459. Evaluation of Miltenyi ExpAct and TransAct CD3/28 Beads for CAR-T Cell Manufacturing

Adoptive transfer of chimeric antigen receptor (CAR) engineered T cells is a promising emerging strategy to treat cancer patients. Large-scale manufacturing of cGMP-grade CAR T cells using patient T cells selected and activated by CTS™ Dynabeads® CD3/CD28 (Dynabeads) followed by transduction with retroviral vectors is being used in the context of many clinical trials by our laboratory and others. Although we have established a robust protocol using Dynabeads, it is important to explore alternative sources to pre-empt supply chain limitations of this critical reagent. To this end, we evaluated T cell activation with either Miltenyi TransAct CD3/28 (TransAct) beads or Miltenyi ExpAct Treg (ExpAct) beads. In small-scale experiments, PBMCs were directly activated with TransAct or ExpAct beads and compared with our standard T cell selection and activation using Dynabeads. Overall, the transduction efficiency and expansion of T cells were comparable upon activation with all three reagents. The TransAct bead-stimulated cells exhibited comparable effector memory (EM)/central memory (CM) phenotype to that of the Dynabeads stimulated cells. In line with the EM/CM phenotype, CAR T cells stimulated with either TransAct or Dynabeads and tranduced with CD19-targeted CAR demonstrated robust and comparable antitumor activity in a systemic NSG/CD19+ NALM6 tumor mouse model. We further tested the efficacy of TransAct beads using positively or negatively selected T cells in a large-scale cGMP grade CAR-T cell manufacturing setting. Both the transduction efficiency and expansion of selected CD3+ cells activated with TransAct beads and Dynabeads were comparable. CD19-targeted CAR T cells activated by either TransAct or Dynabead were subjected to an in vivo stress test by using decreasing amount of CAR-T cells to treat systemic CD19+NALM6 tumors in NSG mice. In this experimental setting, T cells stimulated with TransAct beads demonstrated equivalent if not better anti-tumor activity than T cells stimulated with Dynabeads. In conclusion, our pre-clinical results suggest that TransAct beads support efficient transduction and expansion of CAR T cells. TransAct activated T cells exhibit antitumor activity equivalent to Dynabeads activated T cells in our NSG/CD19+NAML6 stress test. Therefore, Miltenyi TransAct beads can be used as an alternative to Dynabeads to stimulate T cells in clinical trials aiming at evaluating CAR T cell safety and antitumor activity.