Cell-based BSEP trans-inhibition: A novel, non-invasive test for diagnosis of antibody-induced BSEP deficiency

TY - JOUR

T1 - Cell-based BSEP trans-inhibition: A novel, non-invasive test for diagnosis of antibody-induced BSEP deficiency

AU - Stindt, Jan

AU - Dröge, Carola

AU - Lainka, Elke

AU - Kathemann, Simone

AU - Pfister, Eva-Doreen

AU - Baumann, Ulrich

AU - Stalke, Amelie

AU - Grabhorn, Enke

AU - Shagrani, Mohammad Ali

AU - Mozer-Glassberg, Yael

AU - Hartley, Jane

AU - Wammers, Marianne

AU - Klindt, Caroline

AU - Philippski, Paulina

AU - Liebe, Roman

AU - Herebian, Diran

AU - Mayatepek, Ertan

AU - Berg, Thomas

AU - Schmidt-Choudhury, Anjona

AU - Wiek, Constanze

AU - Hanenberg, Helmut

AU - Luedde, Tom

AU - Keitel, Verena

N1 - © 2023 The Authors.

PY - 2023/7

Y1 - 2023/7

N2 - BACKGROUND & AIMS: Antibody-induced bile salt export pump deficiency (AIBD) is an acquired form of intrahepatic cholestasis, which may develop following orthotopic liver transplantation (OLT) for progressive familial intrahepatic cholestasis type 2 (PFIC-2). Approximately 8-33% of patients with PFIC-2 who underwent a transplant develop bile salt export pump (BSEP) antibodies, which trans-inhibit this bile salt transporter from the extracellular, biliary side. AIBD is diagnosed by demonstration of BSEP-reactive and BSEP-inhibitory antibodies in patient serum. We developed a cell-based test directly measuring BSEP trans-inhibition by antibodies in serum samples to confirm AIBD diagnosis.METHODS: Sera from healthy controls and cholestatic non-AIBD or AIBD cases were tested (1) for anticanalicular reactivity by immunofluorescence staining of human liver cryosections, (2) for anti-BSEP reactivity by immunofluorescence staining of human embryonic kidney 293 (HEK293) cells expressing BSEP-enhanced yellow fluorescent protein (EYFP) and immunodetection of BSEP-EYFP on Western blot, and (3) for BSEP trans-inhibition using HEK293 cells stably expressing Na+/taurocholate cotransporting polypeptide (NTCP)-mCherry and BSEP-EYFP. The trans-inhibition test uses [3H]-taurocholate as substrate and is divided into an uptake phase dominated by NTCP followed by BSEP-mediated export. For functional analysis, sera were bile salt depleted.RESULTS: We found BSEP trans-inhibition by seven sera containing anti-BSEP antibodies, but not by five cholestatic or nine control sera, all lacking BSEP reactivity. Prospective screening of a patient with PFIC-2 post OLT showed seroconversion to AIBD, and the novel test method allowed monitoring of treatment response. Notably, we identified a patient with PFIC-2 post OLT with anti-BSEP antibodies yet without BSEP trans-inhibition activity, in line with asymptomatic presentation at serum sampling.CONCLUSIONS: Our cell-based assay is the first direct functional test for AIBD and allows confirmation of diagnosis as well as monitoring under therapy. We propose an updated workflow for AIBD diagnosis including this functional assay.IMPACT AND IMPLICATIONS: Antibody-induced BSEP deficiency (AIBD) is a potentially serious complication that may affect patients with PFIC-2 after liver transplantation. To improve its early diagnosis and thus immediate treatment, we developed a novel functional assay to confirm AIBD diagnosis using a patient's serum and propose an updated diagnostic algorithm for AIBD.

AB - BACKGROUND & AIMS: Antibody-induced bile salt export pump deficiency (AIBD) is an acquired form of intrahepatic cholestasis, which may develop following orthotopic liver transplantation (OLT) for progressive familial intrahepatic cholestasis type 2 (PFIC-2). Approximately 8-33% of patients with PFIC-2 who underwent a transplant develop bile salt export pump (BSEP) antibodies, which trans-inhibit this bile salt transporter from the extracellular, biliary side. AIBD is diagnosed by demonstration of BSEP-reactive and BSEP-inhibitory antibodies in patient serum. We developed a cell-based test directly measuring BSEP trans-inhibition by antibodies in serum samples to confirm AIBD diagnosis.METHODS: Sera from healthy controls and cholestatic non-AIBD or AIBD cases were tested (1) for anticanalicular reactivity by immunofluorescence staining of human liver cryosections, (2) for anti-BSEP reactivity by immunofluorescence staining of human embryonic kidney 293 (HEK293) cells expressing BSEP-enhanced yellow fluorescent protein (EYFP) and immunodetection of BSEP-EYFP on Western blot, and (3) for BSEP trans-inhibition using HEK293 cells stably expressing Na+/taurocholate cotransporting polypeptide (NTCP)-mCherry and BSEP-EYFP. The trans-inhibition test uses [3H]-taurocholate as substrate and is divided into an uptake phase dominated by NTCP followed by BSEP-mediated export. For functional analysis, sera were bile salt depleted.RESULTS: We found BSEP trans-inhibition by seven sera containing anti-BSEP antibodies, but not by five cholestatic or nine control sera, all lacking BSEP reactivity. Prospective screening of a patient with PFIC-2 post OLT showed seroconversion to AIBD, and the novel test method allowed monitoring of treatment response. Notably, we identified a patient with PFIC-2 post OLT with anti-BSEP antibodies yet without BSEP trans-inhibition activity, in line with asymptomatic presentation at serum sampling.CONCLUSIONS: Our cell-based assay is the first direct functional test for AIBD and allows confirmation of diagnosis as well as monitoring under therapy. We propose an updated workflow for AIBD diagnosis including this functional assay.IMPACT AND IMPLICATIONS: Antibody-induced BSEP deficiency (AIBD) is a potentially serious complication that may affect patients with PFIC-2 after liver transplantation. To improve its early diagnosis and thus immediate treatment, we developed a novel functional assay to confirm AIBD diagnosis using a patient's serum and propose an updated diagnostic algorithm for AIBD.

U2 - 10.1016/j.jhepr.2023.100690

DO - 10.1016/j.jhepr.2023.100690

M3 - SCORING: Journal article

C2 - 37425215

VL - 5

SP - 100690

JO - JHEP REP

JF - JHEP REP

SN - 2589-5559

IS - 7

ER -