Development of clinical biomarkers of ADPKD progression

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Final report from the investigators: Our hypothesis is that the proteins contained in the small vesicles (called exosomes) contained in the urine of ADPKD (Autosomal Dominant Polycystic Kidney Disease) patients reflect the disease stage and severity and can be used to identify biomarkers and evaluate risk of progression.

Aim 1: to isolate of urinary exosomes has been completed

The laboratory methodology for optimising the isolation of urinary exosomes has been finished. Membrane vesicles (30-100nm diameter) have been characterized by scanning electron microscopy and marker protein profiling by Western blot (illustrated below).

Exosome Proteomics

Aim 2: to collect and archive at least 200 urine and blood samples from ADPKD patients has been completed and exceeded

More than 350 urine and blood samples have been collected at 6-monthly intervals from ADPKD patients visiting the Royal Free Hospital specialist ADPKD clinic. This cohort of patients includes those at all pre-dialysis stages of progression (i.e. CKD stages 1, 2, 3, and 4). Total urine samples and aliquots suitable for exosome analysis have been processed, archived and stored at -80oC in the PKD Charity-sponsored Bioresource Bank. Sample archiving details have been incorporated into our linked ADPKD clinical information database (Vital Data). Ethics, R&D and LCRN branding NIHR (National Institute for Health Research) applications are being processed for approval to allow for the addition of Royal London and Evelina ADPKD patient sample collection and addition to the Royal Free sample set to provide a confirmatory cohort and to add childhood onset ADPKD to our area of study.

Aim 3: Proteomic profiling of urinary exosomes has been started and proof-of-principle established

A pilot analysis of proteomic profiling has been conducted on a control set of urine samples demonstrating the feasibility of unbiased, comprehensive protein 'fingerprinting' analysis in our NHS (NIHR)-supported UCL/GOSH/Biomedical Research Centre state-of-the-art biological mass spectrometry facility (Dr Mills). A total of 200 patient samples are now ready for analysis for Phase 1 of our cross-sectional study, subject to availability of machine time. This will provide a profile and comparison of early- (stages 1 and 2) and late- stages (3 and 4) of ADPKD. Subsequent prospective samples collected longitudinally every 6 months will be collected and analysed over the next 3 years. Combination of the final results with patients' clinical data and validation in an independent cohort (from Royal London Hospital) will provide an individual progression risk score assessment and the basis for a predictive test.

PI (Principal Investigator):    Dr Jill Norman, UCL/Royal Free Centre for Nephrology, London
Co-PIs:   Dr David Wheeler, Prof Pat Wilson, Dr Danny Gale

Submitted to the PKD Charity in February 2015