Figure 2
B-CLL-specific immunity associated with elimination of B-CLL SP cells and subsequent decrease in leukemic cell counts in one patient treated with hCD40L/IL-2. (A) Vaccination with hCD40L/IL-2 gene-modified tumor cells resulted in the stable elimination of SP cells in the peripheral blood of one patient (P1300), in whom SP cells did not return when immunization was complete. Analysis of leukemic cell number in the peripheral blood of patient P1300 showed a delayed decline in circulating B-CLL cells, beginning six months after loss of circulating SP cells. Absolute cell number was calculated from the white blood cell (WBC) count multiplied by the percentage of B-CLL (CD5+CD19+), normal B cell (CD5-CD19+) and T cells (CD3+), and SP (CD5+CD19+SP+) as determined by flow cytometry. Arrows indicate when patient received vaccination (6 vaccine injections total at 0, 1, 2, 6, 8 and 10 weeks). (B) P1300 received 6 subcutaneous injections of hCD40L/IL-2 gene-modified autologous tumor cells at 0, 1, 2, 6, 8 and 10 weeks (indicated by arrows). B-CLL-specific immunity was measured by selecting CD4 and CD8 T cells from PBMC by magnetic separation and measuring IFN-γ and IL-5 by ELIspot following a 36 hour co-culture with autologous tumor cells (SFC; spot-forming cells per 1 × 105 T cells). ELIspot analysis was performed on weeks 0 (pre-vaccine), 3, 12 and 16 (two months after last vaccination).