卵巢上皮性癌中干扰素介导的跨膜蛋白1的表达意义

目的:探讨干扰素介导的跨膜蛋白1(Interferon-induced transmembrane protein 1,IFITM1)基因在卵巢上皮性癌中表达的相关性及其意义。方法:应用Western blotting检测正常卵巢、卵巢良性肿瘤、卵巢交界性肿瘤和卵巢上皮性癌组织中IFITM1蛋白表达。免疫组织化学检测12例正常卵巢、21例卵巢良性肿瘤、18例卵巢交界性肿瘤和85例卵巢上皮性癌组织中IFITM1的蛋白表达,同时分析IFITM1表达状况与临床病理因素之间的相关性。结果:Western blotting显示卵巢上皮性癌和卵巢交界性肿瘤中IFITM1表达水平明显高于正常卵巢组织和卵巢良性肿瘤。免疫组化显示在正常卵巢组织中IFITM1阳性表达率为41.7%(5/12),在卵巢良性肿瘤组织中71.4%(15/21),在卵巢交界性肿瘤组织中为72.2%(13/18),在卵巢上皮性癌中为77.6%(66/85),IFITM1蛋白表达强度在正常卵巢、良性卵巢肿瘤、交界性卵巢肿瘤、上皮性卵巢癌间的比较有统计学意义(P0.05)。IFITM1蛋白表达与病理类型、肿瘤分化程度、肿瘤FIGO分期有关(P0.05),与淋巴结转移、腹水无明显相关性。化疗敏感组和耐药组的IFITM1表达强度间差异有统计学意义(P0.05)。结论:IFITM1在正常卵巢、卵巢良性肿瘤、卵巢交界性肿瘤和卵巢上皮性癌组织中的表达依次升高,并与卵巢癌以铂类为基础的化疗耐药性产生有相关性,为进一步研究IFITM1在卵巢癌诊治及化疗中的应用前景提供依据。     

uPA、PAI-1 的表达和MVD 计数与浸润性乳腺癌侵袭性的关系

目的:探讨尿激酶型纤溶酶原激活剂(uPA)、纤溶酶原激活物抑制剂(PAI-1)的表达和血管形成与浸润性乳腺癌侵袭性的关系。方法:应用免疫组化SP法检测80例浸润性乳腺癌、20例良性乳腺肿瘤组织中uPA、PAI-1的表达情况并进行微血管计数。结果:uPA和PAI-1在浸润性乳腺癌组的阳性表达显著高于良性肿瘤组,且其高表达率与乳腺癌的临床病理参数密切相关(P<0.05);微血管计数在乳腺癌组和良性肿瘤组分别为30.87±7.64、20.28±8.72,两组比较有显著性差异(P<0.01)。uPA与PAI-1在浸润性乳腺癌中的表达呈正相关(P<0.05)。结论:uPA、PAI-1的高表达与浸润性乳腺癌的浸润、转移相关,uPA、PAI-1的高表达和MVD计数可作为评价浸润性乳腺癌侵袭性、评估预后和确定治疗方案的生物学指标。     

uPA、PAI-1的表达和MVD计数与浸润性乳腺癌侵袭性的关系

目的：探讨尿激酶型纤溶酶原激活剂（uPA）、纤溶酶原激活物抑制剂（PAI-1）的表达和血管形成与浸润性乳腺癌侵袭性的关系。方法：应用免疫组化SP法检测80例浸润性乳腺癌、20例良性乳腺肿瘤组织中uPA、PAI-1的表达情况并进行微血管计数。结果：uPA和PAI-1在浸润性乳腺癌组的阳性表达显著高于良性肿瘤组,且其高表达率与乳腺癌的临床病理参数密切相关（P〈0.05）;微血管计数在乳腺癌组和良性肿瘤组分别为30.87±7.64、20.28±8.72,两组比较有显著性差异（P〈0.01）。uPA与PAI-1在浸润性乳腺癌中的表达呈正相关（P〈0.05）。结论：uPA、PAI-1的高表达与浸润性乳腺癌的浸润、转移相关,uPA、PAI-1的高表达和MVD计数可作为评价浸润性乳腺癌侵袭性、评估预后和确定治疗方案的生物学指标。     

EGFR和KRAS在上皮性卵巢癌组织中的表达及临床意义

目的:探讨表皮生长因子受体(EGFR)和鼠Kirsten肉瘤病毒致癌基因(KRAS)蛋白在上皮性卵巢癌组织中的表达水平及临床意义。方法:利用免疫组化SP法对上皮性卵巢癌组织(病例组,n=57)、卵巢良性肿瘤组织(良性组,n=50)以及正常卵巢组织(对照组,n=50)中的EGFR、KRAS蛋白水平进行检测,并分析其在上皮性卵巢癌发生发展中的作用。结果:病例组的EGFR、KRAS蛋白阳性率高于良性组和对照组(P0.05),良性组和对照组的EGFR、KRAS蛋白阳性率差异无统计学意义(P0.05)。浆液性腺癌组EGFR、KRAS蛋白的阳性表达率高于非浆液性腺癌组,Ⅲ~Ⅳ期的上皮性卵巢癌组织中EGFR、KRAS蛋白阳性表达率高于Ⅰ~Ⅱ期,中、低分化组中EGFR、KRAS蛋白阳性表达率高于高分化组,差异均具有统计学意义(P0.05)。上皮性卵巢癌组织中EGFR与KRAS的蛋白的阳性表达率呈正相关关系(r=0.469,P0.05)。结论:EGFR及KRAS蛋白在上皮性卵巢癌组织中的表达水平明显升高,可能参与了上皮性卵巢癌的发生发展过程,且两者之间呈正相关关系,联合检测可作为早期诊断上皮性卵巢癌的重要指标。     

HE4、NF-κBp65和MMP-9在卵巢上皮性肿瘤中的表达及其临床意义

目的探讨HE4、NF-κBp65以及MMP-9的表达与上皮性卵巢肿瘤临床病理生物学行为的关系以及对患者预后的影响。方法应用免疫组化对80例卵巢上皮性癌、10例交界性上皮性肿瘤及10例良性上皮性肿瘤组织进行HE4、NF-κB p65及MMP-9蛋白的检测。结果 HE4、NF-κBp65及MMP-9蛋白的阳性表达率在卵巢癌组均高于交界性及良性肿瘤组（P〈0.05）。三个指标的表达与EOC的组织学分级、腹腔脏器及淋巴结转移以及PTNM临床分期有关（P〈0.05）;在EOC中,NF-κBp65分别与HE4、MMP-9的表达呈正相关（r1=0.673,P〈0.05;r2=0.775,P〈0.05）。多因素分析,PTNM分期、MMP-9的表达是影响卵巢癌术后患者预后的独立因素（P〈0.05）;HE4阳性组与阴性组5年生存率分别为12.7%和84.0%,MMP-9阳性组与阴性组5年生存率分别为8.7%和70.6%,差异均有统计学意义（P〈0.05）。结论 NF-κBp65可能通过上调HE4、MMP-9的表达促进EOC的浸润和转移,联合检测HE4、NF-κBp65以及MMP-9的表达可能作为预测和评价患者预后的生物学指标。     

趋化因子受体CCR3在上皮性卵巢癌中的表达及临床意义

目的:探讨趋化因子受体CCR3在上皮性卵巢癌组织中的表达情况,以及其与卵巢癌临床病理特征的关系。方法:收集上皮性卵巢癌组织、良性上皮性卵巢肿瘤组织以及正常卵巢组织标本各30例,采用多聚腺苷酸加尾实时荧光定量反转录聚合酶链反应[poly(A)-RT-qPCR]检测其CCR3的表达,并分析上皮性卵巢癌组织中CCR3的表达与患者临床病理特征之间的关系。结果:上皮性卵巢癌组织中CCR3的表达显著高于良性卵巢肿瘤组和正常卵巢组,差异有统计学意义(P0.05);且上皮性卵巢癌组织中CCR3的表达与患者的分期、组织分级及淋巴结转移均显著相关(P0.05)。结论:CCR3在上皮性卵巢癌组织中呈高表达,且在上皮性卵巢癌的发生和发展过程中均起着十分重要的作用。     

Med1/TRAP220和uPAR在非小细胞肺癌中表达的临床意义

目的:探讨Med1/TRAP220和uPAR在非小细胞肺癌中表达的临床病理意义.方法:用免疫组织化学技术检测120例非小细胞肺癌组织中Med1/TRAP220和uPAR的表达,并分析其与患者临床病理指标的关系及其和预后的关系.结果:免疫组化结果显示在120例非小细胞肺癌中Med1/TRAP220的阳性表达率为53.3％,uPAR的阳性表达率为70％,癌旁正常肺组织uPAR阳性表达率为12.5％(15/120),Med1/TRAP220的阳性表达率为80％(96/120).uPAR在肺癌中的阳性表达率显著高于癌旁正常肺组织(P＜0.01),Med1/TRAP220在癌旁正常肺组织中的阳性表达率显著高于肺癌组织(P＜0.01).Med1/TRAP220和uPAR在肺癌中的表达与性别,年龄,肿瘤大小无关(P＞0.05).有淋巴结转移组uPAR的阳性表达率显著高于无淋巴结转移组(P＜0.05),而无淋巴结转移组Med1/TRAP220的阳性表达率显著高于有淋巴结转移组(P＜0.05).uPAR的阳性表达率与非小细胞肺癌的临床分期无显著相关(P＞0.05),而Ⅰ-Ⅱ期非小细胞肺癌组Med1/TRAP220的阳性表达率显著高于Ⅲ-Ⅳ期组(P＜0.05).uPAR在肺鳞癌和腺癌中的表达率有显著差异(P＜0.05),而Med1/TRAP220在肺鳞癌和腺癌中的表达率无显著差异(P＞0.05).uPAR阳性组生存率显著低于uPAR阴性组(P＜0.05),而Med1/TRAP220阳性组生存率显著高于Med1/TRAP220阴性组(P＜0.05).Cox比例危险度模型显示uPAR阳性表达反映不良预后的危险性最大,风险度exp(β)为2.012(P＜0.01),其次为肿瘤的TNM分期,风险度exp(β)为1.521,而Med1/TRAP220为预后有利因素,风险度exp(β)为0.386(P＜0.01).结论:Med1/TRAP220和uPAR联合应用在非小细胞肺癌预后预测中有重要意义.     

雌激素受体GPR30在上皮性卵巢癌中的表达及其与MMP-9表达的相关性

本文旨在研究新型雌激素受体——G蛋白耦联受体30(G protein-coupled receptor30,GPR30)在上皮性卵巢癌中的表达,及其与基质金属蛋白酶9(matrix metalloproteinases-9,MMP-9)表达的相关性。收集39例上皮性卵巢肿瘤组织标本,包括上皮性卵巢癌30例(浆液性囊腺癌19例、粘液性囊腺癌5例和子宫内膜样癌6例)、良性上皮性卵巢肿瘤9例,并取4例正常卵巢组织进行对照,应用免疫组织化学法检测GPR30和MMP-9的表达,使用?2检验、Fisher’s精确检验和Spearman等级相关分析等统计学方法对所得数据进行统计分析。结果显示,上皮性卵巢癌中GPR30的高表达率显著高于良性卵巢肿瘤(P0.01)和正常卵巢组织(P0.05);上皮性卵巢癌中MMP-9的高表达率显著高于正常卵巢组织(P0.05),但与良性卵巢肿瘤无明显差异。肿瘤病理类型、分期和患者年龄因素与GPR30高表达率的相关性分析显示:不同病理类型上皮性卵巢癌的GPR30高表达率之间存在显著性差异(P0.05),而各病理类型的MMP-9高表达率之间没有差异;临床III~IV期上皮性卵巢癌的GPR30高表达率显著高于I~II期(P0.05),二者MMP-9高表达率之间没有差异;三个年龄组病例中的GPR30高表达率之间未见明显差异,三者MMP-9高表达率之间亦没有差异。Spearman等级相关分析显示上皮性卵巢癌组织中GPR30与MMP-9的表达之间具有正向相关性(P0.01)。以上结果提示,GPR30可能参与上皮性卵巢癌的发生发展,并与其侵袭和转移具有一定的相关性,可作为卵巢癌早期诊断和恶性程度判断的指标之一。     

微小RNA-21与PTEN在上皮性卵巢癌组织中的表达及临床意义

目的:检测微小RNA-21(miR-21)和抑癌基因PTEN在上皮性卵巢癌组织中的表达,探讨二者的相关性及其与临床病理特征间的关系.方法:采用茎环实时荧光逆转录聚合酶链反应(Real-time RT-PCR)检测48例上皮性卵巢癌组织、24例良性上皮性卵巢肿瘤及15例正常卵巢组织标本中miR-21表达.采用链霉菌抗生物素蛋白一过氧化酶(SP)免疫组化方法检测上述组织中PTEN蛋白表达,分析mir-21表达和PTEN表达的相关性及其与临床病理特征的关系.结果:茎环Real-time RT-PCR检测miR-21表达的敏感性和特异性良好;miR-21在卵巢癌组织中的相对表达量(4.849±1.813)显著高于良性卵巢肿瘤(1.133±0.291)及正常卵巢组织(P<0.01),PTEN在卵巢癌组织中阳性表达率为(41.66%),明显低于卵巢良性肿瘤(83.33%)及正常卵巢(100%).卵巢癌组织中miR-21表达量与PTEN阳性表达率存在显著负相关(r=-0.447,P<0.01).miR-21及PTEN表达随卵巢癌的临床分期和组织分级的进展分别呈上升及下降趋势,而且二者表达均与淋巴结转移相关(P均<0.01),而与组织病理类型无关.结论:miR-21在上皮性卵巢癌组织中的表达升高,可能通过负性调节PTEN表达在卵巢癌的发生发展中发挥致癌基因的作用,并且与卵巢癌的演进及不良预后密切相关.     

Skp2和P27在上皮性卵巢癌中表达及其临床意义

目的:检测Skp2和P27在上皮性卵巢癌中的表达,分析Skq2和P27与临床病理特征间的关系,探讨二者在上皮性卵巢癌发生发展中的作用.方法:应用免疫组织化学SP法,检测69例上皮性卵巢癌、15例良性卵巢肿瘤以及15例正常卵巢组织中Skp2和P27的表达,分析Skp2和P27表达与上皮性卵巢癌临床病理参数的关系,及两者的相关性.结果:上皮性卵巢癌、良性卵巢肿瘤及正常卵巢组织Skp2和P27表达阳性率各为42.02%,26.67%、6.67%和47.82%,66.67%、86.67%,差异有统计学意义(P<0.05).Skp2的表达与临床分期和病理组织学分级显著相关,与患者年龄、病理类型、有无淋巴结转移和肿瘤大小无显著相关.P27的表达与临床分期、有无淋巴结转移和病理组织学分级显著相关,与患者年龄、病理类型和肿瘤大小无显著相关.Skp2和P27在上皮性卵巢癌中表达呈显著负相关,相关系数r=-0.463,(P<0.01).结论:Skp2与P27呈负相关,二者表达可能与上皮性卵巢癌的发生发展有关.     

Urokinase-targeted recombinant bacterial protein toxins — a rationally designed and engineered anticancer agent for cancer therapy

Urokinase-targeted recombinant bacterial protein toxins are a sort of rationally designed and engineered anticancer recombinant fusion proteins representing a novel class of agents for cancer therapy. Bacterial protein toxins have long been known as the primary virulence factor(s) for a variety of pathogenic bacteria and are the most powerful human poisons. On the other hand, it has been well documented that urokinase-type plasminogen activator (uPA) and urokinase plasminogen activator receptor (uPAR), making up the uPA system, are over-expressed in a variety of human tumors and tumor cell lines. The expression of uPA system is highly correlated with tumor invasion and metastasis. To exploit these characteristics in the design of tumor cell-selective cytotoxins, two prominent bacterial protein toxins, i.e., the diphtheria toxin and anthrax toxin are deliberately engineered through placing a sequence targeted specifically by the uPA system to form anticancer recombinant fusion proteins. These uPA system-targeted bacterial protein toxins are activated selectively on the surface of uPA system-expressing tumor cells, thereby killing these cells. This article provides a review on the latest progress in the exploitation of these recombinant fusion proteins as potent tumoricidal agents. It is perceptible that the strategies for cancer therapy are being innovated by this novel therapeutic approach.     

Urokinase-targeted recombinant bacterial protein toxins-a rationally designed and engineered anticancer agent for cancer therapy

Urokinase-targeted recombinant bacterial protein toxins are a sort of rationally designed and engineered anticancer recombinant fusion proteins representing a novel class of agents for cancer therapy.Bacterial protein toxins have long been known as the primary virulence factor(s) for a variety of pathogenic bacteria and are the most powerful human poisons.On the other hand,it has been well documented that urokinase-type plasminogen activator (uPA) and urokinase plasminogen activator receptor (uPAR),making up the uPA system,are overexpressed in a variety of human tumors and tumor cell lines.The expression of uPA system is highly correlated with tumor invasion and metastasis.To exploit these characteristics in the design of tumor cell-selective cytotoxins,two prominent bacterial protein toxins,i.e.,the diphtheria toxin and anthrax toxin are deliberately engineered through placing a sequence targeted specifically by the uPA system to form anticancer recombinant fusion proteins.These uPA system-targeted bacterial protein toxins are activated selectively on the surface of uPA systemexpressing tumor cells,thereby killing these cells.This article provides a review on the latest progress in the exploitation of these recombinant fusion proteins as potent tumoricidal agents.It is perceptible that the strategies for cancer therapy are being innovated by this novel therapeutic approach.     

Proteomic profiling of proteins associated with urokinase plasminogen activator receptor in a colon cancer cell line using an antisense approach

Expression of urokinase plasminogen activator (uPA) and its receptor (uPAR) strongly correlates with a malignant tumour cell phenotype. In the multistep process of metastasis, uPA binding to uPAR influences different cellular functions. In the present study, a highly metastatic colon cancer cell line, HCT116 was transfected with an expression vector containing a 5' uPAR cDNA fragment in an antisense orientation. This construct was most effective in reducing uPAR cell surface expression as confirmed by flow cytometry analysis. Antisense transfection of HCT116 cells had no effect on proliferation but the following effects were observed: (1) a 1.3-fold decreased adhesion; (2) a two-fold decreased Erk MAP kinase activity; (3) a 2.7-fold decrease in Src kinase activity; (4) a 1.5- and two-fold decrease in uPA cell surface expression and secretion; (5) abrogation of promatrix metalloproteinase-9 secretion; and (6) a complete suppression of plasminogen-dependent matrix degradation. Using proteomic analysis, we demonstrate loss of approximately 200 proteins and quantitative differences in the expression of 141 other proteins in an antisense-clone compared to wild-type and mock-transfected control. Such changes in protein expression with the down-regulation of uPAR may be an important contributor in colon cancer progression and metastasis and may not only provide a basis to develop a proteomic data bank of uPAR-mediated signaling molecules but may also lead to the development of therapeutic approaches for the cure and better management of colon cancer.     

The effects of vitronectin on specific interactions between urokinase-type plasminogen activator and its receptor: ab initio molecular orbital calculations

Binding of urokinase-type plasminogen activator (uPA) to its receptor (uPAR) on the surface of a cancer cell is considered to be a trigger for starting cancer invasions. In addition, the somatomedin B (SMB) domain of vitronectin binds simultaneously to uPAR to construct a ternary complex of uPAR–uPA–SMB. Here we present stable structures of the solvated complexes of uPAR–uPA and uPAR–uPA–SMB obtained by classical molecular mechanics simulations, and the specific interactions between uPAR, uPA and SMB are investigated by ab initio fragment molecular orbital calculations. The result indicates that the SMB binding enhances the binding affinity between uPAR and uPA, although there is no direct contact between SMB and uPA. In particular, the specific interaction between uPAR and the Lys36 residue of uPA is significantly affected by the SMB binding. The positively charged Lys23, Lys46 and Lys61 residues of uPA have strong attractive interactions to uPAR in both the uPAR–uPA and uPAR–uPA–SMB complexes, demonstrating the importance of these residues in the specific binding between uPAR and uPA. The current results on the specific interactions are informative for proposing potent antagonists, which block the uPA and SMB bindings to uPAR.     

Noncatalytic domain of uPA stimulates human extravillous trophoblast migration by using phospholipase C,phosphatidylinositol 3-kinase and mitogen-activated protein kinase

The serine protease urokinase-type plasminogen activator (uPA) promotes matrix degradation by many cell types, including the invasive extravillous trophoblast (EVT) of the human placenta. The noncatalytic amino-terminal end of uPA binds to uPA receptors (uPARs) expressed by these cells. A highly polarized expression of uPAR-bound uPA at the migration front of EVT cells in situ suggests a functional role of uPA:uPAR interaction in EVT cell migration. The present study examined whether uPA stimulates EVT cell migration, independent of proteolytic function, and investigated some of the signaling pathways involved. Using in vitro-propagated EVT cells in Transwell migration assays, both uPA and its noncatalytic amino-terminal fragment (ATF) were shown to stimulate migration through multiporous polycarbonate (pore size 8 microm) membranes. A uPAR-blocking antibody inhibited basal and ATF-stimulated migration. Migration was found to be stimulated by hypoxic conditions, which upregulates uPAR expression; this stimulation was abrogated with the uPAR-blocking antibody, indicating the role of endogenous uPA in EVT cell migration. Spectrofluorometric measurement of cytosolic calcium in cells treated with uPA and ATF demonstrated a rapid rise in [Ca2+](i), which was prevented by pretreatment of cells with thapsigargin, indicating a release from intracellular stores. Both basal and ATF-mediated migratory responses were suppressed in the presence of selective pharmacological inhibitors LY294002, U73122, and U0126, implicating the respective roles of phosphatidinylinositol 3-kinase (PI 3-K), phospholipase C (PLC), and MEK1/2 in basal and ATF-stimulated migratory capacity. Taken together, these results demonstrate that uPA:uPAR interaction stimulates EVT cell migration, independent of uPA enzymatic activity, using the mitogen-activated protein kinase pathway and calcium signaling events including the participation of PI 3-K and PLC. These findings are relevant to clinical conditions of aberrant trophoblast migration, including spontaneous abortion, preeclampsia, and choriocarcinoma.     

子宫内膜癌组织中uPA及Cath-D的表达及意义(英文)

目的:检测子宫内膜癌组织中尿激酶型纤溶酶原激活物(uPA)及组织蛋白酶(Cath-D)的表达并探讨相关性及其临床意义。方法:采用免疫组织化学方法(PV-6000二步法)检测31例子宫内膜癌组织(内膜癌组),17例子宫内膜增生组织(增生组)及10例正常子宫内膜组织(对照组)中uPA及Cath-D的表达,并研究其相关性。结果:1.内膜癌组中uPA和Cath-D的表达均高于增生组及对照组中的表达,差异均有统计学意义(P0.05);在增生组中的表达与对照组差异无统计学意义(P0.05)。2.uPA和Cath-D的阳性表达与子宫内膜癌的临床病理分期、组织学分级及肌层浸润深度有关,差异均具有统计学意义(P0.05)。3.内膜癌组中uPA与Cath-D的表达呈正相关(r=0.673,P0.05)。结论:uPA和Cath-D在子宫内膜癌发生发展及侵袭转移过程中起着协同作用,Cath-D可诱导产生活化的uPA,促进癌细胞的浸润转移,因此,两者的联合检测可有助于成为判断子宫内膜癌的发展及预后的重要指标。     

High level synthesis of recombinant soluble urokinase receptor (CD87) by ovarian cancer cells reduces intraperitoneal tumor growth and spread in nude mice

Focussing of the serine protease urokinase (uPA) to the tumor cell surface via interaction with its receptor (uPAR) is an important step in tumor invasion and metastasis. The human ovarian cancer cell line OV-MZ-6#8 was stably transfected with expression plasmids either encoding cell-associated uPAR (GPI-uPAR) or a soluble form of uPAR (suPAR) lacking its glycan lipid anchor. In vitro, high level synthesis of functionally active recombinant suPAR inhibited cell proliferation and led to reduced cell-associated fibrin matrix degradation, whereas fibrinolytic activity was increased in OV-MZ-6#8 cells overexpressing GPI-uPAR. Both OV-MZ-6#8-derived clones were inoculated into the peritoneum of nude mice and tested for tumor growth and spread. High level synthesis of recombinant suPAR (without altering the physiological expression levels of GPI-uPAR and uPA in these cells) resulted in a significant reduction of tumor burden (up to 86%) in the xenogeneic mouse model. In contrast, overexpression of GPI-uPAR in tumor cells did not affect tumor growth. Our results demonstrate that high levels of suPAR in the ovarian cancer cell vicinity can act as a potent scavenger for uPA, thereby significantly reducing tumor cell growth and cancer progression in vivo.     

High-affinity urokinase-derived cyclic peptides inhibiting urokinase/urokinase receptor-interaction: effects on tumor growth and spread

Urokinase-type plasminogen activator (uPA) binds with high affinity to its specific cell surface receptor (uPAR) (CD87) via a well-defined sequence within the N-terminal region of uPA (uPA(19-31)). Since this uPA/uPAR-interaction plays a significant role in tumor cell invasion and metastasis, it has become an attractive therapeutic target. Two small peptidic cyclic competitive antagonists of uPA/uPAR-interaction have been developed, based on the uPAR binding site in uPA: WX-360 (cyclo(21,29)[D-Cys21]-uPA(21-30)[S21C;H29C]) and its norleucine (Nle) derivative WX-360-Nle (cyclo(21,29)[D-Cys21]-uPA(21-30)[S21C;K23Nle;H29C]). These peptides display an only five to 10-fold lower affinity to uPAR as compared to the naturally occurring uPAR-ligand uPA. In this study, WX-360 and WX-360-Nle were tested in nude mice for their potency to inhibit tumor growth and intraperitoneal spread of lacZ-tagged human ovarian cancer cells. Intraperitoneal administration of either cyclic peptide (20 mg peptide/kg; 1x daily for 37 days) into the tumor-bearing nude mice resulted in a significant reduction of tumor weight and spread within the peritoneum as compared to the untreated control group. This is the first report demonstrating effective reduction of tumor growth and spread of human ovarian cancer cells in vivo by small synthetic uPA-derived cyclic peptides competitively interfering with uPA/uPAR-interaction. Thus, both WX-360 and WX-360-Nle are promising novel compounds to reduce dissemination of human ovarian carcinoma.     

MConfocal Fluorescence Microscopy of Urokinase Plasminogen Activator Receptor and Cathepsin D in Human MDA-MB-231 Breast Cancer Cells Migrating in Reconstituted Basement Membrane

Using confocal fluorescence microscopy with a monoclonal antibody, we have localized the receptor for urokinase plasminogen activator (uPAR) in MDA-MB-231 human breast cancer cells migrating into a reconstituted basement membrane. Patchy and polarized uPAR immunoreactivity was found at the cell membrane, and strong staining was found both in the ruffled border or leading edge of the cells and at pseudopodia penetrating into the membrane. Intracellular uPAR staining was localized in the paranuclear region and in rounded granule-like structures: some of these were identified as lysosomes by double staining for uPAR and the lysosomal enzyme cathepsin D. Urokinase plasminogen activator (uPA) activity has previously been shown to play a role in migration of cells into basement membranes, and it has been proposed that uPAR also is involved in this process. uPA is known to be internalized and degraded after complex formation with the inhibitor PAI-1. Lysosomal uPAR immunoreactivity may result from concomitant internalization of the receptor.