Objective The goal of these studies is to discover novel urinary biomarkers of lupus nephritis (LN).
Methods Urine from systemic lupus erythematosus (SLE) patients was interrogated for 1000 proteins using a novel, quantitative planar protein microarray. Hits were validated in an independent SLE cohort with inactive, active non-renal (ANR) and active renal (AR) patients, in a cohort with concurrent renal biopsies, and in a longitudinal cohort. Single-cell renal RNA sequencing data from LN kidneys were examined to deduce the cellular origin of each biomarker.
Results Screening of 1000 proteins revealed 64 proteins to be significantly elevated in SLE urine, of which 17 were ELISA validated in independent cohorts. Urine Angptl4 (area under the curve (AUC)=0.96), L-selectin (AUC=0.86), TPP1 (AUC=0.84), transforming growth factor-β1 (TGFβ1) (AUC=0.78), thrombospondin-1 (AUC=0.73), FOLR2 (AUC=0.72), platelet-derived growth factor receptor-β (AUC=0.67) and PRX2 (AUC=0.65) distinguished AR from ANR SLE, outperforming anti-dsDNA, C3 and C4, in terms of specificity, sensitivity and positive predictive value. In multivariate regression analysis, urine Angptl4, L-selectin, TPP1 and TGFβ1 were highly associated with disease activity, even after correction for demographic variables. In SLE patients with serial follow-up, urine L-selectin (followed by urine Angptl4 and TGFβ1) were best at tracking concurrent or pending disease flares. Importantly, several proteins elevated in LN urine were also expressed within the kidneys in LN, either within resident renal cells or infiltrating immune cells, based on single-cell RNA sequencing analysis.
Conclusion Unbiased planar array screening of 1000 proteins has led to the discovery of urine Angptl4, L-selectin and TGFβ1 as potential biomarker candidates for tracking disease activity in LN.
- lupus nephritis
- systemic lupus erythematosus
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MP and CM are joint senior authors.
Handling editor Josef S Smolen
Contributors KV, SS, BS, TZ and MN undertook experiments needed for this study, and also read and approved the final manuscript. ClP, PC, CG, SD, SM, NT and VTTT undertook data analysis, and also read and approved the final manuscript. NJ, RS, ChP and MP contributed samples, discussed clinical data interpretation, and also read and approved the manuscript. CM conceived the study, helped design the experiments, interpret data, and read and approved the manuscript.In addition, KV, BS, TZ, SD and CM also wrote the manuscript.
Funding This work is supported by NIH funding AR074096, AR69572, and the George M. O’Brien Kidney Research Core Center P30DK079328. The Hopkins Lupus Cohort is supported by AR69572.
Competing interests None declared.
Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.
Patient consent for publication Not required.
Ethics approval The study was approved by the institutional review boards of John Hopkins University School of Medicine, Albert Einstein College of Medicine, UTSW and the University of Houston.
Provenance and peer review Not commissioned; externally peer reviewed.
Data availability statement All data relevant to the study are included in the article or uploaded as online supplementary information.