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β 2-adrenergic receptor ( β 2-AR) agonists are widely applied in the clinic for treatment of respiratory disease. Although their therapeutic effect mainly results from their spasmolytic activity, β -agonists have also been attributed with anti-inflammatory properties. We studied the effect of β -agonist cotreatment on TNF- α -induced activation of inflammatory signalling in astrocytes, focusing on Nuclear Factor- κ B (NF- κ B), a master regulator of inflammatory gene expression. Unexpectedly, we observed that, unlike other anti-inflammatory drugs, β -agonists did not inhibit the activity of NF- κ B. Instead, β -agonist cotreatment enhanced TNF- α -induced expression of a subset of prototypical NF- κ B target genes. The most pronounced effect was apparent for Interleukin-6 (IL-6) and the CXC chemokines CXCL2 and CXCL3. We established that cooperativity between NF- κ B and β 2-AR-induced transcriptional regulators is at the basis of the synergy. Our current research aims at further defining the molecular players regulating IL-6 transcription using a novel approach combining DNA affinity purification and mass spectrometry. Importantly, the β 2-agonist-induced potentiation of inflammatory responses was also recapitulated in vivo , in rats treated intracerebroventricularly with a combination of TNF- α and clenbuterol. Gene expression changes were furthermore reflected in a redistribution of the leukocyte subsets present in the brain. More specifically, a skewing of the T-cell population towards a double-negative phenotype, and a shift towards neutrophilic expansion in the myeloid subset were apparent. Although functional characterization of the infiltrating leukocytes remains to be done, these results suggest β -agonists can catalyze inflammatory reactions in vivo and warrant further study into their mode of action. [ABSTRACT FROM AUTHOR]

The activating JAK2-V617F mutation is found in many patients suffering from Myeloproliferative neoplasms such as Polycythemia Vera and Essential Thrombocythemia. JAK2-V617F mutation confers cytokine hypersensitivity, constitutive activation of the JAK-STAT pathway, and cytokine-independent growth. We demonstrate uncontrolled membrane binding of the multi-site docking protein Gab1 in JAK2-V617F expressing cells. We elaborated the molecular mechanism and regulation of Gab1-membrane recruitment. In line with the regulatory function of Gab1 on STAT-independent signalling we show that PI3K signalling regulates MAPK activation in JAK2-V617F-positive cells. This cross-regulation of the MAPK pathway by PI3K affects JAK2-V617F-specific target gene induction, erythroid colony formation, and regulates proliferation of JAK2-V617F-positive patient cells in a synergistically manner. In summary, our study demands to consider cross-talk mechanisms as potential new targets. This may increase the specificity for intervention approaches compared to the interference in very upstream signalling elements such as receptor kinases or receptor-associated kinases. [ABSTRACT FROM AUTHOR]

Systemic toxicity currently prevents full clinical application of type I interferons (IFNs) for the treatment of cancers and viral infections. Although the use of IFNs fused to antibodies resulted in a 10-fold increased activity towards targeted cells, off-target side effects experienced by the patients are expected to remain a major obstacle. We present a novel strategy to engineer IFNα with high in vitro and in vivo activity towards cells expressing a specific surface marker while having an up to 1000-fold reduced activity towards non-relevant cells. The concept lies in the use of immunocytokines based on IFN mutants with strongly reduced binding affinity for the IFN receptor. Such a targeted IFN recovers almost maximally its antiviral activity uniquely on the targeted cells and acts to selected cell populations in mice with targeting efficiencies up to 3 orders of magnitude. This novel concept holds promise to revitalize the clinical potential of IFNα and other cytokines. [ABSTRACT FROM AUTHOR]