The 1 vs 2 fragment pair is top-ranked when using the ESP-Sim (B3LYP/6-31G**) metric, but not by the 2D-based methods. The Morgan fingerprints rank the 1 vs 2 pair among the top five (Table ), which is reasonably high. However, given the challenges and resource investments required to establish new synthetic routes, our experience is that very few alternative ring analogs are explored in real-life projects. Typically, only a couple of ring replacements are made and tested, essentially enforcing that only top-ranked scaffolds would be followed-up. Two other observations provide further support for the use of the ESP-Sim method. First, the MACCS keys fingerprint resulted in very similar values for many scaffolds (e.g., the Tanimoto similarity value for five scaffolds against scaffold of compound 1 show identical values -0.87), suggesting that the MACCS keys similarity metric is not sufficient for capturing such subtle differences. Second, there are a couple of clearly structurally dissimilar fragments in Figure As a final observation, deriving ESP-similarities with methods of lower theory for calculating the underlying partial charges (Gasteiger, MMFF and HF/3-21G, data not shown) also yielded the 1 vs 2 pair as top-ranked, suggesting that such partial charges may be sufficient and a cost-effective alternative for this data set. We recommend using a higher level of theory, although computationally more demanding. For example, RESP partial charges derived using the B3LYP method and the 6-31G** basis set or AM1-BCC partial charges, which were found to reproduce the QM electrostatic potentials best in our benchmark.