Background: Recently, it has been demonstrated that immunostimulatory DNA sequences (ISS) containing CpG motifs prevent the development of allergic airway responses in murine models of disease. However, few studies have addressed the issue of whether these agents will reverse established Tm_H2-driven allergic airway responses. Objective: The aim of this study was to determine whether intradermal delivery of an immunogenic protein of ragweed pollen linked to an immunostimulatory DNA sequence could reverse an established allergic response in the mouse lung. Methods: Mice sensitized and challenged with ragweed pollen extract were treated intradermally twice at 1-week intervals with an ISS chemically linked to Amb a 1 (Amb a 1-ISS). One week after the Amb a 1-ISS treatment, mice were rechallenged intratracheally with ragweed extract, and airway responses were assessed. Results: Amb a 1-ISS treatment of ragweed-sensitized and ragweed-challenged mice significantly reversed allergen-induced airway hyperresponsiveness and suppressed the total number of eosinophils in bronchoalveolar lavage fluid. The inhibitory effect of Amb a 1-ISS was associated with a marked increase in IFN-γ levels by Amb a 1-stimulated splenocytes and a shift in the antibody profile from a T_H2-directed IgG1 response to a T_H1-directed IgG2a response. Interestingly, the inhibitory effect of Amb a 1-ISS on allergen-driven airway hyperresponsiveness was independent of suppression of T_H2 cytokine production. Conclusion: These results demonstrate that intradermal delivery of allergen-specific DNA conjugates can reverse established allergic responses in the murine lung, supporting their potential use in the treatment of human asthma. (J Allergy Clin Immunol 2002;109:455-62.)