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Abstract

Diabetic nephropathy (DN) is the main reason for end-stage renal disease. Microalbuminuria as
the non-invasive available diagnosis marker lacks specificity and gives high false positive rates. To identify
and validate biomarkers for DN, we used in the present study urine samples from four patient groups:
diabetes without nephropathy, diabetes with microalbuminuria, diabetes with macroalbuminuria and
proteinuria without diabetes. For the longitudinal validation, we recruited 563 diabetic patients and
collected 1363 urine samples with the clinical data during a follow-up of 6 years. Comparative urinary
proteomics identified four proteins Apolipoprotein A-I (APOA1), Beta-2-microglobulin (B2M), E-cadherin
(CDH1) and Lithostathine-1-alpha (REG1A), which differentiated with high statistical strength (p < 0.05)
between DN patients and the other groups. Label-free mass spectrometric quantification of the candidates
confirmed the discriminatory value of E-cadherin and Lithostathine-1-alpha (p < 0.05). Immunological
validation highlighted E-cadherin as the only marker able to differentiate significantly between the
different DN stages with an area under the curve (AUC) of 0.85 (95%-CI: [0.72, 0.97]). The analysis of the
samples from the longitudinal study confirmed the prognostic value of E-cadherin, the critical increase
in urinary E-cadherin level was measured 20 ± 12.5 months before the onset of microalbuminuria and
correlated significantly (p < 0.05) with the glomerular filtration rate measured by estimated glomerular
filtration rate (eGFR)