Anti-Human 4-1BB Recombinant Antibody (TAB-179) (CAT#: TAB-179)

Figure 1 Administration of anti-hPD-1 mAb (nivolumab) as a single agent or in combination with anti-hCD137 (urelumab) exacerbates xGVHD mediated by human PBMCs in Rag2−/−IL2Rγnull mice.

A, Rag2−/−IL2Rγnull mice were injected with 107 human PBMCs i.p. on day 0. Treatments with anti-hCD137 (urelumab), anti-hPD-1 (nivolumab), combination (Combo; urelumab+nivolumab), or saline control i.v. were injected on days 4, 7, and 10. Survival was monitored up to 100 days. B, the Kaplan–Meier plot depicts overall survival among the four groups of treatment urelumab (n = 22), nivolumab (n = 23), combination (Combo; n = 13), and control (n = 19). Experiments were independently repeated three times and results pooled. *, P < 0.5; ***, P < 0.001. C, microphotographs of H&E and hCD3 IHC stainings in representative tissue sections of spleen, liver, and lung in one representative mouse from each group of treatment. Bar, 400 μm.

Sanmamed, M. F., Rodriguez, I., Schalper, K. A., Onate, C., Azpilikueta, A., Rodriguez-Ruiz, M. E., ... & Alfaro, C. (2015). Nivolumab and urelumab enhance antitumor activity of human T lymphocytes engrafted in Rag2−/− IL2Rγnull immunodeficient mice. Cancer research.

Figure 2 Evidence for enhanced xenoreactive human lymphocyte activity upon treatment with anti-hCD137 (urelumab), anti-hPD-1 (nivolumab), and their combination.

Experiments performed as in Fig. 2A. A, human IFNγ plasma levels of mice were studied before the first mAb dose and after the first (day 6) and second (day 9) doses of antibodies. B and C, mice were sacrificed at week 3 after human PBMCs transfer and single-cell suspensions were prepared from the lung, liver, spleen, and peritoneal lavage to be studied by flow cytometry. B, absolute numbers of hCD3+ (right) and hCD8+ (left) per gram of tissue in the liver. C, perforin mean fluorescence intensity (MFI) of hCD3+hCD8+ (left) and hCD3+hCD4+ (right) in the lung lymphocyte infiltrates. Data were analyzed by the Mann–Whitney U test. *, P < 0.05; **, P < 0.01. Error bars represent SEM.

Sanmamed, M. F., Rodriguez, I., Schalper, K. A., Onate, C., Azpilikueta, A., Rodriguez-Ruiz, M. E., ... & Alfaro, C. (2015). Nivolumab and urelumab enhance antitumor activity of human T lymphocytes engrafted in Rag2−/− IL2Rγnull immunodeficient mice. Cancer research.

Figure 3 Immunostimulatory mAbs anti-hCD137 (urelumab) and anti-hPD-1 (nivolumab) alone or in combination show antitumor activity against a xenografted human colon cancer mediated by transferred allogeneic human PBMC.

A, Rag2−/−IL2Rγnull mice were injected with 7 × 106 human PBMCs i.p. and challenged with 3.5 × 106 HT-29 cells s.c. in the right flank on day 0 of the experiment. Thereafter, the mice were treated with anti-hCD137 (urelumab), anti-hPD-1 (nivolumab), combination (urelumab+nivolumab), or isotype control (hIgG4) i.v. on days 6, 14, and 20. Tumor volumes were measured twice per week until mice were sacrificed on day 22 of experiment. Tumors were removed after sacrifice and studied by flow cytometry and IHC. B, results depict mean ± SEM of the progression of subcutaneous tumor volumes (n = 5 mice per group). Arrows, time of treatment administration. Experiments were repeated four times, rendering similar results. C, human IFNγ plasma levels were studied previous to the first mAb dose, after the first and second antibody dose. D, tumor TILs from each treatment group were studied by flow cytometry and results of number of human CD8 T lymphocytes per mg of tumor (left), number of human regulator T cells per gram of tumor (center graph), and ratio of number of human CD8 T cells per number of human Treg (right) are depicted. *, P < 0.05; **, P < 0.01. Error bars represent SEM. T, tumor; p< hPBMCs.

Sanmamed, M. F., Rodriguez, I., Schalper, K. A., Onate, C., Azpilikueta, A., Rodriguez-Ruiz, M. E., ... & Alfaro, C. (2015). Nivolumab and urelumab enhance antitumor activity of human T lymphocytes engrafted in Rag2−/− IL2Rγnull immunodeficient mice. Cancer research.

Figure 4 hCD137, hPD-1, and PD-L1/B7-H1 are expressed in the xenografted tumors.

Representative multiplexed immunofluorescence microphotographs showing the protein expression of hCD137 (red fluorescence channel, left), hPD-1 (red channel, middle), and PD-L1 (red channel, right) in the tumor tissues (cytokeratin positive, green channel) from mice subjected to treatment with hIgG4 (control), anti-hCD137 (urelumab), anti-hPD-1 (nivolumab), or the combination (Combo). hCD137 and hPD-1 positivity was distributed predominantly toward the periphery of the tumor surrounding the cytokeratin-positive tumor areas. In contrast, PD-L1 was allocated predominantly toward the tumor center/core, in close association with the tumor nests. Nuclei were stained with DAPI (blue fruorescence channel). Bar, 100 μm.

Sanmamed, M. F., Rodriguez, I., Schalper, K. A., Onate, C., Azpilikueta, A., Rodriguez-Ruiz, M. E., ... & Alfaro, C. (2015). Nivolumab and urelumab enhance antitumor activity of human T lymphocytes engrafted in Rag2−/− IL2Rγnull immunodeficient mice. Cancer research.

Figure 1 Anti-Human 4-1BB Therapeutic Antibody (Urelumab) (TAB-179) in WB

Western blot analysis of TAB-179 was performed with Recombinant Human CD137 Protein (His Tag).
Lane 1: Reducing antigen (0.1μg)
Lane 2: Reducing antigen (0.3μg)
Lane 3: Reducing antigen (0.6μg)
The secondary antibody: HRP Goat Anti-Human IgG (H+L)

Figure 2 Anti-Human 4-1BB Therapeutic Antibody (Urelumab) (TAB-179) in ELISA

ELISA analysis of TAB-179 was performed by coating with Recombinant Human CD137 Protein (His Tag).
The secondary antibody: HRP Goat Anti-Human IgG (H+L)

Figure 2 Anti-Human 4-1BB Therapeutic Antibody (Urelumab) (TAB-179) in Dot Blot

Dot Blot analysis of TAB-179 was performed with Recombinant Human CD137 Protein (His Tag).
The secondary antibody: HRP Goat Anti-Human IgG (H+L)