Urine Proteomics and Renal Single‐Cell Transcriptomics Implicate Interleukin‐16 in Lupus Nephritis

Abstract

Objective Current lupus nephritis (LN) treatments are effective in only 30% of patients, emphasizing the need for novel therapeutic strategies. We undertook this study to develop mechanistic hypotheses and explore novel biomarkers by analyzing the longitudinal urinary proteomic profiles in LN patients undergoing treatment. Methods We quantified 1,000 urinary proteins in 30 patients with LN at the time of the diagnostic renal biopsy and after 3, 6, and 12 months. The proteins and molecular pathways detected in the urine proteome were then analyzed with respect to baseline clinical features and longitudinal trajectories. The intrarenal expression of candidate biomarkers was evaluated using single‐cell transcriptomics of renal biopsy sections from LN patients. Results Our analysis revealed multiple biologic pathways, including chemotaxis, neutrophil activation, platelet degranulation, and extracellular matrix organization, which could be noninvasively quantified and monitored in the urine. We identified 237 urinary biomarkers associated with LN, as compared to controls without systemic lupus erythematosus. Interleukin‐16 (IL‐16), CD163, and transforming growth factor β mirrored intrarenal nephritis activity. Response to treatment was paralleled by a reduction in urinary IL‐16, a CD4 ligand with proinflammatory and chemotactic properties. Single‐cell RNA sequencing independently demonstrated that IL16 is the second most expressed cytokine by most infiltrating immune cells in LN kidneys. IL‐16–producing cells were found at key sites of kidney injury. Conclusion Urine proteomics may profoundly change the diagnosis and management of LN by noninvasively monitoring active intrarenal biologic pathways. These findings implicate IL‐16 in LN pathogenesis, designating it as a potentially treatable target and biomarker.

Interleukin (IL) 16: a candidate urinary biomarker for proliferative lupus nephritis

“…Our findings thus suggest that the discriminatory potential of dt-u-IL-16 for PLN should be further investigated and validated. Our findings extend recent data reported by Fava et al , 10 who identified u-IL-16 as a marker of histological activity in PLN.…”

“…This goes in line with findings by Fava et al, where no correlation with proteinuria was found. 10 We found that more than half of the kidney infiltrating cells in LN express IL-16 protein. Numerically higher numbers of IL-16 positive cells were observed in patients without ongoing treatment.…”

Urine proteomic insights from the belimumab in lupus nephritis trial

“…With respect to renal response, longitudinal reduction in urinary CD163 abundance was strongly associated with complete response (per the definition used in the Methods) in the BLISS-LN trial regardless of treatment arm. Urinary CD163 is a macrophage marker which correlates with histological activity index on renal biopsy and has been previously associated with renal response to LN therapy in at least two independent studies, namely our prior findings in the Accelerating Medicines Partnership in Rheumatoid Arthritis/SLE 4 and also those of Mejia-Vilet et al . 2 As shown in figure 1 , reduction in activated leucocyte cell adhesion molecule (ALCAM) and interferon beta (IFNb) were also seen in renal responders versus non-responders.…”

One-third of patients with lupus nephritis classified as complete responders continue to accrue progressive renal damage despite resolution of proteinuria

“…CKD progression in patients with proteinuric response may be secondary to persistent intrarenal inflammation,8 9 profibrotic processes18 or other mechanisms not yet discovered. We speculate that specific urinary proteins that correlate with intrarenal inflammation or active fibrosis–rather than the total amount of protein in the urine–could potentially be superior predictors of the development of CKD in LN 19 20. Urine proteomic biomarkers of intrarenal matrix remodelling and inflammation can successfully detect early CKD in various other nephropathies and predict risk of CKD progression in diabetic nephropathy 21–23.…”