FIGURE 2.
Characterization of CD4+ T cell line A9895. A, CD4+ T cell line A9895 exhibits a Th1-like cytokine profile. Secretion of cytokines IFN-γ, GM-CSF, IL-2, and TNF-α, but not IL-5, is seen against target melanoma line (1102 Mel). There was no reactivity seen against the control target (1102 EBV-B). The line also produces significant amounts of RANTES against melanoma target (1102 Mel), but not against control target (*, 888 Mel). Background reactivity for each cytokine and chemokine (baseline tumor production at 24 h) were subtracted from individual total values, respectively. B, Reactivity of line A9895 is MHC class II restricted. T cells were coincubated with targets in the presence of anti-HLA-A, -B, -C, anti-DR, and anti-murine CD4 mAbs. As a control for mAb blocking, the h-gp100209–217-specific, HLA-A*0201-restricted, human CD8+ T cell clone (K4/H5) was cocultured with identical targets. Recognition of 697 Mel and 1102 Mel by line A9895 was blocked by mAbs anti-DR and anti-CD4, but unaffected by mAb anti-HLA-A, -B, -C. In the control group, mAb anti-HLA-A, -B, -C blocked recognition of 697 Mel and 1102 Mel (both express HLA-A*0201), but was unaffected by mAbs anti-DR and anti-CD4. C, Line A9895 specifically lyses DR4+ human melanoma (1102 Mel), but not control melanoma (888 Mel), in a 15-h 51Cr release assay. For A and B, T cells and targets were cocultured for 24 h and supernatants were measured for individual cytokine and chemokine secretion by ELISA. All experiments were repeated twice with similar results using different but comparable targets, thus demonstrating the generalizability of the experimental findings.