Federico Bussolino

Università degli Studi di Torino, Torino, Piedmont, Italy

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Publications (346)1915.96 Total impact

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    ABSTRACT: Neuronal pentraxins (NPTX) and their corresponding receptors (NPTXR) have been studied as synapse-associated proteins in the nervous system, but their role in cancer is largely unknown. By applying a multidisciplinary, high-throughput proteomic approach, we have recently identified a peptide ligand motif for targeted drug delivery to neuroblastoma. Here we report the sequence similarity between this peptide and a conserved portion of the pentraxin domain that is involved in the homo- and hetero-oligomerization of NPTX2 and NPTXR. We show that, in comparison to normal tissues, NPTX2 and NPTXR are overexpressed in vivo in mouse models, as well as in human Schwannian stroma-poor, stage IV neuroblastoma. Both proteins are concentrated in the vicinity of tumor blood vessels, with NPTXR also present on neuroblastic tumor cells. In vivo targeting of NPTX2 and NPTXR with the selected peptide or with specific antibodies reduces tumor burden in orthotopic mouse models of human neuroblastoma. In vitro interference with this ligand/receptor system inhibits the organization of neuroblastoma cells in tumor-like masses in close contact with vascular cells, as well as their adhesion to normal microenvironment-derived cells, suggesting a role in the crosstalk between tumor and normal cells in the early steps of neuroblastoma development. Finally, we show that NPTX2 is a marker of poor prognosis for neuroblastoma patients. Copyright © 2015, American Association for Cancer Research.
    Cancer Research 08/2015; DOI:10.1158/0008-5472.CAN-15-0649 · 9.28 Impact Factor
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    ABSTRACT: Cellular protrusions are highly dynamic structures involved in fundamental processes, including cell migration and invasion. For a cell to migrate, its leading edge must form protrusions, and then adhere or retract. The spatial and temporal coordination of protrusions and retraction is yet to be fully understood. The study of protrusion dynamics mainly relies on live-microscopy often coupled to fluorescent labeling. Here we report the use of an alternative, label-free, quantitative and rapid assay to analyze protrusion dynamics in a cell population based on the real-time recording of cell activity by means of electronic sensors. Cells are seeded on a plate covered with electrodes and their shape changes map into measured impedance variations. Upon growth factor stimulation the impedance increases due to protrusive activity and decreases following retraction. Compared to microscopy-based methods, impedance measurements are suitable to high-throughput studies on different cell lines, growth factors and chemical compounds. We present data indicating that this assay lends itself to dissect the biochemical signaling pathways controlling adhesive protrusions. Indeed, we show that the protrusion phase is sustained by actin polymerization, directly driven by growth factor stimulation. Contraction instead mainly relies on myosin action, pointing at a pivotal role of myosin in lamellipodia retraction.
    Scientific Reports 05/2015; 5:10206. DOI:10.1038/srep10206 · 5.58 Impact Factor
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    ABSTRACT: Aberrant cholesterol homeostasis and biosynthesis has been observed in different tumour types. This paper investigates the role of the post-squalenic enzyme of cholesterol biosynthesis, oxidosqualene cyclase (OSC), in regulating tumour angiogenesis and metastasis dissemination in mouse models of cancer. We showed that Ro 48-8071, a selective inhibitor of OSC, reduced vascular density and increased pericyte coverage, with a consequent inhibition of tumour growth in a spontaneous mouse model of pancreatic tumour (RIP-Tag2) and two metastatic mouse models of human colon carcinoma (HCT116) and pancreatic adenocarcinoma (HPAF-II). Remarkably, the inhibition of OSC hampered metastasis formation in HCT116 and HPAF-II models. Ro 48-8071 induced tumour vessel normalization and enhanced the anti-tumoral and anti-metastatic effects of 5-fluorouracil (5-FU) in HCT116 mice. Ro 48-8071 exerted a strong anti-angiogenic activity by impairing endothelial cell adhesion and migration, and by blocking vessel formation in angiogenesis assays. OSC inhibition specifically interfered with the PI3K pathway. According to in vitro results, Ro 48-8071 specifically inhibited Akt phosphorylation in both cancer cells and tumour vasculature in all treated models. Thus, our results unveil a crucial role of OSC in the regulation of cancer progression and tumour angiogenesis, and indicate Ro 48-8071 as a potential novel anti-angiogenic and anti-metastatic drug.
    Scientific Reports 03/2015; 5(9054):1-12. DOI:10.1038/srep09054 · 5.58 Impact Factor
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    ABSTRACT: Aberrant cholesterol homeostasis and biosynthesis has been observed in different tumour types. This paper investigates the role of the post-squalenic enzyme of cholesterol biosynthesis, oxidosqualene cyclase (OSC), in regulating tumour angiogenesis and metastasis dissemination in mouse models of cancer. We showed that Ro 48-8071, a selective inhibitor of OSC, reduced vascular density and increased pericyte coverage, with a consequent inhibition of tumour growth in a spontaneous mouse model of pancreatic tumour (RIP-Tag2) and two metastatic mouse models of human colon carcinoma (HCT116) and pancreatic adenocarcinoma (HPAF-II). Remarkably, the inhibition of OSC hampered metastasis formation in HCT116 and HPAF-II models. Ro 48-8071 induced tumour vessel normalization and enhanced the anti-tumoral and anti-metastatic effects of 5-fluorouracil (5-FU) in HCT116 mice. Ro 48-8071 exerted a strong anti-angiogenic activity by impairing endothelial cell adhesion and migration, and by blocking vessel formation in angiogenesis assays. OSC inhibition specifically interfered with the PI3K pathway. According to in vitro results, Ro 48-8071 specifically inhibited Akt phosphorylation in both cancer cells and tumour vasculature in all treated models. Thus, our results unveil a crucial role of OSC in the regulation of cancer progression and tumour angiogenesis, and indicate Ro 48-8071 as a potential novel anti-angiogenic and anti-metastatic drug
    Scientific Reports 03/2015; 5(9054):1-12. · 5.58 Impact Factor
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    ABSTRACT: The need for decentralized clinical tests together with the concept of time and cost saving are pushing the development of portable, miniaturized, compact biosensors with diagnostic and prognostic purpose. Here, we propose an innovative detection system based on a Single Photon Avalanche Diode (SPAD) with high sensitivity and low noise, crucial features for an efficient chemiluminescence biosensor. The SPAD detector, having 60µm diameter, has a Photon Detection Efficiency higher than 55% at 425nm and a Dark Count Rate lower than 100Hz at room temperature. Our design allows a good optical coupling efficiency between sample and detector. A specific biofunctional surface was implemented taking advantage of aptamers, short DNA sequences having high selectivity and affinity toward their targets. We successfully detected physiological levels of Vascular Endothelial Growth Factor (VEGF), a circulating protein biomarker highly correlated with cancer. The SPAD aptasensor showed a Limit of Detection (LoD) in the pM range, stability (up to 42 days) and re-usability (up to seven cycles). This compact biosensor is therefore a promising step toward the actual use of portable microdevices in diagnostics. Copyright © 2015 Elsevier B.V. All rights reserved.
    Biosensors & Bioelectronics 01/2015; 68C:500-507. DOI:10.1016/j.bios.2015.01.042 · 6.45 Impact Factor
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    ABSTRACT: Non-amoeboid cell migration is characterised by dynamic competition among multiple protrusions to establish new adhesion sites at the cell's leading edge. However, the mechanisms that regulate the decision to disassemble or to grow nascent adhesions are not fully understood. Here we show that in endothelial cells (EC) 3-phosphoinositide-dependent protein (PDK1) promotes focal adhesions (FA) turnover by controlling endocytosis of integrin αvβ3 in a PI3K-dependent manner. We demonstrate that PDK1 binds and phosphorylates integrin αvβ3. Down-regulation of PDK1 increases FA size and slows down their disassembly. This process requires both PDK1 kinase activity and PI3K activation but does not involve Akt. Moreover, PDK1 silencing stabilizes FA in membrane protrusions decreasing EC migration on vitronectin. These results indicate that modulation of integrin endocytosis by PDK1 hampers EC adhesion and migration on extracellular matrix, thus unveiling a novel role for this kinase.
    Journal of Cell Science 01/2015; DOI:10.1242/jcs.149294 · 5.33 Impact Factor
  • Laura di Blasio · Federico Bussolino · Luca Primo
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    ABSTRACT: In vitro assays with endothelial cells (EC) cultured on three-dimensional gel recapitulate several aspects of vascular morphogenesis and pathological angiogenesis. The two most used in vitro assays of vascular morphogenesis are the tube formation on extracellular matrix gel and the sprouting from EC spheroids. Tube formation assay measures the ability of EC, plated on gel derived from reconstituted basement membrane, to form capillary-like structures. Sprouting assay is based on spheroids of EC, embedded in collagen gel and stimulated with angiogenic factors, which originate a complex network of capillary-like structures invading the gel. Both these assays can be exploited for antiangiogenic drug screening and gene function analysis during vascular morphogenesis.
    Methods in molecular biology (Clifton, N.J.) 01/2015; 1214:41-7. DOI:10.1007/978-1-4939-1462-3_4 · 1.29 Impact Factor
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    ABSTRACT: The introduction of biodrugs, e.g. the anti-EGFR antibodies, was initially seen as a promise to radically change the landscape for patients with metastatic colorectal cancer. However, although EGFR-targeted therapies, combined with chemotherapy, have prolonged survival expectancy to 24 months, cure remains anecdotal. Target therapies suffer of high costs, important side effects, and low response rates: it is now clear that this approach as it was originally conceived failed to meet the majority of its expectations. Importantly, because of novel prescription guidelines, patients with KRAS-mutated tumors are excluded from EGFR-targeted therapies: for these patients alternative treatments are urgently needed. We here propose an innovative strategy based on the identification of molecular targets specifically associated with the microenvironment of metastatic colorectal cancer in patients carrying oncogenic KRAS. For this purpose, we set up mice models of metastatic colorectal cancer by intrasplenic (to evaluate liver homing) and intrahepatic (to investigate liver colonization) implantation of human colorectal cancer cell lines (SW48 and LIM1215) in which oncogenic KRAS (G12D, G12V, G13D) variants have been somatically knocked-in. We based our “target fishing” strategy on high-throughput, phage display-mediated proteomic screenings of deriving tumor samples ex vivo. Proteome signatures from the cognate cell lines in vitro served as a subtractive reference for the ex vivo biopanning, aimed at the identification of peptide ligands specific for non-epithelial components. By this combined biological-genetic-bioinformatics approach, we identified peptide/protein signatures selectively associated to metastasis microenvironments with controlled genetic backgrounds in vitro and in vivo. We selected 2 target proteins and 2 targeting peptides, which were exploited for diagnostic and therapeutic purposes. First, we evaluated by IHC the presence and localization of the target proteins in samples (tumor, healthy liver) from a panel of human biopsies and from the described in vivo models. This analysis confirmed a specific enrichment in KRAS-mutated microenvironments. Second, we tested the capability of dye-conjugated targeting peptides to home to these same tumor microenvironments, observing a specific accumulation in target tissues, compared to their scrambled versions and to control tissues. Third, we investigated a potential anti-metastatic effect of these peptides when orthotopically co-injected with colorectal cancer cell lines; preliminary experiments revealed that the targeting peptides, but not the scrambled variants, inhibit the onset of liver metastases from KRAS-mutated cell lines. In summary, we obtained proof-of-concept results in preclinical studies and produced prototype compounds to provide innovative tools that can be translated into the clinical practice with a mid-term perspective.
    Cancer Research 10/2014; 74(19 Supplement):1163-1163. DOI:10.1158/1538-7445.AM2014-1163 · 9.28 Impact Factor
  • Cancer Research 10/2014; 74(19 Supplement):4807-4807. DOI:10.1158/1538-7445.AM2014-4807 · 9.28 Impact Factor
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    ABSTRACT: The mechanism by which angiogenic endothelial cells break the physical barrier of the vascular basement membrane and consequently sprout to form new vessels in mature tissues is unclear. Here, we show that the angiogenic endothelium is characterized by the presence of functional podosome rosettes. These extracellular-matrix-degrading and adhesive structures are precursors of de novo branching points and represent a key feature in the formation of new blood vessels. VEGF-A stimulation induces the formation of endothelial podosome rosettes by upregulating integrin α6β1. In contrast, the binding of α6β1 integrin to the laminin of the vascular basement membrane impairs the formation of podosome rosettes by restricting α6β1 integrin to focal adhesions and hampering its translocation to podosomes. Using an ex vivo sprouting angiogenesis assay, transgenic and knockout mouse models and human tumour sample analysis, we provide evidence that endothelial podosome rosettes control blood vessel branching and are critical regulators of pathological angiogenesis.
    Nature Cell Biology 09/2014; DOI:10.1038/ncb3036 · 20.06 Impact Factor
  • Journal of Biological Chemistry 09/2014; 289(37):25475-25475. DOI:10.1074/jbc.A113.530972 · 4.57 Impact Factor
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    Davide Corà · Elena Astanina · Enrico Giraudo · Federico Bussolino
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    ABSTRACT: During organogenesis, patterning is primarily achieved by the combined actions of morphogens. Among these, semaphorins represent a general system for establishing the appropriate wiring architecture of biological nets. Originally discovered as evolutionarily conserved steering molecules for developing axons, subsequent studies on semaphorins expanded their functions to the cardiovascular and immune systems. Semaphorins participate in cardiac organogenesis and control physiological vasculogenesis and angiogenesis, which result from a balance between pro- and anti-angiogenic signals. These signals are altered in several diseases. In this review, we discuss the role of semaphorins in vascular biology, emphasizing the mechanisms by which these molecules control vascular patterning and lymphangiogenesis, as well as in genetically inherited and degenerative vascular diseases.
    Trends in Molecular Medicine 08/2014; 10(2):1-10. DOI:10.1016/j.molmed.2014.07.005 · 10.11 Impact Factor
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    ABSTRACT: Directional cell migration is of paramount importance in both physiological and pathological processes, such as development, wound healing, immune response, and cancer invasion. Here, we report that 3-phosphoinositide-dependent kinase 1 (PDK1) regulates epithelial directional migration and invasion by binding and activating myotonic dystrophy kinase-related CDC42-binding kinase α (MRCKα). We show that the effect of PDK1 on cell migration does not involve its kinase activity but instead relies on its ability to bind membrane phosphatidylinositol (3,4,5)-trisphosphate. Upon epidermal growth factor (EGF) stimulation, PDK1 and MRCKα colocalize at the cell membrane in lamellipodia. We demonstrate that PDK1 positively modulates MRCKα activity and drives its localization within lamellipodia. Likewise, the retraction phase of lamellipodia is controlled by PDK1 through an MRCKα-dependent mechanism. In summary, we discovered a functional pathway involving PDK1-mediated activation of MRCKα, which links EGF signaling to myosin contraction and directional migration.
    The Journal of Cell Biology 08/2014; 206(3):415-34. DOI:10.1083/jcb.201312090 · 9.69 Impact Factor
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    ABSTRACT: Angiopoietin-like (ANGPTL) proteins are secreted proteins showing structural similarity to members of the angiopoietin family. Some ANGPTL proteins possess pleiotropic activities, being involved in cancer lipid, glucose energy metabolisms, and angiogenesis. ANGPTL7 is the less characterized member of the family whose functional role is only marginally known. In this study, we provide experimental evidences that ANGPTL7 is over-expressed in different human cancers. To understand the role played by ANGPTL7 in tumor biology, we asked whether ANGPTL7 is endogenously expressed by malignant cells or in response to environmental stimuli. We found that ANGPTL7 is marginally expressed under standard growth condition while it is specifically up-regulated by hypoxia. Interestingly, the protein is secreted and partially associated with the exosomal fraction, suggesting that it could be found in the systemic circulation of oncologic patients and act in an endocrine way. Moreover, we found that ANGPTL7 exerts a pro-angiogenetic effect on human differentiated endothelial cells by stimulating their proliferation, motility, invasiveness, and capability to form capillary-like networks while it does not stimulate progenitor endothelial cells. Finally, we showed that ANGPTL7 promotes vascularization in vivo in the mouse Matrigel sponge assay, thereby accrediting this molecule as a pro-angiogenic factor.
    Angiogenesis 06/2014; 17(4). DOI:10.1007/s10456-014-9435-4 · 4.41 Impact Factor
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    ABSTRACT: The synaptic protein Neuroligin 1 (NLGN1), a cell adhesion molecule, is critical for the formation and consolidation of synaptic connectivity and is involved in vascular development. The mechanism through which NLGN1 acts, especially in vascular cells, is unknown. Here, we aimed at deepening our knowledge on the cellular activities and molecular pathways exploited by endothelial NLGN1 both in vitro and in vivo. We analyzed the phenotypic consequences of NLGN1 expression modulation in endothelial cells through in vitro angiogenesis assays and the mouse postnatal retinal angiogenesis model. We demonstrate that NLGN1, whereas not affecting endothelial cell proliferation or migration, modulates cell adhesion to the vessel stabilizing protein laminin through cooperation with the α6 integrin, a specific laminin receptor. Finally, we show that in vivo, NLGN1 and α6 integrin preferentially colocalize in the mature retinal vessels, whereas NLGN1 deletion causes an aberrant VE-cadherin, laminin and α6 integrin distribution in vessels, along with significant structural defects in the vascular tree.
    Journal of Biological Chemistry 05/2014; DOI:10.1074/jbc.M113.530972 · 4.57 Impact Factor
  • Molecular Cancer Therapeutics 01/2014; 12(11_Supplement):A164-A164. DOI:10.1158/1535-7163.TARG-13-A164 · 6.11 Impact Factor
  • Federico Bussolino · Enrico Giraudo · Guido Serini
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    ABSTRACT: Semaphorins were originally identified as axon guidance molecules involved in the development of the neuronal system. However, accumulating evidences have clearly demonstrated that the semaphorin system is not restricted to the brain but supports functions of other organs. Here, we review the rapidly emerging functions of sempahorins and, in particular class 3 semaphorin, in vascular and lymphatic systems during the development, tumor angiogenesis and ischemic revascularization.
    Clinical Reviews in Allergy & Immunology 01/2014; 99:71-88. DOI:10.1159/000353315 · 4.73 Impact Factor
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    ABSTRACT: Supplementary Figure for "Diffusion limited phase separation in eukaryotic chemotaxis", P.N.A.S. 2005, 102, 16927
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    ABSTRACT: Supplementary Text for "Diffusion limited phase separation in eukaryotic chemotaxis", P.N.A.S. 2005, 102, 16927

Publication Stats

15k Citations
1,915.96 Total Impact Points

Institutions

  • 1982–2015
    • Università degli Studi di Torino
      • • Dipartimento di Scienza e Tecnologia del Farmaco
      • • Department of Medical Science
      • • Dipartimento di Scienze Cliniche e Biologiche
      Torino, Piedmont, Italy
  • 2014
    • Cancer Research Institute
      New York, New York, United States
  • 2013
    • University of Toronto
      Toronto, Ontario, Canada
  • 2012
    • Bioindustry Park Silvano Fumero
      Torino, Piedmont, Italy
  • 1999–2012
    • Institute for Cancer Research and Treatment
      Torino, Piedmont, Italy
    • Università degli Studi di Siena
      Siena, Tuscany, Italy
  • 2007
    • Max Planck Institute of Biochemistry
      München, Bavaria, Germany
    • University of Naples Federico II
      • Department of Physical Sciences
      Napoli, Campania, Italy
    • University of Milan
      Milano, Lombardy, Italy
  • 2005
    • San Raffaele Scientific Institute
      Milano, Lombardy, Italy
  • 2002
    • Istituto Superiore di Sanità
      • Laboratory of Virology
      Roma, Latium, Italy
    • Università degli Studi di Bari Aldo Moro
      • Dipartimento di Scienze Biomediche ed Oncologia Umana (DIMO)
      Bari, Apulia, Italy
  • 2000
    • University of Amsterdam
      Amsterdamo, North Holland, Netherlands
  • 1987–1999
    • Mario Negri Institute for Pharmacological Research
      • Laboratory: Vascular Biology
      Milano, Lombardy, Italy
    • University at Buffalo, The State University of New York
      • Department of Medicine
      Buffalo, NY, United States
  • 1997
    • Università degli Studi di Brescia
      • Department of Clinical and Experimental Sciences
      Brescia, Lombardy, Italy
  • 1995–1997
    • University of Pavia
      Ticinum, Lombardy, Italy
    • Universität Erfurt
      Erfurt, Thuringia, Germany
  • 1996
    • University of Buenos Aires
      • Biological Chemistry Department
      Buenos Aires, Buenos Aires F.D., Argentina
  • 1987–1993
    • Polo d'Innovazione di Genomica Genetica e Biologia
      Perugia, Umbria, Italy
  • 1988
    • Università degli Studi del Sannio
      Benevento, Campania, Italy
  • 1987–1988
    • Eawag: Das Wasserforschungs-Institut des ETH-Bereichs
      Duebendorf, Zurich, Switzerland