Autoantigen-based immunotherapeutics have been shown to activate regulatory responses capable of inhibiting T cell-mediated autoimmune disease in animal models. However, their efficacy generally declines, as treatment occurs later in the disease process, and their mechanism of action is a matter of intense debate. Here, we report that the early administration of beta cell autoantigens (betaCAAs) to nonobese diabetic (NOD) mice broadly diverts the natural development of potentially pathogenic Thl-biased autoimmune responses toward the Th2 phenotype through Th2 spreading. With disease progression, there was a steady decline in the ability of betaCAA treatment to promote Th2-type cellular and humoral autoimmunity. Late in the disease process, some betaCAAs were still able to induce Th2 responses and Th2 spreading (although to a much lesser extent), while other autoantigens were not. This attenuation of inducible Th2 immunity with disease progression is likely to reflect a reduction in the availability of uncommitted autoantigen-reactive precursor T cells. These findings suggest that there are inherent differences in the frequency of betaCAA-reactive T cells and that, in advanced stages of autoimmune disease, regulatory responses may be best elicited with target tissue Ags against which large uncommitted T cell pools are still available. Since individuals presenting the first signs of autoimmune disease are likely to already have an advanced disease process, these findings may be useful for the rational design of Ag-based immunotherapeutics.