1758 DJ Flavell et al Figure 2 Lysis of target HSB-2 cells by ADCC elicited by SCID ( ) or NOD/SCID mouse ( ) effector splenocytes at a 100:1 E:T ratio in the presence of increasing concentrations of the anti-CD7 antibody HB2. Lysis of target HSB-2 cells in the presence of the irrelevant anti-CD19 isotype matched control antibody BU-12 with CB.17 SCID ( ) or NOD/SCID ( ) splenocytes is also shown. Bars, SD their ability to lyse target YAC-1 or HSB-2 cells in a chromium release assay at target to effector (T:E) ratios ranging from 1:200 to 1:20. Figure 1A shows that at T:E ratios of 1:200 and 1:100 SCID and NOD/SCID splenocytes performed comparably, elic- iting a similar degree of lysis of YAC-1 cells. However, at T:E ratios of 1:50 and 1:20 NOD/SCID splenocytes declined in their ability to lyse YAC-1 cells while SCID splenocytes maintained their capacity at a level similar to that seen at the higher T:E ratios. Neither SCID or NOD/SCID splenocytes were capable of lysing HSB-2 cells at any of the T:E ratios studied (Figure 1B). Antibody-dependent cellular cytotoxicity (ADCC) Both SCID and NOD/SCID splenocytes were tested for their ability to lyse target HSB-2 cells in an antibody-dependent cellular cytotoxicity (ADCC) assay at a T:E ratio of 1:100 in the presence of increasing concentrations of the anti-CD7 antibody HB2. The results of this study are shown in Figure 2. The greatest degree of target cell lysis was achieved with SCID mouse splenocytes at the two highest antibody concentrations of 660 and 66 pM being 55% and 52%, respectively compared with 31% and 37% lysis obtained with NOD/SCID splenocytes at these same antibody concentra- tions. In the same assay the irrelevant isotype matched control antibody BU-12 (anti-CD19) used at the two highest concentra- tions did not cause lysis of HSB-2 cells when either SCID or NOD/SCID splenocytes were employed. Therapeutic effects of murine anti-CD7 antibody treatment in SCID-HSB-2 and NOD/SCID-HSB-2 mice SCID mice injected i.v with two million HSB-2 cells on day 1 and treated with three 10 µ g i.v. doses of HB2 antibody on days 7, 9 and 11 showed a significant (P = 0.0135) prolongation in the time taken to develop symptoms compared with PBS sham-treated SCID mice, with 37% of these animals alive and disease-free at elective termina- tion of the experiment at 135 days (Figure 3A). In contrast HB2 antibody showed no significant therapeutic effect (P = 0.243) in identically treated NOD/SCID mice, with antibody-treated animals developing symptoms at virtually the same rate as PBS sham-treated British Journal of Cancer (2000) 83(12), 1755–1761 Figure 3 Kaplan-Meyer plots for groups of SCID (closed symbols) or NOD/SCID mice (open symbols) injected i.v. on day 1 with two million HSB-2 cells and treated with (A) three 10 µ g i.v. injections of HB2 antibody given on days 7, 9 and 11 ( , ) or sham treated at the same times with 200 µ l of PBS ( , ) or (B) treated with a single 10 µ g i.v. injection of HB2-SAPORIN on day 7 ( , ) or sham treated at the same time with 200 µ l PBS ( , ) NOD/SCID control animals and with all animals in the antibody- treated group dead by 102 days (Figure 3A). In this instance NOD/SCID PBS sham-treated controls succumbed to disease at a slightly faster rate overall than the equivalent SCID control animals, though the difference between them was not significant (Figure 3A). Therapeutic effects of HB2-SAPORIN and HB2-F(ab)2- SAPORIN in SCID-HSB-2 and NOD/SCID- HSB-2 mice To determine the therapeutic effect of the anti-CD7 HB2-SAPORIN IT in SCID-HSB-2 and NOD/SCID-HSB-2 mice we injected a single 10 µ g i.v. dose of HB2-SAPORIN IT into mice 7 days after i.v. challenge with two million HSB-2 cells. The IT had a highly significant therapeutic effect (P = 0.0002) in SCID-HSB-2 mice compared with PBS sham-treated SCID control animals, with 80% of the IT treated animals alive and disease-free for the 135 day dura- tion of the experiment (Figure 3B). In identically treated NOD/SCID mice the IT was therapeutically ineffective in the majority of animals, with 70% of these animals developing signs and symptoms of leukaemia growth at the same rate as the PBS sham-treated control animals (Figure 3B) but with an eventual 30% of animals disease-free at elective termination of the experiment. However, despite this 30% survival rate in the IT-treated NOD/SCID group © 2000 Cancer Research Campaign