Abstract 1547: Defining the optimal source of allogeneic γ δ T cells for novel cellular therapy approaches in neuroblastoma
Authors: Jonus Hunter Connell , Burnham Rebecca E. , Spencer Trent H. , Goldsmith Kelly C. 
Published: July 2021

Adoptive cell therapy for aggressive pediatric solid tumors like neuroblastoma (NB) has focused on autologous products of αβ T cells, that so far, have been uniformly unsuccessful. γδ T cells offer a potentially superior, off-the-shelf product as they are MHC independent, directly cytotoxic to tumor cells, and lack the alloreactivity of αβ T cells. Expanded γδ T cells also express high CD16 (FcγRIII) necessary for antibody dependent cellular cytotoxicity. We have shown that γδ T cells expanded from NB patient apheresis products potently kill NB cell models. However, NB patient marrows are heavily pretreated and apheresis products are often depleted. Thus, we propose to expand γδ T cells from healthy donor peripheral blood mononuclear cells (PBMCs), yielding a robust product for our upcoming first-in-child phase I evaluation of ex vivo expanded γδ T cells in combination with dinutuximab (anti-GD2) and temozolomide for recurrent NB.

Our focus now is to identify biomarkers of the most potent γδ T cell products for use in this trial. PBMCs were isolated from whole blood of healthy adults and expanded using zoledronate and IL-2 via our GMP-compliant protocol. Across 25 cell expansions from 12 independent donors, the final γδ TCR+ percentage of the expanded product was highly variable (range 21.3%-98.4%). In addition, the γδ TCR+ percentage of PBMCs on day 0 did not correlate to the final percentage of γδ T cells in the expanded product. The other major cellular constituent present in expanded products were identified as Natural Killer (NK) lymphocytes (range 2.30%-77.60%). Interestingly, the NK content of PBMCs used to seed the expansion on day 0 inversely correlated with the final γδ T cell content of the expanded product (p = 0.0137). The cytotoxicity of products with varying γδ/NK content was evaluated against NB cell lines in the presence and absence of dinutuximab. When NB cells were incubated with immune cells alone, expanded products with greater NK content induced significantly higher levels of tumor cell death at low effector to target ratios (1:1). However, when the expanded γδ/NK product was combined with dinutuximab, all products demonstrated enhanced cytotoxic potential with no significant variation due to NK content. Functional activation (TNFα) and degranulation (CD107a) of isolated NK or γδ T cells co-incubated with NB cells supports that NK cells exert cytolytic effects independently, while γδ T cells require dinutuximab to mediate robust tumor cell death.

In conclusion, not all healthy donors expand cytolytic lymphocytes equally, nor are their products equally potent against NB in the absence of dinutuximab. Greater NK content predicts a more effective product given alone, yet antitumor potency when combined with dinutuximab is agnostic to γδ T cell or NK content. Studies are ongoing to investigate the influence of cellular subsets on tumor trafficking and in vivo potency.

Citation Format: Hunter Connell Jonus, Rebecca E. Burnham, Trent H. Spencer, Kelly C. Goldsmith. Defining the optimal source of allogeneic γ δ T cells for novel cellular therapy approaches in neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1547.