Application of Holistic Liquid Chromatography-High Resolution Mass Spectrometry Based Urinary Metabolomics for Prostate Cancer Detection and Biomarker Discovery

Abstract

Human exhibit wide variations in their metabolic profiles because of differences in genetic factors, diet and lifestyle. Therefore in order to detect metabolic differences between individuals robust analytical methods are required. A protocol was produced based on the use of Liquid Chromatography- High Resolution Mass Spectrometry (LC-HRMS) in combination with orthogonal Hydrophilic Interaction (HILIC) and Reversed Phase (RP) liquid chromatography methods for the analysis of the urinary metabolome, which was then evaluated as a diagnostic tool for prostate cancer (a common but highly heterogeneous condition). The LC-HRMS method was found to be robust and exhibited excellent repeatability for retention times (<±1%), and mass accuracy (<±1 ppm). Based on normalised data (against creatinine levels, osmolality or MS total useful signals/MSTUS) coupled with supervised multivariate analysis using Orthogonal Partial Least Square-Discriminant Analysis (OPLS-DA), we were able to discriminate urine samples from men with or without prostate cancer with R2Y(cum) >0.9. In addition, using the receiver operator characteristics (ROC) test, the area under curve (AUC) for the combination of the four best characterised biomarker compounds was 0.896. The four biomarker compounds were also found to differ significantly (P<0.05) between an independent patient cohort and controls. This is the first time such a rigorous test has been applied to this type of model. If validated, the established protocol provides a robust approach with a potentially wide application to metabolite profiling of human biofluids in health and disease.

Metabolomic Biomarkers of Prostate Cancer: Prediction, Diagnosis, Progression, Prognosis, and Recurrence

“…developed their signatures in blood samples, and it is of note that there was little crossover in terms of the specific constituent metabolites with the urine-based diagnostic signatures proposed by Zhang et al . (38), Struck-Lewick et al (42) or Wu et al . (36).…”

“…The selected studies indicated that distinct clusters based on metabolite profiles could be identified in a variety of biological media including blood (40, 41, 51), urine (36, 38, 42), tissue (16, 28, 31, 32, 45, 47, 52) and expressed prostatic secretions (39). These clusters were able to distinguish cancer from benign (28, 31, 32, 36, 38-42, 47), tumors by their degree of aggressiveness (16, 32, 47), cases who recurred (45), and those who responded well to therapy (51).…”

“…These clusters were able to distinguish cancer from benign (28, 31, 32, 36, 38-42, 47), tumors by their degree of aggressiveness (16, 32, 47), cases who recurred (45), and those who responded well to therapy (51). The distinction between the groups of interest on the basis of their metabolic profiles suggested that the development of metabolomics-based biomarkers may be possible.…”

<strong>A short overview on the use of metabolomic multi-platforms in biomedical research</strong>

“…On this basis, the combination of reversed-phase and hydrophilic interaction liquid chromatography (HILIC) is also an interesting strategy, since the coupling HILIC-MS allows analyzing polar compounds in biological samples without the recourse of the derivatization step needed in gas chromatography [4][5][6]. Alternatively, capillary electrophoresis (CE) can also resolve the polar metabolome, but its combination with RP-LC-MS has only been proposed in a few works [7][8].…”

Translational Metabolomics in Cancer Research

“…The field of lipidomics has evolved to fill this gap and lower abundance metabolites are analyzed in more specialized assays such as those described for estrogens [3]. Cancer metabolic studies are generally conducted using either NMR spectroscopy or LC-MS using three major approaches: first, untargeted LC-MS methods to profile metabolites in order to discover those that are dysregulated using sophisticated software packages to reveal differences in the chromatographic profiles [4–7]. Characterizing the differentially regulated metabolites that are found remains a major challenge.…”