Urinary Metabolic Profiling of Liver Fluke-Induced Cholangiocarcinoma-A Follow-Up Study

Munirah Alsaleh; Paiboon Sithithaworn; Narong Khuntikeo; Watcharin Loilome; Puangrat Yongvanit; Thomas Hughes; Thomas O'Connor; Ross H Andrews; Christopher A Wadsworth; Roger Williams; Larry Koomson; Isobel Jane Cox; Elaine Holmes; Simon D Taylor-Robinson

doi:10.1016/j.jceh.2022.11.012

Urinary Metabolic Profiling of Liver Fluke-Induced Cholangiocarcinoma-A Follow-Up Study

J Clin Exp Hepatol. 2023 Mar-Apr;13(2):203-217. doi: 10.1016/j.jceh.2022.11.012. Epub 2022 Nov 26.

Authors

Munirah Alsaleh¹, Paiboon Sithithaworn², Narong Khuntikeo², Watcharin Loilome², Puangrat Yongvanit², Thomas Hughes¹, Thomas O'Connor¹, Ross H Andrews^{1

2}, Christopher A Wadsworth¹, Roger Williams^{3

4}, Larry Koomson¹, Isobel Jane Cox^{3

4}, Elaine Holmes¹, Simon D Taylor-Robinson¹

Affiliations

¹ Department of Metabolism, Digestion and Reproduction, Imperial College London, St Mary's Hospital Campus, London W2 INY, United Kingdom.
² Cholangiocarcinoma Research Institute, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
³ The Roger Williams Institute of Hepatology, Foundation for Liver Research, 111 Coldharbour Lane, London SE5 9NT, United Kingdom.
⁴ Faculty of Life Sciences & Medicine, King's College London, United Kingdom.

Abstract

Background/aims: Global liquid chromatography mass spectrometry (LC-MS) profiling in a Thai population has previously identified a urinary metabolic signature in Opisthorchis viverrini-induced cholangiocarcinoma (CCA), primarily characterised by disturbance in acylcarnitine, bile acid, steroid, and purine metabolism. However, the detection of thousands of analytes by LC-MS in a biological sample in a single experiment potentially introduces false discovery errors. To verify these observed metabolic perturbations, a second validation dataset from the same population was profiled in a similar fashion.

Methods: Reverse-phase ultra-performance liquid-chromatography mass spectrometry was utilised to acquire the global spectral profile of 98 spot urine samples (from 46 healthy volunteers and 52 CCA patients) recruited from Khon Kaen, northeast Thailand (the highest incidence of CCA globally).

Results: Metabolites were differentially expressed in the urinary profiles from CCA patients. High urinary elimination of bile acids was affected by the presence of obstructive jaundice. The urine metabolome associated with non-jaundiced CCA patients showed a distinctive pattern, similar but not identical to published studies. A panel of 10 metabolites achieved a diagnostic accuracy of 93.4% and area under the curve value of 98.8% (CI = 96.3%-100%) for the presence of CCA.

Conclusions: Global characterisation of the CCA urinary metabolome identified several metabolites of biological interest in this validation study. Analyses of the diagnostic utility of the discriminant metabolites showed excellent diagnostic potential. Further larger scale studies are required to confirm these findings internationally, particularly in comparison to sporadic CCA, not associated with liver fluke infestation.

Keywords: ANOVA, analysis of variance; BCAA, branched chain amino acids; CCA, cholangiocarcinoma; CID, collision-induced dissociation; CT, computed tomography; CV-ANOVA, ANOVA of cross-validated residuals; DDA, data-dependent acquisition; ESI −, electrospray ionisation negative mode; ESI, electrospray ionisation; ESI +, electro spray ionisation positive mode; LC-MS, liquid chromatography mass spectroscopy; MRI, magnetic resonance imaging; NMR, nuclear magnetic resonance; OPLS-DA, orthogonal projections to latent structures discriminant analysis; QC, quality control; ROC, receiver operating characteristic; RP, reverse phase; TOF, time of flight; UPLC, ultra-performance liquid chromatography; biomarkers; cholangiocarcinoma; dCCA, distal cholangiocarcinoma; iCCA, intrahepatic cholangiocarcinoma; liver fluke; mass spectroscopy; pCCA, perihilar cholangiocarcinoma.