Postgrad LIVE! Study Fairs

Southampton | Bristol

European Molecular Biology Laboratory (Heidelberg) Featured PhD Programmes
University of Huddersfield Featured PhD Programmes
Birkbeck, University of London Featured PhD Programmes
University of Portsmouth Featured PhD Programmes
Karlsruhe Institute of Technology Featured PhD Programmes

Targeting extracellular vesicle-mediated intercellular communication in cancer through the use of 3D tissue engineered models

  • Full or part time
  • Application Deadline
    Thursday, February 28, 2019
  • Competition Funded PhD Project (European/UK Students Only)
    Competition Funded PhD Project (European/UK Students Only)

Project Description

Extracellular vesicles (EVs) are nanoscale sized, membrane-enclosed particles that are produced by all cells of the body. Cancer cells release more EVs, with an altered cargo, compared to normal cells. The primary supervisor has recently shown that EVs from head and neck cancer cells contain a specific RNA signature indicative of the parent cell[1]. Cancer-derived EVs recruit normal cells surrounding a tumour by transferring molecular cargo (including nucleic acids) to recipient cells, which converts them to a pro-tumorigenic phenotype. Hence, EVs are an attractive target for novel therapeutics. The majority of in vitro EV research is achieved by isolating EVs from cells cultured as 2D monolayers and then treatment of a recipient cell type in 2D culture. However, this does not represent the in vivo situation where cells exist in a 3D environment. Here we will use 3D tissue engineered models of head and neck cancer to test the hypothesis that ’targeted reduction of extracellular vesicle release will inhibit cancer progression’. The hypothesis will be tested by addressing the following research aims:

Aim 1: Establishment of a fluorescent-reporter 3D cancer model
3D tissue engineered in vitro cancer models will be created using protocols established by the secondary supervisor[2]. The resulting models will replicate carcinoma in situ surrounded by normal epithelium, with normal stromal cells (such as fibroblasts) in the underlying connective tissue. The cancer cells will stably express CD63-RFP that will label the EV membrane with red fluorescent protein. The cancer cells will also stably express an RNA molecule (highly enriched in EVs) fused with a green fluorescent tag. Both fluorescent constructs are already available in the lab of the primary supervisor. The resulting models will be sectioned and analysed by immunohistochemistry to ensure normal tissue architecture in the reporter model and transmission electron microscopy to visualise cellular ultrastructure (including EV precursors). Confocal fluorescence microscopy will be used to visualise the uptake of fluorescent cancer EVs by adjacent and distant normal cells (epithelial cells and fibroblasts).

Aim 2: Reduction of EV release by chemical inhibitors
The models generated in Aim 1 will be treated with chemical inhibitors: Amiloride (a drug used to treat high blood pressure) and GW4869 (a neutral sphingomyelinase inhibitor) reduce EV release in vitro. Models will be analysed as above to determine if treatments have reduced EV release and therefore transmission of fluorescent markers to neighbouring cells. Immunohistochemistry will be used to examine the phenotype of neighbouring cells (proliferation, activation etc). The depth of cancer cell invasion will also be measured to determine if inhibiting EV release reduces cancer progression.


Aim 3: Perturbing EV release by CRISPR/Cas9 gene editing
We have data showing that there is widespread dysregulation of EV biogenesis machinery in head and neck cancer. CRISPR/Cas9 gene editing will be utilised (in collaboration with the industrial supervisor at AstraZeneca[3]) to reduce EV release in the fluorescent reporter cancer cell line. Models will be created with the EV deficient cell line and analysed as above to determine if EVs are required for cancer progression.

Enquiries:
Interested candidates should in the first instance contact Dr Stuart Hunt ()

How to Apply:
Please complete a University Postgraduate Research Application form available here: http://www.shef.ac.uk/postgraduate/research/apply

Please clearly state the prospective main supervisor in the respective box and select Dentistry as the department.

Interviews are due to take place on Monday 25th March 2019.

Funding Notes

The Faculty of Medicine, Dentistry and Health has received an allocation of three EPSRC studentships for 2019 entry from the Doctoral Training Partnership grant that is awarded to the University of Sheffield to fund PhD studentships in the EPSRC remit. These studentships will be 42 months in duration, and include home fee, stipend at RCUK rates and a research training support grant (RTSG) of £4,500.

Home/EU students must have spent the 3 years immediately preceding the start of their course in the UK to receive the full funding.

References

1. Peacock B, Rigby A, Bradford J, Pink R, Hunter K, Lambert D and Hunt S (2018). Extracellular vesicle microRNA cargo is correlated with HPV status in oropharyngeal carcinoma. Journal of Oral Pathology and Medicine: 47(10): 954-963.
2. Colley HE, Hearnden V, Jones AV, Weinreb PH, Violette SM, Macneil S, Thornhill MH and Murdoch C (2011). Development of tissue-engineered models of oral dysplasia and early invasive oral squamous cell carcinoma. British Journal of Cancer: 105(10):1582-92.
3. Heath N, Grant L, De Oliveira TM, Rowlinson R, Osteikoetxea X, Dekker N and Overman R (2018). Rapid isolation and enrichment of extracellular vesicle preparations using anion exchange chromatography. Scientific Reports: 8: 5730.

Email Now

Insert previous message below for editing? 
You haven’t included a message. Providing a specific message means universities will take your enquiry more seriously and helps them provide the information you need.
Why not add a message here
* required field
Send a copy to me for my own records.

Your enquiry has been emailed successfully





FindAPhD. Copyright 2005-2019
All rights reserved.