Selective oral ROCK2 inhibitor down-regulates IL-21 and IL-17 secretion in human T cells via STAT3-dependent mechanism

Abstract

Rho-associated kinase 2 (ROCK2) regulates the secretion of proinflammatory cytokines and the development of autoimmunity in mice. Data from a phase 1 clinical trial demonstrate that oral administration of KD025, a selective ROCK2 inhibitor, to healthy human subjects down-regulates the ability of T cells to secrete IL-21 and IL-17 by 90% and 60%, respectively, but not IFN-γ in response to T-cell receptor stimulation in vitro. Pharmacological inhibition with KD025 or siRNA-mediated inhibition of ROCK2, but not ROCK1, significantly diminished STAT3 phosphorylation and binding to IL-17 and IL-21 promoters and reduced IFN regulatory factor 4 and nuclear hormone RAR-related orphan receptor γt protein levels in T cells derived from healthy subjects or rheumatoid arthritis patients. Simultaneously, treatment with KD025 also promotes the suppressive function of regulatory T cells through up-regulation of STAT5 phosphorylation and positive regulation of forkhead box p3 expression. The administration of KD025 in vivo down-regulates the progression of collagen-induced arthritis in mice via targeting of the Th17-mediated pathway. Thus, ROCK2 signaling appears to be instrumental in regulating the balance between proinflammatory and regulatory T-cell subsets. Targeting of ROCK2 in man may therefore restore disrupted immune homeostasis and have a role in the treatment of autoimmunity.T he immune response is a delicate balancing act, protecting the integrity of the host organism from foreign invaders while not causing autoimmune reactivity (1). IL-21 and IL-17 are proinflammatory cytokines produced by T-helper 17 (Th17) cells that are involved in the pathogenesis of many autoimmune diseases (2-5). The generation of Th17 cells is induced by a combination of several cytokines including transforming growth factor-β (TGF-β1), IL-1β, IL-6, and IL-23, and involves the activation of transcription factors, such as RAR-related orphan receptor (ROR) γt, RORα, IFN regulatory factor (IRF) 4, and signal transducer and activator of transcription 3 (STAT3) (2, 6, 7). However, the signaling pathways that lead to activation of this transcriptional profile are poorly understood and remain unclear.Rho GTPase-mediated signaling pathways play a central role in the coordination and balancing of T-cell-mediated immune responses, including T-cell receptor (TCR)-mediated signaling, cytoskeletal reorganization, and the acquisition of the appropriate T-cell effector program (8). The Rho kinase family members, consisting of Rho-associated kinase 1 (ROCK1) and ROCK2, are serine-threonine kinases that are activated by Rho GTPases and mediate the phosphorylation of downstream targets in cells (9). Recent studies have demonstrated that ROCK2 regulates the production of both IL-21 and IL-17 and plays an essential role in the development of autoimmunity in mice (10, 11). Indeed, pan ROCK inhibition was reported to effectively down-regulate ongoing autoimmune response in animal models (11,12). Additionally, ROCK activity was found to be up-regulated in patients w...

Cutting Edge: Selective Oral ROCK2 Inhibitor Reduces Clinical Scores in Patients with Psoriasis Vulgaris and Normalizes Skin Pathology via Concurrent Regulation of IL-17 and IL-10

“…At week 12, 42% of patients in the 400 mg once a day cohort and 29% of patients in the 400 mg twice a day cohort achieved a PASI 50 response (Fig. 1B), suggesting a potential clinical benefit of using a lower dosage regimen (200 mg twice a day) of the selective ROCK2 inhibitor KD025, which is 75-to 100-fold more selective for ROCK2 than for ROCK1 (15,21). Interestingly, a recent study demonstrated that inhibition of both ROCK1 and ROCK2 isoforms with pan-ROCK inhibitors potently induces proliferation of primary human keratinocytes in vitro, and might have a negative impact in proinflammatory settings such as psoriasis compared with selective ROCK2 inhibition (22).…”

“…Additionally, several groups demonstrated that ROCK2 specifically contributes to regulation of IL-17 secretion in mice and humans (13,15). To explore whether plasma IL-17 levels could be used to monitor response to the KD025 treatment, peripheral blood samples were collected on a monthly basis and cytokine levels were measured by using a Quanterix Simoa platform for biomolecule detection.…”

“…KD025 (formerly Slx-2119) is ATP competitive and 100-fold more selective for the ROCK2 isoform compared with ROCK1, with no significant activity against 300 intracellular kinases and surface receptors (15,19). A phase 2, open-label, dose-finding study was initiated to evaluate the safety, tolerability, and activity of KD025 in subjects with psoriasis vulgaris (NCT02317627 at ClinicalTrials.gov).…”

“…Previous findings demonstrated that oral administration of a selective ROCK2 inhibitor (KD025) in healthy subjects decreases IL-17 and IL-21 secretion induced by ex vivo stimulation (15 immunosuppressive T cell subsets via concurrent regulation of STAT3/STAT5 phosphorylation (16)(17)(18). Thus, we have hypothesized that targeting of ROCK2 may restore disrupted immune homeostasis and could have a role in the treatment of Th17-mediated inflammatory disorders such as psoriasis.…”

“…Rho-associated kinase (ROCK)2 was shown recently to be implicated in the regulation of autoimmunity in mice and humans (13)(14)(15). Previous findings demonstrated that oral administration of a selective ROCK2 inhibitor (KD025) in healthy subjects decreases IL-17 and IL-21 secretion induced by ex vivo stimulation (15 immunosuppressive T cell subsets via concurrent regulation of STAT3/STAT5 phosphorylation (16)(17)(18).…”