LAUSR

Although advances in multiple myeloma (MM) therapy such as stem cell transplantation and newly approved biological agents have shown promising results, MM remains a terminal disease. Thus, new strategies to eliminate or control MM cells are needed. Elotuzumab, a humanized monoclonal antibody approved for treatment of MM, targets myeloma cells expressing CS1/SLAMF7. We heteroconjugated anti-CD3 (OKT3) and elotuzumab to develop a BiAb, CS1Bi, to target activated T cells (ATC) to CS1+ MM cells. To examine the potential therapeutic activity of BATs against MM, we tested the ability of ATC armed with CS1Bi to bind to MM cell lines, specifically kill MM cells lines expressing different levels of CS1, and produce Th1 cytokines upon engaging MM tumor targets. Initial studies showed that CS1-BATs bound to MM cells by flow cytometry and specifically killed CS1+ targets as measured in a 6 hour 51Cr-release assay. Cytokine secretion by CS1-BATs vs. unarmed ATC (n=2) of interferon-γ (IFN- γ: 512 pg/mL vs. 260pg/mL), tumor necrosis factor-α (TNF- α: 450pg/mL vs. 110pg/mL), granulocyte-macrophage colony-stimulating factor (GM-CSF: 217 pg/mL vs. 135 pg/mL), macrophage inflammatory protein-1α (MIP-1α: 8250 pg/mL vs. 400 pg/mL), macrophage inflammatory protein-1β (MIP-1β: 14750 pg/mL vs. 600 pg/mL), and RANTES (3550 pg/mL vs. 2600 pg/mL) were induced when CS1-BATs were co-cultured with CS1+ cell line, L-363 for 24h at E:T of 25:1. A more sensitive quantitative flow cytometry-based cytotoxicity assay was developed based on comparing of the number of eFluor 450-labeled target cells pre- and post-incubation with either CS1Bi-BATs or unarmed ATC prepared from healthy donors, or culture media alone. The number of eFluor450+/7AAD- viable cells/unit volume post-incubation were compared to the number of viable cells in untreated wells to calculate the percent cytotoxicity. This assay reproducibly demonstrated significant killing of five MM cell lines by CS1-BATS (armed with 50ng CS1Bi/106 ATC at an E:T ratio of 1:1) ranging up to 60%, depending on the target line, (n=8). Between 78- 95% of MM.1S cells were eliminated at an E:T ratio of 3:1 (n=2). These data demonstrate the clinical relevance of this anti-MM strategy for use in a Ph1/II clinical trial in patients with relapsed and/or refractory MM. Citation Format: Manley Huang, Archana Thakur, Lawrence G. Lum. Anti-CS1 x anti-CD3 bispecific antibody (BiAb)-armed anti-CD3 activated T cells (CS1-BATs) specifically kill CS1+ myeloma cells and release type-1 cytokines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2312.