Selective inhibitors of tumor progression loci-2 (Tpl2) kinase with potent inhibition of TNF-alpha production in human whole blood.

Tpl2 (cot/MAP3K8) is an upstream kinase of MEK in the ERK pathway. It plays an important role in Tumor Necrosis Factor-alpha (TNF-alpha) production and signaling. We have discovered that 8-halo-4-(3-chloro-4-fluoro-phenylamino)-6-[(1H-[1,2,3]triazol-4-ylmethyl)-amino]-quinoline-3-carbonitriles (4) are potent inhibitors of this enzyme. In order to improve the inhibition of TNF-alpha production in LPS-stimulated human blood, a series of analogs with a variety of substitutions around the triazole moiety were studied. We found that a cyclic amine group appended to the triazole ring could considerably enhance potency, aqueous solubility, and cell membrane permeability. Optimization of these cyclic amine groups led to the identification of 8-chloro-4-(3-chloro-4-fluorophenylamino)-6-((1-(1-ethylpiperidin-4-yl)-1H-1,2,3-triazol-4-yl)methylamino)quinoline-3-carbonitrile (34). In a LPS-stimulated rat inflammation model, compound 34 showed good efficacy in inhibiting TNF-alpha production.

PD-1/PD-L1, but not PD-1/PD-L2, interactions regulate the severity of experimental autoimmune encephalomyelitis.

Interactions between PD-1 and its two differentially expressed ligands, PD-L1 and PD-L2, attenuate T cell activation and effector function. To determine the role of these molecules in autoimmune disease of the CNS, PD-1-/-, PD-L1-/- and PD-L2-/- mice were generated and immunized to induce experimental autoimmune encephalomyelitis (EAE). PD-1-/- and PD-L1-/- mice developed more severe EAE than wild type and PD-L2-/- mice. Consistent with this, PD-1-/- and PD-L1-/- cells produced elevated levels of the pro-inflammatory cytokines IFN-gamma, TNF, IL-6 and IL-17. These results demonstrate that interactions between PD-1/PD-L1, but not PD-1/PDL-2, are crucial in attenuating T cell responses in EAE.

Human bronchial epithelial cells express and secrete MMP-12.

Matrix metalloproteinases (MMPs) degrade extracellular matrix proteins, which may be responsible for enlargement of alveoli in chronic obstructive pulmonary disease (COPD) and remodeling of pulmonary tissue associated with chronic asthma. Here, we provide novel evidence that MMP-12 is expressed and secreted by normal human bronchial epithelial cell cultures (NHBECs) and reveal the regulation of MMP-12 gene expression by tumor necrosis factor-alpha (TNF-alpha), epidermal growth factor (EGF), and interferon gamma (IFN-gamma). Reverse transcription-polymerase chain reaction analyses demonstrated MMP-12 mRNA presence in unstimulated differentiated NHBEC cultures. Cultures stimulated independently with EGF or IFN-gamma failed to alter MMP-12 mRNA abundance, while TNF-alpha, TNF-alpha+EGF, or TNF-alpha+IFN-gamma elicited relatively early (6 h) peak increases in MMP-12 mRNA levels. Western blot analyses specifically indicated the presence of MMP-12 in differentiated NHBEC-conditioned media. These findings indicate that the bronchial epithelium may be an important source of elastolytic activity in COPD and tissue remodeling in chronic asthma.

Local delivery of granulocyte macrophage colony-stimulating factor by retrovirally transduced antigen-specific T cells leads to severe, chronic experimental autoimmune encephalomyelitis in mice.

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease of the central nervous system (CNS) that can be induced in susceptible mice by the transfer of autoreactive T cells that recognize myelin basic protein (MBP). The onset and subsequent recovery from disease are associated with distinct patterns of cytokine and chemokine expression within the inflammatory lesions of the CNS. Given the likely importance of the local cytokine milieu in regulating the disease process, it would be preferable to administer cytokines locally to the CNS and reduce systemic delivery in order to evaluate their immunoregulatory roles in EAE. For this purpose, we have used retrovirally transduced T cells from MBP-specific T cell receptor transgenic mice in an attempt to target cytokine delivery to the CNS where MBP is primarily expressed. We have found that T cells expressing granulocyte macrophage colony-stimulating factor (GM-CSF) induce severe, chronic EAE from which mice fail to recover. Our results indicate that increased local GM-CSF expression could play an important role in inducing chronic EAE.