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- Bockermann, Robert, et al.
(författare)
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Imlifidase-generated Single-cleaved IgG : Implications for Transplantation
- 2022
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Ingår i: Transplantation. - : Ovid Technologies (Wolters Kluwer Health). - 0041-1337 .- 1534-6080. ; 106:7, s. 1485-1496
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Tidskriftsartikel (refereegranskat)abstract
- Background. Imlifidase is an immunoglobulin G (IgG)-specific protease conditionally approved in the EU for desensitization in highly sensitized crossmatch positive kidney transplant patients. Imlifidase efficiently cleaves both heavy chains of IgG in a 2-step process. However, low levels of the intermediate cleavage product, single-cleaved IgG (scIgG), may persist in the circulation. The study objective was to investigate Fc-mediated effector functions of scIgG and its potential impact on common clinical immunologic assays used to assess transplant eligibility.Methods. Imlifidase-generated scIgG, obtained by in vitro cleavage of HLA-sensitized patient serum or selected antibodies, was investigated in different complement- and Fc gamma R-dependent assays and models, including clinical tests used to evaluate HLA-specific antibodies.Results. ScIgG had significantly reduced Fc-mediated effector function compared with intact IgG, although some degree of activity in complement- and Fc gamma R-dependent models was still detectable. A preparation of concentrated scIgG generated from a highly HLA-sensitized individual gave rise to a positive signal in the anti-HLA IgG LABScreen, which uses anti-Fc detection, but was entirely negative in the C1qScreen. The same high-concentration HLA-binding scIgG preparation also generated positive complement-dependent cytotoxicity responses against 80%-100% of donor T and B cells, although follow-up titrations demonstrated a much lower intrinsic activity than for intact anti-HLA IgG.Conclusions. ScIgG has a significantly reduced capacity to mediate Fc-dependent effector functions. However, remaining HLA-reactive scIgG in plasma after imlifidase treatment can cause positive assay results equivalent to intact IgG in clinical assays. Therefore, complete IgG cleavage after imlifidase treatment is essential to allow correct decision-making in relation to transplant eligibility.
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- Runstrom, Anna, et al.
(författare)
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IgM single antigen bead HLA-assay is affected by imlifidase through the cleavage of IgG but not IgM
- 2021
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Ingår i: Transplant Immunology. - : Elsevier. - 0966-3274 .- 1878-5492. ; 68
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Tidskriftsartikel (refereegranskat)abstract
- Aim: The aim of this study was to investigate if human IgM is a cleavable substrate for imlifidase and to explain an observed effect in anti-HLA IgM single antigen bead (SAB) assays in sensitized patients. Methods: Serum samples collected pre-and 24 h post-imlifidase administration from sensitized patients enrolled in a phase II trial were investigated for anti-HLA IgG and IgM using SAB assays, with and without in vitro IgG depletion using a CaptureSelectTM affinity matrix. In addition, pre-dose samples and purified human IgM samples were treated with imlifidase in vitro and evaluated by SDS-PAGE, Western blot (PE-conjugated anti-human IgM) and SAB (IgG, IgM) assays. Results: By comparing the mean fluorescence intensity (MFI) of HLA-beads, pre-and post-imlifidase administration, three IgM-related patterns were observed; IgM-specific HLA-SABs with an increased MFI post-imlifidase, IgM-specific HLA-SABs with a decreased MFI post-imlifidase, and IgM-specific HLA-SABs with a marginal MFI difference between the pre-and post-imlifidase administration. These IgM signal patterns were observed despite neither purified IgM nor serum IgM could be cleaved by imlifidase. After removing IgG, the effects observed on anti-HLA IgM was largely eliminated with the biggest differences seen in patients with very high anti-HLA IgG in pre-dose samples. Conclusion: We demonstrate that imlifidase does not cleave human IgM, including HLA-specific IgM antibodies from highly sensitized subjects. Observed decreases of SAB-HLA IgM signals after imlifidase treatment may result from the cleavage of IgG-IgM complexes which are bound to SAB-HLA. Serum analysis of patients with high levels of anti-HLA IgG will result in a more accurate SAB-HLA IgM reading after IgG depletion.
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