A novel cell surface pancreatic and cholangiocarcinoma biomarker that promotes resistance to Gemcitabine


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Jonathan Kay1,Frances Richards2,Laura Bucklow2,Rebecca Brais3,Aarthi Gopinathan2,Hayley Luxton4,Eva Papachristou2,Clive D'Santos2,Duncan Jodrell2,Hayley Whitaker1
1University College London,2CRUK Cambridge Institute,3Addenbrooke's Hospital,4University College london

Abstract

Background

BI-010 has been identified in our laboratory as a membrane bound biomarker that is overexpressed in a number of tumours including lung, breast, bladder, ovarian, pancreatic cancers and cholangiocarcinoma. Its expression can distinguish between cases of pancreatic cancer and chronic pancreatitis that are often at risk of developing cancer. In cholangiocarcinoma cases, BI-010 expression can be combined with IMP3 expression, the current gold standard, to improve the sensitivity of diagnosis. Work is now underway to elucidate its function.

Method

Expression of BI-010 at the cell surface has been confirmed using immunohistochemistry, flow cytometry and cell surface biotinylation. RNAseq analysis was used to determine differentially expressed genes in stably knocked-down shBI-010-Panc-1 and MiaPaCa2 shBI-010-cell lines. These cells were also used to generate shBI-010 knock-down xenograft mouse models. Proteins that interacted with BI-010 were determined using RIME (rapid immunoprecipitation mass spectrometry of endogenous proteins). Activation of multiple kinases was also assessed via a proteome profile array following BI-010 knockdown.

Results

BI-010 knockdown reduced expression of numerous cell surface receptors including TGF-β, WNT/β-Catenin, Notch, EGFR and integrin’s. Independent network analysis of differentially expressed genes and interacting protein revealed an association of BI-010 with components of cell junctions, the extra cellular matrix and cell cycle regulation which indicated a role in endothelial to mesenchymal transition (EMT). ShBI-010 Panc-1 xenograft tumours had significantly reduced expression of EMT markers including Notch 2 and fibronectin. ShBI-010-Panc-1 cells show significantly increased sensitivity (p<0.01) to the standard chemotherapy agent Gemcitabine due to perturbed activation of CHK2, a cell cycle regulator that is activated following DNA damage.

Conclusion

BI-010 is a novel biomarker that is overexpressed in multiple tumours that influences the expression of cell surface receptors. In pancreatic cells and cholangiocarcinoma it shows potential as a future therapeutic target.