Human Anti-CTLA4 Recombinant Antibody (clone 10D1)

Figure 1 shows melanoma-specific antibody responses in animals treated with a melanoma vaccine and the anti CTLA-4 antibody 10D1.

Antibody reactivity in pooled plasma samples from treated animals was measured by flow cytometry.

Figure 2 shows the pharmacokinetic profile of the anti CTLA-4 antibody 10D1 during chronic dosing of primates.

Antibody was administered on days 0, 28, 56.84 and 140 and plasma concentrations of 10D1 were analyzed by ELISA. The mean +/-SEM of six treated animals is shown.

Figure 3 shows the pharmacokinetic profile of anti-CTLA-4 antibody 10D1 in prostate cancer patients treated with a single dose of 10D1 at day 0

Plasma concentration (ug/ml) is shown. The mean +/-SEM for 14 treated patients is shown.

Figure 4 Ligand blocking by mAb 10D1. Inhibition of recombinant B7.1 (A) and B7.2 (B) binding to CTLA-4-expressing T cells.

B7-Ig fusion proteins were incubated with huCTLA4 transfectants in the presence of varying concentrations of 10D1 F(ab)2 or isotype control F(ab)2. The mouse anti-human CTLA-4 mAb BNI-3 was also included as a positive control. B7-Ig binding was detected using goat anti-human IgG (Fc-specific)-PE probe. The cell-associated fluorescence was determined by flow cytometry, and percent inhibition of binding was determined.

Davis, T., Lonberg, N., Korman, A., Blanset, S. L., Srinivasan, M., & Graziano, R. F. (2003). Activity and Safety of CTLA-4 Blockade. J Immunol, 171, 6251-6259.

Figure 5 CTLA-4 blockade with mAb 10D1 enhances Ab response to a melanoma vaccine in monkeys

Groups of six cynomolgus monkeys were given s.c. injections of 5106 SKmel-GM cells alone (arrows) or in combination with a bolus i.v. administration of mAb 10D1 (10 mg/kg). Analysis of Ab responses to SKmel-3 cells by flow cytometry (A) or ELISA (B) was determined from plasma samples taken 2 wk after each immunization. The values represent the mean SE from plasma samples, 6/group, tested at a 1/1000 dilution for flow cytometry or 1/200 for ELISA. Statistically significant differences by the Student t test are noted; p= 0.05,p=0.01,p=0.005. C, The individual animal responses by flow cytometry or ELISA (pretreatment values subtracted) at day 97.

Davis, T., Lonberg, N., Korman, A., Blanset, S. L., Srinivasan, M., & Graziano, R. F. (2003). Activity and Safety of CTLA-4 Blockade. J Immunol, 171, 6251-6259.

Figure 6 Ab responses to melanoma and nonmelanoma cell lines in animals treated with melanoma vaccine and mAb 10D1.

Pooled plasma from six cynomolgus monkeys given SKmel-3-GM-CSF vaccine and mAb 10D1 (day 41) was tested for reactivity to a variety of melanoma and nonmelanoma cell lines by flow cytometry (1/1000 dilution).p 0.05human melanoma cell binding vs human nonmelanoma cells.

Davis, T., Lonberg, N., Korman, A., Blanset, S. L., Srinivasan, M., & Graziano, R. F. (2003). Activity and Safety of CTLA-4 Blockade. J Immunol, 171, 6251-6259.

Figure 7 CTLA-4 blockade enhances ADCC activity in response to the melanoma vaccine.

Pooled plasma samples from vaccinated animals with or without 10D1 mAb were taken prior to immunization (Pre) or 2 wk after the second immunization (day 41). The samples were tested for ADCC activity on three melanoma cell lines at a 1/500 dilution.

Davis, T., Lonberg, N., Korman, A., Blanset, S. L., Srinivasan, M., & Graziano, R. F. (2003). Activity and Safety of CTLA-4 Blockade. J Immunol, 171, 6251-6259.