NFE2L2 mutations enhance radioresistance in head and neck cancer by modulating intratumoral myeloid cells. NFE2L2 mutations enhance radioresistance in head and neck cancer by modulating intratumoral myeloid cells

To identify the mechanism by which Nrf2E79Q mediates RT resistance and CB-839 overcomes RT resistance in Nrf2E79Q, we performed RNA sequencing of SAS cells stably expressing either Nrf2WT or Nrf2E79Q treated with RT±CB-839 and analyze the differentially expressed genes in Nrf2E79Q vs. Nrf2WT cells. Overall design: Nrf2WT or Nrf2E79Q SAS cells were pretreated with 0.1μM CB-839 or vehicle for 24 hours, then irradiated (4 Gy). RT was performed as previously described(14) using a 225kVp cabinet x-ray irradiator filtered with 0.5 mm Cu (IC-250, Kimtron Inc.). Thus 4 groups were created, including Nrf2WT+RT, Nrf2E79Q+RT, Nrf2WT+RT+CB-839, Nrf2E79Q+RT+CB-839. All the experiments were performed in triplicate, generating a total of 12 samples. Next, RNA was isolated using Trizol (Catalog# 15596026, Invitrogen) 24 hours after RT. Library preparation and RNA sequencing were performed by Novogene via Illumina platforms based on mechanism of SBS (Sequencing by synthesis).