2-Hydroxy-3-methylanthraquinone inhibits homologous recombination repair in osteosarcoma through the MYC-CHK1-RAD51 axis.

BACKGROUND: Osteosarcoma is a malignant bone tumor that usually affects adolescents aged 15-19 y. The DNA damage response (DDR) is significantly enhanced in osteosarcoma, impairing the effect of systemic chemotherapy. Targeting the DDR process was considered a feasible strategy benefitting osteosarcoma patients. However, the clinical application of DDR inhibitors is not impressive because of their side effects. Chinese herbal medicines with high anti-tumor effects and low toxicity in the human body have gradually gained attention. 2-Hydroxy-3-methylanthraquinone (HMA), a Chinese medicine monomer found in the extract of Oldenlandia diffusa, exerts significant inhibitory effects on various tumors. However, its anti-osteosarcoma effects and defined molecular mechanisms have not been reported. METHODS: After HMA treatment, the proliferation and metastasis capacity of osteosarcoma cells was detected by CCK-8, colony formation, transwell assays and Annexin V-fluorescein isothiocyanate/propidium iodide staining. RNA-sequence, plasmid infection, RNA interference, Western blotting and immunofluorescence assay were used to investigate the molecular mechanism and effects of HMA inhibiting osteosarcoma. Rescue assay and CHIP assay was used to further verified the relationship between MYC, CHK1 and RAD51. RESULTS: HMA regulate MYC to inhibit osteosarcoma proliferation and DNA damage repair through PI3K/AKT signaling pathway. The results of RNA-seq, IHC, Western boltting etc. showed relationship between MYC, CHK1 and RAD51. Rescue assay and CHIP assay further verified HMA can impair homologous recombination repair through the MYC-CHK1-RAD51 pathway. CONCLUSION: HMA significantly inhibits osteosarcoma proliferation and homologous recombination repair through the MYC-CHK1-RAD51 pathway, which is mediated by the PI3K-AKT signaling pathway. This study investigated the exact mechanism of the anti-osteosarcoma effect of HMA and provided a potential feasible strategy for the clinical treatment of human osteosarcoma.

GPR78 Regulates Autophagy and Drug Resistance in Non-small Cell Lung Cancer.

Context: Lung cancer is one of the most common forms of cancer. Autophagy and apoptosis play an important role in the development of lung cancer. Researchers have found upregulation of GRP78 expression in cancer cells of various types. Objective: The study intended to explore the mechanism of G protein-coupled receptor 78(GPR78) in regulating autophagy and drug resistance in non-small cell lung cancer (NSCLC). Design: The research team performed a laboratory study. Setting: The study took place in the Department of Thoracic Surgery at Hainan General Hospital of the Hainan Affiliated Hospital of Hainan Medical University in Haikou, Hainan, China. Intervention: The research team cultured immortalized, normal, human bronchial epithelial cells C3 (HBEC3) lines and HBEC4 lines in a serum medium without keratinocytes and infected the expression of GPR78 in knockdown A549 cells using lentiviral agents. The team divided the cells into a control group and a shRNA-GPR78 group, the intervention group. The lentiviral silencing vector expressing shRNA targets human GPR78#1 and GPR78 #2aadam10. Outcome Measures: The research team analyzed the mRNA expression of GPR78 in the NSCLC cell lines H1975, H1299, and A549 and in HBEC3 and HBEC4 using a real time-polymerase chain reaction (RT-PCR) and measured the proliferation of A549 cells at 0h, 24h, 48h, 72h, and 96h using yellow tetrazolium salt (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The team also analyzed the migration and invasion ability of cells using wound healing and Transwell tests as well as measured the protein expression of the autophagy-related factors Beclin-1, microtubule-associated protein light chain 3-I/II (LC3-I/LC3-II), ubiquitin-binding protein p62 and c-Jun N-terminal kinase (JNK) using a Western blot test. The team also analyzed the protein expressions of caspase-9, caspase-3, and caspase-12 related to apoptosis using a Western blot. To detect the cell viability induced by cisplatin, the team used a Cell Counting Kit 8 (CCK-8) at the concentrations of 1µM, 3µM and 10µM. Results: The mRNA expression of GPR78 in the H1975, H1299, and A549 cell lines was significantly higher than that in the HBEC3 and HBEC4 cell lines (P < .05). At 48h, 72h, and 96h, the A549 cell proliferation in the shRNA-GPR78 group was significantly lower than that of the control group (P < .05). The cell migration and invasion of cells in the shRNA-GPR78 group was significantly lower than that in the control group (P < .05), and the cell viability of the shRNA-GPR78 group was significantly lower than that of control group (P < .05). The expression of Beclin-1 and JNK protein in shRNA-GPR78 group was significantly higher than that in the control group (P < .05), and the expression of LC3-I/LC3-II and p62 protein in shRNA-GPR78 group was significantly lower than that in the control group (P < .05). The protein expressions of caspase-9, caspase-3, and caspase-12 in the shRNA-GPR78 group were significantly higher than those of the control group (P < .05), and the protein activities of RhoA and Rac1 in the shRNA-GPR78 group were significantly lower than those in the control group (P < .05). Conclusion: NSCLC upregulated GPR78. The knockdown of GPR78 can attenuate the proliferation, migration, and invasion of NSCLC cells and increase the apoptosis and autophagy of NSCLC cells that cisplatin has induced. Therefore, targeting GPR78 may be a promising treatment strategy for NSCLC patients.

Ferroptosis as a novel form of regulated cell death: Implications in the pathogenesis, oncometabolism and treatment of human cancer.

The treatment of cancer mainly involves surgical excision supplemented by radiotherapy and chemotherapy. Chemotherapy drugs act by interfering with tumor growth and inducing the death of cancer cells. Anti-tumor drugs were developed to induce apoptosis, but some patient's show apoptosis escape and chemotherapy resistance. Therefore, other forms of cell death that can overcome the resistance of tumor cells are important in the context of cancer treatment. Ferroptosis is a newly discovered iron-dependent, non-apoptotic type of cell death that is highly negatively correlated with cancer development. Ferroptosis is mainly caused by the abnormal increase in iron-dependent lipid reactive oxygen species and the imbalance of redox homeostasis. This review summarizes the progression and regulatory mechanism of ferroptosis in cancer and discusses its possible clinical applications in cancer diagnosis and treatment.

The synergistic anticancer effect of cisplatin combined with Oldenlandia diffusa in osteosarcoma MG-63 cell line in vitro.

BACKGROUND: Oldenlandia diffusa (OD) is a well-known traditional Chinese medicine, which is used to prevent and treat many disorders, especially cancers. However, its role in osteosarcoma has not been well understood. Here, we used OD and cisplatin individually and combined in osteosarcoma MG-63 cell to explore whether OD could induce cellular apoptosis and suppress the ability of proliferation and invasion of osteosarcoma MG-63 cell. METHODS: The changes of cellular shape were analyzed by optical microscopy. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide assay was used to analyze cell survival rate in vitro. Flow cytometry was performed to detect cell cycle and cell death. Scratch migration assay was used to evaluate cell migration and invasion. Western blot was performed to determine the expression levels of pro-apoptotic and anti-apoptotic protein. RESULTS: In this study, we found that the survival rate reduced significantly in the combined group compared with the individual group and control group. The apoptosis-inducing effect of combined application was much more significant than that of individual application. The invasion ability of combined application was significantly lower than that of the individual application. In the combined group, there were high expression levels of pro-apoptotic protein and low expression of anti-apoptotic protein. Cell-cycle analysis showed a change in the cell-cycle distribution and arrested cells in G2-M phase. CONCLUSION: In this study, we found that OD inhibited proliferation and induced apoptosis in the human osteosarcoma MG-63 cell line in a time-dependent and dose-dependent manner. In addition, OD displayed inhibitory activity on MG-63 cell proliferation and invasion and the study also showed that OD activity might be mediated by caspase activation. These data suggest that OD might represent a novel, efficient candidate agent for further experimentation in osteosarcoma treatment.

A CONSORT-compliant, randomized, double-blind, placebo-controlled pilot trial of purified anthocyanin in patients with nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is a common liver disease that can progress to cirrhosis and liver failure. Anthocyanin, a member of the flavonoid family, has been shown to ameliorate NAFLD-associated pathologies in rodents.The aim of this CONSORT-compliant pilot study is to evaluate the effects of anthocyanin supplementation on insulin resistance and liver injury biomarkers in patients with NAFLD.A total of 74 subjects with NAFLD were divided into 2 groups in this double-blind, randomized study. Patients received either purified anthocyanin (320 mg/d) derived from bilberry and black currant or placebo for 12 weeks. Diet, physical activity, anthropometric parameters, glucose tolerance, and a set of biomarkers related to NAFLD were evaluated before and after intervention.No significant differences were observed in nutrient intake, physical activity, anthropometric parameters, or plasma lipid profile between patients receiving anthocyanin or placebo. Compared to controls, the anthocyanin group exhibited significant decreases (P < 0.05, all comparisons) in plasma alanine aminotransferase (-19.1% vs 3.1%), cytokeratin-18 M30 fragment (-8.8% vs 5.6%) and myeloperoxidase (-75.0% vs -44.8%). Significant decreases from baseline in fasting blood glucose and homeostasis model assessment for insulin resistance were observed in the anthocyanin group; however, these differences were not significant relative to placebo controls. In addition, the oral glucose tolerance test indicated that anthocyanin supplementation significantly decreased the 2-hour loading glucose level compared to control (-18.7% vs -3.8%, P = 0.02).A 12-week supplement of purified anthocyanin improved insulin resistance, indicators of liver injury, and clinical evolution in NAFLD patients. Further studies are warranted to determine the clinical applications of anthocyanin in NAFLD.This trial was registered at clinicaltrials.gov as NCT01940263.