If on one hand it is indeed encouraging that MD replicates are able to collectively sample the two ligand poses with the same probability obtained in HREM simulations investing the same amount of CPU time, on the other hand its quite disheartening that we needed, in total, 288 ns of standard MD simulation for such result. In a FEP approach, there is no reason to believe that such amount of time is required only to sample effectively the initial state of the ligand-receptor complex. If we change the nature of the ligand receptor interactions, as it occurs in the alchemical intermediate states of a standard FEP calculation, we very likely alter the probability ratio of the two poses, with an impact of the free energy change. For example, the oxygen of the isoxazole moiety of ODR is engaged in a strong hydrogen bond with a hydrogen of the NH 2 group of ASN100 in the primary pose (see Figure ). This H-bond is lost when the ligand switches to the secondary pose.