Urinary C-X-C Motif Chemokine 10 Independently Improves the Noninvasive Diagnosis of Antibody-Mediated Kidney Allograft Rejection

Abstract

Urinary levels of C-X-C motif chemokine 9 (CXCL9) and CXCL10 can noninvasively diagnose T cell-mediated rejection (TCMR) of renal allografts. However, performance of these molecules as diagnostic/prognostic markers of antibody-mediated rejection (ABMR) is unknown. We investigated urinary CXCL9 and CXCL10 levels in a highly sensitized cohort of 244 renal allograft recipients (67 with preformed donor-specific antibodies [DSAs]) with 281 indication biopsy samples. We assessed the benefit of adding these biomarkers to conventional models for diagnosing/prognosing ABMR. Urinary CXCL9 and CXCL10 levels, normalized to urine creatinine (Cr) levels (CXCL9:Cr and CXCL10:Cr) or not, correlated with the extent of tubulointerstitial (i+t score; all P<0.001) and microvascular (g+ptc score; all P<0.001) inflammation. CXCL10:Cr diagnosed TCMR (area under the curve [AUC]=0.80; 95% confidence interval [95% CI], 0.68 to 0.92; P<0.001) and ABMR (AUC=0.76; 95% CI, 0.69 to 0.82; P<0.001) with high accuracy, even in the absence of tubulointerstitial inflammation (AUC=0.70; 95% CI, 0.61 to 0.79; P<0.001). Although mean fluorescence intensity of the immunodominant DSA diagnosed ABMR (AUC=0.75; 95% CI, 0.68 to 0.82; P<0.001), combining urinary CXCL10:Cr with immunodominant DSA levels improved the diagnosis of ABMR (AUC=0.83; 95% CI, 0.77 to 0.89; P<0.001). At the time of ABMR, urinary CXCL10:Cr ratio was independently associated with an increased risk of graft loss. In conclusion, urinary CXCL10:Cr ratio associates with tubulointerstitial and microvascular inflammation of the renal allograft. Combining the urinary CXCL10:Cr ratio with DSA monitoring significantly improves the noninvasive diagnosis of ABMR and the stratification of patients at high risk for graft loss.

Keywords: antibody-mediated rejection; biomarker; kidney transplantation; noninvasive diagnosis.

Figure 1.

Urinary CXCL9 and CXCL10 levels, normalized or not by urine creatinine, correlated well with the extent of tubulointerstitial inflammation and microvascular inflammation. (A) Dendrogram representations of unsupervised hierarchical clustering analysis of acute and chronic Banff elementary scores of 281 biopsies and CXCL9 and CXCL10 levels normalized or not with urine creatinine. The vertical axis of the dendrogram represents the distance or dissimilarity between clusters. The dendrograms show a high association of CXCL9 and CXCL10, normalized or not with urine creatinine, with all acute Banff scores (i, t, g, ptc, and v). CXCL10 and CXCL10:Cr were found to be most significantly correlated with the microcirculation scores (g and ptc), whereas CXCL9 and CXCL9:Cr correlated most significantly with the cellular inflammation scores (i and t). (B) Spearman correlation matrix of Banff elementary lesions and urinary biomarkers. The color of each box indicates the P value. (C) Correlation of four urinary biomarker levels with the tubulointerstitial inflammation burden (indicated by the g+ptc score) with or without concomitant tubulointerstitial inflammation. Box-and-whisker plots show the log (natural)–transformed urinary biomarker levels. The horizontal line within each box represents the median, the bottom and top of each box represent the 25th and 75th percentile values, respectively, and the I bars represent the 10th and 90th percentile values. Spearman’s correlation coefficient (r_s) and corresponding P values are shown.

Figure 2.

Urinary biomarkers are diagnostic of both TCMR and ABMR. (A) Box-and-whisker plots show the log (natural)–transformed urinary biomarker levels in 203 matched urine/biopsy samples from patients with allograft DNR, 10 matched urine/biopsy samples from patients with TCMR, 37 matched urine/biopsy samples from patients with pure ABMR, and 31 matched urine/biopsy samples from patients with mixed rejection (mixed). P values are on the basis of the Kruskal–Wallis test. Asterisks depict pairwise group comparisons by means of Dunn’s post-test. **P<0.01; ***P<0.001. (B–E) ROC curves for the urinary chemokines. The fraction of true-positive results (sensitivity) and the fraction of false-positive results (1− specificity) for urinary CXCL9 and CXCL10 levels, normalized or not by urine creatinine, as diagnostic biomarkers of AR (both TCMR and ABMR) compared with the fractions in the group of patients with (B) DNR, (C) pure ABMR compared with DNR, (D) mixed rejection compared with DNR, and (E) TCMR compared with DNR. 95% CIs were generated by 2000 stratified bootstrap replicates.

Figure 3.

Urinary CXCL10:Cr ratio improves noninvasive diagnosis of ABMR. Shown are the ROC curves of a conventional noninvasive diagnostic model on the basis of the MFI of the iDSA and a model integrating urine CXCL10:Cr ratio into the previous model. The provided P value indicates the significant difference between the two AUCs.

Figure 4.

CXCL10:Cr ratio is associated with death-censored graft loss after ABMR. Death–censored graft survival postbiopsy showing ABMR on the basis of the median urinary CXCL10:Cr value at the time of biopsy. Estimates were obtained using the Kaplan–Meier method and compared using the log-rank test.