bims-nurfca 2023-07-16 papers

bims-nurfca

Biomed News

on NRF2 and Cancer

Issue of 2023–07–16
seven papers selected by
Caner Geyik, Istinye University

Nuclear localization of NRF2 in stroma of HER2 positive and triple-negative breast cancer.
Fisetin induces apoptosis in colorectal cancer cells by suppressing autophagy and down-regulating nuclear factor erythroid 2-related factor 2 (Nrf2).
Impact of Nrf2 overexpression on cholangiocarcinoma treatment and clinical prognosis.
Co-targeting autophagy and NRF2 signaling triggers mitochondrial superoxide to sensitize oral cancer stem cells for cisplatin-induced apoptosis.
Necroptosis inhibits autophagy by regulating the formation of RIP3/p62/Keap1 complex in shikonin-induced ROS dependent cell death of human bladder cancer.
Huangqin Decoction Delays Progress of Colitis-Associated Carcinogenesis by Regulating Nrf2/HO-1 Antioxidant Signal Pathway in Mice.
Glutamine antagonist DRP-104 suppresses tumor growth and enhances response to checkpoint blockade in KEAP1 mutant lung cancer.

Pathol Res Pract. 2023 Jul 05. pii: S0344-0338(23)00362-X. [Epub ahead of print]248 154662

Nuclear localization of NRF2 in stroma of HER2 positive and triple-negative breast cancer.

Tomislav Orešić, Sanda Bubanović, Snježana Ramić, Božena Šarčević, Ana Čipak Gašparović.

  Breast cancer is one of the leading causes of cancer-related mortality in women. During tumor growth, periods of hypoxia are followed by reoxygenation due to neovascularisation leading to disturbed redox homeostasis. ROS (Reactive Oxygen Species) produced under hypoxia activate HIF1α. ROS can also activate the major antioxidant transcription factor NRF2, but also cause damage to biomolecules. Lipids are susceptible to peroxidation, as evidenced by the formation of reactive aldehydes, among which, HNE (4-hydroxynonenal) is the most studied one. Knowing that HIF1α (Hypoxia Inducing Factor 1α) is associated with breast cancer malignancy, we aimed to investigate its correlation with HNE and NRF2 (Nuclear factor erythroid 2-related factor 2). Our results show that HIF1α is activated in breast cancer, indicating an increase in ROS but not followed by HNE production. On the other hand, NRF2 was increased in all types of breast cancer suggesting that oxidative stress is present in these pathologies, but also supporting HIF1α. Interestingly, NRF2 was activated in HER2 positive and TNBC, indicating the role of stromal NRF2 in breast cancer malignancy.

Keywords:  4-hydroxynonenal; Breast cancer; HIF1α; NRF2

DOI:  https://doi.org/10.1016/j.prp.2023.154662
J Cell Biochem. 2023 Jul 14.

Fisetin induces apoptosis in colorectal cancer cells by suppressing autophagy and down-regulating nuclear factor erythroid 2-related factor 2 (Nrf2).

Akanksha Pandey, Surendra Kumar Trigun.

  Modulation of autophagy is evolving as a relevant strategy in cancer pathogenesis and therapeutic intervention and hence, needs to be examined as a target for the promising anticancer agents. Fisetin, a dietary flavanol, is emerging as a potent anticancer agent, however, its tumour-specific pharmacological targets remain largely unexplored. This article describes correlative profiles of autophagy and apoptotic markers versus nuclear factor erythroid 2-related factor 2 (Nrf2) and reactive oxygen species (ROS) in the colorectal cancer (CRC) cell line SW-480. As compared to the untreated cells, significantly less number of fluorescent detected autophagic vacuoles (AVOs) in the fisetin-treated cells coincided with a similar decline of the autophagy flux markers, Beclin 1 and microtubule-associated protein-1 light chain-3 and accumulation of p62 in those cells. The significantly increased number of annexin-V/propidium iodide (+/+) positive and acridine orange/ethidium bromide-stained apoptotic cells coincided with the enhanced signals for the cleaved caspase 3 and nuclear PARP-1 in those fisetin-treated cells. This was consistent with the collapse of mitochondrial membrane potential and release of cytochrome c. The fisetin-treated cells showed increased ROS level and a significant decline in nuclear Nrf2 immunosignal versus recovery in nuclear Nrf2 due to the treatment with curcumin and resveratrol (Nrf2 activators) and thus, suggesting a role of Nrf2 suppression in fisetin-mediated apoptosis in SW-480 cells. The effect of chloroquine, an autophagy inhibitor, resulted into declined number of AVOs and enhanced apoptosis, similar to that of the fisetin effect. Also, regaining of AVOs number and reduced apoptosis of CRC cells due to the treatment with rapamycin, an autophagy inducer, could be observed. These loss and gain of functions experiments thus suggested a correlation between fisetin-mediated autophagy suppression and apoptotic induction in a colorectal cell line.

Keywords:  Nrf2; ROS; apoptosis; autophagy; colorectal cells; fisetin

DOI:  https://doi.org/10.1002/jcb.30447
Korean J Clin Oncol. 2023 Jun;19(1): 18-26

Impact of Nrf2 overexpression on cholangiocarcinoma treatment and clinical prognosis.

Huisong Lee, Seog Ki Min, Min-Sun Cho, Hyeon Kook Lee.

   PURPOSE: Nrf2 regulates antioxidant protein expression and protects against drug toxicity and oxidative stress, whereas Keap1 controls Nrf2 activity. The Keap1-Nrf2 pathway affects the prognosis of various cancers, however, its effect on cholangiocarcinoma chemoresistance and prognosis remains unclear. This study aimed to determine whether the Keap1-Nrf2 pathway affects chemoresistance and prognosis of distal cholangiocarcinoma.
METHODS: We investigated the correlation between Nrf2 and Keap1 expression and clinical characteristics and prognosis in 91 patients with distal cholangiocarcinoma who underwent curative surgery. Immunohistochemical staining was performed on paraffin blocks using primary antibodies against Nrf2 and Keap1. The relationship between Keap1 and Nrf2 protein expression levels, and clinical characteristics and prognosis was examined.
RESULTS: Nrf2 expression was not associated with overall survival in patients who did not receive adjuvant chemotherapy (P=0.994). Among patients receiving adjuvant chemotherapy, the Nrf2 low expression group had a significantly longer median overall survival than the Nrf2 high expression group in Kaplan-Meier survival analysis (P=0.019). In multivariate analysis, high expression of Nrf2 was confirmed as an independent poor prognostic factor in the group receiving adjuvant chemotherapy (P=0.041).
CONCLUSION: This study suggests that Nrf2 overexpression reduces the efficacy of adjuvant chemotherapy in distal cholangiocarcinoma.

Keywords:  Adjuvant chemotherapy; Cholangiocarcinoma; Keap1 protein; Nrf2 protein; Pancreatoduodenectomy

DOI:  https://doi.org/10.14216/kjco.23004
Free Radic Biol Med. 2023 Jul 07. pii: S0891-5849(23)00526-9. [Epub ahead of print]

Co-targeting autophagy and NRF2 signaling triggers mitochondrial superoxide to sensitize oral cancer stem cells for cisplatin-induced apoptosis.

Prakash P Praharaj, Amruta Singh, Srimanta Patra, Sujit K Bhutia.

  Cancer stem cell (CSC) populations are regulated by autophagy, which in turn modulates tumorigenicity and malignancy. In this study, we demonstrate that cisplatin treatment enriches the CSCs population by increasing autophagosome formation and speeding up autophagosome-lysosome fusion by recruiting RAB7 to autolysosomes. Further, cisplatin treatment stimulates lysosomal activity and increases autophagic flux in oral CD44+ cells. Interestingly, both ATG5-and BECN1-dependent autophagy are essential for maintaining cancer stemness, self-renewal, and resistance to cisplatin-induced cytotoxicity in oral CD44+ cells. Moreover, we discovered that autophagy-deficient (shATG5 and/or shBECN1) CD44+ cells benefited from cisplatin treatment because it activated NRF2 (nuclear factor, erythroid 2 like 2) signaling, which in turn reduces the elevated ROS level enhancing cancer stemness. Genetic inhibition of NRF2 (siNRF2) in autophagy-deficient CD44+ cells increases mtROS level, reducing cisplatin-resistance CSCs, and pre-treatment with mitoTEMPO [a mitochondria-targeted SOD (superoxide dismutase) mimetic] lessened the cytotoxic effect enhancing cancer stemness. We also found that inhibiting autophagy (with CQ) and NRF2 signaling (with ML-385) combinedly increased cisplatin cytotoxicity, thereby suppressing the expansion of oral CD44+ cells; this finding has the potential to be clinically applicable in resolving CSC-associated chemoresistance and tumor relapse in oral cancer.

Keywords:  Apoptosis; Autophagy; Cancer stem cell; NRF2; Oral cancer; mtROS

DOI:  https://doi.org/10.1016/j.freeradbiomed.2023.07.008
Phytomedicine. 2023 Jun 28. pii: S0944-7113(23)00304-5. [Epub ahead of print]118 154943

Necroptosis inhibits autophagy by regulating the formation of RIP3/p62/Keap1 complex in shikonin-induced ROS dependent cell death of human bladder cancer.

Xiaojie Liu, Lu Liu, Xu Wang, Yubin Jin, Shuang Wang, Qin Xie, Yanhe Jin, Mengli Zhang, Yunhe Liu, Jinfeng Li, Zhenya Wang, Xiangjing Fu, Cheng-Yun Jin.

   BACKGROUND: Shikonin, a natural naphthoquinone compound, has a wide range of pharmacological effects, but its anti-tumor effect and underlying mechanisms in bladder cancer remain unclear.
PURPOSE: We aimed to investigate the role of shikonin in bladder cancer in vitro and in vivo in order to broaden the scope of shikonin's clinical application.
STUDY DESIGN AND METHODS: We performed MTT and colony formation to detect the inhibiting effect of shikonin on bladder cancer cells. ROS staining and flow cytometry assays were performed to detect the accumulation of ROS. Western blotting, siRNA and immunoprecipitation were used to evaluate the effect of necroptosis in bladder cancer cells. Transmission electron microscopy and immunofluorescence were used to examine the effect of autophagy. Nucleoplasmic separation and other pharmacological experimental methods described were used to explore the Nrf2 signal pathway and the crosstalk with necroptosis and autophagy. We established a subcutaneously implanted tumor model and performed immunohistochemistry assays to study the effects and the underlying mechanisms of shikonin on bladder cancer cells in vivo.
RESULTS: The results showed that shikonin has a selective inhibitory effect on bladder cancer cells and has no toxicity on normal bladder epithelial cells. Mechanically, shikonin induced necroptosis and impaired autophagic flux via ROS generation. The accumulation of autophagic biomarker p62 elevated p62/Keap1 complex and activated the Nrf2 signaling pathway to fight against ROS. Furthermore, crosstalk between necroptosis and autophagy was present, we found that RIP3 may be involved in autophagosomes and be degraded by autolysosomes. We found for the first time that shikonin-induced activation of RIP3 may disturb the autophagic flux, and inhibiting RIP3 and necroptosis could accelerate the conversion of autophagosome to autolysosome and further activate autophagy. Therefore, on the basis of RIP3/p62/Keap1 complex regulatory system, we further combined shikonin with late autophagy inhibitor(chloroquine) to treat bladder cancer and achieved a better inhibitory effect.
CONCLUSION: In conclusion, shikonin could induce necroptosis and impaired autophagic flux through RIP3/p62/Keap1 complex regulatory system, necroptosis could inhibit the process of autophagy via RIP3. Combining shikonin with late autophagy inhibitor could further activate necroptosis via disturbing RIP3 degradation in bladder cancer in vitro and in vivo.

Keywords:  Autophagy; Necroptosis; RIP3/p62/Keap1; ROS; Shikonin

DOI:  https://doi.org/10.1016/j.phymed.2023.154943
Chin J Integr Med. 2023 Jul 12.

Huangqin Decoction Delays Progress of Colitis-Associated Carcinogenesis by Regulating Nrf2/HO-1 Antioxidant Signal Pathway in Mice.

Li-Mei Gu, He-Zhong Li, Lei Gao, Hui Li, Lan-Fu Wei, Cheng-Yu Pan, Ke-Xuan Wu, Yao-Zhou Tian.

   OBJECTIVE: To investigate the effect of Huangqin Decoction (HQD) on nuclear factor erythroid 2 related-factor 2 (Nrf2)/heme oxygenase (HO-1) signaling pathway by inducing the colitis-associated carcinogenesis (CAC) model mice with azoxymethane (AOM)/dextran sodium sulfate (DSS).
METHODS: The chemical components of HQD were analyzed by liquid chromatography-quadrupole-time-of-flight mass spectrometry (LC-Q-TOF-MS/MS) to determine the molecular constituents of HQD. Totally 48 C57BL/6J mice were randomly divided into 6 groups by a random number table, including control, model (AOM/DSS), mesalazine (MS), low-, medium-, and high-dose HQD (HQD-L, HQD-M, and HQD-H) groups, 8 mice in each group. Except for the control group, the mice in the other groups were intraperitoneally injected with AOM (10 mg/kg) and administrated with 2.5% DSS orally for 1 week every two weeks (totally 3 rounds of DSS) to construct a colitis-associated carcinogenesis mouse model. The mice in the HQD-L, HQD-M and HQD-H groups were given HQD by gavage at doses of 2.925, 5.85, and 11.7 g/kg, respectively; the mice in the MS group was given a suspension of MS at a dose of 0.043 g/kg (totally 11 weeks). The serum levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were measured by enzyme-linked immunosorbent assay. The mRNA and protein expression levels of Nrf2, HO-1, and inhibitory KELCH like ECH-related protein 1 (Keap1) in colon tissue were detected by quantitative real-time PCR, immunohistochemistry, and Western blot, respectively.
RESULTS: LC-Q-TOF-MS/MS analysis revealed that the chemical constituents of HQD include baicalin, paeoniflorin, and glycyrrhizic acid. Compared to the control group, significantly higher MDA levels and lower SOD levels were observed in the model group (P<0.05), whereas the expressions of Nrf2 and HO-1 were significantly decreased, and the expression of Keap1 increased (P<0.01). Compared with the model group, serum MDA level was decreased and SOD level was increased in the HQD-M, HQD-H and MS groups (P<0.05). Higher expressions of Nrf2 and HO-1 were observed in the HQD groups.
CONCLUSION: HQD may regulate the expression of Nrf2 and HO-1 in colon tissue, reduce the expression of MDA and increase the expression of SOD in serum, thus delaying the progress of CAC in AOM/DSS mice.

Keywords:  Chinese medicine; Huangqin Decoction; colitis-associated carcinogenesis; nuclear factor erythroid 2 related-factor 2/heme oxygenase signaling pathway; oxidative stress

DOI:  https://doi.org/10.1007/s11655-023-3554-y
bioRxiv. 2023 Jun 28. pii: 2023.06.27.546750. [Epub ahead of print]

Glutamine antagonist DRP-104 suppresses tumor growth and enhances response to checkpoint blockade in KEAP1 mutant lung cancer.

Ray Pillai, Sarah E LeBoeuf, Yuan Hao, Connie New, Jenna L E Blum, Ali Rashidfarrokhi, Shih Ming Huang, Christian Bahamon, Warren L Wu, Burcu Karadal-Ferrena, Alberto Herrera, Ellie Ivanova, Michael Cross, Jozef P Bossowski, Hongyu Ding, Makiko Hayashi, Sahith Rajalingam, Triantafyllia Karakousi, Volkan I Sayin, Kamal M Khanna, Kwok-Kin Wong, Robert Wild, Aristotelis Tsirigos, John T Poirier, Charles M Rudin, Shawn M Davidson, Sergei B Koralov, Thales Papagiannakopoulos.

Loss-of-function mutations in KEAP1 frequently occur in lung cancer and are associated with resistance to standard of care treatment, highlighting the need for the development of targeted therapies. We have previously shown that KEAP1 mutant tumors have increased glutamine consumption to support the metabolic rewiring associated with NRF2 activation. Here, using patient-derived xenograft models and antigenic orthotopic lung cancer models, we show that the novel glutamine antagonist DRP-104 impairs the growth of KEAP1 mutant tumors. We find that DRP-104 suppresses KEAP1 mutant tumor growth by inhibiting glutamine-dependent nucleotide synthesis and promoting anti-tumor CD4 and CD8 T cell responses. Using multimodal single-cell sequencing and ex vivo functional assays, we discover that DRP-104 reverses T cell exhaustion and enhances the function of CD4 and CD8 T cells culminating in an improved response to anti-PD1 therapy. Our pre-clinical findings provide compelling evidence that DRP-104, currently in phase 1 clinical trials, offers a promising therapeutic approach for treating patients with KEAP1 mutant lung cancer. Furthermore, we demonstrate that by combining DRP-104 with checkpoint inhibition, we can achieve suppression of tumor intrinsic metabolism and augmentation of anti-tumor T cell responses.

DOI: https://doi.org/10.1101/2023.06.27.546750