The OX40 (CD134) molecule is induced primarily during T cell activation and, as we show in this study, is also expressed on CD25+CD4+ regulatory T (Treg) cells. A necessary role for OX40 in the development and homeostasis of Treg cells can be inferred from the reduced numbers of the cells present in the spleens of OX40-deficient mice, and their elevated numbers in the spleens of mice that overexpress the OX40 ligand (OX40L). The homeostatic proliferation of Treg cells following transfer into lymphopenic mice was also found to be potentiated by the OX40-OX40L interaction. Suppression of T cell responses by Treg cells was significantly impaired in the absence of OX40, indicating that, in addition to its homeostatic functions, OX40 contributes to efficient Treg-mediated suppression. However, despite this, we found that CD25−CD4+ T cells became insensitive to Treg-mediated suppression when they were exposed to OX40L-expressing cells, or when they were treated with an agonistic OX40-specific mAb. OX40 signaling could also abrogate the disease-preventing activity of Treg cells in an experimental model of inflammatory bowel disease. Thus, although the data reveal important roles for OX40 signaling in Treg cell development, homeostasis, and suppressive activity, they also show that OX40 signals can oppose Treg-mediated suppression when they are delivered directly to Ag-engaged naive T cells.

The transplantation of human cancer cells into immunodeficient NOD/SCID/IL‐2Rγcnull (NOG) mice often causes highly malignant cell populations like cancer stem cells to emerge. Here, by serial transplantation in NOG mice, we established two highly tumorigenic adult T‐cell leukemia‐derived cell lines, ST1‐N6 and TL‐Om1‐N8. When transplanted s.c., these cells formed tumors significantly earlier and from fewer initial cells than their parental lines ST1 and TL‐Om1. We found that protein kinase B (AKT) signaling was upregulated in ST1‐N6 and TL‐Om1‐N8 cells, and that this upregulation was due to the decreased expression of a negative regulator, INPP5D. Furthermore, the introduction of a constitutively active AKT mutant expression vector into ST1 cells augmented the tumorigenicity of the cells, whereas treatment with the AKT inhibitor MK‐2206 attenuated the progression of tumors induced by ST1‐N6 cells. Collectively, our results reveal that the AKT signaling pathway plays a critical role in the malignancy of adult T‐cell leukemia‐derived cells.

Carbonic anhydrase IX (CA9) is a membrane‐associated carbonic anhydrase that regulates cellular pH, is upregulated in various solid tumors, and is considered to be a therapeutic target. Here, we describe the essential role of CA9 in the tumorigenicity of cells derived from human adult T‐cell leukemia/lymphoma (ATL). We previously established the highly tumorigenic ST1‐N6 subline from the ATL‐derived ST1 cell line by serial xenotransplantation in NOG mice. In the present study, we first show that CA9 expression is strongly enhanced in ST1‐N6 cells. We then sorted ST1 cells by high or low CA9 expression and established ST1‐CA9high and ST1‐CA9low sublines. ST1‐CA9high cells, like ST1‐N6 cells, were more strongly tumorigenic than ST1‐CA9low or parental ST1 cells when injected into NOG mice. Knockdown of CA9 with shRNAs suppressed the ability of ST1‐CA9high cells to initiate tumors, and the tumorigenicity of ST1 cells was significantly enhanced by introducing wild‐type CA9 or a CA9 mutant with deletion of an intracytoplasmic domain. However, a CA9 with point mutations in the catalytic site did not increase the tumorigenicity of ST1 cells. Furthermore, we detected a small population of CA9+ CD25+ cells in lymph nodes of ATL patients. These findings suggest that CA9, and particularly its carbonic anhydrase activity, promotes the tumorigenicity of ATL‐derived cells and may be involved in malignant development of lymphoma‐type ATL.