Solid Phase-Based Cross-Matching as Solution for Kidney Allograft Recipients Pretreated with Therapeutic Antibodies

Abstract

In order to select recipients without donor-specific anti-HLA antibodies, the complement-dependent cytotoxicity crossmatch (CDC-CM) was established as the standard procedure about 40 years ago. However, the interpretability of this functional assay strongly depends on the vitality of isolated donors' lymphocytes. Since the application of therapeutic antibodies for the immunosuppressive regimen falsifies the outcome of the CDC-crossmatch as a result of these antibodies' complement-activating capacity in the recipients' sera, we looked for an alternative methodical approach. We here present 27 examples of AB0 blood group-incompatible living kidney allograft recipients who, due to their treatment with the humanized chimeric monoclonal anti-CD20 antibody Rituximab, did not present valid outcomes of CDC-based pretransplant cross-matching. Additionally, four cases of posttransplant cross-matching after living kidney allografting and consequent treatment with the therapeutic anti-CD25 antibody Basiliximab (Simulect) due to acute biopsy-proven rejection episodes are presented and compared regarding CDC- and ELISA-based crossmatch outcomes. In all cases, it became evident that the classical CDC-based crossmatch was completely unfeasible for the detection of donor-specific anti-HLA antibodies, whereas ELISA-based cross-matching not requiring vital cells was not artificially affected. We conclude that ELISA-based cross-matching is a valuable tool to methodically circumvent false positive CDC-based crossmatch results in the presence of therapeutically applied antibodies.

ELISA-Based Crossmatching Allowing the Detection of Emerging Donor-Specific Anti-HLA Antibodies through the Use of Stored Donors’ Cell Lysates

“…Consequently these artefacts falsify this assay's outcome by simulating DSA-mediated positive reactions. Pharmaceutical treatment, especially the use of therapeutic monoclonal antibodies of complement-activating isotypes such as Rituximab and Basiliximab both of which belong to the IgG1 isotype, is noteworthy in this context [14–17]. Furthermore, autoimmune diseases especially those of the immune complex type III such as Systemic Lupus Erythematosus and Rheumatoid Arthritis just as much lead to false positive results of the CDC-CM [4, 16, 18–20].…”

“…Several publications dealing with the superiority of these solid-phase-based assays over the classical CDC-CM have hitherto been published especially in the context of various factors falsifying the outcome of the CDC-CM. These factors comprise the use of pharmaceutical treatment such as cytostatic agents or therapeutic antibodies [14–17] and accompanying autoimmune diseases [4, 11, 12, 16, 18–20] of a given recipient. Furthermore, the procedure of ELISA-based crossmatching was successfully implemented using the outer corneal rim, that is, highly acellular tissue as the only material available from given donors instead of vital blood lymphocytes [11, 21].…”

Approaches for the characterization of clinically relevant pre‐transplant human leucocyte antigen (HLA) antibodies in solid organ transplant patients

“…The SPAs offer advantages of increased specificity, sensitivity, higher throughput, ease of use and independence from lymphocyte suspensions (Nguyen et al, 2015;Picascia et al, 2014;South & Grimm, 2016;Süsal, Opelz, et al, 2013;Tait et al, 2013). A further advantage is their ability to provide DSA assessments for patients who have received therapeutic antibodies that would interfere with a cellular crossmatch (Schlaf et al, 2015).…”

Improved flow cytometry crossmatching in kidney transplantation

“…Prospective crossmatching (P‐XM) is used to detect DSAs prior to clinical organ transplantation. Conventional, complement‐dependent cytotoxicity‐XM (CDC‐XM), ELISA XM, bead assay XM and flow cytometry crossmatching (FC‐XM) assays are variously used in the P‐XM process . It has been clearly established that positive CDC crossmatches (due to preformed DSA) are associated with early AMR and allograft loss .…”

“…Conventional, complementdependent cytotoxicity-XM (CDC-XM), ELISA XM, bead assay XM and flow cytometry crossmatching (FC-XM) assays are variously used in the P-XM process. [1][2][3][4][5] It has been clearly established that positive CDC crossmatches (due to preformed DSA) are associated with early AMR and allograft loss. 6 However, the CDC assay only detects complement-fixing antibodies and this assay presents a number of weaknesses such as low sensitivity and nonspecific responses.…”