Discovery of METTL3 Small Molecule Inhibitors by Virtual Screening of Natural Products
Yue Du1,2, Yongliang Yuan1,2, Le Xu3
1Department of Pharmacy, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.
Abstract:
N6-Methyladenosine (m6A) is the most prevalent mRNA modification in mammalian cells that is mainly catalyzed by the methyltransferase complex of methyltransferase-like 3 and methyltransferase-like 14 (METTL3-METTL14). Many lines of evidence suggest that METTL3 plays important roles in several diseases such as cancers and viral infection. In the present study, 1,042 natural products from commercially available sources were chosen to establish a screening library, and docking-based high-throughput screening was performed to discover potential METTL3 inhibitors. The selected compounds were then further validated by an in vitro methyltransferase inhibition assay in which m6A content was determined by LC-MS/MS. A cellular assay of the inhibition of m6A methylation was performed to determine the METTL3 inhibitory activity of the selected compound. CCK-8 assay was applied to evaluate the effects of the selected compound on tumor cell viability. Additionally, binding mode analysis, molecular dynamics (MD) simulation, and binding free energy analysis were performed to study the process and characteristics of inhibitor binding. Finally, quercetin was identified as a METTL3 inhibitor with an IC50 value of 2.73 μM. The cellular assay of m6A methylation inhibition showed that quercetin decreased m6A level in a dose-dependent manner in MIA PaCa-2 pancreatic cancer cells. CCK-8 assay showed quercetin efficiently inhibited the proliferation of MIA PaCa-2 and Huh7 tumor cells, with IC50 values 73.51 ± 11.22 μM and 99.97 ± 7.03 μM, respectively. Molecular docking studies revealed that quercetin filled the pocket of the adenosine moiety of SAM but not the pocket of the SAM methionine in the METTL3 protein, and hydrogen bonds, hydrophobic interactions, and pi-stacking were formed. The values of the root mean square deviation (RMSD), the root mean square fluctuations (RMSF), and binding free energy suggested that quercetin can efficiently bind to the pocket of the METTL3 protein and form a stable protein-ligand complex. The present study is the first to identify METTL3 inhibitors from natural products, thus providing a basis for subsequent research and facilitating the development of METTL3-targeting drugs for diseases.
Insights
Researchers identified quercetin as a natural product inhibitor of METTL3, an enzyme crucial in diseases like cancer. This discovery offers a foundation for developing new METTL3-targeting cancer drugs.
Area of Science:
- Biochemistry
- Molecular Biology
- Pharmacology
Background:
- N⁶-Methyladenosine (m⁶A) is the most abundant mRNA modification, primarily regulated by the METTL3-METTL14 complex.
- METTL3 is implicated in various diseases, including cancers and viral infections, making it a potential therapeutic target.
Purpose of the Study:
- To screen natural products for potential METTL3 inhibitors.
- To validate and characterize the identified inhibitors for their therapeutic potential.
Main Methods:
- High-throughput screening of 1,042 natural products using molecular docking.
- In vitro methyltransferase inhibition assays (LC-MS/MS) and cellular m⁶A methylation assays.
- Cell viability assays (CCK-8), molecular dynamics (MD) simulations, and binding free energy analysis.
Main Results:
- Quercetin identified as a potent METTL3 inhibitor with an IC₅₀ of 2.73 μM.
- Quercetin dose-dependently reduced m⁶A levels in MIA PaCa-2 pancreatic cancer cells.
- Quercetin inhibited proliferation of MIA PaCa-2 and Huh7 tumor cells (IC₅₀ values 73.51 ± 11.22 μM and 99.97 ± 7.03 μM, respectively).
- Molecular docking and MD simulations confirmed stable binding of quercetin to METTL3.
Conclusions:
- This study is the first to identify natural product-derived METTL3 inhibitors.
- Quercetin demonstrates significant potential as a METTL3 inhibitor for cancer therapy.
- Provides a basis for developing novel METTL3-targeting drugs.
