A simple method for detecting oncofetal chondroitin sulfate glycosaminoglycans in bladder cancer urine

Thomas Mandel Clausen; Gunjan Kumar; Emilie K Ibsen; Maj S Ørum-Madsen; Antonio Hurtado-Coll; Tobias Gustavsson; Mette Ø Agerbæk; Francesco Gatto; Tilman Todenhöfer; Umberto Basso; Margaret A Knowles; Marta Sanchez-Carbayo; Ali Salanti; Peter C Black; Mads Daugaard

doi:10.1038/s41420-020-00304-z

A simple method for detecting oncofetal chondroitin sulfate glycosaminoglycans in bladder cancer urine

Cell Death Discov. 2020 Jul 27:6:65. doi: 10.1038/s41420-020-00304-z. eCollection 2020.

Authors

Thomas Mandel Clausen^#^{1

2

3}, Gunjan Kumar^#^{1

2

4}, Emilie K Ibsen³, Maj S Ørum-Madsen^{1

2}, Antonio Hurtado-Coll^{1

2}, Tobias Gustavsson³, Mette Ø Agerbæk^{3

5}, Francesco Gatto^{6

7}, Tilman Todenhöfer^{8

9}, Umberto Basso¹⁰, Margaret A Knowles¹¹, Marta Sanchez-Carbayo¹², Ali Salanti³, Peter C Black^{1

2}, Mads Daugaard^{1

2}

Affiliations

¹ Department of Urologic Sciences, University of British Columbia, Vancouver, BC Canada.
² Vancouver Prostate Centre, Vancouver, BC Canada.
³ Centre for Medical Parasitology at Department for Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen and Department of Infectious Disease, Copenhagen University Hospital, Copenhagen, Denmark.
⁴ Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BC Canada.
⁵ VarCT Diagnostics ApS, Copenhagen, Denmark.
⁶ Department of Biology and Biological Engineering, Chalmers University of Technology, Göteborg, Sweden.
⁷ Present Address: Elypta AB, Stockholm, Sweden.
⁸ Department of Urology, University Hospital Tübingen, Eberhard-Karls University Tübingen, Tübingen, Germany.
⁹ Studienpraxis Urologie, Clinical Trial Unit, Steinengrabenstr. 17, Nürtingen, Germany.
¹⁰ Medical Oncology Unit 1, Istituto Oncologico Veneto IOV - IRCCS, Padova, Italy.
¹¹ Division of Molecular Medicine, Leeds Institute of Medical Research at St James's, St James's University Hospital, Beckett Street, Leeds, UK.
¹² Health Sciences Department, Universidad Pontificia de Salamanca, Salamanca, Spain.

^# Contributed equally.

Abstract

Proteoglycans in bladder tumors are modified with a distinct oncofetal chondroitin sulfate (ofCS) glycosaminoglycan that is normally restricted to placental trophoblast cells. This ofCS-modification can be detected in bladder tumors by the malarial VAR2CSA protein, which in malaria pathogenesis mediates adherence of parasite-infected erythrocytes within the placenta. In bladder cancer, proteoglycans are constantly shed into the urine, and therefore have the potential to be used for detection of disease. In this study we investigated whether recombinant VAR2CSA (rVAR2) protein could be used to detect ofCS-modified proteoglycans (ofCSPGs) in the urine of bladder cancer patients as an indication of disease presence. We show that ofCSPGs in bladder cancer urine can be immobilized on cationic nitrocellulose membranes and subsequently probed for ofCS content by rVAR2 protein in a custom-made dot-blot assay. Patients with high-grade bladder tumors displayed a marked increase in urinary ofCSPGs as compared to healthy individuals. Urine ofCSPGs decreased significantly after complete tumor resection compared to matched urine collected preoperatively from patients with bladder cancer. Moreover, ofCSPGs in urine correlated with tumor size of bladder cancer patients. These findings demonstrate that rVAR2 can be utilized in a simple biochemical assay to detect cancer-specific ofCS-modifications in the urine of bladder cancer patients, which may be further developed as a noninvasive approach to detect and monitor the disease.

Keywords: Bladder cancer; Diagnostic markers.