Exploiting differential Wnt target gene expression to generate a molecular biomarker for colorectal cancer stratification

Abstract

ObjectivePathological 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.DesignWe 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.ResultsWnt 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).ConclusionsEpigenetic 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.

Activation of WNT7b Autocrine Eases Metastasis of Colorectal Cancer via Epithelial to Mesenchymal Transition and Predicts Poor Prognosis

“…They suggest that distinguishing between LD and ligand-independent (LI) tumor types is important; patients with LD tumors retained sensitivity to WNT ligand inhibition and may be strati ed at diagnosis to clinical trials of porcupine inhibitors [17]. As more attention turned to WNTs, there were studies of WNT isoform mechanisms in CRC.…”

An “Expressionistic” Look at Serrated Precancerous Colorectal Lesions

“…8) suggests that a variant form of Wnt signaling, likely resembling that reported for alveolar epithelial cells, is active during serrated tumorigenesis. Kleeman et al [38] reported that the Wnt signaling activation observed in CRCs arising through the serrated pathway is ligand-dependent, i.e., resulting from mutations in genes encoding RNF43 or RSPOs proteins, which amplify Wnt signal transmembrane transduction. Increased expression of AQP5 mRNA has also been demonstrated in MMR-de cient CRCs arising via the serrated pathway [62], suggesting that such variant Wnt signaling might be upregulating the expression of this gene in the bases of serrated crypts.…”

“…In contrast, Wnt signaling activation is ligand-independent in CRCs arising along the conventional tumorigenic process acting in cADNs, i.e., tumors with mutations in APC or CTNNB1 genes encoding the intracellular signal transduction proteins Adenomatous polyposis coli or β-catenin, respectively. The constitutive activation in cADNs of this canonical Wnt signaling at the base of normal colorectal crypts upregulates the expression of well-known Wnt target genes, such as CMYC, CD44, NKD1 and AXIN2 [39] [38]. NKD1 (naked cuticle homolog 1) encodes a protein that negatively regulates canonical Wnt signaling via mechanisms that are still incompletely understood [63] [64].…”

“…As for TSAs, they are generally thought to progress along the KRAS-mutated molecular pathway, but progression also appears to occur along the BRAFV600E or KRAS/BRAF-wild type pathway [37]. Recent molecular studies have revealed that signaling pathways typically involved in colorectal tumorigenesis (e.g., Wnt signaling) have different pathogenic trajectories during the evolution of different CRC precursor types [38][39] [40]. For example, somatic APC mutations lead to early constitutive activation of canonical Wnt signaling in cADNs, whereas aberrant Wnt signaling occurs later in SSA/P and TSA tumorigenesis and is triggered by epigenetic silencing (via CIMP) or genetic mutations affecting Wnt-signaling modulators or antagonists (e.g., SFRPs, AXIN2, RNF43, or RSPOs) [18][41] [42] [43].…”

MicroRNA: Another Pharmacological Avenue for Colorectal Cancer?

“…So far as the status, the mechanisms of colorectal carcinogenesis remain still elusive. Recent research has demonstrated that a series of signalings/pathways are involved in the development and oncogenesis of colorectal cancer, such as Wnt ( Kleeman et al, 2019 ), TGF-β ( Calon et al, 2012 ), K-ras ( Codacci-Pisanelli et al, 2009 ), PI3K ( Zhang et al, 2011 ), and p53 ( Seton-Rogers, 2015 ). miR can regulate some pivotal oncogenes and anticancer genes, thus, dysregulation of miR may contribute to the carcinogenesis of tumors.…”