Afuco™ Anti-Human IL2RB ADCC Recombinant Antibody (MiK-Beta-1), ADCC Enhanced (CAT#: AFC-408CL)

Figure 1 Hu-Mikβ1 inhibits IL-2– and IL-15–induced proliferation of 32Dβ cells that express human IL-2/IL-15Rβ and the murine common γ-c.

The dose-response effects of addition of Hu-Mikβ1 Ab were evaluated on IL-2 (1.0 ng/mL)– and IL-15 (20 ng/mL)–induced proliferation of 32Dβ cells. In these cells, which express only human IL-2/IL-15Rβ and the common γ-c, but not IL-2Rα or IL-15Rα, IL-2– and IL-15–induced proliferations were inhibited by the addition of Hu-Mikβ1.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.

Figure 2 Hu-Mikβ1 inhibits IL-15–induced proliferation of wild-type Kit 225 cells but not Kit 225 7.10 cells that had been transfected with IL-15Rα.

(A) Hu-Mikβ1 inhibited IL-15– but not IL-2–mediated proliferation of wild-type Kit 225 cells that express IL-2Rα, IL-2/IL-15Rβ, and γc but not IL-15Rα. (B) Anti-Tac (anti–IL-2Rα) inhibited IL-2– but not IL-15–mediated proliferation of wild-type Kit 225 cells. (C) In contrast neither anti-Tac (♦) nor Hu-Mikβ1 (■) inhibited the IL-15 (1 ng/mL)–induced proliferation of Kit 225 7.10 cells that had been transfected with IL-15Rα and that expressed IL-15Rα, IL-2/IL-15Rβ, and γc chains in cis. Anti–IL-15 (○) did inhibit IL-15–mediated proliferation in these cells.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.

Figure 3 A 0.5 mg/kg dose of Hu-Mikβ1 does not maintain saturation of the IL-2/IL-15Rβ, whereas 1.5 mg/kg is sufficient to maintain saturation for 3 weeks.

(A) Three patients with T-LGL leukemia were evaluated for saturation of their IL-2/IL-15R between 6 and 72 hours after administration of 0.5 mg/kg of Hu-Mikβ1 and were shown to have saturated receptors. However this receptor saturation was not maintained in 2 patients at 7, 14, and 21 days. To evaluate saturation, FACS analysis was performed at these time points with fluorochrome-labeled Hu-Mikβ1 and fluorochrome-labeled Mikβ3, which define non-cross-competing epitopes on IL-2/IL-15Rβ, as described in "Methods." Different symbols represent the results for different patients. (B) Three patients with T-LGL leukemia were evaluated for saturation of their receptors after a 1.5 mg/kg IV infusion of Hu-Mikβ1. This dose of Hu-Mikβ1 was sufficient to provide prolonged saturation of the IL-2/IL-15R. In particular, saturation of CD122 was achieved at the 6- to 72-hour time point and was maintained at 100% saturation at the 7- and 14-day time points and at 80% saturation for the cells at the 21-day time point. Different symbols reflect the results for different patients.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.

Figure 4 FACS analyses of Hu-Mikβ1 saturation was performed on ex vivo CD8+ T cells after administration of 1.5 mg/kg of Hu-Mikβ1.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.

Figure 5 Pharmacokinetic analysis of Hu-Mikβ1 was performed after administration of 0.5, 1.0, and 1.5 mg/kg of Hu-Mikβ1 Ab to patients with T-LGL leukemia.

After administration of 0.5 (○), 1.0 (■), and 1.5 (▾) mg/kg of Hu-Mikβ1, the serum concentration of the Ab was quantitated at subsequent time points in patients with T-LGL leukemia. Hu-Mikβ1 was not demonstrable in the serum after the 2-week time point after 0.5 mg/kg administration, but was still present in the circulation at day 22 at concentrations of 160, 400, and 2530 pg/mL in the 3 patients receiving 1.5 mg/kg of Hu-Mikβ1.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.

Figure 6 PBMCs from patients with T-LGL leukemia do not manifest ex vivo spontaneous proliferation in a 6-day ex vivo culture.

PBMCs from patients with T-LGL leukemia were placed in a 6-day culture in1640 media containing 10% FCS and proliferation was assessed by the evaluation of the uptake of thymidine added during the last 4 hours of culture. There was no meaningful thymidine uptake, indicating that there was essentially no ex vivo proliferation of PBMCs from the 6 patients studied with monoclonal T-LGL leukemia. When 20 000 pg/mL of IL-15 was added to the T-LGL of 5 patients, there was an increase in the geometric mean proliferation from 472-5 565 cpm per 1 millicurie of 3H thymidine added. This proliferation was inhibited to baseline levels of 728 cpm on addition of 10 μg/mL of Hu-Mikβ1 at the onset of the cultures.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.

Figure 7 The expression of IL-2 and IL-15 mRNA was evaluated in the PBMCs from 6 patients with LGL.

There was either no or only minimal IL-2 and IL-15 mRNA expression observed in the patients examined. This contrasts with the high IL-15 expression in the adult T-cell leukemia/lymphoma cell line HuT-102 or in normal PBMCs after ionomycin/phorbol 12-myristate 13-acetate addition.

Waldmann, T. A., Conlon, K. C., Stewart, D. M., Worthy, T. A., Janik, J. E., Fleisher, T. A., ... & Decker, J. R. (2013). Phase 1 trial of IL-15 trans presentation blockade using humanized Mik-Beta-1 mAb in patients with T-cell large granular lymphocytic leukemia. Blood, 121(3), 476-484.