Model of P-Glycoprotein Ligand Binding and Validation with Efflux Substrate Matched Pairs

The blood–brain barrier (BBB) poses a significant obstacle in developing therapeutics for neurodegenerative diseases and central nervous system (CNS) disorders. P-glycoprotein (P-gp), a multidrug resistance protein, is a critical gatekeeper in the BBB and plays a role in cancer chemoresistance. This paper uses cryo-EM P-gp structures as starting points with an induced fit docking (IFD) model to evaluate 19 pairs of compounds with known P-gp efflux data. The study reveals significant differences in binding energy and sheds light on structural modifications’ impact on efflux properties. In the cases examined, fluorine incorporation influences the efflux by altering the molecular conformation rather than proximal heteroatom basicity. Although there are limitations in addressing covalent interactions or when binding extends into the more flexible vestibule region of the protein, the results provide valuable insights and potential strategies to overcome P-gp efflux, contributing to the advancement of drug development for both CNS disorders and cancer therapies.


Table of Contents
The difference in the torsional angle between the 3-F-phenyl and the tetrazole of -67° versus -37°, respectively, for the related torsion in 6b as well as the difference in steric bulk between the methyltetrazole and the methylcarboxylate largely account for the differences in the scores and the poses.

Figure S2 .
Figure S2.Compound 2a: (A) The pose for compound 2a displays no room for the -CF2H group in compound 2b.The surface of the adjacent residues F983 and M986 shows that the -CF2H substitution could not be accommodated by compound 2b in the same conformation or pose.(B) The 2D ligand-interaction diagram of compound 2a.

Figure S4 .Figure S5 .Figure S6 .BFigure S7 .Figure S8 .
Figure S4.Compound 2c: (A) F substitution in compound 2d would not be accommodated in this pose due to the positions of the adjoining residues M986 and F983.The orientation of the pocket is rotated approximately 90° around the vertical axis compared to previous orientations.(B) The 2D ligand-interaction diagram for compound 2c.

Figure S9 .
Figure S9.Model compounds used to calculate the torsion differences between a methyltetrazole and a methylester.The difference supports the observed pose difference between compounds 6a and 6b.

Figure S10 .
Figure S10.Compounds 6a and 6b are from a series of bradykinin B1 receptor antagonists.The lowest energy pose for 6a (orange, -46064) and the second-lowest energy pose for 6b (cyan, -46039) display a similar binding mode.The pose for 6a is stabilized by both NH groups H-bonded to the side chain amide C=O of Q725 as well as by pi-pi stacking between the biphenyl groups and the surrounding phenylalanine rings.The difference in the torsional angle between the 3-F-phenyl and the tetrazole of -67° versus -37°, respectively, for the related torsion in 6b as well as the difference in steric bulk between the methyltetrazole and the methylcarboxylate largely account for the differences in the scores and the poses.

Figure S11 .
Figure S11.Compounds 7a and 7b are from a series of serotonin and noradrenaline monoamine reuptake inhibitors.Compound 7a (orange) with a 4-tetrahyropyranyl group versus compound 7b (cyan) with an isobutyl group show different lowest energy poses.Compound 7a displays pi-stacking between the dichlorophenyl group and W232 and Hbonds with Q990.Compound 7b, with a more hydrophobic isobutyl, displays a cation-pi interaction with F343 and a pi-pi interaction between the dichlorophenyl group and F983.

Figure S12 .BFigure S13 .
Figure S12.(A)Compounds 8a (orange) and 8b (cyan) were synthesized as phosphodiesterase 10A (PDE10A) inhibitors.Chirality for compound 8a was not specified, and both enantiomers were used in the procedure.The R-enantiomer is displayed.The S-enantiomer shows a similar pose.(B) 8b lacks the phenyl-piperazine moiety and hence has a lower MW.Pi-pi interactions contribute heavily to the lowest energy pose for 8a (S).The quinoline ring in 8b interacts with multiple residues in the vestibule region, mainly via pi-pi interactions.