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Mice deficient in the E3 ubiquitin ligase Cbl-b have CD28-independent T cells and develop autoimmunity. We previously reported that Cbl-b−/− CD4+CD25− T effector cells are resistant in vitro to the antiproliferative effects of CD4+CD25+ regulatory T cells and TGF-β. We have now asked whether the resistance noted in Cbl-b−/− T cells is restricted solely to TGF-β’s antiproliferative effects, whether the TGF-β resistance has in vivo relevance, and whether a defect can be identified in the TGF-β signaling pathway. We now demonstrate the following: 1) in vitro, Cbl-b deficiency prevents the TGF-β-mediated induction of Foxp3+ functional regulatory T cells; 2) in vivo, Cbl-b−/− mice show a significantly enhanced response to a tumor that is strictly TGF-β regulated; and 3) Cbl-b−/− T effector cells have defective TGF-β-mediated Smad2 phosphorylation. These studies are the first to document that the E3 ubiquitin ligase Cbl-b plays an integral role in T cell TGF-β signaling, and that its absence results in multifunctional TGF-β-related defects that have important disease-related implications.

In this study, we show that engagement of CTLA-4 on tumor-infiltrating lymphocytes from low-dose melphalan (l-phenylalanine mustard (l-PAM))-treated MOPC-315 tumor bearers led to IL-10 secretion. In addition, the inhibitory activity of CTLA-4 ligation for IFN-γ secretion following stimulation with anti-CD3 plus anti-CD28 mAb depended on IL-10 production. Consistent with the importance of IL-10 for CTLA-4-mediated inhibition, administration of neutralizing anti-IL-10 mAb to low-dose l-PAM-treated MOPC-315 tumor bearers (administration of blocking anti-CTLA-4 mAb) resulted in enhanced tumor-infiltrating lymphocyte-mediated anti-MOPC-315 cytotoxicity and led to complete tumor eradication in a higher percentage of mice than that observed with low-dose l-PAM alone. The percentage of MOPC-315 tumor-bearing mice cured following administration of neutralizing anti-IL-10 mAb to low-dose l-PAM-treated MOPC-315 tumor bearers was comparable to that observed following administration of blocking anti-CTLA-4 mAb. Moreover, IL-10 neutralization together with CTLA-4 blockade did not provide added therapeutic benefits to low-dose l-PAM-treated MOPC-315 tumor bearers. Taken together, these results indicate that CTLA-4 blockade improves the therapeutic outcome of low-dose l-PAM for MOPC-315 tumor bearers by inhibiting IL-10 secretion as a consequence of blocking CTLA-4 ligation.

Herein we demonstrate that B cell-activating factor of the TNF family (BAFF), a B cell survival factor, also regulates CD21/35 and CD23 expression. BAFF blockade in wild-type mice down-modulates CD21/35 and CD23 on B cells while survival remains intact, and BAFF exposure causes elevated CD21/35 and CD23 expression. Similar down-modulation is observed in bcl-2-transgenic mice treated with a BAFF inhibitor. This is the first evidence that BAFF has a function independent of B cell survival. Reports using CD21/35 and CD23 expression to assess splenic B cell subsets in BAFF-null mice concluded a lack of B cells beyond the immature stage. Since CD21/35 and CD23 are inadequate for delineating B cell subpopulations in BAFF-null mice, we used expression of BAFF-R and several B cell markers to identify more mature splenic B cells in these mice. These data broaden our understanding of BAFF function and correct the view that BAFF-null mice lack mature B cells.