility that small molecules which stabilise these mutant proteins may be of therapeutic value.

scholarly journals Pharmacological Chaperone Therapy for Pompe Disease

Molecules ◽  
2021 ◽  
Vol 26 (23) ◽  
pp. 7223
Author(s):  
Marc Borie-Guichot ◽  
My Lan Tran ◽  
Yves Génisson ◽  
Stéphanie Ballereau ◽  
Cécile Dehoux
Keyword(s):  
Motor Neurons ◽  
Pompe Disease ◽  
Storage Disease ◽  
Protein Stabilization ◽  
Lysosomal Storage ◽  
Pharmacological Chaperone ◽  
Pharmacological Chaperones ◽  
Enzyme Replacement ◽  
Alpha Glucosidase ◽  
Gaa Gene

Pompe disease (PD), a lysosomal storage disease, is caused by mutations of the GAA gene, inducing deficiency in the acid alpha-glucosidase (GAA). This enzymatic impairment causes glycogen burden in lysosomes and triggers cell malfunctions, especially in cardiac, smooth and skeletal muscle cells and motor neurons. To date, the only approved treatment available for PD is enzyme replacement therapy (ERT) consisting of intravenous administration of rhGAA. The limitations of ERT have motivated the investigation of new therapies. Pharmacological chaperone (PC) therapy aims at restoring enzymatic activity through protein stabilization by ligand binding. PCs are divided into two classes: active site-specific chaperones (ASSCs) and the non-inhibitory PCs. In this review, we summarize the different pharmacological chaperones reported against PD by specifying their PC class and activity. An emphasis is placed on the recent use of these chaperones in combination with ERT.

scholarly journals Structural snapshots of GALC: The Krabbe Disease enzyme

2014 ◽  
Vol 70 (a1) ◽  
pp. C451-C451
Author(s):  
Chris Hill ◽  
Stephen Graham ◽  
Samantha Spratley ◽  
Randy Read ◽  
Janet Deane
Keyword(s):  
Conformational Changes ◽  
Active Site ◽  
Rational Design ◽  
Specific Binding ◽  
Genetic Disorder ◽  
Krabbe Disease ◽  
Missense Mutations ◽  
Pharmacological Chaperone ◽  
Trafficking Defects ◽  
Key Steps

Glycosphingolipids are ubiquitous components of mammalian cell membranes, and defects in their catabolism by lysosomal enzymes cause a diverse array of diseases. Deficiencies in the enzyme β-galactocerebrosidase (GALC) cause Krabbe disease, a devastating genetic disorder characterized by widespread demyelination and rapid, fatal neurodegeneration[1]. Many disease-causing mutations throughout the GALC gene have been identified; rather than rendering the enzyme catalytically incompetent, many of these are predicted to destabilise the protein, compromising lysosomal delivery by causing retention in the endoplasmic reticulum. In these cases, pharmacological chaperone therapy (PCT) may be an appropriate treatment strategy, whereby the specific binding of a small molecule may stabilise folding of the mutant protein sufficiently to correct trafficking defects. Here we present a series of high resolution crystal structures that illustrate key steps in the catalytic cycle of GALC, including enzyme-substrate, enzyme-intermediate and enzyme-product complexes[2]. These structures reveal active site conformational changes accompanying the catalytic steps and provide key mechanistic insights. Building on this, we identify two candidate molecules that stabilise GALC by specific binding to the active site. The nature of several disease-causing missense mutations is also explored using secretion and activity assays, and we show that mutations can be classified into two groups based on defects in catalysis or trafficking. Overall, this study provides an atomic framework for the rational design of GALC pharmacological chaperones, and helps to identify which mutations may be suitable for PCT, bringing the prospect of a drug-based therapy one step closer.

scholarly journals Investigation of the Stability of Yeast rad52 Mutant Proteins Uncovers Post-translational and Transcriptional Regulation of Rad52p

Genetics ◽  
2003 ◽  
Vol 163 (1) ◽  
pp. 91-101 ◽  
Author(s):  
Erin N Asleson ◽  
Dennis M Livingston
Keyword(s):  
Half Life ◽  
Translational Regulation ◽  
Cellular Level ◽  
Missense Mutations ◽  
Wild Type ◽  
Mutant Proteins ◽  
Gal1 Promoter ◽  
Rad52 Protein ◽  
The Stability ◽  
Mutant Forms

Abstract We investigated the stability of the Saccharomyces cerevisiae Rad52 protein to learn how a cell controls its quantity and longevity. We measured the cellular levels of wild-type and mutant forms of Rad52p when expressed from the RAD52 promoter and the half-lives of the various forms of Rad52p when expressed from the GAL1 promoter. The wild-type protein has a half-life of 15 min. rad52 mutations variably affect the cellular levels of the protein products, and these levels correlate with the measured half-lives. While missense mutations in the N terminus of the protein drastically reduce the cellular levels of the mutant proteins, two mutations—one a deletion of amino acids 210-327 and the other a missense mutation of residue 235—increase the cellular level and half-life more than twofold. These results suggest that Rad52p is subject to post-translational regulation. Proteasomal mutations have no effect on Rad52p half-life but increase the amount of RAD52 message. In contrast to Rad52p, the half-life of Rad51p is >2 hr, and RAD51 expression is unaffected by proteasomal mutations. These differences between Rad52p and Rad51p suggest differential regulation of two proteins that interact in recombinational repair.

scholarly journals Mutation-specific non-canonical pathway of PTEN as a distinct therapeutic target for glioblastoma

2021 ◽  
Vol 12 (4) ◽  
Author(s):  
Seung Won Choi ◽  
Yeri Lee ◽  
Kayoung Shin ◽  
Harim Koo ◽  
Donggeon Kim ◽  
...  
Keyword(s):  
Functional Properties ◽  
Therapeutic Target ◽  
Medical Center ◽  
Dominant Negative ◽  
The Cancer Genome Atlas ◽  
Dominant Negative Effect ◽  
Cell Periphery ◽  
Missense Mutations ◽  
Phosphatase Domain ◽  
Mutant Proteins

AbstractPTEN is one of the most frequently altered tumor suppressor genes in malignant tumors. The dominant-negative effect of PTEN alteration suggests that the aberrant function of PTEN mutation might be more disastrous than deletion, the most frequent genomic event in glioblastoma (GBM). This study aimed to understand the functional properties of various PTEN missense mutations and to investigate their clinical relevance. The genomic landscape of PTEN alteration was analyzed using the Samsung Medical Center GBM cohort and validated via The Cancer Genome Atlas dataset. Several hotspot mutations were identified, and their subcellular distributions and phenotypes were evaluated. We established a library of cancer cell lines that overexpress these mutant proteins using the U87MG and patient-derived cell models lacking functional PTEN. PTEN mutations were categorized into two major subsets: missense mutations in the phosphatase domain and truncal mutations in the C2 domain. We determined the subcellular compartmentalization of four mutant proteins (H93Y, C124S, R130Q, and R173C) from the former group and found that they had distinct localizations; those associated with invasive phenotypes (‘edge mutations’) localized to the cell periphery, while the R173C mutant localized to the nucleus. Invasive phenotypes derived from edge substitutions were unaffected by an anti-PI3K/Akt agent but were disrupted by microtubule inhibitors. PTEN mutations exhibit distinct functional properties regarding their subcellular localization. Further, some missense mutations (‘edge mutations’) in the phosphatase domain caused enhanced invasiveness associated with dysfunctional cytoskeletal assembly, thus suggesting it to be a potent therapeutic target.

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