Hydroxyurea Is Most Suitable for Cytoreduction of AML Prior to CD33/CD3 Bispecific BiTE® Antibody (AMG 330) Therapy: Uncompromised T-Cell Proliferation Ex-Vivo and CD33 Upregulation on AML Cells

• Published in: Blood 2014-12, Vol.124 (21), p.986-986
• Main Authors: Krupka, Christina, Brauneck, Franziska, Lichtenegger, Felix S, Kufer, Peter, Kischel, Roman, Zugmaier, Gerhard, Köhnke, Thomas, Altmann, Torben, Schneider, Stephanie, Fiegl, Michael, Spiekermann, Karsten, Sinclair, Angus M, Newhall, Kathryn J, Frankel, Stanley R, Baeuerle, Patrick, Hiddemann, Wolfgang, Riethmüller, Gert, Subklewe, Marion
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V124.21.986.986

Bispecific T-cell engager (BiTE®) antibodies represent a promising tool for anti-leukemic immunotherapy. The CD19/CD3-bispecific antibody blinatumomab was shown to be active in refractory and relapse patients with B-precursor acute lymphoblastic leukemia (Topp et al, ASCO 2014). Transient, blinatumomab-mediated cytokine release syndrome has been linked to target cell numbers as this phenomenon is predominantly observed within the first treatment cycle. In our previous work, we demonstrated that the bispecific CD33/CD3 BiTE® antibody AMG 330 is able to induce activation and proliferation of residual autologous T-cells and effectively mediates lysis of primary acute myeloid leukemia (AML) cells (Krupka et al, Blood 2014; 123(3):356-65). We hypothesize that in AML patients with high initial leukocyte counts (WBC > 30.000/μl) a cytoreductive phase prior to AMG 330 therapy might be beneficial to reduce the incidence and severity of cytokine mediated toxicity. Ideally, the cytoreductive drug does not impair T-cell function or reduce target antigen expression level. In the current study, we evaluated the effect of cytarabine (20 µM), decitabine (5 µM), azacitidine (1 µM and 5 µM) and hydroxyurea (10 µM and 100 µM) on T-cell proliferation and function in close analogy to potential treatment algorithms for AML. Healthy donor (HD) T-cells were pre-incubated with the cytoreductive drugs for 72 hours. T-cells were CFSE-labeled and co-cultured with either HL60 or MV4-11 cells (effector cell:target (E:T) ratio 1:1) in the presence or absence of AMG 330 (5 ng/ml). After 3 days of co-culture, lysis of HL60 cells and T-cell proliferation was assessed by flow cytometry. Pretreatment of T-cells with cytarabine completely abrogated T-cell function (lysis of HL60 cells: untreated (UT): 96.9% vs 20 µM: 4.2%) and significantly impaired T-cell proliferation (UT: 31.2% vs 20 µM: 4.6%). These findings correlated to data using primary AML samples collected 3 and 6 days after discontinuation of cytarabine treatment. After a 3-day chemotherapy-free interval, we observed no relevant T-cell proliferation and lysis of AML cells upon the addition of AMG 330 to the ex-vivo long-term culture system (lysis of AML cells on day 12: 30%; fold change T-cell expansion 0.9). After a 6-day treatment-free interval, high T-cell proliferation and cytotoxicity against primary AML cells were observed (lysis of AML cells on day 12: 61%; fold change T-cell expansion: 3.1). In contrast to cytarabine, decit