Investigating heterocellular signalling in the micrometastatic stage of colorectal cancer recurrence in the liver

Background: Despite adjuvant chemotherapy, over 30% of Stage III (node-positive) colorectal cancer (CRC) patients suffer post-operative recurrence, which is most frequently seen in the liver. In these cases, micro-metastatic disease has not been effectively cleared, leading to persistence and establishment of colorectal liver metastases. Certain molecular variants (CMS-4) have worse prognosis and these will be our initial focus. Aims: The aims of this project are to: (i) model the micro-metastatic stage of post-surgical CRC recurrence in the liver; (ii) map the signalling cross-talk between CRC cells, hepatocytes and hepatic stellate cells (HStCs); (iii) predict targeted inhibitors which disrupt growth and survival signals conferred on CRC cells by the liver microenvironment; (iv) determine the signalling effects of selected inhibitors compared to standard adjuvant therapy, in co-culture systems, using patient-derived organoids (PDOs). Methods: Micro-metastatic CRC recurrence in the liver will be modelled using 2D and 3D co-cultures of CRC cells/ PDOs, hepatocytes and HStCs. Initially, the effect of resident liver cells on CRC growth and morphology will be assessed by microscopy. Next, reciprocal changes in cell state and signalling will be identified by subjecting co-cultures to mass cytometry (CyTOF). These data will guide the selection and testing of signalling inhibitors (small molecule inhibitors/ blocking antibodies), which abrogate growth and survival benefits conferred upon CRC by the liver microenvironment. Finally, co-cultures comprising PDOs from Stage III CRC will be used to assess signal-directed interventions (e.g. EGFR/ MEK/ PI3K inhibitors) in comparison to and combined with standard of care (e.g. capecitabine/ FOLFOX). How the results of this research will be used: The study will generate a network of signalling between CRC cells and the liver microenvironment, which can be utilised in biomarker development studies and clinical trials for post-surgical Stage III CRC. Signalling nodes and pathways which are activated in co-culture assays will be put forward as predictive biomarkers for selection of targeted adjuvant treatments. Furthermore, clinically approved inhibitors already exist for the majority of signalling nodes being assessed, allowing for potentially rapid translation to clinical trials, or co-clinical trials using PDOs.