Introduction: Lymphodepletion is a key factor for responses in both autologous and allogeneic cell therapies. We are currently evaluating in a Phase I trial, the combination of Temozolomide (TMZ) chemotherapy with methylguanine-DNA methyltransferase (MGMT) modified γδ T cells (Drug Resistant Immunotherapy, or DRI) during maintenance TMZ in patients receiving the Stupp protocol. In this report, we examine both DRI graft and circulating lymphocyte phenotype and function during the treatment phase.

Methods: Patients meeting enrollment criteria undergo tumor resection, placement of a Rickham catheter in the tumor cavity, and an apheresis collection immediately prior to standard chemoradiotherapy (CRT). DRI follows TMZ on Day 1 of maintenance and consists of 150mg/m2 IV TMZ and concurrent intracranial injection of 1.0 x 107 DRI γδT cells. Peripheral blood (PB) was collected at apheresis and immediately prior to the start of each 28-day maintenance cycle. Flow cytometric (FACS) analysis of PB and DRI grafts using antibodies to CD3, CD4, CD8, CD16/56, CD19, TCR-γδ, Vδ1, Vδ2, CD197, CD27, CD28, CD57, CD45RA, Th1/Th2/Th17 PB serum cytokines, and DRI cytotoxic graft function were obtained. When applicable, FFPE sections of recurrent resected GBM tissues were examined for immune infiltration.

Results: DRI grafts contained 74-93% activated γδ T cells with 0.3-0.5 MGMT copies/cell. Cytotoxicity against K562 ranged from 30% to 75% at 40:1 (E:T). Peripheral lymphodepletion was evident throughout TMZ treatment in the Stupp protocol. T cell counts fell precipitously after CRT and cycle 1 (1647+774 vs 609+214 cells/mL) and remained low throughout maintenance phase (range 214-1450 T cells/mL). NK counts remained normal and showed uneven recovery through the first three cycles but failed to recover for the remainder of maintenance phase. γδ T cells also recovered modestly through the first two cycles but failed to recover after additional cycles. Interestingly, CD45RA+CD27- effector T cells showed only slight increases for each patient throughout TMZ maintenance. Cytokine analysis did not show a clear trend except for consistent T cell expression of perforin and IP-10. One patient with recurrent GBM was successfully resected 110d following DRI. Histopathology revealed widespread necrosis with significant infiltration of CD4+ and CD8+ T cells and γδ T cells in the tumor parenchyma.

Conclusions: Standard of care TMZ is lymphodepleting and DRI therapy can be conducted in an environment favorable to T cell persistence and sustained immune response. One treated patient demonstrated infiltration of both αβ and γδ T cells 110 days following a single dose of DRI γδ T cells. TMZ maintenance during the Stupp regimen prolongs lymphodepletion, thereby presenting a favorable setting for adoptive cell therapy.

Citation Format: Lawrence S. Lamb, Lei Ding, Ryan C. Miller, Mariska ter Haak, Caitlyn Lucas, Becca Weekley, Samantha Youngblood, Cathy Langford, Guoling Chen, Louis B. Nabors. Maintenance-phase Temozolomide as a lymphodepletion platform for intracranialadoptiveγδ T cell-basedtherapy in primary high-grade gliomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1173.