Asymmetric synthesis of novel N-(1-phenyl-2,3-dihydroxypropyl)arachidonylamides and evaluation of their anti-inflammatory activity.

AIMS: To design and synthesize novel N-(1-phenyl-2,3-dihydroxypropyl)arachidonylamides and evaluate their analgesic and anti-inflammatory potential. MAIN METHODS: The murine macrophage cell line RAW 264.7 has been widely used as a model for inflammatory responses in vitro. Our model consists of cultured monolayers of RAW 264.7 cells in which media concentrations of 15-deoxy-Δ(13,14)-PGJ2 (PGJ) are measured by ELISA following LPS (10ng/ml) stimulation and treatment with 0.1, 0.3, 1.0, 3.0 and 10µM concentrations of the compounds. KEY FINDINGS: Our data indicate that several of our compounds have the capacity to increase production of PGJ and may also increase the occurrence of programmed cell death (apoptosis). SIGNIFICANCE: Thus these agents are potential candidates for the therapy of conditions characterized by ongoing (chronic) inflammation and its associated pain.

N-Amino acid linoleoyl conjugates: anti-inflammatory activities.

Several N-linked amino acid-linoleic acid conjugates were studied for their potential as anti inflammatory agents. The parent molecule, N-linoleoylglycine was tested in an in vivo model, the mouse peritonitis assay where it showed activity in reducing leukocyte migration at doses as low as 0.3mg/kg when administered by mouth in safflower oil. Harvested peritoneal cells produced elevated levels of the inflammation-resolving eicosanoid 15-deoxy-Δ(13,14)-PGJ(2). These results are similar to those obtained in earlier studies with N-arachidonoylglycine. An in vitro model using mouse macrophage RAW cells was used to evaluate a small group of structural analogs for their ability to stimulate 15-deoxy-Δ(13,14)-PGJ(2) production. The d-alanine derivative was the most active while the d-phenylalanine showed almost no response. A high degree of stereo specificity was observed comparing the d and l alanine isomers; the latter being the less active. It was concluded that linoleic acid conjugates could provide suitable templates in a drug discovery program leading to novel agents for promoting the resolution of chronic inflammation.

Resolution of inflammation by N-arachidonoylglycine.

N-arachidonoylglycine (NAgly) is an endogenous signaling lipid that is a member of the eicosanoid super family and is related to anandamide. It shows anti-inflammatory activity in vivo in the mouse peritonitis model where it reduces migration of inflammatory leukocytes following injection of pro-inflammatory agents into the peritoneal cavity. Using cell culture models, including GPR18 transfected HEK-293 cells, evidence is presented that the orphan receptor GPR18 is involved in this action. Increases in free arachidonic acid, and robust stimulation of anti-inflammatory eicosanoids were observed at low micromolar concentrations. These included 15-deoxy-delta-13,14-PGJ(2) and lipoxin A(4) both of which are believed to mediate the resolution stage of inflammation. It was further shown that NAgly might act via GPR18 activation in promoting the number of Trypan Blue stained cells, a possible indicator of programmed cell death. Thus, we hypothesize that NAgly induces the death of inflammatory cells, a process that is considered to be important for the resolution of inflammation.

Coexpression of oncostatin M and its receptors and evidence for STAT3 activation in human ovarian carcinomas.

The expression of oncostatin M and leukemia inhibitory factor (LIF), JAK-STAT activators and members of the interleukin-6 family of cytokines, were examined in a series of primary ovarian carcinomas using immunohistochemistry. The malignant epithelial cells of all 29 ovarian carcinomas examined expressed oncostatin M; none expressed LIF. Oncostatin M can activate two related receptors, one consisting of a low-affinity LIF receptor subunit, LIFR beta, which forms a heterocomplex with the gp130 signal transducing protein and can recognize both oncostatin M and LIF, and a second heterocomplex consisting of a subunit that specifically recognizes oncostatin M, OSMR beta, and the gp130 protein. By immunohistochemistry, 25 of 25 ovarian carcinomas examined expressed the LIFR beta subunit in the malignant epithelial cells (all samples express gp130), and two-thirds the ovarian carcinomas studied expressed OSMR beta mRNA as determined by RT-PCR. Thus oncostatin M and its receptors are commonly coexpressed in malignant ovarian epithelial cells, and represent a potential autocrine loop in this tumor type. STAT3, of one the signaling proteins downstream of the oncostatin M/LIF receptors, was found in its phosphorylated, activated form (phosphotyrosine 705 STAT3) in the malignant epithelial cells of 17 of 23 ovarian carcinomas examined (74%) as determined by immunohistochemistry; this suggests that this protein is constitutively activated in most ovarian carcinomas, as it is in many other human malignancies. Recombinant human Oncostatin M (rhOSM) can induce the transient tyrosine 705 phosphorylation of STAT3 in serum-starved LIFR beta/OSMR beta expressing ovarian carcinoma cell lines, but does not alter cell growth and effects only a modest increase in the apoptotic rate in these cultured cells. Oncostatin M and its receptors may be part of a network of cytokine systems within ovarian carcinomas that may act to maintain STAT3 in its activated form, a phenomenon associated with the malignant phenotype.