Peripheral Blood Transcriptome Study
Research type
Research Study
Full title
Developing a blood test of immunity in illness: A study examining the peripheral blood transcriptome in patients with cancer, autoimmune disease, immunodeficiency or iatrogenic immune suppression.
IRAS ID
150485
Contact email
Sponsor organisation
University of Leeds
Duration of Study in the UK
5 years, 1 months, 0 days
Research summary
Summary of Research
The immune system is vital for good health. In many diseases, changes in the immune system either cause the disease or determine how well the patient "deals with" his/her disease. This is true of cancer, a common skin disease called psoriasis, and in auto-immune diseases such as systemic lupus erythematosus and rheumatoid arthritis. Suppression of immunity is important in organ transplantation. It would be extremely helpful to have a blood test to monitor the immune system to help in diagnosis of the above conditions, to estimate severity (prognosis) and to test whether the treatment is effective or whether over treatment has actually occurred. In this application we seek approval to use a measure of the immune system called the peripheral blood transcriptome in a variety of different diseases and in health to determine if this test could be developed for use as a general measure of immune function. We will compare the value of the test with a commercial test called ImmunTracker and will perform immunophenotyping using flow cytometry (FCM), which is a more widely used test of immunity albeit primarily in a research setting.Summary of Results
Hypothesis to be tested
The hypothesis to be tested in this project was that gene expression patterns in immune cells circulating in the blood reflect functional differences between people, which might be identified and developed as biomarkers to predict prognosis and response to treatment in the future. Moreover ,that the gut microbiome and protein variation in the blood may inform outcomes.Background
Melanoma of the skin is an increasingly frequent tumour in the UK and many western countries. Whilst most patients are cured by prompt excision of the primary tumour, treatment of secondary tumours has been very much more difficult. Without treatment with modern immunotherapy only about 7% of stage IV melanoma patients survive. That modern immunotherapy however is of significant benefit for only around 50% and there is therefore a need to better understand the immune responses to melanoma and to test the hypothesis that measurement of those responses (as measured using the gene expression of circulating immune cells) will be of prognostic and predictive value. The toxicity of the drugs is considerable and much remains to be understood around anticipating that toxicity.The side effects of immunotherapy are essentially because the treatment works by removing breaks on the immune system, and the increased activity of that immune system may then damage healthy tissues. This is very much like the spontaneous emergence of an overactive immune system in autoimmune diseases such as psoriasis. We therefore took samples from participating patients with psoriasis and indeed healthy controls to compare with those from melanoma patients.
Recruitment
Power calculations carried out in 2015 suggested that a total of 1500 participants should be recruited, and the intent was that this study would recruit from multiple centres. In the event the recruitment was almost entirely from Leeds with a very small number from Nottingham. The total number recruited up till March 2020 and COVID was 210. The reasons for poor recruitment are described in the closure of study form but essentially was as follows:-
• Potential participants were usually quite sick and were certainly anxious about their health and considering optional additional tasks such as completing albeit brief questionnaires might understandably seem too much to contemplate.
• If a biomarker was identified, then it would be most useful before the immunotherapy begins so that alternative treatments could be tried instead of a treatment unlikely to be of benefit. Therefore it was important to recruit during that very difficult time just before treatment commences ie when the patients are most anxious and speed is critical.
• Other centres were starting similar studies and although we tried to set up a network of independent centres collecting similar data and pooling data this was not very successful initially. When it became clear that no centres were recruiting easily then this collaboration was established.
• During the pandemic we requested approval for continued recruitment but in fact recruitment ceased completely during the pandemic and we were never able to recommence because the clinical services were so overwhelmed. Whilst the research group study manager was funded by research grants during the period of active recruitment, the grant funding ended during COVID. Further recruitment was therefore dependent upon the clinical services who were overwhelmed.Samples
• Blood samples were collected as described in the protocol. The whole blood was passed through an RNA filter (LeukoLOCKTM) after collection which both catches cells on the filter and fixes their RNA. RNA was then extracted from those cells.
• Blood was also processed in NHS laboratories to measure inflammatory markers, and serum stored for proteomic examination of the blood.
• Stool samples were collected at home by the patient before treatment and before the second treatment and were then mailed to the lab.Generation of peripheral blood transcriptomes
Whole genome expression data were generated from RNA (200ng) using the GeneChip™ Human Transcriptome Array 2.0 (Applied Biosystems™, ThermoFisher Scientific) and the GeneChip™ WT PLUS Reagent Kit according to supplied protocols by a service provider (Hologic, UK). Quality control was carried out in house and small numbers of samples were discarded. The data were analyses using differential gene expression and the biological relevance of differences between diagnoses were explored using MetacoreTM and DAVID.Collaborations
1. We collaborated primarily with the PRIMM consortium directed by Professor Tim Spector and colleagues at Kings College. The aims of the consortium were essentially the same as ours and we provided anonymized data and samples for centralised quality control and processing. Samples of faecal DNA and serum were provided and these formed the basis of two studies referenced below (1, 2). The collaboration continues.
2. We also collaborated with Dr Clare Palles from the University of Birmingham to look at inherited variation impacting on toxicity as a result of immunotherapy. These data are currently in analysis.
3. We will continue to collaborate with the University of Nottingham and other agencies, academic and commercial where the aims of the work are consistent with the consent obtained.REC name
North West - Greater Manchester West Research Ethics Committee
REC reference
15/NW/0933
Date of REC Opinion
21 Dec 2015
REC opinion
Further Information Favourable Opinion