Highlights d High-binding-efficiency ligands of METTL3-14-WTAP are identified in silico d The binding mode of compounds overlaps with the METTL3-14-WTAP active site d The compounds are experimentally characterized as METTL3-14-WTAP activators d The compounds affect m 6 A level in mRNA and rRNA in cells
The RNA 6-N-methyladenosine
(m6A) demethylase ALKBH5 has been shown
to be oncogenic in several cancer types, including leukemia and glioblastoma.
We present here the target-tailored development and first evaluation
of the antiproliferative effects of new ALKBH5 inhibitors. Two compounds,
2-[(1-hydroxy-2-oxo-2-phenylethyl)sulfanyl]acetic acid (3) and 4-{[(furan-2-yl)methyl]amino}-1,2-diazinane-3,6-dione
(6), with IC50 values of 0.84 μM and 1.79 μM,
respectively, were identified in high-throughput virtual screening
of the library of 144 000 preselected compounds and subsequent
verification of hits in an m6A antibody-based enzyme-linked immunosorbent
assay (ELISA) enzyme inhibition assay. The effect of these compounds
on the proliferation of selected target cancer cell lines was then
measured. In the case of three leukemia cell lines (HL-60, CCRF-CEM,
and K562) the cell proliferation was suppressed at low micromolar
concentrations of inhibitors, with IC50 ranging from 1.38
to 16.5 μM. However, the effect was low or negligible in the
case of another leukemia cell line, Jurkat, and the glioblastoma cell
line A-172. These results demonstrate the potential of ALKBH5 inhibition
as a cancer-cell-type-selective antiproliferative strategy.
The fat mass and obesity-associated protein (FTO), an RNA N6-methyladenosine (m6A) demethylase, is an important regulator of central nervous system development, neuronal signaling and disease. We present here the target-tailored development and biological characterization of small-molecule inhibitors of FTO. The active compounds were identified using high-throughput molecular docking and molecular dynamics screening of the ZINC compound library. In FTO binding and activity-inhibition assays the two best inhibitors demonstrated Kd = 185 nM; IC50 = 1.46 µM (compound 2) and Kd = 337 nM; IC50 = 28.9 µM (compound 3). Importantly, the treatment of mouse midbrain dopaminergic neurons with the compounds promoted cellular survival and rescued them from growth factor deprivation induced apoptosis already at nanomolar concentrations. Moreover, both the best inhibitors demonstrated good blood-brain-barrier penetration in the model system, 31.7% and 30.8%, respectively. The FTO inhibitors demonstrated increased potency as compared to our recently developed ALKBH5 m6A demethylase inhibitors in protecting dopamine neurons. Inhibition of m6A RNA demethylation by small-molecule drugs, as presented here, has therapeutic potential and provides tools for the identification of disease-modifying m6A RNAs in neurogenesis and neuroregeneration. Further refinement of the lead compounds identified in this study can also lead to unprecedented breakthroughs in the treatment of neurodegenerative diseases.
The N6-methyladenosine
(m6A) modifications in both viral
and host cell RNAs play an important role in HIV-1 virus genome transcription
and virus replication. We demonstrate here that activators of the
METTL3/METTL14/WTAP RNA methyltransferase complex enhance the production
of virus particles in cells harboring HIV-1 provirus. In parallel,
the amount of m6A residues in the host cell mRNA was increased
in the presence of these activator compounds. Importantly, the m6A methylation of the HIV-1 RNA was also enhanced significantly
(about 18%). The increase of virus replication by the small-molecule
activators of the METTL3/METTL14/WTAP complex excludes them as potential
anti-HIV-1 drug candidates. However, the compounds may be of large
interest as activators for the latent HIV-1 provirus copies deposited
in host cells’ genome and the subsequent virus eradication
by an antiviral compound.
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