Flavonoids have been reported to play an essential role in modulating processes of cellular redox homeostasis such as scavenging ROS. Meanwhile, they also induce oxidative stress that exerts potent antitumor bioactivity. However, the contradiction between these two aspects still remains unclear. In this study, four typical flavonoids were selected and studied. The results showed that low-dose flavonoids slightly promoted the proliferation of breast cancer cells under normal growth via gradually reducing accumulated oxidative products and demonstrated a synergistic effect with reductants NAC or VC. Besides, low-dose flavonoids significantly reduced the content of ROS and MDA induced by LPS or Rosup but restored the activity of SOD. However, high-dose flavonoids markedly triggered the cell death via oxidative stress as evidenced by upregulated ROS, MDA and downregulated SOD activity that could be partly rescued by NAC pretreatment, which was also confirmed by antioxidative gene expression levels. The underlying mechanism of such induced cell death was pinpointed as apoptosis, cell cycle arrest, accumulated mitochondrial superoxide, impaired mitochondrial function and decreased ATP synthesis. Transcriptomic analysis of apigenin and quercetin uncovered that high-dose flavonoids activated TNF-α signaling, as verified through detecting inflammatory gene levels in breast cancer cells and RAW 264.7 macrophages. Moreover, we identified that BRCA1 overexpression effectively attenuated such oxidative stress, inflammation and inhibited ATP synthesis induced by LPS or high dose of flavonoids possibly through repairing DNA damage, revealing an indispensable biological function of BRCA1 in resisting oxidative damage and inflammatory stimulation caused by exogenous factors.

Breast cancer, the most common cancer in women, usually exhibits intrinsic insensitivity to drugs, even without drug resistance. MUC1 is a highly glycosylated transmembrane protein, overexpressed in breast cancer, contributing to tumorigenesis and worse prognosis. However, the molecular mechanism between MUC1 and drug sensitivity still remains unclear. Here, natural flavonoid apigenin was used as objective due to the antitumor activity and wide availability. MUC1 knockout (KO) markedly sensitized breast cancer cells to apigenin cytotoxicity in vitro and in vivo. Both genetical and pharmacological inhibition significantly enhanced the chemosensitivity to apigenin and clinical drugs whereas MUC1 overexpression conversely aggravated such drug resistance. Constitutively re-expressing wild type MUC1 in KO cells restored the drug resistance; however, the transmembrane domain deletant could not rescue the phenotype. Notably, further investigation discovered that membrane-dependent drug resistance relied on the extracellular glycosylated modification since removing O-glycosylation via inhibitor, enzyme digestion, or GCNT3 (MUC1 related O-glycosyltransferase) knockout markedly reinvigorated the chemosensitivity in WT cells, but had no effect on KO cells. Conversely, inserting O-glycosylated sites to MUC1-N increased the drug tolerance whereas the O-glycosylated deletant (Ser/Thr to Ala) maintained high susceptibility to drugs. Importantly, the intracellular concentration of apigenin measured by UPLC and fluorescence distribution firmly revealed the increased drug permeation in MUC1 KO and BAG-pretreated cells. Multiple clinical chemotherapeutics with small molecular were tested and obtained the similar conclusion. Our findings uncover a critical role of the extracellular O-glycosylation of MUC1-N in weakening drug sensitivity through acting as a barrier, highlighting a new perspective that targeting MUC1 O-glycosylation has great potential to promote drug sensitivity and efficacy.