Discovery of Clinical Candidate GLPG3970: A Potent and Selective Dual SIK2/SIK3 Inhibitor for the Treatment of Autoimmune and Inflammatory Diseases.

The salt-inducible kinases (SIKs) SIK1, SIK2, and SIK3 belong to the adenosine monophosphate-activated protein kinase (AMPK) family of serine/threonine kinases. SIK inhibition represents a new therapeutic approach modulating pro-inflammatory and immunoregulatory pathways that holds potential for the treatment of inflammatory diseases. Here, we describe the identification of GLPG3970 (32), a first-in-class dual SIK2/SIK3 inhibitor with selectivity against SIK1 (IC50 of 282.8 nM on SIK1, 7.8 nM on SIK2 and 3.8 nM on SIK3). We outline efforts made to increase selectivity against SIK1 and improve CYP time-dependent inhibition properties through the structure-activity relationship. The dual activity of 32 in modulating the pro-inflammatory cytokine TNFα and the immunoregulatory cytokine IL-10 is demonstrated in vitro in human primary myeloid cells and human whole blood, and in vivo in mice stimulated with lipopolysaccharide. Compound 32 shows dose-dependent activity in disease-relevant mouse pharmacological models.

Safety, tolerability, pharmacokinetics, and pharmacodynamics of GLPG1690, a novel autotaxin inhibitor, to treat idiopathic pulmonary fibrosis (FLORA): a phase 2a randomised placebo-controlled trial.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) causes irreversible loss of lung function. People with IPF have increased concentrations of autotaxin in lung tissue and lysophosphatidic acid (LPA) in bronchoalveolar lavage fluid and exhaled condensate. GLPG1690 (Galapagos, Mechelen, Belgium) is a novel, potent, selective autotaxin inhibitor with good oral exposure. We explored the effects of GLPG1690 in patients with IPF. METHODS: This was a randomised, double-blind, placebo-controlled phase 2a study done in 17 centres in Italy, Ukraine and the UK. Eligible patients were aged 40 years or older, non-smokers, not taking pirfenidone or nintedanib, and had a centrally confirmed diagnosis of IPF. We used a computer-generated randomisation schedule to assign patients 1:3 to receive placebo or 600 mg oral GLPG1690 once daily for 12 weeks. The primary outcomes were safety (adverse events), tolerability, pharmacokinetics, and pharmacodynamics. Spirometry was assessed as a secondary outcome. This trial is registered with ClinicalTrials.gov, number NCT02738801. FINDINGS: Between March 24, 2016, and May 2, 2017, 72 patients were screened., of whom 49 were ineligible and 23 were enrolled in eight centres (six in Ukraine and two in the UK). Six patients were assigned to receive placebo and 17 to receive GLPG1690. 20 patients completed the study after one in each group discontinued because of adverse events and one in the GLPG1690 group withdrew consent. Four (67%) patients in the placebo group and 11 (65%) in the GLPG1690 group had treatment-emergent adverse events, most of which were mild to moderate. The most frequent events in the GLPG1690 group were infections and infestations (ten events) and respiratory, thoracic, and mediastinal disorders (eight events) with no apparent differences from the placebo group. Two (12%) patients in the GLPG1690 group had events that were judged to be related to treatment. Serious adverse events were seen in two patients in the placebo group (one had a urinary tract infection, acute kidney injury, and lower respiratory tract infection and the other had atrioventricular block, second degree) and one in the GLPG1690 group (cholangiocarcinoma that resulted in discontinuation of treatment). No patients died. The pharmacokinetic and pharmacodynamic profiles of GLPG1690 were similar to those previously shown in healthy controls. LPA C18:2 concentrations in plasma were consistently decreased. Mean change from baseline in forced vital capacity at week 12 was 25 mL (95% CI -75 to 124) for GLPG1690 and -70 mL (-208 to 68 mL) for placebo. INTERPRETATION: Our findings support further development of GLPG1690 as a novel treatment for IPF. FUNDING: Galapagos.

Neutralization of IL-17C Reduces Skin Inflammation in Mouse Models of Psoriasis and Atopic Dermatitis.

IL-17C is a functionally distinct member of the IL-17 family that was believed to play a role in the pathogenesis of psoriasis. Here we confirmed that IL-17C is involved in psoriasis and explored potential roles for IL-17C in atopic dermatitis (AD). An anti-IL-17C antibody, MOR106, was generated that potently and selectively binds to human and mouse IL-17C, thereby inhibiting the binding of IL-17C to its IL-17RE receptor. The antibody inhibited cutaneous inflammation in an IL-23-induced psoriatic-like skin inflammation model. In lesional skin of patients with AD, IL-17C expression levels were increased and localized to keratinocytes and infiltrating immune cells. To determine the contribution of IL-17C to AD pathogenesis, MOR106 was tested in two distinct in vivo models. In the calcipotriol-induced AD model, ear skin inflammation, TSLP, and IL-33 protein production in ears was suppressed by MOR106. Consistently, in the flaky tail strain mouse model, spontaneous development of AD-like skin inflammation was reduced by MOR106. Moreover, serum IgE levels, number of mast cells in skin and T helper type 2-related cytokines IL-4 and CCL17 in serum were all reduced. Overall, our results indicate that IL-17C is a central mediator of skin inflammation beyond psoriasis and is relevant in particular in AD.

Estrogen receptor-alpha mediates the brain antiinflammatory activity of estradiol.

Beyond the key role in reproductive and cognitive functions, estrogens have been shown to protect against neurodegeneration associated with acute and chronic injuries of the adult brain. Current hypotheses reconcile this activity with a direct effect of 17beta-estradiol (E2) on neurons. Here we demonstrate that brain macrophages are also involved in E2 action on the brain. Systemic administration of hormone prevents, in a time- and dose-dependent manner, the activation of microglia and the recruitment of peripheral monocytes induced by intraventricular injection of lipopolysaccharide. This effect occurs by limiting the expression of neuroinflammatory mediators, such as the matrix metalloproteinase 9 and lysosomal enzymes and complement C3 receptor, as well as by preventing morphological changes occurring in microglia during the inflammatory response. By injecting lipopolysaccharide in estrogen receptor (ER)-null mouse brains, we demonstrate that hormone action is mediated by activation of ERalpha but not of ERbeta. The specific role of ERalpha is further confirmed by comparing the effects of ERs on the matrix metalloproteinase 9 promoter activity in transient transfection assays. Finally, we report that genetic ablation of ERalpha is associated with a spontaneous reactive phenotype of microglia in specific brain regions of adult ERalpha-null mice. Altogether, these results reveal a previously undescribed function for E2 in brain and provide a mechanism for its beneficial activity on neuroinflammatory pathologies. They also underline the key role of ERalpha in brain macrophage reactivity and hint toward the usefulness of ERalpha-specific drugs in hormone replacement therapy of inflammatory diseases.

Estradiol enhances primary antigen-specific CD4 T cell responses and Th1 development in vivo. Essential role of estrogen receptor alpha expression in hematopoietic cells.

It is widely accepted that females have superior immune responses than males, but the ways by which sex hormones may enhance T cell responses are still poorly understood. In the present study, we analyzed the effect of estrogens on CD4 T cell activation and differentiation after immunization with exogenous antigens. We show that administration of low doses of 17beta-estradiol (E2) to castrated female mice results in a striking increase of antigen-specific CD4 T cell responses and in the selective development of IFN-gamma-producing cells. Quantitative assessment of the frequency of T cells bearing a public TCR beta chain CDR3 motif demonstrated that the clonal size of primary antigen-specific CD4 T cells was dramatically increased in immune lymph nodes from E2-treated mice. By using mice with disrupted estrogen receptor (ER) alpha or beta genes, we show that ERalpha, but not ERbeta, was necessary for the enhanced E2-driven Th1 cell responsiveness. Furthermore, ERalpha expression in hematopoietic cells was essential, since E2 effects on Th1 responses were only observed in mice reconstituted with bone marrow cells from ERalpha+/+, but not ERalpha-deficient mice. These results demonstrate that estrogen administration promotes strong antigen-specific Th1 cell responses in a mechanism that requires functional expression of ERalpha in hematopoietic cells.

Expression of Sox9 in granulosa cells lacking the estrogen receptors, ERalpha and ERbeta.

Ovaries from adult mice lacking both estrogen receptors ERalpha and ERbeta (ERalphabetaKO mice) contain abnormal cells sharing morphologic features with Sertoli cells, which are located mainly in the interstitial compartment. We show here that these cells express the Sertoli cell markers TIF1beta, TIF2, and Sox9. In ERalphabetaKO ovaries, Sox9 is expressed by granulosa cells before the morphologic appearance of Sertoli cells, but neither by granulosa cell precursors nor by non-Sertolian interstitial cells. These findings suggest that functional Sertoli cells can transdifferentiate from mature granulosa cells devoid of estrogen receptors as a result of Sox9 expression.

Estrogen receptor-alpha mediates the protective effects of estrogen against vascular injury.

Blood vessel cells express the 2 known estrogen receptors, alpha and beta (ERalpha, ERbeta), which are thought to mediate estrogen inhibition of vascular injury and atherosclerosis, but the relative role of ERalpha and ERbeta in these events is controversial. Estrogen inhibits the vascular injury response to the same extent in ovariectomized female wild-type mice and in the original single gene knockout mice for ERalpha (ERalphaKO(Chapel Hill) [ERalphaKO(CH)]) and ERbeta (ERbetaKO(Chapel Hill) [ERbetaKO(CH)]). In double gene knockout mice generated by crossing these animals (ERalpha,betaKO(CH)), estrogen no longer inhibits medial thickening after vascular injury, but still inhibits vascular smooth muscle cell proliferation and increases uterine weight. The partial retention of estrogen responsiveness in ERalpha,betaKO(CH) mice could be due either to the presence of a novel, unidentified estrogen receptor or to functional expression of an estrogen receptor-alpha splice variant in the parental ERalphaKO(CH) mice. To distinguish between these possibilities, we studied recently generated mice fully null for estrogen receptor alpha (ERalphaKO(Strasbourg) [ERalphaKO(St)]) and examined the effect of estrogen on the response to vascular injury. In the present study, we show that after vascular injury in ovariectomized ERalphaKO(St) mice, estrogen has no detectable effect on any measure of vascular injury, including medial area, proteoglycan deposition, or smooth muscle cell proliferation. These data demonstrate that estrogen receptor-alpha mediates the protective effects of estrogen on the response to vascular injury.