Lung cancer (LC) is one of the most common malignancies worldwide and has a high mortality rate (1). Previous studies have shown that lung carcinogenesis is attributed to the gain-functional mutation of several cancer-associated genes, including the epidermal growth factor receptor (EGFR), Kirsten rat sarcoma viral oncogene homolog (KRAS), and v-raf murine sarcoma viral oncogene homolog B1 (BRAF) (2–4). Actually, therapeutic drugs targeting these molecules have been demonstrated to prolong the survival of LC patients, particularly for non-small cell lung cancer (NSCLC) patients. However, therapeutic efficacy of these drugs is limited due to rapid development of drug resistance in LC patients (4–6). Therefore, other effective treatments are urgently needed. Currently, cancer has been thought not to be a genetic disease, rather than a metabolic disease, which is associated with tumor immune escape (7, 8). It is well known that tumor cells usually undergo aerobic glycolysis for their glucose metabolism, known as the Warburg effect (9). Moreover, extensive studies have revealed that alternations in metabolisms are not only for glucose, but also for amino acid, lipid, nucleotide, and others in cancer (10). Notably, tryptophan (Try) metabolism is a particularly compelling physiological context in LC because of its complex and multifaceted effect on LC cells and cancer-associated cells in immune escape (11).

Try cannot be synthesized directly by the human body and has the lowest levels in the human body among 20 essential amino acids such that it depends on food protein. Similar to other essential amino acids, Try is essential for biosynthesizing cellular protein and formatting cytoskeleton (12). In the circulation, most Try binds to albumin for transportation and only 10%–20% of it remain free amino acid (13, 14). The free Try is mainly degraded through the kynurenine (KYN) pathway and is metabolized to form serotonin or other metabolites (15). Try plays a significantly physiological role in synthesizing proteins. However, the metabolic formation of serotonin and the KYN pathway-mediated metabolism, together with the lack of its endogenous production, may make Try shortage that can impair the protein synthesis. In the KYN pathway, Try is firstly converted to formyl-kynurenine, which is rapidly degraded to KYN by key enzymes of indoleamine-pyrrole 2,3-dioxygenase (IDO)1, IDO2, and tryptophan 2,3-dioxygenase (TDO), particularly by IDO1 (14, 16). Next, KYN is catalyzed into a series of metabolites, including anthranilic acid (AA), kynurenic acid (KA), xanthurenic acid (XA), 3-hydroxyanthranilic (3-HAA), quinolinic acid (QA), and NAD+ (17). In the lung, Try degradation is mainly catalyzed by IDO1 because IDO1 is constitutively expressed in many organs while TDO is predominantly expressed in the liver (18). Previous studies have shown that most Try metabolites in the KYN pathway are associated with the development of many diseases, including cancer. Actually, the IDO1-related Try metabolites are associated with lung cancer development (19, 20). This review aims to summarize the research advance in how Try and its metabolites contribute to the development and progression of LC.

The immune escape is a “hallmark of cancer” (70, 71). Tumor immune escape refers to the phenomenon, in which tumor cells can grow and metastasize by avoiding recognition and attack by the immune system through various mechanisms (72). Currently, IDO1 has been suggested to be important for immune escape of LC. First, upregulated IDO1 expression promotes the degradation of Try and the accumulation of its metabolites (such as KYN, 3-HAA and others) in LC. These metabolites act on various immune cells, including T cells (naive CD4+ T cells, Th17, and Treg), antigen-presenting cells (APC, DCs, and macrophages), and NK cells, and lead to immune escape. The promising mechanisms by which Try metabolites induce cancer immune tolerance and immunosuppression are summarized in Figure 1 .

First, Try is an essential amino acid for immune cell proliferation, and Try depletion results in T-cell apoptosis, which is one major reason for cancer immunosuppression (73). The decreased Try levels can inhibit T-cell proliferation by activating general control over nonderepressible 2 (GCN2) kinase and suppressing the mTOR signaling, a target of rapamycin (74–76). The GCN2 is a serine/threonine kinase and can phosphorylate eukaryotic initiation factor 2a kinase (eIF2a) in the presence of low concentration of Try, inhibiting protein synthesis and T cell proliferation (74). Activated GCN2 can also promote the differentiation of naïve CD4+ T cells into Tregs (16, 74, 77). Furthermore, GCN2 can alter the phenotype of DCs and macrophages (75, 76, 78), making them prone to immunosuppression to promote tumorigenesis. In contrast, other studies argue that GCN2 is a sensor of amino acid starvation and its activation is not dependent on a low Try level, rather than deficiency in many amino acids (79, 80). Actually, T cells with GCN2 deficiency have similar activity to wild-type T cells in B16 melanoma-bearing mice (79), which contradicts the tumor promotion of GCN2. Apparently, there may be another mechanism that senses Try-deprived condition to regulate T cell immunity against tumor. The mTOR signaling appears to be a possible candidate (81, 82) because inhibition of mTOR complex 1 (mTORC1) can induce T-cell autophagy and anergy in the tumor microenvironment (83). Moreover, mTORC1 inhibition can also induce Treg cells to suppress anti-tumor immune responses (82).

Second, increased KYN can lead to immune tolerance by inhibiting T cell proliferation and inducing T cell apoptosis to promote tumor growth (38). The KYN is a ligand of AhR, and its activation promotes Treg cell differentiation that can directly inhibit anti-tumor immune responses, contributing to cancer immune escape (77, 84). The AhR activation can also direct DCs and macrophages toward an immune-suppressive phenotype (85–87). The AhR activation enhances IL-10 synthesis and secretion, and inhibits the IFN-β signaling in DCs, but induces IL-10 and IFN-α production in NK cells, respectively. Consistently, higher frequency of Tregs is detected in mice bearing cisplatin-resistant tumors than those bearing cisplatin-sensitive tumors (26).

Third, the downstream metabolites (such as 3-HAA and QA) of KYN can also induce T-cell apoptosis (88), contributing to immune tolerance. Recent studies have shown that QA can inhibit the proliferation of cancer-specific CD8+ cytotoxic T and NK cells to promote tumor growth (89). Furthermore, LC patients with lower plasma 3-HAA, the precursor of QA, benefit more from ICI treatment, suggesting that plasma 3-HAA levels may be a biomarker for predicting the response of LC patients to ICIs (23). The lower plasma 3-HAA may reflect less immunosuppression in patients because 3-HAA can promote Treg responses to produce high levels of TGF-β that decrease effector T-cell function, leading to immunosuppression (90). However, its precise mechanism in tumor immunity is not clear.

Next, IDO1 expression can be regulated by cytokines, such as IL-10 and IFN-γ (91, 92) while IDO1 inhibition can enhance T-cell proliferation and infiltration in the tumor environment and IL-2 production (93). Furthermore, IDO1 or IDO2 deficiency can modulate the tumor microenvironment by reducing KTR, enhancing immune cell infiltration and IFN-γ production (67). TDO and IDO2 act as the Try-metabolizing enzymes and can also promote Try degradation, resulting in immune regulation similar to IDO1. However, there are few reports and further studies are needed.

Last, IDO1 and TDO catalyze the production of several downstream Try metabolites, such as KYN (64, 84), KA (94), and XA (66), which can activate the AhR and may contribute to the immune modulation of IDO1 and TDO. Interestingly, KYN can directly bind and activate the AhR, with a high affinity at low picomolar levels (95). However, whether similar mechanisms also apply to other polar metabolites that activate the AhR, such as KA, remains to be investigated. In addition, AhR can regulate IDO-related regulatory phenotype in DCs (96). Here, an autocrine IDO−KYN/AhR−IDO feedback loop may contribute to the immune modulation (97, 98).

Enhanced IDO1 expression and activity can evade immunosurveillance and are associated with poor prognosis of LC. Therefore, inhibition of IDO1 may be an ideal strategy for intervention of LC. There are several direct IDO1 inhibitors available, including epacadostat and navoximod that neither directly kill tumor cells, nor spontaneously initiate an immune response (99). Unlike epacadostat, the Try mimetic indoximod (D-1MT, 1-methyl-D-tryptophan) is the first non-enzyme inhibitory drug that targets the IDO1 pathway and can inhibit lung tumor growth in vivo (100–102). Indoximod can directly act on immune cells by creating an artificially Try-related signal, relieving the IDO1-mediated immunosuppression (99). There are ongoing clinical trials that investigate anti-IDO1 agents as monotherapy or adjuvant therapies with other drugs for various solid tumors. The clinical trials of anti-IDO1 agents for different combination strategies, such as combination with ICIs, other immunomodulators, and chemotherapy, are summarized in Table 1 .

Epacadostat, a small-molecule IDO1 inhibitor, was developed by Incyte and is being tested for its therapeutic efficacy and safety in an advanced stage of clinical trial (103). The phase I/II KeyNote 037/ECHO 301 trial to test the safety and efficacy of different doses of epacadostat combined with 200 mg pembrolizumab (i.e., an anti-PD1 agent) every 3 weeks (Q3W) in 62 patients with different types of cancers has achieved promising results (104). There were 24% of patients experiencing high-grade toxicities but no treatment-related death and 12 out of 22 patients obtained objective responses. Unfortunately, a further phase III clinical trial with epacadostat 100 mg twice a day (BID) and pembrolizumab 200 mg (Q3W) failed to improve progression-free survival (PFS) in patients with metastatic melanoma (105). Because of the limitations of this trial, further clinical trials are necessary to test its therapeutic efficacy and safety.

The phase I ECHO-110 study was designed to test epacadostat at different doses combined with atezolizumab (i.e., an anti-PD-L1 agent) 1,200 mg Q3W in 29 patients with stage IIIB/IV NSCLC, who had previously been treated with ≥1 prior line of platinum-based chemotherapy (≥2 cycles), but not with checkpoint/IDO inhibitors. Similarly, 7 out of 29 patients displayed high-grade toxicities but no treatment-related death. Epacadostat at a dose up to 300 mg BID combined with atezolizumab 1,200 mg Q3W was well-tolerated in patients with previously treated NSCLC (103). However, only one patient achieved objective response. The low therapeutic efficacy may stem from the fact of almost all patients with negative PD-L1 expression. Similarly, the single-arm combination of the ECHO-301 trial also failed, lining with the results from other Phase II and III trials conducted in different settings (17) and was converted into the randomized phase II trials of epacadostat combined with pembrolizumab in LC patients. In addition, the KEYNOTE-715-06/ECHO-306-06 trials with the combination of epacadostat, pembrolizumab, and platinum-based chemotherapy did not obtain promising benefit in overall response rate in NSCLC patients (Clinicaltrial.gov.). These observations suggest that the combination of Epacadostat and a PD-1/PD-L1 blockade may not be valuable for patients with PD-L1 negative LC. However, whether this treatment strategy can achieve positive responses in PD-L1 expressing NSCLC or whether combination with platinum-based chemotherapy can achieve a better outcome in NSCLC patients has not been clarified. The ongoing, randomized, phase 2 ECHO-305 (NCT03322540) and ECHO-306 (NCT03322566) trials may give promising results.

New IDO inhibitors, such as navoximod (NLG-919/GDC-0919) and BMS-986205, are also being tested in clinical trials (106). In a phase I study of the IDO1 inhibitor, combination of navoximod and atezolizumab displayed acceptable safety, tolerability, and pharmacokinetics, but not clear beneficial evidence of navoximod in patients with advanced solid tumors (107).

There are questions on whether epacadostat doses used in the ECHO-301 trial could effectively inhibit IDO1 activity in the tumor microenvironment and whether targeting multiple enzymes in the KYN pathway to control Try metabolism would benefit to these patients (58).

There are also ongoing trials testing IDO1 and TDO dual inhibitors such as HTI-1090 (SHR9146) as a monotherapy for solid tumors (NCT03208959). The dual inhibitor of DN1406131 is being tested for its safety in healthy subjects (NCT03641794) while RG70099 from Curadev/Roche is still in preclinical development (108).

In a word, most researchers have focused on IDO/TDO inhibitors for the treatment of LC, and some of them have already been tested in clinical trials. However, the current therapeutic efficacy appears limited. Thus, further studies are necessary to understand the biological functions of Try and its metabolites in the development and progression of LC. Given that the KYN downstream metabolites have profound functions in regulating T cell immunity against LC, these metabolites and their catalyzing enzymes may be explored for development of therapies for LC. Similarly, combination of IDO/TDO inhibitors and other therapies (chemotherapy, radiotherapy, targeted therapy, and immunotherapy) should be pursued to determine the safety and therapeutic efficacy in LC. Previous studies have demonstrated that patient’s metabolism (BMI variation and hypercholesterolemia) has a significant impact on the outcome of PD1 inhibitor treatment in LC patients (109, 110). Some drugs can regulate body metabolism and are significantly related to clinical outcomes of ICI treatment in LC patients (111, 112). Metformin, an effective agent for the management of type 2 diabetes mellitus, in combination with ICI treatment can improve the anticancer effects of ICIs (113, 114). Statins can inhibit cholesterol production (115) and is associated with better clinical outcome of anti-PD1 treatment in advanced NSCLC patients in an intensity-dependent manner (111). However, IDO1 as an immune checkpoint is not as well studied as PD1, and the role of patient metabolism and drugs involved in its regulation on the outcome of patients treated with IDO/TDO inhibitors needs to be further confirmed. If demonstrated, IDO/TDO inhibitors may benefit many patients with LC.

Currently, modulation of Try metabolism has been used for diagnosis, prognosis, and therapies for LC. The levels of circulating IDO activity and downstream metabolites (3-HAA, QA, KA, etc.) can be used to predict the efficacy of different treatments in LC (116, 117). However, the results are inconsistent, which may be caused by limitations, such as small sample size, inconsistent measurement methods, influence of the gender, tumor stage, and tumor heterogeneity. Hence, further studies are needed in multi-centers with a larger population, standardized measurement methods, paired samples, and detailed analysis for different stages and pathological types of LC. Currently, some metabolites, enzyme inhibitors targeting immune checkpoints, and modulators have been developed for the diagnosis and treatment of LC. Because the change in metabolomics is one of the factors for the development of cancer, it will be wise to integrate the role of metabolomic changes in the pathogenesis of LC and consider other factors together for the development of therapeutic strategies for LC. Therefore, further studies are necessary to understand the process of complicated Try metabolism and its regulation in different types and stages of LC.

CL performed the literature search and drafted the manuscript and figures. HZ edited and revised the manuscript. All authors contributed to the article and approved the submitted version.

This work was supported by grants from the National Natural Science Foundation of China 81703087, the United Fund of the Second Hospital of Dalian Medical University and Dalian Institute of Chemical Physics, Chinese Academy of Sciences (UF-ZD-202011), and the Project of Education Department of Liaoning Province (LZ2020009).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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