Elevated XRCC5 expression level can promote temozolomide resistance and predict poor prognosis in glioblastoma

Drug resistance and disease recurrence are important contributors for the poor prognosis of glioblastoma multiforme (GBM). Temozolomide (TMZ), the standard chemotherapy for GBM treatment, can methylate DNA and cause the formation of double-strand breaks (DSBs). X-ray repair cross complementing 5 (XRCC5), also known as Ku80 or Ku86, is required for the repair of DSBs. The present study identified novel determinants that sensitize cells to TMZ, using an array-based short hairpin (sh)RNA library. Then, cBioportal, Oncomine, and R2 databases were used to analyze the association between gene expression levels and clinical characteristics. Subsequently, lentiviral shRNA or pCMV was used to knockdown or overexpress the gene of interest, and the effects on TMZ sensitivity were determined using a MTT assay and western blot analysis. TMZ-resistant cells were also established and were used in in vitro and in vivo experiments to analyze the role of the gene of interest in TMZ resistance. The results indicated that XRCC5 was effective in enhancing TMZ cytotoxicity. The results from the bioinformatics analysis revealed that XRCC5 mRNA expression levels were associated with clinical deterioration and lower overall survival rates. In addition, XRCC5 knockdown could significantly increase TMZ sensitivity in GBM cells, while XRCC5 overexpression caused the cancer cells to be resistant to TMZ. Both the in vivo and in vitro experiments showed that TMZ treatment could induce expression of XRCC5 in TMZ-resistant cells. Taken together these findings suggested that XRCC5 could be a promising target for GBM treatment and could also be used as a diagnostic marker for refractory GBM.     

Human chondrosarcoma secretes vascular endothelial growth factor to induce tumor angiogenesis and stores basic fibroblast growth factor for regulation of its own growth.

Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are well-known factors that induce neovascularization in many tumors. The molecular mechanisms that regulate tumor angiogenesis in human chondrosarcoma are not clear. We assessed in this work the angiogenic activities of a human chondrosarcoma cell line (OUMS-27) in vivo and determined the efficacies of angiogenic factors derived from OUMS-27 cells on human umbilical vein endothelial cells (HUVECs) in vitro. Tumor xenografts induced an increase in the formation of neovessels, but the distributions of Ki-67 antigen, VEGF and bFGF were unaffected. We also demonstrated that OUMS-27 cells secreted VEGF(165) into the culture medium and that it was the maximal angiogenic factor to stimulate endothelial proliferation and migration in chondrosarcoma. Anti-VEGF antibodies induced an approximately 70% inhibition of these responses of HUVECs, but did not have any effect on OUMS-27 cells. Anti-bFGF antibodies suppressed not only the activities of HUVECs but also the growth of tumor cells in vitro. We indicate that angiogenesis is principally elicited by VEGF(165) and that tumorigenesis is mainly regulated by bFGF stored in the extracellular matrix of OUMS-27 cells. The present study may offer the availability of combination therapies for inhibition of VEGF and bFGF action on vascular endothelial cells and chondrosarcoma cells, respectively.     

Influence of intratumoral basic fibroblast growth factor concentration on survival in ovarian cancer patients

Since basic fibroblast growth factor (bFGF) is considered as a potent mitogen that stimulates the growth of ovarian cancer cells, we evaluated the role of bFGF as a prognostic marker in patients with epithelial ovarian cancer. bFGF was quantified from the tumor cytoplasm of 76 patients with FIGO stage I–III ovarian cancer by a human FGF basic immunoassay (R&D Systems). After a mean follow-up period of 42 months, 50 patients were found to be free of tumor while 26 patients had died of the disease. The median bFGF concentration was 352.9 pg/mg (range 27.4–26 600 pg/mg). After dichotomization cytoplasmic expression of bFGF was found to be low in 44 tumors (≤500 pg/mg) and high in 32 tumors (>500 pg/mg). The probability of overall survival was 38.8 and 58.5% in the low bFGF and high bFGF groups, respectively (log-rank P=0.0066). In multivariate analysis, residual tumor after initial surgery and bFGF, but not histologic grade or stage of the disease, independently influenced the overall survival probability. Furthermore, tumors with high cytoplasmic expression of bFGF revealed a much greater stromal content. Therefore, we hypothesize that bFGF may induce a fibroblastic response which causes tumors with a high bFGF to be less aggressive than those with less stromal tissue.     

Role of transforming growth factor beta in the growth inhibition of human breast cancer cells by basic fibroblast growth factor

Recent studies from our laboratory have revealed that basic fibroblast growth factor (bFGF) selectively inhibits the proliferation of human MCF-7 breast cancer cells. It has also been shown to enhance cis-platinum-induced apoptosis, decrease levels of the anti-apoptotic gene product bcl-2, and increase levels of the cyclin-dependent protein kinase inhibitor p21/WAF1/Cip1. Transforming growth factor beta-1 (TGF₁), a cell growth regulator has been found to have an inhibitory effect on breast cancer cells. The aim of the present study was to evaluate the possible role of TGF₁ in the antiproliferative effects of bFGF in MCF-7 breast cancer cells. We found that exogenous, as well as endogenous (overexpressed) bFGF increased TGF₁ mRNA expression in the cells and enhanced the secretion of TGF₁ into culture medium. However, exogenous addition of TGF₁ neither led to a decrease in bcl-2 nor induced an increase in the levels of p21/WAF1/Cip1 and neutralizing antibodies to TGF₁, did not reverse bFGF-induced G₁ arrest nor the increase in p21/WAF1/Cip1 level. In contrast, antisense oligonucleotides to TGF₁ abrogated the antiproliferative effects and inhibited the induction of p21/WAF1/Cip1 by bFGF in MCF-7 cells. These data suggest that the anti-proliferative effects of bFGF in human MCF-7 breast cancer cells are mediated by endogenous TGF₁, while exogenous TGF₁ does not mimic all the effects of bFGF on these breast cancer cells. These findings provide an important basis for further investigations into the autocrine and paracrine processes that control the growth of breast cancer cells.     

碱性成纤维细胞生长因子治疗白血病化疗后口腔溃疡

目的　探讨碱性成纤维细胞生长因子在白血病化疗后口腔溃疡治疗中的疗效和安全性。方法　 60例病人随机分为两组 ,各 3 0例。治疗组口腔局部应用碱性成纤维细胞生长因子 ,配合常规处理 ;对照组单纯应用常规处理。结果　治疗组总有效率 86.7% ,对照组总有效率 46.7%。两组总有效率比较差异有非常显著意义 (P <0 .0 1) ,无严重不良反应。结论　碱性成纤维细胞生长因子治疗白血病化疗后口腔溃疡疗效好 ,安全可靠。     

碱性成纤维细胞生长因子在前列腺癌中的表达

目的探讨碱性成纤维细胞生长因子(bFGF)在前列腺癌中表达的临床意义。方法应用半定量RT-PCR技术和免疫组织化学方法,检测22例前列腺癌组织中bFGF的表达,并与正常的前列腺组织进行比较。结果前列腺癌组织中的bFGF表达高于正常的前列腺组织(P<0.05);bFGF表达升高的程度与前列腺癌病理分级和临床分期均无明显关系。结论前列腺癌组织中bFGF的表达高于正常前列腺组织,提示bF-GF与前列腺癌的发生发展密切相关。     

Expression of basic fibroblast growth factor is associated with poor outcome in non-Hodgkin's lymphoma

It is now clear that angiogenesis and angiogenesis factors are important in the pathogenesis of haematological malignancies. High pretreatment levels of serum basic fibroblast growth factor have been shown to be associated with poor prognosis in patients with non-Hodgkin's lymphoma. The aim of this study was to evaluate whether non-Hodgkin's lymphoma cells express basic fibroblast growth factor and/or its receptor (fibroblast growth factor receptor-1) and whether basic fibroblast growth factor expression correlates with basic fibroblast growth factor serum levels, intratumoral microvessel density, and patient outcome. We measured basic fibroblast growth factor by enzyme-linked immunosorbent assay in sera taken from 58 patients with non-Hodgkin's lymphoma before treatment and in 19 of them also after treatment. Pathological specimens at diagnosis were evaluated by immunohistochemistry staining using polyoclonal antibody against factor-VIII-related antigen, basic fibroblast growth factor and fibroblast growth factor receptor-1 to determine the expression of the microvessel count and basic fibroblast growth factor and fibroblast growth factor receptor-1. The lymphoma specimens demonstrated positive staining for basic fibroblast growth factor (in 23%) and fibroblast growth factor receptor-1 (in 58.5%). The patients who expressed basic fibroblast growth factor had a significantly worse progression-free and overall survival than those who did not (P=0.003 and P=0.03 respectively), while patients expressing fibroblast growth factor receptor-1 were less likely to achieve complete remission than those lacking the receptor (33% vs 65%, P=0.047). There was no correlation of basic fibroblast growth factor staining with either serum basic fibroblast growth factor levels or microvessel count. Basic fibroblast growth factor serum levels did not change significantly after treatment These results suggest that non-Hodgkin's lymphoma specimens express basic fibroblast growth factor and its receptor (fibroblast growth factor receptor-1) and this expression is associated with poor patient outcome.     

PI3K/AKT is involved in mediating survival signals that rescue Ewing tumour cells from fibroblast growth factor 2-induced cell death

While in vitro studies had shown that fibroblast growth factor 2 (FGF2) can induce cell death in Ewing tumours, it remained unclear how Ewing tumour cells survive in vivo within a FGF2-rich microenvironment. Serum- and integrin-mediated survival signals were, therefore, studied in adherent monolayer and anchorage-independent colony cell cultures. In a panel of Ewing tumour cell lines, either adhesion to collagen or exposure to serum alone only had a minor protective effect against FGF2. However, both combined led to complete resistance to 5 ng ml(-1) FGF2 in three of four FGF2-sensitive cell lines (RD-ES, RM-82 and WE-68), and to an increased survival as compared to other culture conditions in TC-71 cells. Inhibition studies with LY294002 demonstrated that the serum signal is mediated via the phosphoinositide 3-OH kinase/AKT pathway. Thus, Ewing tumour cells escape FGF2-induced cell death by modulating FGF2 signalling. The tumour microenvironment provides the necessary survival signals by integrin-mediated adhesion and soluble serum factor(s). These survival signals warrant further investigation as a potential resistance mechanism to other apoptosis-inducing agents in vivo.     

Tumour formation by single fibroblast growth factor receptor 3-positive rhabdomyosarcoma-initiating cells

Background:

The hypothesis that malignant tumours are generated by rare populations of cancer stem cells that are more tumourigenic than other cancer cells has gained increasing credence. The objective of this study was to identify and characterise a subpopulation of human sarcoma-initiating cells.

Methods:

We examined established rhabdomyosarcoma cell lines by flow cytometry. Tumourigenesis was examined by xenograft models. Real-time PCR and immunohistochemistry were performed to examine the gene expression using cell lines and biopsy specimens.

Results:

Rhabdomyosarcoma cell lines included small populations of fibroblast growth factor receptor 3 (FGFR3)-positive cells. FGFR3-positive KYM-1 and RD cells were more strongly tumourigenic than FGFR3-negative cells. In addition, xenoengraftment of 33% of single FGFR3-positive KYM-1 cells yielded tumour formation. Stem cell properties of FGFR3-positive cells were further established by real-time PCR, which demonstrated upregulation of undifferentiated cell markers and downregulation of differentiation markers. We showed that in the absence of serum, addition of basic fibroblast growth factor maintained and enriched FGFR3-positive cells. On the other hand, ciliary neurotrophic factor reduced the proportion of FGFR3-positive cells. Real-time PCR and immunohistochemical examination revealed that embryonal rhabdomyosarcoma patient biopsy specimens were found to over-express FGFR3.

Conclusions:

Our findings suggest that rhabdomyosarcoma cell lines include a minor subpopulation of FGFR3-positive sarcoma-initiating cells, which can be maintained indefinitely in culture and which is crucial for their malignancy.     

γ-干扰素对喉癌细胞株碱性成纤维细胞生长因子表达的影响

目的:研究γ-干扰素(IFN-γ)对人喉癌细胞株Hep-2细胞碱性成纤维细胞生长因子(bFGF)基因及蛋白表达的影响。方法:将IFN-γ以不同浓度[(1、10、100、1000、10000)U/ml]、不同作用时间[(0、12、24、36、48、60、72)h]作用于Hep-2细胞,用四甲基偶氮唑蓝(MTT)比色法测定细胞增殖;用实时荧光定量PCR(Realtime-PCR)法测定IFN-γ(1000U/ml)不同作用时间[(0、2、6、12、24、36、48、60、72)h]细胞bFGFmRNA含量;用双抗体夹心酶联免疫吸附试验(ELISA)和免疫细胞化学方法测定与Realtime-PCR相同药物剂量和作用时间细胞培养上清液和细胞浆中bFGF蛋白含量。结果:细胞生长受到抑制,以高剂量组表现最为明显。不同浓度(1、10、100、1000、10000U/ml)IFN-γ作用不同时间[(0、12、24、36、48、60、72)h]后,从48h开始,(100、1000、10000)U/ml的IFN-γ对Hep-2细胞有明显的抑制作用,差异有显著性(P<0.05),但不随着浓度的增加而成正比。1000U/mlIFN-γ作用后,在2h后与对照组比,bFGFmRNA表达上调,差异有显著性(P<0.05),特别是48h后,有显著性差异(P<0.01)。1000U/mlIFN-γ作用48h后,与对照组比较,Hep-2细胞浆中的棕色颗粒明显增多。结论:IFN-γ抑制Hep-2细胞分泌bFGF却,不抑制其合成。     

前列腺癌细胞株VEGF及其受体KDR,bFGF及其受体FGFR2的表达及意义

目的:探讨人前列腺癌细胞株血管内皮细胞生长因子(VEGF)及其受体(KDR),碱性成纤维细胞生长因子(bFGF)及其受体(FGFR2)的表达,进一步阐明前列腺癌细胞中VEGF和bFGF的自分泌机制。方法:以小鼠成纤维细胞系L929作为对照,选取三种前列腺癌细胞株(PC3、LNcap和DU145),采用免疫组化染色、RT-PCR及Westernblot,检测VEGF及其受体KDR,bFGF及其受体FGFR2的表达。结果:三种前列腺癌细胞株(PC3、LNcap和DU145)中均有VEGF、KDR及bFGF、FGFR2的表达,但表达水平略有差别。结论:在前列腺癌的血管形成中可能存在VEGF和bFGF的自分泌机制。     

Upregulation of basic fibroblast growth factor in breast carcinoma and its relationship to vascular density, oestrogen receptor, epidermal growth factor receptor and survival

Background: Angiogenesis, the process whereby endothelial cells divide and migrate to form new blood capillaries, has been assessed in tumours by measuring microvessel density. High microvessel density is a significant adverse prognostic factor in breast cancer. The angiogenic factor, basic fibroblast growth factor (bFGF), has been associated with tumourigenesis and metastasis in several human cancers. There are few quantitative studies of bFGF expression in normal tissues compared to cancer.Patients and methods: We have measured bFGF levels in 149 human primary breast carcinomas and assessed the findings in relation to microvessel density, oestrogen receptor (ER) and epidermal growth factor receptor (EGFR).Basic FGF levels were measured by ELISA. Western blotting and immunohistochemistry were carreid out to confirm the presence of bFGF.Results: Levels of bFGF were more than 10-fold higher in tumour cytosols compared to reduction mammoplasty tissue and 3-fold compared to non neoplastic cytosols from the same breast as the tumour (P < 0.0001). Immunohistochemistry showed bFGF protein was localised exclusively in the stroma whereas no bFGF staining was observed in the epithelial cells. High bFGF levels were significantly related to high ER (P = 0.01). Similarly, high bFGF levels were significantly related to low grade (P = 0.046) and to small tumour size (P = 0.04). No significant relationship was observed between bFGF and microvessel count, EGFR or age. In univariate analysis and in a Cox proportional hazard model bFGF did not reach significance for overall or relapse free survival.Conclusions: Our results show that although bFGF is elevated in breast carcinomas compared to normal breast tissue it is not related to microvessel density and it is not an independent predictor of survival in breast cancer patients. Basic FGF may be one of multiple factors that synergise with other growth factors such as VEGF to enhance angiogenesis.     

Epstein-Barr virus latent membrane protein 1 promotes concentration in multivesicular bodies of fibroblast growth factor 2 and its release through exosomes

FGF-2, a potent angiogenic factor that is involved in tumor invasion, is known to be released extracellularly by a nonclassical secretory pathway. Recently it has become clear that Epstein-Barr virus, specifically its oncoprotein LMP1, can induce expression of angiogenic factors. Among these factors is FGF-2. LMP1 not only promotes expression of FGF-2, but also the release extracellularly of its 18-kDa isoform. We analyzed the mechanism of FGF-2 release induced by LMP1. Confocal immunofluorescence microscopy revealed colocalization of FGF-2 with LMP1 in small dots also stained positively for CD63 and cathepsin D, markers of late endosomes or multivesicular bodies. Biochemical analysis and immunoelectron microscopy of purified exosomal fractions from cotransfected cells demonstrated increased release of exosomes and the concentration of LMP1 and FGF-2 in these structures. Moreover, cotransfection appeared to induce partial redistribution of the Na(+)/K(+)-ATPase, which participates in FGF-2 release, from the plasma membrane to the intracellular LMP1/FGF-2 positive dots. Treatment with ouabain, which inhibits Na(+)/K(+)-ATPase activity, partially suppressed FGF-2 secretion via exosomes in a dose-dependent manner. The results suggest that exosomes may represent a previously unrecognized mechanism for FGF-2 release mediated by LMP1, and that this pathway involves the activity of Na(+)/K(+)-ATPase.     

Fatty acid binding protein 5 promotes metastatic potential of triple negative breast cancer cells through enhancing epidermal growth factor receptor stability

Fatty acid binding protein 5 (FABP5), an intracellular lipid binding protein, has been shown to play a role in various cancers, including breast cancer. However, FABP5 and its role in triple negative breast cancer (TNBC) have not been studied. We show FABP5 protein expression correlates with TNBC, high grade tumors, and worse disease-free survival in a tissue microarray containing 423 breast cancer patient samples. High FABP5 expression significantly correlates with epidermal growth factor receptor (EGFR) expression in these samples. Decreased tumor growth and lung metastasis were observed in FABP5^−/− mice othotopically injected with murine breast cancer cells. FABP5 loss in TNBC tumor cells inhibited motility and invasion. Mechanistic studies revealed that FABP5 knockdown in TNBC cells results in decreased EGFR expression and FABP5 is important for EGF-induced metastatic signaling. Loss of FABP5 leads to proteasomal targeting of EGFR. Our studies show that FABP5 has a role in both host and tumor cell during breast cancer progression. These findings suggest that FABP5 mediates its enhanced effect on TNBC metastasis, in part, through EGFR, by inhibiting EGFR proteasomal degradation. These studies show, for the first time, a correlation between FABP5 and EGFR in enhancing TNBC metastasis through a novel mechanism.     

Role of fibroblast growth factor receptor 4 in cancer

Fibroblast growth factor receptors (FGFR) play a significant role in both embryonic development and in adults. Upon binding with ligands, FGFR signaling is activated and triggers various downstream signal cascades that are implicated in diverse biological processes. Aberrant regulations of FGFR signaling are detected in numerous cancers. Although FGFR4 was discovered later than other FGFR, information on the involvement of FGFR4 in cancers has significantly increased in recent years. In this review, the recent findings in FGFR4 structure, signaling transduction, physiological function, aberrant regulations, and effects in cancers as well as its potential applications as an anticancer therapeutic target are summarized.     

成纤维细胞生长因子受体1和血管内皮生长因子在肺鳞癌中的表达及与预后的关系

目的 观察成纤维细胞生长因子受体1(FGFR1)和血管内皮生长因子(VEGF)在肺鳞癌中的表达,并分析其与预后的相关性.方法 收集肺鳞癌组织标本135例和癌旁组织标本125例.采用免疫组织化学染色法检测不同肺组织中FGFR1和VEGF的表达水平,分析两者表达与临床病理参数的关系.肺鳞癌患者预后的影响因素采用Cox多因素分析.结果 肺鳞癌组织标本中FGFR1和VEGF的阳性表达率和表达水平均高于癌旁组织(P﹤0.05);在肺鳞癌组织中,FGFR1阳性表达与肿瘤分化程度、淋巴结转移、远处器官转移及TNM分期有关(P﹤0.05),VEGF阳性表达与肿瘤分化程度、淋巴结转移及TNM分期有关(P﹤0.05);Cox多因素分析结果显示,肿瘤分化程度、淋巴结转移、TNM分期、FGFR1及VEGF均为肺鳞癌预后的独立因素(P﹤0.05).结论 肺鳞癌患者的FGFR1和VEGF表达水平升高,并在肿瘤分化、淋巴结转移、TNM分期及预后中发挥重要作用.     

非小细胞肺癌血管生成因子与耐药相关基因关系的研究

目的:探讨血管生成因子(VEGF、bFGF)和耐药相关基因(MDR1、MRP、LRP)在非小细胞肺癌(non-smallcelllungcancer,NSCLC)中的表达及相互关系。方法:应用免疫组化技术检测96例NSCLC组织中VEGF、bFGF、MDR1、MRP、LRP蛋白表达,其中36例应用RT-PCR技术检测上述基因mRNA表达。结果:VEGF、bFGF、MDR1、MRP、LRPmRNA表达率分别为69.5%(25/36)、52.8%(19/36)、33.3%(12/36)、52.8%(19/36)、50.0%(18/36);蛋白表达率分别为51.0%(49/96)、58.3%(56/96)、45.8%(44/96)、59.4%(57/96)、64.6%(62/96),各基因mRNA表达与蛋白表达基本一致。统计分析表明:VEGF表达与MDR1、LRP表达相关(P=0.025,P=0.022),与MRP表达无关(P=0.428);bFGF表达与MDR1、LRP表达相关(P=0.001,P=0.012),并与MRP+LRP共表达相关(P=0.001)。结论:在非小细胞肺癌中血管生成因子与耐药相关基因具有一定的相关性。     

EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR AND BASIC FIBROBLAST GROWTH FACTOR IN HUMAN NON-SMALL CELL LUNG CANCER AND THEIR CLINICAL SIGNIFICANCE

Solid tumors contain tumor cells and vascular systems[1]. The growth and metastasis of solid tumors beyond 1-2mm in diameter depends on neovascularization. So the study of neovascularization helps to explain the biologic behavior of tumor. Angiogenesis is a complicated process mediated by a variety of angiogenic factors, which include vascular endothelial growth factor(VEGF) and basic fibroblast growth factor(bFGF). VEGF is an endothelial cell-specific mitogen with a pivotal role and may a…     

KHDRBS3 promotes multi-drug resistance and anchorage-independent growth in colorectal cancer

5-Fluorouracil (5-FU) is one of the most frequently used pharmacological agents in the treatment of colorectal cancer (CRC). Resistance to chemotherapy is a major cause of treatment failure of CRC, and it is a well known fact that cancer stem cells play a significant role in the acquisition of drug resistance. In this study, we focused on the KHDRBS3 gene that encodes KH RNA Binding Domain Containing, Signal Transduction Associated 3. We first clarified the relationship between KHDRBS3 and 5-FU resistance. We then observed higher expression levels of KHDRBS3 in KRAS-mutant organoids and cell lines in comparison with KRAS wild-type organoids and cell lines. Immunohistochemical analysis using CRC cases revealed that the prognosis of KHDRBS3-positive patients was significantly worse compared with that of KHDRBS3-negative patients. Univariate and multivariate Cox proportional hazards analyses showed that KHDRBS3 was an independent prognostic factor in patients with CRC. We determined that KHDRBS3 might play a crucial role in the acquisition of stem cell properties, such as drug resistance and spheroid/organoid formation, by regulating CD44 variant expression and the Wnt signaling pathway. In an immunodeficient mouse model, KHDRBS3-positive cells showed efficient tumor formation and formed metastatic lesions in the lungs. These results indicated that KHDRBS3 plays a crucial role in drug resistance and anchorage-independent growth by maintaining stem cell-like features in CRC cells. KHDRBS3 could be a promising candidate marker for predicting chemotherapeutic effect and prognosis in CRC patients.