## 3 Results and Discussion
### Molecular Dynamics Simulations  
The molecules that exhibited an ADMET risk \(\leq\)6 (**01**, **03**, **2**, **5**, **9**, **24**, and **48**) were subjected to MD simulations. Molecules **01** and **03** are matched molecular pairs of Scaffold 1 identified using our previously designed virtual screening workflow. The MD simulation results for **01** and **03** were reported in our previously published work [14].  
For the top hits from this work (**2**, **5**, **9**, **24**, and **48**), the interactions that were maintained during the 100 ns MD simulations between the ligands and FmIH are given in Figure 12. It was observed in the previous work by [PERSON] et al. that the most potent co-crystallized ligand did not show a \(\pi\)-\(\pi\) interaction with Y46 [14]. The novel scaffold in **2** exhibited \(\pi\)-\(\pi\) stacking with Y46, which was maintained for 63% of the simulation time (Figure 12A), a significant improvement over the most potent co-crystallized ligand [14]. In addition, a hydrogen bonding interaction was observed with the neighboring S10 residue. One of the sulfonamide oxygens of the terminal aglycone fragment interacted with the sidechain of the S10 residue in the previously reported research on FmIH-binding small molecule glycomimetics [14; 18]. In molecule **5**, the central novel scaffold is a non-aromatic moiety; therefore, a \(\pi\)-\(\pi\) stacking interaction with Y46 was not possible, but instead, there was hydrogen bonding of the carbonyl group with Y46. (Figure 12B). In molecule **9**, a water-mediated hydrogen bonding interaction was observed between the glycosidic linkage connecting the GalNAc and the novel Scaffold 1 moiety, which was maintained for 52% of the simulation time (not shown for clarity). A hydrogen bonding interaction was found between the primary amine group in the central scaffold in molecule **24** and residue D45 (Figure 12D). D45 was previously identified as an important residue involved in polar  
Figure 11: Numerical values of the predicted (**A**) hERG toxicity, mutagenicity risk, (**B**) acute rat toxicity (LD\({}_{50}\)), and Rat TD\({}_{50}\) and Mouse TD\({}_{50}\) values for the hits. The red line in (**A**) represents all the hits that exhibited hERG toxicity lower than the cut-off. The negative control (**00**) used in this study exhibited higher hERG toxicity. hERG_PLC\({}_{50}\) is given in mol/L. LD\({}_{50}\) for rat acute toxicity = mg/kg in the oral dose that would be lethal to 50% of the rats. Rat_TD\({}_{50}\) = mg/kg/day in the oral dose. Mouse_TD\({}_{50}\) = mg/kg/day in the oral dose.  
interactions with strong FmIH-binding glycomimetics [14; 18]. Finally, for molecule **48** (obtained after replacing Scaffold 2), the novel aglycone terminal scaffold exhibited a hydrogen bonding interaction with the neighboring R142 residue, which was maintained for 43% of the simulation time (Figure 12E). In all cases, a complex network of hydrogen bonding interactions between the GalNAc moiety and FmIH residues were maintained. During the molecular dynamics (MD) simulations, the top-ranked compounds did not dissociate from the binding site. The root-mean-square deviations (RMSD) of the five hits are shown in Figure S5. Compounds **5** and **9** exhibited lower RMSD variations than **2**, **24**, and **48** (Figure S5). All the ligands maintained low RMSD values throughout the MD simulation trajectories, indicating that they were stably bound in the FmIH binding pocket.  
Figure 12: MD simulation analysis of the 100 ns trajectories of the FmIH-hit complexes. (**A–D**) Hit glycomimetics from Scaffold 1 hopping, which exhibited improved ADMET profiles. (**E**) Hit glycomimetic from Scaffold 2 hopping, which showed an improved ADMET profile. The molecule number is indicated inside each box panel in bold. Blue N, red O, yellow S. Pink dashed lines indicate FmIH-ligand polar interactions, with the percent occupancy listed for one case in each image.