(C) Tumor progression (= 8C10). T cell activities. Therefore, our results have recognized creatine as an important metabolic regulator controlling antitumor T cell immunity, underscoring the potential of creatine supplementation to improve T cellCbased malignancy immunotherapies. Graphical Abstract Open in a separate window Introduction T cells play a central role in mediating and orchestrating immune responses against malignancy; therefore, they are attractive therapeutic targets for treating malignancy (Couzin-Frankel, 2013; Page et al., 2014; Ribas, 2015; Rosenberg and Restifo, 2015; Baumeister et al., 2016; Lim and June, 2017). The maintenance and activation of T cells are energy-demanding activities, requiring the use of bioenergy in the form of ATP (Fox et al., 2005). Unique metabolic programs are used by T cells to generate ATP to support their diverse homeostatic and effector Thapsigargin functions (Fox et al., 2005; ONeill et al., 2016; Kidani and Bensinger, 2017; Zeng and Chi, 2017). In the tumor microenvironment, T cells face the special challenge of competing with fast-growing tumor cells for metabolic gas such as glucose, amino acids, and lipids, which can be limiting (McCarthy et al., 2013). Therefore, an efficient and economical bioenergy metabolism is needed for tumor-infiltrating T cells to mount and sustain effective anticancer responses (Siska Thapsigargin and Rathmell, 2015). However, the study of metabolic regulators controlling antitumor T cell immunity has just begun (Chang and Pearce, 2016; Ho and Kaech, 2017; Kishton et al., 2017; Patel and Powell, 2017). Here we show that creatine is usually a critical molecule buffering ATP levels in cancer-targeting CD8 T cells through maintaining a readily available high-energy phosphate reservoir (Wyss and Kaddurah-Daouk, 2000). We found that tumor-infiltrating immune cells (TIIs) up-regulated their expression of the creatine transporter gene (or (is an X-linked gene encoding a surface transporter (creatine transporter [CrT]) that controls the uptake of creatine into a cell in an Na+/K+-dependent manner, Rabbit Polyclonal to RPS12 where creatine is used to store high-energy phosphates and to buffer intracellular ATP levels through a CK/PCr/Cr (creatine kinase/phospho-creatine/creatine) system (Fig. 1 B; Wyss and Kaddurah-Daouk, 2000). Open in a separate window Physique 1. or = 3C4) measured by qPCR. Cells were collected on day 14 after tumor challenge. (B) Diagram showing creatine uptake and creatine-mediated bioenergy buffering in cells with high-energy demand. Cr, creatine; PCr, phospho-creatine; Crn, creatinine; CK, creatine kinase. (CCG) Study of B16-OVA tumor growth in = 3). (ECG) On day 14, tumors were collected from experimental mice, and TIIs were isolated for further analysis. (E) FACS plots showing the detection of tumor-infiltrating CD4 and CD8 T cells (gated as TCR+CD4+ and TCR+CD8+ cells, respectively). (F) FACS plot showing PD-1 expression on tumor-infiltrating CD8 T cells. (G) Quantification of F (= 3). Representative of two (A) and three (CCG) experiments, respectively. Data are offered as the mean SEM. *, P 0.05; **, P 0.01 by one-way ANOVA (A) or Students test (D and G). See also Fig. S1. Creatine is usually a nitrogenous organic acid that naturally occurs in vertebrates. It is mainly produced in the liver and kidneys but predominantly stored in skeletal muscle mass (Wyss and Kaddurah-Daouk, 2000). For humans, diet is also a major source of creatine (Wyss and Kaddurah-Daouk, 2000). Expression of CrT is usually important for cells demanding high energy, such as muscle mass cells and brain cells; in humans, CrT deficiency has been associated with muscle mass diseases and neurological disorders (Wyss and Kaddurah-Daouk, 2000). On the other hand, oral creatine supplements have been broadly used by bodybuilders and athletes to gain muscle mass and to improve overall performance (Kreider et al., 2017). However, the Thapsigargin function of CrT/creatine outside of the muscle mass and brain tissues is largely unknown. Since we found up-regulated gene expression in TIIs, we asked if the CrT/creatine system might also Thapsigargin regulate the energy metabolism of tumor-fighting immune cells, in particular CD8 cytotoxic T cells, which have a massive demand for Thapsigargin energy and can benefit from an energy storage/ATP buffering system (Fig. 1 B). gene expression in tumor-infiltrating WT CD8 T cell subsets showed an up-regulation of gene expression that was more significant in the PD-1hi subset than in the PD-1lo subset, suggesting a possible opinions loop in PD-1hi CD8 T cells that compensates for bioenergy insufficiency by increasing creatine uptake (Fig. S1 K). In particular, the PD-1hiTim-3hiLAG-3hi tumor-infiltrating CD8 T cells, which are considered to be the most worn out, expressed the.