表皮生长因子受体3在乳腺癌中的研究进展

表皮因子受体3(ErbB3/HER3)是表皮生长因子跨膜受体家族的成员之一。近年来证实ErbB3/HER3与乳腺癌的发病、复发转移、化疗以及内分泌治疗的疗效密切相关,已成为非常有前景的治疗候选靶点。本文简要综述近年来ErbB3/HER3在乳腺癌中的研究进展。     

Deltex-3-like (DTX3L) stimulates metastasis of melanoma through FAK/PI3K/AKT but not MEK/ERK pathway

Deltex-3-like (DTX3L), an E3 ligase, is a member of the Deltex (DTX) family and is also called B-lymphoma and BAL-associated protein (BBAP). Previously, we established RFP/RET-transgenic mice, in which systemic hyperpigmented skin, benign melanocytic tumor(s) and melanoma(s) develop stepwise. Here we showed that levels of Dtx3l/DTX3L in spontaneous melanoma in RFP/RET-transgenic mice and human melanoma cell lines were significantly higher than those in benign melanocytic cells and primarily cultured normal human epithelial melanocytes, respectively. Immunohistochemical analysis of human tissues showed that more than 80% of the melanomas highly expressed DTX3L. Activity of FAK/PI3K/AKT signaling, but not that of MEK/ERK signaling, was decreased in Dtx3l/DTX3L-depleted murine and human melanoma cells. In summary, we demonstrated not only increased DTX3L level in melanoma cells but also DTX3L-mediated regulation of invasion and metastasis in melanoma through FAK/PI3K/AKT but not MEK/ERK signaling. Our analysis in human BRAF^V600E inhibitor-resistant melanoma cells showed about 80% decreased invasion in the DTX3L-depleted cells compared to that in the DTX3L-intact cells. Thus, DTX3L is clinically a potential therapeutic target as well as a potential biomarker for melanoma.     

4‘,5,7—三羟基异黄酮对Balb／c 3T3小鼠成纤维细胞恶性转化的抑制作用

目的４’,５,７－三羟基异黄酮与多种肿瘤的低发生风险呈正相关,此文旨在探讨这一化合物的癌化学预防作用。方法 用３－ＭＣＡ和ＴＰＡ诱导Ｂａｌｂ／ｃ３Ｔ３细胞,建立体外二阶段转化模型,用这一模型来观察ｇｅｎｉｓｔｅｉｎ对Ｂａｌｂ／ｃ３Ｔ３－Ａ３１小鼠成纤维细胞恶性转化的影响。结果 在对细胞没有明显细胞毒作用的浓度下,ｇｅｎｉｓｔｅｉｎ明显抑制细胞的恶性转化,机理研究表明,ｇｅｎｉｓｔｅｉｎ抑制ＴＰＡ刺     

Complement C3a activates osteoclasts by regulating the PI3K/PDK1/SGK3 pathway in patients with multiple myeloma

Objective:Myeloma bone disease (MBD) is the most common complication of multiple myeloma (MM). Our previous study showed that the serum levels of C3/C4 in MM patients were significantly positively correlated with the severity of bone disease. However, the mechanism of C3a/C4a in osteoclasts MM patients remains unclear.Methods:The formation and function of osteoclasts were analyzed after adding C3a/C4a in vitro. RNA-seq analysis was used to screen the potential pathways affecting osteoclasts, and the results were verified by Western blot, qRT-PCR, and pathway inhibitors.Results:The osteoclast area per view induced by 1 μg/mL (mean ± SD: 50.828 ± 12.984%) and 10 μg/mL (53.663 ± 12.685%) of C3a was significantly increased compared to the control group (0 μg/mL) (34.635 ± 8.916%) (P < 0.001 and P < 0.001, respectively). The relative mRNA expressions of genes, OSCAR/TRAP/RANKL/cathepsin K, induced by 1 μg/mL (median: 5.041, 3.726, 1.638, and 4.752, respectively) and 10 μg/mL (median: 5.140, 3.702, 2.250, and 5.172, respectively) of C3a was significantly increased compared to the control group (median: 3.137, 2.004, 0.573, and 2.257, respectively) (1 μg/mL P = 0.001, P = 0.003, P < 0.001, and P = 0.008, respectively; 10 μg/mL: P < 0.001, P = 0.019, P < 0.001, and P = 0.002, respectively). The absorption areas of the osteoclast resorption pits per view induced by 1 μg/mL (mean ± SD: 51.464 ± 11.983%) and 10 μg/mL (50.219 ± 12.067%) of C3a was also significantly increased (33.845 ± 8.331%) (P < 0.001 and P < 0.001, respectively) compared to the control. There was no difference between the C4a and control groups. RNA-seq analysis showed that C3a promoted the proliferation of osteoclasts using the phosphoinositide 3-kinase (PI3K) signaling pathway. The relative expressions of PIK3CA/phosphoinositide dependent kinase-1 (PDK1)/serum and glucocorticoid inducible protein kinases (SGK3) genes and PI3K/PDK1/p-SGK3 protein in the C3a group were significantly higher than in the control group. The activation role of C3a in osteoclasts of MM patients was reduced by the SGK inhibitor (EMD638683).Conclusions:C3a activated osteoclasts by regulating the PI3K/PDK1/SGK3 pathways in MM patients, which was reduced using a SGK inhibitor. Overall, our results identified potential therapeutic targets and strategies for MBD patients.     

洛铂诱导顺铂耐药卵巢癌SKOV3/ DDP细胞的凋亡

 目的 探讨洛铂体外诱导人卵巢癌顺铂耐药SKOV3／DDP细胞凋亡及其机制。方法 采用四甲基偶氮唑盐（MTT）法测定不同浓度和时间洛铂对SKOV3／DDP细胞的生长抑制作用，并与其亲本细胞SKOV3（敏感株）相比较；进一步通过光镜及透射电镜观察SKOV3／DDP细胞的形态学改变；并采用流式细胞仪检测细胞周期变化。结果 洛铂作用SK0V3／DDP细胞的增殖呈时间剂量依赖模式受到抑制，与SKOV3细胞相比，差异无统计学意义（P〉0．05）；一定浓度的SKOV3／DDP作用一定时间，可诱导SKOV3／DDP细胞发生凋亡；光镜和透射电镜下可见典型的凋亡细胞形态学改变；流式细胞技术分析发现洛铂作用首先引起细胞周期分布的改变，随用药时间的延长，凋亡率逐渐升高。结论 洛铂通过诱导凋亡而抑制体外培养的SKOV3／DDP细胞生长，其机制可能与细胞G0／G1期阻滞有关。     

In vitro Transformation of BALB/c 3T3 Cells by 1,1,2,2-Tetrachloroethane

1,1,2,2-Tetrachloroethane (1,1,2,2-TTCE) was shown to be capable of inducing in vitro transformation of BALB/c 3T3 cells (clone A-31) either in the presence or in the absence of S9 activating system using an amplification-transformation (level-II) assay by reseeding confluent cells from each treatment and allowing additional rounds of cell replication. In the absence of metabolic activation, the highest assayed dose (1000 μg/ml), exerting the highest toxicity, was the only transforming dose. Lower doses of 1,1,2,2-TTCE were capable of transforming BALB/c cells in the presence of S9 activating system, the dose of 500 μg/ml exerting the highest transforming activity. The number and size of transformed foci recognized in the level-II plates were a function of the number of cells reseeded in the amplification assay. Foci obtained in the presence of S9 activating systems were larger in size, more deeply basophilic, and exhibited denser multilayering of constituent cells than foci recognized in the absence of exogenous metabolic activation.     

PI3K/Akt/mTOR信号传导通路与恶性肿瘤浸润和转移的研究进展

PI3K/Akt/mTOR信号通路作为细胞内重要信号传导通路之一,通过影响下游多种效应分子的活化状态,在细胞内发挥着抑制凋亡、促进增殖的关键作用,它与人类多种肿瘤的发生发展密切相关.本文综述了PI3K/Akt/mTOR信号通路的组成与功能、调节以及其抗肿瘤细胞凋亡作用机理等方面的研究进展,并就其抗细胞凋亡作用在肿瘤治疗中的应用作了评述,期待为以PI3K/Akt/mTOR信号通路中关键分子为靶点的肿瘤治疗研究提供参考.     

RhoE/Rnd3蛋白在乳腺癌组织中的表达及意义

目的:探讨RhoE/Rnd3蛋白在乳腺癌组织中的表达情况及意义.方法:采用免疫组化方法检测60例乳腺浸润性导管癌、30例正常乳腺组织中RhoE/Rnd3蛋白的表达情况.结果:RhoE/Rnd3蛋白在正常乳腺组织和乳腺癌中的表达率分别为73.3%和40.0%,二者的表达差异具有显著性意义(P＜0.05)RhoE/Rnd3蛋白在乳腺癌中表达与恶性程度呈负相关(P＜0.05).RhoE/Rnd3的表达与与患者年龄和淋巴结转移呈负相关.与生育史呈正相关.结论:RhoE/Rnd3在乳腺癌的发生中呈负调控作用,RhoE/Rnd3可能通过抑制Cyclin B1来抑制乳腺癌的发生;RhoE/Rnd3过表达与乳腺癌预后较好有关.     

Multipoint targeting of the PI3K/mTOR pathway in mesothelioma

Background:

Mesothelioma is a notoriously chemotherapy-resistant neoplasm, as is evident in the dismal overall survival for patients with those of asbestos-associated disease. We previously demonstrated co-activation of multiple receptor tyrosine kinases (RTKs), including epidermal growth factor receptor (EGFR), MET, and AXL in mesothelioma cell lines, suggesting that these kinases could serve as novel therapeutic targets. Although clinical trials have not shown activity for EGFR inhibitors in mesothelioma, concurrent inhibition of various activated RTKs has pro-apoptotic and anti-proliferative effects in mesothelioma cell lines. Thus, we hypothesised that a coordinated network of multi-RTK activation contributes to mesothelioma tumorigenesis.

Methods:

Activation of PI3K/AKT/mTOR, Raf/MAPK, and co-activation of RTKs were evaluated in mesotheliomas. Effects of RTK and downstream inhibitors/shRNAs were assessed by measuring mesothelioma cell viability/growth, apoptosis, activation of signalling intermediates, expression of cell-cycle checkpoints, and cell-cycle alterations.

Results:

We demonstrate activation of the PI3K/AKT/p70S6K and RAF/MEK/MAPK pathways in mesothelioma, but not in non-neoplastic mesothelial cells. The AKT activation, but not MAPK activation, was dependent on coordinated activation of RTKs EGFR, MET, and AXL. In addition, PI3K/AKT/mTOR pathway inhibition recapitulated the anti-proliferative effects of concurrent inhibition of EGFR, MET, and AXL. Dual targeting of PI3K/mTOR by BEZ235 or a combination of RAD001 and AKT knockdown had a greater effect on mesothelioma proliferation and viability than inhibition of individual activated RTKs or downstream signalling intermediates. Inhibition of PI3K/AKT was also associated with MDM2-p53 cell-cycle regulation.

Conclusions:

These findings show that PI3K/AKT/mTOR is a crucial survival pathway downstream of multiple activated RTKs in mesothelioma, underscoring that PI3K/mTOR is a compelling target for therapeutic intervention.     

Inhibition of PI3K/Akt/mTOR signaling pathway enhances the sensitivity of the SKOV3/DDP ovarian cancer cell line to cisplatin in vitro

The activation of the PI3K/AKT/m TOR pathway plays a key role in ovarian cancer tumorigenesis, progression and chemotherapy resistance. This study aimed to explore the possible mechanism that PI-103, a dual inhibitor of phosphatidylinositide 3-kinase and m TOR, enhances the sensitivity of SKOV3/DDP ovarian cancer cell line to cisplatin chemotherapy. The results showed that PI-103 could significantly increase the sensitivity of SKVO3/DDP cells to cisplatin through inhibiting the activation of PI3K/Akt/m TOR signaling pathway and inducing cell cycle arrest and apoptosis.     

Induction of metastatic potential by TrkB via activation of IL6/JAK2/STAT3 and PI3K/AKT signaling in breast cancer

In metastatic breast cancers, the acquisition of metastatic ability, which leads to clinically incurable disease and poor survival, has been associated with acquisition of epithelial-mesenchymal transition (EMT) program and self-renewing trait (CSCs) via activation of PI3K/AKT and IL6/JAK2/STAT3 signaling pathways. We found that TrkB is a key regulator of PI3K/AKT and JAK/STAT signal pathway-mediated tumor metastasis and EMT program. Here, we demonstrated that TrkB activates AKT by directly binding to c-Src, leading to increased proliferation. Also, TrkB increases Twist-1 and Twist-2 expression through activation of JAK2/STAT3 by inducing c-Src-JAK2 complex formation. Furthermore, TrkB in the absence of c-Src binds directly to JAK2 and inhibits SOCS3-mediated JAK2 degradation, resulting in increased total JAK2 and STAT3 levels, which subsequently leads to JAK2/STAT3 activation and Twist-1 upregulation. Additionally, activation of the JAK2/STAT3 pathway via induction of IL-6 secretion by TrkB enables induction of activation of the EMT program via induction of STAT3 nuclear translocation. These observations suggest that TrkB is a promising target for future intervention strategies to prevent tumor metastasis, EMT program and self-renewing trait in breast cancer.     

JAK/STAT3和MAPK/ERK在乳腺癌细胞侵袭转移中的交互作用及意义

目的研究应用JAK酶抑制剂AG490对乳腺癌细胞MDA-MB-231 STAT3和ERK磷酸化的影响,初步探讨JAK/STAT3和MAPK/ERK两条信号转导通路的交互作用以及在乳腺癌细胞侵袭转移中的调控意义。方法以JAK酶抑制剂AG490处理乳腺癌细胞MDA-MB-231,Western blot检测细胞中P-STAT3、P-ERK蛋白水平变化;RT-PCR检测细胞中STAT3、ERK1、ERK2mRNA的变化;明胶酶谱法检测细胞分泌MMP-2、MMP-9的变化,Transwell小室进行人工重组基底膜侵袭和运动实验。结果应用JAK酶抑制剂AG490后人乳腺癌细胞MDA-MB-231中P-STAT3、P-ERK蛋白均减少,STAT3、ERK1、ERK2mRNA表达下降,同时可使细胞分泌MMP-2、MMP-9减少,使细胞侵袭、迁移能力降低。结论JAK/STAT3和MAPK/ERK两条信号转导通路之间存在交互作用,通过JAK酶抑制可改变转录因子STAT3和激酶ERK磷酸化水平,进而可以交互影响其基因转录的表达。JAK酶抑制对两条信号转导通路的激活有阻断作用而可以抑制乳腺癌细胞的侵袭转移。     

Gastrin acting on the cholecystokinin2 receptor induces cyclooxygenase-2 expression through JAK2/STAT3/PI3K/Akt pathway in human gastric cancer cells

Gastrin, cholecystokinin2 receptor (CCK2R), and cyclooxygenase-2 (COX-2) have been implicated in the carcinogenesis and progression of gastric cancer. Our study demonstrated that antagonist or siRNA against CCK2R blocked amidated gastrin (G17)-induced activation of STAT3 and Akt in gastric cancer cell lines. G17-increased COX-2 expression and cell proliferation were effectively blocked by CCK2R antagonist and inhibitors of JAK2 and PI3K. In addition, knockdown of STAT3 expression significantly attenuated G17-induced PI3K/Akt activation, COX-2 expression, and cell proliferation. These results suggest that CCK2R-mediated COX-2 up-regulation via JAK2/STAT3/PI3K/Akt pathway is involved in the proliferative effect of G17 on human gastric cancer cells.     

C-erbB-3/HER3在肺腺癌中的表达及其与预后的相关性

目的 :探讨C erbB 3 HER 3在肺腺癌中的表达及其与临床病理特征、生存期的相关性。方法 :2 3例手术切除的肺腺癌组织 ,15例相应的交界区组织 ,10例肿瘤远端正常肺组织标本 ,采用免疫组织化学SP法进行半定量检测C erbB 3 HER3的表达。结果 :C erbB 3 HER3在常肺组织中呈弱表达 ,交界区组织与腺癌组织相比 ,临床Ⅰ～ⅢA与ⅢB～Ⅳ相比 ,病理分化Ⅰ级与Ⅱ、Ⅲ级相比 ,其过表达率均呈增高趋势 ,过表达组生存期相对短于弱表达组。但统计学处理均未达到显著性水准。结论 :C erbB 3 HER3可能参与了肺正常组织细胞的生长发育 ,以及肺腺癌的发生发展 ,该基因受体蛋白对估计肺腺癌预后可能有意义 ,但仍需积累更多的资料研究 ,以明确其生物学作用。     

抑制PI3K/Akt通路提高HeLa细胞化疗效果的实验研究

目的探讨抑制PI3K/Akt信号转导通路提高抗癌药物对人宫颈癌HeLa细胞的杀伤作用。方法采用MTT法检测Celecoxib、DDP及Docetaxel单独或联合PI3K抑制剂LY294002对人宫颈癌HeLa细胞的抑制率;采用流式细胞技术检测药物单独或联合作用对HeLa细胞凋亡的影响。结果(1)联合LY294002能够显著提高Celecoxib、DDP及Docetaxel对HeLa细胞抑制率;(2)LY294002与Celecoxib、DDP及Docetaxel的协同治疗指数均小于1,二者起协同治疗作用;(3)联合LY294002能够增加HeLa细胞的凋亡水平。结论抑制PI3K/Akt信号转导通路能够显著提高Celecoxib、DDP及Docetaxel对HeLa细胞杀伤作用。     

Isolation and Characterization of ras-Transfected BALB/3T3 Clone Showing Morphological Transformation by 12-O-Tetradecanoyl-phorbol-13-acetate

The transformation frequency of mouse BALB/3T3 cells was significantly enhanced after transfection with an activated ras oncogene (v-Ha- ras ) followed by treatment with a tumor promoter, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), suggesting that the ras oncogene acted as an initiator in two-stage carcinogenesis. A cell clone (Bhas42) containing the ras oncogene was isolated from the ras -transfected BALB/3T3 cells. Bhas42 cells were flat and showed contact inhibition, but the addition of TPA to quiescent Bhas42 cultures resulted in a dramatic change of cell morphology to spindle shape, doubling of the cell population, and increased DNA synthesis.     

PI3K／Akt／inTOR信号传导通路与恶性肿瘤浸润和转移的研究进展

PI3K／Akt／mTOR信号通路作为细胞内重要信号传导通路之一,通过影响下游多种效应分子的活化状态,在细胞内发挥着抑制凋亡、促进增殖的关键作用,它与人类多种肿瘤的发生发展密切相关。本文综述了PI3K／Akt／mTOR信号通路的组成与功能、调节以及其抗肿瘤细胞凋亡作用机理等方面的研究进展,并就其抗细胞凋亡作用在肿瘤治疗中的应用作了评述,期待为以PI3K／Akt／mTOR信号通路中关键分子为靶点的肿瘤治疗研究提供参考。     

Compensatory ErbB3/c-Src signaling enhances carcinoma cell survival to ionizing radiation

Summary EGFR and ErbB2 are two members of the ErbB family of receptor Tyr Kinases identified as therapeutic targets for treating carcinomas. Breast carcinoma cells express different complements and variable proportions of ErbB receptor Tyr kinases, which activate unique and redundant signaling cascades that are essential for cell survival. Previously it was shown that a COOH-terminal truncation mutant of the EGFR (EGFR-CD533) blocks EGFR dependent signals and radiosensitizes breast carcinoma cells. In this study the effects of EGFR-CD533 and an analogous truncation mutant of ErbB2 (ErbB2-CD572) on ErbB receptor family dimerization and signaling are further investigated. Using adenoviral vectors in breast carcinoma cell lines with variable ErbB expression profiles, we demonstrate different effects for each deletion mutant. EGFR-CD533 blocks ligand stimulation of EGFR, ErbB2, and ErbB4, but is associated with a compensatory Tyr kinase activity resulting in phosphorylation of ErbB3. In contrast, ErbB2-CD572 produces a weaker, non-specific pattern of ErbB receptor family inhibition, based upon the ErbB expression pattern of the cell type. Investigation of the compensatory Tyr kinase activity associated with EGFR-CD533 expression identified an ErbB3/c-Src signaling pathway that regulates expression of anti-apoptotic Bcl family proteins. This signaling is active in the T47D cell line, which inherently over-express ErbB3, absent in MDA-MB231 cells, which have low ErbB3 expression levels, and is restored in a MDA-MB231 cell line engineered to over-express ErbB3. Furthermore we demonstrate that ErbB3/c-Src signaling is radio-protective, and that its elimination through pharmacologic inhibition of c-Src enhances radiation-induced apoptosis. In summary, these studies identify a novel ErbB3/c-Src survival signal and point to ErbB3 expression levels as an important variable in therapeutic targeting of ErbB receptors in breast carcinoma cells. ^†(Rupert K. Schmidt-Ullrich) Deceased December 20, 2004 Address for offprints and correspondence: Joseph N. Contessa, Department of Radiation Oncology, University of Michigan, 1500 East Center Drive, UH B2 C490, Box 0010, Ann Arbor, MI 48109; E-mail: jcontess@med.umich.edu