We used the popular 'single trajectory protocol' in this work, which involves simulating only the protein-ligand complex and re-purposing the bound conformations of the protein and ligand for the unbound calculations. It is assumed that the protein and the ligand sample similar conformations in the bound and unbound states which might not be always valid. We do not expect to see a big difference in predicted affinities when using the alternative 'multiple trajectory protocol'. However, it may be worth trying this in the future for the macrocycles in the current dataset since the macrocycles do not have much flexibility in the binding pocket due to their size, and might possibly sample other conformations when simulated in pure solvent.