Background: Many autoimmune conditions are driven by T cell responses with dominant Th17 or Th1 activity. There is plasticity in the differentiation of Th17 and regulatory T cells (Tregs), which relies in part on STAT3 signaling. Previous work in other autoimmune models has shown that inhibition of STAT3 alters the balance of Th17 and Treg cells, favoring a less inflammatory T cell repertoire and ameliorating inflammation. though janus kinase (JAK) signaling has arisen as an important pharmaceutical target in many autoimmune disorders, including lupus and myositis, no STAT inhibitors are currently approved for treatment of autoimmune disease.

Objectives: We quantified T cell subset distribution in vitro from peripheral blood mononuclear cells (PBMC) isolated from adult patients with inflammatory diseases or healthy controls as well as in vivo in murine models of myositis and lupus to determine responses to STAT3 inhibition and whether this is a viable therapeutic target.

Methods: Peripheral blood mononuclear cells (PBMC) from healthy controls and patients were harvested by Ficoll separation and cultured in vitro for 3 days with anti-CD3/anti-CD28 stimulatory beads and IL2 ± TGFβ and IL6 with or without the small molecule STAT3 inhibitor LLL12b at varying concentrations. Th17 and Treg cells were quantified by flow cytometry by intra-nuclear transcription factor staining for FoxP3 and RORγt. Differences in population frequencies were compared between pairs of groups via the Mann Whitney U test. Similar studies were performed on murine PBMC and splenocytes isolated from experimental animals treated with histidyl tRNA synthetase (HRS) or imiquimod to induce inflammatory myositis or systemic lupus erythematosus, respectively.

Results: Myositis and lupus patients with clinically active disease demonstrated increased RORγt+ Th17 T cells, but this was not universal across all patients. STAT3 inhibition with LLL12b in vitro decreased the frequency of Th17 T cells in these patients but not in healthy controls. In mice, LLL12b treatment in vivo did not appear to ameliorate disease nor prevent immune cell infiltration into diseased tissues in the myositis model; parallel histological analyses in the lupus model are currently pending.

Conclusions: STAT3 inhibition was sufficient to decrease the proportion of Th17 cells in vitro , but other factors may have limited LLL12B’s in vivo effects. Important considerations when interpreting these data is that Th17 and Treg quantitation was based on transcription factor expression in the peripheral blood rather than T cell functional assays. Patients were also receiving treatment for their lupus and myositis at the time PBMC were collected, which might alter T cell profiling results. Additional studies are in progress to assess T cell function (secretion of IL17, IL10, and TGFβ) as well as proliferation both in vitro and in vivo to better characterize the effect of STAT3 inhibition on Th17 and Treg cells in autoimmune disease. These studies have implications for treatment development and may help shed additional light on the pathogenesis of autoimmune disorders.

REFERENCES: NIL.

Acknowledgments: NIL.

Disclosure of Interests: None declared.