Abstract
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.
Highlights
Chemical modifications of RNA have a critical impact on many cellular functions, such as proliferation, survival, and differentiation [1,2]
These include m6 A writers such as the methyltransferase-like protein 16 (METTL16) [5] as well as the RNA methyltransferase enzyme complex METTL3/METTL14/WTAP usually composed of three components: METTL3, METTL14
We identified unique small-molecule ligands that bind to fat mass and obesity-associated protein (FTO)
Summary
Chemical modifications of RNA have a critical impact on many cellular functions, such as proliferation, survival, and differentiation [1,2]. The precise role of the RNA demethylases FTO and ALKBH5 in the regulation of survival and regeneration of DA neurons has remained enigmatic One reason for this is the lack of highly specific inhibitors of these enzymes. Two ALKBH5 inhibitors that we have described earlier [32], were less potent in rescuing DA neurons This is the first demonstration that inhibitors of FTO, and in general m6 A regulators can support the survival and protect dopamine neurons from growth factor deprivation induced death in vitro. These compounds may further serve as lead molecules for development of novel drugs for neurodegenerative diseases such as PD. These compounds may further serve as lead molecules for development of novel drugs for neurodegenerative dis of 15 eases such as PD
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