Targeting ONECUT2 inhibits tumor angiogenesis via down-regulating ZKSCAN3/VEGFA.

OC-2 plays a vital role in tumor growth, metastasis and angiogenesis, but molecular mechanism how OC-2 regulates angiogenic factors is unclear. We found that OC-2 was highly expressed in HepG2, COLO, MCF-7, SKOV3 cells and rectum carcinoma tissues, and angiogenic factors levels were positively related to OC-2. Then OC-2 KD inhibited the tumor growth, metastasis and angiogenesis process in vitro and vivo. ChIP-Seq showed that 228 target genes of OC-2 were identified and they were associated with tumor growth, metastasis, angiogenesis and signal transduction; OC-2 bound to ZKSCAN3 at promoter region. Luciferase assays showed that ZKSCAN3 was identified as target gene of OC-2 and VEGFA was identified as target gene of ZKSCAN3; OC-2 promoted VEGFA expression via activating ZKSCAN3 transcriptional program. Importantly, OC-2 KD down-regulated VEGFA secretion to suppress tumor angiogenesis of HUVECs. Besides VEGFA, OC-2 was positively correlated with other angiogenic factors HIF-1α, FGF2, EGFL6 and HGF. Meanwhile, ERK1/2 and Smad1 signaling pathways might be related to function of OC-2 driving tumor aggressiveness. We revealed that OC-2 might regulate tumor growth, metastasis, angiogenesis via ERK1/2, Smad1 signaling pathways and regulate VEGFA expression for tumor angiogenesis via activating ZKSCAN3 transcriptional program, indicating that OC-2 was a convincing target to develop novel anti-tumor drugs based on angiogenesis.

Current state of research on copper complexes in the treatment of breast cancer.

Breast cancer, a malignancy originating from the epithelium or ductal epithelium of the breast, is not only highly prevalent in women but is also the leading cause of cancer-related deaths in women worldwide. Research has indicated that breast cancer incidence is increasing in younger women, prompting significant interest from scientists actively researching breast cancer treatment. Copper is highly accumulated in breast cancer cells, leading to the development of copper complexes that cause immunogenic cell death, apoptosis, oxidative stress, redox-mediated cell death, and autophagy by regulating the expression of key cell death proteins or assisting in the onset of cell death. However, they have not yet been applied to clinical therapy due to their solubility in physiological buffers and their different and unpredictable mechanisms of action. Herein, we review existing relevant studies, summarize the detailed mechanisms by which they exert anti-breast cancer effects, and propose a potential mechanism by which copper complexes may exert antitumor effects by causing copper death in breast cancer cells. Since copper death in breast cancer is closely related to prognosis and immune infiltration, further copper complex research may provide an opportunity to mitigate the high incidence and mortality rates associated with breast cancer.

A Morpholine Derivative N-(4-Morpholinomethylene)ethanesulfonamide Induces Ferroptosis in Tumor Cells by Targeting NRF2.

Small molecule drugs containing morpholine-based moieties have become crucial candidates in the tumor targeted therapy strategies, but the specific molecular mechanisms of these drugs causing tumor cell death require further investigation. The morpholine derivative N-(4-morpholinomethylene)ethanesulfonamide (MESA) was used to stimulate prostate and ovarian cancer cells and we focused on the ferroptosis effects, including the target molecule and signal pathways mediated by MESA. The results showed that MESA could induce ferroptosis to cause the proliferation inhibition and apoptosis effects of tumor cells according to the identification of ferroptosis inhibitor fer-1 and other cell death inhibitors. Further MESA could significantly increase the intracellular malondialdehyde (MDA), reactive oxygen species (ROS) and Fe2+ levels in tumor cells and mediate the dynamic changes of ferroptosis-relative molecules GPX4, nuclear factor erythroid2-related factor 2 (NRF2), ACSL4, SLC7A11 and P62-Kelch-like ECH-associated protein 1 (KEAP1)-NRF2-antioxidant response element (ARE) signal pathways. Further, NRF2 overexpression could reduce the tumor cell death and ROS levels exposure to MESA. Most importantly, it was confirmed that MESA could bind to NRF2 protein through molecular docking and thermal stability assays and NRF2 was a target molecule of MESA for inducing ferroptosis effects in tumor cells. Collectively, our findings indicated the ferroptosis effects of the morpholine derivative MESA in prostate and ovarian cancer cells and its function mechanism including targeted molecule and signal pathways, which would be helpful for developing MESA as a prospective small molecule drug for cancer therapy based on cell ferroptosis.

miR-129-2-3p mediates LPS-induced macrophage polarization and ferroptosis by targeting the SMAD3-GPX4 axis.

Macrophages has become a promising target of sepsis treatment because macrophages dysfunction contributes to the progress of sepsis. The targeted therapy of sepsis based on macrophages ferroptosis is drawing more and more attention, but the molecular mechanism involved is poorly understood. In this study, Mus musculus-derived macrophages were used for in-vitro experiments. We found that LPS could induce ferroptosis in macrophages via the detection of apoptosis, GSH, lipid peroxide and GPX4 levels. Meanwhile, miR-129-2-3p was up-regulated in macrophages exposure to LPS. Next, we confirmed that miR-129-2-3p promoted the LPS-induced polarization of M1 phenotype in macrophages via the detection of Arg-1 and iNOS levels; miR-129-2-3p promoted the LPS-induced ferroptosis in macrophages. Further, luciferase assay showed that SMAD3 was identified as a target gene of miR-129-2-3p and its expression was negatively regulated by miR-129-2-3p and LPS. SMAD3 could inhibit the LPS-induced polarization of M1 phenotype and ferroptosis in macrophages by targeting GPX4. Collectively, we demonstrated the target gene and molecular mechanism of miR-129-2-3p mediating LPS-induced polarization and ferroptosis in macrophages by targeting the SMAD3-GPX4 axis, which would provide a novel strategy for sepsis targeted therapy based on macrophages polarization and ferroptosis.

Universal point-of-care detection of proteins based on proximity hybridization-mediated isothermal exponential amplification.

We have developed a point-of-care (POC) lateral flow biosensor (LFB) for universal protein detection based on a proximity hybridization-mediated protein-to-DNA signal transducer, isothermal exponential amplification (EXPAR) and catalytic hairpin assembly (CHA) with high sensitivity and specificity. In this assay, a protein signal transducer was employed to convert the input protein to the output DNA signal. Antibody conjugated DNA1 was firstly hybridized with the output DNA (DNA3). The binding of antibody conjugated DNA1 and DNA2 to the same protein was able to increase the local concentrations, resulting in strand displacement between DNA3 and DNA2. DNA3 with nicking endonuclease recognition sequences at the 5' end then hybridized with hairpin probe 1 to mediate EXPAR in the presence of nicking endonuclease and polymerase. A large number of single strand DNA were produced in the circle of nicking, polymerization and strand displacement. The resulting ssDNA products were further amplified by CHA to generate double-stranded DNA products. The double-stranded DNA products were detected with a lateral flow biosensor within 5 min. This proposed assay has very high sensitivity and selectivity with a dynamic response ranging from 1 fM to 100 nM, and the detection limit was 0.74 fM. This work provides a universal and simple method for protein detection.

The metastasizing mechanisms of lung cancer: Recent advances and therapeutic challenges.

Lung cancer is one of the common malignant tumors that threaten human life with serious incidence and high mortality. According to the histopathological characteristics, lung cancer is mainly divided into non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC). NSCLC accounts for about 80-85% of lung cancers. In fact, lung cancer metastasis is a major cause of treatment failure in clinical patients. The underlying reason is that the mechanisms of lung cancer metastasis are still not fully understood. The metastasis of lung cancer cells is controlled by many factors, including the interaction of various components in the lung cancer microenvironment, epithelial-mesenchymal transition (EMT) transformation, and metastasis of cancer cells through blood vessels and lymphatics. The molecular relationships are even more intricate. Further study on the mechanisms of lung cancer metastasis and in search of effective therapeutic targets can bring more reference directions for clinical drug research and development. This paper focuses on the factors affecting lung cancer metastasis and connects with related molecular mechanisms of the lung cancer metastasis and mechanisms of lung cancer to specific organs, which mainly reviews the latest research progress of NSCLC metastasis. Besides, in this paper, experimental models of lung cancer and metastasis, mechanisms in SCLC transfer and the challenges about clinical management of lung cancer are also discussed. The review is intended to provide reference value for the future research in this field and promising treatment clues for clinical patients.

Imaging genomics for accurate diagnosis and treatment of tumors: A cutting edge overview.

Imaging genomics refers to the establishment of the connection between invasive gene expression features and non-invasive imaging features. Tumor imaging genomics can not only understand the macroscopic phenotype of tumor, but also can deeply analyze the cellular and molecular characteristics of tumor tissue. In recent years, tumor imaging genomics has been a key in the field of medicine. The incidence of cancer in China has increased significantly, which is the main reason of disease death of urban residents. With the rapid development of imaging medicine, depending on imaging genomics, many experts have made remarkable achievements in tumor screening and diagnosis, prognosis evaluation, new treatment targets and understanding of tumor biological mechanism. This review analyzes the relationship between tumor radiology and gene expression, which provides a favorable direction for clinical staging, prognosis evaluation and accurate treatment of tumors.

CRISPR/Cas9-Mediated OC-2 Editing Inhibits the Tumor Growth and Angiogenesis of Ovarian Cancer.

Ovarian cancer is the leading cancer-related cause of death in women worldwide. It is of great relevance to understand the mechanism responsible for tumor progression and identify unique oncogenesis markers for a higher chance of preventing this malignant disease. The high-expression OC-2 gene has been shown to be a potential candidate for regulating oncogenesis and angiogenesis in ovarian cancer. Hence, we wished to investigate the impact of OC-2 gene on ovarian cancer aggressiveness. CRISPR/Cas9, a gene editing tool, allows for direct ablation of OC-2 at the genomic level, and we successfully generated OC-2 KO cell lines from SKOV3 and CAOV3 cells. In an apoptosis assay, OC-2 KO induced the apoptosis activation of tumor cells, with the up-regulation of Bax/Caspase-8 and the down-regulation of Bcl-2. Consequently, the proliferation, migration, and invasion of OC-2 KO cell lines were significantly inhibited. Assays of qRT-PCR and Western blotting showed that the expression levels of pro-angiogenic growth factors VEGFA, FGF2, HGF, and HIF-1α and the activation of Akt/ERK pathways were significantly down-regulated at the loss of OC-2. In the xenograft model, OC-2 KO potently suppressed the subcutaneous tumor growth, with the inhibition exceeding 56%. The down-regulation of CD31 and relevant pro-angiogenic growth factors were observed in OC-2 KO tumor tissues. Taken together, OC-2 depletion negatively regulated the ovarian cancer progression possibly by apoptosis activation and angiogenesis inhibition. This work revealed a pivotal regulator of apoptosis and angiogenesis networks in ovarian cancer, and we applied the CRISPR/Cas9 system to the transcription factor pathway for developing a broad-acting anti-tumor gene therapy.

miR-6086 inhibits ovarian cancer angiogenesis by downregulating the OC2/VEGFA/EGFL6 axis.

miRNAs have emerged as a pivotal component of gene regulatory networks, mediating cytokines secretion, cell cycle, and differentiation regulation. However, how miRNAs collaborate with transcription factors and downstream effector proteins that determine the fate of ovarian cancer cells remains to be understood, especially regarding to mechanism of tumor angiogenesis regulation. Based on the qRT-PCR and IHC analysis, we found that miR-6086 was maintained a very low level both in ovarian cancer cell lines and tissues. Further, we identified OC2 and EGFL6 as the direct targets of miR-6086 by luciferase assay and we observed an inverse relationship between the expression of miR-6086 and the OC2/VEGFA/EGFL6 axis. The Western blotting analysis suggested that OC2 could directly upregulate VEGFA and indirectly up-regulate EGFL6 through VEGFA. Moreover, miR-6086 could indirectly downregulate VEGFA through OC2. In addition, miR-6086, siOC2 and siEGFL6 could negatively regulate the tumor growth and angiogenesis of ovarian cancer (Skov3) in the animal studies, with the inhibition rates of 77.07%, 69.89%, and 73.62%, respectively (**p < 0.01). Moreover, the tumor cell proliferation, migration, and invasion of ovarian cancer cell lines (Caov3 and Skov3) and vascular formation (HUVECs) were significantly suppressed in vitro, by decreasing the AKT/MAPK pathways (*p < 0.05). Taken together, our results reveal that miR-6086 can suppress the angiogenesis networks in ovarian cancer by down-regulating the OC2/VEGFA/EGFL6 axis, directly or indirectly, which may provide potential targets for tumor therapeutics.

Blockade of ONECUT2 expression in ovarian cancer inhibited tumor cell proliferation, migration, invasion and angiogenesis.

One cut homeobox 2 (ONECUT2 or OC-2) is a newly discovered transcription factor. Aberrant expression of OC-2 is closely related to cell proliferation, migration, invasion, and angiogenesis. In this study, we found that OC-2 expression was upregulated in ovarian adenocarcinoma cells, by Western blot analysis. The results of immunohistochemistry showed that the expression of OC-2 was also increased in malignant ovarian cancer tissue. In order to explore the role of OC-2 in the development of ovarian cancer, siRNAs that specifically targets OC-2 were designed. The siRNA targeting OC-2 could effectively inhibit the vascular endothelial growth factor A (VEGFA) expression, but silence and overexpression of VEGFA did not affect OC-2 expression. In addition, OC2-siRNA could block the proliferation, migration, and invasion, and inhibit epithelial-mesenchymal transition and the AKT/ERK signaling pathway, of human ovarian cancer cells in vitro. In a mouse model of ovarian cancer xenograft tumors, OC2-siRNA could significantly inhibit tumor cell growth and the tumor inhibition rate reached approximately 73%. The results of immunohistochemistry showed that the densities of microvessels stained with CD31, the expression of OC-2 and VEGFA were significantly decreased in tumors. These data indicated that OC-2 was an upstream regulator of VEGFA and silencing OC-2 could inhibit ovarian cancer angiogenesis and tumor growth.

Assessing the role of IL-35 in colorectal cancer progression and prognosis.

Despite the recent realization of Interleukin (IL)-35 in tumorigenesis, its exact impact on colorectal cancer (CRC) progression and prognosis, however, is yet to be elucidated clearly. We thus in the present report conducted comparative analysis of IL-35 levels between CRC patients and matched control subjects. IL-35 is highly expressed in all CRC tissues, which can be detected in vast majority of colorectal cancer cells. IL-35 levels in CRC lysates and serum samples are highly correlated to the severity of malignancy and the clinical stage of tumor. Particularly, a significant reduction for serum IL-35 was noted in patients after surgical resection, indicating that IL-35 promotes CRC progression associated with poor prognosis. Mechanistic study demonstrated a significant correlation between serum IL-35 levels and the number of peripheral regulatory T (Treg) cells in CRC patients, suggesting that IL-35 implicates in CRC pathogenesis probably by inducing Treg cells, while cancer cell-derived IL-35 may also recruit Treg cells into the tumor microenvironment in favor of tumor growth. Together, our data support that IL-35 could be a valuable biomarker for assessing CRC progression and prognosis in clinical settings.

[Differentiation of human promyelocytic leukemia HL-60 cells induced by proanthocyanidin and its mechanism].

This study was purposed to investigate the proliferation, differentiation and apoptosis of human promyelocytic leukemia HL-60 cells induced by proanthocyanidin (PAC). HL-60 cells were incubated with 20 mg/L PAC for 24 h, the cell growth was evaluated by CCK-8 assay. the effect of PAC on HL-60 cells was evaluated and the cells morphology was observed by optical microscopy. Expression of CD14 and CD11b, and cell cycle were analyzed by flow cytometry. The results showed that the growth of HL-60 cells was inhibited after treatment with PAC of different concentration in a dose-dependent manner (P < 0.05). 20 mg/L PAC displayed significant effect on HL-60 cells with inhibition ratio (72.3 ± 1.8)% for 24 h. Microscopy displayed that some cells differentiated to relative mature cells after treating for 48 h. Expression of CD14 increased and the expression of CD11b increased a little after treating with 20 mg/L PAC for 24 h, the ratio of cells in G0/G1 phase increased, but the ratio of cells in S phase decreased. The mRNA and protein expression of P21 gene increased, but the protein expression of CDK4 and Cyclin D1 decreased. It is concluded that PAC may inhibit the proliferation of HL-60 cells in vitro, induces the differentiation of HL-60 cells, and arrests the cells in G0/G1 phase. The possible mechanism may be related to up-regulation of P21 gene expression and down-regulation of the protein expression of CDK4 and Cyclin D1.

Overexpression of PTEN induces cell growth arrest and apoptosis in human breast cancer ZR-75-1 cells.

Phosphatase and tensin homolog (PTEN) is a tumor suppressor gene located at human chromosome 10q23, might play an important role in cell proliferation, cell cycle and apoptosis of cancer cells. In this study, the eukaryotic expression vectors pBP-wt-PTEN (containing a wild-type PTEN gene) and pBP-G129R-PTEN (containing a mutant PTEN gene) were used to transfect breast cancer ZR-75-1 cells. After transfection, ZR-75-1 cells expressing PTEN were obtained and tested. The blue exclusion assay showed the growth rate of the cells transfected with pBP-wt-PTEN was significantly lower than that of the control cells transfected with pBP-G129R-PTEN. Analysis of the cell cycle by flow cytometry showed that the progression from the G(1) to the S phase was arrested in cells expressing wild-type PTEN. Some typical morphological changes of apoptosis were also observed in cells transfected with pBP-wt-PTEN, but not in those transfected with pBP-G129R-PTEN. This study shows that overexpression of PTEN in ZR-75-1 cells leads to cell growth arrest and apoptosis.