Senior Lecturer Transfusion Medicine

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Research themes:

Megakaryocyte and platelet biology, In vitro production of cellular components for transfusion in humans

Description of research

My group is based at the Cambridge Blood Centre, NHS Blood and Transplant (NHSBT) and its programme of research is based on platelet and megakaryocyte biology. It focuses on understanding the key biological players in platelet production all the way from haemopoietic stem cells (hSCs) or pluripotent stem cells, through the process of megakaryopoiesis and proplatelet formation, and finally platelets themselves. The group has a keen interest in translating biological discoveries into applications for transfusion medicine and uses a multidisciplinary approach that encompasses cell biology, engineering, computational biology and population genetics, the latter in close collaboration with the group of Prof Ouwehand.

1. Megakaryopoiesis and platelet production in vitro

Post-doc: Dr Meera Arumugam

This project which is funded by the NIHR looks at the feasibility of producing platelet in an in vitro system that would ultimately be applicable to transfusion into patients. The approach combines the use of bioreactors and specialized matrices with protein biology and nanotechnology to deliver growth signals to the megakaryocytes (MKs) and optimize platelet production in a controlled system.

2. New stem cell resources for platelet production

Post-doc: Dr Thomas Moreau

The aim of this project, also funded through the NIHR, is to create a bank of pluripotent stem cells (iPSCs) and a method to differentiate these cells into MKs that is applicable to transfusion medicine in patients. The approach is based on the integration of several projects: 1. Functional annotation of the genome in primary human megakaryocytes in collaboration with Prof Bertie Göttgens’ group to establish the network of transcription factors in megakaryopoiesis; 2. Discovery of new genetic mutations in pedigrees affected by platelet disorders in order to identify key players in platelet production; 3. Population studies in healthy individuals to identify regulators of platelet volume and count (work carried out by Prof Ouwehand’s group).

3. Model organism of platelet disorders:

a. Mouse model of platelet disorders:

Jak2 V617F model. This project is funded by the British Heart Foundation. The JAK2 V617F mutation is present in 50% of patients with Essential Thrombocythaemia and the clinical hallmark in this population is of an increased platelet count and risk of cardiovascular events. This project is based on the study of a knock-in model produced by Prof Anthony Green’s group and aims at identifying how the mutant JAK2 leads to increase platelet production and alters platelet function through a range of in vivo and cell biology assays.

Other mouse models of megakaryocyte/platelet disorders are currently under investigations stemming from either gene knock-down studies (based at the Wellcome Trust Sanger Institute, WTSI) or from targeting genes identified in studies of pedigrees with inherited platelet disorders.

b. iPSC modeling of MK disorder

The efficient megakaryocyte production from iPSCs allows us to study in detail the biological role of hitherto unknown proteins identified from genetic studies in patients with inherited platelet disorders. We derive iPSC cells from the patient and use the iPSC-derived megakaryocytes as a platform for cell-based assays. Furthermore new molecular techniques developed in collaboration with the WTSI allow us to place a tag specifically at the end of proteins of interest within the iPSC cell lines and using MKs derived from the cell lines identify the binding partner and cellular localization of these proteins.

4. Development of new therapeutics for platelet alloimmune disorders.

Co-ordinator: Dr Cedric Ghevaert

This project funded by the NHSBT, looks at developing new diagnostic and therapeutic approaches in the context of fetomaternal alloimmune thrombocytopenia. This disorder affects 1/1200 pregnancies in the UK and leads in 10-20% of fetus/neonates affected to severe brain haemorrhage. We have developed a recombinant blocking antibody that is currently being tested in human studies that would potentially be a much safer and effective alternative therapy to the current use of intra-uterine transfusion or immunomodulation.

5. Influence of donor platelet phenotype and platelet concentrate in clinical transfusion in neonates

Up to 75% of neonates on intensive care units will receive platelet transfusion support. Bleeding in these patients tends to often manifest itself as intracranial bleed with potentially devastating neurological consequences. As shown by clinical trials (including the PlaNeT study run by the NHSBT) risk of bleeding does not necessarily correlate with platelet count and therefore the correct transfusion support for neonates is difficult to assess. In collaboration with the Neonatal Intensive Care Unit at Addenbrookes, we are developing new techniques based on flow cytometry technology to be able to assess platelet function more accurately, in particular in the group of premature newborns where the risk of bleeding is markedly increased, to be able to more effectively deliver transfusion support in a bid to reduce the risk of bleeding.

Research focus

Keywords: Platelet transfusion, Megakaryocyte, Pluripotent stem cells

Clinical conditions: Inherited Platelet Disorders

Methodologies: In vitro culture and differentiation by forward programming Bioengineering Flow cytometry Protein signalling Molecular biology and genetic engineering Platelet survival studies

Cambridge collaborators

Department of Material Sciences

Neonatal Intensive Care Unit, Addenbrookes Hospital

Department of Nuclear Medicine, Addenbrookes Hospital

Other collaborators

NHS Blood and Transplant

Wellcome Trust Sanger Institute

Key Publications

Low-affinity FcγR interactions can decide the fate of novel human IgG-sensitised red blood cells and platelets. Armour KL, Smith CS, Turner CP, Kirton CM, Wilkes AM, Hadley AG, Ghevaert C, Williamson LM, Clark MR. Eur J Immunol. 2014 Mar;44(3):905-14. doi: 10.1002/eji.201343825. Epub 2014 Feb 16.

JAK2V617F leads to intrinsic changes in platelet formation and reactivity in a knock-in mouse model of essential thrombocythemia. Hobbs CM, Manning H, Bennett C, Vasquez L, Severin S, Brain L, Mazharian A, Guerrero JA, Li J, Soranzo N, Green AR, Watson SP, Ghevaert C. Blood. 2013 Nov 28;122(23):3787-97. doi: 10.1182/blood-2013-06-501452. Epub 2013 Oct 1.

Megakaryopoiesis through the ages: from the twinkle in the eye to the fully grown adult. Ghevaert C. J Thromb Haemost. 2013 Sep;11(9):1727-9. doi: 10.1111/jth.12349.

Recombinant HPA-1a antibody therapy for treatment of fetomaternal alloimmune thrombocytopenia: proof of principle in human volunteers. Ghevaert C, Herbert N, Hawkins L, Grehan N, Cookson P, Garner SF, Crisp-Hihn A, Lloyd-Evans P, Evans A, Balan K, Ouwehand WH, Armour KL, Clark MR, Williamson LM. Blood. 2013 Jul 18;122(3):313-20. doi: 10.1182/blood-2013-02-481887. Epub 2013 May 8.

New insights into the genetic basis of TAR (thrombocytopenia-absent radii) syndrome. Albers CA, Newbury-Ecob R, Ouwehand WH, Ghevaert C. Curr Opin Genet Dev. 2013 Jun;23(3):316-23. doi: 10.1016/j.gde.2013.02.015. Epub 2013 Apr 17. Review.

Transcription factors in late megakaryopoiesis and related platelet disorders. Tijssen MR, Ghevaert C. J Thromb Haemost. 2013 Apr;11(4):593-604. doi: 10.1111/jth.12131. Review.

Compound inheritance of a low-frequency regulatory SNP and a rare null mutation in exon-junction complex subunit RBM8A causes TAR syndrome. Albers CA, Paul DS, Schulze H, Freson K, Stephens JC, Smethurst PA, Jolley JD, Cvejic A, Kostadima M, Bertone P, Breuning MH, Debili N, Deloukas P, Favier R, Fiedler J, Hobbs CM, Huang N, Hurles ME, Kiddle G, Krapels I, Nurden P, Ruivenkamp CA, Sambrook JG, Smith K, Stemple DL, Strauss G, Thys C, van Geet C, Newbury-Ecob R, Ouwehand WH, Ghevaert C. Nat Genet. 2012 Feb 26;44(4):435-9, S1-2. doi: 10.1038/ng.1083

Developing recombinant HPA-1a-specific antibodies with abrogated Fcgamma receptor binding for the treatment of fetomaternal alloimmune thrombocytopenia. Ghevaert C, Wilcox DA, Fang J, Armour KL, Clark MR, Ouwehand WH, Williamson LM. J Clin Invest. 2008 Aug;118(8):2929-38.

Dasatinib enhances megakaryocyte differentiation but inhibits platelet formation. Mazharian A, Ghevaert C, Zhang L, Massberg S, Watson SP. Blood. 2011 May 12;117(19):5198-206. Epub 2011 Mar 8.

A nonsynonymous SNP in the ITGB3 gene disrupts the conserved membrane-proximal cytoplasmic salt bridge in the alphaIIbbeta3 integrin and cosegregates dominantly with abnormal proplatelet formation and macrothrombocytopenia. Ghevaert C, Salsmann A, Watkins NA, Schaffner-Reckinger E, Rankin A, Garner SF, Stephens J, Smith GA, Debili N, Vainchenker W, de Groot PG, Huntington JA, Laffan M, Kieffer N, Ouwehand WH. Blood. 2008 Apr 1;111(7):3407-14. Epub 2007 Dec 7