The stress kinase GCN2 does not mediate suppression of antitumor T cell responses by tryptophan catabolism in experimental melanomas

Abstract

Tryptophan metabolism is a key process that shapes the immunosuppressive tumor microenvironment. The two rate-limiting enzymes that mediate tryptophan depletion, indoleamine-2,3-dioxygenase (IDO) and tryptophan-2,3-dioxygenase (TDO), have moved into the focus of research and inhibitors targeting IDO and TDO have entered clinical trials. Local tryptophan depletion is generally viewed as the crucial immunosuppressive mechanism. In T cells, the kinase general control non-derepressible 2 (GCN2) has been identified as a molecular sensor of tryptophan deprivation. GCN2 activation by tryptophan depletion induces apoptosis and mitigates T cell proliferation. Here, we investigated whether GCN2 attenuates tumor rejection in experimental B16 melanoma using T cell-specific knockout mice. Our data demonstrate that GCN2 in T cells did not affect immunity to B16 tumors even when animals were treated with antibodies targeting cytotoxic T lymphocyte antigen-4 (CTLA4). GCN2-deficient gp100 TCR-transgenic T cells were equally effective as wild-type pmel T cells against gp100-expressing B16 melanomas after adoptive transfer and gp100 peptide vaccination. Even augmentation of tumoral tryptophan metabolism in B16 tumors by lentiviral overexpression of did not differentially affect GCN2-proficient vs. GCN2-deficient T cells . Importantly, GCN2 target genes were not upregulated in tumor-infiltrating T cells. MALDI-TOF MS imaging of B16 melanomas demonstrated maintenance of intratumoral tryptophan levels despite high tryptophan turnover, which prohibits a drop in tryptophan sufficient to activate GCN2 in tumor-infiltrating T cells. In conclusion, our results do not suggest that suppression of antitumor immune responses by tryptophan metabolism is driven by local tryptophan depletion and subsequent GCN2-mediated T cell anergy.

Reimagining IDO Pathway Inhibition in Cancer Immunotherapy via Downstream Focus on the Tryptophan–Kynurenine–Aryl Hydrocarbon Axis

“…Recent studies have cast doubt on the Trp-depletion mechanisms (32,33) and highlighted the potent immunosuppressive activity of intratumoral Kyn and AhR signaling. To this end, in the evaluation of Trp-Kyn-AhR inhibitors, reduction of extracellular Kyn within the TME or downstream AhR transcriptional programs should be emphasized as major pharmacodynamic endpoints.…”

“…[29][30][31]. However, recent studies have questioned the significance of this mechanism (32,33). Accumulation of kynurenines induce effector T-cell arrest and lead to binding of AhR.…”

Characterization of the Selective Indoleamine 2,3-Dioxygenase-1 (IDO1) Catalytic Inhibitor EOS200271/PF-06840003 Supports IDO1 as a Critical Resistance Mechanism to PD-(L)1 Blockade Therapy

“…GCN2 is activated by elevations in uncharged tRNAs and phosphorylates eIF2a leading to inhibition of protein translation (6). Recent data however did not confirm the role of GCN2 (31). Tryptophan depletion may also inactivate the mTOR pathway, although this was not demonstrated in lymphocytes so far (8).…”

Upregulation of tryptophanyl-tRNA synthethase adapts human cancer cells to nutritional stress caused by tryptophan degradation

“…On the other hand, Trp depletion has been implicated in the proliferation arrest of CD8 + T cells 10 by the accumulation of uncharged transfer RNA (tRNA) and subsequent activation of the general control non-derepressible-2 (GCN2) kinase pathway 11,12 , while Gcn2 in T cells has recently been shown to be dispensable for the suppression of antitumor immune responses in experimental melanomas 13 .…”