Taken together, our results indicate great potential for the use of S28463 as an antiinflammatory therapeutic agent for the management of chronic asthma.

Asthma is one of the leading causes of childhood hospitalization, and its incidence is on the rise throughout the world. Currently, the standard treatment for asthma is the use of corticosteroids to try to suppress the inflammatory reaction taking place in the bronchial tree. Using a murine model of atopic allergic asthma employing a methacholine-hyperresponsive (A/J) as well as a hyporesponsive (C57BL/6) strain of mice sensitized and challenged with ovalbumin, we show that treatment with a synthetic Toll-like receptor 7 (TLR7) ligand (S-28463, a member of the imidazoquinoline family) prevents development of the asthmatic phenotype. Treatment with S-28463 resulted in a reduction of airway resistance and elastance following ovalbumin sensitization and challenge. This was accompanied by a dramatic reduction in infiltration of leukocytes, especially eosinophils, into the lungs of both C57BL/6 and A/J mice following OVA challenge. Treatment with S-28463 also abolished both the elevation in serum IgE level as well as the induction of IL-4, IL-5, and IL-13 by OVA challenge. The protective effects of S-28463 were also observed in MK2 knockout, but not MYD88 knockout, mice. We did not observe a switch in cytokine profile from T(H)2 to T(H)1, as both IL-12p70 and IFN-gamma levels were reduced following S-28463 treatment. These results clearly demonstrate the anti-inflammatory effect of imidazoquinolines in an allergic asthma model as well as the clinical potential of TLR7 ligands in the treatment of allergic diseases.

Secretory leukocyte protease inhibitor (SLPI) is an anti-inflammatory protein that is observed at high levels in asthma patients. Resiquimod, a TLR7/8 ligand, is protective against acute and chronic asthma, and it increases SLPI expression of macrophages in vitro. However, the protective role played by SLPI and the interactions between the SLPI and resiquimod pathways in the immune response occurring in allergic asthma have not been fully elucidated. To evaluate the role of SLPI in the development of asthma phenotypes and the effect of resiquimod treatment on SLPI, we assessed airway resistance and inflammatory parameters in the lungs of OVA-induced asthmatic SLPI transgenic and knockout mice and in mice treated with resiquimod. Compared with wild-type mice, allergic SLPI transgenic mice showed a decrease in lung resistance (p < 0.001), airway eosinophilia (p < 0.001), goblet cell hyperplasia (p < 0.001), and plasma IgE levels (p < 0.001). Allergic SLPI knockout mice displayed phenotype changes significantly more severe compared with wild-type mice. These phenotypes included lung resistance (p < 0.001), airway eosinophilia (p < 0.001), goblet cell hyperplasia (p < 0.001), cytokine levels in the lungs (p < 0.05), and plasma IgE levels (p < 0.001). Treatment of asthmatic transgenic mice with resiquimod increased the expression of SLPI and decreased inflammation in the lungs; resiquimod treatment was still effective in asthmatic SLPI knockout mice. Taken together, our study showed that the expression of SLPI protects against allergic asthma phenotypes, and treatment by resiquimod is independent of SLPI expression, displayed through the use of transgenic and knockout SLPI mice.

Resiquimod is a compound belonging to the imidazoquinoline family of compounds known to signal through Toll-like receptor 7. Resiquimod treatment has been demonstrated to inhibit the development of allergen induced asthma in experimental models. The aim of the present study was to elucidate the molecular processes that were altered following resiquimod treatment and allergen challenge in a mouse model of allergic asthma. Employing microarray analysis, we have characterized the "asthmatic" transcriptome of the lungs of A/J and C57BL/6 mice and determined that it includes genes involved in the control of cell cycle progression, the complement and coagulation cascades, and chemokine signaling. Our results demonstrated that resiquimod treatment resulted in the normalization of the expression of genes involved with airway remodeling, and generally, chemokine signaling. Resiquimod treatment also altered the expression of cell adhesion molecules, and molecules involved in natural killer (NK) cell-mediated cytotoxicity. Furthermore, we have demonstrated that systemic resiquimod administration resulted in the recruitment of NK cells to the lungs and livers of the mice, although no causal relationship between NK cell recruitment and treatment efficacy was found. Overall, our findings identified several genes, important in the development of asthma pathology, that were normalized following resiquimod treatment, thus improving our understanding of the molecular consequences of resiquimod treatment in the lung milieu. The recruitment of NK cells to the lungs may also have application in the treatment of virally induced asthma exacerbations.

Airway responsiveness is the ability of the airways to respond to bronchoconstricting stimuli by reducing their diameter. Airway hyperresponsiveness has been associated with asthma susceptibility in both humans and murine models, and it has been shown to be a complex and heritable trait. In particular, the A/J mouse strain is known to have hyperresponsive airways, while the C57BL/6 strain is known to be relatively refractory to bronchoconstricting stimuli. We analyzed recombinant congenic strains (RCS) of mice generated from these hyper- and hyporesponsive parental strains to identify genetic loci underlying the trait of airway responsiveness in response to methacholine as assessed by whole-body plethysmography. Our screen identified 16 chromosomal regions significantly associated with airway hyperresponsiveness (genome-wide P 73 cM), chromosome 7 (>63 cM), chromosome 8 (52-67 cM), chromosome 10 (3-7 cM and >68 cM), and chromosome 12 (25-38 cM and >52 cM). Our data identify several likely candidate genes from the 16 regions, including Ddr2, Hc, Fbn1, Flt3, Utrn, Enpp2, and Tsc.