title: N-terminus-modified Hec1 suppresses tumour growth by interfering with kinetochore-microtubule dynamics.
abstract: Mitotic proteins are attractive targets to develop molecular cancer therapeutics due to the intimate interdependence between cell proliferation and mitosis. In this work, we have explored the therapeutic potential of the kinetochore (KT) protein Hec1 (Highly Expressed in Cancer protein 1) as a molecular target to produce massive chromosome missegregation and cell death in cancer cells. Hec1 is a constituent of the Ndc80 complex, which mediates KT-microtubule (MT) attachments at mitosis and is upregulated in various cancer types. We expressed Hec1 fused with enhanced green fluorescent protein (EGFP) at its N-terminus MT-interaction domain in HeLa cells and showed that expression of this modified Hec1, which localized at KTs, blocked cell proliferation and promoted apoptosis in tumour cells. EGFP-Hec1 was extremely potent in tumour cell killing and more efficient than siRNA-induced Hec1 depletion. In striking contrast, normal cells showed no apparent cell proliferation defects or cell death following EGFP-Hec1 expression. Live-cell imaging demonstrated that cancer cell death was associated with massive chromosome missegregation within multipolar spindles after a prolonged mitotic arrest. Moreover, EGFP-Hec1 expression was found to increase KT-MT attachment stability, providing a molecular explanation for the abnormal spindle architecture and the cytotoxic activity of this modified protein. Consistent with cell culture data, EGFP-Hec1 expression was found to strongly inhibit tumour growth in a mouse xenograft model by disrupting mitosis and inducing multipolar spindles. Taken together, these findings demonstrate that stimulation of massive chromosome segregation defects can be used as an anti-cancer strategy through the activation of mitotic catastrophe after a multipolar mitosis. Importantly, this study represents a clear proof of concept that targeting KT proteins required for proper KT-MT attachment dynamics constitutes a powerful approach in cancer therapy.
authors.affiliation: Institute of Biology, Molecular Medicine and Nanobiotechnology, CNR National Research Council, Rome, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Orticello
authors.affiliation: Institute of Biology, Molecular Medicine and Nanobiotechnology, CNR National Research Council, Rome, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Fiore
authors.affiliation: Institute of Biology, Molecular Medicine and Nanobiotechnology, CNR National Research Council, Rome, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Totta
authors.affiliation: Experimental Chemotherapy Laboratory, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Desideri
authors.affiliation: Chromosome Instability and Dynamics Laboratory, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal.
authors.forename: M
authors.initials: M
authors.lastname: Barisic
authors.affiliation: Anatomic Pathology Institute, Tor Vergata University, Rome, Italy.
authors.forename: D
authors.initials: D
authors.lastname: Passeri
authors.affiliation: Department of Biology and Biotechnology 'Charles Darwin', Sapienza University, Rome, Italy.
authors.forename: J
authors.initials: J
authors.lastname: Lenzi
authors.affiliation: Department of Biology and Biotechnology 'Charles Darwin', Sapienza University, Rome, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Rosa
authors.affiliation: Anatomic Pathology Institute, Tor Vergata University, Rome, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Orlandi
authors.affiliation: 1] Chromosome Instability and Dynamics Laboratory, Instituto de Biologia Molecular e Celular, Universidade do Porto, Porto, Portugal [2] Cell Division Unit, Department of Experimental Biology, Faculdade de Medicina, Universidade do Porto, Porto, Portugal.
authors.forename: H
authors.initials: H
authors.lastname: Maiato
authors.affiliation: Experimental Chemotherapy Laboratory, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: D
authors.initials: D
authors.lastname: Del Bufalo
authors.affiliation: Institute of Biology, Molecular Medicine and Nanobiotechnology, CNR National Research Council, Rome, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Degrassi
mesh_terms: D000818:Animals; D017209:Apoptosis; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic; D003598:Cytoskeletal Proteins; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism*; D008297:Male; D051379:Mice; D008870:Microtubules / Q000378:metabolism*; D008938:Mitosis; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D064047:Spindle Poles / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D003598:Cytoskeletal Proteins; C120529:NDC80 protein, human; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D025521:Tumor Suppressor Proteins