Investigation of gene-environment interactions between Vitamin D and colorectal cancer susceptibility genetic variants in large bowel epithelium.

Our genes make up a large part of who we are: our gender or the colour of our skin for example. We know that our genetic code also makes some people more susceptible to certain diseases, such as cancer, than others. Until recently, the evidence for this has been the fact that some diseases run in families. However, with increasing knowledge of genetic biology and the completion of human genome project in 2007 we are now beginning to uncover the actual genes that increase the chances of getting a disease. In the long term this research could be used to predict what diseases an individual is prone to and at what age they are likely to develop them. With this information drugs and preventative treatments could be targeted for that individual.
We also know that it is not only our genes, but our environment (for example smoking) that can cause disease, and that often these two factors interact with each other. Sometimes it takes a certain gene and also a certain factor in the environment to cause a disease. Geneticists are beginning to look at how genetic risk factors combine with environmental risk factors to cause disease.
Colorectal cancer is common, with over 1 million people across the world getting the disease every year. Although treatments including surgery and chemotherapy have improved the chance of survival, still only about half those diagnosed are alive 5 years later. Furthermore, the available treatments can cause severe side effects and sometimes death. Therefore work to try and prevent colorectal cancer before it happens could save millions of lives and significantly improve the health and wellbeing of the population.
Studies have shown that vitamin D is an example of an environmental factor that means people are less likely to develop colorectal cancer. For example, Vitamin D is produced in response to sunlight and colorectal cancer is less common in areas with greater amounts of sunlight. However, no one has identified why vitamin D affects colorectal cancer and how our genes are involved. The aim of this project is to investigate how vitamin D and risk genes interact to cause colorectal cancer. Ultimately this knowledge could be used to help develop drugs or supplements that can be given to people with risk genes for colorectal cancer to help prevent the disease. Furthermore, the methods used in this project could be applied to the study of how other diseases with a known genetic component might be prevented.
This project will be conducted at the Institute of Genetics and Molecular Medicine, University of Edinburgh. We have already begun to grow cancer cells in the presence and absence of vitamin D and have shown that it affects some important genes. We will investigate the link between vitamin D and colorectal cancer genes in several other ways:
We will ask permission and then take some cells from cancer specimens removed from patients with bowel cancer and take a vitamin D level from each patient through a simple blood test. Through this we can look at how the vitamin D level is associated with the expression of risk genes in these patients' cancer cells.
Secondly we will set up a study and give cancer patients and healthy volunteers vitamin D supplements every day for 12 weeks. In this way we can control exactly how much vitamin D each patient has. We will take a simple biopsy from the rectum which can be done in the clinic without the need for a general anaesthetic. We can then compare the expression of risk genes in these cells before and after the patients are given vitamin D.
This is a potentially a new and exciting area of research. If we can successfully demonstrate the mechanism by which vitamin D affects colorectal cancer development our future aim would be to trial vitamin D as a targeted chemopreventive agent within the next 10 years.

Aims and Objectives
The host laboratory has demonstrated a high prevalence of vitamin D deficiency in the Scottish population and a highly significant association between vitamin D deficiency and CRC risk. Pilot gene-environment interaction studies have identified vitamin D - SNP interaction signals associated with CRC risk. Homozygotes for one such SNP and low vitamin D levels had 4 fold increased risk of CRC while the tagged gene ('Gene A') is also biologically relevant in human bowel epithelium. The aim of the fellowship is to determine whether vitamin D influences expression of cancer-relevant genes and whether this effect is genotype specific.

Methodology
A series of in vitro, ex vivo and in vivo studies will be conducted. Firstly CRC cells lines and colorectal epithelial organoids will be cultured in the presence and absence of calcitriol and candidate gene expression (including Gene A) investigated. Secondly colorectal epithelium harvested from 100 patients undergoing CRC resection will be collected and the association between plasma vitamin D levels, genotype at known risk loci and expression of candidate genes within the colorectal epithelial tissue will be studied. Finally 40 participants (with and without CRC) will be recruited to a vitamin D intervention study. Rectal biopsies (by rigid sigmoidoscopy) and plasma vitamin D levels will be taken before and after Vitamin D3 supplementation (3200iu/day for 12 weeks; ethical approval pending). Genotype at CRC risk loci will be assessed and comparative gene expression investigated.

Outcomes
Combined analysis of genotype and gene expression from the four arms of the study will aim to determine whether vitamin D impacts on expression of genes tagged by common genetic variance linked to CRC susceptibility. The ultimate aim of this work is the development of targeted chemoprevention and a decrease in the incidence of colorectal cancer with significant attendant health, societal and economic impacts.

We acknowledge the need to consider the social and economic implications of all proposed research. The potential benefits of this project are far-reaching and methods of maximising delivery of the impact will be tailored accordingly.

Clinical and societal impact
This project aims to investigate genetic mechanisms associating two highly incident diseases; vitamin D deficiency and colorectal cancer. The detrimental effects of CRC on wellbeing and quality of life, significant healthcare and welfare costs, and loss of productivity contribute to the impact of this disease. The ultimate aim of this work is the development of targeted chemoprevention and a decrease in the incidence of colorectal cancer with significant attendant health, societal and economic impacts. Furthermore, methods utilised within this project could be applied to the study of other environmental factors that are known to offer chemopreventive potential for colorectal cancer and other common cancers where putative risk loci have already or will be identified.
With potential to improve risk profiling and prediction models, clinicians will be of further importance to help to validate and improve these models. The online risk prediction algorithm (URL: hnpccpredict.hgu.mrc.ac.uk) developed by the CCCG has been used extensively in clinical genetics' practice, with validation in other independent analyses. Our more recent data has also led to the development of risk profiling algorithms and these will ultimately be layered into the current risk model and also lead to the development of newer on-line profiling models.
Development of possible therapeutic targets will also involve discussion and collaboration not only with pharmaceutical companies but clinical departments. Collaborations will be important in facilitating clinical validation of improved risk models and therapeutic agent development.
MGD has previously and remains pivotal to developing clinical screening guidelines, policy, information sheets and molecular interpretation groups, which will be important in maximising the future impact of improved genetic profiling and chemopreventive agents.
Finally, through pathways to impact aimed at the general public, this project will improve public awareness of the importance of colorectal cancer and vitamin D deficiency. In the area local to the host laboratory, this is of particular relevance given the striking incidence of vitamin D deficiency in the Scottish population. Through engagement with the general public, early CRC symptom recognition and presentation will be encouraged enabling earlier diagnosis and improved outcomes. Meanwhile, members of the public may take steps to improve their personal dietary or pharmacological intake of vitamin D.
Public Sector
Development of patentable material will ultimately benefit the University of Edinburgh, both from the patents themselves and also increasing funding to the University. The University has onsite contacts for discussion of such patents and therefore the impact can be realised efficiently. As stated above, the results could lead to improved screening and cancer management, which can lead to reduced NHS, social care and welfare costs.

Economic impact
There is the potential that findings from this research may lead to be patentable discoveries such as:
i) development of targeted preventive or therapeutic agents
ii) risk profiling and prediction models
iii) new CRC risk allele tests

Development of targeted preventive or therapeutic agents and even risk allele tests will interest various industrial companies and we will lead and instigate interactions between Industry and grant giving bodies, to influence and increase similar research at the National level. Furthermore, the realisation of targeted preventive or therapeutic agents would also encourage a major expansion in the commercial private sector developing rapid cost-effective genome sequencing for direct clinical utility.