Objective To discover and confirm blood-based colon cancer early-detection markers.
Design We created a high-density antibody microarray to detect differences in protein levels in plasma from individuals diagnosed with colon cancer <3 years after blood was drawn (ie, prediagnostic) and cancer-free, matched controls. Potential markers were tested on plasma samples from people diagnosed with adenoma or cancer, compared with controls. Components of an optimal 5-marker panel were tested via immunoblotting using a third sample set, Luminex assay in a large fourth sample set and immunohistochemistry (IHC) on tissue microarrays.
Results In the prediagnostic samples, we found 78 significantly (t-test) increased proteins, 32 of which were confirmed in the diagnostic samples. From these 32, optimal 4-marker panels of BAG family molecular chaperone regulator 4 (BAG4), interleukin-6 receptor subunit beta (IL6ST), von Willebrand factor (VWF) and CD44 or epidermal growth factor receptor (EGFR) were established. Each panel member and the panels also showed increases in the diagnostic adenoma and cancer samples in independent third and fourth sample sets via immunoblot and Luminex, respectively. IHC results showed increased levels of BAG4, IL6ST and CD44 in adenoma and cancer tissues. Inclusion of EGFR and CD44 sialyl Lewis-A and Lewis-X content increased the panel performance. The protein/glycoprotein panel was statistically significantly higher in colon cancer samples, characterised by a range of area under the curves from 0.90 (95% CI 0.82 to 0.98) to 0.86 (95% CI 0.83 to 0.88), for the larger second and fourth sets, respectively.
Conclusions A panel including BAG4, IL6ST, VWF, EGFR and CD44 protein/glycomics performed well for detection of early stages of colon cancer and should be further examined in larger studies.
- COLORECTAL CANCER SCREENING
- COLORECTAL ADENOMAS
- COLORECTAL CANCER
- TUMOUR MARKERS
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Contributors J-hR designed the work, conducted experiments, interpreted the data and wrote the manuscript. JJL conducted experiments, interpreted the data and wrote the manuscript. PDL conceived, designed and established the project, interpreted the data and wrote the manuscript. YZ performed statistical analyses and approved the manuscript. DS conducted experiments and approved the manuscript. SMH provided a subset of preliminary data for potential biomarkers and critically reviewed and approved the manuscript. CIL contributed the CHS colon cancer prediagnostic plasma samples and matched controls and critically reviewed and approved the manuscript. DEB collected and provided colon cancer diagnostic plasma samples from the EDRN project and approved the manuscript. HY provided the Japanese samples and critically reviewed the manuscript. TT, HT and TK supplied the Japanese samples and approved the manuscript. DS supplied the colon adenoma and cancer TMAs, helped interpret the results and approved the manuscript. DC performed the TMA staining, assigned the Allred scores, wrote the sections concerning this work and approved the manuscript. JDP provided plasma samples from the CPRU studies conducted at the University of Minnesota and edited and approved the manuscript.
Funding This work was funded in part by grants U01 CA152746 (PDL and SMH), U01 CA152637 (CIL and PDL) and U01CA086400 (DEB) from the National Institutes of Health as part of the EDRN, grant P50 CA130810 (GI SPORE (DEB)), the Kutsche Family Memorial Chair in Internal Medicine (DEB) and the Geriatric Research Education and Clinical Center at the Ann Arbor VA Medical Center. Assaying of the Japanese sample cohort was funded in part by Wako Diagnostics.
Competing interests Fred Hutchinson Cancer Research Center has filed patent applications on the results of this study. HY is an employee of Wako Life Sciences, Inc.
Ethics approval Fred Hutchinson Cancer Research Center institutional review board.
Provenance and peer review Not commissioned; externally peer reviewed.