Emerging urinary biomarkers continue to show promise in evaluating lupus nephritis (LN). Here, we screen urine from active LN patients for 1129 proteins using an aptamer-based platform, followed by ELISA validation in two independent cohorts comprised of 127 inactive lupus, 107 active LN, 67 active non-renal lupus patients and 74 healthy controls, of three different ethnicities. Urine proteins that best distinguish active LN from inactive disease are ALCAM, PF-4, properdin, and VCAM-1 among African-Americans, sE-selectin, VCAM-1, BFL-1 and Hemopexin among Caucasians, and ALCAM, VCAM-1, TFPI and PF-4 among Asians. Most of these correlate significantly with disease activity indices in the respective ethnic groups, and surpass conventional metrics in identifying active LN, with better sensitivity, and negative/positive predictive values. Several elevated urinary molecules are also expressed within the kidneys in LN, based on single-cell RNAseq analysis. Longitudinal studies are warranted to assess the utility of these biomarkers in tracking lupus nephritis.

Objectives We investigated the cell adhesion molecules (CAMs) Vascular CAM 1 (VCAM-1) and Activated Leucocyte CAM (ALCAM) as urinary biomarkers in SLE patients with and without renal involvement. Methods Female SLE patients (n = 111) and non-SLE population-based controls (n = 99) were enrolled. We measured renal activity using the renal domain of the BILAG index and urine (U) and plasma (P) concentrations of soluble (s)VCAM 1 and U-sALCAM using ELISA. U-sCAM levels were next corrected by U-creatinine. Results U-sVCAM-1/creatinine and U-sALCAM/creatinine ratios were higher in SLE patients vs non-SLE controls (P < 0.001 for both), as well as in patients with active/low-active (BILAG A–C; n = 11) vs quiescent (BILAG D; n = 19) LN (P = 0.023 and P = 0.001, respectively). U-sALCAM/creatinine but not U-sVCAM-1/creatinine ratios were higher in patients with nephritis history (BILAG A–D; n = 30) vs non-renal SLE (BILAG E; n = 79) (P = 0.014). Patients with baseline U-sVCAM-1/creatinine ratios ≥75th percentile showed a 23-fold increased risk of a deterioration in estimated glomerular filtration rate by ≥25% during a 10-year follow-up (odds ratio: 22.9; 95% CI: 2.8, 189.2; P = 0.004); this association remained significant after adjustments for age, disease duration and organ damage. Traditional markers including anti-dsDNA antibodies did not predict this outcome. Conclusion While high U-sVCAM-1 levels appear to reflect SLE disease activity, sALCAM might have particular importance in renal SLE. Both U-sVCAM-1 and U-sALCAM showed ability to distinguish SLE patients with active renal involvement from patients with quiescent or no prior nephritis. High U-sVCAM-1 levels may indicate patients at increased risk for long-term renal function loss.

Objective. To investigate the utility of a sensitive platform using electrochemiluminescence (ECL) for the identification of low-abundance urinary protein biomarkers in lupus nephritis (LN).Methods. Forty-eight urine samples were obtained from subjects in 2 independent cohorts, each consisting of 3 groups (matched for age, sex, and race) of 8 patients with active LN (renal Systemic Lupus Erythematosus Disease Activity Index [SLEDAI] >0), 8 patients with inactive SLE (renal SLEDAI 0), and 8 healthy controls. Samples were tested using a preexisting 40-plex ECL panel. A custom 5-plex ECL panel was then developed for further validation studies and used to test 140 urine samples (from 44 patients with active LN, 41 patients with inactive SLE, 28 healthy controls, and 27 patients with other kidney diseases).Results. Levels of 17 urinary proteins were elevated (P < 0.05 by 2-tailed Mann-Whitney U test) in samples from patients with active LN compared to samples from patients with inactive SLE and healthy controls in cohort 1, while 9 were similarly elevated in cohort 2. Of these, interleukin-7 (IL-7), IL-12p40, IL-15, interferonγ-inducible protein 10 (IP-10), and thymus and activation-regulated chemokine (TARC) were chosen for further validation. These 5 proteins were undetectable by enzyme-linked immunosorbent assay (ELISA). Hence, a custom 5-plex ECL panel was developed and used to validate the results from the initial 40-plex screening panel. Urinary IL-7, IL-12p40, IL-15, IP-10, and TARC levels were again significantly elevated in patients with active LN compared to those with inactive SLE and healthy controls, and correlated well with the renal SLEDAI and physician's global assessment of disease activity (R > 0.67, P < 0.05). All 5 urinary proteins were more frequently elevated in LN compared to controls with other chronic kidney diseases, although overall group differences attained significance only for urinary IL-7 and IL-15.Conclusion. Urinary levels of IL-7, IL-12p40, IL-15, IP-10, and TARC are potentially useful diagnostic tools in LN. The use of ECL assays may allow detection of urinary biomarkers that are below ELISA detection limits. ECL ASSAYS FOR LOW-ABUNDANCE BIOMARKERS IN LUPUS NEPHRITIS| 745 and others (6) are revolutionizing biomarker discovery, allowing researchers to screen for new biomarkers in an unbiased manner. These new technologies vary in sensitivity and cannot handle the high-volume sample throughput needed for subsequent biomarker validation studies or routine clinical diagnostics.Newer technologies have attempted to reduce the levels of background noise seen in a traditional enzyme-linked immunosorbent assay (ELISA) or antibody-based arrays by replacing an enzymatic-based detection signal with an alternative signal. One of the most promising techniques is the use of an electrode-coupled immunoassay with an electrochemiluminescent (ECL) signal. In this arrangement, the capture antibody is coupled to an electrode, and the detection antibody is labeled with a tag that emits a chemiluminescent...

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Background Lupus affects millions of people worldwide and is difficult to diagnose early in patients. By utilizing enzyme-linked immunosorbent assays (ELISA), this study seeks to identify urinary biomarkers that may detect lupus in its earlier stages, allowing for better, earlier treatment regimens. Methods 197 urine samples from a Swedish SLE cohort were divided into four groups, each matched for demographics: 23 active non-renal SLE samples (SLEDAI&gt;0, rSLEDAI=0), 29 active renal SLE samples (SLEDAI&gt;0, rSLEDAI&gt;0), 46 inactive SLE samples (SLEDAI=0), and 99 healthy controls. These samples were screened for 2 human proteins using ELISA, selected because they exhibited biomarker potential in our earlier screens. Results VCAM was noted to be significantly elevated (2-tailed t-test, p&lt;0.0001) in SLE patients compared to healthy controls. More specifically, VCAM was elevated in active renal samples when compared to healthy samples (p&lt;0.0001). An ROC curve shows that VCAM can be used to differentiate between active renal samples and control samples (AUC=0.8889, p-value=0.0250). ALCAM was also elevated (p&lt;0.0001) in SLE patients compared to healthy patients. However, ALCAM was not as highly elevated in active renal samples compared to healthy samples (p&lt;0.001). Additionally, an ROC curve does not show ALCAM strongly differentiating between active renal samples and control samples. Based on these results, VCAM is the more promising biomarker of the two. Conclusions Urinary VCAM and ALCAM are both potentially viable biomarkers for lupus that warrant further investigation. Studies are in progress to compare the performance of these urinary markers to conventional yardsticks and their relationship to other clinical parameters.