BIX 02189 of the van der Waals differences footprints shows

xamination that for all inhibitors a methionine at position 790 is energetically accommodated in the pocket BIX 02189 and steric packing interactions localized to this position in fact become more favorable as a result of the double mutation. Increased packing as a result of T790M is physically reasonable and occurs as a result of the hydrophilic to hydrophobic substitution. Although other van der Waals changes are less readily explained, the H805 increase with AEE788 coincides with the previously noted piperazine H bond. Compellingly, erlotinib and AEE788 show significant losses in ΔΔEvdw in contrast to gefitinib, which likely contributes to these compounds being more affected by the double mutations.
Water Mediated Interactions Examination of the underlying explicit solvent TIP3P MD trajectories, used subsequently for continuum based free energy calculations, revealed water molecules which appear to be important for positioning of ligands in the binding pocket. High water occupancy is observed at two primary positions, AM-1241 termed site 1 and site 2, as shown in Figure 12a for erlotinib with L858R and the double mutant, which are representative. Figure 12b quantifies S1 and S2 populations for all six inhibitor simulations with averages total count/5000 frames. Site waters were defined as present if a water hydrogen was within 2.5 Å of each ligand,s relevant nitrogen acceptor or residue Q791 at O. Importantly, the MD simulations reproduce the crystallographically observed water at S1 for all ligands.
The water at both sites are observed in the crystal structure of AEE788 with EGFR . For all ligands with L858R, waters are present 50 90% at S1 and 80% at S2 which indicates these are long lived significant interactions. As shown in Figure 12a, these waters are involved in a quadrifurcated H bonding network involving the ligands with three nearby residues, including the site of the known drug resistance mutation T790M. Notably, in all cases, occupancy at S1 and S2 is reduced as a result of L858R&T790. As an alternative metric, energy calculations reveal favorable Coulombic interactions between pocket waters and amino acids in the H bond network including the ligands. Here, the two waters closest to each ligand at N were used define key pocket waters.
Interestingly the L858R&T790M mutant leads to changes in bridging water interactions with each ligand that roughly mirror trends in the experimental FR data with erlotinib and AEE788 both being Balius and Rizzo Page 10 Biochemistry. Author manuscript, available in PMC 2010 September 8. NIH PA Author Manuscript NIH PA Author Manuscript NIH PA Author Manuscript adversely affected compared to gefitinib. Favorable electrostatic interactions between these waters and residue 790 are similarly reduced as a result of the double mutant, particularly for erlotinib, and thus expected to lead to weaker protein ligand binding. Further, despite the fact that some water mediated H bonding with M790 is observed, an overall weaker network would be expected due to the fact that sulfur is a weaker H bond acceptor than oxygen. Overall, the energetic description is consistent with the reduced population counts suggesting weaker interactions in the drug resistant mutant. H bonding between quinazoline based inhibitors and binding site waters were previously predicted by Wissner et al.