Modulation of the ATM/autophagy pathway by a G-quadruplex ligand tips the balance between senescence and apoptosis in cancer cells

Abstract

G-quadruplex ligands exert their antiproliferative effects through telomere-dependent and telomere-independent mechanisms, but the inter-relationships among autophagy, cell growth arrest and cell death induced by these ligands remain largely unexplored. Here, we demonstrate that the G-quadruplex ligand 20A causes growth arrest of cancer cells in culture and in a HeLa cell xenografted mouse model. This response is associated with the induction of senescence and apoptosis. Transcriptomic analysis of 20A treated cells reveals a significant functional enrichment of biological pathways related to growth arrest, DNA damage response and the lysosomal pathway. 20A elicits global DNA damage but not telomeric damage and activates the ATM and autophagy pathways. Loss of ATM following 20A treatment inhibits both autophagy and senescence and sensitizes cells to death. Moreover, disruption of autophagy by deletion of two essential autophagy genes ATG5 and ATG7 leads to failure of CHK1 activation by 20A and subsequently increased cell death. Our results, therefore, identify the activation of ATM by 20A as a critical player in the balance between senescence and apoptosis and autophagy as one of the key mediators of such regulation. Thus, targeting the ATM/autophagy pathway might be a promising strategy to achieve the maximal anticancer effect of this compound.

Triarylpyridine Compounds and Chloroquine Act in Concert to Trigger Lysosomal Membrane Permeabilization and Cell Death in Cancer Cells

“…Another factor that regulates the lysosomal size and number is the TFEB transcription factor, which is responsible for the induction of genes encoding lysosomal proteins through a mechanism that depends on mTOR inhibition [32,33]. The fact that 20A causes the upregulation of several genes encoding for lysosomal proteins and inhibition of mTOR (as shown in our previous works [24,34] and Figure S1) argues in favor of the activation of the TFEB protein in response to 20A. It would be interesting to determine if and how TFEB regulates lysosome functions and cell death in cells exposed to 20A alone or with chloroquine.…”

“…We have previously shown that 20A, a G-quadruplex ligand belonging to the triarylpyridine family, exerts potent cytotoxic and cytostatic effects on several cancer cells. We found that one of the gene classes most significantly upregulated by 20A corresponds to the autophagy lysosomal pathway [24].…”

“…The human cervical cancer cell lines HeLa and U20S were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA) and the human lung carcinoma A549 cell line is a generous gift of Prof. Jean-François Riou (CNRS UMR 7196, Paris, France). The autophagy-deficient (ATG5 and ATG7 depleted) HeLa cell lines were generated as previously described [24]. U2OS expressing mCherry-Galectin3 cell line is a generous gift from Dr. A H. Woldrich (CNRS UMR5, Bordeaux, France) [41].…”

“…For some experiments, cells were stained with 150 nM tetramethylrhodamine methyl ester (TMRM, from Molecular Probes-Life Technologies) and 5 µg/mL 4 ,6-diaminidino-2-phenylindole (DAPI, from Molecular Probes-Life Technologies) for 30 min at 37 • C. Cytofluorometric determinations of early and late apoptotis were carried out on a MACSQuant cytometer (Miltenyi Biotec, Bergisch Gladbach, Germany) upon gating on events displaying normal forward scatter and side scatter parameters. Apoptosis was also evaluated by western blotting analyses of cleaved forms of either PARP 1 or caspase 3 [24]. Where indicated, the caspase inhibitor QV-D-Oph (20 µM) was added to cells to evaluate the possible implication of caspases in cell demise.…”

G‐Quadruplex‐Binding Small Molecule Induces Synthetic Lethality in Breast Cancer Cells by Inhibiting c‐MYC and BCL2 Expression

“…Small molecules capable of stabilizing G4 can trigger DNA double-strand breaks and induce DNA damage response (DDR) and cell death. [5] Because G4s contain the G-quartet as a common structural feature, ligands can stabilize more than one G4 target present in different promoters. One such example is quarfloxin (CX-3543), which shows selectivity for parallel G4s, such as c-MYC [6] and VEGF.…”