Systemic depletion of CD4+CD25+ T regulatory cells (Tregs) enhances the therapeutic anti-tumor effects of a recombinant vaccinia virus vaccine encoding tumor-specific MHC Class I and Class II dominant epitopes and the GM-CSF gene

1416 MB49, a highly aggressive C57BL/6 bladder tumor expressing the male minor histocompatibility antigen HY, grows progressively in syngeneic female mice (fB6) without inducing immunity to HY. This lack of responsiveness is associated with an IL-10-dependent reduction of dendritic cell function. While optimal immunization of tumor-bearing mice using recombinant vaccinia virus encoding the MHC Class I dominant epitope from the Uty gene (VV-Uty) plus VV-GM-CSF can result in systemic immunity, it does not significantly affect tumor growth. CD4+CD25+ Treg cells are associated with in vivo tumors expressing IL-10 and TGF-β (both expressed in MB49). Prophylactic depletion of Tregs using the anti-CD25 monoclonal antibody (MAb) PC61 resulted in complete rejection of MB49 by fB6 mice; however, Treg depletion in MB49-bearing mice (therapeutic) failed to elicit tumor regression. To determine the effects of Treg depletion on the ability to productively immunize tumor-bearing mice to HY, we treated MB49-bearing fB6 mice with combined Treg depletion plus VV-Uty, VV-GM-CSF. While treatment failed to result in tumor regression, Treg depletion significantly enhanced the levels of systemic immunity to HY as measured by splenic CTL, increased numbers of CD8+ Uty-tetramer positive T cells, and enhanced HY-specific IFN-γ production by splenocytes. Because studies have shown that rejection of male B6 skin by female B6 mice is dependent on a CD4 response, we generated a recombinant vaccinia virus encoding the MHC Class II dominant epitope of HY from the Dby gene and examined its ability to immunize and affect tumor growth in MB49-bearing fB6 mice. Treg depletion plus immunization with VV-Dby, VV-GM-CSF resulted in complete tumor regression in 8/24 fB6 and significant prolongation of survival with stable disease in an additional 3/24 fB6 compared to controls. Regression was accompanied by significantly increased CTL activity and IFN-γ production. We conclude that Treg depletion significantly enhances the therapeutic anti-tumor effect of immunization using recombinant vaccinia vaccines. We stress that this is the first antigen-based vaccine strategy that we have found to reduce the growth of this highly aggressive murine bladder tumor. In addition, combining Treg depletion with immunization using a dominant MHC Class II epitope may result in an even greater overall therapeutic response. These results indicate that combination therapy with Treg depletion and vaccinia-based antigen-specific immunization using MHC Class I and II epitopes may represent a more effective approach to tumor therapy and, we believe, warrants consideration as a clinical candidate for therapy. Supported by USPHS CA42908.