Computer-aided design. With our first three series of POP inhibitors illustrated by compounds 2, 3 and 4, we have demonstrated the accuracy of our docking program FITTED in predicting binding modes of POP covalent inhibitors. When 2 and stereoisomers of 2 (adhering to the [3.3.0] bicyclic system) were evaluated, we found that the stereochemistry corresponding to that of Damino acids was optimal (hydrogen atom highlighted in blue in Figure , compound 2). The resultant stereochemistry upon cyclization (hydrogen atom highlighted in green in Figure ) at the ring junction fortunately imposed a shape that fit best in the binding site. In this previous report, computational studies also indicated that this [3.3.0] bicyclic system was less optimal for binding to POP, and that a [4.3.0]-ring system with a specific stereochemistry should exhibit better affinity (Figure , compound 2a). Unfortunately, our synthetic efforts were vain, as the epimer at the ring junction (hydrogen atom highlighted in green in Figure , compound 2a) was the only isomer observed experimentally but was not predicted to bind optimally in the active site of POP. Further computational predictions indicated that the affinity for POP could be improved by increasing the size of the western ring and inverting two stereocenters, both the carbon at the cyclic fusion (C7a in 2, C8a in 2a) and the carbon alpha to the cyclic amide (C6). To do so, we decided to prepare a first series of analogues built around a [4.3.0]-ring system similar to that of 2a but which could be accessible synthetically. After several rounds of virtual modifications and docking predictions, inhibitor structure 10a was discovered. As can be seen in Figure , the predicted binding mode of nitrile 10a is highly favored, featuring the same key interactions as potent aldehyde 1. Our previous inhibitor 2 and this newly-designed scaffold resemble previously-reported potent inhibitor 1 (Figure ). The bicyclic scaffolds 2 and 10 were introduced by virtually rigidifying 1 and introducing heterocyclic alkanes to both optimize the docking pose and ensure synthetic feasibility. The valine-based side chain of Talabostat (Figure ), a POP-FAP inhibitor that reached Phase III clinical trials, inspired the introduction of methyl groups into 10a and incorporation of the boronic acid warhead, leading to 12c and 13b. The complete list of new analogues selected for synthesis is provided in Table . Many POP inhibitors feature nitriles, activated nitriles (i.e., with proximal fluorine atoms), or boronic acids, the latter two of which are more electrophilic and lead to more potent FAP inhibition. Our current version of our docking program FITTED does not consider either the reactivity of the catalytic residue nor the reactive warhead. Nevertheless, a computational study from our group on the reactivity of the catalytic serine residues in both POP and FAP suggests that the catalytic serine in POP is more nucleophilic than that in FAP and that, as a result, nitrile derivatives are unlikely to act as potent covalent inhibitors of FAP, while boronic acids are promising alternatives. As a result, in our quest to develop dual POP/FAP inhibitors, the boronic ester or acid derivatives were also considered.