Following organ transplantation, lifelong immunosuppressive therapy is required to prevent the host immune system from destroying the allograft. This can cause severe side effects and increased recipient morbidity and mortality. Complete cessation of immunosuppressive drugs has been successfully accomplished in selected transplant recipients, providing proof of principle that operational allograft tolerance is attainable in clinical transplantation. The intra-graft molecular pathways associated with successful drug withdrawal, however, are not well defined. In this study, we analyzed sequential blood and liver tissue samples collected from liver transplant recipients enrolled in a prospective multicenter immunosuppressive drug withdrawal clinical trial. Before initiation of drug withdrawal, operationally tolerant and non-tolerant recipients differed in the intra-graft expression of genes involved in the regulation of iron homeostasis. Furthermore, as compared with non-tolerant recipients, operationally tolerant patients exhibited higher serum levels of hepcidin and ferritin and increased hepatocyte iron deposition. Finally, liver tissue gene expression measurements accurately predicted the outcome of immunosuppressive withdrawal in an independent set of patients. These results point to a critical role for iron metabolism in the regulation of intra-graft alloimmune responses in humans and provide a set of biomarkers to conduct drug-weaning trials in liver transplantation.
Felix Bohne, Marc Martínez-Llordella, Juan-José Lozano, Rosa Miquel, Carlos Benítez, María-Carlota Londoño, Tommaso-María Manzia, Roberta Angelico, Dorine W. Swinkels, Harold Tjalsma, Marta López, Juan G. Abraldes, Eliano Bonaccorsi-Riani, Elmar Jaeckel, Richard Taubert, Jacques Pirenne, Antoni Rimola, Giuseppe Tisone, Alberto Sánchez-Fueyo
Stable liver transplant recipients with follow-up beyond 3 years after transplantation were enrolled in an immunosuppression withdrawal clinical trial. Drugs were gradually discontinued over a 6- to 9-month period, and patients were followed for an additional 12-month period. Protocol liver biopsies were obtained at study onset, at any time rejection was suspected, and at the end of the study in patients who did not reject. Patients who maintained stable graft function during the entire duration of the study and in whom no signs of rejection were noticed in protocol biopsies were considered operationally tolerant (TOL). Patients who underwent rejection at any time during the follow-up were labeled as non-tolerant (Non-TOL). Pre-weaning liver biopsy differential gene expression was assessed in a training set of TOL and Non-TOL recipients (all from Hospital Clinic Barcelona) employing whole-genome microarrays. A panel of selected genes was then analyzed by qPCR, followed by a search for predictive genetic classifiers. This was accomplished employing a random splitting strategy that incorporated an independent set of TOL and Non-TOL recipients (from University of Rome “Tor Vergata” and University Hospitals Leuven).