Background. Biopsy-based transcripts associated with antibody-mediated rejection (AMR) hold promise as substitutes for C4d positivity. However, their utility in cases with additional diagnoses other than rejection remains inadequately studied. Methods. In our comprehensive analysis of 326 kidney allograft biopsies, assessed by histology and the Molecular Microscope Diagnostic System, we identified 68 cases characterized by additional pathologies, including pyelonephritis (n = 15), BK nephropathy (n = 20), acute interstitial nephritis (n = 5), and glomerular diseases (n = 28). Results. Among cases with pyelonephritis, 7 of 15 cases (46%) showed a rejection-like signal, 4 above (16%) and 3 (20%) below diagnostic thresholds. Notably, the T cell–mediated rejection (TCMR) archetype score R2 (median, 0.13; interquartile range [IQR], 0.04–0.34) predominantly contributed to this observation. In BK nephropathy, 13 of 20 cases (65%) showed a rejection-like signal, 10 (50%) above and 3 (15%) below diagnostic thresholds. Elevated TCMR R2 (median, 0.07; IQR, 0.00–0.41) and all AMR archetype scores R4–6 (median, 0.23; IQR, 0.07–0.53) were driving factors. Among cases with acute interstitial nephritis, 3 of 5 cases (60%) showed TCMR-like signal with elevated R2 scores (median, 0.13; IQR, 0.00–0.54). Conversely, only 5 of 28 cases (18%) showed a rejection-like signal in glomerular disease cases, whereas 57% displayed all AMR archetype scores of ≥0.30. Conclusions. Additional pathologies can affect the Molecular Microscope Diagnostic System output, giving a molecular rejection-like signal. The prevalence of rejection-like signals below diagnostic thresholds is noteworthy, warranting caution and prompting further investigation.

Abstract Background and Aims Biopsy based transcript diagnostics using the molecular microscope diagnostic system (MMDx) have the potential to add diagnostic clarification in the antibody mediated rejection (AMR) continuum. While de novo donor specific antibody (dnDSA) or preformed DSA accompanied by allograft function deterioration are accepted indications for allograft biopsy with therapeutic implications, management strategy for subclinical DSA lacks consensus. MMDx might show clinical utility in this controverse scenario by helping with therapeutic decisions. Method In this single-center cohort of 326 indication kidney transplant biopsies assessed by histology and MMDx at the University Hospital Zurich, we analyzed 58 cases with subclinical DSA as biopsy indication. Cases with (n = 44) and without (n = 14) histopathologic features of AMR according to Banff 2022 were compared for molecular AMR rejection transcripts. Results When biopsied for subclinical DSA, all 14 cases without histopathologic features of AMR showed absence of molecular AMR. In contrast, molecular AMR was detected in 6/19 (32%) cases with histopathologic diagnosis of AMR and 2/25 (8%) with probable AMR (p = 0.059), Fig. 1. Microvascular inflammation below threshold with C4d negativity was significantly associated with absence of molecular AMR (p < 0.0001). Higher number of calculated epitope mismatch load for HLA class II as measured by PIRCHE II Score was significantly associated with molecular AMR (p = 0.01). In cases with molecular AMR, early AMR scores predominated (R4 vs R6, 0.0002 and R4 vs R5 0.051). Conclusion No molecular AMR was observed in absence of histopathologic AMR. Yet, in cases with histopathologic features of AMR, only 8/44 showed molecular AMR. Predominance of early AMR rejection phenotype in cases with molecular AMR transcripts allows hope for MMDx to identify treatment amenable cases with subclinical DSA. Discrepant cases call for re-biopsy to guide therapeutic decisions.

Biopsy-based transcript diagnostics may identify molecular antibody-mediated rejection (AMR) when microvascular inflammation (MVI) is absent. In this single-center cohort, biopsy-based transcript diagnostics were validated in 326 kidney allograft biopsies. A total of 71 histological AMR and 35 T cell-mediated rejection (TCMR) cases were identified as molecular AMR and TCMR in 55% and 63%, respectively. Among 121 cases without MVI (glomerulitis + peritubular capillaritis = 0), 45 (37%) donor-specific antibody (DSA)-positive and 76 (63%) DSA-negative cases were analyzed. Twenty-one out of the 121 (17%) cases showed borderline changes, or TCMR, while BK nephropathy was excluded. None of the 45 DSA-positive patients showed molecular AMR. Among 76 DSA-negative patients, 2 had mixed molecular AMR/TCMR. All-AMR phenotype scores (sum of R4-R6) exhibited median values of 0.13 and 0.12 for DSA-positive and DSA-negative patients, respectively (P = .84). A total of 13% (6/45) DSA-positive and 11% (8/76) DSA-negative patients showed an all-AMR phenotype score > 0.30 (P = .77). Patients with a higher all-AMR phenotype score showed 33% more histologic TCMR (P = .005). The median all-AMR phenotype scores of glomerular basement membrane double contours = 0 and glomerular basement membrane double contours > 0 biopsies were 0.12 and 0.10, respectively (P = .35). Biopsy-based transcript diagnostics did not identify molecular AMR in cases without MVI. Follow-up biopsies and outcome data should evaluate the clinical relevance of subthreshold molecular alterations.

Background Isolated tubulitis, borderline changes and isolated arteritis suspicious for histologic T-cell-mediated rejection (hTCMR) remain findings of uncertain significance. Although the Molecular Microscope Diagnostics System (MMDx) has not been trained on those lesions, it was suggested that MMDx might reclassify a subgroup to molecular TCMR (mTCMR). Methods In this single-center cohort of 326 consecutive, unselected kidney allograft biopsies assessed by histology and MMDx, we analyzed 249 cases with isolated tubulitis (i0, t1-3, v0; n = 101), borderline changes (according to Banff 2022, v0; n = 9), isolated arteritis (no borderline, v1; n = 37), no inflammation (i0, t0, v0; n = 67) and a positive control cohort (hTCMR, n = 27; mixed histologic rejection, n = 8; both according to Banff 2022; total n = 35). The first three groups were summarized as TCMR-suspicion (n = 147). Subcategorization included the presence and absence of microvascular inflammation (MVI); g+ptc ptc >= 2. Molecular rejection rates and differentiation were investigated. Results Molecular rejection rates were 37/147 cases (25.2%; 32 with MVI) in TCMR-suspicion, 6/67 (9%; 4 with MVI) in no inflammation and 30/35 (85.7%; 19 with MVI) in the positive control cohort. Molecular antibody-mediated rejection (mAMR) was present in 39/73 (53.4%) of cases. The presence of donor-specific antibodies at the time of the biopsy was high (127/249, 51%). Only 3 mAMR/TCMR and 0 pure mTCMR cases were detected in TCMR-suspicion and no inflammation, compared with 12 mAMR/TCMR and 10 mTCMR cases in the positive control cohort (P < .001). Even though the TCMR-specific molecular (Classifier) score differentiated between TCMR-suspicion and no inflammation (P = 0.005), rejection phenotype scores (R2 and R3) did not (P = .157 and .121). Conclusions MMDx did not identify pure mTCMR among isolated tubulitis, borderline changes or isolated arteritis, likely due to low sensitivity for TCMR lesions. However, it identified mAMR or mAMR/TCMR, especially in cases with MVI. Subthreshold findings remain to be further studied.

Abstract Background and Aims Biopsy-based transcripts associated with antibody-mediated rejection (AMR) hold promise as substitutes for C4d positivity, according to the latest Banff Meeting Report of 2022. However, patterns and strength of rejection phenotype scores remain inadequately studied within the new subgroups along the AMR continuum. Method We analyzed 365 kidney allograft biopsies by histology and the Molecular Microscope Diagnostic System (MMDx) at the University Hospital of Zurich from July 2021 to November 2023. Histologic findings were classified according to Banff 2022 into (1) probable AMR (n = 32), (2) donor-specific antibodies (DSA)-negative C4d-negative microvascular inflammation (MVI, n = 32), and (3) AMR (n = 60). A control group of 118 cases without MVI was used for comparison. Results 10 out of 32 cases (31%) with probable AMR, 20 of 32 cases (63%) with DSA-negative C4d-negative MVI, and 31 of 60 cases (52%) with AMR showed molecular AMR. Among cases with molecular AMR the sum of the TCMR-related (R2) and late-stage AMR-related (R6) phenotype scores was significantly higher in probable AMR and DSA-negative MVI cases compared to AMR cases (p = 0.029). In contrast, the sum of the mixed rejection (R3), early-stage AMR (R4), fully-developed AMR (R5) phenotype scores was higher in AMR compared to probable AMR (p = 0.063). Among cases without molecular AMR the all AMR phenotype score was significantly higher in AMR and DSA-negative MVI compared to probable AMR and the control group without MVI (p = 0.012, p < 0.001). Conclusion Our results identify different molecular rejection phenotype patterns between cases with probable AMR, DSA-negative MVI, and AMR. Sub-threshold findings appear relevant along the AMR continuum.

Schachtner, Thomas; Weidmann, Lukas; Harmacek, Dusan; López, Kai C.; Korach, Raphael; Bortel, Nicola; Mueller, Thomas F. Author Information

Schachtner, Thomas; Moos, Seraina C. Von; López, Kai C.; Weidmann, Lukas; Harmacek, Dusan; Bortel, Nicola; Korach, Raphael; Mueller, Thomas F. Author Information

Schachtner, Thomas; López, Kai C.; Weidmann, Lukas; Harmacek, Dusan; Bortel, Nicola; Korach, Raphael; Mueller, Thomas F. Author Information

Schachtner, Thomas; Weidmann, Lukas; Harmacek, Dusan; Bortel, Nicola; López, Kai C.; Korach, Raphael; Mueller, Thomas F. Author Information