MYCN-Amplified Neuroblastoma Is Addicted to Iron and Vulnerable to Inhibition of the System Xc-/Glutathione Axis

Abstract

MYCN is amplified in 20% to 25% of neuroblastoma, and MYCN-amplified neuroblastoma contributes to a large percent of pediatric cancer–related deaths. Therapy improvements for this subtype of cancer are a high priority. Here we uncover a MYCN-dependent therapeutic vulnerability in neuroblastoma. Namely, amplified MYCN rewires the cell through expression of key receptors, ultimately enhancing iron influx through increased expression of the iron import transferrin receptor 1. Accumulating iron causes reactive oxygen species (ROS) production, and MYCN-amplified neuroblastomas show enhanced reliance on the system Xc- cystine/glutamate antiporter for ROS detoxification through increased transcription of this receptor. This dependence creates a marked vulnerability to targeting the system Xc-/glutathione (GSH) pathway with ferroptosis inducers. This reliance can be exploited through therapy with FDA-approved rheumatoid arthritis drugs sulfasalazine (SAS) and auranofin: in MYCN-amplified, patient-derived xenograft models, both therapies blocked growth and induced ferroptosis. SAS and auranofin activity was largely mitigated by the ferroptosis inhibitor ferrostatin-1, antioxidants like N-acetyl-L-cysteine, or by the iron scavenger deferoxamine (DFO). DFO reduced auranofin-induced ROS, further linking increased iron capture in MYCN-amplified neuroblastoma to a therapeutic vulnerability to ROS-inducing drugs. These data uncover an oncogene vulnerability to ferroptosis caused by increased iron accumulation and subsequent reliance on the system Xc-/GSH pathway. Significance: This study shows how MYCN increases intracellular iron levels and subsequent GSH pathway activity and demonstrates the antitumor activity of FDA-approved SAS and auranofin in patient-derived xenograft models of MYCN-amplified neuroblastoma.

MYCN upregulates the transsulfuration pathway to suppress the ferroptotic vulnerability in MYCN-amplified neuroblastoma

“…We employed our syngeneic SK-N-SH neuroblastoma cell pair expressing exogenous MYCN or GFP [11] and performed untargeted liquid chromatography/tandem mass spectrometry of cell lysates to determine changes in a broad array of polar metabolites. Our data indicated decreased glutamate levels, increased cysteine levels and increased cysteinyl-glycine (Cys-Gly) levels, consistent with an activated system Xc-antiporter [7]. 'Confirming our previously published data from colorimetric analyses of GSH levels [7], GSH levels were increased over 5-fold in the presence of MYCN (Fig.…”

MYCN mediates TFRC-dependent ferroptosis and reveals vulnerabilities in neuroblastoma

“…This argument can be further supported by the study from Floros et al For instance, SAS is capable of inducing~70% cell death in MYCN-amplified NB cells. However, knockdown of two System X c (−) components SLC7A11 and SLC3A2 induce only~25% cell death in the same cell line 31 , confirming that SAS harbors considerable off-target effects. In conclusion, the complex effects of SAS treatments leading to ferroptosis cannot be solely linked to the sensitivity of NB cells to System X c (−) inhibition.…”

“…Notably, Floros et al recently reported that MYCNamplified NB cells are more sensitive to the system X c (−) inhibitor sulfasalazine (SAS) 31 . Indeed, MYCN-amplified NB cells are slightly sensitive to SAS as reflected by their smaller IC50 values compared to non-amplified cells, although no statistical significance was achieved in our hands (Fig.…”

Genetic Profiles of Ferroptosis in Malignant Brain Tumors and Off-Target Effects of Ferroptosis Induction

“…In about 25% of neuroblastoma cases MYCN is amplified ( 19 ). In patient-derived xenografts of these cases, the xCT-driven antioxidant response after sulfasalazine application is increased compared to controls, which leads to an increase in ferroptosis and subsequently limited tumor growth ( 19 ).…”

“…In about 25% of neuroblastoma cases MYCN is amplified ( 19 ). In patient-derived xenografts of these cases, the xCT-driven antioxidant response after sulfasalazine application is increased compared to controls, which leads to an increase in ferroptosis and subsequently limited tumor growth ( 19 ). Further studies revealed that the transferrin receptor 1 was upregulated in response to such MYCN amplification, leading to increased GPx4 sensitivity and rendering neuroblastoma cells vulnerable to ferroptosis induction ( 20 ).…”