Pathogenic role of autoantibodies in SSc

Aab are present in most of SSc patients and constitute important diagnosis and prognosis biomarkers (ANA such as ATA but also functional antibodies such as EC). Yet, their pathophysiological role remains unclear. We previously reported that: (i) purified IgG from SSc patients could modify the whole FB proteome/transcriptome profiles, (ii) purified IgG from SSc with ATA group induced a singular and profibrotic profile on FB when compared to other serotypes. These data suggested a possible direct pathogenic role of Aab in SSc, especially ATA. Given that this initial unbiased work on the multiomic profiles induced by purified IgG enabled us to isolate a possible endotype represented by the ATA serotype, and that ANA are used in the daily routine management of patients, we will focus more specifically on the potential pathogenic role of ATA on different cell types targeted by SSc. Additionally, we will develop complementary approaches including other serotypes or functional antibodies such as anti-endothelin or anti-angiotensin receptor. We aim to understand the pathogenic role of SSc purified IgG or isolated Aab on keys effector cells, i.e. FB, EC, keratinocytes and digestive tract cells (in collaboration with WP2).

To further study the FB and IgG interaction in SSc, we will assess 1. the biological pathway activation in FB by phosphoproteomics and kinetics approaches and the secretome of FB after incubation with total purified IgG from ATA positive SSc patients or purified ATA IgG and monoclonal ATA (collaboration Eric Meffre, Stanford), 2. the attachment to membrane and the intra-cellular trafficking. Dermal FB will be cultured in the presence of purified total IgG from SSc patients with or without ATA or from healthy controls or in the presence of purified ATA IgG. The consequences of purified total IgG or purified ATA on FB will be assessed by analyzing the phosphoproteome, cell compartments and exosomes by mass spectrometry coupled with liquid chromatography (LC-MS/MS). Determination of the interaction between ATA and FB will be performed using confocal microscopy.

We will set up an in vitro model of human umbilical vein EC on which we will apply the same experimental conditions as on FB. Our cohort will be composed of patients with ATA, patients with anti-centromere antibodies, patients with anti-RNA polymerase III and healthy subjects. The effect of the antibodies will be measured by high-throughput omics techniques (LC-MS/MS and RNA-sequencing) and complemented by conventional approaches (ELISA, WB, IF) to confirm the identified targets.