The receptor-binding sequence of urokinase. A biological function for the growth-factor module of proteases.

Previous studies have shown that the region of human urokinase-type plasminogen activator (uPA) responsible for receptor binding resides in the amino-terminal fragment (ATF, residues 1-135) (Stoppelli, M.P., Corti, A., Soffientini, A., Cassani, G., Blasi, F., and Assoian, R.K. (1985) Proc. Natl. Acad. Sci. U.S. A. 82, 4939-4943). The area within ATF responsible for specific receptor binding has now been identified by the ability of different synthetic peptides corresponding to different regions of the amino terminus of uPA to inhibit receptor binding of 125I-labeled ATF. A peptide corresponding to human [Ala19]uPA-(12-32) resulted in 50% inhibition of ATF binding at 100 nM. Peptides uPA-(18-32) and [Ala13]uPA-(9-20) inhibit at 100 and 2000 microM, respectively. The human peptide uPA-(1-14) and the mouse peptide [Ala20]uPA-(13-33) have no effect on ATF receptor binding. This region of uPA is referred to as the growth factor module since it shares partial amino acid sequence homology (residues 14-33) to epidermal growth factor (EGF). Furthermore, this region of EGF is responsible for binding of EGF to its receptor (Komoriya, A. Hortsch, M., Meyers, C., Smith, M., Kanety, H., and Schlessinger, J. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 1351-1355). However, EGF does not inhibit ATF receptor binding. Comparison of the sequences responsible for receptor binding of uPA and EGF indicate that the region of highest homology is between residues 13-19 and 14-20 of human uPA and EGF, respectively. In addition, there is a conservation of the spacings of four cysteines in this module whereas there is no homology between residues 20-30 and 21-33 of uPA and EGF. Thus, residues 20-30 of uPA apparently confer receptor binding specificity, and residues 13-19 provide the proper conformation to the adjacent binding region

Separation and characterization of nonphosphorylated and serine-phosphorylated urokinase. Catalytic properties and sensitivity to plasminogen activator inhibitor type 1.

Urokinase synthesized by human A431 epidermoid carcinoma cells is phosphorylated on serine (Mastronicola, M. R., Stoppelli, M. P., Migliaccio, A., Auricchio, F., and Blasi, F. (1990) FEBS Lett. 266, 109-114). To test the possibility that phosphorylation may have specific effects on urokinase function, the phosphorylated and nonphosphorylated forms of urokinase were separated by Fe(3+)-Sepharose chromatography. Both forms exhibit indistinguishable Km and kcat for plasminogen activation. On the other hand, their sensitivity toward the specific plasminogen activator inhibitor type 1 is different as assessed by measuring both the stability of the covalent complex and the residual enzymatic activity. Phosphorylated urokinase was 50% inhibited at a concentration of plasminogen activator inhibitor type 1 4-fold higher than nonphosphorylated urokinase (0.7 versus 0.15 nM). Furthermore about 10% of phosphorylated urokinase was resistant to plasminogen activator inhibitor type 1 at a concentration as high as 20 nM. Thus, phosphorylation affects urokinase sensitivity to plasminogen activator inhibitor type 1, therefore resulting in a net, although indirect, increase of urokinase activity. These results suggest the existence of a novel cellular regulatory mechanism of extracellular proteolysis

Breast tumor cell invasion and pro-invasive activity of cancer-associated fibroblasts co-targeted by novel urokinase-derived decapeptides

Among peritumoral cells, cancer-associated fibroblasts (CAFs) are major facilitators of tumor progression. This study describes the effects of two urokinase-derived, novel decapeptides, denoted as Pep 1 and its cyclic derivative Pep 2. In a mouse model of tumor dissemination, using HT1080 fibrosarcoma cells, Pep 2 reduced the number and size of lung metastases. Specific binding of fluoresceinated Pep 2 to HT1080 and telomerase immortalised fibroblasts (TIF) cell surfaces was enhanced by αv overexpression or abolished by excess vitronectin, anti-αv antibodies or silencing of ITGAV αv gene, identifying αv-integrin as the Pep 2 molecular target. In 3D-organotypic assays, peptide-exposed TIFs and primary CAFs from breast carcinoma patients both exhibited a markedly reduced pro-invasive ability of either HT1080 fibrosarcoma or MDA-MB-231 mammary carcinoma cells, respectively. Furthermore, TIFs, either exposed to Pep 2, or silenced for αv integrin, were impaired in their ability to chemoattract cancer cells and to contract collagen matrices, exhibiting reduced α-smooth muscle actin (α-SMA) levels. Finally, peptide exposure of αv-expressing primary CAFs led to the downregulation of α-SMA protein and to a dramatic reduction of their pro-invasive capability. In conclusion, the ability of the novel decapeptides to interfere with tumor cell invasion directly and through the down-modulation of CAF phenotype suggests their use as lead compounds for co-targeting anti-cancer strategies

The effects of poetry-writing SANTEL on erotic body image in remission of cancer in women: a pilot study

International audienceAbstract Aim: Our pilot study aims to describe the effects ofa new specific and structured protocol focused on poetic/erotic writing (named SANTEL) on the (re)sexualization ofbody image in women, who have experienced cancer.Procedure: The protocol consists of four steps: to choose alist of erotic verses focused on the body parts, to fill a semistructuredpoetic text, to write sentences after target phraseson the body; and in the end, to write a free poem. Mrs V.suffered from breast cancer, and one breast was removed.She and her husband participated in this poetic writing protocol,separately. We analyzed the linguistic metaphors ofthe body by QSR Nvivo10 software.Results: Using this protocol, we showed discourse variationsof metaphors before and after the experience of writing.Patient V used “I feel like an alien” as a starting metaphorto describe her cancer experience and after poetic writingsessions, she used other bodily metaphors like “My body isa flower” and “My sensual and white flesh”.Conclusion: This poetic perspective promises a type of“perceptive-literary surgery”, characterized by a sensualinvestment process after remission: a poetic reconstructionof erotic body image.Les effets d'un protocole d'écriture poétique SANTEL sur l'image érotique du corps dans le traitement du cancer féminin : étude pilote The effects of poetry-writing SANTEL on erotic body image in remission of cancer in women: a pilot study A. Santarpia · J. Tellène · M. Carrier Résumé Objectif : Cette étude pilote de type qualitative et exploratoire vise à décrire les effets d'un nouveau protocole d'écriture poético-érotique (nommée SANTEL) sur la rééro-tisation de l'image du corps chez une femme, ayant vécu un cancer. Matériel et méthodes : Il s'agit d'un protocole composé de quatre étapes : une liste des phrases à caractères poétiques et érotiques à choisir, un texte à trous à remplir, des amorces de phrases ciblées sur le corps et en fin un poème libre. Madame V. a subi un cancer du sein nécessitant une ablation complète. Madame V. et son conjoint exécutent le protocole d'écriture séparément. Nous montrons les variations discursives des métaphores utilisées avant et après l'expérience de l'écriture, à travers le logiciel d'analyse qualitative QSR NVivo10. Résultats : Madame V. passera de la métaphore initiale « je me sens une extraterrestre » vers la plus atténuée « Non. Je me dis qu'extraterrestre c'était peut-être un peu énorme ». En plus, elle utilisera de nouvelles métaphores linguistiques du corps pour raconter son image du corps telles que « ce corps de chair blanche » et « une fleur qui s'ouvre délicatement ». Conclusion : Cet exercice spécifique d'écriture promet un type de « chirurgie perceptive-littéraire » dans le processus d'investissement sensuel et affectif après la rémission, une reconstruction perceptive et poétique de l'image érotique du corps. Mots clés Métaphores perceptives · Image du corps · Cancer féminin · Corps érotique · Écriture poétique · Chirurgie perceptive-littéraire · Logiciel QSR NVivo10. Abstract Aim: Our pilot study aims to describe the effects of a new specific and structured protocol focused on poetic/ erotic writing (named SANTEL) on the (re)sexualization of body image in women, who have experienced cancer. Procedure: The protocol consists of four steps: to choose a list of erotic verses focused on the body parts, to fill a semi-structured poetic text, to write sentences after target phrases on the body; and in the end, to write a free poem. Mrs V. suffered from breast cancer, and one breast was removed. She and her husband participated in this poetic writing protocol , separately. We analyzed the linguistic metaphors of the body by QSR Nvivo10 software. Results: Using this protocol, we showed discourse variations of metaphors before and after the experience of writing. Patient V used " I feel like an alien " as a starting metaphor to describe her cancer experience and after poetic writing sessions, she used other bodily metaphors like " My body is a flower " and " My sensual and white flesh ". Conclusion: This poetic perspective promises a type of " perceptive-literary surgery " , characterized by a sensual investment process after remission: a poetic reconstruction of erotic body image. Keywords Bodily metaphors · Body image · Feminine cancer · Erotic body · Poetry writing · Perceptive-literary surgery · QSR Nvivo10 software

Identification and primary sequence of an unspliced human urokinase poly(A)+ RNA.

Human urokinase cDNA clones have been identified from a cDNA library prepared from total RNA of human fibroblasts transformed by simian virus 40 [Okayama, H. & Berg, P. (1983) Mol. Cell. Biol. 3, 280-289]. Synthetic oligonucleotides, corresponding to urokinase protein sequence, were used as probes. The cloned cDNA covers most of the coding sequence and the entire 3' untranslated region. The nucleotide sequence of one of the clones identifies this as a copy of a partially spliced polyadenylylated precursor to urokinase mRNA. The introns separate functionally different domains of the enzyme. Human urokinase mRNA has been identified by RNA blot and its size was estimated at 2500 nucleotides

Analysis of secretome changes uncovers an autocrine/paracrine component in the ability of c -Myc to modulate cell proliferation and motility

Proteins secreted by cancer cells are a major component of tumor microenvironment. However, little is known on the impact of single oncogenic lesions on the expression of secreted proteins at early stages of tumor development. Because c-Myc over-expression is among the most frequent alterations in cancer, here we investigated the effect of sustained c-Myc expression on the secretome of a non-transformed human epithelial cell line (hT-RPE). By using a quantitative proteomic approach we have identified 125 proteins in conditioned media of hT-RPE/MycER cells upon c-Myc induction. Analysis of the 49 proteins significantly down-regulated by c-Myc, revealed a marked enrichment of factors associated with growth inhibition and cellular senescence. Accordingly, media conditioned by hT-RPE cells expressing c-Myc show an increased ability to sustain hT-RPE cellular proliferation/viability. We also find a marked down-regulation of several structural and regulatory components of the extracellular matrix (ECM), which correlates with an increased chemotactic potency of the conditioned media towards fibroblasts, a major cellular component of tumor stroma. In accordance with these data, the expression of the majority of the genes encoding proteins down-regulated in hT-RPE was significantly reduced also in colorectal adenomatous polyps, early tumors in which c-Myc is invariably over-expressed. These findings help to elucidate the significance of c-Myc over-expression at early stages of tumor development and uncover a remarkable autocrine/paracrine component in the ability of c-Myc to stimulate proliferation, sustain tumor maintenance and modulate cell migration

αV-Integrin-Dependent Inhibition of Glioblastoma Cell Migration, Invasion and Vasculogenic Mimicry by the uPAcyclin Decapeptide

Among the deadliest human cancers is glioblastoma (GBM) for which new treatment approaches are urgently needed. Here, the effects of the cyclic decapeptide, uPAcyclin, are investigated using the U87-MG, U251-MG, and U138-MG human GBM and C6 rat cell models. All GBM cells express the αV-integrin subunit, the target of uPAcyclin, and bind specifically to nanomolar concentrations of the decapeptide. Although peptide exposure affects neither viability nor cell proliferation rate, nanomolar concentrations of uPAcyclin markedly inhibit the directional migration and matrix invasion of all GBM cells, in a concentration- and αV-dependent manner. Moreover, wound healing rate closure of U87-MG and C6 rat glioma cells is reduced by 50% and time-lapse videomicroscopy studies show that the formation of vascular-like structures by U87-MG in three-dimensional matrix cultures is markedly inhibited by uPAcyclin. A strong reduction in the branching point numbers of the U87-MG, C6, and U251-MG cell lines undergoing vasculogenic mimicry, in the presence of nanomolar peptide concentrations, was observed. Lysates from matrix-recovered uPAcyclin-exposed cells exhibit a reduced expression of VE-cadherin, a prominent factor in the acquisition of vascular-like structures. In conclusion, these results indicate that uPAcyclin is a promising candidate to counteract the formation of new vessels in novel targeted anti-GBM therapies