Oncogenic RAS Mutants Confer Resistance of RMS13 Rhabdomyosarcoma Cells to Oxidative Stress-Induced Ferroptotic Cell Death

Abstract

Recent genomic studies revealed a high rate of recurrent mutations in the RAS pathway in primary rhabdomyosarcoma (RMS) samples. In the present study, we therefore investigated how oncogenic RAS mutants impinge on the regulation of cell death of RMS13 cells. Here, we report that ectopic expression of NRAS12V, KRAS12V, or HRAS12V protects RMS13 cells from oxidative stress-induced cell death. RMS13 cells engineered to express NRAS12V, KRAS12V, or HRAS12V were significantly less susceptible to loss of cell viability upon treatment with several oxidative stress inducers including the thioredoxin reductase inhibitor Auranofin, the glutathione (GSH) peroxidase 4 inhibitor RSL3 or Erastin, an inhibitor of the cysteine/glutamate amino acid transporter system xcMathClass-bin− that blocks GSH synthesis. Notably, addition of Ferrostatin-1 confers protection against Erastin- or RSL3-induced cytotoxicity, indicating that these compounds trigger ferroptosis, an iron-dependent form of programed cell death. Furthermore, RMS13 cells overexpressing oncogenic RAS mutants are significantly protected against the dual PI3K/mTOR inhibitor PI103, whereas they are similarly sensitive to DNA-damaging drugs such as Doxorubicin or Etoposide. This suggests that oncogenic RAS selectively modulates cell death pathways triggered by cytotoxic stimuli in RMS13 cells. In conclusion, our discovery of an increased resistance to oxidative stress imposed by oncogenic RAS mutants in RMS13 cells has important implications for the development of targeted therapies for RMS.

Targeting ferroptosis in rhabdomyosarcoma cells

“…In the present study, we observed no correlation between RAS mutational status and response to Erastin‐induced cell death in the investigated RMS cell lines. By comparison, ectopic overexpression of oncogenic RAS mutants such as NRAS12V, KRAS12V or HRAS12V in RAS wild‐type RMS13 cells reduced Erastin‐mediated cell death . Recently, the susceptibility of RMS cell lines to ferroptosis has been linked to a high proliferation status via ERK signaling …”

“…By comparison, ectopic overexpression of oncogenic RAS mutants such as NRAS12V, KRAS12V or HRAS12V in RAS wild-type RMS13 cells reduced Erastin-mediated cell death. 35 Recently, the susceptibility of RMS cell lines to ferroptosis has been linked to a high proliferation status via ERK signaling. 36 Besides PKC, our study points to an involvement of NOX in ROS signaling and ferroptotic cell death in RMS, since the broad-spectrum NOX inhibitor DPI as well as the selective NOX1/4 inhibitor GKT137831 rescued RMS cells from Erastin-stimulated ROS production, lipid peroxidation and cell death.…”

Ferroptosis, a new form of cell death, and its relationships with tumourous diseases

“…Ferroptosis can occur in rhabdomyosarcoma cells, as demonstrated by an experimental observation that erastin and RSL3 treatments can induce ferroptosis in these cells . Sun et al .…”

Molecular mechanisms of ferroptosis and its role in cancer therapy

“…Furthermore, the rhabdomyosarcoma engineered to express the mutated RAS even confers resistance to erastin-or RSL3-induced cell death. 15 Leukaemia cells without RAS mutation also showed great sensitivity to ferroptosis induction. 1 Further studies showed the different ferroptosis sensitivities Cell lines derived from the oesophagus, upper respiratory tract, stomach, pancreas, breast, skin and large intestine were generally insensitive to the four ferroptotic reagents.…”