Objective Pathological Wnt pathway activation is a conserved hallmark of colorectal cancer. Wnt-activating mutations can be divided into: i) ligand-independent (LI) alterations in intracellular signal transduction proteins (Adenomatous polyposis coli, β-catenin), causing constitutive pathway activation and ii) ligand-dependent (LD) mutations affecting the synergistic R-Spondin axis (RNF43, RSPO-fusions) acting through amplification of endogenous Wnt signal transmembrane transduction. Our aim was to exploit differential Wnt target gene expression to generate a mutation-agnostic biomarker for LD tumours.
Design We undertook harmonised multi-omic analysis of discovery (n=684) and validation cohorts (n=578) of colorectal tumours collated from publicly available data and the Stratification in Colorectal Cancer Consortium. We used mutation data to establish molecular ground truth and subdivide lesions into LI/LD tumour subsets. We contrasted transcriptional, methylation, morphological and clinical characteristics between groups.
Results Wnt disrupting mutations were mutually exclusive. Desmoplastic stromal upregulation of RSPO may compensate for absence of epithelial mutation in a subset of stromal-rich tumours. Key Wnt negative regulator genes were differentially expressed between LD/LI tumours, with targeted hypermethylation of some genes (AXIN2, NKD1) occurring even in CIMP-negative LD cancers. AXIN2 mRNA expression was used as a discriminatory molecular biomarker to distinguish LD/LI tumours (area under the curve >0.93).
Conclusions Epigenetic suppression of appropriate Wnt negative feedback loops is selectively advantageous in LD tumours and differential AXIN2 expression in LD/LI lesions can be exploited as a molecular biomarker. Distinguishing between LD/LI tumour types is important; patients with LD tumours retain sensitivity to Wnt ligand inhibition and may be stratified at diagnosis to clinical trials of Porcupine inhibitors.
- colorectal cancer
- Wnt signalling
- molecular biomarker
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Contributors SOK, VHK, TM and SJL conceived and designed the project. Experiments were conducted by SOK, VHK, HJSJ, EGV, HD, VP, MAJK, TRML, SLW and DW. Data provision and bioinformatic analysis were carried out by ADB, SOK, RA and ED. Pathology support, tissue provision and intellectual input were from VHK, L-MW, JEE, CC, MBL, PDD and IT. Manuscript written by SOK and SJL.
Funding This work was supported by Wellcome Trust Senior Clinical Research Fellowship (206314/Z/17/Z) and Worldwide Cancer Research grant (16-0042) to SJL, the Stratification in Colorectal Cancer (S:CORT) Consortium, which is funded by the Medical Research Council and Cancer Research UK and the National Institute for Health Research (NIHR) Oxford Biomedical Research Centre. VHK was funded by the Swiss National Science Foundation (P2SKP3_168322/1 and P2SKP3_168322/2). HJSJ was supported by a Bowel Disease Research Foundation (BDRF) research grant. ADB is funded by a Cancer Research UK Advanced Clinician Scientist award (C31641/A23923) and the Wellcome Trust (102732/Z/13/Z). TRML, SLW and DW supported by the NHMRC, Beat Cancer SA & Cure Cancer Australia. JEE was funded by the National Institute for Health Research (NIHR) Oxford Biomedical Research Centre (BRC). Core funding to the Wellcome Trust Centre for Human Genetics was provided by the Wellcome Trust (090532/Z/09/Z).
Competing interests SJL has received grant income from UCB Pharma. VHK is participant of a patent application co-owned by the Netherlands Cancer Institute (NKI-AVL) and the University of Basel on the assessment of cancer immunotherapy biomarkers by digital pathology and has served as an invited speaker on behalf of Indica Labs. ADB has received grant funding from InCyte for molecular pathology research; speaker fees and travel costs from Illumina UK for immuno-oncology research and consultancy fees from Bristol-Myers-Squibb for immuno-oncology research. All other authors have nothing to disclose.
Patient consent for publication Not required.
Provenance and peer review Not commissioned; externally peer reviewed.
Data availability statement Data relating to transcriptome profiling of relevant genes in polyps cohort and validation cohorts A/B are available online (https://www.s-cort.org/exploiting-differential-wnt). All other data relevant to the study are referenced, included in the article or uploaded as supplementary information.
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