Limited importance of CD40/CD40L interaction in the B7-dependent generation of anti-MOPC-315 cytotoxic T lymphocyte activity by tumor bearer splenic cells stimulated in vitro in the presence of tumor necrosis factor

 We have previously illustrated the importance of B7-2 expression for the enhanced generation of cytotoxic T lymphocyte (CTL) activity by stimulation cultures of tumor bearer splenic cells to which tumor necrosis factor α (TNFα) has been added. Here we show that the B7-1 molecule is also important for CTL generation by such stimulation cultures, although to a much lesser extent than the B7-2 molecule. In addition, we show the importance of CD40/CD40L interaction for the expression of the B7-2 molecule, but not the B7-1 molecule, by tumor bearer splenic cells stimulated in vitro in the presence of TNF. The CD40/CD40L interaction is also shown to be important for the generation of CTL activity by tumor bearer splenic cells stimulated in vitro in the presence of exogenous TNF. However, the CD40/CD40L interaction is less important for the generation of enhanced CTL activity than for the expression of an elevated level of B7-2. Specifically, blockade of CD40/CD40L interaction, which reduced the level of B7-2 expressed by tumor bearer splenic cells stimulated in vitro in the presence of TNF to the level of B7-2 expressed by tumor bearer splenic cells stimulated in vitro in the absence of exogenous TNF, failed to reduce the level of CTL generated to the level generated by tumor bearer splenic cells stimulated in the absence of exogenous TNF. Finally, blockade of CD40/CD40L interaction was inferior to blockade of B7-2/CD28 interaction in inhibiting the generation of CTL activity by tumor bearer splenic cells stimulated in the presence of exogenous TNF. Thus, although CD40/CD40L interaction is important for the generation of enhanced CTL activity by stimulation cultures of tumor bearer splenic cells to which TNF has been added, TNF also mediates its potentiating effect for CTL generation by such stimulation cultures via other mechanisms that are independent of CD40/CD40L interaction but dependent on B7-2 expression. Received: 31 December 1997 / Accepted: 27 March 1998     

Therapeutic effectiveness of the immunity elicited by P815 tumor cells engineered to express the B7-2 costimulatory molecule

 It is well accepted that inoculation of B7-1-transfected tumor cells into normal mice leads to tumor rejection and subsequent resistance to challenge. However, the effectiveness of B7-2-transfected tumor cells in eliciting protective antitumor immunity is less clear. Here we show that B7-2-transfected P815 tumor cells (B7-2⁺) are as effective as B7-1-transfected P815 tumor cells (B7-1⁺) in eliciting protective immunity in normal DBA/2 mice. In addition, B7-2⁺ cells were found to be at least as effective as B7-1⁺ cells in retarding tumor progression when admixed with parental P815 tumor cells prior to inoculation into normal mice. Moreover, the B7-2⁺ cells and the B7-1⁺ cells were equivalent in their ability to retard tumor growth when administered peritumorally into mice bearing established (approx. 3 mm in diameter) parental P815 tumors. Finally, P815 tumor cells infected with a recombinant replication-defective adenovirus encoding the murine B7-2 gene were effective in retarding the growth of established parental P815 tumors. Thus, B7-1 and B7-2 are comparable in terms of their ability to stimulate the generation of tumor-eradicating immunity in normal mice as well as in mice bearing established parental tumors. Moreover, adenovirus vectors can be used to generate B7-2-expressing tumor cells effective in the immunotherapy of established parental tumors. Received: 10 January 1996 / Accepted: 23 February 1996     

CD80 (B7-1) expression on human acute myeloid leukaemic cells cultured with GM-CSF, IL-3 and IL-6

Acute myeloid leukaemia (AML) blasts rarely express the B7 family of co-stimulatory molecules and do not elicit a clinically significant autologous T-lymphocyte anti-tumour response. The aim of this study was the in vitro modification of AML blasts to an antigen-presenting cell phenotype characterised by upregulated expression of the co-stimulatory molecule CD80 (B7-1). Circulating AML cells were induced to undergo partial differentiation in culture with the cytokines IL-3, IL-6 and GM-CSF; they exhibited variable upregulation of CD80 and continued to express MHC class I and II. These cells remained viable to day 20, in contrast with normal peripheral blood mononuclear cells (PBMNC), which did not survive under the culture conditions. In contrast to unmanipulated blasts, cultured leukaemic cells expressed B7-1. Where initial cytogenetic abnormalities were present, they were also seen in flow-sorted CD80-expressing cells after culture in cytokines, indicating their malignant origin. The immunogenic potential of these cultured cells was highlighted by allogeneic and autologous mixed lymphocyte reactions, in which both differentiated, but not unmanipulated, blasts produced expansion of T-lymphocyte numbers. Autologous cytotoxic T-lymphocyte (CTL) assays indicated specific killing of B7-1+ leukaemic cells, which was greatly enhanced after priming of the T-lymphocytes by B7-1+ blasts prior to the CTL assay, then enabling the CTL to lyse both unmanipulated and differentiated leukaemic cells.