Background Systemic Lupus Erythematosus (SLE) and Lupus Nephritis (LN) are prototypical autoimmune conditions. Autoantibodies (AutoAb) play a central role in disease initiation and progression.

Methods In the present work, we investigated sera of SLE and LN patients obtained through the ZEUS consortium, using a high-density peptide array covering the coding sequences of 464 proteins involved in rheumatologic and immunologic processes encompassing 17,402 tiled peptides. Results were validated by ELISA in patients affected by SLE (n=113) or LN (n=210). Immunohistochemistry and electron microscopy was performed in a subset of LN patients (n=12). The prognostic value of selected identified AutoAbs was tested in LN patients with at least two paired serum and urine samples (T0–12). Predictive value of baseline anti-FMNL1 AutoAbs was tested in subset of patients treated with Belimumab.

Results A panel of 133 proteins reactive to SLE and LN sera were identified.

Among those proteins, peptides coding for Formin-like 1 Protein (FMNL1) scored with the highest probability suggestive of FMNL1 being a potential antigen in both SLE and LN patients. High levels of anti-FMNL1 IgG2 were detected by ELISA in serum of LN and SLE patients and baseline FMNL1 values predicted proteinuria at 12-month follow-up in LN patients. Patients with high circulating anti-FMNL1 IgG2 maintained high levels of proteinuria after 1-year follow- up and failed to respond to Belimumab, suggestive of potential use of anti-FMNL1 as clinical predictive biomarker.

In kidney LN biopsies, FMNL1 protein co-stained with macrophage marker CD68, its high expression being associated with proliferative LN stage III-IV. In glomeruli, FMNL1 co-stained with IgG2 suggestive of specific interaction and potentially modulation of macrophages by circulating anti-FMNL1 AutoAbs.

Conclusions Deployment of peptide array technology was pivotal to identify novel autoantibodies present in in SLE and LN; among those AutoAbs, high circulating levels of anti-FMNL1 IgG2 were detected in both SLE and LN. Anti-FMNL1 AutoAbs target a protein expressed in glomeruli by macrophages and associated with proliferative LN. That evidence is potentially suggestive of a role of anti-FMNL1 AutoAbs in modulating cells involved in tissue repair determining the disease outcome.

Zeus Consortium: Gabriella Moroni⁸, Renato Alberto Sinico⁹, Franco Franceschini¹⁰, Micaela Fredi¹⁰⁰, Augusto Vaglio¹¹, Lorenzo Cavagna¹², Federico Pratesi¹³, Paola Migliorini¹³, Francesco Locatelli¹¹, Giulia Pazzola¹⁴, Giampaola Pesce¹⁵, Angelo Manfredi¹⁶, Giuseppe A Ramirez¹⁶, Pasquale Esposito¹⁷, Giuseppe Murdaca¹⁸, Simone Negrini¹⁸, Leda Cipriani¹⁷, BarbaraTrezzi⁸, Giacomo Emmi¹⁹, Ilaria Cavazzana⁹, Valentina Binda⁹, Paride Fenaroli²⁰, Isabella Pisani²⁰, Giacomo Garibotto¹⁷, Carlomaurizio Montecucco¹², Domenico Santoro²¹, Francesco Scolari²², Stefano Volpi²³, Marta Mosca ²³, Angela Tincani¹⁰, Andrea Petretto²⁴

8 Department of Biomedical Sciences, Humanitas University and IRCCS Humanitas Research Hospital,Milan, Italy

9 Department of Medicine and surgery, University of Milan, Bicocca, Italy;

10 Rheumatology and Clinical Immunology, ASST SpedaliCivili and Università of Brescia, Italy;

11Department of Biomedical, Experimental and Clinical Sciences ‘Mario Serio’, University of Firenze, and Nephrology and Dialysis Unit, Meyer Children’s Hospital, Firenze, Italy;

12 Division of Rheumatology, University and IRCCS Policlinico S. Matteo, Pavia, Italy;

13 Clinical Immunology Unit, Department of Internal Medicine, University of Pisa, Italy;

14 Nephrology and Dialysis, Arciospedale Santa Maria nuova, Reggio Emilia, Italy;