Dimeric but Not Monomeric Soluble CD40 Prolongs Allograft Survival and Generates Regulatory T Cells That Inhibit CTL Function

Background: We previously reported that adenovirus mediated CD40Ig gene therapy (AdCD40Ig) induced long-term acceptance of fully allogeneic rat cardiac allografts, however, the underlying mechanism has not been fully clarified. To address this we have compared the ability of dimeric and monomeric soluble CD40 to prolong allograft survival in vivo and generate regulatory T cells in vitro.

Methods: The ability of CD40Ig (soluble dimmer, containing an Fc region) or CD40/Myc/His (soluble monomer, lacking an Fc region) therapy to generate CD4CD25 regulatory T cells in vitro and to prevent rejection of rat cardiac allografts (ACI to LEWIS) was compared. Immunoregulatory capacity of regulatory T cells generated was determined by suppression of alloantigen specific proliferation and cytotoxicity.

Results: Dimeric soluble CD40Ig did not inhibit CD4 T cell proliferation but rather promoted IL-2 production and the generation of CD4CD25 T cells, which regulated alloantigen-specific cytotoxic T lymphocyte activity. Treatment with either AdCD40Ig or purified soluble CD40Ig prolonged the survival of rat cardiac allografts. In contrast, although monomeric soluble CD40/Myc/His suppressed IL-12 production in a similar manner to that achieved by CD40Ig, it did not augment IL-2 production. Moreover, while CD40/Myc/His also generated CD4CD25 T cells, they did not exhibit regulatory activity and administration of soluble CD40/Myc/His failed to prolong cardiac allograft survival.

Conclusions: These results suggest signaling through CD154 in addition to blocking of CD154-CD40 interaction is important for the immunomodulatory effects of soluble CD40Ig. Taken together, our results provide new insight into the mechanism of immunomodulation by soluble CD40 constructs.