Ligands stimulating antitumour immunity as the next G-quadruplex challenge

Abstract

G-quadruplex (G4) binders have been investigated to discover new anticancer drugs worldwide in past decades. As these ligands are generally not highly cytotoxic, the discovery rational was mainly based on increasing the cell-killing potency. Nevertheless, no G4 binder has been shown yet to be effective in cancer patients. Here, G4 binder activity at low dosages will be discussed as a critical feature to discover ligands with therapeutic effects in cancer patients. Specific effects of G4 binders al low doses have been reported to occur in cancer and normal cells. Among them, genome instability and the stimulation of cytoplasmic processes related to autophagy and innate immune response open to the use of G4 binders as immune-stimulating agents. Thus, we propose a new rational of drug discovery, which is not based on cytotoxic potency but rather on immune gene activation at non-cytotoxic dosage.

Keywords: Cancer immunotherapy; G-loop; G-quadruplex; G4 ligands; Genome instability; Innate immunity.

Fig. 1

G-quadruplex and other non-B DNA structures. G-quadruplexes can form in G-rich strand of both RNA and DNA strands (A). They can fold into different conformations depending on strand stoichiometry (intra- or inter-strand topologies), strand direction (parallel, antiparallel, or mixed) and length of intervening nucleotides (bulges and loops) (B). C-rich DNA strands can fold into i-motif structures (C). R-loops can favour G4 folding on the displaced DNA strand resulting into another non-B structure known as G-loop (D)

Fig. 2

Cytotoxicity potency of anticancer  DNA-interacting agents. Violin plots showing IC₅₀ values of selected DNA-interacting agents with established anticancer activity in standard chemotherapy of human tumours. Data were obtained from “Genomics of Drug Sensitivity in Cancer” (GDSC) database [40], which includes information from around one thousands cancer cell lines (www.cancerrxgene.org). Drugs are classified on the base of their mechanism of action and ranked by cytotoxicity (left to right). G4 stabilizers have a lower cytotoxic potency than topoisomerase poisons, microtubule interactors, Mitomycin C and Dactinomycin (Actinomycin D). Alkylating agents and cross-linkers can react with cytoplasmic nucleophilic groups in cells, effectively lowering their nuclear concentrations [41]. Moreover, PARP inhibitors are mainly used in BRCAness tumours where their cell killing potency is much increased due to mutations or deletion of BRCA genes [42]. Therefore, G4 binders (Pyridostatin and CX-5461) have a low cytotoxic potency in comparison to other effective DNA-interacting anticancer agents

Fig. 3

G4 binders increase R-loop-dependent genome instability activating then the cGAS/STING/IRF3 pathway in cancer cells. Mechanism of G4-binders activity in cancer cells. A) G4 binders target G4 structures by stabilizing the G-loop. B) G-loops can induce double strand breaks through either replication- and transcription-dependent mechanisms, resulting in cell senescence or micronuclei induction. C) Micronuclei are recognized by DNA sensing protein cGAS, leading to activation of cGAS/STING pathway and to IRF3- regulated gene expression. D) The phosphorylated form of IRF3 translocates to the nucleus and promotes IFNB expression in cancer cells. IFNB secretion allows its binding to a membrane interferon receptor and consequent IFNB-dependent transcriptional cascade activation.

Fig. 4

G4 binder-mediated cellular effects. G4-binder mechanisms of action and their pleiotropic cellular outcome as discussed in the review. Created with BioRender.com