PARP inhibitors suppress tumours via centrosome error-induced senescence independent of DNA damage response.

BACKGROUND: Poly(ADP-ribose) polymerase (PARP) inhibitors have emerged as promising chemotherapeutic drugs primarily against BRCA1/2-associated tumours, known as synthetic lethality. However, recent clinical trials reported patients' survival benefits from PARP inhibitor treatments, irrelevant to homologous recombination deficiency. Therefore, revealing the therapeutic mechanism of PARP inhibitors beyond DNA damage repair is urgently needed, which can facilitate precision medicine. METHODS: A CRISPR-based knock-in technology was used to establish stable BRCA1 mutant cancer cells. The effects of PARP inhibitors on BRCA1 mutant cancer cells were evaluated by biochemical and cell biological experiments. Finally, we validated its in vivo effects in xenograft and patient-derived xenograft (PDX) tumour mice. FINDINGS: In this study, we uncovered that the majority of clinical BRCA1 mutations in breast cancers were in and near the middle of the gene, rather than in essential regions for DNA damage repair. Representative mutations such as R1085I and E1222Q caused transient extra spindle poles during mitosis in cancer cells. PAR, which is synthesized by PARP2 but not PARP1 at mitotic centrosomes, clustered these transient extra poles, independent of DNA damage response. Common PARP inhibitors could effectively suppress PARP2-synthesized PAR and induce cell senescence by abrogating the correction of mitotic extra-pole error. INTERPRETATION: Our findings uncover an alternative mechanism by which PARP inhibitors efficiently suppress tumours, thereby pointing to a potential new therapeutic strategy for centrosome error-related tumours. FUNDING: Funded by National Natural Science Foundation of China (NSFC) (T2225006, 82272948, 82103106), Beijing Municipal Natural Science Foundation (Key program Z220011), and the National Clinical Key Specialty Construction Program, P. R. China (2023).

Protocol for identifying metabolite biomarkers in patient uterine fluid for early ovarian cancer detection.

High mortality of ovarian cancer (OC) is primarily attributed to the lack of effective early detection methods. Uterine fluid, pooling molecules from neighboring ovaries, presents an organ-specific advantage over conventional blood samples. Here, we present a protocol for identifying metabolite biomarkers in uterine fluid for early OC detection. We describe steps for uterine fluid collection from patients, metabolite extraction, metabolomics experiments, and candidate metabolite biomarker screening. This standardized workflow holds the potential to achieve early OC diagnosis in clinical practice. For complete details on the use and execution of this protocol, please refer to Wang et al.1.

Profiling the metabolome of uterine fluid for early detection of ovarian cancer.

Ovarian cancer (OC) causes high mortality in women because of ineffective biomarkers for early diagnosis. Here, we perform metabolomics analysis on an initial training set of uterine fluid from 96 gynecological patients. A seven-metabolite-marker panel consisting of vanillylmandelic acid, norepinephrine, phenylalanine, beta-alanine, tyrosine, 12-S-hydroxy-5,8,10-heptadecatrienoic acid, and crithmumdiol is established for detecting early-stage OC. The panel is further validated in an independent sample set from 123 patients, discriminating early OC from controls with an area under the curve (AUC) of 0.957 (95% confidence interval [CI], 0.894-1). Interestingly, we find elevated norepinephrine and decreased vanillylmandelic acid in most OC cells, resulting from excess 4-hydroxyestradiol that antagonizes the catabolism of norepinephrine by catechol-O-methyltransferase. Moreover, exposure to 4-hydroxyestradiol induces cellular DNA damage and genomic instability that could lead to tumorigenesis. Thus, this study not only reveals metabolic features in uterine fluid of gynecological patients but also establishes a noninvasive approach for the early diagnosis of OC.

A 1,2,3-Triazole Derivative of Quinazoline Exhibits Antitumor Activity by Tethering RNF168 to SQSTM1/P62.

Quinazoline and its derivatives have drawn much attention in the development of potential antitumor agents. Here, we synthesized a series of 1,2,3-triazole derivatives of quinazoline at the C6 position and evaluated for their cytotoxic activity in various human cancer cell lines. We found that compound 5a was the most cytotoxic to HCT-116 cells (IC50, 0.36 µM). Target profiling found that 5a directly binds to both the autophagy-associated protein SQSTM1/P62 and the E3 ligase RNF168, promoting their interaction. Consistently, 5a treatment induces a decrease in RNF168-mediated H2A ubiquitination and compromises homologous recombination-mediated DNA repair, thus increasing the sensitivity of HCT-116 to X-ray radiation. Moreover, 5a suppressed xenografted tumor growth in mice in a dose-dependent manner. Taken together, the 1,2,3-triazole derivative of quinazoline 5a may serve as a novel compound for tumor therapy based on its role in promoting a P62/RNF168 interaction.

The Apoptotic Resistance of BRCA1-Deficient Ovarian Cancer Cells is Mediated by cAMP.

Breast cancer type 1 susceptibility protein (BRCA1) is essential for homologous recombination repair of DNA double-strand breaks. Loss of BRCA1 is lethal to embryos due to extreme genomic instability and the activation of p53-dependent apoptosis. However, the apoptosis is resisted in BRCA1-deficient cancer cells even though their p53 is proficient. In this study, by analysis of transcriptome data of ovarian cancer patients bearing BRCA1 defects in TCGA database, we found that cAMP signaling pathway was significantly activated. Experimentally, we found that BRCA1 deficiency caused an increased expression of ADRB1, a transmembrane receptor that can promote the generation of cAMP. The elevated cAMP not only inhibited DNA damage-induced apoptosis through abrogating p53 accumulation, but also suppressed the proliferation of cytotoxic T lymphocytes by enhancing the expression of immunosuppressive factors DKK1. Inhibition of ADRB1 effectively killed cancer cells by abolishing the apoptotic resistance. These findings uncover a novel mechanism of apoptotic resistance in BRCA1-deficient ovarian cancer cells and point to a potentially new strategy for treating BRCA1-mutated tumors.

Cinnamaldehyde prevents intergenerational effect of paternal depression in mice via regulating GR/miR-190b/BDNF pathway.

Paternal stress exposure-induced high corticosterone (CORT) levels may contribute to depression in offspring. Clinical studies disclose the association of depressive symptoms in fathers with their adolescent offspring. However, there is limited information regarding the intervention for intergenerational inheritance of depression. In this study we evaluated the intervention of cinnamaldehyde, a major constituent of Chinese herb cinnamon bark, for intergenerational inheritance of depression in CORT- and CMS-induced mouse models of depression. Depressive-like behaviors were induced in male mice by injection of CORT (20 mg·kg-1·d-1, sc) for 6 weeks or by chronic mild stress (CMS) for 6 weeks. We showed that co-administration of cinnamaldehyde (10, 20, or 40 mg·kg-1·d-1, ig) for 6 weeks in F0 males prevented the depressive-like phenotypes of F1 male offspring. In addition, co-administration of cinnamaldehyde (20 mg·kg-1·d-1, ig) for 4 weeks significantly ameliorated depressive-like behaviors of chronic variable stress (CVS)-stimulated F1 offspring born to CMS mice. Notably, cinnamaldehyde had no reproductive toxicity, while positive drug fluoxetine showed remarkable reproductive toxicity. We revealed that CMS and CORT significantly reduced testis glucocorticoid receptor (GR) expression, and increased testis and sperm miR-190b expression in F0 depressive-like models. Moreover, pre-miR-190b expression was upregulated in testis of F0 males. The amount of GR on miR-190b promoter regions was decreased in testis of CORT-stimulated F0 males. Cinnamaldehyde administration reversed CORT-induced GR reduction in testis, miR-190b upregulation in testis and sperm, pre-miR-190b upregulation in testis, and the amount of GR on miR-190b promoter regions of F0 males. In miR-190b-transfected Neuro 2a (N2a) cells, we demonstrated that miR-190b might directly bind to the 3'-UTR of brain-derived neurotrophic factor (BDNF). In the hippocampus of F1 males of CORT- or CMS-induced depressive-like models, increased miR-190b expression was accompanied by reduced BDNF and GR, which were ameliorated by cinnamaldehyde. In conclusion, cinnamaldehyde is a potential intervening agent for intergenerational inheritance of depression, probably by regulating GR/miR-190b/BDNF pathway.

A synergetic effect of BARD1 mutations on tumorigenesis.

To date, a large number of mutations have been screened from breast and ovarian cancer patients. However, most of them are classified into benign or unidentified alterations due to their undetectable phenotypes. Whether and how they could cause tumors remains unknown, and this significantly limits diagnosis and therapy. Here, in a study of a family with hereditary breast and ovarian cancer, we find that two BARD1 mutations, P24S and R378S, simultaneously exist in cis in surviving cancer patients. Neither of the single mutations causes a functional change, but together they synergetically impair the DNA damage response and lead to tumors in vitro and in vivo. Thus, our report not only demonstrates that BARD1 defects account for tumorigenesis but also uncovers the potential risk of synergetic effects between the large number of cis mutations in individual genes in the human genome.

Pilot study of nitrogen removal from landfill leachate by stable nitritation-denitrification based on zeolite biological aerated filter.

A pilot (about 1 m3/d) process consisting of pre-denitrification and zeolite biological aerated filter (ZBAF) was established and run for nitrogen removal of landfill leachate. The results showed that stable nitritation and denitrification was achieved for landfill leachate with removal efficiency of Chemical Oxygen Demand (CODCr), ammonium and total nitrogen (TN) of 53.2 ±â€¯3.0%, 93.5 ±â€¯2.4% and 74.7 ±â€¯9.4%, respectively. Based on the ammonium adsorption equilibrium by zeolite, stable free ammonia could be maintained for inhibition of nitrite oxidizing bacteria (NOB) and dominance of ammonia oxidizing bacteria (AOB) in ZBAF, resulting in efficient nitritation with a nitrite accumulation ratio higher than 90.0% and an average nitrite production rate of 1.387 kg NO2--N m-3 day-1. High-throughput sequencing analysis further revealed enrichment of AOB and elimination of NOB in ZBAF. Compared to two-stage anoxic-oxic process, the pilot-scale process could save approximate 5000 mg/L glucose (about 3.10 US dollar/m3) with almost similar TN removal performance. All results obtained demonstrated the feasibility of the pilot process, which might be highly promising for the nitritation and denitrification of low C/N landfill leachate in the future.

An in vitro model of foam cell formation induced by a stretchable microfluidic device.

Although a variety of animal models of atherosclerosis have been developed, these models are time-consuming and costly. Here, we describe an in vitro model to induce foam cell formation in the early stage of atherosclerosis. This model is based on a three-dimension co-culture system in a stretchable microfluidic device. An elastic membrane embedded in the microfluidic device is capable of delivering nonuniform strain to vascular smooth muscle cells, endothelial cells and monocytes adhering thereto, which are intended to mimic the biological environment of blood vessels. Under low-density lipoprotein and stretch treatment, foam cell formation was successfully induced in co-culture with changes in mRNA and protein expression of some related key factors. Subsequently, the model was used to assess the inhibitory effect of atorvastatin on foam cell formation. The results obtained indicate that atorvastatin has a significantly dose-dependent inhibition of foam cell formation, which can be explained by the changes in mRNA and protein expression of the related factors. In principle, the model can be used to study the role of different types of cells in the formation of foam cells, as well as the evaluation of anti-atherosclerotic drugs.

Rapid start-up and performance of denitrifying granular sludge in an upflow sludge blanket (USB) reactor treating high concentration nitrite wastewater.

Denitrifying granular sludge reactor holds better nitrogen removal efficiency than other kinds of denitrifying reactors, while this reactor commonly needs seeding anaerobic granular sludge and longer period for start-up in practice, which restricted the application of denitrifying granular sludge reactor. This study presented a rapid and stable start-up method for denitrifying granular sludge. An upflow sludge blanket (USB) reactor with packings was established with flocculent activated sludge for treatment of high concentration nitrite wastewater. Results showed mature denitrifying granular sludge appeared only after 15 days with highest nitrogen removal rate of 5.844 kg N/(m3 day), which was much higher than that of compared anoxic sequencing batch reactor (ASBR). No significant nitrite inhibition occurred in USB and denitrification performance was mainly influenced by hydraulic retention time, influent C/N ratio and internal reflux ratio. Hydraulic shear force created by upflow fluid, shearing of gaseous products and stable microorganisms adhesion on the packings might be the reasons for rapid achievement of granular sludge. Compared to inoculated sludge and ASBR, remarkable microbial communitiy variations were detected in USB. The dominance of Proteobacteria and Bacteroidetes and enrichment of species Pseudomonas_stutzeri should be responsible for the excellent denitrification performance, which further verified the feasibility of start-up method.