Fig. 1.

Cytolytic activity of ESK1-BiTE against ovarian cancer cells. A Expression of WT1 protein was measured by intracellular staining of the SKOV3 Muc16 low cells using mAb to WT1 protein. B Expression of the cell surface WT1 epitope RMF/HLA-A2 complex was measured by staining the cells with mouse ESK1 mAb or its control at 3ug/ml and analyzed by flow cytometry. C and D ESK1.BiTE-mediated T cell cytotoxicity against SKOV3/A2 (C) or HL-60 (D). PBMCs from a healthy donor were incubated with SKOV3 Muc16 low target cells at a E:T ratio of 30:1 in the presence or absence of BiTEs at the indicated concentrations for 5 h. The BiTE-mediated T cell cytotoxicity was measured by standard ⁵¹Cr-release assay. The HL-60 AML cell line (HLA-A2-) was used as a control. E Binding of ESK1 to fresh tumor cells in ascites from an ovarian cancer patient. Ascites cells were stained with mouse ESK1 versus anti-CD45 or its isotype control, to distinguish lymphocytes from tumor cells. Smaller lymphocytes and large tumor cells were gated and the CD45 versus ESK1 staining was shown in each gate (lower panels)