Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2024 May 20;19(5):e0303789.
doi: 10.1371/journal.pone.0303789. eCollection 2024.

Discovery of allosteric regulators with clinical potential to stabilize alpha-L-iduronidase in mucopolysaccharidosis type I

Elena Cubero  1 , Ana Ruano  1 , Aida Delgado  1 , Xavier Barril  1   2   3 , Sara Morales  1 , Ana Trapero  1 , Lorenzo Leoni  4 , Manolo Bellotto  4 , Roberto Maj  4 , Beatriz Calvo-Flores Guzmán  4 , Natalia Pérez-Carmona  1 , Ana Maria Garcia-Collazo  1
Affiliations

Discovery of allosteric regulators with clinical potential to stabilize alpha-L-iduronidase in mucopolysaccharidosis type I

Elena Cubero et al. PLoS One. .

Abstract

Mucopolysaccharidosis type I (MPS I) is an inherited lysosomal disease caused by lowered activity of the enzyme alpha-L-iduronidase (IDUA). Current therapeutic options show limited efficacy and do not treat some important aspects of the disease. Therefore, it may be advantageous to identify strategies that could improve the efficacy of existing treatments. Pharmacological chaperones are small molecules that protect proteins from degradation, and their use in combination with enzyme replacement therapy (ERT) has been proposed as an alternative therapeutic strategy. Using the SEE-Tx® proprietary computational drug discovery platform, a new allosteric ligand binding cavity in IDUA was identified distal from the active site. Virtual high-throughput screening of approximately 5 million compounds using the SEE-Tx® docking platform identified a subset of small molecules that bound to the druggable cavity and functioned as novel allosteric chaperones of IDUA. Experimental validation by differential scanning fluorimetry showed an overall hit rate of 11.4%. Biophysical studies showed that one exemplary hit molecule GT-01803 bound to (Kd = 22 μM) and stabilized recombinant human IDUA (rhIDUA) in a dose-dependent manner. Co-administration of rhIDUA and GT-01803 increased IDUA activity in patient-derived fibroblasts. Preliminary in vivo studies have shown that GT-01803 improved the pharmacokinetic (PK) profile of rhIDUA, increasing plasma levels in a dose-dependent manner. Furthermore, GT-01803 also increased IDUA enzymatic activity in bone marrow tissue, which benefits least from standard ERT. Oral bioavailability of GT-01803 was found to be good (50%). Overall, the discovery and validation of a novel allosteric chaperone for rhIDUA presents a promising strategy to enhance the efficacy of existing treatments for MPS I. The compound's ability to increase rhIDUA activity in patient-derived fibroblasts and its good oral bioavailability underscore its potential as a potent adjunct to ERT, particularly for addressing aspects of the disease less responsive to standard treatment.

PubMed Disclaimer

Conflict of interest statement

Elena Cubero, Ana Ruano, Aida Delgado, Xavier Barril, Ana Trapero, Manolo Bellotto, Beatriz Calvo-Flores Guzmán, Natalia Pérez-Carmona and Ana Maria Garcia-Collazo are employees of Gain Therapeutics Sucursal en España or GT Gain Therapeutics SA. Sara Morales, Roberto Maj, past employees, and Lorenzo Leoni, scientific adviser, declare no conflicts of interest.

Figures

Fig 1
Fig 1. The SEE-Tx® methodology uses a structure-based approach to discover non-competitive pharmacological allosteric regulators of alpha-L-iduronidase (IDUA).
Self-explanatory graphic of the procedure used to discover new allosteric regulators for alpha-L-iduronidase.
Fig 2
Fig 2. Binding of small molecule hit compounds to rhIDUA as determined by DSF.
Difference in melting temperature (ΔTm) relative to rhIDUA in the presence of the compounds #1 to #21 at 30 μM. The mean ΔTm values ± standard deviations are from 2 independent experiments (n = 2). Most compounds have a shift in Tm relative to the baseline suggesting protein stabilization. The dotted line shows the threshold value for the DSF screening, and it was ΔTm ≥ 0.5ºC.
Fig 3
Fig 3. Chemical structure of hit compound #18 (GT-01803).
Fig 4
Fig 4. Differential scanning fluorimetry curve shows the thermal shift of recombinant human alpha-L-iduronidase (rhIDUA) with increasing doses of compound GT-01803.
Dose-dependent effect on thermal stability of alpha-L-iduronidase (rhIDUA) in the presence of compound #18 (GT-01803). The mean ΔTm values ± standard deviations are from 2 independent experiments (n = 2).
Fig 5
Fig 5. Isothermal denaturation assay to determine ligand-induced stabilisation of recombinant human alpha-L-iduronidase (rhIDUA) by GT-01803 (30 μM) at 37 °C.
The percentage of IDUA folded values ± standard errors of the mean are from 2 independent experiments (n = 2). The best-fit value for the half-life of rhIDUA in the absence and presence of the compound was determined using GraphPad Prism software.
Fig 6
Fig 6. IDUA cell activity following co-administration of rhIDUA without or with 50 μM GT-01803.
(A) at different concentrations of rhIDUA in Hurler-Scheie (GM02845) fibroblasts at 96 h; (B) in Hurler (GM00798) and Hurler/Scheie (GM02845, GM00512, GM01898, GM00963) patient-derived fibroblasts at 96 h; and (C) in Hurler-Scheie fibroblasts (GM02845) at increasing incubation times. Individual values of IDUA activity (nmol/h·mg) are represented in the graphs. Repeated Measured Two-way ANOVA with the Geisser-Greenhouse correction and Šídák’s multiple comparisons test were used to compare the groups. Statistically significant differences are represented *p≤0.05; **p≤0.01 ***p≤0.001; ****p≤0.0001. h, hours.
Fig 7
Fig 7. Mean plasma concentration-time profiles data of GT-01803.
Plasma pharmacokinetics parameters are calculated using the non-compartmental analysis tool of Phoenix WinNonlin (Version 7.0) following single intravenous (i.v.) and oral (p.o.) administration doses in male C57BL/6 mice.
Fig 8
Fig 8
Enzymatic activity of rhIDUA with or without GT-01803 in plasma (A) and in bone marrow (B). (A) mice administered with rhIDUA + vehicle (black circles) and rhIDUA + GT-01803 (at 5 mg/kg, 10 mg/kg and 20 mg/kg i.v. q.d.) (blue square, triangle, and rhombus, respectively), in plasma at 0.5, 1 and 2 h after treatment with GT-01803. (B) untreated naïve control mice (grey bar), mice administered with rhIDUA + vehicle (black bars) and rhIDUA + GT-01803 (at 5 mg/kg, 10 mg/kg and 20 mg/kg i.v. q.d.) (blue bars), in bone marrow at 6, 8 and 10 h after treatment with GT-01803. Data expressed as mean ± SEM, n = 6.; ***p<0.01 vs vehicle-treated mice; Vehicle: NMP (5%) + Solutol HS-15 (5%) + saline solution (45%) + PEG-400 (45%) for GT-01803.
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Beck M, Arn P, Giugliani R, et al.. The natural history of MPS I: global perspectives from the MPS I Registry. Genet Med. 2014; 16(10):759–65. doi: 10.1038/gim.2014.25 - DOI - PMC - PubMed
    1. Giugliani R, Federhen A, Rojas MV, et al.. Mucopolysaccharidosis I, II, and VI: Brief review and guidelines for treatment. Genet Mol Biol. 2010; 33(4):589–604. doi: 10.1590/S1415-47572010005000093 - DOI - PMC - PubMed
    1. Hampe CS, Eisengart JB, Lund TC, et al.. Mucopolysaccharidosis Type I: A Review of the Natural History and Molecular Pathology. Cells. 2020; 9(8). doi: 10.3390/cells9081838 - DOI - PMC - PubMed
    1. Çelik B, Tomatsu SC, Tomatsu S, Khan SA. Epidemiology of Mucopolysaccharidoses Update. Diagnostics (Basel). 2021; 11(2). doi: 10.3390/diagnostics11020273 - DOI - PMC - PubMed
    1. de Ru MH, Boelens JJ, Das AM, et al.. Enzyme replacement therapy and/or hematopoietic stem cell transplantation at diagnosis in patients with mucopolysaccharidosis type I: results of a European consensus procedure. Orphanet J Rare Dis. 2011; 6:55. doi: 10.1186/1750-1172-6-55 - DOI - PMC - PubMed

Substances

Grants and funding

This study was funded by the Eurostars-2 joint program with co-funding from the European Union Horizon 2020 research and Innosuisse – Swiss Innovation Agency [E!113321_CHAPERONE]. The funders had no role in study design, data collection and analysis, publication decisions, or manuscript preparation.