Jun He

A regulatory circuit of miR-148a/152 and DNMT1 in modulating cell transformation and tumor angiogenesis through IGF-IR and IRS1

Dysregulation of microRNAs is a common feature in human cancers, including breast cancer (BC). Here we describe the epigenetic regulation of miR-148a and miR-152 and their impact on BC cells. Due to the hypermethylation of CpG island, the expression levels of both miR-148a and miR-152 (miR-148a/152) are decreased in BC tissues and cells. DNMT1, the DNA methyltransferase 1 for the maintenance methylation, is aberrantly up-regulated in BC and its overexpression is responsible for hypermethylation of miR-148a and miR-152 promoters. Intriguingly, we found that DNMT1 expression, which is one of the targets of miR-148a/152, is inversely correlated with the expression levels of miR-148a/152 in BC tissues. Those results lead us to propose a negative feedback regulatory loop between miR-148a/152 and DNMT1 in BC. More importantly, we demonstrate that IGF-IR and IRS1, often overexpressed in BC, are two novel targets of miR-148a/152. Overexpression of miR-148a or miR-152 significantly inhibits BC cell proliferation, colony formation, and tumor angiogenesis via targeting IGF-IR and IRS1 and suppressing their downstream AKT and MAPK/ERK signaling pathways. Our results suggest a novel miR-148a/152-DNMT1 regulatory circuit and reveal that miR-148a and miR-152 act as tumor suppressors by targeting IGF-IR and IRS1, and that restoration of miR-148a/152 expression may provide a strategy for therapeutic application to treat BC patients.

Roles and Mechanism of miR-199a and miR-125b in Tumor Angiogenesis

MicroRNAs (miRNAs) have been shown to be involved in different aspects of cancer biology including tumor angiogenesis. In this study, we identified that two miRNAs, miR-199a and miR-125b were downregulated in ovarian cancer tissues and cell lines. Overexpression of miR-199a and miR-125b inhibited tumor-induced angiogenesis associated with the decrease of HIF-1α and VEGF expression in ovarian cancer cells. Moreover, the levels of miR-199a and miR-125b were negatively correlated with VEGF mRNA levels in ovarian tissues. We further showed that direct targets of miR-199a and miR-125b HER2 and HER3 were functionally relevant. Forced expression of HER2 and HER3 rescued miR-199a- and miR-125b-inhibiting angiogenesis responses and Akt/p70S6K1/HIF-1α pathway. This study provides a rationale for new therapeutic approach to suppress tumor angiogenesis using miR-199a, miR-125b, or their mimics for ovarian cancer treatment in the future.

MicroRNA-143 inhibits tumor growth and angiogenesis and sensitizes chemosensitivity to oxaliplatin in colorectal cancers.

Colorectal cancer (CRC) is one of the leading cancer-related causes of death in the world. Recently, downregulation of microRNA-143 (miR-143) has been observed in CRC tissues. Here in this study, we found that miR-143 expression was downregulated both in CRC patients’ blood samples and tumor specimens. MiR-143 expression levels were strongly correlated with clinical stages and lymph node metastasis. Furthermore, insulin-like growth factor-I receptor (IGF-IR), a known oncogene, was a novel direct target of miR-143, whose expression levels were inversely correlated with miR-143 expression in human CRC specimens. Overexpression of miR-143 inhibited cell proliferation, migration, tumor growth and angiogenesis and increased chemosensitivity to oxaliplatin treatment in an IGF-IR-dependent manner. Taken together, these results revealed that miR-143 levels in human blood and tumor tissues are associated with CRC cancer occurrence, metastasis and drug resistance, and miR-143 levels may be used as a new diagnostic marker and therapeutic target for CRC in the future.

Reactive oxygen species regulate ERBB2 and ERBB3 expression via miR-199a/125b and DNA methylation.

Overexpression of ERBB2 or ERBB3 is associated with cancer development and poor prognosis. In this study, we show that reactive oxygen species (ROS) induce both ERBB2 and ERBB3 expression in vitro and in vivo. We also identify that miR‐199a and miR‐125b target ERBB2 and/or ERBB3 in ovarian cancer cells, and demonstrate that ROS inhibit miR‐199a and miR‐125b expression through increasing the promoter methylation of the miR‐199a and miR‐125b genes by DNA methyltransferase 1. These findings reveal that ERBB2 and ERBB3 expression is regulated by ROS via miR‐199a and miR‐125b downregulation and DNA hypermethylation.

Downregulation of ATG14 by EGR1-MIR152 sensitizes ovarian cancer cells to cisplatin-induced apoptosis by inhibiting cyto-protective autophagy

Cisplatin is commonly used in ovarian cancer treatment by inducing apoptosis in cancer cells as a result of lethal DNA damage. However, the intrinsic and acquired resistance to cisplatin in cancer cells remains a big challenge for improving overall survival. The cyto-protective functions of autophagy in cancer cells have been suggested as a potential mechanism for chemoresistance. Here, we reported MIR152 as a new autophagy-regulating miRNA that plays a role in cisplatin-resistance. We showed that MIR152 expression was dramatically downregulated in the cisplatin-resistant cell lines A2780/CP70, SKOV3/DDP compared with their respective parental cells, and in ovarian cancer tissues associated with cisplatin-resistance. Overexpression of MIR152 sensitized cisplatin-resistant ovarian cancer cells by reducing cisplatin-induced autophagy, enhancing cisplatin-induced apoptosis and inhibition of cell proliferation. A mouse subcutaneous xenograft tumor model using A2780/CP70 cells with overexpressing MIR152 was established and displayed decreased tumor growth in response to cisplatin. We also identified that ATG14 is a functional target of MIR152 in regulating autophagy inhibition. Furthermore, we found that EGR1 (early growth response 1) regulated the MIR152 gene at the transcriptional level. Ectopic expression of EGR1 enhanced efficacy of chemotherapy in A2780/CP70 cells. More importantly, these findings were relevant to clinical cases. Both EGR1 and MIR152 expression levels were significantly lower in ovarian cancer tissues with high levels of ERCC1 (excision repair cross-complementation group 1), a marker for cisplatin-resistance. Collectively, these data provide insights into novel mechanisms for acquired cisplatin-resistance. Activation of EGR1 and MIR152 may be a useful therapeutic strategy to overcome cisplatin-resistance by preventing cyto-protective autophagy in ovarian cancer.

Chronic arsenic exposure and angiogenesis in human bronchial epithelial cells via the ROS/miR-199a-5p/HIF-1α/COX-2 pathway.

Background: Environmental and occupational exposure to arsenic is a major public health concern. Although it has been identified as a human carcinogen, the molecular mechanism underlying the arsenic-induced carcinogenesis is not well understood. Objectives: We aimed to determine the role and mechanisms of miRNAs in arsenic-induced tumor angiogenesis and tumor growth. Methods: We utilized an in vitro model in which human lung epithelial BEAS-2B cells were transformed through long-term exposure to arsenic. A human xenograft tumor model was established to assess tumor angiogenesis and tumor growth in vivo. Tube formation assay and chorioallantoic membranes assay were used to assess tumor angiogenesis. Results: We found that miR-199a-5p expression levels were more than 100-fold lower in arsenic-transformed cells than parental cells. Re-expression of miR-199a-5p impaired arsenic-induced angiogenesis and tumor growth through its direct targets HIF-1α and COX-2. We further showed that arsenic induced COX-2 expression through HIF-1 regulation at the transcriptional level. In addition, we demonstrated that reactive oxygen species are an upstream event of miR-199a-5p/ HIF-1α/COX-2 pathway in arsenic-induced carcinogenesis. Conclusion: The findings establish critical roles of miR-199a-5p and its downstream targets HIF-1/COX-2 in arsenic-induced tumor growth and angiogenesis. Citation: He J, Wang M, Jiang Y, Chen Q, Xu S, Xu Q, Jiang BH, Liu LZ. 2014. Chronic arsenic exposure and angiogenesis in human bronchial epithelial cells via the ROS/miR-199a-5p/HIF-1α/COX-2 Pathway. Environ Health Perspect 122:255–261; http://dx.doi.org/10.1289/ehp.1307545

Arsenite induces cell transformation by reactive oxygen species, AKT, ERK1/2, and p70S6K1

Arsenic is naturally occurring element that exists in both organic and inorganic formulations. The inorganic form arsenite has a positive association with development of multiple cancer types. There are significant populations throughout the world with high exposure to arsenite via drinking water. Thus, human exposure to arsenic has become a significant public health problem. Recent evidence suggests that reactive oxygen species (ROS) mediate multiple changes to cell behavior after acute arsenic exposure, including activation of proliferative signaling and angiogenesis. However, the role of ROS in mediating cell transformation by chronic arsenic exposure is unknown. We found that cells chronically exposed to sodium arsenite increased proliferation and gained anchorage-independent growth. This cell transformation phenotype required constitutive activation of AKT, ERK1/2, mTOR, and p70S6K1. We also observed these cells constitutively produce ROS, which was required for the constitutive activation of AKT, ERK1/2, mTOR, and p70S6K1. Suppression of ROS levels by forced expression of catalase also reduced cell proliferation and anchorage-independent growth. These results indicate cell transformation induced by chronic arsenic exposure is mediated by increased cellular levels of ROS, which mediates activation of AKT, ERK1/2, and p70S6K1.

Downregulation of miR-145 associated with cancer progression and VEGF transcriptional activation by targeting N-RAS and IRS1

MicroRNA-145 (miR-145) is downregulated in various tumor types. However, its mechanism in inhibiting tumor growth and angiogenesis remains to be elucidated. In this study, we found that miR-145 was significantly downregulated in the plasma and cancer tumor tissues of colorectal cancer (CRC) patients, and overexpression of miR-145 inhibited cell proliferation, migration and invasion. To understand the potential mechanism of miR-145 in inhibiting tumor growth, we showed that miR-145 blocked the activation of AKT and ERK1/2 pathways, and the expression of HIF-1 and VEGF via directly targeting N-RAS and IRS1, and VEGF is an important effector for tumor growth. Forced expression of N-RAS and IRS1 restored VEGF expression via transcriptional activation. MiR-145 also inhibited N-RAS and IRS1 expression to suppress AKT and ERK1/2 activation, and VEGF expression in mouse xenograft tumors. To test the clinical relevance of these results, we used 60 pairs of colorectal cancer tissues and adjacent normal tissues, analyzed the levels of miR-145, N-RAS and IRS1 expression in these tissues, and found that miR-145 levels were significantly inversely correlated with N-RAS and IRS1 levels in these colorectal cancer tissues, suggesting the important implication of our findings in translational application for colorectal cancer diagnostics and treatment in the future.

Repression of miR-143 mediates Cr (VI)-induced tumor angiogenesis via IGF-IR/IRS1/ERK/IL-8 pathway

Hexavalent chromium [Cr (VI)] is a well-known human carcinogen associated with the increased risk of lung cancer. However, the mechanism underlying the Cr (VI)-induced carcinogenesis remains unclear due to the lack of suitable experimental models. In this study, we developed an in vitro model by transforming nontumorigenic human lung epithelial BEAS-2B cells through long-term exposure to Cr (VI). By utilizing this model, we found that miR-143 expression levels were dramatically repressed in Cr (VI)-transformed cells. The repression of miR-143 led to Cr (VI)-induced cell malignant transformation and angiogenesis via upregulation of insulin-like growth factor-1 receptor (IGF-IR) and insulin receptor substrate-1 (IRS1) expression. Moreover, we found that interleukin-8 is the major upregulated angiogenesis factor induced by Cr (VI) through activation of IGF-IR/IRS1 axis followed by activation of downstream ERK/hypoxia-induced factor-1α/NF-κB signaling pathway. These findings establish a causal role and mechanism of miR-143 in regulating Cr (VI)-induced malignant transformation and tumor angiogenesis.

NADPH oxidase subunit p22phox-mediated reactive oxygen species contribute to angiogenesis and tumor growth through AKT and ERK1/2 signaling pathways in prostate cancer

Excessive generation of reactive oxygen species (ROS) in cancer cells is associated with cancer development, but the underlying mechanisms and therapeutic significance remain elusive. In this study, we reported that levels of ROS and p22(phox) expression are greatly increased in human prostate cancer tissues, and knockdown of p22(phox) by specific small interfering RNA (siRNA) decreased ROS levels in prostate cancer cells. We also showed that stable downregulation of p22(phox) in prostate cancer cells inhibited cell proliferation and colony formation, which was mediated by AKT and extracellular signal-regulated kinase (ERK)1/2 signaling pathways and their downstream molecules hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF). The NADPH oxidase subunit NOX1 was also elevated in prostate cancer cells, and was involved in activation of AKT/ERK/HIF-1/VEGF pathway and regulation of cell proliferation. Knockdown of p22(phox) resulted in inhibition of tumor angiogenesis and tumor growth in nude mice. These findings reveal a new function of p22(phox) in tumor angiogenesis and tumor growth, and suggest that p22(phox) is a potential novel target for prostate cancer treatment.