免疫组化标记物在子宫内膜样腺癌与宫颈腺癌鉴别诊断中的表达及意义

目的探讨免疫组化标记物波形蛋白(Vimentin)、癌胚抗原(CEA)、雌激素受体(ER)、孕激素受体(PR)和p16蛋白在子宫内膜样腺癌与宫颈腺癌鉴别诊断中的表达和意义。方法采用免疫组织化学染色法检测48例子宫内膜样腺癌和21例宫颈腺癌患者肿瘤组织中Vimentin、CEA、ER、PR和p16的表达,并分析其临床意义。结果子宫内膜样腺癌患者肿瘤组织中Vimentin、CEA、ER、PR和p16阳性表达率分别为79.2%、31.3%、87.5%、81.3%和20.8%。宫颈腺癌患者肿瘤组织中Vimentin、CEA、ER、PR和p16阳性表达率分别为14.3%、90.5%、23.8%、42.9%和81.0%。经2检验,Vimentin(2=25.50)、CEA(2=20.50)、ER(2=27.29)、PR(2=10.17)和p16(2=22.17)在子宫内膜样腺癌和宫颈腺癌患者肿瘤组织中阳性表达率的差异均有统计学意义(P<0.05)。结论免疫组化法检测Vimentin、CEA、ER、PR和p16的表达有助于临床鉴别诊断宫颈腺癌和子宫内膜样腺癌。     

Well-differentiated endometrial adenocarcinomas and poorly differentiated mixed mullerian tumors have altered ER and PR isoform expression.

Both the estrogen receptor (ER) and the progesterone receptor (PR) have two subtypes: ER-alpha and beta, and PR-A and -B, respectively. These subtypes differ in function and expression, and recent reports have correlated changes in the normal proportions of these isoforms with neoplastic states. We investigated ER and PR isoform expression in normal pre- and post-menopausal endometrium, well-differentiated endometrial adenocarcinoma, and poorly differentiated malignant mixed mullerian tumors (MMMTs). Semi-quantitative RT-PCR and immunoblotting were used to measure receptor mRNA and protein expression. Estrogen receptor-alpha/beta mRNA ratios were significantly higher in postmenopausal (27.3) compared to premenopausal endometrium (4.9) mainly as a result of lower ER-beta expression in the former. Compared to age-matched postmenopausal controls, the ER-alpha/beta ratio was reduced in both grade I adenocarcinoma and MMMT specimens (3.3 and 6.8, respectively), due to a selective loss of ER-alpha. The relative abundance of PR-A and PR-B mRNA remained unchanged between all tissue subtypes. Total PR protein, however, was significantly reduced in MMMTs compared to all other groups. Thus, sex steroid receptor expression is significantly and differentially altered in well-differentiated and poorly-differentiated endometrial cancers. Both cancers exhibit decreased ER-alpha expression and the MMMTs also demonstrate a significant loss of PR protein.     

HER-2/neu overexpression and hormone dependency in endometrial cancer: analysis of cohort and review of literature

BACKGROUND: Overexpression of HER-2/neu (HER) is associated with unfavorable prognoses in both endometrial and breast cancer. MATERIALS AND METHODS: To determine whether an association exists between HER expression and markers of hormone dependency in endometrial cancers, we subjected hysterectomy specimens from 125 patients to immunohistochemical staining for HER. HER was visually interpreted as negative/weakly positive (HER-) versus strongly positive (HER+). Estrogen receptor (ER) and progesterone receptor (PR) levels were quantitated on fresh tissue using a dextran-coated charcoal assay. RESULTS: HER+ was observed in 12% of endometrioid tumors and 22% of nonendometrioid tumors (p = 0.07). Mean ER and PR levels were 255 fmol/mg and 457 fmol/mg in endometrioid tumors, compared with 74 and 104 in nonendometrioid tumors (p < 0.01). Hyperplasia associated with the tumor was related to high levels of both ER and PR (p < 0.05), but not with HER expression. Age was significantly related to high levels of ER (p = 0.007). Both recurrence and death rates were significantly associated with low levels of ER and PR (p < 0.01). Mean ER and PR levels were 270 and 466 fmol/mg, respectively, in HER-tumors, compared with 95 (p = 0.14) and 138 fmol/mg (p = 0.02) in HER+ tumors. CONCLUSION: HER overexpression may be an important step in hormone-independent growth and proliferation in a subgroup of endometrial cancers.     

子宫内膜癌组织雌、孕激素受体测定及其临床意义

目的:探讨子宫内膜癌组织雌激素受体(ER)和孕激素受体(PR)与临床病理特征的关系。方法:采用葡聚糖-活性炭吸附法(DCC法)对68例子宫内膜癌组织进行ER、PR测定,同时采用免疫组化法对其中30例进行ER、PR检测。结果:DCC法检测ER、PR的阳性率分别为79·4%和77·9%,免疫组化法均为86·7%。两种测定方法比较,ER与PR的总符合率分别为82·4%和85·7%。免疫组化法可在DCC法基础上进一步明确组织来源。ER、PR水平与组织学分级呈负相关。组织类型中腺癌(包括乳头状腺癌)与腺棘癌的ER、PR水平高于其他癌。ER水平与肥胖呈正相关。结论:ER、PR水平与组织学分级、组织学类型均反映了子宫内膜癌的生物学行为,ER、PR的测定对估计预后和临床选择激素治疗具有重要意义。     

Activation of downstream epidermal growth factor receptor (EGFR) signaling provides gefitinib-resistance in cells carrying EGFR mutation

Patients with pulmonary adenocarcinoma carrying the epidermal growth factor receptor (EGFR) mutation tend to display dramatic clinical response to treatment with the EGFR tyrosine kinase inhibitor gefitinib. Unfortunately, in many cases the cancer cells eventually acquire resistance, and this limits the duration of efficacy. To gain insight into these acquired resistance mechanisms, we first prepared HEK293T cell line stably transfected with either wild-type (WT) or mutant (L858R) EGFR, and then expressed oncogenic K-Ras12V mutant in the latter transfectant. Although 293T cells expressing wild-type EGFR did not show any growth inhibition by gefitinib treatment similarly to the non-transfected cells, the cells expressing the EGFR-L858R were exquisitely sensitive. Consistently, phospho-Akt levels were decreased in response to gefitinib in cells expressing EGFR-L858R but not in cells with EGFR-WT. In contrast, 293T cells expressing both EGFR-L858R and oncogenic K-Ras were able to proliferate even in the presence of high concentration of gefitinib probably by inducing Erk1/2 activation. We also expressed K-Ras12V in the gefitinib-sensitive pulmonary adenocarcinoma cell line PC-9, which harbors an in-frame deletion in the EGFR gene. The activated K-Ras inhibited the effects of gefitinib treatment on cell growth, cell death induction and levels of phospho-Akt, as well as phospho-Erk. These data indicate that activated Ras could substitute most of the upstream EGFR signal, and are consistent with the hypothesis that mutational activation of targets immediately downstream from the EGFR could induce the secondary resistance to gefitinib in patients with lung cancer carrying EGFR mutation.     

宫腔镜结合ER、PR表达对早期诊断子宫内膜样腺癌的临床价值

目的探讨宫腔镜结合子宫内膜组织雌激素受体(ER)、孕激素受体(PR)表达对早期诊断子宫内膜样腺癌的临床价值。方法选择2007年1月至2012年1月于我院妇科行宫腔镜检查及诊刮术的840例患者,应用免疫组化EnVision法检测各类子宫内膜组织中ER、PR的表达情况。结果在本组840例患者中,经宫腔镜检查诊断子宫内膜样腺癌25例,经组织病理诊断子宫内膜样腺癌26例。宫腔镜诊断的敏感度为96.2%,特异度为100.0%,阳性预测值为100.0%,阴性预测值为99.9%。在本组的10例正常子宫内膜、20例子宫内膜不典型增生和26例子宫内膜样腺癌的子宫内膜组织中,ER、PR阳性表达率逐渐降低,有显著性差异(P0.01)。随着病理分级及FIGO分期增加,ER、PR阳性表达降低,且降低趋势具有统计学意义(P0.01),但在肌层浸润深度和有无淋巴结转移方面,ER、PR的表达组间无显著性差异(P0.05)。结论宫腔镜检查结合子宫内膜组织ER、PR的表达检测对子宫内膜样腺癌早期诊断有重要的意义。     

STAT3与雌、孕激素受体在子宫内膜癌中表达的相关性

目的探讨孕激素受体（PR）、雌激素受体（ER）与信号转导及转录活化因子3（STAT3）在子宫内膜癌患者中的表达情况及其相关性。方法选取30例经病理组织学证实为子宫内膜癌的患者为研究对象,并选取30例健康体检妇女为对照组,采用免疫组化法测定2组子宫内膜标本中ER、PR、STAT3表达情况,并分析三者相关性。结果子宫内膜癌组织中ER、PR、STAT3阳性表达率显著高于对照组,差异有统计学意义（P〈0．05）。子宫内膜癌中STAT3、ER表达均与肿瘤临床分期、病理分级、淋巴结转移有关（均P〈0．05）;而PR则与临床分期及病理分级有关（均P〈0．05）。经相关性分析可知,STAT3与ER激素呈正相关（γ=0．452,P=0．000）,与PR激素无关（γ=0．1378,P=0．071）。结论子宫内膜癌的发生与ER、PR、STAT3过度表达有关,而STAT3的表达与雌激素调控有密切关系。     

子宫内膜癌雌、孕激素受体和C-erbB-2检测的临床意义

目的 检测子宫内膜癌组织中雌激素受体(ER)、孕激素受体(PR)及癌基因蛋白C-erbB-2表达的阳性率并探讨其与预后的关系.方法 用免疫组织化学法对32份子宫内膜癌标本进行了ER、PR及C-erbB-2的检测.结果 子宫内膜癌组织中ER、PR、C-erbB-2的阳性率分别为53.1%、50.0%、46.9%.ER、PR的阳性表达率与癌组织的细胞分化程度有关,随着子宫内膜癌组织学分级的增高,ER、PR阳性表达率逐渐降低,C-erbB-2的阳性表达率与肿瘤病理分级呈正相关,与ER、PR表达呈负相关.结论 ER、PR、C-erbB-2均反映了子宫内膜癌的生物学行为,其测定对预测预后、指导选择内分泌治疗具有重要意义.     

K-Ras and H-Ras activation promote distinct consequences on endometrial cell survival

A considerable amount of evidence indicates that Ras signaling contributes to the development of endometrial cancer. We previously demonstrated that endometrial cancer cells carrying oncogenic [(12)Val]K-ras were susceptible to apoptosis. The present study examined the role of K-and H-Ras in the induction of apoptosis using rat endometrial cells (RENT4 cells). We found that constitutively activated K-Ras promoted apoptotic cell death, whereas the H-Ras mutant rescued rat endometrial cells from apoptosis. Expression of a constitutively active form of Raf-1 (Raf-CAAX) promoted apoptosis, whereas expression of a constitutively active catalytic subunit of phosphoinositide 3-kinase, p110K227E, allowed cells to escape from apoptosis. Moreover, inhibition of the MEK-MAPK pathway by the specific inhibitor, UO126, rescued the cells from apoptosis, whereas the inhibition of phosphoinositide 3-kinase by its specific inhibitor, LY294002, promoted apoptosis in RENT4 cells expressing activated K-Ras. However, both inhibitors promoted apoptosis in RENT4 cells expressing activated H-Ras. This difference in the regulation of apoptosis by the MEK inhibitor between K-Ras- and H-Ras-expressing cells depended on the interaction of effector proteins downstream of each Ras isoform. Finally, to elucidate the role of downstream K-Ras signal pathways, we generated K-Ras effector domain mutants (K12V35S, K12V40C). We examined the incidence of apoptotic cell death induced by the K-Ras effector domain mutants (K12V35S, K12V40C). The relative ratio of phospho-MAPK to phospho-Akt compared with that of mock cells was higher in K12V35S cells than in K12V40C cells. Ectopic expression of K12V35S protein increased the proportion of apoptotic cells, and in turn, the expression of K12V40C protein decreased compared with the expression of K12V protein without the effector domain mutant. These results demonstrate that K- and H-Ras-mediated signaling pathways exert distinct effects on apoptosis and that K-Ras downstream Raf/MEK/MAPK pathway is required for the induction of apoptosis in endometrial cells. Coordination of the two pathways contributes to endometrial cell survival.     

人类组织激肽释放酶10在子宫内膜腺癌组织中的表达及其与雌激素、孕激素受体表达的相关性

 目的　检测人类组织激肽释放酶10（KLK10）在不同子宫内膜组织中的表达,并探讨其与雌激素受体（ER）和孕激素受体（PR）之间的相关性。方法　应用免疫组织化学方法测定KLK10在12例正常子宫内膜、19例子宫内膜增生症、34例子宫内膜腺癌组织中的表达,并分析KLK10与ER、PR之间的相关性。结果　KLK10在正常子宫内膜组织、子宫内膜增生症组织、子宫内膜腺癌组织中阳性率分别为91.7 %（11／12）、78.9 %（15／19）、38.2 %（13／34）,两两比较差异均有统计学意义（均P＜0.05）。KLK10在Ⅰ、Ⅱ、Ⅲ期子宫内膜腺癌组织中表达阳性率分别为64.3 %（9／14）、30.0 %（3／10）、10.0 %（1／10）,两两比较差异均有统计学意义（均P＜0.05）。KLK10在高、中、低分化的内膜腺癌组织中表达阳性率分别为66.7 %（8／12）、33.3 %（4／12）、10.0 %（1／10）,两两比较差异均有统计学意义（均P＜0.05）。34例子宫内膜腺癌组织中,KLK10表达阳性率为38.2 %,ER、PR表达阳性率为67.6 %、55.9 %,KLK10与ER、PR表达均呈正相关（χ2＝0.448,P＜0.01）。结论　KLK10表达缺失或下调可能促进子宫内膜腺癌的发生和发展。KLK10在子宫内膜腺癌组织中表达降低,ER、PR的表达也降低,呈正相关。KLK10阳性者预后相对较好。     

Expression of estrogen receptor (ER) (beta)cx protein in ER(alpha)-positive breast cancer: specific correlation with progesterone receptor

Estrogen receptor (ER) (beta)cx, a splice variant of ERbeta, is a dominant repressor of ER(alpha) function. In this study we investigated the possibility that because the progesterone receptor (PR) gene is a downstream target of activated ER(alpha), in ER(alpha)-positive breast cancers, expression of ER(beta)cx would result in repression of PR. In ER(alpha)-positive MCF-7 cells, stable transfection of an ER(beta)cx expression vector resulted in reduced expression of PR without affecting ER(alpha) expression. In breast cancers, immunohistochemical evaluation of ER(alpha)-positive foci for the expression of PR and ER(beta)cx revealed a significant correlation between a PR-negative phenotype and the presence of ER(beta)cx within the foci. However, when entire lesions were evaluated by Allred scoring in 115 ER(alpha)-positive breast cancer specimens, the presence of two distinct groups of patients could be discerned. One group expressed ER(beta)cx and had very reduced levels of PR expression, as expected. The second group showed both ER(beta)cx and high levels of PR. To evaluate the role of ER(beta)cx in sensitivity to tamoxifen, 18 core needle biopsies, obtained before preoperative treatment with tamoxifen, were investigated. The results show that expression of ER(beta)cx in primary lesions correlated with a poor response to tamoxifen, especially in cancers with a low PR expression in Allred score. This is the first evidence that evaluation of ER(beta)cx along with PR may contribute to a better characterization of ER(alpha)-positive breast cancers.     

更年期子宫内膜增殖症与内膜腺癌ER、PR、PNA免疫组化研究

本文用免疫组化检测更年期子宫内膜增殖症与子宫内膜腺癌雌激素受体（ＥＲ）、孕激素受体（ＰＲ）、花生凝集素（ＰＮＡ）水平。结果显示：更年期子宫内膜增殖症和子宫内膜腺癌中均含有ＥＲ、ＰＲ，二者在受体表达强度方面无明显差异（Ｐ＞０．０５），提示在更年期子宫内膜增殖症和内膜腺癌存在相似的雌激素环境。但ＰＮＡ的表达在二者之间具有差异（Ｐ＜０．０５），从而揭示ＰＮＡ在子宫内膜腺癌中的表达特征及其所代表的细胞膜结构中精蛋白的变化对于子宫内膜腺体增生和癌变研究具有的重要价值。     

K-ras Asp12 mutant neither interacts with Raf, nor signals through Erk and is less tumorigenic than K-ras Val12

Different mutant amino acids in the Ras proteins lead to distinct transforming capacities and different aggressiveness in human tumors. K-Ras Asp12 (K12D) is more prevalent in benign than in malignant human colorectal tumors, whereas K-Ras Val12 (K12V) associates with more advanced and metastatic carcinomas, higher recurrence and decreased survival. Here, we tested, in a nude mouse xenograft model, whether different human K-Ras oncogenes mutated at codon 12 to Val, Asp or Cys would confer NIH3T3 fibroblasts distinct oncogenic phenotypes. We studied tumor histology and growth, apoptotic and mitotic rates, activation of signal transduction pathways downstream of Ras and regulation of the cell cycle and apoptotic proteins in tumors derived from the implanted transformants. We found that the K12V oncogene induces a more aggressive tumorigenic phenotype than the K12D oncogene, whereas K12C does not induce tumors in this model. Thus, K12V mutant tumors proliferate about seven times faster, and have higher cellularity and mitotic rates than the K12D mutant tumors. A molecular analysis of the induced tumors shows that the K12V mutant protein interacts with Raf-1 and transduces signals mainly through the Erk pathway. Unexpectedly, in tumors induced by the K12D oncogene, the K-Ras mutant protein does not interact with Raf-1 nor activates the Erk canonical pathway. Instead, it transduces signals through the PI3K/Akt, JNK, p38 and FAK pathways. Finally, the higher growth rate of the K12V tumors associates with enhanced Rb phosphorylation, and PCNA and cyclin B upregulation, consistent with faster G1/S and G2/M transitions, without alteration of apoptotic regulation.     

Expression of steroid receptor coactivators and corepressors in human endometrial hyperplasia and carcinoma with relevance to steroid receptors and Ki-67 expression

BACKGROUND: To examine the steroid hormone dependent growth mechanism of human endometrial hyperplasia and carcinoma, expression levels of steroid receptor cofactors, such as coactivators (steroid receptor coactivator 1 [SRC-1] and p300/cyclic AMP-response element-binding protein (p300/CBP]) and corepressors (nuclear receptor corepressor [NCoR] and silencing mediator for retinoid and thyroid-hormone receptors [SMRT]), were investigated. METHODS: The expression levels of cofactors were examined immunohistochemically using 20 samples of normal endometria, 36 samples of hyperplastic endometria, and 58 of malignant endometria and were compared with the expression levels of estrogen receptor (ER), progesterone receptor (PR), and a proliferation marker, Ki-67. RESULTS: In samples of normal endometria, the expression of coactivators was observed diffusely in glandular cells in the proliferative phase, with a mean positivity index (PI) of 81.8 for SRC-1 and 91.3 for p300/CBP, whereas expression levels decreased in endometrial hyperplasia (PI: SRC-1, 58.9; p300/CBP, 83.8) and endometrial carcinoma (PI: SRC-1, 45.0; p300/CBP, 55.4). In endometrial hyperplasia, there was a significant correlation between the expression of ER and SRC-1 or p300/CBP. In contrast, there were no significant statistical or topologic correlations between the expression of coactivators and the expression of ER/PR in endometrial carcinoma. The expression of corepressors generally was limited, except for elevated expression of NCoR in endometrial hyperplasia (PI, 23.8). CONCLUSIONS: The current study showed that expression levels of the steroid receptor coactivators SRC-1 and p300/CBP were reduced in endometrial carcinoma compared with normal and hyperplastic endometrium. In addition, topologic coexpression of both coactivators and ER/PR was lost in endometrial carcinoma. Accordingly, limited response to sex steroids in patients with endometrial carcinoma may be ascribed to the dissociation of cofactors and ER/PR.     

Chronic pancreatitis is essential for induction of pancreatic ductal adenocarcinoma by K-Ras oncogenes in adult mice

Pancreatic ductal adenocarcinoma (PDA), one of the deadliest human cancers, often involves somatic activation of K-Ras oncogenes. We report that selective expression of an endogenous K-Ras(G12V) oncogene in embryonic cells of acinar/centroacinar lineage results in pancreatic intraepithelial neoplasias (PanINs) and invasive PDA, suggesting that PDA originates by differentiation of acinar/centroacinar cells or their precursors into ductal-like cells. Surprisingly, adult mice become refractory to K-Ras(G12V)-induced PanINs and PDA. However, if these mice are challenged with a mild form of chronic pancreatitis, they develop the full spectrum of PanINs and invasive PDA. These observations suggest that, during adulthood, PDA stems from a combination of genetic (e.g., somatic K-Ras mutations) and nongenetic (e.g., tissue damage) events.     

Progesterone Stimulates Proliferation and Promotes Cytoplasmic Localization of the Cell Cycle Inhibitor p27 in Steroid Receptor Positive Breast Cancers

Progestins are reported to increase the risk of more aggressive estrogen receptor positive, progesterone receptor positive (ER+ PR+) breast cancers in postmenopausal women. Using an in vivo rat model of ER+ PR + mammary cancer, we show that tumors arising in the presence of estrogen and progesterone exhibit increased proliferation and decreased nuclear expression of the cell cycle inhibitor p27 compared with tumors growing in the presence of estrogen alone. In human T47D breast cancer cells, progestin increased proliferation and decreased nuclear p27 expression. The decrease of nuclear p27 protein was dependent on activation of Src and PI3K by progesterone receptor isoforms PRA or PRB. Importantly, increased proliferation and decreased nuclear p27 expression were observed in invasive breast carcinoma compared with carcinoma in situ. These results suggest that progesterone specifically regulates intracellular localization of p27 protein and proliferation. Therefore, progesterone-activated pathways can provide useful therapeutic targets for treatment of more aggressive ER+ PR+ breast cancers.     

Targeting Ras-PI3K/mTOR pathway and the predictive biomarkers in endometrial cancer

The Ras-PI3K (phosphatidylinositol-3-kinase)/mTOR (mammalian Target of Rapamycin) pathway is frequently activated in various types of cancers. A number of inhibitors targeting the PI3K/mTOR pathway and MAPK pathway (another Ras effector pathway) are under development. PI3K/AKT activating mutations, including mutations in PTEN (50%), PIK3CA (30%), and K-Ras (20%), are frequently observed in endometrial cancer. A coexistence of these mutations is also commonly observed. We classified 13 endometrial cancer cell lines into three groups according to their mutational status in these genes: Group A (n=9); K-Ras wild-type and PTEN mutant, Group B (n=2); K-Ras mutant, and Group C (n=2) without any mutations in K-Ras, PTEN or PIK3CA. We determined the effects a dual PI3K/mTOR inhibitor (Inhibitor P) on these cell lines. MTT assay revealed that all the nine cell lines in Group A were sensitive to the inhibitor P (IC50<100 nM), whereas the other four cell lines in Group B or C were less sensitive to it(IC50>100 nM). Daily oral administration of inhibitor P showed anti-tumor effects in the mice bearing Group A tumors. Our data suggest that dual inhibition of the PI3K/mTOR is a promising molecular-targeted therapeutic for certain endometrial cancers, and that the mutational status of K-Ras and PI3K pathway-related genes, like PTEN and PIK3CA, could be useful for predicting sensitivities to such agents.