Background: Immunotherapy using checkpoint blockade has demonstrated impressive, durable responses in select patients with solid malignancies. This is especially true for the blockade of programmed death 1 (PD1). Currently, efforts are focused on understanding non-response. One hypothesis is that multiple inhibitory receptors (IRs) are upregulated on hypofunctional tumor-infiltrating lymphocytes (TILs) and necessitates blocking multiple IRs to increase the response rate. We describe data from a unique xenograft model of human lung cancer where tumor-reactive human T cells become hypofunctional and upregulate multiple IRs including PD1 and TIM3 (T cell immunoglobulin and mucin-domain containing-3). Combining a single IV dose of tumor-reactive T cells with blockade of both PD1 and TIM3 led to greater control of tumor growth than combining it with blockade of either alone.

Materials/Methods: The A549 human lung cancer cell line was transduced to stably express the cancer-testis antigen, NYESO1, expressed in the contact of HLA-A2 (A549-A2-ESO). 5x106 A549-A2-ESO cells were injected subcutaneously into the flanks NSG mice. Activated T cells from healthy donors were lentivirally transduced with a TCR targeting NYESO1 (Ly95). After two weeks when tumors were established and measured ~150mm3, the mice were randomly assigned to one of the following treatments: 1) untreated, 2) non-transduced (NTD) T cells plus anti-PD1 and anti-TIM3 Ab, 3) Ly95 T cells, 4) Ly95 T cells + anti-PD1 Ab, 5) Ly95 T cells + anti-TIM3 Ab, 6) Ly95 T cells + anti-PD1 and anti-TIM3 Ab. T cells were injected IV at a single dose of 10x106/mouse. Abs were injected intraperitonealy (IP) at 10mg/kg every 5 days from the time of T cell injection. Tumor volumes were measured serially. At the end, the mice were sacrificed and the tumors were harvested, digested, and processed into single cell suspension. Flow cytometry was performed to look at degree of TIL infiltration and expression of surface markers. Ficoll gradient was used to isolate the T cells to conduct functional analyses.

Results/Conclusion: By twenty days post T cell injection, the tumors in the Ly95 T cell group (3) were ~30% smaller than that of the untreated (1) and the NTD T cell + Ab group (2). Anti-PD1 Ab augmented Ly95 T cell control of tumor (4) (further decrease in size by ~30%) (4). Anti-TIM3 Ab had no effect on Ly95 T cell tumor control (5). Anti-PD1 plus anti-TIM3 Abs had the greatest augmentation on Ly95 T cell control of tumor size (6) (greater than 50% reduction in tumor size compared Ly95 T cells alone) The greatest TIL infiltration and ex-vivo TIL function were seen in tumors that received Ly95 T cell + anti-PD1 and anti-TIM3 Abs. Flow cytometry analysis revealed dynamic, adaptive upregulation of one IR when another IR was blocked. This explained why augmentation of Ly95 T cell control of tumor was greatest with combinatorial Ab blockade.

Citation Format: Edmund Moon, Soyeon Kim, Shaun O'Brien, Naomi Saint Jean, Raluca Verona, Linda Snyder, Yangbing Zhao, Steven Albelda. Dual antibody blockade of TIM3 and PD1 on NYESO1 redirected human T cells leads to augmented control of lung cancer tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1641. doi:10.1158/1538-7445.AM2017-1641