Late antibody-mediated rejection (ABMR) is a leading cause of transplant failure. Blocking IL-6 has been proposed as a promising therapeutic strategy.
We performed a phase 2 randomized pilot trial to ...evaluate the safety (primary endpoint) and efficacy (secondary endpoint analysis) of the anti-IL-6 antibody clazakizumab in late ABMR. The trial included 20 kidney transplant recipients with donor-specific, antibody-positive ABMR ≥365 days post-transplantation. Patients were randomized 1:1 to receive 25 mg clazakizumab or placebo (4-weekly subcutaneous injections) for 12 weeks (part A), followed by a 40-week open-label extension (part B), during which time all participants received clazakizumab.
Five (25%) patients under active treatment developed serious infectious events, and two (10%) developed diverticular disease complications, leading to trial withdrawal. Those receiving clazakizumab displayed significantly decreased donor-specific antibodies and, on prolonged treatment, modulated rejection-related gene-expression patterns. In 18 patients, allograft biopsies after 51 weeks revealed a negative molecular ABMR score in seven (38.9%), disappearance of capillary C4d deposits in five (27.8%), and resolution of morphologic ABMR activity in four (22.2%). Although proteinuria remained stable, the mean eGFR decline during part A was slower with clazakizumab compared with placebo (-0.96; 95% confidence interval 95% CI, -1.96 to 0.03 versus -2.43; 95% CI, -3.40 to -1.46 ml/min per 1.73 m
per month, respectively,
=0.04). During part B, the slope of eGFR decline for patients who were switched from placebo to clazakizumab improved and no longer differed significantly from patients initially allocated to clazakizumab.
Although safety data indicate the need for careful patient selection and monitoring, our preliminary efficacy results suggest a potentially beneficial effect of clazakizumab on ABMR activity and progression.
BACKGROUND.Late antibody-mediated rejection (AMR) is a major cause of transplant failure. Potential therapeutic targets are plasma cells (PC) and/or natural killer (NK) cells, both expressing high ...levels of CD38.
METHODS.Here, we report the use of CD38 monoclonal antibody daratumumab (9-month course) in a kidney allograft recipient diagnosed with smoldering myeloma and anti-HLA class II donor-specific antibody (DSA)-positive chronic active AMR 13 years after transplantation. Patient monitoring included serial HLA single antigen testing, peripheral blood immune cell phenotyping, as well as follow-up allograft and bone marrow biopsies at 3 and 9 months, including analyses of rejection-related gene expression patterns.
RESULTS.Daratumumab led to persistent CD138 cell depletion in the bone marrow and blood, and substantially decreased NK cells counts in blood and graft tissue. At the same time, DSA in serum disappeared, and in vitro alloantibody production by CD138 cells enriched from bone marrow aspirates was abrogated. A 3-month follow-up biopsy revealed a complete resolution of microcirculation inflammation (g+ptc3 to 0) and molecular AMR activity (AMR score0.79 to <0.2). The same biopsy showed (subclinical) tubulo-interstitial inflammation, which prompted steroid treatment. Over an observation period of 12 months, graft function stabilized.
CONCLUSIONS.Targeting CD38 for PC and NK cell depletion may be an effective strategy to counteract AMR. Our results may encourage the design of future trials to clarify the role of this innovative treatment concept in organ transplantation.
ABSTRACTThe complement system may contribute in many ways to transplant injury, being a promising target for specific therapeutic interventions. There is evidence that the monoclonal anti-C5 antibody ...eculizumab is effective in the prevention and treatment of early antibody-mediated rejection, but terminal complement blockade might be of limited efficiency in chronic rejection. Given the diversity of immunological events triggered by activation steps upstream to C5, in particular, opsonin and anaphylatoxin formation through C3 cleavage, one may argue that, in the specific context of antibody-mediated rejection, inhibition of antibody-triggered classical pathway (CP) activation might be beneficial. Strategies to interfere with key CP component C1 are currently under clinical evaluation and include the therapeutic use of purified C1-inhibitor, which, besides targeting the integrity and function of the C1 complex, also affects components of the LP, the contact system, the coagulation cascade or surface molecules mediating leukocyte-endothelial interactions. In addition, a monoclonal anti-C1s antibody (BIVV009) has now entered clinical evaluation and was shown to effectively block antibody-triggered CP activation in rejecting kidney allografts. Moreover, modified apheresis techniques for preferential removal of macromolecules, including C1q, may allow for efficient complement depletion, in addition to antibody removal. The availability of effective strategies to interfere with the CP, as well as innovative approaches targeting other pathways, some of them already being tested in clinical trials, will help us figure out how complement contributes to acute and chronic graft injury, and hopefully provide us with new ways to more efficiently counteract rejection.
During human pregnancy, placenta-derived extravillous trophoblasts (EVTs) invade the decidua and communicate with maternal immune cells. The decidua distinguishes into basalis (decB) and parietalis ...(decP). The latter remains unaffected by EVT invasion. By defining a specific gating strategy, we report the accumulation of macrophages in decB. We describe a decidua basalis-associated macrophage (decBAM) population with a differential transcriptome and secretome compared with decidua parietalis-associated macrophages (decPAMs). decBAMs are CD11chi and efficient inducers of Tregs, proliferate in situ, and secrete high levels of CXCL1, CXCL5, M-CSF, and IL-10. In contrast, decPAMs exert a dendritic cell-like, motile phenotype characterized by induced expression of HLA class II molecules, enhanced phagocytosis, and the ability to activate T cells. Strikingly, EVT-conditioned media convert decPAMs into a decBAM phenotype. These findings assign distinct macrophage phenotypes to decidual areas depending on placentation and further highlight a critical role for EVTs in the induction of decB-associated macrophage polarization.
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•Decidua basalis-associated macrophages (decBAMs) accumulate during human pregnancy•decBAMs are CD11c+HLA-DRlo, have a specific transcriptome, and proliferate in situ•decBAMs are inefficient APCs, induce Tregs, and secrete IL-10 and M-CSF•Invasive trophoblasts induce a decBAM-associated phenotype
Vondra et al. report the accumulation of CD163+CD206+CD11chiHLA-DRlo macrophages termed decidua basalis-associated macrophages (decBAMs). decBAMs proliferate, secrete pregnancy-sustaining factors, and induce Tregs but show diminished phagocytosis and T cell activation efficiency. The secretome of placenta-derived trophoblasts partly induces this phenotype, suggesting that placentation co-regulates macrophage polarization during human pregnancy.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Introduction
Antibody mediated rejection (ABMR) is a major factor limiting outcome after organ transplantation. Anti-HLA donor-specific antibodies (DSA) of the IgG isotype are mainly responsible for ...ABMR. Recently DSA of the IgE isotype were demonstrated in murine models as well as in a small cohort of sensitized transplant recipients. In the present study, we aimed to determine the frequency of pre-existing and
de novo
anti-HLA IgE antibodies in a cohort of 105 solid organ transplant recipients.
Methods
We prospectively measured anti-HLA IgE antibodies in a cohort of kidney (n=60), liver, heart and lung (n=15 each) transplant recipients before and within one-year after transplantation, employing a single-antigen bead assay for HLA class I and class II antigens. Functional activity of anti-HLA IgE antibodies was assessed by an
in vitro
mediator release assay. Antibodies of the IgG1-4 subclasses and Th1 and Th2 cytokines were measured in anti-HLA IgE positive patients.
Results
Pre-existing anti-HLA IgE antibodies were detected in 10% of renal recipients (including 3.3% IgE-DSA) and in 4.4% of non-renal solid organ transplant recipients (heart, liver and lung cohort). Anti-HLA IgE occurred only in patients that were positive for anti-HLA IgG, and most IgE positive patients had had a previous transplant. Only a small fraction of patients developed
de novo
anti-HLA IgE antibodies (1.7% of kidney recipients and 4.4% of non-renal recipients), whereas no
de novo
IgE-DSA was detected. IgG subclass antibodies showed a distinct pattern in patients who were positive for anti-HLA IgE. Moreover, patients with anti-HLA IgE showed elevated Th2 and also Th1 cytokine levels. Serum from IgE positive recipients led to degranulation of basophils
in vitro
, demonstrating functionality of anti-HLA IgE.
Discussion
These data demonstrate that anti-HLA IgE antibodies occur at low frequency in kidney, liver, heart and lung transplant recipients. Anti-HLA IgE development is associated with sensitization at the IgG level, in particular through previous transplants and distinct IgG subclasses. Taken together, HLA specific IgE sensitization is a new phenomenon in solid organ transplant recipients whose potential relevance for allograft injury requires further investigation.
Late antibody-mediated rejection (ABMR) triggered by donor-specific antibodies (DSA) is a cardinal cause of kidney allograft dysfunction and loss. Diagnostic criteria for this rejection type are well ...established, but effective treatment remains a major challenge. Recent randomized controlled trials (RCT) have failed to demonstrate the efficacy of widely used therapies, such as rituximab plus intravenous immunoglobulin or proteasome inhibition (bortezomib), reinforcing a great need for new therapeutic concepts. One promising target in this context may be interleukin-6 (IL-6), a pleiotropic cytokine known to play an important role in inflammation and adaptive immunity.
This investigator-driven RCT was designed to assess the safety and efficacy of clazakizumab, a genetically engineered humanized monoclonal antibody directed against IL-6. The study will include 20 DSA-positive kidney allograft recipients diagnosed with ABMR ≥ 365 days after transplantation. Participants will be recruited at two study sites in Austria and Germany (Medical University of Vienna; Charité University Medicine Berlin). First, patients will enter a three-month double-blind RCT (1,1 randomization, stratification according to ABMR phenotype and study site) and will receive either clazakizumab (subcutaneous administration of 25 mg in monthly intervals) or placebo. In a second open-label part of the trial (months 4-12), all patients will receive clazakizumab at 25 mg every month. The primary endpoint is safety and tolerability. Secondary endpoints are the pharmacokinetics and pharmacodynamics of clazakizumab, its effect on drug metabolism in the liver, DSA characteristics, morphological ABMR lesions and molecular gene expression patterns in three- and 12-month protocol biopsies, serum/urinary biomarkers of inflammation and endothelial activation/injury, Torque Teno viral load as a measure of overall immunosuppression, kidney function, urinary protein excretion, as well as transplant and patient survival.
Currently, there is no treatment proven to be effective in halting the progression of late ABMR. Based on the hypothesis that antagonizing the effects of IL-6 improves the outcome of DSA-positive late ABMR by counteracting DSA-triggered inflammation and B cell/plasma cell-driven alloimmunity, we suggest that our trial has the potential to provide proof of concept of a novel treatment of this type of rejection.
ClinicalTrials.gov, NCT03444103 . Registered on 23 February 2018 (retrospective registration).
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
The kidney is especially sensitive to diseases associated with overactivation of the complement system. While most of these diseases affect kidney glomeruli and the microvasculature, there is also ...evidence for tubulointerstitial deposition of complement factors. Complement inactivating factors on cell membranes comprise CD55, CD59 and CD46, which is also termed membrane cofactor protein (MCP). CD46 has been described as localized to glomeruli, but especially also to proximal tubular epithelial cells (RPTECs). However, human cell culture models to assess CD46 function on RPTECs are still missing. Therefore, we here performed gene editing of RPTEC/TERT1 cells generating a monoclonal CD46-/- cell line that did not show changes of the primary cell like characteristics. In addition, factor I and CD46-mediated cleavage of C4b into soluble C4c and membrane deposited C4d was clearly reduced in the knock-out cell line as compared to the maternal cells. Thus, human CD46-/- proximal tubular epithelial cells will be of interest to dissect the roles of the epithelium and the kidney in various complement activation mediated tubulointerstitial pathologies or in studying CD46 mediated uropathogenic internalization of bacteria. In addition, this gives proof-of-principle, that telomerized cells can be used in the generation of knock-out, knock-in or any kind of reporter cell lines without losing the primary cell characteristics of the maternal cells.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Natural killer (NK) cells may contribute to antibody-mediated rejection (ABMR) of renal allografts. The role of distinct NK cell subsets in this specific context, such as NK cells expressing the ...activating receptor NKG2C, is unknown. Our aim was to investigate whether
gene deletion variants which determine NKG2C expression affect the pathogenicity of donor-specific antibodies (DSA) and, if so, influence long-term graft survival. We genotyped the
variants for two distinct kidney transplant cohorts, (i) a cross-sectional cohort of 86 recipients who, on the basis of a positive post-transplant DSA result, all underwent allograft biopsies, and (ii) 1,860 recipients of a deceased donor renal allograft randomly selected from the Collaborative Transplant Study (CTS) database. In the DSA+ patient cohort,
(80%) was associated with antibody-mediated rejection (ABMR; 65% versus 29% among
subjects;
=0.012), microvascular inflammation MVI; median g+ptc score: 2 (interquartile range: 0-4) versus 0 (0-1),
=0.002, a molecular classifier of ABMR 0.41 (0.14-0.72) versus 0.10 (0.07-0.27),
=0.001, and elevated NK cell-related transcripts (
=0.017). In combined analyses of
variants and a functional polymorphism in the Fc gamma receptor IIIA gene (
-V/F158), ABMR rates and activity gradually increased with the number of risk genotypes. In DSA+ and CTS cohorts, however, the
variant did not impact long-term death-censored graft survival, also when combined with the
V158 risk variant.
may be associated with DSA-triggered MVI and ABMR-associated gene expression patterns, but the findings observed in a highly selected cohort of DSA+ patients did not translate into meaningful graft survival differences in a large multicenter kidney transplant cohort not selected for HLA sensitization.
The functional Fc gamma receptor (FcγR) IIIA polymorphism
-V/F158 was earlier suggested to determine the potential of donor-specific HLA antibodies to trigger microcirculation inflammation, a key ...lesion of antibody-mediated renal allograft rejection. Associations with long-term transplant outcomes, however, have not been evaluated to date. To clarify the impact of
-V/F158 polymorphism on kidney transplant survival, we genotyped a cohort of 1,940 recipient/donor pairs. Analyzing 10-year death-censored allograft survival, we found no significant differences in relation to
-V/F158. There was also no independent survival effect in a multivariable Cox model. Similarly, functional polymorphisms in two other activating FcγR,
-H/R131 (FcγRIIA) and
-NA1/NA2 (FcγRIIIB), were not associated with outcome. There were also no significant survival differences among patient subgroups at increased risk of rejection-related injury, such as pre-sensitized recipients (> 0% panel reactivity; n = 438) or recipients treated for rejection within the first year after transplantation (n = 229). Our study results suggest that the earlier shown association of FcγR polymorphism with microcirculation inflammation may not be strong enough to exert a meaningful effect on graft survival.
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