BIRB 796 (Doramapimod)

EC50: 18 nM for TNF-α in THP-1 cells

The signal transduction pathway leading to the production of TNF-α from stimulated inflammatory cells, while not fully understood, has been shown to be partially regulated by p38 mitogen activated protein (MAP) kinase. The p38 MAP kinase plays a crucial role in regulating the production of proinflammatory cytokines. Blocking this kinase may offer an effective therapy for the treatment of many inflammatory diseases. BIRB 796 is a highly potent inhibitor of p38 MAPK.

In vitro: BIRB 796 is a picomolar inhibitor of human p38 MAP kinase with a 12,000-fold increase in binding affinity. Moreover, BIRB 796 behavors as one of the most potent and slowest dissociating human p38 MAP kinase inhibitors now known [1].

In vivo: In a LPS-stimulated TNF-α synthesis mouse model, a 65% inhibition of TNF-α synthesis was observed when BIRB 796 was dosed orally at 10 mg/kg. In a model of established collagen-induced arthritis using B10.RIII mice, BIRB 796 showed a 63% inhibition of arthritis severity when dosed orally at 30 mg/kg qd [2].

Clinical trial: No clinical efficacy (Crohn’s Disease Endoscopic Index of Severity) was seen for BIRB 796 in comparison with placebo. A significant and dose-dependent decrease of C-reactive protein level was seen transiently after BIRB 796 after 1 week with a return to baseline level over time [3].

References:
[1] Pargellis C, Tong L, Churchill L, Cirillo PF, Gilmore T, Graham AG, Grob PM, Hickey ER, Moss N, Pav S, Regan J.  Inhibition of p38 MAP kinase by utilizing a novel allosteric binding site. Nat Struct Biol. 2002 Apr;9(4):268-72.
[2] Regan J, Breitfelder S, Cirillo P, Gilmore T, Graham AG, Hickey E, Klaus B, Madwed J, Moriak M, Moss N, Pargellis C, Pav S, Proto A, Swinamer A, Tong L, Torcellini C.  Pyrazole urea-based inhibitors of p38 MAP kinase: from lead compound to clinical candidate. J Med Chem. 2002 Jul 4;45(14):2994-3008.
[3] Schreiber S, Feagan B, D'Haens G, Colombel JF, Geboes K, Yurcov M, Isakov V, Golovenko O, Bernstein CN, Ludwig D, Winter T, Meier U, Yong C, Steffgen J; BIRB 796 Study Group.  Oral p38 mitogen-activated protein kinase inhibition with BIRB 796 for active Crohn's disease: a randomized, double-blind, placebo-controlled trial. Clin Gastroenterol Hepatol. 2006 Mar;4(3):325-34.


Synthesis and p38 Inhibitory Activity of Some Novel Substituted N,N'-Diarylurea Derivatives.[Pubmed:27223276]

Molecules. 2016 May 23;21(5). pii: molecules21050677.

We have identified a novel series of substituted N,N'-diarylurea p38alpha inhibitors. The inhibitory activity of the target compounds against the enzyme p38alpha, MAPKAPK2 in BHK cells, TNF-alpha release in LPS-stimulated THP-1 cells and p38alpha binding experiments were tested. Among these compounds, 25a inhibited the p38alpha enzyme with an IC50 value of 0.47 nM and a KD value of 1.54 x 10(-8) and appears to be the most promising one in the series.

Mechanisms of corticosteroid insensitivity in COPD alveolar macrophages exposed to NTHi.[Pubmed:28420398]

Respir Res. 2017 Apr 18;18(1):61.

BACKGROUND: Non-typeable Haemophilus influenza (NTHi) infection is common in COPD. Corticosteroids can have limited therapeutic effects in COPD patients. NTHi causes corticosteroid insensitive cytokine production from COPD alveolar macrophages. We investigated the mechanisms by which NTHi causes corticosteroid insensitive inflammatory responses, and the effects of NTHi exposure on COPD macrophage polarisation. METHOD: Alveolar macrophages from COPD patients and controls were exposed to NTHi in conjunction with the corticosteroid dexamethasone and/or the p38 MAPK inhibitor BIRB-796. Cytokine release, GR phosphorylation and modulation and macrophage phenotype were analysed. RESULTS: Dexamethasone significantly inhibited NTHi induced TNF-alpha, IL-6 and IL-10 from COPD macrophages but, CXCL8 was not suppressed. BIRB-796 combined with dexamethasone caused significantly greater inhibition of all cytokines than either drug alone (p < 0.05 all comparisons). NTHi caused phosphorylation of GR S226 reducing GR nuclear localisation, an effect regulated by p38 MAPK. NTHi altered macrophage polarisation by increasing IL-10 and decreasing CD36, CD206, CD163 and HLA-DR. CONCLUSION: NTHi exposure causes p38 MAPK dependent GR phosphorylation associated with decreased GR function in COPD alveolar macrophages. Combining a p38 MAPK inhibitor with corticosteroids can enhance anti-inflammatory effects during NTHi exposure of COPD alveolar macrophages. NTHi causes macrophage polarisation that favours bacterial persistence.

Stress-dependent phosphorylation of myocardin-related transcription factor A (MRTF-A) by the p38(MAPK)/MK2 axis.[Pubmed:27492266]

Sci Rep. 2016 Aug 5;6:31219.

Myocardin-related transcription factor A (MRTF-A) is a known actin-regulated transcriptional coactivator of serum response factor (SRF). Stimulation of actin polymerization activates MRTF-A by releasing it from G-actin and thus allowing it to bind to and activate SRF. Here, we compared protein phosphorylation in MK2/3-deficient cells rescued or not by ectopic expression of MK2 in two independent phosphoproteomic approaches using anisomycin-treated MEF cells and LPS-stimulated mouse macrophages, respectively. Two MRTF-A sites, Ser(351) (corresponding to Ser(312) in human) and Ser(371) (Ser(333) in human), showed significantly stronger phosphorylation (12-fold and 6-fold increase) in the cells expressing MK2. MRTF-A is phosphorylated at these sites in a stress-, but not in a mitogen-induced manner, and p38(MAPK)/MK2 catalytic activities are indispensable for this phosphorylation. MK2-mediated phosphorylation of MRTF-A at Ser(312) and Ser(333) was further confirmed in an in vitro kinase assay and using the phospho-protein kinase-D (PKD)-consensus motif antibody (anti-LXRXXpS/pT), the p38(MAPK) inhibitor BIRB-796, MK2/3-deficient cells and MRTF-A phospho-site mutants. Unexpectedly, dimerization, subcellular localization and translocation, interaction with actin, SRF or SMAD3 and transactivating potential of MRTF-A seem to be unaffected by manipulating the p38(MAPK)/MK2-dependent phosphorylations. Hence, MRTF-A is stress-dependently phosphorylated by MK2 at Ser(312) and Ser(333) with so far undetected functional and physiological consequences.

microRNA-200a silencing protects neural stem cells against cerebral ischemia/reperfusion injury.[Pubmed:28222148]

PLoS One. 2017 Feb 21;12(2):e0172178.

Neural stem cells (NSCs) play major roles in neurological recovery after cerebral infarction (CI). This study was trying to investigate whether miR-200a, a vital regulator in cell proliferation, migration and apoptosis, also has a role in oxygen-glucose deprivation/reperfusion (OGD/R) injured NSCs. In this study, primary NSCs were subjected to OGD/R conditions to mimic an in vitro CI model. Before OGD/R induction, NSCs were transfected with vector or shRNA against miR-200a to overexpress or suppress miR-200a expression. The changes in cell viability, apoptosis, migration, the expression of c-Myc, and the phosphorylation of STAT1, STAT3 and MAPK were respectively assessed. Inhibitors of STAT1/3 and MAPK, i.e., Nifuroxazide and BIRB 796, were used to administrate miR-200a-silenced NSCs, and the expressions of above mentioned proteins were detected. After OGD/R exposure, miR-200a was up-regulated in NSCs (P < 0.001). miR-200a silencing alleviated OGD/R-induced the decrease of cell viability and migration (P < 0.01); meanwhile, alleviated OGD/R-induced apoptosis via reducing Bax/Bcl-2 ratio and down-regulating p53 and cytochrome c (P < 0.01 or P < 0.001). c-Myc, p-STAT1, p-STAT3, p-MAPK were all negatively regulated by miR-200a (P < 0.01 or P < 0.001); more important, the increase of c-Myc induced by miR-200a silencing was abolished by Nifuroxazide or BIRB 796 (P < 0.01 or P < 0.001). These data indicate miR-200a silencing protects NSCs from OGD/R-induced injury, possibly via regulating the STATs/c-Myc and MAPK/c-Myc signalings.

Evaluating the Role of p38 MAPK in the Accelerated Cell Senescence of Werner Syndrome Fibroblasts.[Pubmed:27136566]

Pharmaceuticals (Basel). 2016 Apr 28;9(2). pii: ph9020023.

Progeroid syndromes show features of accelerated ageing and are used as models for human ageing, of which Werner syndrome (WS) is one of the most widely studied. WS fibroblasts show accelerated senescence that may result from p38 MAP kinase activation since it is prevented by the p38 inhibitor SB203580. Thus, small molecule inhibition of p38-signalling may be a therapeutic strategy for WS. To develop this approach issues such as the in vivo toxicity and kinase selectivity of existing p38 inhibitors need to be addressed, so as to strengthen the evidence that p38 itself plays a critical role in mediating the effect of SB203580, and to find an inhibitor suitable for in vivo use. In this work we used a panel of different p38 inhibitors selected for: (1) having been used successfully in vivo in either animal models or human clinical trials; (2) different modes of binding to p38; and (3) different off-target kinase specificity profiles, in order to critically address the role of p38 in the premature senescence seen in WS cells. Our findings confirmed the involvement of p38 in accelerated cell senescence and identified p38 inhibitors suitable for in vivo use in WS, with BIRB 796 the most effective.

Design, synthesis, and biological evaluation of novel Tri- and tetrasubstituted imidazoles as highly potent and specific ATP-mimetic inhibitors of p38 MAP kinase: focus on optimized interactions with the enzyme's surface-exposed front region.[Pubmed:18578517]

J Med Chem. 2008 Jul 24;51(14):4122-49.

The synthesis, biological testing, and SAR of novel 2,4,5- and 1,2,4,5-substituted 2-thioimidazoles are described. Amino, oxy, or thioxy substituents at the 2-position of the pyridinyl moiety were evaluated for their contributions to inhibitor potency and selectivity against p38 mitogen activated protein kinase (p38 MAPK) as well as for the ability to minimize cytochrome P450 (CYP450) inhibition. Incorporation of polar substituted (cyclo)aliphatic amino substituents (e.g., tetrahydropyranylamino), which positively interacted with the surface-exposed front region (hydrophobic region II) of the enzyme led to the identification of extremely potent p38 MAPK inhibitors with p38 IC 50 values in the low nanomolar range. Approximately 90 pyridinylimidazole-based compounds with a range of potencies against p38alpha MAP kinase were further investigated for their ability to inhibit the release of tumor necrosis factor-alpha (TNFalpha) and/or interleukin-1beta (IL-1beta) from human whole blood. Some of the most promising drug candidates underwent selectivity profiling against a panel of 17 different kinases besides p38alpha and/or were tested for their interaction potential toward a number of metabolically relevant CYP450 isozymes.

BIRB796 inhibits all p38 MAPK isoforms in vitro and in vivo.[Pubmed:15755732]

J Biol Chem. 2005 May 20;280(20):19472-9.

The compound BIRB796 inhibits the stress-activated protein kinases p38alpha and p38beta and is undergoing clinical trials for the treatment of inflammatory diseases. Here we report that BIRB796 also inhibits the activity and the activation of SAPK3/p38gamma. This occurs at higher concentrations of BIRB796 than those that inhibit p38alpha and p38beta and at lower concentrations than those that inhibit the activation of JNK isoforms. We also show that at these concentrations, BIRB796 blocks the stress-induced phosphorylation of the scaffold protein SAP97, further establishing that this is a physiological substrate of SAPK3/p38gamma. Our results demonstrate that BIRB796, in combination with SB203580, a compound that inhibits p38alpha and p38beta, but not the other p38 isoforms, can be used to identify physiological substrates of SAPK3/p38gamma as well as those of p38alpha and p38beta.

Structure-activity relationships of the p38alpha MAP kinase inhibitor 1-(5-tert-butyl-2-p-tolyl-2H-pyrazol-3-yl)-3-[4-(2-morpholin-4-yl-ethoxy)naph- thalen-1-yl]urea (BIRB 796).[Pubmed:14561087]

J Med Chem. 2003 Oct 23;46(22):4676-86.

We report on the structure-activity relationships (SAR) of 1-(5-tert-butyl-2-p-tolyl-2H-pyrazol-3-yl)-3-[4-(2-morpholin-4-yl-ethoxy)naphthal en-1-yl]urea (BIRB 796), an inhibitor of p38alpha MAP kinase which has advanced into human clinical trials for the treatment of autoimmune diseases. Thermal denaturation was used to establish molecular binding affinities for this class of p38alpha inhibitors. The tert-butyl group remains a critical binding element by occupying a lipophilic domain in the kinase which is exposed upon rearrangement of the activation loop. An aromatic ring attached to N-2 of the pyrazole nucleus provides important pi-CH(2) interactions with the kinase. The role of groups attached through an ethoxy group to the 4-position of the naphthalene and directed into the ATP-binding domain is elucidated. Pharmacophores with good hydrogen bonding potential, such as morpholine, pyridine, and imidazole, shift the melting temperature of p38alpha by 16-17 degrees C translating into K(d) values of 50-100 pM. Finally, we describe several compounds that potently inhibit TNF-alpha production when dosed orally in mice.

Doramapimod (BIRB 796) is an orally active, highly potent p38 MAPK inhibitor, which has an IC50 for p38α=38 nM, for p38β=65 nM, for p38γ=200 nM, and for p38δ=520 nM. Doramapimod (BIRB 796) has picomolar affinity for p38 kinase (Kd=0.1 nM). Doramapimod (BIRB 796) also inhibits B-Raf with an IC50 of 83 nM.

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