Abstract 4236: Temporal inhibition of BCAT1 alters metal homeostasis leading to reversal of terminal exhaustion of CD8 T cells, increased cytotoxicity, and increased efficacy of checkpoint inhibition in cancer

作者: Vu, Hong A. ; Li, Weixing ; Papathanassiu, Adonia E.

AbstractWe have previously demonstrated that BCAT1, the enzyme responsible for the reversible transamination of leucine in the cytosol, is a druggable target in immune-oncology: the small molecule inhibitor ERG245 dramatically increases the efficacy of anti-PD-1 treatment in the moderately immunogenic CT26 colon cancer model. We have also shown that Bcat1 gene expression is enriched in exhausted CD4+ and CD8+ intratumoral cells in cancer patients from diverse cancers. Here, we illustrate that the adoptive cell transfer (ACT) of murine CD8+ T cells treated with ERG245 increased the median survival of CT26 tumor-bearing mice from 25 days, observed with the ACT of untreated CD8+ T cells, to 32 days. This suggested that inhibition of BCAT1 prevented the known exhaustion of adoptively transferred CD8+ T cells, induced by the tumor microenvironment of CT26 tumors. To confirm this hypothesis, we developed an in vitro model of generating terminally exhausted (PD-1+TIM-3+GZMB+CXCL13+) CD8+ T cells by subjecting the cells to consecutive cycles of αCD3/αCD28 activation in the presence of TGF-β. Addition of ERG245 in the last activation cycle decreased terminal exhaustion of the cells. Mechanistically, bulk RNAseq experiments indicated and ICP analysis confirmed that inhibition of BCAT1 interfered with metal homeostasis leading to increased concentrations of Fe in the mitochondria and decreased concentrations of Zn in the nucleus of newly activated CD8+ T cells. The immediate impact of ERG245 treatment on the cells included a general decrease in the genes of processes depending on proper metal equilibrium such as cholesterol biosynthesis and ribosome biogenesis. Withdrawal of BCAT1 restored or reversed the cell phenotype leading to epigenetically altered CD8+ T cells with decreased histone acetyltransferase and histone demethylase presence, increased cytotoxicity, and increased ribosome biogenesis. Collectively, the data suggest that temporal inhibition of BCAT1 inhibition induces profound effects on CD8+ T cells including increased cytotoxicity and suppression of terminal exhaustion, thus rendering the cells rescuable by anti-PD-1 therapy.Citation Format: Adonia E. Papathanassiu, Weixing Li, Hong A. Vu. Temporal inhibition of BCAT1 alters metal homeostasis leading to reversal of terminal exhaustion of CD8 T cells, increased cytotoxicity, and increased efficacy of checkpoint inhibition in cancer [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 4236.

2021-07-01·Cancer Research

Abstract 1202: ERG344: A novel IRG1 inhibitor for the treatment of colon cancer

作者: Lambrechts, Diether ; Vu, Hong A. ; Lodi, Francesca ; Papathanassiu, Adonia E.

AbstractImmune responsive gene 1 (Irg1) is an LPS-inducible gene within macrophages (MF) whose protein product is responsible for the synthesis of itaconate from aconitate through a break in the TCA cycle. Although primarily present in proinflammatory macrophages, IRG1 expression has been linked with certain cancers (glioblastoma, ovarian carcinoma) and its tumor-driven upregulation in peritoneal resident macrophages (pResMF) has been shown to contribute to peritoneal tumor growth. Here, we report Irg1 gene expression at the single cell level (scRNAseq) in samples obtained from colorectal cancer patients (n=7; ~11,000 single cells). Transcriptomic analysis showed that Irg1 was primarily expressed by cancer cells, although other tumoral cell types also showed substantial levels of the gene. To determine if IRG1 is a druggable target in cancer, we synthesized a series of small MW inhibitors and selected ERG344 for in vivo testing based on its ability to inhibit itaconate synthesis in a rat glioblastoma cell line (C6) engineered to stably express Irg1. The IC50 value of inhibition was ~150 μM. In vivo, ERG344 suppressed tumor growth and increased survival probability in a syngeneic colon cancer model (CT26). Drug administration was initiated when the tumors reached an average tumor volume of ~100 mm3 and involved daily ERG344 doses of 0.2 mg/kg ip or 0.3 mg/kg po. Complete tumor regressions (~17%) were observed in the same model when ERG344 was administered ip at 3 mg/kg. Immunophenotyping of CT26 tumors treated with either vehicle or 0.2 mg/kg ERG344 ip indicated that IRG1 inhibition renders the tumor microenvironment less immunosuppressive by decreasing the frequency of monocytic myeloid suppressor cells (m-MDSCs), increasing the frequency of cytotoxic CD8+ T cells, and decreasing the CD8+ expression of the negative co-stimulatory molecule PD-1. To determine if ERG344 is also able to reduce pResMF-driven peritoneal tumor growth, murine melanoma B16F15 cells were injected into the peritoneal cavity of C57BL/6 mice treated with vehicle or 1 mg/kg ERG344 (n=8/group). Seven days after tumor inoculation, mice treated with ERG344 showed reduced tumor growth in the peritoneal cavity, which led to increased survival probability of the animals. The studies collectively suggest that IRG1 is a novel target for the treatment of primary and metastatic tumor growth in colorectal cancer and highlight the therapeutic potential of ERG344.Citation Format: Adonia E. Papathanassiu, Francesca Lodi, Hong A. Vu, Diether Lambrechts. ERG344: A novel IRG1 inhibitor for the treatment of colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1202.

2020-11-15·Journal of Cell Science2区 · 生物学

BCAT1 affects mitochondrial metabolism independently of leucine transamination in activated human macrophages

2区 · 生物学

Article

作者: Frezza, Christian ; Park, Kwon-Sik ; Buang, Norzawani ; Ko, Jeong-Hun ; Mauro, Claudio ; Papathanassiu, Adonia E. ; Olona, Antoni ; Behmoaras, Jacques ; Costa, Ana S. H.

ABSTRACTIn response to environmental stimuli, macrophages change their nutrient consumption and undergo an early metabolic adaptation that progressively shapes their polarization state. During the transient, early phase of pro-inflammatory macrophage activation, an increase in tricarboxylic acid (TCA) cycle activity has been reported, but the relative contribution of branched-chain amino acid (BCAA) leucine remains to be determined. Here, we show that glucose but not glutamine is a major contributor of the increase in TCA cycle metabolites during early macrophage activation in humans. We then show that, although uptake of BCAAs is not altered, their transamination by BCAT1 is increased following 8 h lipopolysaccharide (LPS) stimulation. Of note, leucine is not metabolized to integrate into the TCA cycle in basal or stimulated human macrophages. Surprisingly, the pharmacological inhibition of BCAT1 reduced glucose-derived itaconate, α-ketoglutarate and 2-hydroxyglutarate levels without affecting succinate and citrate levels, indicating a partial inhibition of the TCA cycle. This indirect effect is associated with NRF2 (also known as NFE2L2) activation and anti-oxidant responses. These results suggest a moonlighting role of BCAT1 through redox-mediated control of mitochondrial function during early macrophage activation.

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