PS1302 THE CD58-CD2 AXIS CONTROLS KILLING OF B CELLS BY CYTOTOXIC T CELLS

Abstract

Background: DuoBody-CD3xCD20 (GEN3013) is a bispecific antibody (bsAb) recognizing the T-cell antigen CD3 and the B-cell antigen CD20, that triggers potent T-cell-mediated lysis of CD20-expressing cells. DuoBody-CD3xCD20 is a full-length bispecific IgG1 generated by controlled Fab-arm exchange (Labrijn et al. PNAS 2013, Nat Prot 2014), with an effector function-silenced Fc region. Aims: Preclinical characterization of DuoBody-CD3xCD20. Methods: T-cell activation and cytotoxicity induced by DuoBody-CD3xCD20 in vitro, in comparison with a known CD3xCD20 bsAb, was measured by flow cytometry, 48 hours after incubation of healthy donor T cells with CD20-expressing cell lines (E:T ratio 2:1). Anti-tumor activity of DuoBody-CD3xCD20 in vivo was assessed in xenograft models. In a co-engraftment model, Raji tumor cells were co-inoculated with healthy donor PBMCs subcutaneously (SC) in NOD-SCID mice, followed by intravenous (IV) antibody treatment. In a MiXeno model, JEKO-1 tumor cells were inoculated SC in NCG mice, followed by intraperitoneal injection of human PBMCs. Treatment was initiated when tumors reached an average size of 100 mm3. Human immune system (HIS mice) were inoculated with Daudi lymphoma cells IV or with Raji lymphoma cells SC, followed by treatment from day 3. Activity of DuoBody-CD3xCD20 after SC or IV administration to cynomolgus monkeys was assessed as part of nonclinical safety studies. Results: DuoBody-CD3xCD20 induced potent activation and cytotoxic activity of CD4+ and CD8+ T cells in the presence of CD20-expressing cells in vitro. T-cell-mediated cytotoxicity was observed in a diverse panel of diffuse large B-cell and mantle cell lymphoma cell lines, with EC50 values ranging from 0.2 to 5.0 pM. Direct comparison in vitro demonstrated that DuoBody-CD3xCD20 was ∼100-fold more potent than another CD3xCD20 bsAb that is currently tested in clinical trials. DuoBody-CD3xCD20 potently inhibited outgrowth of CD20-expressing tumors (Raji) in vivo, even in the presence of high doses of Fc-inactive rituximab, demonstrating that the anti-tumor activity of DuoBody-CD3xCD20 was not inhibited by an excess of circulating CD20 antibodies that compete for target binding. In addition, DuoBody-CD3xCD20 induced regression of established mantle cell lymphoma (JEKO-1) tumors and delayed tumor growth and enhanced survival in the HIS xenograft models. In cynomolgus monkeys, profound and long-lasting B-cell depletion (≥ 70 days) from peripheral blood and lymphoid organs was induced by DuoBody-CD3xCD20 at dose levels > 0.1 mg/kg. B-cell depletion was reversible, with time to recovery correlating with the treatment dose. A transient decline in peripheral T cells was observed, as well as T cell activation. At the same dose levels, SC and IV administration of DuoBody-CD3xCD20 induced comparable B cell depletion. Importantly, peak plasma levels were reduced and delayed after SC administration, which was associated with reduced plasma cytokine levels. Summary/Conclusion: DuoBody-CD3xCD20 induced potent anti-tumor activity in vitro and in vivo, also in the presence of competing CD20 antibodies. DuoBody-CD3xCD20 was ∼100-fold more potent in vitro than another CD3xCD20 bsAb that is in clinical development. SC administration in cynomolgus monkeys induced profound B-cell depletion, which was associated with reduced plasma cytokine levels compared to IV administration. A First-in-Human trial to evaluate the safety and preliminary efficacy of DuoBody-CD3xCD20 by SC administration in patients with B-cell malignancies is currently enrolling (NCT03625037).