The classical pathway (CP) of complement may contribute to the pathogenesis of antibody‐mediated rejection (ABMR). Selective CP blockade may be a promising strategy to counteract rejection. The ...objective of this first‐in‐patient phase 1b trial was to evaluate the safety/tolerability and CP‐blocking potential of 4 weekly doses (60 mg/kg) of the anti‐C1s antibody BIVV009 in complement‐mediated disorders. Here we describe the results in a cohort of 10 stable kidney transplant recipients (median of 4.3 years posttransplantation) with late active ABMR and features of CP activation, such as capillary C4d or complement‐fixing donor‐specific antibodies (DSA). During 7 weeks follow‐up, no severe adverse events were reported, and BIVV009 profoundly inhibited overall and DSA‐triggered CP activation in serum. Five of 8 C4d‐positive recipients turned C4d‐negative in 5‐week follow‐up biopsies, while another 2 recipients showed a substantial decrease in C4d scores. There was, however, no change in microcirculation inflammation, gene expression patterns, DSA levels, or kidney function. In conclusion, we demonstrate that BIVV009 effectively blocks alloantibody‐triggered CP activation, even though short‐course treatment had no effect on indices of activity in late ABMR. This initial trial provides a valuable basis for future studies designed to clarify the therapeutic value of CP blockade in transplantation. ClinicalTrials.gov NCT#02502903.
This first‐in‐patient phase 1 trial demonstrates safety, tolerability, and profound complement inhibition efficacy of a novel humanized anti‐C1s monoclonal antibody in a cohort of 10 kidney allograft recipients with late antibody‐ mediated rejection.
The 10th Banff Conference on Allograft Pathology was held in Banff, Canada from August 9 to 14, 2009. A total of 263 transplant clinicians, pathologists, surgeons, immunologists and researchers ...discussed several aspects of solid organ transplants with a special focus on antibody mediated graft injury. The willingness of the Banff process to adapt continuously in response to new research and improve potential weaknesses, led to the implementation of six working groups on the following areas: isolated v‐lesion, fibrosis scoring, glomerular lesions, molecular pathology, polyomavirus nephropathy and quality assurance. Banff working groups will conduct multicenter trials to evaluate the clinical relevance, practical feasibility and reproducibility of potential changes to the Banff classification. There were also sessions on quality improvement in biopsy reading and utilization of virtual microscopy for maintaining competence in transplant biopsy interpretation. In addition, compelling molecular research data led to the discussion of incorporation of omics‐technologies and discovery of new tissue markers with the goal of combining histopathology and molecular parameters within the Banff working classification in the near future.
The 10th Banff Conference on Allograft Pathology held in Banff, Canada August 9–14, 2009 specifically focused on antibody mediated graft injury, and resulted in implementation of six Banff working groups to evaluate clinical relevance and reproducibility of potential changes to the Banff classification.
The 9th Banff Conference on Allograft Pathology was held in La Coruna, Spain on June 23–29, 2007. A total of 235 pathologists, clinicians and scientists met to address unsolved issues in ...transplantation and adapt the Banff schema for renal allograft rejection in response to emerging data and technologies. The outcome of the consensus discussions on renal pathology is provided in this article. Major updates from the 2007 Banff Conference were: inclusion of peritubular capillaritis grading, C4d scoring, interpretation of C4d deposition without morphological evidence of active rejection, application of the Banff criteria to zero‐time and protocol biopsies and introduction of a new scoring for total interstitial inflammation (ti‐score). In addition, emerging research data led to the establishment of collaborative working groups addressing issues like isolated ‘v’ lesion and incorporation of omics‐technologies, paving the way for future combination of graft biopsy and molecular parameters within the Banff process.
The Ninth Banff Conference on Allograft Pathology held in La Coruna, Spain June 23‐29, 2007 resulted in new developments pertaining to C4d staining, peritubular capillaritis, protocol biopsies, lesion scoring, and eventual incorporation of “omics” technologies into kidney transplant rejection diagnosis.
B7.1/2‐targeted costimulation blockade (CTLA4 immunoglobulin CTLA4‐Ig) is available for immunosuppression after kidney transplantation, but its potentially detrimental impact on regulatory T cells ...(Tregs) is of concern. We investigated the effects of CTLA4‐Ig monotherapy in a fully mismatched heart transplant model (BALB/c onto C57BL/6). CTLA4‐Ig was injected chronically (on days 0, 4, 14, and 28 and every 4 weeks thereafter) in dosing regimens paralleling clinical use, shown per mouse: low dose (LD), 0.25 mg (≈10 mg/kg body weight); high dose (HD), 1.25 mg (≈50 mg/kg body weight); and very high dose (VHD), 6.25 mg (≈250 mg/kg body weight). Chronic CTLA4‐Ig therapy showed dose‐dependent efficacy, with the LD regimen prolonging graft survival and with the HD and VHD regimens leading to >95% long‐term graft survival and preserved histology. CTLA4‐Ig's effect was immunosuppressive rather than tolerogenic because treatment cessation after ≈3 mo led to rejection. FoxP3‐positive Tregs were reduced in naïve mice to a similar degree, independent of the CTLA4‐Ig dose, but recovered to normal values in heart recipients under chronic CTLA4‐Ig therapy. Treg depletion (anti‐CD25) resulted in an impaired outcome under LD therapy but had no detectable effect under HD therapy. Consequently, the immunosuppressive effect of partially effective LD CTLA4‐Ig therapy is impaired when Tregs are removed, whereas CTLA4‐Ig monotherapy at higher doses effectively maintains graft survival independent of Tregs.
Chronic CTLA4Ig monotherapy in a murine model of cardiac transplantation results in dose‐dependent immunosuppressive efficacy that is Treg‐dependent at low, but not high, doses.
Immunohistochemistry (IHC) is the gold standard for diagnosing (positive vs. negative) polyomavirus BK (BKV) nephropathy and has the potential for disease staging based on staining intensity and ...quantification of infected cells. This multicenter trial evaluated the reproducibility of BKV IHC among 81 pathologists at 60 institutions. Participants stained tissue microarray slides and scored them for staining intensity and percentage of positive nuclei. Staining protocol details and evaluation scores were collected online. Slides were returned for centralized panel re‐evaluation and kappa statistics were calculated. Individual assessment of staining intensity and percentage was more reproducible than combined scoring. Inter‐institutional reproducibility was moderate for staining intensity (κ = 0.49) and percentage (κ = 0.42), fair for combined (κ = 0.25) and best for simple positive/negative scoring (κ = 0.78). Inter‐observer reproducibility was substantial for intensity (κ = 0.74), percentage (κ = 0.66), positive/negative (κ = 0.78) and moderate for combined scoring (κ = 0.43). Inter‐laboratory reproducibility was fair for intensity (κ = 0.37), percentage (κ = 0.40) and combined (κ = 0.24), but substantial for positive/negative scoring (κ = 0.67). BKV RNA copies/cell correlated with staining intensity (r = 0.56) and percentage (r = 0.62). These results indicate that BKV IHC is reproducible between observers but scoring should be simplified to a single‐feature schema. Standardization of tissue processing and staining protocols would further improve inter‐laboratory reproducibility.
This Banff multicenter study reveals that polyomavirus immunohistochemistry is reproducible between institutions if scoring is simplified to a single feature (preferably percent‐stained nuclear) and staining protocols are standardized.
At the Centre for Proton Therapy at the Paul Scherrer Institute, cancer patients are treated with a fixed beamline and in two gantries for ocular and nonocular malignancies, respectively. For the ...installation of a third gantry, a new patient safety system (PaSS) was developed and is subsequently being rolled out to update the existing areas. The aim of the PaSS is to interrupt the treatment whenever any subsystem detects a hazardous condition. To ensure correct treatment delivery, this system needs to be thoroughly tested as part of the regular quality assurance (QA) protocols as well as after any upgrade. In the legacy safety systems, unit testing required extensive use of resources: 2 weeks of work per area in the laboratory in addition to QA beam time. In order to significantly reduce the time, an automated PaSS test stand for unit testing was developed based on a PCI eXtensions for Instrumentation chassis with virtually unlimited I/Os that are synchronously stimulated or sampled at 1 MHz. It can emulate the rest of the facility using adapters to connect each type of interface. With it, the PaSS can be tested under arbitrary conditions. A VHSIC (very high speed integrated circuit) hardware description language-based formal language was developed to describe stimuli, expected behavior, and specific measurements, interpreted by a LabView runtime environment. This paper describes the tools and methodology being applied for unit testing and QA release tests for the new PaSS. It shows how automation and formalization made possible to increase the test coverage while significantly reducing the laboratory testing time and facility's beam usage.
Endothelial deposition of the complement split product C4d is an established marker of antibody-mediated acute renal allograft rejection. A contribution of alloantibody-dependent immune reactions to ...chronic rejection is under discussion. In this study, the association of immunohistochemically detected endothelial C4d deposition in peritubular capillaries (PTC) with morphologic features of chronic renal allograft injury was investigated in a large study cohort. C4d deposits in PTC were detected in 73 (34%) of 213 late allograft biopsies performed in 213 patients more than 12 mo after transplantation (median, 4.9 yr) because of chronic allograft dysfunction. Endothelial C4d deposition was found to be associated with chronic transplant glomerulopathy (CG) (P < 0.0001), with basement membrane multilayering in PTC (P = 0.01), and with an accumulation of mononuclear inflammatory cells in PTC (P < 0,001). Furthermore, C4d deposits in PTC (in biopsies with normal glomerular morphology) were associated with development of CG in follow-up biopsies. Other morphologic features of chronic allograft nephropathy (with exception of tubular atrophy) were not associated with C4d deposits in PTC. Analyses of previous and follow-up biopsies revealed that C4d deposits may occur de novo and may also disappear at any time after transplantation. In conclusion, the data suggest that complement activation in renal microvasculature, indicating humoral alloreactivity, contributes to chronic rejection characterized by chronic transplant glomerulopathy and basement membrane multilayering in PTC.
Therapeutic administration of regulatory T cells (Tregs) leads to engraftment of conventional doses of allogeneic bone marrow (BM) in nonirradiated recipient mice conditioned with costimulation ...blockade and mammalian target of rapamycin inhibition. The mode of action responsible for this Treg effect is poorly understood but may encompass the control of costimulation blockade–resistant natural killer (NK) cells. We show that transient NK cell depletion at the time of BM transplantation led to BM engraftment and persistent chimerism without Treg transfer but failed to induce skin graft tolerance. In contrast, the permanent absence of anti–donor NK reactivity in mice grafted with F1 BM was associated with both chimerism and tolerance comparable to Treg therapy, implying that NK cell tolerization is a critical mechanism of Treg therapy. Indeed, NK cells of Treg‐treated BM recipients reshaped their receptor repertoire in the presence of donor MHC in a manner suggesting attenuated donor reactivity. These results indicate that adoptively transferred Tregs prevent BM rejection, at least in part, by suppressing NK cells and promote tolerance by regulating the appearance of NK cells expressing activating receptors to donor class I MHC.
NK cell depletion can replace Treg therapy to achieve bone marrow engraftment without irradiation under costimulation blockade and rapamycin while the permanent absence of NK cell donor reactivity is required for robust tolerance.
Lipocalin 2 (Lcn2) is rapidly produced by damaged nephron epithelia and is one of the most promising new markers of renal injury, delayed graft function and acute allograft rejection (AR); however, ...the functional importance of Lcn2 in renal transplantation is largely unknown. To understand the role of Lcn2 in renal AR, kidneys from Balb/c mice were transplanted into C57Bl/6 mice and vice versa and analyzed for morphological and physiological outcomes of AR at posttransplantation days 3, 5, and 7. The allografts showed a steady increase in intensity of interstitial infiltration, tubulitis and periarterial aggregation of lymphocytes associated with a substantial elevation in serum levels of creatinine, urea and Lcn2. Perioperative administration of recombinant Lcn2:siderophore:Fe complex (rLcn2) to recipients resulted in functional and morphological amelioration of the allograft at day 7 almost as efficiently as daily immunosuppression with cyclosporine A (CsA). No significant differences were observed in various donor–recipient combinations (C57Bl/6 wild‐type and Lcn2−/−, Balb/c donors and recipients). Histochemical analyses of the allografts showed reduced cell death in recipients treated with rLcn2 or CsA. These results demonstrate that Lcn2 plays an important role in reducing the extent of kidney AR and indicate the therapeutic potential of Lcn2 in transplantation.
The authors demonstrate the function of lipocalin 2 as an upregulated marker during acute rejection and as a therapeutic agent to reduce acute rejection in a murine kidney transplantation model, supporting a possible future application of lipocalin 2 as an early biomarker as well as an immunosuppressive agent in kidney transplantation.
Humoral alloreactivity is well established to predict adverse allograft outcomes. However, in some recipients, alloantibodies may also occur in the absence of graft dysfunction. We evaluated if and ...how often complement‐ and noncomplement‐fixing alloantibodies are detectable in stable recipients and whether, in this context, they affect long‐term outcomes. Sera obtained from 164 kidney transplant recipients at 2, 6 and 12 months were evaluated by FlowPRA screening and single‐antigen testing for detection of IgG‐ or C4d‐fixing HLA panel reactivity and donor‐specific antibodies (DSA). Applying stringent criteria, we selected 34 patients with an uneventful 1‐year course (no graft dysfunction or rejection) and excellent graft function at 12 months estimated glomerular filtration rate (eGFR) ≥60 mL/min and proteinuria ≤0.5 g/24 h. Nine (27%) and 5 (15%) of these recipients tested positive by IgG and C4dFlowPRA screening, respectively. In five cases, DSA were identified. Frequencies of positive test results and DSA binding intensities were not significantly lower than those documented for patients who did not fulfill the above criteria. In recipients with an excellent 1‐year course, FlowPRA reactivity was not associated with lower eGFR or increased protein excretion during 68‐month median follow‐up. Our results suggest cautious interpretation of antibody monitoring in patients with normal‐functioning grafts.
This study demonstrates that, in kidney transplant recipients with excellent 1‐year graft performance, the occurrence of complement‐ and noncomplement‐fixing anti‐HLA alloreactivity may not be associated with adverse long‐term graft function, suggesting cautious interpretation of post‐transplant antibody monitoring in the absence of graft dysfunction.
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