Inhibition studies of DNA methyltransferases by maleimide derivatives of RG108 as non-nucleoside inhibitors

Our plain-English summary of Inhibition studies of DNA methyltransferases by maleimide derivatives of RG108 as non-nucleoside inhibitors, by G. Rondelet, L. Fleury, C. Faux, et al (Future Medicinal Chemistry, 2017).

The study

• Authors: G. Rondelet, L. Fleury, C. Faux, V. Masson, J. Dubois, P.B. Arimondo, L. Willems, J. Wouters
• Journal: Future Medicinal Chemistry
• Published: 2017;9(13):1465-1481
• Cited by: 23 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Aim: DNA methyltransferases (DNMTs) are important drug targets for epigenetic therapy of cancer. Nowadays, non-nucleoside DNMT inhibitors are in development to address high toxicity of nucleoside analogs. However, these compounds still have low activity in cancer cells and mode of action of these compounds remains unclear.

Materials & Methods: In this work, we studied maleimide derivatives of RG108 by biochemical, structural and computational approaches to highlight their inhibition mechanism on DNMTs.

Results: Findings demonstrated a correlation between cytotoxicity on mesothelioma cells of these compounds and their inhibitory potency against DNMTs. Noncovalent and covalent docking studies, supported by crystallographic (apo structure of M.HhaI) and differential scanning fluorimetry assays, provided detailed insights into their mode of action and revealed essential residues for the stabilization of such compounds inside DNMTs. [Formula: see text].

In plain English

This laboratory study examined two maleimide derivatives of the non-nucleoside DNA methyltransferase (DNMT) inhibitor RG108, called RG108-1 and RG119-1, using enzyme assays, cell viability tests, X-ray crystallography, differential scanning fluorimetry and molecular docking. Against the enzymes tested, RG119-1 gave IC50 values of 19 ± 2 micromolar for human DNMT1, 3.2 ± 0.5 for human DNMT3A and 4.5 ± 0.7 for bacterial M.SssI, while RG108-1 gave 28 ± 5 (DNMT3A) and 28 ± 27 (M.SssI), with no value determined for DNMT1. In the mesothelioma cell lines M14K and H28, RG119-1 gave TC50 values of 5.4 ± 1.5 and 6.2 ± 1.4 micromolar, whereas RG108-1 had no marked inhibitory effect. Fluorimetry showed RG119-1 stabilised DNMT1 in the presence of DNA (change in Tm about 2.8 degrees C), while RG108-1 did not stabilise DNMT1 in any form. The authors solved an apo crystal structure of M.HhaI at 1.9 Angstrom, but neither compound appeared in the electron density map. The authors concluded their cytotoxicity on mesothelioma cells correlated with their DNMT inhibitory activity, and that docking identified residues orienting the maleimide warhead toward the catalytic cysteine.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) study combining enzyme inhibition assays, cell viability assays, protein crystallography, differential scanning fluorimetry and molecular docking
• Who was studied: Purified enzymes (full-length human DNMT1, catalytic human DNMT3A, bacterial M.SssI and M.HhaI) and two malignant mesothelioma cell lines, M14K and H28; work carried out in Belgium and France
• Exposure: Two maleimide derivatives of RG108: RG108-1 (racemic, purchased) and RG119-1 (a tryptamine derivative lacking the carboxylate), tested over concentration ranges of 0–10 mM (RG108-1) and 0–1 mM (RG119-1) on cells
• Main results:
  • Enzyme inhibition: RG119-1 IC50 19 ± 2 micromolar (DNMT1), 3.2 ± 0.5 (DNMT3A), 4.5 ± 0.7 (M.SssI); RG108-1 28 ± 5 (DNMT3A), 28 ± 27 (M.SssI), not defined for DNMT1
  • In mesothelioma cells, RG119-1 had TC50 values of 5.4 micromolar (M14K) and 6.2 micromolar (H28)
  • RG108-1 showed no marked inhibitory effect on either mesothelioma cell line
  • Differential scanning fluorimetry: RG108-1 did not stabilise DNMT1 in any form
  • Differential scanning fluorimetry: RG119-1 produced a thermal shift of about 2.8 degrees C for DNMT1 in the presence of DNA
  • Crystallography yielded an apo structure of M.HhaI at 1.90 Angstrom resolution (space group P 1 21 1; deposited as PDB 5LOD), with a vacant SAM-binding pocket and slight backbone displacement of Glu40, Trp41, Asp60 and Ile 61
  • Docking: both compounds preferentially occupied the cofactor pocket with the maleimide group directed toward the catalytic cysteine; the carboxylate of RG108-1 formed a salt bridge with Lys1247 and a hydrogen bond with Gln1230 in DNMT1
• The authors' conclusion: The authors concluded that cytotoxicity of these maleimide analogs on mesothelioma cells correlated with their DNMT inhibitory activity, that the apo M.HhaI structure suggested a mode of action competitive with the SAM cofactor, and that docking highlighted residues stabilising the covalent complex
• Limitations the authors state:
  • The authors could not obtain a crystal structure of M.HhaI in complex with either inhibitor, and neither compound was visible in the electron density map, which they attributed to high mobility and low occupancy
  • The IC50 of RG108-1 against human DNMT1 could not be determined because dose-response curves were incomplete
  • No preferential docking pose was obtained for the catalytic domain of mouse DNMT3A

Topics

Medical terms in this study, explained in our Lexicon: Mesothelioma.

Related studies

• Gemcitabine reactivates epigenetically silenced genes and functions as a DNA methyltransferase inhibitor — S.G. Gray, A.M. Baird, et al. (2012)
• Inhibition of EZH2 methyltransferase decreases immunoediting of mesothelioma cells by autologous macrophages through a PD-1-dependent mechanism — M. Hamaidia, H. Gazon, et al. (2019)
• A first-in-human phase I dose-escalation, pharmacokinetic, and pharmacodynamic evaluation of intravenous LY2090314, a glycogen synthase kinase 3 inhibitor, administered in combination with pemetrexed and carboplatin — J.E. Gray, J.R. Infante, et al. (2015)
• A Succinate Ether Derivative of Tocotrienol Enhances Dickkopf-1 Gene Expression through Epigenetic Alterations in Malignant Mesothelioma Cells — A. Sato, H. Ueno, et al. (2018)
• Decitabine, differently from DNMT1 silencing, exerts its antiproliferative activity through p21 upregulation in malignant pleural mesothelioma (MPM) cells — S. Amatori, F. Papalini, et al. (2009)
• A concise synthesis of pyrazole analogues of combretastatin A1 as potent anti-tubulin agents — R. Zaninetti, S.V. Cortese, et al. (2013)

Browse return to medical research.