Abstract A039: Lower cellular activation of cMET signaling network is associated with reduced recurrence risk in stage II colorectal cancer
Authors: McDonough Elizabeth , Sood Anup , Ginty Fiona , Cho Sanghee , Graf John , Prehn Jochen , Dunne Philip , Lindner Andreas , Salvucci Manuela , Longley Daniel , Lawler Mark , Hollmann Travis , Shia Jinru 
Published: March 2020

Abstract
Colorectal cancer (CRC) has one of the highest worldwide incidences and mortality rates. About 25% of stage II patients develop recurrence within 5 years. However, use of adjuvant chemotherapy yields minimal therapeutic benefit. Clinical decision-making in this population would be informed by prognostic and predictive biomarkers that help indicate more targeted interventions and/or intensified disease monitoring. Higher cMET expression has been shown to be a prognostic marker in CRC and has been evaluated as a drug target in several cancers, including CRC. cMET acts as the receptor for hepatocyte growth factor (HGF), and is associated with increased proliferation, migration and morphogenesis of epithelial cells via activation of multiple downstream pathways including PI3K/AKT, MAPK and the NF-kB pathways. Here, we investigated whether increased cellular expression of multiple markers in the cMET/adjacent pathways was correlated with clinical outcome. The study population consisted of 283 patients with stage II CRC, without neoadjuvant treatment. Using a multiplexed immunofluorescence method (MxIF) with cell-level quantification (Cell DIVETM, GE Healthcare), 41 biomarkers in the cMET, mTOR, MAPK and associated pathways, and lymphocyte markers were analyzed in an iterative sequence of staining, imaging and dye inactivation, followed by image registration, cell segmentation and biomarker intensity-quantitation. Poor quality images and cells were filtered based on poor segmentation and tissue quality following manual review. Images that did not perfectly register were also excluded. For epithelial cell analysis, all stromal cells were filtered, resulting in a total of 559,952 epithelial cells. K-means clustering was conducted on cMET and related pathways (cMET, pGSK3β, 4EBP1, p4EBP1, S6, pS6, pStat3, pp38MAPK, pNFkBp65, pNFkBp105, EGFR, pERK1/2, HER2, IGF1R, CA-IX, Glut1, SLC7A5 and Ki67). Cluster analysis identified 6 cell clusters with varying expression levels of cMET and other pathway markers. A cluster corresponding to low cMET network activation (cMETLow) was correlated with reduced risk of recurrence (cox pH model Likelihood ratio test p-value = 0.056). Furthermore, MMR proficient patients (who had a higher recurrence rate) had a significantly greater proportion of cells with elevated cMET and related pathway marker (cMETHigh) s(p= 0.007). They also had a significantly lower fraction of infiltrating helper T cells (CD3+CD4+, pvalue 0.076) and CD68+ cells in the epithelial region, compared to MMR deficient patients (p=0.005). Further research will include continuing in-depth analysis of MMR status, cMET pathway and lymphocyte response as well as the role of cellular heterogeneity. In summary, comprehensive cMET pathway analysis using a multiplexed single cell approach indicates for the first time an association between low cMET network activation and superior clinical outcomes.
Citation Format: Elizabeth McDonough, Anup Sood, Fiona Ginty, Sanghee Cho, John Graf, Jochen Prehn, Philip Dunne, Andreas Lindner, Manuela Salvucci, Daniel Longley, Mark Lawler, Travis Hollmann, Jinru Shia. Lower cellular activation of cMET signaling network is associated with reduced recurrence risk in stage II colorectal cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A039. doi:10.1158/1535-7163.TARG-19-A039