A robust strategy is reported to build perfectly monodisperse star polycations combining a trehalose‐based cyclooligosaccharide (cyclotrehalan, CT) central core onto which oligoethyleneimine radial arms are installed. The architectural perfection of the compounds is demonstrated by a variety of physicochemical techniques, including NMR, MS, DLS, TEM, and GPC. Key to the strategy is the possibility of customizing the cavity size of the macrocyclic platform to enable/prevent the inclusion of adamantane motifs. These properties can be taken into advantage to implement sequential levels of stimuli responsiveness by combining computational design, precision chemistry and programmed host‐guest interactions. Specifically, it is shown that supramolecular dimers implying a trimeric CT‐tetraethyleneimine star polycation and purposely designed bis‐adamantane guests are preorganized to efficiently complex plasmid DNA (pDNA) into transfection‐competent nanocomplexes. The stability of the dimer species is responsive to the protonation state of the cationic clusters, resulting in dissociation at acidic pH. This process facilitates endosomal escape, but reassembling can take place in the cytosol then handicapping pDNA nuclear import. By equipping the ditopic guest with a redox‐sensitive disulfide group, recapturing phenomena are prevented, resulting in drastically improved transfection efficiencies both in vivo and in vitro.

The nuclear export receptor exportin-1 (XPO1, CRM1) mediates the nuclear export of proteins that contain a leucine-rich nuclear export signal (NES) towards the cytoplasm. XPO1 is considered a relevant target in different human diseases, particularly in hematological malignancies, tumor resistance, inflammation, neurodegeneration and viral infections. Thus, its pharmacological inhibition is of significant therapeutic interest. The best inhibitors described so far (leptomycin B and SINE compounds) interact with XPO1 through a covalent interaction with Cys528 located in the NES-binding cleft of XPO1. Based on the well-established feature of chalcone derivatives to react with thiol groups via hetero-Michael addition reactions, we have synthesized two series of chalcones. Their capacity to react with thiol groups was tested by incubation with GSH to afford the hetero-Michael adducts that evolved backwards to the initial chalcone through a retro-Michael reaction, supporting that the covalent interaction with thiols could be reversible. The chalcone derivatives were evaluated in antiproliferative assays against a panel of cancer cell lines and as XPO1 inhibitors, and a good correlation was observed with the results obtained in both assays. Moreover, no inhibition of the cargo export was observed when the two prototype chalcones 9 and 10 were tested against a XPO1-mutated Jurkat cell line (XPO1C528S), highlighting the importance of the Cys at the NES-binding cleft for inhibition. Finally, their interaction at the molecular level at the NES-binding cleft was studied by applying the computational tool CovDock.