Afuco™ Anti-Human CD70 ADCC Recombinant Antibody, ADCC Enhanced (CAT#: AFC-130CL)

Figure 1 Binding and blocking activity of ARGX-110 on U266 (multiple myeloma), Granta519 and Mino-1 (Mantle cell lymphoma) and Raji (Burkitt lymphoma) cell lines.

(A) binding of varying concentrations of ARGX-110 to CD70-positive cell lines and the CD70 negative cell line SUP-T1 as determined by flow cytometry.Data presented as mean fluorescent intensity (MFI) of duplicate measurements and SD (B)blocking of CD27 signaling by ARGX-110.CD70-positive cell lines, HEK293 cells with or without transfection with CD70 and crosslinked recombinant CD70 were co-cultured with HT1080-CD27-positive cells in the presence of varying concentrations of ARGX-110.IL-8 was measured by ELISA as a marker for CD27 signaling.Data presented as mean of triplicate measurements and SD.

Silence,K.,Dreier, T.,Moshir, M.,Ulrichts, P.,Gabriels, S. M., Saunders, M.,& Thibault, A.(2014, March). ARGX-110,a highly potent antibody targeting CD70,eliminates tumors via both enhanced ADCC and immune checkpoint blockade.In MAbs(Vol. 6, No. 2, pp.523-532).Taylor & Francis.

Figure 2 Effector functions of ARGX-110.

(A) ADCC activity measured with Cr51 release assay on 786-O cells (RCC). ARGX-110 is associated with enhanced ADCC activity compared with the fucosylated version of the antibody,41D12.(B) De-fucosylation does not impair CDC on U266 cells (multiple myeloma) nor (C)ADCP activity of ARGX-110(on 786-O cells).All data are presented as mean of triplicate measurements and SD.

Silence,K.,Dreier, T.,Moshir, M.,Ulrichts, P.,Gabriels, S. M., Saunders, M.,& Thibault, A.(2014, March). ARGX-110,a highly potent antibody targeting CD70,eliminates tumors via both enhanced ADCC and immune checkpoint blockade.In MAbs(Vol. 6, No. 2, pp.523-532).Taylor & Francis.

Figure 3 Depletion studies on cell lines representing different histologies.

(A-B) Cell lines were admixed with freshly isolated PBMCs from healthy donors and ARGX-110 or 41D12 was added at 5 µg/ml.The samples were incubated for 2 d and analyzed by FACS for the lysis of the cell lines by looking at specific cell line markers.All data are presented as mean of duplicate measurements.(C) % lysis is plotted against CD70 signal in FACS using anti-CD70-PE demonstrating that depletion is independent of CD70 copy number.

Silence,K.,Dreier, T.,Moshir, M.,Ulrichts, P.,Gabriels, S. M., Saunders, M.,& Thibault, A.(2014, March). ARGX-110,a highly potent antibody targeting CD70,eliminates tumors via both enhanced ADCC and immune checkpoint blockade.In MAbs(Vol. 6, No. 2, pp.523-532).Taylor & Francis.

Figure 4 Blocking of Treg activity by ARGX-110 demonstrated in an in vitro model.

(A) PBMCs from a healthy donor were admixed with CD70-positive tumor cell lines (SUDHL6, Raji and U266) in the presence of soluble anti-CD3 mAb and ARGX-110.ARGX-110 lysed the cell lines during the experiment.(B) ARGX-110 blocks Treg proliferation. More Tregs are formed in the wells where the CD70-positive cell lines were added than in the wells without addition of cell lines and this can be blocked with ARGX-110. Results are shown as mean and SD of duplicates.(C) Proliferation of the Tregs is dependent on CD27 signaling and can be blocked with ARGX-110.After two days of incubation, ARGX-110 reduces sCD27 (as determined by ELISA) levels further witnessing of a role for CD70 in Treg proliferation.

Silence,K.,Dreier, T.,Moshir, M.,Ulrichts, P.,Gabriels, S. M., Saunders, M.,& Thibault, A.(2014, March). ARGX-110,a highly potent antibody targeting CD70,eliminates tumors via both enhanced ADCC and immune checkpoint blockade.In MAbs(Vol. 6, No. 2, pp.523-532).Taylor & Francis.

Figure 5 In vivo efficacy study using Raji xenograft model.

(A) Kaplan-Meier plot showing the antitumor effect of 41D12 in a disseminated Burkitt lymphoma xenograft model. SCID mice were injected intravenously with 106 Raji B lymphoma cells,then treated with 41D12 or isotype control at the doses indicated. Mice were treated two times per week thereafter and received a total of five doses (n = 9 mice per group).(B) A plasma sample was obtained for each mouse at the time of sacrifice.sCD27 levels were determined by ELISA. Tumor bearing mice (n = 28) had higher levels of sCD27 than non-tumor bearing mice (n = 42).Statistics was done comparing groups treated with 41D12 vs.the isotype control.

Silence,K.,Dreier, T.,Moshir, M.,Ulrichts, P.,Gabriels, S. M., Saunders, M.,& Thibault, A.(2014, March). ARGX-110,a highly potent antibody targeting CD70,eliminates tumors via both enhanced ADCC and immune checkpoint blockade.In MAbs(Vol. 6, No. 2, pp.523-532).Taylor & Francis.