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Leukocyte distribution in the developing tumour and the effect of vitamin C

A 2018/2019 Summer Studentship research project

There has not been a systematic study looking at the types of immune cells recruited by the tumour, and how this profile changes over the course of tumour development. Therefore, it would be beneficial to the field of cancer immunotherapy and cancer research to provide a more robust understanding of tumour related immune recruitment over time. This study would also determine if vitamin C may have an impact on immune cell recruitment in cancer.

Student: Thomas Williams
Supervisors: Dr Abel Ang, Associate Professor Gabi Dachs, Associate Professor Margaret Currie
Sponsor: Cancer Society of New Zealand Canterbury/West Coast Division

Project brief


It is now established that tumours are capable of recruiting and subverting immune and inflammatory cells to support tumour growth and expansion while suppressing the immune response. Various cells of the immune and inflammatory response have been shown to be present throughout the tumour and tumour margin, however there is little information of distribution of these cell types during tumour growth. Such information would provide a better understanding of which immune or inflammatory cell types are the initial responders and how the immune or inflammatory response evolves over time.

Vitamin C is a co-factor for numerous proteins with diverse functions, but its effect on the immune response in the context of cancer is poorly understood. Inflammatory and immune cells are among the cell types that actively accumulate vitamin C to high levels, up to 20× higher than plasma levels. This suggests an important role for vitamin C in the function of these cells. Vitamin C is consumed during disease states and cancer patients are reported to often have suboptimal plasma levels, bordering on scurvy. This may have an impact on the function and distribution of immune and inflammatory cells within the tumour and its microenvironment.


Our proposed study will investigate the distribution of various immune and inflammatory cells within the tumour and its microenvironment over time as the tumour grows. It will also determine if this distribution is affected by different tissue levels of vitamin C.


Vitamin C-dependent GULO-/- mice will be used for this study. Female mice will be supplemented with vitamin C in their drinking water (changed twice weekly) at 200mg/L (adequate) or 3300mg/L (high dose). Animals will be maintained on these concentrations for 1 month prior to tumour implantation and throughout the study. Lewis lung tumours will be implanted sc on the flank. Once tumours reach ~50, 200 or 1000 mm3 (caliper measurements daily) mice are injected with a hypoxia probe prior to euthanasia and tissue harvest. Tumours will be processed for frozen and paraffin sectioning. Tumour sections will be stained to determine leukocyte distribution and type by immunohistochemistry and immunofluorescence.

Student researcher’s component of the study

The student will perform immunohistochemistry and immunofluorescence on sectioned tumour samples. Vitamin C content of tumours will be measured using high performance liquid chromatography (HPLC). The student will analyse leukocyte localization and numbers in the stained tumour sections and correlate these with tumour size and vitamin C content of the tumour (all animal work will be completed prior to the summer studentship).