Predictors of Efficacy for Blinatumomab in BCP-ALL Patients: Non-Responders Show Impaired CD19-BiTE®-Mediated Cytotoxicity in Vitro

Bispecific T-cell engaging (BiTE®) antibody constructs recruit T cells to target antigens independent of their T-cell receptor specificity. Blinatumomab, a CD19xCD3 BiTE® antibody construct, is approved for the treatment of relapsed/refractory (r/r) B-cell precursor acute lymphoblastic leukemia (BCP-ALL), including patients with persistence or reoccurrence of measurable residual disease (MRD). Despite superior anti-leukemic efficacy compared to cytostatic agents, a majority of patients do not respond to treatment. Biomarkers for the identification of non-responders prior to or early during treatment are unknown. However, the definition of robust biomarkers for efficacy is of high importance for clinical decision-making and might also indicate ways to improve response rates. We therefore established a comprehensive immune-monitoring program for blinatumomab patients. We sequentially analysed peripheral blood of patients with r/r or MRD-positive disease receiving blinatumomab prior to the start of infusion and once weekly over the course of the first 28-day cycle. We determined CD3+ T cell counts and their subset distribution (CD4, CD8, naïve [TN], central memory [TCM], effector memory [TEM], and effector memory CD45RA-positive [TEMRA] T cells) by multiparameter flow cytometry (MPFC). Additionally, patient-derived T cells were cocultered with BCP-ALL cell lines (SEM and REH) at an effector: target ratio of 1:3 in presence of blinatumomab or a control BiTE® (0.5 ng/ml, respectively) for 3 days to assess their cytotoxic capacity. Blinatumomab-mediated cytotoxicity was determined by MPFC. T-cell proliferation was evaluated by MPFC (Far Red Cell tracer) after incubation with CD3/CD28 beads for 6 days. A total of 16 patients were enrolled. Four of these patients suffered from MRD disease, whereas the remaining 12 had overt relapse of BCP-ALL. Response rates for patients with morphological relapse were 50% (4 CRMRD- and 2 CRMRD+ after the first cycle) and 75% for patients with MRD disease (3 MRD conversions, 1 MRD persistence). Absolute lymphocyte counts were not significantly different between responders and non-responders before treatment initiation (0.9 G/l and 0.7 G/l, respectively). Whereas the percentage of CD3+ T cells (of all lymphocytes) did not significantly differ between responders and non-responders on day 0, non-responders had a significantly reduced CD3+ percentage on day 7 (81.2% vs 92.9%, p=0.03). Until day 28 of the first cycle, CD3+ percentages