Kai Ma

Upregulation of KLF4 by methylseleninic acid in human esophageal squamous cell carcinoma cells: Modification of histone H3 acetylation through HAT/HDAC interplay.

Esophageal squamous cell carcinoma (ESCC) occurs at a very high frequency in certain areas of China. Supplementation with selenium‐containing compounds was associated with a significantly lower cancer mortality rate in a study conducted in Linxia, China. Thus, selenium could be a potential anti‐esophageal cancer agent. In this study, methylseleninic acid (MSA) could inhibit cell growth of ESCC cells in vitro and in vivo. Upon treated with MSA, the activity of histone deacetylases (HDACs) was decreased and general control nonrepressed protein 5 (GCN5) was upregulated in ESCC cells. Meanwhile, a significant increase of H3K9 acetylation (H3K9ac) was detected. Upregulation of Krüppel‐like factor 4 (KLF4) was also observed after MSA treatment. Additionally, the acetylated histone H3 located more at KLF4 promoter region after MSA treatment, shown by chromatin immunoprecipitation (ChIP) assay. Moreover, knockdown of GCN5 decreased the protein level of both H3K9ac and KLF4, along with less cell growth inhibition. Taken all, our results indicated that MSA could inhibit ESCC cell growth, at least in part, by MSA‐HDAC/GCN5‐H3K9ac‐KLF4 axis. To our best knowledge, this is the first report that MSA induced acetylation of histone H3 at Lys9, which might depend on the activities and the balance between HDACs and HATs.

Methylseleninic acid activates Keap1/Nrf2 pathway via up-regulating miR-200a in human oesophageal squamous cell carcinoma cells.

Methylseleninic acid (MSA), as a potent second-generation selenium compound, could activate KLF4/miR-200a/Keap1/Nrf2 pathway in oesophageal squamous cell carcinoma cells.

Nuclear accumulation of Yes-Associated Protein (YAP) maintains the survival of doxorubicin-induced senescent cells by promoting survivin expression

Although chemotherapeutic drugs can induce senescence to prohibit further division of tumor cells, senescence could also promote tumorigenesis mainly through a senescence-associated secretory phenotype. Therefore, senescent tumor cells should be eliminated immediately to prevent drug resistance and recurrence. Here, we used a doxorubicin-induced senescence model to explore the mechanism underlying the survival of therapy-induced senescent cells. After low-dose doxorubicin treatment, tumor cells turned on a senescence program and became large and flattened, increasing their contact area with the extracellular matrix (ECM). Furthermore, Yes-associated protein (YAP) accumulated in the nucleus and YAP activity was increased in doxorubicin-induced senescent cells. Knockdown of YAP increased the sensitivity of cells to low-dose doxorubicin treatment, causing apoptosis rather than senescence. Moreover, the anti-apoptotic gene survivin, a YAP target gene, was overexpressed in senescent cells. Inhibition of survivin could lead to selective elimination of senescent cells through apoptosis. Our study indicates that nuclear accumulation of YAP could promote the survival of senescent cells by increasing survivin expression. Therefore, targeting YAP or survivin might be a new strategy for clearing senescent cancer cells during drug treatment.

Polarization of macrophages in the tumor microenvironment is influenced by EGFR signaling within colon cancer cells

Epidermal growth factor receptor (EGFR) is a target of colon cancer therapy, but the effects of this therapy on the tumor microenvironment remain poorly understood. Our in vivo studies showed that cetuximab, an anti-EGFR monoclonal antibody, effectively inhibited AOM/DSS-induced, colitis-associated tumorigenesis, downregulated M2-related markers, and decreased F4/80+/CD206+ macrophage populations. Treatment with conditioned medium of colon cancer cells increased macrophage expression of the M2-related markers arginase-1 (Arg1), CCL17, CCL22, IL-10 and IL-4. By contrast, conditioned medium of EGFR knockout colon cancer cells inhibited expression of these M2-related markers and induced macrophage expression of the M1-related markers inducible nitric oxide synthase (iNOS), IL-12, TNF-α and CCR7. EGFR knockout in colon cancer cells inhibited macrophage-induced promotion of xenograft tumor growth. Moreover, colon cancer-derived insulin-like growth factor-1 (IGF-1) increased Arg1 expression, and treatment with the IGF1R inhibitor AG1024 inhibited that increase. These results suggest that inhibition of EGFR signaling in colon cancer cells modulates cytokine secretion (e.g. IGF-1) and prevents M1-to-M2 macrophage polarization, thereby inhibiting cancer cell growth.

IL-6 influences the polarization of macrophages and the formation and growth of colorectal tumor

Macrophages play a crucial role in tumorigenesis depending upon the phenotype of macrophages found in tumor microenvironments. To date, how the tumor microenvironment affects the phenotypes of macrophages is not yet fully understood. In this study, we constructed a NIH3T3/Src cell line stably overexpresses the Src protein and found that conditioned medium from this cell line was able to induce polarization towards the M2 phenotype in primary bone marrow-derived macrophages (BMDM) and Ana-1 macrophages. Further investigation revealed that IL-6 produced by NIH3T3/Src cells plays a key role in M2 polarization. During the development of colorectal cancer in C57BL/6J-ApcMin/+ mice, increased IL-6 secretion in the interstitial fluid of the colorectal tissues was observed. Furthermore, tumorigenesis in IL-6tm1Kopf mice treated with AOM-DSS, an IL-6 knockout mouse strain, was significantly inhibited compared with the control group, suggesting the important role of IL-6 in promoting tumorigenicity. Our findings identify the target molecules and proinflammatory cytokines responsible for promoting polarization towards the M2 phenotype in macrophages present in tumor microenvironment, which may be useful for the design of novel therapeutic strategies for colorectal cancer.

Overexpression of KLF4 promotes cell senescence through microRNA-203-survivin-p21 pathway

Krüppel-like factor 4 (KLF4) is a transcription factor and functions as a tumor suppressor or tumor promoter in different cancer types. KLF4 regulates many gene expression, thus affects the process of cell proliferation, differentiation, and apoptosis. Recently, KLF4 was reported to induce senescence during the generation of induced pluripotent stem (iPS) cells, but the exact mechanism is still unclear. In this study, we constructed two doxycycline-inducing KLF4 cell models, and demonstrated overexpression of KLF4 could promote cell senescence, detected by senescence-associated β-galactosidase activity assay. Then we confirmed that p21, a key effector of senescence, was directly induced by KLF4. KLF4 could also inhibit survivin, which could indirectly induce p21. By miRNA microarray, we found a series of miRNAs regulated by KLF4 and involved in senescence. We demonstrated that KLF4 could upregulate miR-203, and miR-203 contributed to senescence through miR-203-survivin-p21 pathway. Our results suggest that KLF4 could promote cell senescence through a complex network: miR-203, survivin, and p21, which were all regulated by overexpression of KLF4 and contributed to cell senescence.

TAZ promotes cell growth and inhibits Celastrol-induced cell apoptosis.

TAZ could promote cell proliferation and inhibit Celastrol-induced cell apoptosis. Up-regulation of B-cell lymphoma-2 (Bcl-2), down-regulation of Bcl-2 associated X protein (Bax) and activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway may be the mechanism underlying anti-apoptosis of TAZ.

MicroRNA-492 overexpression involves in cell proliferation, migration and radiotherapy response of cervical squamous cell carcinomas.

MicroRNAs (miRNAs) are small non‐coding RNA that target protein‐coding mRNAs at the post‐transcriptional level. The aim of this study was to define the role of miR‐492 in cervical squamous cell carcinomas. After microRNA profiling and comparison, we firstly detected miR‐492 expression in 104 tumor tissues biopsies derived from advanced staged (FIGO IIB‐IIIB) cervical squamous cell carcinoma patients before receiving concomitant chemoradiotherapy and found miR‐492 expression was significantly higher in the specimens that were sensitive to concomitant chemoradiotherapy, as compared with insensitive cancer specimens (Pu2009<u20090.05). Moreover, higher expression of miR‐492 was associated with pelvic lymph node metastasis (LNM) (Pu2009<u20090.05). Further studies illustrated ectopic miR‐492 overexpression in SiHa cells promoted cell proliferation, migration, and enhanced the sensitivity of cervical cancer cells to irradiation by promoting apoptosis. In addition, we identified TIMP2 as a direct miR‐492 target, which has been shown to be critical in modulating cancer cell migration and invasion. We also confirmed that miR‐492 expression levels in positive pelvic LNM were much higher than negative LNM and miR‐492 played a vital role in pelvic lymph node metastasis via regulating miR‐492/TIMP2/MMP10 axis. In particular, miR‐492 was correlated with prognosis in the subgroup of patients with negative pelvic LNM (Pu2009<u20090.05) and had a promising value in predicting treatment response in the subgroup of patients with positive pelvic LNM (an AUC of 85%, 75.00% specificity, and 95.24% sensitivity). Taken together, the results suggested that miR‐492 may serve as a potential biomarker for cervical cancer treatment and prognosis.

Molecular, biological characterization and drug sensitivity of chidamide‑resistant non‑small cell lung cancer cells

Chidamide, a histone deacetylase (HDAC) inhibitor, has been applied in clinical trials for various types of hematological and solid tumors. Although acquired resistance is common in chemotherapy, the mechanism of resistance to chidamide is poorly characterized. The goal of the present study was to explore, in detail, the mechanism for the induced resistance to chidamide, and investigate a potential cross-resistance to other chemotherapeutic drugs. A549 cells were exposed to gradually increasing chidamide concentrations to establish a chidamide-resistant non-small cell lung cancer cell line (A549-CHI-R). The IC50 for chidamide, the proliferation inhibition rate, the total HDAC activity and the HDAC protein level were determined by an MTT assay, colony formation, a fluorometric HDAC activity assay and western blotting, respectively. Overexpression of the HDAC1 gene and HDAC1 gene-knockdown were achieved via plasmid transfection. A549-CHI-R cells demonstrated increased resistance to chidamide (8.6-fold). HDAC1 protein degradation was inhibited and HDAC activity was significantly higher in the A549-CHI-R cells relative to the parental A549 cells. A549-CHI-R cells demonstrated cross-resistance to paclitaxel, vinorelbine and gemcitabine, but not to cisplatin (CDDP) or 5-fluorouracil (5-FU). These results indicated that HDAC1 may be associated with resistance to chidamide, and HDAC1 may therefore be a predictive marker for chidamide sensitivity in cancer. In addition, A549-CHI-R cells remained sensitive to 5-FU and CDDP, indicating a potential strategy for cancer therapy.

Abstract 2253: YAP nuclear accumulation involved in drug-induced cellular senescence in vitro

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CAnnCellular senescence is accompanied by some striking morphological changes, such as becoming large, flat and multinucleated. However, how these changes are established is poorly understood. The Hippo pathway plays an important role in the control of organ size. Previous study reported that YAP, one major downstream effector of Hippo pathway, could be regulated by mechanical signals through F-actin bundles. However, whether YAP participates in the control of cell size through cytoskeleton remains unexplained. In our current study, we found that the senescent cells induced with adriamycin or camptothecin became large and flat, and the stress fibers of the senescent cells seemed to maintain or increase, observed by F-actin staining, though in a way of inappropriate actin clumping. Moreover increased nuclear level and activity of YAP were identified in drug-induced senescent cells. Hence, our data indicated that the increased F-actin bundles and mechanical signals could be acquired in senescent cells induced by adriamycin or camptothecin. In summary, based on our findings here, we proposed that YAP might play some roles during drug-induced cellular senescence in vitro.nnCitation Format: Kai Ma, Shuren Wang, Qing Xu, Mei Liu, Hongxia Zhu, Ningzhi Xu. YAP nuclear accumulation involved in drug-induced cellular senescence in vitro. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2253. doi:10.1158/1538-7445.AM2014-2253