IL4i1 and IDO1: Oxidases that control a tryptophan metabolic nexus in cancer.

Regulated tryptophan metabolism by immune cells has been associated with the promotion of tolerance and poor outcomes in cancer. The main focus of research has centered on local tryptophan depletion by IDO1, an intracellular heme-dependent oxidase that converts tryptophan to formyl-kynurenine. This is the first step of a complex pathway supplying metabolites for de novo NAD+ biosynthesis, 1-carbon metabolism, and a myriad of kynurenine derivatives of which several act as agonists of the arylhydrocarbon receptor (AhR). Thus, cells that express IDO1 deplete tryptophan while generating downstream metabolites. We now know that another enzyme, the secreted L-amino acid oxidase IL4i1 also generates bioactive metabolites from tryptophan. In tumor microenvironments, IL4i1 and IDO1 have overlapping expression patterns, especially in myeloid cells, suggesting the two enzymes control a network of tryptophan-specific metabolic events. New findings about IL4i1 and IDO1 have shown that both enzymes generate a suite of metabolites that suppress oxidative cell death ferroptosis. Thus, within inflammatory environments, IL4i1 and IDO1 simultaneously control essential amino acid depletion, AhR activation, suppression of ferroptosis, and biosynthesis of key metabolic intermediates. Here, we summarize the recent advances in this field, focusing on IDO1 and IL4i1 in cancer. We speculate that while inhibition of IDO1 remains a viable adjuvant therapy for solid tumors, the overlapping effects of IL4i1 must be accounted for, as potentially both enzymes may need to be inhibited at the same time to produce positive effects in cancer therapy.

Kynurenine importation by SLC7A11 propagates anti-ferroptotic signaling.

IDO1 oxidizes tryptophan (TRP) to generate kynurenine (KYN), the substrate for 1-carbon and NAD metabolism, and is implicated in pro-cancer pathophysiology and infection biology. However, the mechanistic relationships between IDO1 in amino acid depletion versus product generation have remained a longstanding mystery. We found an unrecognized link between IDO1 and cell survival mediated by KYN that serves as the source for molecules that inhibit ferroptotic cell death. We show that this effect requires KYN export from IDO1-expressing cells, which is then available for non-IDO1-expressing cells via SLC7A11, the central transporter involved in ferroptosis suppression. Whether inside the "producer" IDO1+ cell or the "receiver" cell, KYN is converted into downstream metabolites, suppressing ferroptosis by ROS scavenging and activating an NRF2-dependent, AHR-independent cell-protective pathway, including SLC7A11, propagating anti-ferroptotic signaling. IDO1, therefore, controls a multi-pronged protection pathway from ferroptotic cell death, underscoring the need to re-evaluate the use of IDO1 inhibitors in cancer treatment.

Gene-selective transcription promotes the inhibition of tissue reparative macrophages by TNF.

Anti-TNF therapies are a core anti-inflammatory approach for chronic diseases such as rheumatoid arthritis and Crohn's Disease. Previously, we and others found that TNF blocks the emergence and function of alternative-activated or M2 macrophages involved in wound healing and tissue-reparative functions. Conceivably, anti-TNF drugs could mediate their protective effects in part by an altered balance of macrophage activity. To understand the mechanistic basis of how TNF regulates tissue-reparative macrophages, we used RNAseq, scRNAseq, ATACseq, time-resolved phospho-proteomics, gene-specific approaches, metabolic analysis, and signaling pathway deconvolution. We found that TNF controls tissue-reparative macrophage gene expression in a highly gene-specific way, dependent on JNK signaling via the type 1 TNF receptor on specific populations of alternative-activated macrophages. We further determined that JNK signaling has a profound and broad effect on activated macrophage gene expression. Our findings suggest that TNF's anti-M2 effects evolved to specifically modulate components of tissue and reparative M2 macrophages and TNF is therefore a context-specific modulator of M2 macrophages rather than a pan-M2 inhibitor.

Neuroblastoma Formation Requires Unconventional CD4 T Cells and Arginase-1-Dependent Myeloid Cells.

Immune cells regulate tumor growth by mirroring their function as tissue repair organizers in normal tissues. To understand the different facets of immune-tumor collaboration through genetics, spatial transcriptomics, and immunologic manipulation with noninvasive, longitudinal imaging, we generated a penetrant double oncogene-driven autochthonous model of neuroblastoma. Spatial transcriptomic analysis showed that CD4+ and myeloid populations colocalized within the tumor parenchyma, while CD8+ T cells and B cells were peripherally dispersed. Depletion of CD4+ T cells or CCR2+ macrophages, but not B cells, CD8+ T cells, or natural killer (NK) cells, prevented tumor formation. Tumor CD4+ T cells displayed unconventional phenotypes and were clonotypically diverse and antigen independent. Within the myeloid fraction, tumor growth required myeloid cells expressing arginase-1. Overall, these results demonstrate how arginine-metabolizing myeloid cells conspire with pathogenic CD4+ T cells to create permissive conditions for tumor formation, suggesting that these protumorigenic pathways could be disabled by targeting myeloid arginine metabolism. SIGNIFICANCE: A new model of human neuroblastoma provides ways to track tumor formation and expansion in living animals, allowing identification of CD4+ T-cell and macrophage functions required for oncogenesis.

Lactate and IL6 define separable paths of inflammatory metabolic adaptation.

Lactate is an end point of Warburg-type metabolism found in inflammatory macrophages. Recently, lactate was shown to modify histones of lipopolysaccharide (LPS)-activated macrophages in a time-dependent way and promote the expression of genes linked to tissue repair, including arginase-1 (Arg1). We tested the interrelationships between histone lactylation (Kla) and tissue reparative gene expression and found that Kla was uncoupled from changes in gene expression linked to resolving M2 macrophage activation but correlated with Arg1 expression. LPS-induced Arg1 was instead dependent on autocrine-paracrine interleukin-6 (IL6) production, the IL6 receptor, and Stat3 signal transduction. We found that Kla increases as macrophages prepare to die under inflammatory stress, and Kla was absent in macrophages that cannot generate reactive nitrogen or have defects in diverse macrophage death pathways. Thus, Kla is a consequence rather than a cause of macrophage activation but occurs coincidently with an IL6- and Arg1-dependent metabolic rewiring under inflammatory duress.

Anti-ferroptotic mechanism of IL4i1-mediated amino acid metabolism.

Interleukin-4-induced-1 (IL4i1) is an amino acid oxidase secreted from immune cells. Recent observations have suggested that IL4i1 is pro-tumorigenic via unknown mechanisms. As IL4i1 has homologs in snake venoms (L-amino acid oxidases [LAAO]), we used comparative approaches to gain insight into the mechanistic basis of how conserved amino acid oxidases regulate cell fate and function. Using mammalian expressed recombinant proteins, we found that venom LAAO kills cells via hydrogen peroxide generation. By contrast, mammalian IL4i1 is non-cytotoxic and instead elicits a cell protective gene expression program inhibiting ferroptotic redox death by generating indole-3-pyruvate (I3P) from tryptophan. I3P suppresses ferroptosis by direct free radical scavenging and through the activation of an anti-oxidative gene expression program. Thus, the pro-tumor effects of IL4i1 are likely mediated by local anti-ferroptotic pathways via aromatic amino acid metabolism, arguing that an IL4i1 inhibitor may modulate tumor cell death pathways.