ABSTRACT
Cytokines, growth factors, and other chemical messengers rely on a class of intracellular nonreceptor tyrosine kinases known as Janus kinases (JAKs) to rapidly transduce intracellular signals. A number of these cytokines are critical for lymphocyte development and mediating immune responses. JAK3 is of particular interest due to its importance in immune function and its expression, which is largely confined to lymphocytes, thus limiting the potential impact of JAK3 inhibition on nonimmune physiology. The aim of this study was to evaluate the potency and selectivity of the investigational JAK3 inhibitor VX-509 (decernotinib) [(R)-2-((2-(1H-pyrrolo[2,3-b]pyridin-3-yl)pyrimidin-4-yl)amino)-2-methyl-N-(2,2,2-trifluoroethyl)butanamide] against JAK3 kinase activity and inhibition of JAK3-mediated signaling in vitro and JAK3-dependent physiologic processes in vivo. These results demonstrate that VX-509 potently inhibits JAK3 in enzyme assays (Ki = 2.5 nM + 0.7 nM) and cellular assays dependent on JAK3 activity (IC50 range, 50-170 nM), with limited or no measurable potency against other JAK isotypes or non-JAK kinases. VX-509 also showed activity in two animal models of aberrant immune function. VX-509 treatment resulted in dose-dependent reduction in ankle swelling and paw weight and improved paw histopathology scores in the rat collagen-induced arthritis model. In a mouse model of oxazolone-induced delayed-type hypersensitivity, VX-509 reduced the T cell-mediated inflammatory response in skin. These findings demonstrate that VX-509 is a selective and potent inhibitor of JAK3 in vitro and modulates proinflammatory response in models of immune-mediated diseases, such as collagen-induced arthritis and delayed-type hypersensitivity. The data support evaluation of VX-509 for treatment of patients with autoimmune and inflammatory diseases such as rheumatoid arthritis.
Subject(s)
Autoimmune Diseases/drug therapy , Heterocyclic Compounds, 2-Ring/pharmacology , Janus Kinase 3/antagonists & inhibitors , Protein Kinase Inhibitors/pharmacology , Valine/analogs & derivatives , Animals , Arthritis, Experimental/drug therapy , Arthritis, Experimental/enzymology , Arthritis, Experimental/immunology , Autoimmune Diseases/enzymology , Autoimmune Diseases/immunology , Disease Models, Animal , Edema/drug therapy , Edema/enzymology , Edema/immunology , Female , Heterocyclic Compounds, 2-Ring/therapeutic use , Humans , Hypersensitivity, Delayed , Inflammation/drug therapy , Janus Kinase 1/antagonists & inhibitors , Janus Kinase 2/antagonists & inhibitors , Male , Mice , Protein Kinase Inhibitors/therapeutic use , Rats , Valine/pharmacology , Valine/therapeutic useABSTRACT
The enzyme inosine monophosphate dehydrogenase (IMPDH) catalyzes the rate-limiting step in the de novo biosynthesis of guanine nucleotides. Inhibition of IMPDH leads to immunosuppression by decreasing guanine nucleotides that are required for the proliferation of lymphocytes. IMPDH activity is mediated by two highly conserved isoforms, type I and type II. We have characterized the mRNA and protein expression of the two isoforms in a variety of human tissues, peripheral blood mononuclear cells (PBMCs), and selected cell lines to investigate their regulation. Type I mRNA was expressed in most tissues with high expression in PBMCs and low expression in thymus. IMPDH type II transcript was also detected in most tissues with low expression in spleen and PBMCs. In PBMCs, induction of both type I and type II mRNAs was observed within 12 hr of mitogenic stimulation. Using type-selective IMPDH antibodies, an increase in the levels of type I and type II proteins was observed after mitogenic stimulation. The effect of two IMPDH inhibitors, MPA and VX-497, was investigated on the expression of type I and type II isoforms. VX-497 is an orally bioavailable, potent and reversible inhibitor of IMPDH, with broad applicability in many viral and immune system-mediated diseases. MPA and VX-497 inhibit both isoforms of IMPDH in vitro. Prolonged treatment of lymphocytes with either VX-497 or MPA did not lead to an increase in type I or type II IMPDH protein levels. These results are discussed in the context of IMPDH being a target for immunosuppressive, anti-viral and anti-cancer therapy.
Subject(s)
IMP Dehydrogenase/metabolism , Isoenzymes/metabolism , Lymphocytes/enzymology , Carbamates/pharmacology , Enzyme Inhibitors/pharmacology , Humans , IMP Dehydrogenase/antagonists & inhibitors , IMP Dehydrogenase/genetics , Isoenzymes/genetics , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/enzymology , Mycophenolic Acid/pharmacology , Phenylurea Compounds/pharmacology , RNA, Messenger/metabolism , Tumor Cells, CulturedABSTRACT
Inosine 5'-monophosphate dehydrogenase (IMPDH) enzyme catalyzes the rate-limiting step in the de novo biosynthesis of guanine nucleotides. Proliferation of lymphocytes is critically dependent on this de novo nucleotide synthesis pathway. Hence, IMPDH is an attractive target for the development of immunosuppressive drugs. VX-148 is a novel, uncompetitive IMPDH inhibitor with a K(i) value of 6 nM against IMPDH type II enzyme. VX-148 is slightly more potent than mycophenolic acid and VX-497 in inhibiting the proliferation of mitogen-stimulated primary human lymphocytes (IC(50) value of ~80 nM). The inhibitory activity of VX-148 is alleviated in the presence of exogenous guanosine. VX-148 does not inhibit proliferation of nonlymphoid cell types such as fibroblasts, indicating selectivity for inhibition of IMPDH activity. VX-148 is orally bioavailable in rats and mice; oral administration of VX-148 inhibits primary antibody response in mice in a dose-dependent manner with an ED(50) value of 38 mg/kg b.i.d. VX-148 significantly prolongs skin graft survival at 100 mg/kg b.i.d. in mice. These results demonstrate that VX-148 is a potent and specific IMPDH inhibitor with a favorable pharmacokinetic profile and good pharmacological activity in mice, and thus support development of VX-148 as an immunosuppressive drug.
