Hexitol nucleic acids (HNA) are oligonucleotides built up from natural nucleobases and a phosphorylated 1,5-anhydrohexitol backbone. Molecular associations between HNA and RNA are more stable than between HNA and DNA and between natural nucleic acids (dsDNA, dsRNA, DNA/RNA). 1H NMR analysis of a HNA dimer confirms the axial orientation of the base moiety with respect to the hexitol ring, and this was used as starting conformation for a molecular dynamics study of HNA/RNA and HNA/DNA duplexes. Both complexes show an A-type geometry and very similar hydrogen bonding patterns between base pairs. The relative stability of HNA/RNA versus HNA/DNA might be explained by a difference in minor groove solvatation.

Fibroblast activation protein (FAP) is a proline-selective serine protease. It is hardly expressed in healthy adult tissue but upregulated in tissue remodeling sites associated with several diseases including epithelial cancer types, atherosclerosis, arthritis and fibrosis. Ongoing research aims at clinical implementation of FAP as a biomarker for these diseases. Several immunochemical methods that quantify FAP expression have been reported. An alternative/complementary approach focuses on quantification of FAP’s enzymatic activity. Developing an activity-based assay for FAP has nonetheless proven challenging because of selectivity issues with respect to prolyl oligopeptidase (PREP). Here, we present substrate-type FAP probes that are structurally derived from a FAP-inhibitor (UAMC1110) that we published earlier. Both cleavage efficiency and FAP-selectivity of the best compounds in the series equal or surpass the most advanced peptide-based FAP substrates reported to date. Finally, proof-of-concept is provided that 4-aminonaphthol containing probes can spatially localize FAP activity in biological samples.

Urokinase plasminogen activator (uPA) is a biomarker and therapeutic target for several cancer types. Its inhibition is regarded as a promising, noncytotoxic approach in cancer therapy by blocking growth and/or metastasis of solid tumors. Earlier, we reported the modified substrate activity screening (MSAS) approach and applied it for the identification of fragments with affinity for uPA's S1 pocket. Here, these fragments are transformed into a novel class of uPA inhibitors with an imidazo[1,2-a]pyridine scaffold. The SAR for uPA inhibition around this scaffold is explored, and the best compounds in the series have nanomolar uPA affinity and selectivity with respect to the related trypsin-like serine proteases (thrombin, tPA, FXa, plasmin, plasma kallikrein, trypsin, FVIIa). Finally, the approach followed for translating fragments into small molecules with a decorated scaffold architecture is conceptually straightforward and can be expected to be broadly applicable in fragment-based drug design.

Vildagliptin is a marketed DPP4 inhibitor, used in the management of type 2 diabetes. The molecule also has notable DPP8/9 affinity, with some preference for DPP9. Therefore, we aimed to use vildagliptin as a starting point for selective DPP8/9 inhibitors, and to engineer out the parent compound's DPP4affinity. In addition, we wanted to identify substructures in the obtained molecules that allow their further optimization into inhibitors with maximal DPP9 selectivity. Various 2S-cyanopyrrolidines and isoindoline were investigated as P1 residues of vildagliptin analogs. The obtained set was expanded with derivatives bearing O-substituted, N-(3-hydroxyadamantyl)glycine moieties at the P2 position. In this way, representatives were discovered with DPP8/9 potencies comparable to the parent molecule, but with overall selectivity towards DPP4, DPP2, FAP, and PREP. Furthermore, the most promising molecules in this series have a 4-to 7-fold preference for DPP9 over DPP8. Finally, a molecular dynamics study was carried out to maximize our insight into experimental selectivity data.