Banff 2022 Kidney Commentary: Reflections and Future Directions.

In September 2022, in Banff, Alberta, Canada, the XVIth Banff meeting, corresponding to the 30th anniversary of the Banff classification, was held, leading to 2 recent publications. Discussions at the Banff meeting focused on proposing improvements to the Banff process as a whole. In line with this, a unique opportunity was offered to a selected group of 16 representatives from the pathology and transplant nephrology community, experts in the field of kidney transplantation, to review these 2 Banff manuscripts. The aim was to provide an insightful commentary, to gauge any prospective influence the proposed changes may have, and to identify any potential areas for future enhancement within the Banff classification. The group expressed its satisfaction with the incorporation of 2 new entities, namely "microvascular inflammation/injury donor-specific antibodies-negative and C4d negative" and "probable antibody-mediated rejection," into category 2. These changes expand the classification, facilitating the capture of more biopsies and providing an opportunity to explore the clinical implications of these lesions further. However, we found that the Banff classification remains complex, potentially hindering its widespread utilization, even if a degree of complexity may be unavoidable given the intricate pathophysiology of kidney allograft pathology. Addressing the histomorphologic diagnosis of chronic active T cell-mediated rejection (CA TCMR), potentially reconsidering a diagnostic-agnostic approach, as for category 2, to inflammation in interstitial fibrosis and tubular atrophy and chronic active T cell-mediated rejection was also an important objective. Furthermore, we felt a need for more evidence before molecular diagnostics could be routinely integrated and emphasized the need for clinical and histologic context determination and the substantiation of its clinical impact through rigorous clinical trials. Finally, our discussions stressed the ongoing necessity for multidisciplinary decision-making regarding patient care.

The 2022 Banff Meeting Lung Report.

The Lung Session of the 2022 16th Banff Foundation for Allograft Pathology Conference-held in Banff, Alberta-focused on non-rejection lung allograft pathology and novel technologies for the detection of allograft injury. A multidisciplinary panel reviewed the state-of-the-art of current histopathologic entities, serologic studies, and molecular practices, as well as novel applications of digital pathology with artificial intelligence, gene expression analysis, and quantitative image analysis of chest computerized tomography. Current states of need as well as prospective integration of the aforementioned tools and technologies for complete assessment of allograft injury and its impact on lung transplant outcomes were discussed. Key conclusions from the discussion were: (1) recognition of limitations in current standard of care assessment of lung allograft dysfunction; (2) agreement on the need for a consensus regarding the standardized approach to the collection and assessment of pathologic data, inclusive of all lesions associated with graft outcome (eg, non-rejection pathology); and (3) optimism regarding promising novel diagnostic modalities, especially minimally invasive, which should be integrated into large, prospective multicenter studies to further evaluate their utility in clinical practice for directing personalized therapies to improve graft outcomes.

Emerging phenotypes in kidney transplant rejection.

PURPOSE OF REVIEW: This review focuses on more recently emerging rejection phenotypes in the context of time post transplantation and the resulting differential diagnostic challenges. It also discusses how novel ancillary diagnostic tools can potentially increase the accuracy of biopsy-based rejection diagnosis. RECENT FINDINGS: With advances in reducing immunological risk at transplantation and improved immunosuppression treatment renal allograft survival improved. However, allograft rejection remains a major challenge and represent a frequent course for allograft failure. With prolonged allograft survival, novel phenotypes of rejection are emerging, which can show complex overlap and transition between cellular and antibody-mediated rejection mechanisms as well as mixtures of acute/active and chronic diseases. With the emerging complexity in rejection phenotypes, it is crucial to achieve diagnostic accuracy in the individual patient. SUMMARY: The prospective validation and adoption of novel molecular and computational diagnostic tools into well defined and appropriate clinical context of uses will improve our ability to accurately diagnose, stage, and grade allograft rejection.

The Banff 2022 Kidney Meeting Report: Reappraisal of microvascular inflammation and the role of biopsy-based transcript diagnostics.

The XVI-th Banff Meeting for Allograft Pathology was held at Banff, Alberta, Canada, from 19th to 23rd September 2022, as a joint meeting with the Canadian Society of Transplantation. To mark the 30th anniversary of the first Banff Classification, premeeting discussions were held on the past, present, and future of the Banff Classification. This report is a summary of the meeting highlights that were most important in terms of their effect on the Classification, including discussions around microvascular inflammation and biopsy-based transcript analysis for diagnosis. In a postmeeting survey, agreement was reached on the delineation of the following phenotypes: (1) "Probable antibody-mediated rejection (AMR)," which represents donor-specific antibodies (DSA)-positive cases with some histologic features of AMR but below current thresholds for a definitive AMR diagnosis; and (2) "Microvascular inflammation, DSA-negative and C4d-negative," a phenotype of unclear cause requiring further study, which represents cases with microvascular inflammation not explained by DSA. Although biopsy-based transcript diagnostics are considered promising and remain an integral part of the Banff Classification (limited to diagnosis of AMR), further work needs to be done to agree on the exact classifiers, thresholds, and clinical context of use.

Deep learning-enabled realistic virtual histology with ultraviolet photoacoustic remote sensing microscopy.

The goal of oncologic surgeries is complete tumor resection, yet positive margins are frequently found postoperatively using gold standard H&E-stained histology methods. Frozen section analysis is sometimes performed for rapid intraoperative margin evaluation, albeit with known inaccuracies. Here, we introduce a label-free histological imaging method based on an ultraviolet photoacoustic remote sensing and scattering microscope, combined with unsupervised deep learning using a cycle-consistent generative adversarial network for realistic virtual staining. Unstained tissues are scanned at rates of up to 7 mins/cm2, at resolution equivalent to 400x digital histopathology. Quantitative validation suggests strong concordance with conventional histology in benign and malignant prostate and breast tissues. In diagnostic utility studies we demonstrate a mean sensitivity and specificity of 0.96 and 0.91 in breast specimens, and respectively 0.87 and 0.94 in prostate specimens. We also find virtual stain quality is preferred (P = 0.03) compared to frozen section analysis in a blinded survey of pathologists.

Perineural Invasion and Spread in Common Abdominopelvic Diseases: Imaging Diagnosis and Clinical Significance.

Malignancies and other diseases may spread by multiple pathways, including direct extension, hematogenous spread, or via lymphatic vessels. A less-well-understood route is the peripheral nervous system, which is known as perineural spread (PNS). In addition to accounting for pain and other neurologic symptoms, PNS affects both disease prognosis and management. Although PNS is commonly discussed in relation to head and neck tumors, there is emerging data regarding PNS in abdominopelvic malignancies and other conditions such as endometriosis. Due to improved contrast and spatial resolution, perineural invasion, a finding heretofore diagnosed only at pathologic examination, can be detected at CT, MRI, and PET/CT. PNS most commonly manifests as abnormal soft-tissue attenuation extending along neural structures, and diagnosis of it is aided by optimizing imaging parameters, understanding pertinent anatomy, and becoming familiar with the typical neural pathways of spread that largely depend on the disease type and location. In the abdomen, the celiac plexus is a central structure that innervates the major abdominal organs and is the principal route of PNS in patients with pancreatic and biliary carcinomas. In the pelvis, the lumbosacral plexus and inferior hypogastric plexus are the central structures and principal routes of PNS in patients with pelvic malignancies. Although the imaging findings of PNS may be subtle, a radiologic diagnosis can have a substantial effect on patient care. Knowledge of anatomy and known routes of PNS and optimizing imaging parameters is of utmost importance in providing key information for prognosis and treatment planning. © RSNA, 2023 Supplemental material and the slide presentation from the RSNA Annual Meeting are available for this article. Quiz questions for this article are available through the Online Learning Center.

Quantitative scoring of progression in transplant glomerulopathy using digital pathology may be superior to Banff cg scoring.

Antibody-mediated rejection (ABMR) is a major cause of kidney allograft failure. Biopsy-based surrogate endpoints reflecting ABMR progression on sequential biopsies that predict long-term outcome offer the potential to make treatment trials for ABMR feasible. However, the Banff transplant glomerulopathy (TG) scoring system (chronic glomerular injury score [cg]) relies on relatively crude and arbitrary ordinal grades and has low inter-observer concordance that currently limits its usefulness as a surrogate endpoint for ABMR progression in clinical drug trials. Here, we describe and validate a novel quantitative method for quantifying progression of TG in ABMR. Using digital pathology in sequential biopsies from 75 patients at various stages of ABMR, we scored all capillaries in the most affected glomeruli for basement membrane duplication that were correlated with allograft function, outcome, Banff lesion scores, and gene expression. Our digital scoring reflected TG progression better than the categorical Banff cg score and correlated with Banff ABMR and chronicity lesions, but not transcript changes. In multivariate analysis, the delta change between biopsies with serum creatinine and mean percent duplicated glomerular basement membranes was significantly associated with graft loss. Neither the delta in any Banff lesion scores (including cg) nor in gene expression was associated with outcome. Receiver operating characteristic curve analysis showed that the digital pathology approach was superior to the conventional score for predicting graft failure. Thus, our digital pathology-based approach for scoring TG accurately assessed progression in TG. However, further validation as a potential surrogate endpoint in clinical trials for the treatment of ABMR is warranted.

Fast hybrid optomechanical scanning photoacoustic remote sensing microscopy for virtual histology.

A rapid scanning microscopy method for hematoxylin and eosin (H&E) like images is sought after for interoperative diagnosis of solid tumor margins. The rapid observation and diagnosis of histological samples can greatly lower surgical risk and improve patient outcomes from solid tumor resection surgeries. Photoacoustic remote sensing (PARS) has recently been demonstrated to provide images of virtual H&E stains with excellent concordance with true H&E staining of formalin-fixed, paraffin embedded tissues. By using PARS with constant velocity and 1D galvanometer mirror scanning we acquire large virtual H&E images (10mm x 5mm) of prostate tissue in less than 3.5 minutes without staining, and over two orders of magnitude faster data acquisition than the current PARS imaging speed.

Bilateral Lung Transplant for Treatment of Idiopathic Pulmonary Fibrosis With Undetected Lung Adenocarcinoma: A Case Report.

BACKGROUND: Lung transplant has become a curative therapy for various forms of progressive lung disease refractory to medical management. Idiopathic pulmonary fibrosis (IPF) is a rare condition characterized by accumulation of activated fibroblasts and secretion of extracellular matrices within the lung parenchyma. End-stage IPF is a fatal condition, with limited medical therapies other than lung transplantation. IPF has been demonstrated as a known risk factor for the development of lung cancer, and current lung transplant standards define history of malignancy within the past five years as an absolute exclusion criterion. CASE REPORT: We present the case of a patient with biopsy-confirmed idiopathic pulmonary fibrosis treated with bilateral lung transplant, discovered to have stage four lung adenocarcinoma in the explanted lungs. The patient subsequently received pseudoadjuvant chemotherapy and remained recurrence-free until 23 months post-transplant. CONCLUSION: This case highlights the challenge of ruling out malignancy in patients with end-stage lung disease. There remains a paucity of clinical studies on lung transplantation for lung cancer and more evidence is required before supporting this clinical decision.

Risk stratification of gastrointestinal stromal tumors by Nanostring gene expression profiling.

PURPOSE: The risk assessment classification schemes for gastrointestinal stromal tumors (GIST) include tumor site, size, mitotic count and variably tumor rupture. Heterogeneity in high-risk GIST poses limitations for current classification schemes. This study aims to demonstrate the clinical utility of risk stratification by gene expression profiling (GEP) using Nanostring technology. METHODS: Fifty-six GIST cases were analyzed using a 231 gene expression panel. GEP results were correlated with clinical and pathological data. The prognostic performance was assessed in 34 patients with available survival data using ROC curves, Kaplan-Meier survival curves and compared with traditional risk assessment schemes. Volcano plot analysis identified seven genes with significantly higher expression (FDR < .0.05) in high-risk than in non-high-risk tumors, namely TYMS, CDC2, TOP2A, CCNA2, E2F1, PCNA, and BIRC5. Together, these transcripts exhibited significantly higher expression in high-risk tumors than in intermediate (P < 0.01), low (P < 0.001), and very low (P = 0.01) risk tumors. Receiver-operating characteristic curve analysis demonstrated area under the curve (AUC) to be 0.858 for the separation of high-risk and non-high-risk tumors. Kaplan-Meier survival analysis demonstrated improved risk stratification (log-rank test P < 0.001) compared to the current risk assessment classification (P = 0.231). CONCLUSION: In addition to current clinical and histology-based risk classification for patients with GIST, gene expression may offer complementary prognostic information.

Diffuse C4d staining of peritubular capillaries in renal allograft following bamlanivimab therapy.

Neutralizing monoclonal antibodies such as bamlanivimab emerged as promising agents in treating kidney transplant recipients with COVID-19. However, the impact of bamlanivimab on kidney allograft histology remains unknown. We report a case of a kidney transplant recipient who received bamlanivimab for COVID-19 with subsequent histologic findings of diffuse peritubular capillary C4d staining. A 33-year-old man with end-stage kidney disease secondary to hypertension who received an ABO compatible kidney from a living donor, presented for his 4-month protocol visit. He was diagnosed with COVID-19 44 days prior to his visit and had received bamlanivimab with an uneventful recovery. His 4-month surveillance biopsy showed diffuse C4d staining of the peritubular capillaries without other features of antibody-mediated rejection (ABMR). Donor-specific antibodies were negative on repeat evaluations. ABMR gene expression panel was negative. His creatinine was stable at 1.3 mg/dl, without albuminuria. Given the temporal relationship between bamlanivimab and our observations of diffuse C4d staining of the peritubular capillaries, we hypothesize that bamlanivimab might bind to angiotensin-converting enzyme 2, resulting in classical complement pathway and C4d deposition. We elected to closely monitor kidney function which has been stable at 6 months after the biopsy. In conclusion, diffuse C4d may present following bamlanivimab administration without any evidence of ABMR.

Acute Kidney Injury in Severe COVID-19 Has Similarities to Sepsis-Associated Kidney Injury: A Multi-Omics Study.

OBJECTIVE: To compare coronavirus disease 2019 (COVID-19) acute kidney injury (AKI) to sepsis-AKI (S-AKI). The morphology and transcriptomic and proteomic characteristics of autopsy kidneys were analyzed. PATIENTS AND METHODS: Individuals 18 years of age and older who died from COVID-19 and had an autopsy performed at Mayo Clinic between April 2020 to October 2020 were included. Morphological evaluation of the kidneys of 17 individuals with COVID-19 was performed. In a subset of seven COVID-19 cases with postmortem interval of less than or equal to 20 hours, ultrastructural and molecular characteristics (targeted transcriptome and proteomics analyses of tubulointerstitium) were evaluated. Molecular characteristics were compared with archived cases of S-AKI and nonsepsis causes of AKI. RESULTS: The spectrum of COVID-19 renal pathology included macrophage-dominant microvascular inflammation (glomerulitis and peritubular capillaritis), vascular dysfunction (peritubular capillary congestion and endothelial injury), and tubular injury with ultrastructural evidence of mitochondrial damage. Investigation of the spatial architecture using a novel imaging mass cytometry revealed enrichment of CD3+CD4+ T cells in close proximity to antigen-presenting cells, and macrophage-enriched glomerular and interstitial infiltrates, suggesting an innate and adaptive immune tissue response. Coronavirus disease 2019 AKI and S-AKI, as compared to nonseptic AKI, had an enrichment of transcriptional pathways involved in inflammation (apoptosis, autophagy, major histocompatibility complex class I and II, and type 1 T helper cell differentiation). Proteomic pathway analysis showed that COVID-19 AKI and to a lesser extent S-AKI were enriched in necroptosis and sirtuin-signaling pathways, both involved in regulatory response to inflammation. Upregulation of the ceramide-signaling pathway and downregulation of oxidative phosphorylation in COVID-19 AKI were noted. CONCLUSION: This data highlights the similarities between S-AKI and COVID-19 AKI and suggests that mitochondrial dysfunction may play a pivotal role in COVID-19 AKI. This data may allow the development of novel diagnostic and therapeutic targets.

Gene Expression Profiling in Kidney Transplants with Immune Checkpoint Inhibitor-Associated Adverse Events.

BACKGROUND AND OBJECTIVES: Immune checkpoint inhibitors are increasingly used to treat various malignancies, but their application in patients with kidney transplants is complicated by high allograft rejection rates. Immune checkpoint inhibitor-associated rejection is a novel, poorly understood entity demonstrating overlapping histopathologic features with immune checkpoint inhibitor-associated acute interstitial nephritis, which poses a challenge for diagnosis and clinical management. We sought to improve the understanding of these entities through biopsy-based gene expression analysis. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: NanoString was used to measure and compare the expression of 725 immune-related genes in 75 archival kidney biopsies, including a 25-sample discovery cohort comprising pure T cell-mediated rejection and immune checkpoint inhibitor-associated acute interstitial nephritis and an independent 50-sample validation cohort comprising immune checkpoint inhibitor-associated acute interstitial nephritis, immune checkpoint inhibitor-associated T cell-mediated rejection, immune checkpoint inhibitor-associated crescentic GN, drug-induced acute interstitial nephritis, BK virus nephropathy, and normal biopsies. RESULTS: Significant molecular overlap was observed between immune checkpoint inhibitor-associated acute interstitial nephritis and T cell-mediated rejection. Nevertheless, IFI27, an IFN-α-induced transcript, was identified and validated as a novel biomarker for differentiating immune checkpoint inhibitor-associated T cell-mediated rejection from immune checkpoint inhibitor-associated acute interstitial nephritis (validation cohort: P<0.001, area under the receiver operating characteristic curve =100%, accuracy =86%). Principal component analysis revealed heterogeneity in inflammatory gene expression patterns within sample groups; however, immune checkpoint inhibitor-associated T cell-mediated rejection and immune checkpoint inhibitor-associated acute interstitial nephritis both demonstrated relatively more molecular overlap with drug-induced acute interstitial nephritis than T cell-mediated rejection, suggesting potential dominance of hypersensitivity mechanisms in these entities. CONCLUSIONS: These results indicate that, although there is significant molecular similarity between immune checkpoint inhibitor-associated rejection and acute interstitial nephritis, biopsy-based measurement of IFI27 gene expression represents a potential biomarker for differentiating these entities.

Intragraft gene expression in native kidney BK virus nephropathy versus T cell-mediated rejection: Prospects for molecular diagnosis and risk prediction.

Novel tools are needed to improve diagnostic accuracy and risk prediction in BK virus nephropathy (BKVN). We assessed the utility of intragraft gene expression testing for these purposes. Eight hundred genes were measured in 110 archival samples, including a discovery cohort of native kidney BKVN (n = 5) vs pure T cell-mediated rejection (TCMR; n = 10). Five polyomavirus genes and seven immune-related genes (five associated with BKVN and two associated with TCMR) were significantly differentially expressed between these entities (FDR < 0.05). These three sets of genes were further evaluated in samples representing a spectrum of BK infection (n = 25), followed by a multicenter validation cohort of allograft BKVN (n = 60) vs TCMR (n = 10). Polyomavirus 5-gene set expression reliably distinguished BKVN from TCMR (validation cohort AUC = 0.992), but the immune gene sets demonstrated suboptimal diagnostic performance (AUC ≤ 0.720). Within the validation cohort, no significant differences in index biopsy gene expression were identified between BKVN patients demonstrating resolution (n = 35), persistent infection (n = 14) or de novo rejection (n = 11) 6 months following a standardized reduction in immunosuppression. These results suggest that, while intragraft polyomavirus gene expression may be useful as an ancillary diagnostic for BKVN, assessment for concurrent TCMR and prediction of clinical outcome may not be feasible with current molecular tools.

Clinicopathologic predictors of renal outcomes in light chain cast nephropathy: a multicenter retrospective study.

Light chain cast nephropathy (LCCN) in multiple myeloma often leads to severe and poorly reversible acute kidney injury. Severe renal impairment influences the allocation of chemotherapy and its tolerability; it also affects patient survival. Whether renal biopsy findings add to the clinical assessment in predicting renal and patient outcomes in LCCN is uncertain. We retrospectively reviewed clinical presentation, chemotherapy regimens, hematologic response, and renal and patient outcomes in 178 patients with biopsy-proven LCCN from 10 centers in Europe and North America. A detailed pathology review, including assessment of the extent of cast formation, was performed to study correlations with initial presentation and outcomes. Patients presented with a mean estimated glomerular filtration rate (eGFR) of 13 ± 11 mL/min/1.73 m2, and 82% had stage 3 acute kidney injury. The mean number of casts was 3.2/mm2 in the cortex. Tubulointerstitial lesions were frequent: acute tubular injury (94%), tubulitis (82%), tubular rupture (62%), giant cell reaction (60%), and cortical and medullary inflammation (95% and 75%, respectively). Medullary inflammation, giant cell reaction, and the extent of cast formation correlated with eGFR value at LCCN diagnosis. During a median follow-up of 22 months, mean eGFR increased to 43 ± 30 mL/min/1.73 m2. Age, ß2-microglobulin, best hematologic response, number of cortical casts per square millimeter, and degree of interstitial fibrosis/tubular atrophy (IFTA) were independently associated with a higher eGFR during follow-up. This eGFR value correlated with overall survival, independently of the hematologic response. This study shows that extent of cast formation and IFTA in LCCN predicts the quality of renal response, which, in turn, is associated with overall survival.

Apelin directs endothelial cell differentiation and vascular repair following immune-mediated injury.

Sustained, indolent immune injury of the vasculature of a heart transplant limits long-term graft and recipient survival. This injury is mitigated by a poorly characterized, maladaptive repair response. Vascular endothelial cells respond to proangiogenic cues in the embryo by differentiation to specialized phenotypes, associated with expression of apelin. In the adult, the role of developmental proangiogenic cues in repair of the established vasculature is largely unknown. We found that human and minor histocompatibility-mismatched donor mouse heart allografts with alloimmune-mediated vasculopathy upregulated expression of apelin in arteries and myocardial microvessels. In vivo, loss of donor heart expression of apelin facilitated graft immune cell infiltration, blunted vascular repair, and worsened occlusive vasculopathy in mice. In vitro, an apelin receptor agonist analog elicited endothelial nitric oxide synthase activation to promote endothelial monolayer wound repair and reduce immune cell adhesion. Thus, apelin acted as an autocrine growth cue to sustain vascular repair and mitigate the effects of immune injury. Treatment with an apelin receptor agonist after vasculopathy was established markedly reduced progression of arterial occlusion in mice. Together, these initial data identify proangiogenic apelin as a key mediator of coronary vascular repair and a pharmacotherapeutic target for immune-mediated injury of the coronary vasculature.

Long-term Kinetics of Intragraft Gene Signatures in Renal Allograft Tolerance Induced by Transient Mixed Chimerism.

BACKGROUND: Renal allograft tolerance (TOL) has been successfully induced in nonhuman primates (NHPs) and humans through the induction of transient mixed chimerism. To elucidate the mechanisms of TOL, we compared local immunologic responses in renal allografts with those in T-cell-mediated rejection (TCMR) and chronic antibody-mediated rejection (CAMR) in NHPs. METHODS: Using the NanoString nCounter platform, we retrospectively studied 52 mRNAs in 256 kidney allograft samples taken from NHP kidney recipients of donor BMT. No immunosuppression was given after 1-month post-donor BMT. Recipients who achieved TOL (n = 13) survived for >1840 ± 1724 days with normal kidney function, while recipients with CAMR (n = 13) survived for 899 ± 550 days with compromised graft function, and recipients with TCMR (n = 15) achieved only short-term survival (132 ± 69 days). RESULTS: The most prominent difference between the groups was FOXP3, which was significantly higher in TOL than in CAMR and TCMR, both early (<1 y, P < 0.01) and late (≥1 y, P < 0.05) after transplant. Other mRNAs related to regulatory T cells (Treg), such as IL10, TGFB, and GATA3, were also high in TOL. In contrast, transcripts of inflammatory cytokines were higher in TCMR, while activated endothelium-associated transcripts were higher in CAMR than in TOL. The receiver operating characteristic analyses revealed that intragraft FOXP3 and CAV1 can reliably distinguish TOL from CAMR. CONCLUSIONS: High FOXP3 and other Treg-related mRNAs together with suppressed inflammatory responses and endothelial activation in renal allografts suggest that intragraft enrichment of Treg is a critical mechanism of renal allograft TOL induced by transient mixed chimerism.