Computational predictions of binding affinities to dihydrofolate reductase: Synthesis and biological evaluation of methotrexate analogues

The relative binding affinities to human dihydrofolate reductase of four ne w potential antifolates, containing ester linkages between the two aromatic systems, were estimated by free energy perturbation simulations. The ester analogue, predicted to exhibit the highest binding affinity to human dihyd rofolate reductase, and a reference ester (more structurally related to met hotrexate) were synthesized. As deduced from the measured IC50 values, the calculated ranking of the ligands was correct although a greater difference in affinity was indicated by the experimental measurements. Among the new antifolates the most potent inhibitor exhibited a similar pharmacokinetic p rofile to methotrexate but lacked activity in a complex antiarthritic model in rat in vivo.