Functionalized microcarriers improve T cell manufacturing by facilitating migratory memory T cell production and increasing CD4/CD8 ratio

Abstract

Adoptive cell therapies (ACT) using chimeric antigen receptor (CAR) T cells have shown promise in treating cancer, but manufacturing large numbers of high quality cells remains challenging. Critically, current T cell expansion technologies only partially recapitulate the in vivo microenvironment found in the human lymph nodes. In these organs, T cells expand at high cell density with autocrine/paracrine signaling, as well as signals from the extracellular matrix (ECM). Here we describe a T cell expansion system using degradable gelatin microcarriers functionalized with anti-CD3 and anti-CD28 monoclonal antibodies (mAbs), which address several of these shortcomings. We show that using this system, we can achieve approximately 2-fold greater expansion compared to functionalized magnetic beads, the current industry standard. Furthermore, carriers generated higher numbers of CCR7+CD62L+ migratory, central memory T cells and CD4+ T cells across multiple donors. Both these phenotypes have emerged as important for establishing durable and effective responses in patients receiving T cell immunotherapies. We further demonstrate that carriers can achieve greater memory cell yield compared to beads across a range of IL2 concentrations from 20 U/mL to 100 U/mL. These differences were greater at lower IL2 concentrations, indicating that the carriers are more efficient. We optimized this system using a design of experiments (DOE) approach and found that the carrier concentration affects the memory cell yield in a quadratic manner, where high or low concentrations are detrimental to memory formation. Finally, we show that carriers do not hinder CAR transduction and can maintain the CD4 and memory phenotype advantages in CAR-transduced T cells.