Skip to main content
Key Specialties
Cardiology Diabetology Endocrinology Gastroenterology Geriatrics and Gerontology Gynecology and Obstetrics Hematology Infectious Disease Internal Medicine Nephrology Neurology Oncology Pediatrics Respiratory Medicine Rheumatology Surgery
Congress Coverage
2024 ACC Congress 2023 ESMO Congress 2023 EASD Congress 2023 ERS Congress 2023 ESC Congress 2023 EHA Congress 2023 ASCO Annual Meeting
Clinical Collections
Advances in Alzheimer’s

At a glance: The ONWARDS insulin icodec trials

A round-up of the ONWARDS phase 3 clinical trials evaluating the novel weekly insulin icodec in people with diabetes.
ADVANCED SEARCH
Log in
Top
Journal of Experimental & Clinical Cancer Research
Published in: Journal of Experimental & Clinical Cancer Research 1/2017

Open Access 01-12-2017 | Research

Targeting the cross-talk between Urokinase receptor and Formyl peptide receptor type 1 to prevent invasion and trans-endothelial migration of melanoma cells

Authors: Concetta Ragone, Michele Minopoli, Vincenzo Ingangi, Giovanni Botti, Federica Fratangelo, Antonello Pessi, Maria Patrizia Stoppelli, Paolo Antonio Ascierto, Gennaro Ciliberto, Maria Letizia Motti, Maria Vincenza Carriero

Published in: Journal of Experimental & Clinical Cancer Research | Issue 1/2017

Login to get access

Abstract

Background

Accumulating evidence demonstrates that the Urokinase Receptor (uPAR) regulates tumor cell migration through its assembly in composite regulatory units with transmembrane receptors, and uPAR88–92 is the minimal sequence required to induce cell motility through the Formyl Peptide Receptor type 1 (FPR1). Both uPAR and FPR1 are involved in melanoma tumor progression, suggesting that they may be targeted for therapeutic purposes. In this study, the role of the uPAR-FPR1 cross-talk to sustain melanoma cell ability to invade extracellular matrix and cross endothelial barriers is investigated. Also, the possibility that inhibition of the uPAR mediated FPR1-dependent signaling may prevent matrix invasion and transendothelial migration of melanoma cells was investigated.

Methods

Expression levels of uPAR and FPR1 were assessed by immunocytochemistry, Western Blot and qRT-PCR. Cell migration was investigated by Boyden chamber and wound-healing assays. Migration and invasion kinetics, trans-endothelial migration and proliferation of melanoma cells were monitored in real time using the xCELLigence technology. The agonist-triggered FPR1 internalization was visualized by confocal microscope. Cell adhesion to endothelium was determined by fluorometer measurement of cell-associated fluorescence or identified on multiple z-series by laser confocal microscopy. The 3D–organotypic models were set up by seeding melanoma cells onto collagen I matrices embedded dermal fibroblasts. Data were analyzed by one-way ANOVA and post-hoc Dunnett t-test for multiple comparisons.

Results

We found that the co-expression of uPAR and FPR1 confers to A375 and M14 melanoma cells a clear-cut capability to move towards chemotactic gradients, to cross extracellular matrix and endothelial monolayers. FPR1 activity is required, as cell migration and invasion were abrogated by receptor desensitization. Finally, melanoma cell ability to move toward chemotactic gradients, invade matrigel or fibroblast-embedded collagen matrices and cross endothelial monolayers are prevented by anti-uPAR84–95 antibodies or by the RI-3 peptide which we have previously shown to inhibit the uPAR84–95/FPR1 interaction.

Conclusions

Collectively, our findings identify uPAR and FPR1 as relevant effectors of melanoma cell invasiveness and suggest that inhibitors of the uPAR84–95/FPR1 cross-talk may be useful for the treatment of metastatic melanoma.
Appendix
Available only for authorised users
Literature
1.
2.
3.
Ascierto PA, Grimaldi AM, Anderson A, Bifulco C, Cochran A, Garbe C, et al. Future perspectives in melanoma research: meeting report from the “melanoma bridge”, Napoli, December 5th-8th 2013. J Transl Med. 2014;12:277.CrossRefPubMedPubMedCentral
4.
Azijli K, Stelloo E, Peters GJ, Van Den Eertwegh AJ. New developments in the treatment of metastatic melanoma: immune checkpoint inhibitors and targeted therapies. Anticancer Res. 2014;34:1493–505.PubMed
5.
Shi H, Hugo W, Kong X, Hong A, Koya RC, Moriceau G, et al. Acquired resistance and clonal evolution in melanoma during BRAF inhibitor therapy. Cancer Discov. 2014;4:80–93.CrossRefPubMed
6.
Van Allen EM, Wagle N, Sucker A, Treacy DJ, Johannessen CM, Goetz EM, et al. The genetic landscape of clinical resistance to RAF inhibition in metastatic melanoma. Cancer Discov. 2014;4:94–109.CrossRefPubMed
7.
Luke JJ, Flaherty KT, Ribas A, Long GV. Targeted agents and immunotherapies: optimizing outcomes in melanoma. Nat Rev Clin Oncol. 2017;14(8):463–82.CrossRefPubMed
8.
Malas S, Harrasser M, Lacy KE, Karagiannis SN. Antibody therapies for melanoma: new and emerging opportunities to activate immunity (review). Oncol Rep. 2014;32(3):875–86.CrossRefPubMedPubMedCentral
9.
De Vries TJ, Quax PH, Denijn M, Verrijp KN, Verheijen JH, Verspaget HW, et al. Plasminogen activators, their inhibitors, and urokinase receptor emerge in late stages of melanocytic tumor progression. Am J Pathol. 1994;144:70–81.PubMedPubMedCentral
10.
Quax PH, van Muijen GN, Weening-Verhoeff EJ, Lund LR, Dano K, Ruiter DJ, et al. Metastatic behavior of human melanoma cell lines in nude mice correlates with urokinase-type plasminogen activator, its type-1 inhibitor, and urokinase-mediated matrix degradation. J Cell Biol. 1991;115:191–9.CrossRefPubMed
11.
Rofstad EK, Rasmussen H, Galappathi K, Mathiesen B, Nilsen K, Graff BA. Hypoxia promotes lymph node metastasis in human melanoma xenografts by up-regulating the urokinase-type plasminogen activator receptor. Cancer Res. 2002;62:1847–53.PubMed
12.
Besch R, Berking C, Kammerbauer C, Degitz K. Inhibition urokinase type plasminogen activator receptor induces apoptosis in melanoma cells by activation of p53. Cell Death Differ. 2007;14(4):818–29.CrossRefPubMed
13.
Matheis F, Heppt MV, Graf SA, Düwell P, Kammerbauer C, Aigner A, et al. A Bifunctional approach of Immunostimulation and uPAR inhibition shows potent antitumor activity in melanoma. J Invest Dermatol. 2016;136(12):2475–84.CrossRefPubMed
14.
Jacobsen B, Ploug M. The urokinase receptor and its structural homologue C4.4A in human cancer: expression, prognosis and pharmacological inhibition. Curr Med Chem. 2008;15:2559–73.CrossRefPubMed
15.
Sidenius N, Blasi F. The urokinase plasminogen activator system in cancer: recent advances and implication for prognosis and therapy. Cancer Metastasis Rev. 2003;22:205–22.CrossRefPubMed
16.
Carriero MV, Stoppelli MP. The urokinase-type plasminogen activator and the generation of inhibitors of urokinase activity and signaling. Curr Pharm Des. 2011;17:1944–61.CrossRefPubMed
17.
Mazar AP. Urokinase plasminogen activator receptor choreographs multiple ligand interactions: implications for tumor progression and therapy. Clin Cancer Res. 2008;14(18):5649–55.CrossRefPubMed
18.
Carriero MV, Del Vecchio S, Franco P, Potena MI, Chiaradonna F, Botti G, et al. Vitronectin binding to urokinase receptor in human breast cancer. Clin Cancer Res. 1997;3(8):1299–308.PubMed
19.
Carriero MV, Del Vecchio S, Capozzoli M, Franco P, Fontana L, Zannetti A, et al. Urokinase receptor interacts with alpha(v)beta5 vitronectin receptor, promoting urokinase-dependent cell migration in breast cancer. Cancer Res. 1999;59(20):5307–14.PubMed
20.
Resnati M, Pallavicini I, Wang JM, Oppenheim J, Serhan CN, Romano M, et al. The fibrinolytic receptor for urokinase activates the G protein-coupled chemotactic receptor FPRL1/LXA4R. Proc Natl Acad Sci U S A. 2002;99(3):1359–64.CrossRefPubMedPubMedCentral
21.
Montuori N, Carriero MV, Salzano S, Rossi G, Ragno P. The cleavage of the urokinase receptor regulates its multiple functions. J Biol Chem. 2002;277(49):46932–9.CrossRefPubMed
22.
Gargiulo L, Longanesi-Cattani I, Bifulco K, Franco P, Raiola R, Campiglia P, et al. Cross-talk between fMLP and vitronectin receptors triggered by urokinase receptor-derived SRSRY peptide. J Biol Chem. 2005;280(26):25225–32.CrossRefPubMed
23.
Blasi F, Sidenius N. The urokinase receptor: focused cell surface proteolysis, cell adhesion and signaling. FEBS Lett. 2010;584(9):1923–30.CrossRefPubMed
24.
Smith HW, Marshall CJ. Regulation of cell signalling by uPAR. Nat Rev Mol Cell Biol. 2010;11(1):23–36.CrossRefPubMed
25.
Ploug M, Ellis V. Structure-function relationships in the receptor for urokinase-type plasminogen activator. Comparison to other members of the Ly-6 family and snake venom alpha-neurotoxins. FEBS Lett. 1994;349(2):163–8.CrossRefPubMed
26.
Gårdsvoll H, Jacobsen B, Kriegbaum MC, Behrendt N, Engelholm L, Østergaard S, et al. Conformational regulation of urokinase receptor function: impact of receptor occupancy and epitope-mapped monoclonal antibodies on lamellipodia induction. J Biol Chem. 2011;286(38):33544–56.CrossRefPubMedPubMedCentral
27.
Huai Q, Mazar AP, Kuo A, Parry GC, Shaw DE, Callahan J, et al. Structure of human urokinase plasminogen activator in complex with its receptor. Science. 2006;311(5761):656–9.CrossRefPubMed
28.
Barinka C, Parry G, Callahan J, Shaw DE, Kuo A, Bdeir K, et al. Structural basis of interaction between urokinase-type plasminogen activator and its receptor. J Mol Biol. 2006;363(2):482–95.CrossRefPubMedPubMedCentral
29.
Xu X, Gardsvoll H, Yuan C, Lin L, Ploug M, Huang M. Crystal structure of the urokinase receptor in a ligand-free form. J Mol Biol. 2012;416(5):629–41.CrossRefPubMed
30.
Fazioli F, Resnati M, Sidenius N, Higashimoto Y, Appella E, Blasi F. A urokinase-sensitive region of the human urokinase receptor is responsible for its chemotactic activity. EMBO J. 1997;16(24):7279–86.CrossRefPubMedPubMedCentral
31.
Bifulco K, Longanesi-Cattani I, Gala M, DI Carluccio G, Masucci MT, Pavone V, et al. The soluble form of urokinase receptor promotes angiogenesis through its ser(88)-Arg-ser-Arg-Tyr(92) chemotactic sequence. J Thromb Haemost. 2010;8(12):2789–99.CrossRefPubMed
32.
33.
Ye RD, Boulay F, Wang JM, Dahlgren C, Gerard C, Parmentier M, et al. International Union of Basic and Clinical Pharmacology. LXXIII. Nomenclature for the formyl peptide receptor (FPR) family. Pharmacol Rev. 2009;61(2):119–61.CrossRefPubMedPubMedCentral
34.
Panaro MA, Acquafredda A, Sisto M, Lisi S, Maffione AB, Mitolo V. Biological role of the N-formyl peptide receptors. Immunopharmacol Immunotoxicol. 2006;28(1):103–27.CrossRefPubMed
35.
Chakravarti N, Peddareddigari VG, Warneke CL, Johnson MM, Overwijk WW, Hwu P, et al. Differential expression of the G-protein-coupled formyl peptide receptor in melanoma associates with aggressive phenotype. Am J Dermatopathol. 2013;35(2):184–90.CrossRefPubMed
36.
Zhang L, Wang H, Yang T, Su Z, Fang D, Wang Y, et al. Formylpeptide receptor 1 mediates the tumorigenicity of human hepatocellular carcinoma cells. Oncoimmunology. 2015;5(2):e1078055.CrossRefPubMedPubMedCentral
37.
Liu M, Zhao J, Chen K, Bian X, Wang C, Shi Y, et al. G protein-coupled receptor FPR1 as a pharmacologic target in inflammation and human glioblastoma. Int Immunopharmacol. 2012;14(3):283–8.CrossRefPubMedPubMedCentral
38.
Bifulco K, Longanesi-Cattani I, Franco P, Pavone V, Mugione P, Di Carluccio G, et al. Single amino acid substitutions in the chemotactic sequence of urokinase receptor modulate cell migration and invasion. PLoS One. 2012;7(9):e44806.CrossRefPubMedPubMedCentral
39.
Bifulco K, Longanesi-Cattani I, Gargiulo L, Maglio O, Cataldi M, De Rosa M, et al. An urokinase receptor antagonist that inhibits cell migration by blocking the formyl peptide receptor. FEBS Lett. 2008;582(7):1141–6.CrossRefPubMed
40.
Carriero MV, Longanesi-Cattani I, Bifulco K, Maglio O, Lista L, Barbieri A, et al. Structure-based design of an urokinase-type plasminogen activator receptor-derived peptide inhibiting cell migration and lung metastasis. Mol Cancer Ther. 2009;8(9):2708–17.CrossRefPubMed
41.
Bifulco K, Longanesi-Cattani I, Liguori E, Arra C, Rea D, Masucci MT, et al. A Urokinase receptor–derived peptide inhibiting VEGF-dependent directional migration and vascular sprouting. Mol Cancer Ther. 2013;12(10):1981–93.CrossRefPubMed
42.
Carriero MV, Bifulco K, Minopoli M, Lista L, Maglio O, Mele L, et al. UPARANT: a urokinase receptor-derived peptide inhibitor of VEGF-driven angiogenesis with enhanced stability and in vitro and in vivo potency. Mol Cancer Ther. 2014;13(5):1092–104.CrossRefPubMed
43.
Carver JA, Esposito G, Viglino P, Fogolari F, Guichard G, Briand JP, et al. Structural comparison between retro-inverso and parent peptides: molecular basis for the biological activity of a retro-inverso analogue of the immunodominant fragment of VP1 coat protein from foot-and-mouth disease virus. Biopolymers. 1997;41(5):569–90.CrossRefPubMed
44.
45.
Peppicelli S, Bianchini F, Toti A, Laurenzana A, Fibbi G, Calorini L. Extracellular acidity strengthens mesenchymal stem cells to promote melanoma progression. Cell Cycle. 2015;14(19):3088–100.CrossRefPubMedPubMedCentral
46.
Franco P, Vocca I, Carriero MV, Alfano D, Cito L, Longanesi-Cattani I, et al. Activation of urokinase receptor by a novel interaction between the connecting peptide region of urokinase and (alpha)v(beta)5 integrin. J Cell Sci. 2006;19:3424–34.CrossRef
47.
Ingangi V, Bifulco K, Yousif AM, Ragone C, Motti ML, Rea D, et al. The urokinase receptor-derived cyclic peptide [SRSRY] suppresses neovascularization and intravasation of osteosarcoma and chondrosarcoma cells. Oncotarget. 2016;7(34):54474–87.CrossRefPubMedPubMedCentral
48.
Timpson P, McGhee EJ, Erami Z, Nobis M, Quinn JA, Edward M, et al. Organotypic collagen I assay: a malleable platform to assess cell behaviour in a 3-dimensional context. J Vis Exp. 2011;56:e3089.
49.
Margheri F, Luciani C, Taddei ML, Giannoni E, Laurenzana A, Biagioni A, et al. The receptor for urokinase-plasminogen activator (uPAR) controls plasticity of cancer cell movement in mesenchymal and amoeboid migration style. Oncotarget. 2014;5(6):1538–53.CrossRefPubMedPubMedCentral
50.
Høyer-Hansen G, Rønne E, Solberg H, Behrendt N, Ploug M, Lund LR, Ellis V, Danø K. Urokinase plasminogen activator cleaves its cell surface receptor releasing the ligand-binding domain. J Biol Chem. 1992;267:18224–9.PubMed
51.
Huang J, Hu J, Bian X, Chen K, Gong W, Dunlop NM, et al. Transactivation of the epidermal growth factor receptor by formylpeptide receptor exacerbates the malignant behavior of human glioblastoma cells. Cancer Res. 2007;67(12):5906–13.CrossRefPubMed
52.
Dupin E, Calloni GW, Le Douarin NM. The cephalic neural crest of amniote vertebrates is composed of a large majority of precursors endowed with neural, melanocytic, chondrogenic and osteogenic potentialities. Cell Cycle. 2010;9(2):238–49.CrossRefPubMed
53.
Friedl P. Prespecification and plasticity: shifting mechanisms of cell migration. Curr Opin Cell Biol. 2004;16(1):14–23.CrossRefPubMed
54.
55.
Albelda SM, Mette SA, Elder DE, Stewart R, Damjanovich L, Herlyn M, et al. Integrin distribution in malignant melanoma: association of the beta 3 subunit with tumor progression. Cancer Res. 1990;50(20):6757–64.PubMed
56.
Voura EB, Ramjeesingh RA, Montgomery AMP, Siu C-H. Involvement of Integrin αvβ3 and cell adhesion molecule L1 in Transendothelial migration of melanoma cells. Mol Biol Cell. 2001;12(9):2699–710.CrossRefPubMedPubMedCentral
57.
Franco P, Carotenuto A, Marcozzi C, Votta G, Sarno C, Iaccarino I, Brancaccio D, De Vincenzo A, Novellino E, Grieco P, Stoppelli MP. Opposite modulation of cell migration by distinct subregions of urokinase connecting peptide. Chembiochem. 2013;14(7):882–9.CrossRefPubMed
58.
Thoma CR, Zimmermann M, Agarkova I, Kelm JM, Krek W. 3D cell culture systems overcome many of the limitations of traditional two-dimensional (2D) monolayer cell culture systems by mimicking more closely the complex cellular heterogeneity and interactions and tumor microenvironmental conditions. Adv Drug Deliv Rev. 2014;69-70:29–41.CrossRefPubMed
59.
Sanz-Moreno V, Gadea G, Ahn J, Paterson H, Marra P, Pinner S, Sahai E, Marshall CJ. Rac activation and inactivation control plasticity of tumor cell movement. Cell. 2008;135:510–23.CrossRefPubMed
60.
61.
Jo M, Thomas KS, Somlyo AV, Somlyo AP, Gonias SL. Cooperativity between the Ras-ERK and rho-rho kinase pathways in urokinase-type plasminogen activator-stimulated cell migration. J Biol Chem. 2002;277(14):12479–85.CrossRefPubMed
62.
Metadata
Title
Targeting the cross-talk between Urokinase receptor and Formyl peptide receptor type 1 to prevent invasion and trans-endothelial migration of melanoma cells
Authors
Concetta Ragone
Michele Minopoli
Vincenzo Ingangi
Giovanni Botti
Federica Fratangelo
Antonello Pessi
Maria Patrizia Stoppelli
Paolo Antonio Ascierto
Gennaro Ciliberto
Maria Letizia Motti
Maria Vincenza Carriero
Publication date
01-12-2017
Publisher
BioMed Central
Published in
Journal of Experimental & Clinical Cancer Research / Issue 1/2017
Electronic ISSN: 1756-9966
DOI
https://doi.org/10.1186/s13046-017-0650-x

Other articles of this Issue 1/2017

Journal of Experimental & Clinical Cancer Research 1/2017 Go to the issue

Research

Downregulation of long noncoding RNA MEG3 is associated with poor prognosis and promoter hypermethylation in cervical cancer

Research

Chrysin inhibited tumor glycolysis and induced apoptosis in hepatocellular carcinoma by targeting hexokinase-2

Research

MiR-203a-3p suppresses cell proliferation and metastasis through inhibiting LASP1 in nasopharyngeal carcinoma

Research

Alantolactone, a natural sesquiterpene lactone, has potent antitumor activity against glioblastoma by targeting IKKβ kinase activity and interrupting NF-κB/COX-2-mediated signaling cascades

Research

MicroRNA-381 inhibits the metastasis of gastric cancer by targeting TMEM16A expression

Research

CLDN6 promotes chemoresistance through GSTP1 in human breast cancer
Webinar | 19-02-2024 | 17:30 (CET)

Keynote webinar | Spotlight on antibody–drug conjugates in cancer

Watch now

Antibody–drug conjugates (ADCs) are novel agents that have shown promise across multiple tumor types. Explore the current landscape of ADCs in breast and lung cancer with our experts, and gain insights into the mechanism of action, key clinical trials data, existing challenges, and future directions.

Dr. Véronique Diéras
Prof. Fabrice Barlesi
Developed by: Springer Medicine
Image Credits