Background
In recent years, T cell-based immunotherapies have shown promising results in hematologic malignancies. However, these strategies seem to be limited in solid cancers, posing more complex challenges including a hostile TME with nutrient deprivation and tissue hypoxia [1]. Additionally, metabolic reprogramming has been identified as a crucial factor for proper cytotoxic T-cell functions upon their activation. Such energy demands are answered by the upregulation of glycolysis, oxidative phosphorylation, and upregulation of nutrient transporters represented by SLCs [2,3]. Within the TME, tumor and immune cells compete for nutrients and shape a distinct metabolic milieu, resulting in an ineffective effector function [4]. Herein, we aim to metabolically engineer T cells to improve their fitness in the glucose-deprived TME and optimize ACT.
Materials and Methods
We retrovirally overexpressed the glucose transporter Slc2a1/GLUT1 in murine CD8+ T cells (CD8+Slc2a1). To assess T-cell fitness we conducted experiments in physiologic (5mM) and hypoglycemic (0.5mM) media conditions. CellTraceTM-based proliferation experiments and killing assays in the OT1-OVA model are used to examine differences to MOCK in functionality and were analyzed via flow cytometry and microscopy, respectively. Furthermore, Seahorse analyses, bulk RNA-Seq, and metabolomic analyses were performed to examine the mechanical background. Murine in vivo studies are performed to approach the translatability of this system into living organisms.
Results
CD8+Slc2a1 cells possessed a higher proliferative capacity in all conditions tested but most prominently in hypoglycemic (0.5mM) media. This better functional activity of CD8+Slc2a1 was also translated to higher killing rates in coculture assays with tumor cells, especially in low-glucose environments. Metabolic flux analyses and multi-omics suggested greater metabolic activity of CD8+Slc2a1 and revealed higher ROS production and upregulation of correlating anti-oxidative pathways, especially the pentose-phosphate pathway. Preliminary in vivo studies support the in vitro killing in a syngeneic tumor model. Furthermore, signs of altered memory formation were visible, expressed in a higher proportion of effector memory cells.
Conclusions
Our data point to the role of GLUT1 overexpression in T cells for improved cytotoxic activity, proliferation, and long-term persistence. Therefore, combinatorial approaches with GLUT1 overexpression could serve as a potential approach to increase efficacy in ACT against solid cancer. We also identified GLUT1-dependent reprogramming in CD8+Slc2a1 cells which is further investigated in ongoing studies. Additionally, we are evaluating the potential risk of this approach to neoplastic formation.
References
Treating hematological malignancies with cell therapy: where are we now? Landoni E, Savoldo B.; Expert Opin Biol Ther. 2018.
Anticancer targets in the glycolytic metabolism of tumors: a comprehensive review; Paolo E. Porporato et al. Frontiers in Pharmacology 2011.
Glucose Metabolism on Tumor Plasticity, Diagnosis, and Treatment; Lin Xiaoping et al. Frontiers in Oncology 2020
Fighting in a wasteland: deleterious metabolites and antitumor immunity. Watson MJ, Delgoffe GM. J Clin Invest. 2022.
M.E. Kirmaier: None. B.L. Cadilha: None. A. Hadzic: None. W. Schmitz: None. M.R. Benmebarek: None. D. Briukhovetska: None. S. Michaelides: None. V. Buschinger: None. B. Tast: None. C.H. Bönigk: None. S. Oganesian: None. L. Vona: None. A. Tischmacher: None. V. Heissmeyer: None. M. Vaeth: None. V.R. Buchholz: None. M. Eilers: None. M. von Bergwelt-Baildon: None. M. Subklewe: None. S. Kobold: None. S. Theurich: None.