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Tumour cell regulation of platelets and thrombosis

Summary of project:

Metastasis is the cause of about 90% of cancer associated deaths, yet the mechanisms governing this clinically important process remain poorly understood. The involvement of blood platelets in tumour dissemination has been recognized since 1865. In addition, cancer patients are at an elevated risk of suffering from thrombotic episodes- a big killer in the western world. The ability of cancer cells to activate platelets is well documented. In turn, platelets aid tumour cells to survive in transit in the blood, as well as aiding in tumour attachment and growth at new sites round the body. The mechanisms by which cancer cells activate platelets to help in their dissemination remain unclear.

Cancer cells express and release mucins, these extensively modified glycoproteins have protective and signalling roles. Many reports have shown that cancer cells expressing large amounts of mucins have a poor prognosis, because of a high rate of metastasis. The reason behind the increased metastatic potential of these cells remains unknown. However, cancer cell derived mucins rapidly induce platelet thrombus formation in vivo in a P-selectin dependent manner. Platelets, once activated, aid in tumour cell induced migration and invasion, both key steps in metastasis.

The aims of this study are to determine the molecular mechanisms by which mucins activate platelets, and to study how mucin-activated platelets may aid in tumour cell adhesion, migration and proliferation. The project will be divided into three main stages.

  1. The student will assess the ability of mucins purified from adenocarcinoma lines to activate platelets. The dose and time dependent ability of mucins to induce platelet aggregation will be determined. It is possible that mucins do not directly activate platelets, but sensitise them to activation by other agonists. The dose dependency for mucins to augment thrombin induced aggregation will be investigated. Platelets will be incubated with sub threshold concentrations of thrombin, which only induce a weak aggregation response. It will be determined if co-incubating with increasing mucins concentrations can enhance the observed aggregation. In addition, the ability of mucins to initiate platelet adhesion and spreading on fibrinogen will be determined by fluorescent microscopy. Adhesion, spreading and aggregation on fibrinogen require the activation of the platelet integrin receptor αIIbβ3.To confirm that mucins can activate integrin αIIbβ3 on adherent platelets, we will use the antibody PAC-1. We have recently developed a range of antagonistic compounds to αIIbβ3, the ability of these compounds to block mucin induced platelet aggregation and spreading will be determined.
  2. The signalling induced by mucins to activate platelets will be investigated by the student. Platelet activation is largely driven by Src kinases, MAP kinases and Rho GTPases. The ability of cancer derived mucins to activate these pathways will be investigated by Western blotting and Rho GTPase assays. If these pathways are activated, chemical inhibitors targeted against these molecules will be used to assess their function in mucin-mediated platelet adhesion and aggregation.
  3. The ability of mucins to up-regulate platelet-cancer interactions will be investigated. A range of cancer lines which we have shown to express integrins will be used. The student will determine if exogenously added mucins stimulate adhesion of platelets to these cancer cells. The ability of these interactions to then stimulate cancer cell migration and invasion will be investigated by the standard scratch assay and through using boyden invasion chambers. In addition, the ability of platelets to stimulate migration of cancer cell lines containing mucins will be compared to cancer cells lines lacking mucins. This will identify the importance of locally produced endogenous mucins.

Entry requirements:

At least 2.i Honours degree or equivalent.

Supervisors:

Title and name:
Dr Wayne Roberts
Position:
Lecturer in Medical Sciences
Email address:
Telephone number :
Work+44 (0) 1274 232131
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Title and name:
Dr Helen Sheldrake
Position:
Lecturer in Organic and Medicinal Chemistry
Email address:
Telephone number :
Work+44 (0) 1274 236858
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Title and name:
Dr Steve Shnyder
Position:
Senior Lecturer in Cancer Biology
Email address:
Telephone number :
Work+44 (0) 1274 235898
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