Ratiometric Cu2+ Binding, Cell Imaging, Mitochondrial Targeting, and Anticancer Activity with Nanomolar IC50 by Spiro-Indoline-Conjugated Calix[4]arene

A triazole-derivatized, spiro-indoline-linked, 1,3-di-derivative of calix[4]arene ( L ) has been synthesized to take advantage of its ion-binding capability in the ring-open form. Indeed, the spiro-indoline moiety is well known for its photochromic, acidochromic, and metallochromic properties. Therefore, the L has been explored for Cu 2+ binding, cell imaging, and anticancer activity of the corresponding complex since Cu 2+ complexes are known for such activity. The conversion from the closed to open form of L is expedited by light or proton, while the metal ion can open as well as stabilize it. The open form of L showed binding of Cu 2+ ratiometrically as demonstrated by absorption and fluorescence spectroscopy. This leads to the formation of 1:1 complex with a binding constant of (6.9 ± 2.3) × 10 5 M -1 , with the lowest detection limit being 1.9 nM. In the complex, the Cu 2+ is bound by two triazole-N and two phenolic-O groups resulting in a distorted tetrahedral coordination core of CuN 2 O 2 as demonstrated based on density functional theory studies. To form such coordination core, the arms underwent considerable changes in some of the dihedral angles. The binding of Cu 2+ to L induces self-assembly of L by varying from simple particles to rodlike structures when bound to Cu 2+ . The on-off fluorescence intensity of L and its Cu 2+ -bound species are responsible for imaging cancer cells. The L shows red fluorescence in MDA-MB-231 cancer cells by targeting mitochondria as proved based on the colocalization study carried out using MitoTracker Green. While the L alone is nontoxic to cancer cells, the presence of Cu 2+ brings cell death to an extent of 90% with an IC 50 value of 165 nM by bringing a substantial quench in the fluorescence of L . A shift of population from G 0 /G 1 and G 2 M phases to the Sub-G 1 phase was observed as the concentration of the complex was increased, indicating cell death as studied by fluorescence-activated cell sorting. Thus, the present work clearly proved that a calix[4]arene functionalized at the lower rim with spiro-indoline moieities when complexed with Cu 2+ acts as an efficient anticancer agent and is capable of imaging cancer cells.