Identification of in-vivo generated human autoantibodies for the screening and early detection of primary colorectal cancer (CRC)

Lead Research Organisation: University of Nottingham
Department Name: School of Medicine

Abstract

CRC is the second highest cause of cancer mortality in the western world: the most important determinant of survival is stage of disease when diagnosed, with over 90% survival for patients with early stage colorectal cancer.
Randomised trials, involving faecal blood testing followed by colonoscopy (use of a surgical endoscope to examine the colon), have shown early detection with appropriate treatment results in mortality reduction of ~16%. Unfortunately the impact of such interventions is limited by a number of factors, including that fact that the majority of patients are not diagnosed until late stage disease, the low patient acceptability of home faecal sampling required for current tests (Uptake: UK 57%; European 34%) and the low sensitivity and specificity of existing faecal tests. Similarly, although an alternative colonic imaging method (sigmoidoscopy) appears to give higher mortality reductions (22%-31%), patient acceptability remains low (45%-58%) with significant social inequalities. Furthermore the examination is usually limited to only part of the colon (distal colon).

Our solution is to provide a blood test that will improve public acceptability while also providing improved sensitivity & specificity compared to faecal screening. This will significantly improve clinical outcomes (improved survival rates) and cost-effectiveness.

The immune system, which protects us from microbes, also mounts a response to molecules overproduced and released by cancer cells within a tumour. This response includes the generation of antibodies to these tumour-associated molecules. Because the molecules recognised by the antibodies originate from the patient, the antibodies are termed autoantibodies. We, and other groups, have good evidence that detecting these autoantibodies in the blood of patients can provide a route to improved methods for early detection of tumours. Our approach will require only a blood sample, avoiding faecal sampling or invasive imaging colon techniques. As such it is highly likely to dramatically increase patient acceptability, which in itself will improve detection rates and at the same time simplify the testing process and provide improved cost-effectiveness for the NHS.

Our approach is to produce an assay enabling us to test for autoantibodies to multiple tumour molecules simultaneously. This in the form of a microarray (regularly spaced spots of selected tumour-associated molecules on a glass slide surface). Serum from patient blood is washed over the array, and if antibodies are present, they bind to one or more of the tumour-associated molecule spots. Bound antibodies are themselves detected with a fluorescent reporter and the signal intensity measured. Comparison to cutoffs based upon examination of known colorectal cancer patients and normal controls allows the identification of samples positive for tumour-molecule autoantibodies, indicating that further investigation is required. The small size of the microarray and spots allows for replication of spots for each tumour-associated molecule on the array, yet requires significantly reduced chemical volumes over tests which examine only one tumour-associated molecule per test.
The potential of this microarray-based autoantibody screening in colorectal cancer is demonstrated in our preliminary work, using a limited panel of 32 tumour-associated molecules and a small sample set of 262 known cancer/normal individuals.
This project will identify the optimal panel from the 32 tumour-associated molecules which are suitable for use as a colorectal cancer screening tool and validate that resultant panel using much larger, clinically well-defined sample sets than our preliminary study.

Our prediction is that with a patient compliance rate of >90%, test specificity of 85% and sensitivity of 65% this would result in detection of 58.5% of the total number of CRCs -more than a doubling of the total number of CRCs currently detected.

Technical Summary

Colorectal cancer (CRC) is the second most common cancer in the western world: the critical survival determinant is disease stage (over 90% survival for Duke's Stage A).
Randomised trials, using the faecal occult blood test (FOBt) followed by colonoscopy, have shown early detection and intervention resulting in ~16% reduced mortality. This impact is limited due to the majority of patients presenting with late disease, low patient compliance for FOBt (Uptake: UK 57%; European 34%) and the relatively low sensitivity (~50%) and specificity (~75%).
Although mortality reduction for one off and repeat flexible sigmoidoscopy appears higher (22%-31%), patient acceptability remains low (45%-58%). Uptake of one-off colonoscopy is even lower (24.6%).
We aim to develop a robust, non-invasive blood test with sensitivity (>65%), specificity(>85%) and a high patient acceptability rate (>90%), with the potential to significantly improve clinical outcomes by detecting early stage CRC.
Immune system recognition of tumour associated antigens (TAAs) elicits autoantibody (AAb) responses during early tumour development; detection of such AAb can support early CRC detection. Microarray technologies for AAb detection offer scalability and economy of sample and reagent use, making them ideal tools for cost-effective, comprehensive CRC screening.
Preliminary work, generated with an MRC CiC award, measured AAb responses to 32 TAAs: identifying that a subset of six TAAs provides >65% sensitivity & >85% specificity using CRC/control samples (262).
This project will identify an optimal panel from the larger group of 32 TAAs and validate that core panel to pre-defined sensitivity and specificity using larger, clinically well-defined sample sets provided by 2 UK and 2 USA centres with known expertise in screening for CRC.
An optimised AAb blood test will, in future prospective studies, be assessed against FOBt or its likely successor the quantitative Faecal Immunochemical Test (qFIT).

Planned Impact

Impact thus Far & Unmet Need: Randomised clinical trials have shown that early detection with appropriate intervention results in reduced mortality of breast, lung and colorectal cancer (CRC). Despite these advances it is generally agreed that there are still significant unmet needs in effective and cost efficient screening and early diagnosis of this common cancer.
Project Goal: A non-invasive blood based test leading to early detection of CRC will dramatically change clinical practice with the potential to significantly improve clinical outcomes. As a blood based test it will have the advantage of being applicable in low and medium income countries.
Dissemination Programme: Dissemination, (publications in peer review journals and Presentations at scientific conferences) plus public and general engagement will play a vital role in raising awareness of the project's results to potential beneficiaries.

This project has the potential to significantly impact on human health worldwide - to the benefit of the population at risk of CRC, patients who develop the disease and also more generally to society and health care providers. The previous lung cancer project (EarlyCDT-Lung) has shown the science is real and clinically robust. This project will show the science is applicable to another common solid cancer and that the improved microarray platform technology is applicable to all solid cancers with potential impact on cancer detection and treatment worldwide.

Dissemination of the results of this project will enable exploitation through downstream validation trials and subsequently rapid uptake by health services, policy makers, patient groups and other supporting organisations. In particular we will inform the Dept of Health National Screening Advisory Board which in turn advises DoH on Screening modalities and policy for Bowel Cancer Screening as well as detailed dissemination and promotion to national bodies throughout Europe and to the International Agency for Research in Cancer.
A comprehensive array of dissemination and engagement activities are planned particularly to patient groups and the general public. The University of Nottingham and the CEAC already have existing precise dissemination programmes to patient groups such as the Global CRC Coalition, the Cancer Information Network (online), The Patient Advocate Foundation (USA), Living With It (online), Patient UK and Macmillan Relief, Beating Bowel Cancer, Digestive Diseases Foundation, Bowel Disease Research Foundation.

Dissemination of the research results will also provide new knowledge to other researchers which they can use to build upon not only in terms of early detection of cancer but also in terms of stratified medicine (in selecting specific treatments based on indirect autoantibody detected tumour antigen arrays) and also other in non-malignant auto-immune diseases. The reputations of the Co-applicants will also facilitate new worldwide collaborations with other research centres of excellence.

Web site & Media: The project's web site will be integrated within the CEAC/UoN website and will have hierarchical levels for public engagement, interested stakeholders and detailed scientific information. It will also be directly linked to additional media coverage. The UoN and CEAC have established press release officers responsible for engagement with TV, radio and the press; with substantial media coverage already arisen as a result of previous research impacts.

Exploitation route: IP arising from this programme will support and enable the exploitation of the science required to bring a CRC AAB test to market and clinical practice.

Publications

10 25 50
 
Description James Tudor Foundation
Amount £60,000 (GBP)
Organisation The James Tudor Foundation 
Start 03/2018 
End 02/2021
 
Title Microarray panel assay 
Description This award has significantly improved the technical characterisation of the microarray assay and reagents such that it can now be used as a reproducible protein micro-array to measure autoantibodies. The initial assay was the basis of out pilot results (10.1371/journal.pone.0156971) which has been significantly improved by this award. By the measurement of autoantibodies to tumour associated antigens the readout provides what is essentially an indirect protein microarray. This technology can be used to understand the immune response to various diseases including cancer. For example assessment of sequential blood samples - eg samples collected from at risk populations prior to the development of cancer - will allow assessment of what tumour antigens are expressed on the cancer and presented to the immune system and the order these antigens are expressed during the carcinogensis process. The technical QC on the microarray platform will allow ojective, reproducible indirect protein array data. We have only just developed this techology and have not yet provided it to other research groups. 
Type Of Material Technology assay or reagent 
Provided To Others? No  
Impact None yet. 
 
Title Autoantibodies for early detection of CRC 
Description Database contains list of samples of CRC, polyps and matched controls along with relevant clinical data. It also has microarray assay autoantibody results from multiple tumour associated antigens. The autoantibdoy results can be correlated with the clinical patient data. We are still in the process of completing the research programme funded by the award with involves constructing the complete database. Untill we have done so and analysed the dataset as part of this award we would not plan to share the database. 
Type Of Material Database/Collection of data 
Provided To Others? No  
Impact None yet. 
 
Description Mount Sinai Hospital, New York 
Organisation Mount Sinai Hospital (USA)
Department Department of Surgery
Country United States 
Sector Hospitals 
PI Contribution Our team have developed the laboratory microarray technology for measuring auto-antibodies to tumour associated antigens. This work has been further supported and developed through this award.
Collaborator Contribution Our partner has provided the blood samples and associated clinical data as one of three collaborating centres - two in the USA and one in the UK - to allow us to construct both a training and a validation dataset.
Impact Publication of pilot data on the use of serum autoantibodies for the detection of early colorectal cancer. PMID:27383396
Start Year 2012
 
Description University of Dundee 
Organisation University of Dundee
Country United Kingdom 
Sector Academic/University 
PI Contribution Our team have developed the laboratory microarray technology for measuring auto-antibodies to tumour associated antigens. This work has been further supported and developed through this award.
Collaborator Contribution Our partner has provided the blood samples and associated clinical data as one of three collaborating centres - two in the USA and one in the UK - to allow us to construct both a training and a validation dataset.
Impact Publication of pilot data on the use of serum autoantibodies for the detection of early colorectal cancer. PMID:27383396
Start Year 2012
 
Description University of Pittsburgh 
Organisation University of Pittsburgh
Department Department of Gastroenterology, Hepatology and Nutrition
Country United States 
Sector Hospitals 
PI Contribution Our team have developed the laboratory microarray technology for measuring auto-antibodies to tumour associated antigens. This work has been further supported and developed through this award.
Collaborator Contribution Our partner has provided the blood samples and associated clinical data as one of three collaborating centres - two in the USA and one in the UK - to allow us to construct both a training and a validation dataset.
Impact Publication of pilot data on the use of serum autoantibodies for the detection of early colorectal cancer. PMID:27383396
Start Year 2012
 
Description Philanthropic funding 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Supporters
Results and Impact I have made numerous presentations on our work on auto-amtobodies in cancer. Since ~2012 I have included our work on colorectal cancer which led to the MRC award and since the award in 2016 I have specifically mentioned the work and results generated by the MRC award thus far. The talks/lectures have also reported on our results using autoantibodies for the early detection of other types of cancer (eg lung, breast, hepatocellular and pancreatic).
Year(s) Of Engagement Activity Pre-2006,2006,2007,2008,