Association of differential expression of genes with survival and relapse in patients treated on the BILCAP clinical trial: Gene expression identification and response to adjuvant chemotherapy in early-stage biliary tract cancer.

Abstract

4142 Background: Adjuvant capecitabine is standard of care based on the results of the BILCAP clinical trial, comparing adjuvant capecitabine with observation for early-stage biliary tract cancer (BTC). Translational work on data from BILCAP aims to identify differentially expressed genes in patients whose tumours relapsed or who died from their cancer, as there are currently no validated biomarkers to predict the risk of relapse or death or response to adjuvant chemotherapy in early-stage BTC. Methods: Bulk RNA sequencing (RNAseq) was performed on archived fixed formalin samples from consented BILCAP patients. Extracted RNAseq data was quantified using salmon, before undergoing differential gene expression (DGE) using DESeq2 to identify differentially expressed genes in patients who died or whose tumours relapsed, compared to those or survived or had no relapse, accounting for batch effect, different anatomical subtypes and adjuvant treatment. Tumour anatomical subtype was highly associated (p < 0.05) with PC2 during principal component analysis and was included as a co-variate during analysis. Results: 200 patient samples were analysed; 104 / 200 (52%) patients received chemotherapy while 96 had observation. 142 / 200 (71%) tumours relapsed and 142 / 200 (71%) patients died by the time of data cut off. DGE analysis identified 146 significantly (p < 0.01) upregulated genes in patients who died, including PRSS2, AMY2A, SPINK1, CTRC and CELA2A, with significantly (p < 0.01) downregulated genes including HP, PHF14, FLI1, NAV3 and NOVA1. 118 genes were significantly (p < 0.01) upregulated through DGE analysis in patients died or relapsed, including PAX5, FAM188B, NET1, DEK and WDR1 with significantly (p < 0.01) downregulated genes including RQCD1, CPA1, FLI1, NAV3 and NOVA1. 41 genes were significantly (p < 0.01) upregulated in both death and relapse, including PAX5, TLK1, ALB, HEATR3 and RAB11FIP4. 171 genes were significantly (p < 0.01) up regulated in patients who died or had their tumours relapse after adjuvant chemotherapy and not in patients undergoing observation, which included THBS1, FAM65B, UBE2W, RPLP0P2 and NPM1, and 66 genes were significantly (p < 0.01) downregulated, which included RP11-521B24.3, FLI1, CACNA1A, KCNH6 and MKKS. Gene ontology enrichment analysis identified the upregulated genes as being associated with cytoplasmic translation and the synthesis of both intra- and extracellular RNA and protein complexes. Conclusions: Differential gene expression of the BILCAP cohort identified genes associated with cancer relapse and death, including genes associated with a lack of response to adjuvant chemotherapy.