Prediction of hERG Channel Inhibition Using In Silico Techniques - Ion Channels and Their Inhibitors - page 222

interactions with the hERG channel. They used 25 3,9-diazaspiro-[5.5]undecane
analogs and 8 hERG mutants. The PLS discriminant analysis (PLS-DA) revealed
that the desolvation component of hERG potency increases the block of the mutants
Tyr652Ala and Tyr652Phe, while the Thr623Ser and Ser624Thr mutants are less
sensitive to inhibitors. These results suggest that the desolvation component of
hERG potency is related to the lipophilicity of the compounds. Within the
nondesolvation-related interactions with the hERG channel the Thr623Ser,
Ser624Thr, Tyr652Ala, Phe656Met, Phe656Thr, and Phe656Trp mutants increase
hERG potency. This indicates that the lipophilic substituents cannot fit into the
inner pore due to clashes with Phe656. This hypothesis is confirmed by docking
studies on the homology model of hERG in the closed state, which predicts that the
molecules cannot fit well at the level of the four Phe656. All together these results
suggest that bulky and rigid substituents are detrimental for the hERG affinity.

5.4 Hydrogen Bonds with Ser624, Thr623 or Val625?

The amino acids Thr623, Ser624 and Val625 lie at the top of the inner cavity.
Dougherty et al. [ 111 ] mutated the amino acids facing the hERG pore to determine
the binding efficacy of astemizol, dofetilide, haloperidol, risperidone, droperidol,
pimozide, loxapine, amoxapine, imipramine, fluphenazine, triflupromazine, cis-
flupenthixol, and amperozide. Astemizole showed an increased IC 50 value with
the double flourinated Phe relative to the single flourinated Phe and the Tyr652Phe
mutants, indicating that Tyr652 interacts with astemizole through p -stacking

and/or -cation interactions. The mutation of Phe656 into the two flourinated

phenylalanines did not affect hERG channel block. This suggests that Phe656
may not be involved in the binding of astemizole, or that it forms hydrophobic
interactions. Mutation of Thr623 to the nonnatural amino acid Thr623-(OH) led
only to slight increase of the binding affinity, while the Thr623Val mutant is less
sensitive to astemizole. The Ser624Thr mutation slightly decreased the astemizole
hERG block, while the affinity for the hERG channel was lost in the Ser624Ala
mutation. These results show the importance of the hydroxy group of Ser624.
Perry et al. [ 12 ] used site-directed mutagenesis to investigate the interactions of
clofilium and ibutilide with hERG channel. The authors performed an alanine
scanning of the amino acids of the S6 domain facing the pore of the hERG channel.
The results show that both blockers were affected by mutation of the amino acids
Thr623, Ser624, Val625, Gly648, Tyr652, Phe656, and Val659 to alanine.
Using the same method, Kamiya et al. [ 8 ] determined that the binding site of the
hERG blockers E-4031 and dofetilide consists of the amino acids Thr623, Ser624,
Val625, Tyr652, Phe656 and Val659. The same effect was obtained for the hERG
blocker nifekalant, except for the Val659Ala mutation that did not affect the
hERG inhibition and for the Ile655Ala that reduced channel block. The Thr623,
Ser624, Val625 and Phe656 mutations reduced the block of the hERG channel by
bepridil.

p