BBSRC iCase 4 Year PhD Studentship (Lee – Fellows)

Department/Location: Wellcome Trust – Medical Research Council Cambridge Stem Cell Institute

Reference: PS11650

Closing date: 31 March 2017

Collaborative PhD Studentship with Astra Zeneca

Elucidating cellular behaviours and potential oncogenicity utilising in vitro organoid culture system Project Description

Department name: Cambridge Stem Cell Institute, University of Cambridge Supervisors: Primary: Dr Joo-Hyeon Lee (University of Cambridge) and Dr Mick Fellows (AstraZeneca)

Assessment of cell transformation (i.e. the acquisition of malignant characteristics in morphology, growth control or function) has proven problematic to model in vitro. The soft agar cell transformation assay has been validated, but only in a limited number of immortalised or mixed population embryonic cells and the specificity of the assay has been questioned. Furthermore, the genetic basis of cell transformation in this model has not been fully delineated (Harvey et al 2015, Creton et al 2012). The use of genetically and phenotypically stable organoids derived from single human stem cells (Schwank et al 2013) should provide a better model for cell proliferation and transformation.

Organoids have the advantage of 3D morphology, being able to be derived from several major organs and can be engineered to demonstrate proliferative effects by introducing a cell type specific ki67 fluorescent protein to tag. Organoid stability is also dependent on several growth factors and inhibitors e.g. EGF, Wnt, BMP inhibitor, TGFb inhibitor. Growth following withdrawal of these factors, which are known pathways implicated in cancer genesis, will be indicative of cell phenotypic and/or transformative changes. It is also proposed that this model could be used to investigate mechanisms of pro-oncogenicity and also the potential off-target effect of genome editing by CRISPR/Cas9, for which there is currently an unmet need for in vitro assays to assess concerns around inappropriate editing in oncogenes.

The project will develop organoids from a variety of tissue sources, including organoids with the ki67 tag, and assessment of proliferative and morphological changes following withdrawal of growth factors after treatment with reference carcinogens and following transfection with specific and promiscuous guide RNA and associated Cas9 protein. Comparative data will be generated from alternative in vitro methodologies to analyse cell transformation e.g. the soft agar assay. Reference carcinogens will be analysed in this assay along with the potential of CRISPR/Cas9 editing to induce cell transformation (which has not been previously assessed). The student will also use next generation sequencing technologies to identify specific genes involved in morphological changes in both cell and organoid cultures. Success will provide new and more relevant model for understanding and assessment of morphological changes and carcinogenesis.

Eligibility and Funding

This studentship covers 4 years’ UK/EU tuition fees (see below for EU eligibility requirements) and a maintenance stipend.

BBSRC funding is available for UK nationals and EU students who meet the residency requirements. Further information about eligibility for funding can be found on the BBSRC website: http://www.bbsrc.ac.uk/documents/studentship-eligibility-pdf/

How to Apply

Please visit our website to find out more, and about the application process itself: http://study.stemcells.cam.ac.uk/study/otheropportunities/#BBSRC-iCASE-ASTRAZENECA.

Deadline for receipt of applications: midnight on Friday 31st March 2017.

References

Harvey, Howe et al. Mutagenesis. 2005 Jan;20(1):51-6. http://mutage.oxfordjournals.org/content/20/1/51.long

Creton, Aardema et al. Mutagenesis. 2012 Jan;27(1):93-101. http://mutage.oxfordjournals.org/content/27/1/93.long

Schwank, Koo et al. Cell Stem Cell . 2013 Dec 5;13(6):653¿58. http://www.sciencedirect.com/science/article/pii/S1934590913004931

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