Abstract 5360: Temozolomide-resistant glioblastoma cells are collaterally sensitive to ferroptosis through NRF2 high expression
Authors: de Souza Izadora , Monteiro Linda Karolynne , Guedes Camila Banca , Latancia Marcela , Andrade Marina Tomaz , Silva Matheus Molina , Contieri Bruna , Ribeiro Bruna Felício Milazzotto , Lazarini Mariana , Gomes Luciana , Rocha Clarissa 
Published: June 2022

Abstract
Glioblastoma patients have a poor prognosis with a low median survival rate mainly due to temozolomide (TMZ) resistance. NRF2 is an important transcript factor involved in chemotherapy resistance due to its ability to regulate genes related to the antioxidant response and to prevent cell death processes, such as ferroptosis. However, the relation between NRF2 and iron-dependent cell death is contradictory and poorly understood. This study aimed to analyze the role of NRF2 in ferroptosis modulation in glioblastoma cells. To this end, it was analyzed two human glioblastoma cell lines (U251MG and T98G) after treatment with TMZ, ferroptosis inducers (Erastin, RSL3, and Sorafenib), and ferroptosis inhibitor (Ferrostatin-1). Also, we performed gene expression analysis of glioma patients. Our results demonstrated that T98G compared to the U251MG was more resistant to chemotherapy and showed elevated levels of NRF2 expression and its targets xCT, HMOX1, and ABCC1. Interestingly, T98G cells revealed higher sensitivity and lipoperoxidation levels after ferroptotic treatment. Next, we established T98G NRF2 silenced cells and we observed a significant reduction in cellular viability after TMZ treatment when compared to wild-type cells. On the other hand, T98G-shNRF2 was more resistant to ferroptosis induction, indicating that NRF2 plays a key role in the modulation of chemoresistance and ferroptosis. After, we showed that NRF2 controls levels of ABCC1/MRP1 in glioblastoma cells, and ABCC1 silencing promotes sensitivity to TMZ and resistance to Erastin. These results support a possible mechanism of ferroptosis modulation by NRF2 on TMZ-resistant gliomas through ABCC1, which has been recently associated with ferroptosis induction by promoting efflux of glutathione out of the cell. Furthermore, we confirmed that NRF2 has a positive correlation with ABCC1 in glioma patients, and higher ABCC1 expression was associated with tumor aggressiveness. Also, we validated ABCC1 as an independent prognostic factor for poor overall survival on glioma. Finally, we observed that T98G cells have sensitivity to the ferroptosis inducer FDA-approved, sorafenib. Altogether our data suggest that high levels of NRF2 may result in ferroptosis sensitivity on glioblastoma through the high levels of expression of its pro-ferroptotic target ABCC1, once the xCT system is blocked by Erastin. Thus, glioblastoma cell vulnerability to ferroptosis by NRF2 and ABCC1 high expression can be an Achilles’ heel to reverse drug resistance on glioblastoma through the treatment with ferroptosis inducers.
Citation Format: Izadora de Souza, Linda Karolynne Monteiro, Camila Banca Guedes, Marcela Latancia, Marina Tomaz Andrade, Matheus Molina Silva, Bruna Contieri, Bruna Felício Milazzotto Ribeiro, Mariana Lazarini, Luciana Gomes, Clarissa Rocha. Temozolomide-resistant glioblastoma cells are collaterally sensitive to ferroptosis through NRF2 high expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5360.