Neolactotetraosylceramide Enables Urinary Detection of Bladder Cancer

Abstract

Glycosphingolipids (GSLs) are potential biomarkers for cancer diagnostics. We applied glycan analytics by multiplexed capillary gel electrophoresis coupled to laser-induced fluorescence detection (xCGE-LIF) to unravel the global GSL profile of tumor tissues and urine samples from bladder cancer patients. We detected neolactotetraosylceramide (nLc4, Galβ1-4GlcNAcβ1-3Galβ1-4Glc-Cer) at statistically significantly increased levels from tumorigenic regions compared to non-malignant adjacent tissue (n = 30), which was validated by immunofluorescence staining. xCGE-LIF analysis of urinary extracellular vesicles showed that nLc4 is increased in bladder cancer patients (n = 16) when compared to controls (n = 50; accuracy, 82%; AUC, 0.75). Finally, we set-up an ELISA targeting nLc4 in a discovery set and an independent validation set, comprising normal, low and high grade bladder cancer urine samples. Increased levels of nLc4 distinguished groups of patients with high grade bladder cancer from control subjects (discovery cohort: n = 18; accuracy, 85%; AUC, 0.94; validation cohort: n = 26; accuracy, 79%; AUC, 0.83). In conclusion, nLc4 has potential as a urinary biomarker for the non-invasive detection of bladder cancer.

Note:
Funding Information: This work was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) for Forschungsgruppe FOR2953 (Projektnummer: 409784463, project P9; BU 2920/4-2 for F.F.R.B.), for Forschungsgruppe FOR2509 (Projektnummer: 289991887, project P3; BU 2920/2-2 for F.F.R.B.) and for the Cluster of Excellence REBIRTH (From Regenerative Biology to Reconstructive Therapy, EXC 62), by the Lower Saxony Ministry of Science and Culture (Niedersächsisches Vorab) for the REBIRTH-Center, by the German Ministry for Education and Research (BMBF 13GW0399B for J.S. and J.H.B.), and by the Wilhelm Sander-Stiftung (for J.S. and J.H.B.).

Declaration of Interests: Patent granted for F. Buettner, C. Rossdam, R. Gerardy-Schahn, A. Oberbeck and H. Tezval: Analytical method and immunological treatment for bladder cancer (EP 20750248.5). All other authors declare that they have no competing interests.

Ethics Approval Statement: Our study was conducted in accordance with the guidelines of the Hannover Medical School Ethics Committee (8619_BO_S_2019 and 10183_BO-K_2022). Informed consents were received from patients/participants who participated in this study.

Keywords: bladder cancer, Biomarker, glycosphingolipid, analytics, liquid biopsy