Biomedical subjects

Jing Jie Yu

Publications and source records attributed to Jing Jie Yu.

7 recordsLinked to original sources

PubMed2006

MZF1 possesses a repressively regulatory function in ERCC1 expression.

ERCC1 is a critical gene within the nucleotide excision repair pathway. Overexpression of ERCC1 through promoter-mediating transcriptional regulation is associated with repair of cisplatin-induced DNA damage and clinical resistance to platinum-chemotherapy. Several transcriptional repressors and activators within the 5'-flanking region of the ERCC1 gene may be involved in the up-regulation of this gene. Minimal sequence within the promoter region required for ERCC1 transcription was analyzed by CAT assay and demonstrated that the region of -220 to -110 is essential to constitutive expression of ERCC1 gene in ovarian cancer cell line A2780/CP70. A more forward upstream region seems to be responsible for cisplatin-induced expression. Study of the functional cis-element in this region by electrophoretic mobility shift assay indicates that a MZF1-like site as well as an AP1-like site responded in a time-dependent manner to cisplatin stimulation with altered binding activities. EMSA with MZF1 ZN1-4 consensus oligonucleotides suggests that the MZF1 N-terminal domain of zinc finger cluster may bind to the MZF1-like site of the ERCC1 promoter region. MZF1 mRNA in A2780/CP70 cells decreased upon cisplatin exposure as analyzed by quantitative PCR, suggesting that MZF1 may mediate cisplatin-invoked gene expression in these cells. Overexpression of MZF1 repressed the ERCC1 promoter activity as determined in co-transfection assay, suggesting that MZF1 might be a repressor of ERCC1 transcription upon cisplatin exposure. In summary, our studies revealed a core promoter region and adjacent drug-responsible region within the ERCC1 promoter. The drug-responsible region contains cis-elements of activator, AP1 and repressor, MZF1. In response to cisplatin treatment, decreased MZF1 and increased AP1 binding activities appear to be the leading mechanism of up-regulation of ERCC1 expression. Our findings imply potential therapeutic strategies to antagonize drug resistant mechanisms in treatment of human ovarian cancer.

Adenocarcinoma↗

PubMed2006

Antisense telomerase RNA inhibits the growth of human glioma cells in vitro and in vivo.

Telomerase is implicated in the development of cellular immortality and oncogenesis. It has been shown that telomerase activity is considerably higher in the tissue of many different cancers than in normal tissue, and that the inhibition or downregulation of telomerase activity can prevent the malignant proliferation of tumor cells. Antisense oligonucleotides have been widely used in suppressing the expression of genes and, therefore, in the present research, we evaluated the effect of antisense human telomerase RNA (hTR) on glioma cell growth in vitro and in vivo. We showed that antisense hTR cDNA significantly inhibited TJ905 human glioma cell proliferation in vitro and tumor growth in vivo, as determined by MTT assay and by measuring the volume of glioma in nude mice. Consistent with these results, we found that telomerase activity and the mRNA levels of hTR and hTERT (human telomerase reverse transcriptase) expression were markedly decreased in tumor cells treated with antisense hTR cDNA, as assessed by TRAP (telomeric repeat amplification protocol) assay and RT-PCR (reverse transcription-polymerase chain reaction) analysis. Our study conclusively demonstrates that antisense hTR effectively inhibits the growth of human glioma cells in vitro and in vivo and, thus, may be potentially used for gene therapy of malignant gliomas and other cancers.

Actins↗

PubMed2006

Protein phosphatase 2A interacts with Chk2 and regulates phosphorylation at Thr-68 after cisplatin treatment of human ovarian cancer cells.

High-fidelity maintenance of genomic integrity in eukaryotes is ensured by cell cycle checkpoints and DNA repair. The checkpoint kinase, Chk2, has been implicated in both of these responses. In response to DNA damage, Chk2 is initially phosphorylated at Thr-68, which leads to its full activation. The fully activated Chk2 then phosphorylates downstream substrates of cell cycle control. However, the mechanism of inactivation of Chk2 is still unknown. Protein phosphatase type 2A (PP2A) plays an essential role in cell cycle regulation and induction of G2 arrest by a mechanism of phosphorylation/dephosphorylation with a variety of protein kinases. Data from our investigation provide evidence that, in response to cisplatin exposure, PP2A associates with Chk2 as a complex in cells and functions as a negative regulator of Chk2 activation by dephosphorylating p-Chk2. Results from immunostaining and coimmunoprecipitation demonstrate that Chk2 and PP2A can colocalize in cells, and the holoenzyme of PP2A (subunits A, B and C) coimmunoprecipitates with p-Chk2. Further, inhibition of PP2A by okadaic acid, an inhibitor of PP2A, and by small interfering RNA (siRNA) to PP2A results in enhanced Chk2 phosphorylation, implicating a direct enzyme-substrate relationship. An in vitro PP2A dephosphorylation assay shows that PP2A dephosphorylates p-Chk2 in a cell-free system. These findings suggest that the protein serine/threonine kinase, Chk2, is activated after cisplatin exposure and negatively regulated by a tightly associated protein serine/threonine phosphatase, PP2A.

Antineoplastic Agents↗

PubMed2004

Combinatorial treatment of ovarian cancer cells with harringtonine and cisplatin results in increased cisplatin-DNA adducts.

The current studies represent the first step in assessing the utility of harringtonine in combination with cisplatin as an improved approach for treating ovarian cancer. Three ovarian cancer cell lines, platinum-sensitive A2780, and platinum-resistant A2780/CP70 and OvCar-3, were exposed to their respective IC(50) dose of cisplatin for 1 h with or without a 24-h pretreatment with harringtonine. The level of platinum-DNA adducts was determined by atomic absorption spectrometry (AAS). These studies show for the first time that harringtonine pretreatment significantly increased the amount of platinum-DNA adducts in all ovarian cancer cell lines by 2-4 fold, immediately following 1-h exposure to cisplatin. Moreover, the level of cisplatin-DNA adducts in harringtonine-pretreated cells remained elevated by 3-4.7-fold for at least 6 h after cisplatin was removed, relative to cells only exposed to cisplatin. In all three cell lines the removal (repair) of platinum-DNA adducts was not significantly altered by harringtonine. In addition, the extent to which harringtonine altered the expression of select DNA damage response genes (p53, P16, ERCC1 and XPB) was determined using RT-PCR and Southern hybridization in A2780 and A2780/CP70 cells. The expression of ERCC1 and XPB RNAs were only modestly altered by harringtonine, consistent with a lack of effect of harringtonine on repair of cisplatin-DNA damage. However, harringtonine altered expression of p53 and P16 RNAs in both cell lines, although the down-regulation of p53 and P16 RNAs by harringtonine were more pronounced in A2780 cells. The novel observation that harringtonine augments platinum-DNA adducts in both platinum-sensitive and -resistant ovarian cancer cells indicates this combination of drugs may have utility in treating ovarian cancer and may be especially useful in managing platinum-resistant cancers. Additional studies are required to determine which sequence of these drugs is most beneficial, as well as the mechanism by which harringtonine increases cisplatin-DNA damage in ovarian cancer cells.

Actins↗

PubMed2004

Confirmation of 42-bp deletion within the ERCC1 5' UTR.

Our previous studies revealed a splicing variant (lacking a 42 base pair segment) within the 5'-UTR of the ERCC1 gene, a critical component of the nucleotide excision repair (NER) pathway that plays an important role in the development of chemoresistance in platinum-based anticancer therapy. This 42-bp segment seems to possess a regulatory function in ERCC1 expression and representing the level of clinical response to platinum-treatment in ovarian cancer patients. To confirm the existence of the 42-bp deletion and to investigate the 42-bp function, we performed several experiments and assays. Northern blot analysis and RNase protection assay provide evidence that the 42-bp deletion occurs at RNA level of ERCC1 5'-UTR in both ovarian cancer cell lines and ovarian cancer tissues. Luciferase assay suggests that this gene fragment possesses a regulatory function as an enhancer of ERCC1 gene expression in ovarian cancer cells. In Electrophoretic Mobility Shift Assay (EMSA), a shift band present in the ovarian cancer cell line extracts is consistent with the presence of an intracellular protein that recognizes this specific 42-bp sequence. Further, specific EMSA results with 42-bp probe mutated at the site of RFX-1 indicate different putative-DNA binding proteins, rather than RFX-1. We conclude that the 42-bp sequence within the 5'-UTR influences the expression of ERCC1 and hence can influence response to cisplatin in ovarian cancer therapy.

5' Untranslated Regions↗

PubMed2004

Excision repair cross complementing-group 1: gene expression and platinum resistance.

Platinum compounds induce their cytotoxic effect by binding to a DNA molecule in the form of a platinum-DNA-adduct. Many previous studies have shown that the level of platinum-DNA-adduct correlats with response to platinum-based chemotherapy. Although the mechanism of platinum resistance in vivo is not clearly understood, laboratory studies on cancer cell lines suggest that nucleotide excision repair (NER) is the main mechanism responsible for this resistance by increased platinum-DNA-adduct removal. NER pathway is a network of many proteins gathered in a DNA-repair system. The excision repair cross complementing-group 1 (ERCC1) gene has the leading role in NER-pathway because of its damage recognition and excision ability. In this report we reviewed the pathway leading to ERCC1 gene transcription and translation in cancer cells when exposed to cisplatin. We summarized data from different cancer cell lines and human cancers showing that the high level of ERCC1-mRNA and/or ERCC1 protein is associated with resistance to platinum compounds with direct impact on cancer patient survival and finally we analyzed drugs interfering with ERCC1 gene expression and causing the reversal of the platinum resistance when given to cancer cells prior to platinum-based chemotherapy.

Animals↗

PubMed2003

Clear cell tumors have higher mRNA levels of ERCC1 and XPB than other histological types of epithelial ovarian cancer.

PURPOSE: The purpose of the present work was to investigate the relationship between mRNA expression of ERCC1 and XPB, two key genes in the nucleotide excision repair pathway, and clinical resistance of platinum-chemotherapy in histological subtypes of epithelial ovarian cancer. EXPERIMENTAL DESIGN: mRNA levels of ERCC1 and XPB in epithelial ovarian cancer specimens from 126 different individuals were assessed using reverse transcription-PCR and followed by Southern hybridization methodology. Data were analyzed by linear regression analyses and by exhaustive regression analyses. RESULTS: Five different histological types of tumors were examined; serous (n = 76), mucinous (n = 11), clear cell (n = 9), poorly differentiated (n = 9), and endometroid (n = 21). Numerical values for mRNA expression levels were based on internal controls for a stable comparative cell line and for beta-actin. Median values for ERCC1 and XPB mRNAs within clear cell tumors were, on average, >2-fold higher than the other histological tumor types. Linear regression analyses suggest a continuum of nucleotide excision repair gene expression among these cell types, and exhaustive regression analyses demonstrate that the higher mRNA levels seen in clear cell tumors are highly statistically significant. CONCLUSIONS: We conclude that mRNA levels of ERCC1 and XPB tend to be higher in clear cell tumors as opposed to other types of epithelial ovarian cancer. This is consistent with the long-standing observation that clear cell tumors are more likely to show de novo drug resistance against DNA damaging agents in the clinic.

Adenocarcinoma, Clear Cell↗