Proc Natl Acad Sci U S A. 2014 Nov 25;111(47):16814-9.
Selective oral ROCK2 inhibitor down-regulates IL-21 and IL-17 secretion in human T cells via STAT3-dependent mechanism
Alexandra Zanin-Zhorov1*, Jonathan Weiss1, Melanie Nyuydzefe1, Wei Chen1, Jose Scher2, Rigen Mo1, David Depoil3, Nishta Rao1, Ben Liu4, Jianlu Wei4, Sarah Lucas1, Matthew Koslow1, Maria Roche1, Olivier Schueller5, Sara Weiss1, Masha Poyurovsky1, James Tonra1, Keli L. Hippen6, Michael L. Dustin3, Bruce R. Blazar6, Chuan-ju Liu4 and Samuel D. Waksal1
1Kadmon Research Institute, New York, NY10016, USA.
2Division of Rheumatology, New York University School of Medicine and New York University Hospital for Joint Diseases, New York, NY 10003, USA.
3Molecular Pathogenesis Program, Skirball Institute of Biomolecular Medicine, Department of Pathology, New York University School of Medicine, New York, NY10016, USA
4Department of Orthopaedic Surgery, New York University School of Medicine and New York University Hospital for Joint Diseases, New York, NY 10003, USA.
5N T Life Sciences, Brighton, MA 02135, USA.
6University of Minnesota Cancer Center and Department of Pediatrics, Division of Blood and Marrow Transplantation, Minneapolis 55455, USA
*Correspondence: Alexandra.Zanin-Zhorov@kadmon.com; Kadmon Research Institute, 450 East 29th street, New York, NY 10016. Tel: 212-308-6000; Fax: 212-683-2105
Rho-associated kinase 2 (ROCK2) regulates the secretion of pro-inflammatory 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 interferon (IFN)-g in response to TCR 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, reduces interferon regulatory factor 4 and steroid receptor-type nuclear receptor RORgt 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 positively regulation of Foxp3 expression. The administration of KD025 in vivo down-regulates the progression of collagen-induced arthritis in mice via targeting of Th17-mediated pathway. Thus, ROCK2 signaling appears to be instrumental in regulating the balance between pro-inflammatory 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.
KEYWORDS: Human T cells; STAT3; STAT5; autoimmunity; proinflammatory cytokines
The immune system must maintain a continuous state of homeostasis in its ability to distinguish foreign enemies and prevent anti-self responses. This complex regulatory pathway between what is foreign and self is maintained by a network of functional cellular interactions between various CD4+ T helper cell subsets (also known as Th cells) and the cytokines they secrete. The ROCK2 signaling pathway was previously shown to be involved in controlling of cellular movement and shape. However, recent work from our group and others has demonstrated a new and important role of ROCK2 in regulating cytokine secretion in T cells(1, 2). We found that ROCK2 promotes pro-inflammatory cytokines such as IL-17 and IL-21 via STAT3-dependent mechanism. Interestingly, while in steady state conditions targeted ROCK2 inhibition has only minimal effect on IFN-g, ROCK2 significantly contributes to regulation of IFN-g secretion in T cells from Rheumatoid Arthritis (RA) patients (3). Both ROCK activity and IFN-g level were previously shown to be increased in systemic lupus erythematosus (SLE) and RA patients (4, 5). Thus, the ROCK2-dependent component of the IFN-g secretion pathway is elevated with RA progression and can be effectively reduced by selective ROCK2 inhibitor. These data suggest that the role of ROCK proteins in the modulation of human cytokine secretion may depend on the disease state of the individual (Figure 1).
Regulatory T cells (Tregs) are subset of T cells that suppress auto-aggressive T-cell responses and prevent autoimmunity (6). The deficiency in Treg function leads to a multi-organ fatal inflammatory disease in mice. In humans, recent studies demonstrated that the suppressive function of Tregs in RA patients is impaired (7, 8). Here we show ROCK2 signaling has a negative impact on IL-2 and IL-10 (Figure 1), cytokines required for appropriate function of Tregs. Moreover, pharmacological inhibition of ROCK2 increases Treg suppressive function, shift Th17/Treg balance and effectively down-regulates ongoing auto-reactive immune responses in vivo as was demonstrated in a mouse model of collagen-induced arthritis. In conclusion, this work provides new insights into molecular mechanism of ROCK2-mediated modulation of the immune response in man and has profound implications for development of selective ROCK2 inhibitor as a new therapeutic target for autoimmunity treatment.
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