Blocking Glycolytic Metabolism Increases Memory T Cells and Antitumor Function

After encountering antigen, naïve CD8<sup>+</sup> T cells undergo an extensive period of proliferation and expansion, and differentiate into effector cells and distinct memory T cell subsets. Increasing glycolytic flux pushes CD8<sup>+</sup> T cells towards a terminally differentiated state that diminishes antitumor activity. In contrast, inhibiting glycolysis using 2-deoxyglucose (2DG) maintains the formation of long-lived memory CD8<sup>+</sup> T cells and enhances antitumor activity

After encountering antigen, naïve CD8+ T cells undergo an extensive period of proliferation and expansion, and differentiate into effector cells and distinct memory T cell subsets. Increasing glycolytic flux pushes CD8+ T cells towards a terminally differentiated state that diminishes antitumor activity. In contrast, inhibiting glycolysis using 2-deoxyglucose (2DG) maintains the formation of long-lived memory CD8+ T cells and enhances antitumor activity.

CD8+ T cells are a major component of the cellular immune response, which is necessary to control a variety of bacterial and viral infections. CD8+ T cells also play a major role in the cell-mediated antitumor immune response. After encountering antigen, naïve CD8+ T cells undergo an extensive period of proliferation and expansion, and differentiate into effector cells and distinct memory T cell subsets. Preclinical studies using adoptive transfer of purified CD8+ T cells have shown that the ability of T cells to proliferate and survive for a long time after transfer is associated with effective antitumor and antiviral responses. Understanding how the formation of long-lived memory T cell subsets is controlled may enable development of more potent immunotherapies against cancer and infectious diseases.

Cellular metabolism is very important for regulating CD8+ T cell differentiation and memory formation. Naïve CD8+ T cells depend on oxidation of fatty acids as a primary source of energy. Following antigen encounter, T cells shift to a glycolytic metabolism to support effector function. It remains unclear whether changes in glucose metabolism promote activated T cells to become long-lived memory cells. Nicholas Restifo, M.D., in CCR’s Surgery Branch and Luca Gattinoni, M.D. in CCR’s Experimental Transplantation and Immunology Branch, and their colleagues set out to investigate whether specific changes in glucose metabolism can influence long-term survival of activated T cells and their ability to enter into the memory pool in vivo.

The researchers used a fluorescent glucose analog, 2-NBDG, to directly measure glucose incorporation in living cells. They found that 2-NBDGlo cells incorporated small amounts of glucose, and had a molecular profile characteristic of memory precursor cells and an increased ability to enter the memory pool compared with 2-NBDGhi cells that took up high amounts of glucose. When mice that were infected with vaccinia virus received a transfer of 2-NBDGhi and 2-NBDGlo cells, the 2-NBDGhi cells exhibited poor engraftment, proliferation, and survival capacities compared with 2-NBDGlo cells. These results indicate that activated CD8+ T cells separated based on intrinsic differences in glucose metabolism have varied outcomes in vivo as CD8+ T cells with high glycolytic metabolism usually become short-lived and cells with low glycolytic metabolism are more likely to become memory cells.  

To determine whether differences in glucose metabolism in CD8+ T cells were associated with differences in transcriptional regulation of T cell differentiation, mRNA levels of key transcription factors and molecules associated with either memory precursors or short-lived, terminally differentiated effector T cells in 2-NBDGhi and 2-NBDGlo cells were measured. Low levels of glycolysis were found to be associated with high expression of transcription factors that support CD8+ T cell memory, such as Tcf7, Lef1, and Bcl6, as well as increased activity of Foxo1, an important regulator of memory T cell differentiation. High levels of glycolysis were associated with an upregulation of transcription factors that regulate terminal effector differentiation, such as Blimp-1, and increased expression of parts of the effector system including perforin and granzyme B. These findings indicate that glucose uptake allowed for the separation of T cells having molecular profiles associated with effector (2-NBDGhi) and memory precursor cells (2-NBDGlo).

The next step was to investigate whether the high glycolytic flux was simply associated with cells with an impaired ability to become memory cells or was the direct cause for the observed defect in CD8+ T cell memory formation. Enforcing glycolytic metabolism by overexpressing the glycolytic enzyme phosphoglycerate mutase-1 severely limited the formation of CD8+ T cell memory in vivo.  On the other hand, when an inhibitor of glycolysis, 2-deoxyglucose (2DG) was used during CD8+ T cell activation, long-lived memory cell generation and antitumor functionality were enhanced.

All together, these results demonstrate that increasing glycolytic flux pushes CD8+ T cells towards a terminally differentiated state, while inhibiting glycolysis maintains the formation of long-lived memory CD8+ T cells. These results may lead to improved design of T cell-based therapies and vaccines for prevention and treatment of chronic infectious diseases and cancer. Reagents, including 2DG, that are currently under evaluation in clinical trials due to their direct negative impact on glycolytic tumor cells might be repurposed to improve CD8+ T cell memory formation.

Summary Posted: 12/2013

Reference

Sukumar M, Liu J, Ji Y, Subramanian M, Crompton JG, Yu Z, Roychoudhuri R, Palmer DC, Muranski P, Karoly E, Mohney R, Klebanoff CA, Lal A, Finkel T, Restifo NP, Gattinoni L. Inhibiting glycolytic metabolism enhances CD8<sup>+</sup> T-cell memory and anti-tumor function. J. Clin. Invest. 2013 PubMed Link