Cellular adaptations impact the biological activity of naphthalene diimide G-quadruplex ligands in ALT-positive...

Telomeric G-quadruplexes (G4s) represent intriguing targets for tumours characterized by the Alternative Lengthening of Telomere (ALT) mechanism. Here we have investigated the effects of two naphthalene diimide (NDI)-based G4 interacting agents (NMe2 and QN-302) in a pair of ALT-positive human osteosarcoma (U-2 OS and Saos-2) cell lines. Both NDIs displayed marked cell growth inhibitory activity associated with the induction of telomere dysfunctions. Moreover, NDI-treated cells were characterized by perturbations at the mitochondrial level as suggested by an increase in the production of reactive oxygen species, the occurrence of changes in mitochondria density and morphology. However, upon initial inhibition of cell growth, U-2 OS cells withstood ligand-induced stress compared to Saos-2 cells. This ability was in part sustained, in a ligand-dependent manner, by the lack of ALT activity inhibition, as indicated by the levels of telomeric C-circle DNA and of Bloom helicase, a member of the RecQ family of helicases. Moreover, marked basal antioxidant capacity, together with the capability to mount an antioxidant response that is in part mediated by the nuclear factor erythroid 2-related factor, has endowed U-2 OS cells with the ability to adapt to NDI exposure. Our data indicate that NDIs rapidly affect the growth of ALT cancer cells by interfering with telomere and mitochondria homeostasis and suggest that small molecule-mediated stabilization of G4s may be a promising therapeutic strategy in ALT-positive tumors. Nonetheless, depending on the individual NDI and the cell's genetic background, cellular adjustment mechanisms may become activated. This, in turn may impinge on the biological activity of G4 interacting agents. Deciphering these mechanisms and the associated molecular determinants will help accelerating the development of G4-based therapeutic interventions in ALT tumors.