Directing the migration of serum-free, ex vivo-expanded V9V2 T cells

V gamma 9V delta 2 T cells represent a promising cancer therapy platform because the implementation of allogenic, off-the-shelf product candidates is possible. However, intravenous administration of human V gamma 9V delta 2 T cells manufactured under good manufacturing practice (GMP)-compliant, serum-free conditions are not tested easily in most mouse models, mainly because they lack the ability to migrate from the blood to tissues or tumors. We demonstrate that these T cells do not migrate from the circulation to the mouse bone marrow (BM), the site of many malignancies. Thus, there is a need to better characterize human gamma delta T-cell migration in vivo and develop strategies to direct these cells to in vivo sites of therapeutic interest. To better understand the migration of these cells and possibly influence their migration, NSG mice were conditioned with agents to clear BM cellular compartments, i.e., busulfan or total body irradiation (TBI), or promote T-cell migration to inflamed BM, i.e., incomplete Freund's adjuvant (IFA), prior to administering gamma delta T cells. Conditioning with TBI, unlike busulfan or IFA, increases the percentage and number of gamma delta T cells accumulating in the mouse BM, and cells in the peripheral blood (PB) and BM display identical surface protein profiles. To better understand the mechanism by which cells migrate to the BM, mice were conditioned with TBI and administered gamma delta T cells or tracker-stained red blood cells. The mechanism by which gamma delta T cells enter the BM after radiation is passive migration from the circulation, not homing. We tested if these ex vivo-expanded cells can migrate based on chemokine expression patterns and showed that it is possible to initiate homing by utilizing highly expressed chemokine receptors on the expanded gamma delta T cells. gamma delta T cells highly express CCR2, which provides chemokine attraction to C-C motif chemokine ligand 2 (CCL2)-expressing cells. IFN gamma-primed mesenchymal stromal cells (MSCs) (gamma MSCs) express CCL2, and we developed in vitro and in vivo models to test gamma delta T-cell homing to CCL2-expressing cells. Using an established neuroblastoma NSG mouse model, we show that intratumorally-injected gamma MSCs increase the homing of gamma delta T cells to this tumor. These studies provide insight into the migration of serum-free, ex vivo-expanded V gamma 9V delta 2 T cells in NSG mice, which is critical to understanding the fundamental properties of these cells.