Update on diagnostic and therapeutic strategies for antibody-mediated rejection in kidney transplantation

Table 1 Chronology of antibody mediated rejection advancements and defining developments

Year	Key developments
1964[11]	Recognition of immediate kidney rejection caused by pre-existing antibodies
1970[12]	Emergence of obliterative vascular lesions in recipients with de novo DSA, leading to poor transplant outcomes
1990[13]	Characterization of AMR through a three-factor framework: (1) Impaired graft function; (2) Presence of neutrophils in peritubular capillaries; and (3) Detection of class I HLA antibodies, independent of conventional T-cell-mediated rejection
1993[14]	Confirmation of the role of DSA in rejection with C4d deposition in PTC
1997[15]	Banff consensus establishes histological guidelines for kidney transplant biopsies: AMR is determined as rejection partially or fully attributed to anti-donor antibodies, categorized as hyperacute (immediate) or accelerated acute (delayed reaction)
2001[16-17]	Classification of AMR. Chronic rejection is defined as progressive deterioration of renal function, marked by hypertension and proteinuria beyond three months post-transplant: C4d staining in PTC distinguished AMR from non-specific CAN. Cases lacking detectable antibodies are classified as “suspicious for AMR.”
2005[18]	Banff classification revised to replace CAN with chronic active AMR. Discovery of AT1R-activating antibodies. Isolated tubulitis under the borderline category
2007[19]	Banff developed grading for PTC inflammation, Focal C4d, C4d scoring, and interpretation of C4d deposition absent overt histological rejection. ti score. Grading of zero-time and protocol biopsies
2009	Subclinical AMR. Creation of Banff Working Groups
2013[20]	Major revision of AMR definition: C4d staining is no longer an absolute necessity; microvascular inflammation or validated GEP is used as an additional diagnostic marker. Cg1a scoring. HLA DSA testing by SA
2015	Specific reports for each organ. i-IFTA. Role of non-HLA-DSA
2017[21]	Banff incorporates C4d staining and molecular classifiers as surrogate indicators of DSA presence. AMR without anti-HLA DSA
2019[22]	Introduction of machine learning-based FFPE-based molecular diagnostics to refine AMR classification. FFPE-based molecular diagnostics
2021	Banff incorporated multi-omics approaches (genomics, transcriptomics, proteomics) to improve AMR detection
2023	Artificial intelligence-assisted digital histopathology was introduced for automated grading of AMR severity. Routine molecular diagnostics as a “companion criterion standard.” Open source analytic pipeline. Machine learning based biopsy contextualization
2025[23-25]	Banff further refines AMR classification with four advancements: (1) C4d and molecular classifiers remain key markers; (2) Single-cell RNA sequencing aids in identifying early AMR signatures; (3) Non-invasive biomarkers (circulating donor-derived cell-free DNA) validated for AMR monitoring; and (4) Personalized immunosuppressive strategies based on molecular profiling introduced

DSA: Donor-specific antibody; AMR: Antibody mediated rejection; HLA: Human leucocyte antigen; PTC: Peritubular capillary; CAN: Chronic allograft nephropathy; AT1R: Angiotensin receptor; GEP: Gene expression profiling; IFTA: Interstitial fibrosis and tubular atrophy; Cg: Chronic glomerulopathy; FFPE: Formalin-fixed paraffin- embedded; SA: Single antigen.

Table 2 Antibody-mediated rejection spectrum according to Banff 2019 classification report

Criterion	Active AMR	Chronic active AMR	Chronic (inactive) AMR
Criterion 1: Histopathology	Acute tissue injury, including one or more of the following: (1) MIV (glomerulitis and/or ptc) in the absence of recurrent or de-novo GN, intimal or transmural arteritis; (2) Acute TMA, in the absence of any other apparent cause; and (3) ATI, in the absence of any other apparent cause	Chronic tissue injury, including one or more of the following: (1) TG (GBM duplication in the absence of subendothelial immune complex deposits) if no evidence of chronic TMA in the absence of recurrent or de-novo glomerulonephritis; (2) Severe peritubular capillary basement membrane multilayering (requires EM); and (3) Transplant arteriopathy (arterial intimal fibrosis of new onset) and mild to moderate acute tissue injury (MIV)	Chronic tissue injury: (1) TG, and/or severe ptc basement membrane multilayering (requires EM) without MVI; and (2) Significant loss of ptc (capillaries simply no longer exist to show capillaritis)
Criterion 2: Evidence of antibody interaction with the endothelium	These include at least one of the following: (1) C4d deposition in ptc, OR; (2) At least moderate MIV, OR; and (3) Increased expression of gene transcripts/classifiers in the biopsy tissue	C4d deposition in ptc, or at least moderate MIV, or increased expression of gene transcripts/classifiers in the biopsy tissue	C4d negative
			There may be prior evidence of antibody interaction with the endothelium
Criterion 3: Serologic evidence of DSAs (HLA or other antigens)	Detectable serum anti-HLA DSA	Detectable serum anti-HLA DSA	Anti-HLA DSA may be undetectable
	If anti-HLA DSA is undetectable, non-HLA antibody testing	If anti-HLA DSA is undetectable, non-HLA antibody testing	However, there should be prior evidence of anti-HLA or non-HLA DSA
	C4d staining or expression of validated transcripts/classifiers may substitute for DSA	C4d staining or expression of validated transcripts/classifiers may substitute for DSA

AMR: Antibody-mediated rejection; ptc: Peritubular capillary; GN: Glomerulonephritis; TMA: Thrombotic microangiopathy; ATI: Acute tubular injury; TG: Transplant glomeruopathy; GBM: Glomerular basement membrane; EM: Electron microscope; HLA: Human leukocyte antigen; DSA: Donor-specific antibody; MVI: Microvascular inflammation.

Table 3 Limitations of biomarkers and their validation status

Biomarker/test	PPV	NPV	Estimated cost	Validation status
dd-cfDNA (> 1%)	61%	84%	About 2800 dollars per test	FDA-approved; medicare reimbursed
kSORT (17-gene PCR panel)	92% (sensitivity)	93% (specificity)	Not commercially available	Research-stage; limited clinical use
TruGraf v1	Combined PPV/NPV with dd-cfDNA	Improved detection	About 2000-2500 dollars per test	Validated in stable graft recipients
Peripheral blood GEP	47%	82%	About 1500-2000 dollars	Clinically available; moderate uptake
ELISPOT (IFN-γ)	Variable	Variable	High (labor-intensive)	Limited scalability; research use
Urinary CXCL10	Moderate	Moderate	Low (about 100-300 dollars)	Research-stage; promising results
Urinary miRNAs	Variable	Variable	Moderate (about 500-1000 dollars)	Experimental; under validation
MMDx	64%	91%	About 3000-4000 dollars per biopsy	Validated; used in specialized centers
Banff transcript panel	Not yet published	Not yet published	TBD	Under evaluation by Banff committee

PPV: Positive predictive value; NPV: Negative predictive value; cfDNA: Cell free DNA; FDA: Food and Drug Administration; kSORT: Kidney Solid Organ Response Test; PCR: Polymerase chain reaction; GEP: Gene expression profiling; ELISPOT: Enzyme linked immuno spot; IFN: Interferon; CXCL10: C-X-C motif chemokine 10; miRNAs: MicroRNAs; MMDx: Molecular Microscope Diagnostic System; TBD: To be determined.

Table 4 Comparison of clinical utility of blood vs urine biomarkers in kidney transplant monitoring

Feature	dd-cfDNA	Urinary CXCL10
Sample type	Peripheral blood	Urine
Target	Fragmented donor DNA from injured graft	Chemokine linked to immune activation
Clinical role	Detects active graft injury and acute rejection	Predicts and monitors acute rejection, especially AMR
Sensitivity/specificity	About 59% sensitivity, about 85% specificity at 1% threshold	High NPV for ruling out rejection
PPV/NPV	PPV: 61%, NPV: 84%	NPV: High; PPV varies
Timing of detection	Early detection due to short half-life (about 30 minutes)	Reflects ongoing inflammation; may lag behind dd-cfDNA
Regulatory status	FDA-approved; reimbursed by Medicare	Research-stage; not yet standardized
Cost	About 2800 dollars per test	About 100-300 dollars per test
Advantages	High specificity; validated in multicenter trials	Noninvasive, inexpensive, easy to collect repeatedly
Limitations	Costly; may be affected by other injuries	Limited standardization; influenced by urine concentration
Use in subclinical rejection	Useful when combined with other assays (e.g., TruGraf)	Promising but less validated for subclinical cases

dd-cfDNA: Donor-derived cell-free DNA; CXCL10: C-X-C motif chemokine 10; AMR: Antibody mediated rejection; NPV: Negative predictive value; PPV: Positive predictive value; FDA: Food and Drug Administration.

Table 5 Approach to prevent the development of antibody-mediated rejection in patients with preexisting donor-specific antibodies

Type of donor	Type of crossmatch	Pre-transplant treatment	Induction and maintenance immunosuppression therapies	Monitoring after transplant
Patients with a potential living donor	Patients with a positive CDC crossmatch or a strongly positive flow crossmatch	Prefer to use KPD programs[74], rather than desensitization[75]	Appropriate for patients at high risk for the development of acute rejection (plasmapharesis/IVIG and glucocorticoids[77]	Routinely monitor DSA levels at months 1, 3, 6, and 12 post-transplant and then annually
			Patients with positive C4d staining, plasmapheresis (two to three sessions), IVIG, and a single dose of rituximab 375 mg/m would be added	Perform kidney allograft biopsies in all patients who develop a de novo DSA
	Patients with a positive virtual crossmatch1 or a mild to moderate flow crossmatch2	Employ HLA desensitization strategies3[76]	Maintenance on a triple therapy4	-
Patients without a potential living donor	-	Employ HLA desensitization strategies strategies3	-	-

¹Antibodies detected by single antigen bead technology.

²Median channel shift of < 200.

³Include treatment with plasmapheresis, recombinant antithymocyte globulin-thymoglobulin, and rituximab.

⁴Regimen include tacrolimus, mycophenolate, and prednisone.

CDC: Complement-dependent cytotoxicity; KPD: Kidney paired donation; IVIG: Intravenous immune globulin; DSA: Donor-specific antibody; HLA: Human leukocyte antigen.

Table 6 The Transplantation Society 2019 consensus-based treatment recommendations in alignment with the Kidney Disease: Improving Global Outcomes 2023 guidelines for the treatment of antibody-mediated rejection

2019 consensus	DSA	Banff 2017 phenotypes	Standard of care	Consider adjunctive therapies	KDIGO 2023 updates
Early acute (< 30 days)	Preexisting DSA (or nonimmunologically naive)	Active AMR	PP/IVIG1 glucocorticoids	Complement	Eculizumab use is restricted to select cases with complement activation; not routine. KDIGO recommends early biopsy confirmation, dd-cfDNA and GEP monitoring, and tailored immunosuppression
				Inhibitor (e.g., eculizumab)
				Rituximab3
				Splenectomy
	Preexisting DSA	Active AMR	PP/IVIG1 glucocorticoids	Rituximab3	KDIGO advises against routine complement blockade unless complement-mediated injury is proven
Late (> 30 days)	De novo DSA	Chronic AMR	Optimize baseline immunosuppression2	-	KDIGO supports noninvasive monitoring (e.g., dd-cfDNA, GEP), and individualized therapy based on graft function
		Active AMR	Optimize baseline immunosuppression2	PP, IVIG	KDIGO encourages molecular diagnostics (e.g., MMDx) and consideration of adherence issues
				Rituximab
		Chronic AMR	Evaluate and manage nonadherence	IVIG	KDIGO highlights importance of patient education, adherence tracking, and long-term graft surveillance

¹Daily or alternate day plasmapheresis sessions × 6 based on donor-specific antibody titer; intravenous immunoglobulin 100 mg/kg after each plasmapheresis treatment or intravenous immunoglobulin 2/kg at the end of plasmapheresis treatments.

²Includes the use of tacrolimus with the goal through levels of > 5 ng/mL and use of maintenance steroid equivalent to prednisone 5 mg/day.

³If age < 70 years, estimated glomerular filtration rate ≥ 20 mL/minute/1.73 m³, lower chronicity scores on biopsy (interstitial fibrosis + tubular atrophy + fibrous intimal thickening + allograft glomerulopathy < 8) and evidence of severe disease [higher donor-specific antibody, diffuse C4d staining, or more extensive microvascular inflammation (i.e., glomerulitis score + peritubular capillary score ≥ 4) on biopsy].

AMR: Antibody-mediated rejection; DSA: Donor-specific antibody; IVIG: Intravenous immunoglobulin; PP: Plasmapheresis; KDIGO: Kidney Disease: Improving Global Outcomes; dd-cfDNA: Donor-derived cell-free DNA; GEP: Gene expression profiling; MMDx: Molecular Microscope Diagnostic System.

Table 7 Evidence-based comparison of acute vs chronic antibody-mediated rejection therapies

Therapy	Use in acute AMR	Use in chronic AMR	Evidence summary	KDIGO 2023 position
PP	First-line for antibody removal; often combined with IVIG	Used selectively; less effective in chronic injury	Supported by multiple case series and consensus guidelines	Recommended in acute AMR; limited role in chronic AMR
IVIG	Adjunct to PP; modulates immune response	Used in chronic AMR with DSA presence	Moderate evidence; variable dosing strategies	Supported in both acute and chronic AMR
Rituximab	Targets CD20+ B cells; used in combination with PP/IVIG	Sometimes used in chronic AMR with active inflammation	Mixed results; better efficacy in early AMR	Considered in both settings; not universally effective
Eculizumab	Complement inhibition in severe or refractory acute AMR	Not recommended for routine chronic AMR	Effective in complement-mediated AMR; high cost, limited trials	Restricted use: Only in complement-driven AMR
Bortezomib	Used in refractory acute AMR; targets plasma cells	Limited efficacy in chronic AMR; poor outcomes in late-stage fibrosis	Early studies showed promise; later trials failed to show consistent benefit[83]	Not recommended for chronic AMR; use in acute AMR remains investigational
Tocilizumab (IL-6 blocker)	Investigational; targets inflammatory cytokine pathways	Emerging therapy in chronic AMR	Phase 2/3 trials ongoing; promising results in chronic inflammation	Under evaluation; not yet standard of care
CD38 antibodies (e.g., felzartamab)	Targets long-lived plasma cells and NK cells	Promising in chronic AMR with persistent DSA	Recent trials show reversal of AMR activity	KDIGO supports further research; not yet routine
Molecular diagnostics (MMDx, dd-cfDNA, GEP)	Used to confirm and monitor acute AMR	Valuable for detecting subclinical chronic AMR	High NPV; improves diagnostic precision	Strongly recommended for both acute and chronic AMR monitoring

PP: Plasmapheresis; IVIG: Intravenous immunoglobulin; IL: Interleukin; MMDx: Molecular Microscope Diagnostic System; dd-cfDNA: Donor-derived cell-free DNA; GEP: Gene expression profiling; AMR: Antibody-mediated rejection; DSA: Donor-specific antibody; NK: Natural killer; NPV: Negative predictive value; KDIGO: Kidney Disease: Improving Global Outcomes.