Targeting tryptophan metabolism in breast cancer immunotherapy: Recent advances and future prospects

Highlights

• •The Trp-Kyn-AhR axis drives immune evasion and "cold" phenotypes in breast cancer.
• •Elevated IDO1 and TDO2 levels are linked to poor prognosis in TNBC and HER2+ breast cancer.
• •Compensatory TDO2 upregulation contributes to the clinical failure of IDO1 inhibitors.
• •Nanomedicine and dual-target inhibitors offer new ways to overcome metabolic barriers.
• •Biomarker-driven stratification is essential for precision metabolic immunotherapy.

Abstract

Breast cancer (BC), a leading cause of cancer-related mortality in women, is predominantly characterized as an immunologically “cold” malignancy. This recalcitrant nature is largely attributed to its profound metabolic landscape, which orchestrates a hostile tumor microenvironment (TME). Central to this metabolic subversion is the kynurenine (Kyn) pathway of tryptophan (Trp) catabolism. Driven by the rate-limiting enzymes indoleamine 2,3-dioxygenase 1/2 (IDO1/2) and tryptophan 2,3-dioxygenase (TDO2), this axis functions as a critical molecular rheostat that promotes immune evasion by depleting essential Trp and accumulating bioactive Kyn metabolites. This review provides a comprehensive analysis of the molecular basis by which the Trp-Kyn-aryl hydrocarbon receptor (AhR) signaling axis impairs T-cell effector function, induces regulatory T-cell (Treg) differentiation, and modulates the plasticity of myeloid-derived suppressor cells (MDSCs) specifically within the BC context. We critically evaluate the clinical trajectory of first-generation IDO1 inhibitors, analyzing the biochemical and compensatory mechanisms—such as TDO2 upregulation—that contributed to recent clinical setbacks. Furthermore, we highlight emerging strategies, including dual IDO1/TDO2 inhibitors, AhR antagonists, and nanomedicine-based delivery systems designed to overcome metabolic barriers. By emphasizing the integration of Trp-targeted agents with immune checkpoint blockade and other conventional therapies, we propose a framework for biomarker-driven patient stratification. Ultimately, we outline future directions to transition from "one-size-fits-all" approaches toward precision metabolic immunotherapy to unlock robust anti-tumor immunity in breast cancer.

Graphical abstract

Introduction

Breast cancer (BC) is the most prevalent malignancy and a leading cause of cancer-related mortality among women worldwide [1]. Conventionally considered as an immunologically “cold” malignancy [2], BC is characterized by scarce infiltration of cytotoxic T lymphocytes [3], abundant immunosuppressive cells and limited expression of immune checkpoint molecules within TME [4]. This "cold" phenotype is a fundamental reason for the poor response of breast cancer to immunotherapy. Its etiology is complex and closely related to the intrinsic biological characteristics of breast cancer. First, breast cancer has a relatively low tumor mutational burden (TMB) [5], resulting in insufficient neoantigen generation and difficulty in eliciting a robust primary T-cell response. Second, breast cancer exhibits high heterogeneity in primary and metastatic disease [6] and molecular subtypes, with four major molecular subtypes including Luminal A [7], Luminal B [8], Human epidermal growth factor receptor 2(HER2)-enriched [9] and TNBC [10], displaying distinct immune microenvironments. Although TNBC typically shows higher lymphocyte infiltration and PD-L1 expression, making it relatively “hotter” [11]. Furthermore, BC can actively establish an immunosuppressive environment through various mechanisms, such as recruiting regulatory T cells (Tregs) [12] , MDSCs [13] and upregulating immunosuppressive metabolic pathways including Trp-Kyn pathway, leading to unchecked immune evasion and ultimately driving tumor progression [14]. Therefore, strategies to effectively inflame the TME and convert immunologically ignorant or excluded tumors into T cell-inflamed, “hot” tumors represent a central pursuit in BC immunotherapy research.

Tumor cells undergo extensive metabolic reprogramming to sustain rapid proliferation, a hallmark known as the “Warburg effect” [15,16]. Beyond fueling anabolic growth, this altered metabolism actively shapes the immune landscape of the TME. Tumor cells compete with immune cells for critical nutrients such as glucose, amino acids [17] (e.g., glutamine, tryptophan), and oxygen, creating a metabolically hostile milieu. This competition leads to T cell exhaustion and dysfunction. Furthermore, the accumulation of specific immunosuppressive metabolites including lactate [18], adenosine [19] and Kyn [20] directly inhibits effector immune cell function while promoting the expansion and activity of regulatory immune populations, such as regulatory Tregs and MDSCs. Thus, tumor metabolic pathways constitute a powerful non-genetic mechanism of immune evasion.

Among various metabolic axes, Trp metabolism is pivotal in tumor immunology. Trp metabolism in human body is primarily catabolized via three major pathways: Kyn, 5-hydroxytryptophan(5-HT) and indole pathways [21]. The Kyn pathway, initiated by the rate-limiting enzymes IDO1, IDO2, and TDO, is frequently overactivated and represents the predominant metabolic pathway in cancer, contributing an immunosuppressive TME [22]. IDO1, in particular, is a focal point in cancer immunotherapy research due to its potent immunosuppressive effects. Its activity depletes local Trp and accumulates Kyn and other downstream metabolites, engaging a dual mechanism that suppresses effector T(Teff) and natural killer (NK) cells while activating immunosuppressive AhR signaling pathways. This establishes the Trp-Kyn pathway as a master regulator of immune tolerance within the TME.

This review aims to comprehensively elucidate how targeting the Trp-Kyn metabolic pathway can potentiate BC immunotherapy. We summarize recent mechanistic insights into how this axis shapes the BC immune microenvironment and critically evaluate preclinical and clinical developments in pharmacological strategies targeting IDO1/TDO and AhR. Finally, we discuss translational challenges, analyze the lessons learned from past trials, and outline future opportunities for integrating metabolic modulation with existing immunotherapeutic regimens to improve outcomes for BC patients.

The Trp-Kyn pathway: mechanisms of immunosuppression in breast cancer

Key metabolic enzymes and their regulation

Trp

Trp, an essential amino acid, is not endogenously synthesized but exclusively obtained from the diet for protein synthesis in humans [21]. >95% of free Trp is metabolized via Kyn pathway, primarily mediated by three different enzymes IDO1 [23], IDO2 [24] or TDO [25], while a minor fraction of free Trp can be utilized for protein and neurotransmitters synthesis [26]. Recent studies have highlighted the dual immunoregulatory roles of Trp metabolism in BC microenvironment. For instance, macrophages exhibit the highest Trp metabolic activity among immune cells; while enhanced Trp metabolism promotes anti-tumor M1 polarization, it simultaneously generates metabolites that suppress Teff cell functions [27]. Notably, high levels of Trp metabolism have been associated with increased immune cell infiltration, particularly CD8+ T cells and enhanced cytolytic activity, highlighting a complex balance between immune activation and suppression [27]. Furthermore, spatial transcriptome analyses reveals elevated macrophage Trp metabolism is associated with better responses to BC immunotherapy, indicating its potential as a predictive biomarker for immunotherapy efficacy in breast cancer. Despite these promising findings, the immunotherapeutic landscape for BC remains fraught with significant challenges, including primary/acquired resistance, limited efficacy and immune evasion [28]. Consequently, understanding the precise immunoregulatory mechanisms of Trp metabolism is essential for developing novel therapeutic approaches.

IDO

IDO, comprising IDO1 and IDO2 [29],is an immune checkpoint marker expressed in tumor and infiltrating immune cells [30,31]that promotes immune escape and exerts immunosuppressive effects in BC [32]. Notably, IDO1 is highly expressed in TNBC and HER2-positive subtypes, but unaltered in luminal breast cancer [31] becauselower IDO1 promoter methylation in TNBC heightens its responsiveness to immune pressure. Mechanistically, CD8+ T cells-derived IFNγ in triggers the JAK/STAT1/IRF1 pathway in TNBC cells, upregulating IDO1 expression to facilitate immune evasion [33]. Similarly, NK cells-derived IFN-γ induces high IDO1 expression in TNBC cells, which also promotes HLA-G expression and subsequently inhibits the anti-tumor activity of NK cells in vivo [34]. Another research reveals that transcription factor IRF5 is highly expressed in TNBC, where it directly drives Trp metabolic reprogramming in tumor cells by transcriptionally activating SLC7A5 (a Trp transporter) and IDO1. Isoliquiritigenin (ISL) has been shown to downregulate IRF5, thereby suppressing the expression of SLC7A5 and IDO1. This effectively blocks Trp metabolism and disrupts the IDO1-mediated immunosuppressive pathway, demonstrating potential immunopotentiating effects [35]. Beyond its cellular source, interstitial IDO regulation within the tumor microenvironment further influences BC progression. For instance, prostaglandin E2 (PGE2) released from COX-2-overexpressing BC cells induces IDO expression in human mammary fibroblasts, activating the Kyn/AhR axis to degrade E-cadherin and enhance tumor invasiveness [36]. Thus, it is indicated that IDO1 inhibitor-based immunotherapy may represent a promising BC therapeutic strategy.

IDO2, also named indoleamine 2,3-dioxygenase-like-protein1(INDOL1), was firstly discovered by Ball et al. [37]. As a paralog of IDO1, IDO2 has negligible catalytic activity in Kyn pathway due to extremely low substrate affinity and catalytic efficiency, essentially lacking functional activity in Kyn synthesis [38,39]. While less frequently overexpressed than IDO1 [40], IDO2 is constitutively co-expressed in several cancers including gastric, colorectal, renal, and brain carcinomas [41,42] and notably overexpressed in pancreatic ductal adenocarcinoma (PDAC) [43] and non-small-cell lung cancer (NSCLC) [44]. In PDAC, IDO2-inactivating single-nucleotide polymorphisms (SNPs) correlates significantly with enhanced disease-free survival (DFS) following adjuvant radiotherapy, making IDO2 genotype a valuable stratification biomarker to guide PDAC treatment decisions [45]. Additionally, IDO2 involvement in PDAC exhibits sexual dimorphism: Ido2⁻/⁻ female mice develop less PDAC, and correspondinglyIDO2 deficiency is rare in female patients [45]. Thus, it is suggested that females are prime candidates for IDO2-targeted therapy. Most notably, a significant positive correlation was identified between high IDO2 expression, high PD-L1 levels and unfavorable prognosis in patients with NSCLC [44]. In BC, IDO2 expression increased relative to normal samples in TCGA and GTEx datasets, and AhR ligands can significantly induce IDO2 expression in MCF-7 cells in an AhR-dependent manner, indicating a tumor-promoting role of IDO2 [46]. However, the precise mechanisms of IDO2 in immune suppression and facilitating tumor progression remain incompletely defined [24], necessitating further research to optimize safe and effective IDO2-targeted immunotherapies.

TDO

Another key component of Kyn pathway dysregulated in BC is TDO. Expression of the TDO2 gene, which encodes the TDO enzyme, is significantly elevated in HER2-positive and TNBC subtypes [47,48]. Furthermore, TDO2 levels are also notably higher in ER-negative than ER-positive patients. Clinically, increased TDO2 correlates with advanced tumor grade, increased malignancy and reduced survival rates [47], making it a robust prognostic biomarker. Mechanistically, TDO-driven Kyn overproduction activates the AhR receptor, promoting BC cell migration and apoptosis resistance [49,50]. Specifically, in TNBC, activation of the NFκB/TDO/Kyn/AhR signaling axis increases resistance to anoikis (detachment-induced apoptosis) in vitro [48]. Conversely, inhibiting this axis suppresses proliferation, migration and invasion. These findings identify the TDO/Kyn/AhR pathway as a promising therapeutic strategy to impede BC progression.

Moreover, TDO2 has also emerged as both a driver of tumor aggressiveness and a potential negative prognostic biomarker, with elevated expression correlating with higher tumor grade and malignancy. In one study, TDO2 levels were reported to correlate with the degree of malignancy and tumor grading in BC, indicating that higher TDO2 expression could serve as a biomarker for poor prognosis [51]. Another study proposes that TDO2-mediated Trp maturation suppresses Tertiary Lymphoid Structure (TLS) maturation by dampening CXCL9 expression and disrupting B cell class switch recombination, thereby facilitating tumor immune evasion [52]. This dual role underscores TDO’s significance beyond metabolism, positioning it as a target for therapies aimed at restoring anti-tumor immunity. The differential expression patterns of IDO1 and TDO2 across BC subtypes underscore the complexity of the Kyn pathway and its therapeutic implications, particularly for immunotherapy [53]. As such, targeting these enzymes could yield a dual benefit: reversing immunosuppression while simultaneously reactivating anti-tumor immunity, particularly in aggressive subtypes like TNBC, where conventional therapies often prove inadequate.

Collectively, the expression profiles of IDO1, IDO2 and TDO2 in BC provide profound insights into tumor biology and patient prognosis. Their upregulation reflects a shift toward a more malignant phenotype and highlights promising therapeutic targets, particularly for immune-based therapies.

Kyn and its immune regulatory role in breast cancer immunotherapy

Kyn, a key component of Trp metabolites, plays a pivotal role in modulating cancer immune responses [54]. Recent findings have illuminated an unforeseen function of the ligand-activated transcription factor AhR in mediating the tumor immune evasion associated with IDO1 and TDO2. Specifically, Kyn-induced AhR activation drives the development of immune-tolerant dendritic cells (DCs) and Tregs fostering an immunosuppressive TME incapable of eliminating malignant cells [54,55]. Moreover, AhR activation is crucial as it influences the expression of immune suppressive genes, thereby facilitating tumor immune evasion [56]. Kyn and its derivatives, such as kynurenic acid, have been implicated in promoting Treg differentiation, which suppresses Teff cell activity and contributes to an immunosuppressive tumor microenvironment. In breast cancer, elevated Kyn levels correlate with increased Treg infiltration and poorer clinical outcomes. Additionally, Kyn directly inhibits the proliferation and cytotoxic activity of CD8⁺ T cells, further dampening anti-tumor immunity. The immunosuppressive effects of Kyn are further amplified by modulating various signaling pathways. For instance, Kyn can enhance the expression of PD-1 on T cells [34]. This upregulation of PD-1, combined with AhR-mediated increases in IDO1 expression, establishes a self-reinforcing feedback loop that augments Kyn production and perpetuates immune evasion [57].

The impact of Kyn on the tumor immune microenvironment extends to innate immune cells like NK cells, which constitute a vital element of the innate immune system and perform the crucial function of immune surveillance against tumors [58]. In gastric cancer, tumor-derived L-Kyn induces ferroptosis in NK cells in an AHR-independent manner, depleting them from the TME and impairing immune surveillance [59]. Consequently, therapeutic interventions targeting this pathway show significant promise. For instance, a dual-pronged “Kynurenine Starvation Therapy” could target Trp-Kyn-AhR axis and lower Kyn/Trp ratio, successfully inhibiting the postoperative breast cancer recurrence and metastasis by reversing the immunosuppressive TME in TNBC [60].

In conclusion, Kyn and its metabolites exert profound immunoregulatory effects that facilitate immune escape in breast cancer. Targeting these mechanisms remains essential for reversing immunosuppression and restoring anti-tumor immunity.

Mechanisms of immunosuppression in tryptophan metabolism

The association between macrophage M1/M2 polarization and tryptophan metabolism

Macrophage polarization is a key determinant of clinical prognosis in BC immunotherapy [61,62]. Generally, M1 macrophages are recognized as effector cells that defend the body against pathogens and tumor cells. In contrast, M2 macrophages function to suppress inflammatory responses and adaptive immunity, while promoting angiogenesis and tumor growth [63]. Trp metabolism correlates with M1 polarization and is primarily linked to the immune microenviroment rather than the stroma. Specifically, high Trp metabolism group enriched immune-activating cells (e.g., M1 macrophages, CD8+ T cells, activated CD4+ memory T cells, follicular helper T cells, and γδ T cells) while depleting multiple immunosuppressive populations (e.g., M2 macrophages, resting CD4+ memory T cells, naïve B cells, and resting mast cells). Consequently, macrophage Trp metabolism may serve as a predictive biomarker for BC immunotherapy response [27], suggesting that targeting tryptophan-metabolizing pathways may represent a potential strategy to manipulate macrophage polarization, promoting an M1 phenotype and thereby strengthening anti-tumor immune responses.

However, the specific role of IDO in macrophage polarization remains controversial. Previous findings have demonstrated that IDO induce the differentiation of macrophages into M2 rather than M1 [64], supporting the counter-regulatory role for IDO to restrain excessive or inappropriate immune activation in inflammatory and tumor microenvironment. Conversely, IDO promotes M1 polarization in non-tumor diseases, such as ulcerative colitis (via ER stress-associated GRP78-XBP1 pathway) [65] and Fungal Keratitis (by blocking CCL2/CCR2 signaling pathway) [66].Taken together, these results fully demonstrate that IDO’s regulatory effects on macrophage polarization exhibit significant differences between tumor and non-tumor diseases, probably as well as across different types of tumors.

Interaction between tryptophan metabolism and immunosuppressive microenvironment: Tregs, MDSCs, PD-1/PD-L1 and CTLA-4

Trp metabolism drives the tumor-associated immunosuppressive microenvironment primarily by elevating Kyn, which impairs Teff cell function and promote the activity of FOXP3+ Tregs and MDSCs. Thus, the increased Treg cell population mediates direct suppression of Teff cells, which in turn compromises the overall anti-tumor immune response [41]. For instance, in colorectal cancer, Kyn upregulates FOXP3 and hinders the activity of Teff cells [67]. In renal cell carcinomas (RCC), PABPC1L induces IDO expression to enhance Trp metabolism and consequently lead to T-cell dysfunction and Treg infiltration, representing a promising target to strengthen the efficacy of immune checkpoint inhibitors (ICIs) therapy [68]. However, the correlation between Trp metabolism-mediated Treg infiltration in BC immunotherapy has rarely been reported.

Furthermore, Trp metabolism shapes MDSC populations. In cisplatin-resistant (CR) non-small cell lung cancer, IDO1-mediated Kyn suppresses the natural killer group 2 member D (NKG2D) receptor on NK cells and CD8⁺ T cells—while simultaneously promoting the expansion of immunosuppressive cell populations Tregs and MDSCs [69]. Importantly, kynurenine also upregulates the expression of immune checkpoint molecules such as PD-L1. Consequently, combining the dual IDO1/TDO2 inhibitor AT-0174 with PD-1 blockade enhances anti-tumor immunity [69].

Clinically, Trp metabolism may predict the efficacy of ICIs in BC patients, stronger baseline Trp metabolic activity predicts PD-1 blockade response, with a notable area under the ROC curve of 73.8% [27]. Additionally, the interplay between gut microbiota and Trp metabolism has been highlighted as a novel biomarker to predict the response to anti-PD-1 immunotherapy. For instance, ginseng polysaccharides (GPs) potentiated the antitumor efficacy of anti-PD-1 immunotherapy by increasing microbial metabolite valeric acid and reducing l-kynurenine levels and the Kyn/Trp ratio. These metabolic changes contributed to the suppression of Tregs and promoted the induction of Teff cells following the combination therapy [70].

Beyond PD-1, IDO impairs the efficacy of anti-CTLA-4 immunotherapy by suppressing Teff cell function and promoting Treg accumulation in melanoma. Notably, the combination of IDO inhibitors (e.g., 1-MT) with CTLA-4 blockade ( e.g., ipilimumab) can significantly enhance antitumor immune responses, even in IDO-negative tumors. Since IDO deficiency also improves anti-PD-1/PD-L1 and anti-GITR therapies, indicating that the immunosuppressive effect of IDO is not limited to the CTLA-4 pathway [71]. Thus, this finding provides a strong rationale for combination therapies with IDO inhibitors and immune checkpoint blockade.

Overall, the interplay between Trp metabolism, Tregs, MDSCs and immune checkpoint blockade drives tumor immune evasion. Targeting this metabolic pathway can enhance immunotherapy efficacy, particularly in cancers characterized by high levels of Trp catabolism and associated immunosuppressive cell populations. However, how Trp metabolism shapes the immunosuppressive microenviroment and impacts immunotherapy outcomes in BCremains to be thoroughly investigated.

Therapeutic strategies targeting tryptophan metabolism

The combination of metabolic enzyme inhibitors targeting IDO1and/or TDO with immune checkpoint inhibitors such as PD-1/PD-L1 or CTLA-4 antibodies has shown promising synergistic antitumor effects in preclinical studies [[71], [72], [73]].

IDO and/or TDO inhibitors and combination therapy

Indoximod, as a novel IDO inhibitor, suppresses TNBC cell viability via apoptosis and G0/G1 phase arrest [74]. Pro-inflammatory cytokines can modulate the efficacy of indoximod, with TNF-α enhancing its cytotoxic and pro-apoptotic effects, while IFN-γ attenuates them. Furthermore, the combination of indoximod with cytokines upregulates PD-L1 expression, suggesting a promising therapeutic strategy for combination of IDO and PD-L1 inhibition. More remarkably, a recent study has reported that addition of indoximod to radiotherapy and anti-PD1 therapy strongly shrink aggressive tumors by synergistically activating both innate and adaptive immunity, with particularly pronounced effects during the early treatment phase, indicating the significant value of this triple combination regimen [75]. Consequently, further research is needed to elucidate the molecular mechanisms of indoximod-induced apoptosis and unravel the complex interplay with cytokines and combination therapies such as radiotherapy.

Epacadostat (INCB24360) is an oral, selective IDO1 inhibitor by competitively binding to the catalytic domain of IDO1 and blocking Trp utilization [76]. While phase I trial of Epacadostat in patients with advanced solid malignancies showed that doses≥100 mg twice daily reduced plasma Kyn levels by 80–90%, no objective responses or immune biomarker alterations were observed when Epacadostat was administered as a monotherapy [77]. Moreover, in the ECHO-301/KEYNOTE-252 phase III trial, adding Epacadostat (100 mg twice daily) to pembrolizumab failed to improve progression‑free survival (PFS) (median PFS 4.7 vs. 4.9 months; HR 1.00, P = 0.52) or overall survival (OS) (median OS not reached; HR 1.13, P = 0.81) in patients with unresectable or metastatic melanoma [78]. Therefore, identifying the optimal dosing of Epacadostat, suitable patient population, and required intratumoral Kyn suppression thresholds remains critical for future investigations.

Navoximod (GDC-0919, previously NLG919) is an orally bioavailable small-molecule IDO1 inhibitor that blocks T-cell suppression and restores immune function by lowering plasma and tissue Kyn levels [79]. In a phase I clinical trial, the combination of Navoximod and atezolizumab demonstrated acceptable safety, tolerability, and PK profiles in advanced cancers including TNBC but failed to provide additional therapeutic benefit [80]. Furthermore, another phase I study in Japanese patients with advanced solid tumors confirmed that both Navoximod monotherapy and its combination with atezolizumab were well-tolerated, although early termination of the study prevented formal determination of a recommended combination dose [81].

NPACT00380 (Calyxins E) is a potential dual inhibitor of IDO1/TDO, whose chromone moiety contributes to its anti-tumor activity. Computational screening and kinetic simulations demonstrate its excellent binding stability and favorable drug-like properties, making it a promising candidate for TNBC treatment [82]. However,further experimental validation of its inhibitory activity and clinical efficacy is warranted.

Linrodostat (BMS-986,205 or F001287) is a selective, once-daily oral IDO1 inhibitor that binds to the heme cofactor–binding site, blocking activation of the IDO1 pathway and reducing Kyn production [83]. In a phase 1/2 trial (ClinicalTrials.gov identifier: NCT02658890) evaluating Linrodostat plus nivolumab in advanced solid tumors or hematologic malignancies, the combination therapy revealed a favorable safety profile in heavily pretreated patients and initial antitumor activity in advanced cervical or bladder cancers. Furthermore, another phase 1/2 study of Linrodostat combined with Nivolumab ± Ipilimumab in advanced solid tumors or hematologic malignancies showed a manageable safety profile. Although reductions in Kyn levels confirmed effective IDO1 inhibition, these changes did not correlate with clinical response. A composite biomarker profile—low TDO expression together with high IFN-γ gene expression—may predict response to Linrodostat plus Nivolumab [84].

EOS200271/PF-06,840,003 is a highly selective, non-heme iron binding IDO1 inhibitor [85] with proven antitumor efficacy. In a phase 1 clinical trial, at doses up to 500 mg BID, PF‑06,840,003 was safe and showed durable clinical benefits in recurrent malignant glioma [86]. IDO1 serves as a critical resistance mechanism to PD-(L)1 block therapy, indicating the synergistic antitumor effect of combination of immunotherapy with PF‑06,840,003. Remarkably, its ability to penetrate the blood-brain barrier makes it a promising treatment candidate for brain metastases and glioblastoma [87]. Other representative clinical studies of IDO and/or TDO inhibitors and AhR antagonists and combination therapy in solid tumors are summarized in Table 1.

Table 1.

Representative clinical studies of IDO and/or TDO inhibitors and AhR antagonists and combination therapy in solid tumors.

Trail ID	Disease type	Drug(s)	Target(s)	Phase	Recruitment Status	Intervention/Treatment
NCT01792050	Metastatic Breast Cancer	Indoximod	IDO	II	Completed	in Combination With a Taxane Chemotherapy (Docetaxel or Paclitaxel)
NCT02048709	Advanced Solid Tumors	GDC-0919	IDO1	I	Completed	GDC-0919
NCT04106414	Recurrent or Persistent Endometrial Cancer or Endometrial Carcinosarcomas	BMS- 986,205	IDO1	II	Active, not recruiting	Nivolumab vs Nivolumab+BMS-986,205
NCT03164603	Advanced Solid Tumors	NLG802	IDO1	I	Completed	NLG8021
NCT03915405	Advanced Bladder Cancer	KHK2455	IDO	I	Terminated	in Combination With Avelumab
NCT02460367	Advanced Previously Treated Non-Small Cell Lung Cancer	Indoximod	IDO	Ib/II	Terminated	in Combination With Tergenpumatucel-L immunotherapy and Docetaxel
NCT00739609	Relapsed or Refractory Solid Tumors	D-1MT	IDO	I	Terminated	D-1MT
NCT01685255	Biochemical-Recurrent-Only Epithelial Ovarian Cancer, Primary Peritoneal Carcinoma, or Fallopian Tube Cancer Following Complete Remission With First-Line Chemotherapy	Epacadostat	IDO	II	Terminated	Epacadostat vs Tamoxifen
NCT03491631	Advanced Solid Tumors	SHR9146	IDO	I	Unknown status	SHR9146+SHR-1210 vs SHR9146+ SHR1210+ Apatinib
NCT02073123	Metastatic Melanoma	Indoximod	IDO	I/II	Completed	Indoximod Plus Immune Checkpoint Inhibitors(Ipilimumab/Pembrolizumab/Nivolumab)
NCT02052648	Primary Malignant Brain Tumors	Indoximod	IDO	I/II	Completed	Temozolomide, Bevacizumab And Stereotactic Radiation
NCT02077881	Metastatic Pancreatic Cancer	Indoximod	IDO	I/II	Completed	in Combination With Gemcitabine and Nab-Paclitaxel
NCT02502708	Pediatric Patients With Progressive Primary Malignant Brain Tumors	Indoximod	IDO	I	Completed	Indoximod and Temozolomide-Based Therapy (Temozolomide, Cyclophosphamide,Etoposide and Conformal Radiation)
NCT03343613	Solid Tumors	LY3381916	IDO-1	Ia/Ib	Terminated	LY3381916 vs LY3381916+LY3300054
NCT03361865	Cisplatin-ineligible Urothelial Carcinoma	Epacadostat	IDO	III	Completed	Pembrolizumab
NCT03374488	Recurrent or Progressive Metastatic Urothelial Carcinoma	Epacadostat	IDO	III	Completed	Pembrolizumab + Epacadostat vs Pembrolizumab + Placebo
NCT03348904	Non-Small Cell Lung Cancer	Epacadostat	IDO	III	Terminated	Nivolumab and Epacadostat With Platinum Doublet Chemotherapy VS.Platinum Doublet Chemotherapy(Pemetrexed/ Paclitaxel/ Gemcitabine+ Cisplatin/ Carboplatin)
NCT03085914	Advanced or Metastatic Solid Tumors	Epacadostat	IDO	I/II	Completed	Epacadostat in Combination With Pembrolizumab and Chemotherapy
NCT03342352	Squamous Cell Carcinoma of the Head and Neck	Epacadostat	IDO	III	Withdrawn	Nivolumab Plus Epacadostat in Combination With Chemotherapy (Platinum + 5-fluorouracil) vs the EXTREME Regimen (Cetuximab + Platinum + 5-fluorouracil)
NCT03347123	Advanced or Metastatic Malignancies	Epacadostat	IDO	I/II	Terminated	Epacadostat and Nivolumab in Combination With Immune Therapies(Ipilimumab/ Lirilumab)
NCT03414229	Metastatic and/​or Locally Advanced Sarcoma	Epacadostat	IDO	II	Active, not recruiting	Epacadostat in Combination With Pembrolizumab
NCT01961115	Stage III-IV Melanoma	Epacadostat	IDO	II	Completed	Epacadostat Plus a Multipeptide Melanoma Vaccine (MELITAC 12.1)
NCT01982487	Epithelial Ovarian, Fallopian Tube, or Primary Peritoneal Cancer Who Are in Remission	INCB024360	IDO1	I/IIb	Withdrawn	Recombinant ALVAC(2)-NY-ESO-1 (M)/TRICOM in Combination With INCB024360
NCT03432676	Advanced Pancreatic Cancer	Epacadostat	IDO	II	Withdrawn	Epacadostat in Combination With Pembrolizumab
NCT02166905	Ovarian, Fallopian Tube, or Primary Peritoneal Cancer in Remission	INCB024360	IDO1	I/IIb	Completed	INCB024360 in combination with DEC-205/NY-ESO-1 fusion protein CDX-1401 and poly ICLC
NCT03459222	Solid Cancers That Are Advanced or Have Spread	BMS-986,205	IDO1	I/II	Completed	Relatlimab Administered in Combination With Both Nivolumab and BMS-986,205 or in Combination With Both Nivolumab and Ipilimumab
NCT03260894	locally advanced/metastatic renal cell carcinoma	Epacadostat	IDO	III	Completed	Pembrolizumab Plus Epacadostat vs standard of Care (Sunitinib or Pazopanib)
NCT03322540	Metastatic Non-Small Cell Lung Cancer	Epacadostat	IDO	II	Completed	Pembrolizumab Plus Epacadostat vs Pembrolizumab Plus Placebo
NCT03358472	Recurrent or Metastatic Head and Neck Squamous Cell Carcinoma	Epacadostat	IDO	III	Completed	pembrolizumab plus epacadostat, pembrolizumab monotherapy, and the EXTREME regimen (cetuximab + cisplatin or carboplatin + 5-fluorouracil)
NCT03322566	Metastatic Non-Small Cell Lung Cancer	Epacadostat	IDO	II	Completed	pembrolizumab plus epacadostat with platinum-based chemotherapy vs pembrolizumab plus platinum-based chemotherapy plus placebo
NCT05472506	Resistant Metastatic or Recurrent Head and Neck Cancer	IK-175	AHR	Ib	Withdrawn	IK-175 + nivolumab
NCT04200963	Locally Advanced or Metastatic Solid Tumors and Urothelial Carcinoma	IK-175	AHR	Ia/b	Completed	IK-175 + nivolumab

Nanomedicine-based combination therapy strategies

Nanomedicines has revolutionized cancer therapeutic approaches by overcoming conventional limitations such as systemic toxicity and inadequate targeting of tumor sites [88]. Nanomedicines have garnered growing interest due to their enhanced tumor targeting capability and favorable biocompatibility [89]. Moreover, by delivering diverse therapeutic agents, nanomedicines enable multimodal synergistic treatment, which holds promise for improving therapeutic outcomes [90]. Importantly, preclinical studies have demonstrated that nanomedicines incorporating IDO1 inhibitors in combination with other treatment modalities such as chemotherapy [91], radiotherapy [92], photodynamic therapy (PDT) [93], and photothermal therapy (PTT) [94] can elicit synergistic antitumor immune responses.1) Chemotherapy: Co-delivery with chemotherapeutic agents like oxaliplatin, doxorubicin [95], or paclitaxel, nanomedicines can promote immunogenic cell death (ICD), thereby enhancing antitumor immunity [96].2) Radiotherapy [97]: Nanomedicines combined with radiotherapy can induce DNA damage and reactive oxygen species (ROS) generation to kill tumor cells and stimulate immune activation, while IDO inhibitors counteract potential immunosuppressive effects triggered by radiation [98].3) PDT: Co‑delivering photosensitizers generates ROS upon light irradiation to induce ICD, while simultaneously IDO inhibition potentiates the immune response [99].4) PTT: Combining nanomedicines with photothermal agents (e.g., gold nanorods, graphene, or organic dyes) leads to thermal ablation‑induced ICD, and concurrent IDO inhibition reverses the immunosuppressive tumor microenvironment [89]. Despite setbacks in Phase III clinical trials (e.g., Epacadostat), IDO nanoformulations still hold promise for future breakthroughs through optimized dosing regimens, novel combination strategies, patient subgroup, and advanced delivery systems. Future research should focus on combining IDO inhibitors with anti-angiogenic agents, developing comprehensive IDO2/TDO inhibitors, designing highly targeted nanocarriers, and identifying predictive biomarkers for patient selection.

Challenges and perspectives

Metabolic heterogeneity and patient stratification strategies

Metabolomics reveals significant heterogeneity in Trp metabolism across breast cancer subtypes, enabling the identification of metabolic biomarkers for patient stratification. These biomarkers, such as N-acetyl-d-tryptophan, can differentiate subtypes (e.g., TNBC vs. non-TNBC) and predict immunotherapy response. Integrating metabolomic data with clinical parameters and machine learning refines stratification and prognostic models, facilitating personalized treatment strategies and improving outcomes.

Systematic biological analysis of metabolic-immune interaction networks

Systems biology and multi-omics approaches elucidate the complex crosstalk between metabolic pathways (especially Trp metabolism) and immune signaling in the TME. Key findings include the association of enhanced Trp metabolism with pro-inflammatory macrophage polarization and the correlation of metabolic states (e.g., hypoxia) with immune evasion. Integrating metabolic and immune data into dynamic models helps predict therapy response and resistance, guiding the development of novel combination strategies.

Clinical translation and exploration of multi-target combination therapy

Optimizing metabolic inhibitors (e.g., targeting IDO1) and combining them with other modalities is a key translational strategy. This includes pairing metabolic modulators with immune checkpoint inhibitors and gut microbiota interventions (e.g., using fucoidan to enhance anti-PD-1 efficacy). Addressing factors like circadian rhythm disruption, which upregulates immunosuppressive Trp pathways, is also important. Successful clinical translation requires rigorous evaluation and biomarker development to personalize these multi-target therapies for improved patient outcomes.

Limitations in current research

Setbacks and limitations in clinical trials

As noted, first-generation IDO1 inhibitor Epacadostat failed to improve PFS or OS in ECHO-301/KEYNOTE-252 clinical trial. This indicates the inherent limitations of a simple, global enzyme inhibition strategy. Furthermore, although Navoximod has demonstrated acceptable safety and pharmacokinetic profiles in clinical trials, there is currently no definitive evidence to suggest that adding them to ICIs provides additional therapeutic benefits. More importantly, despite observed reductions in Kyn levels in Epacadostat clinical trials, these biochemical alterations frequently do not correlate with the clinical responses of patients, indicating the disconnection between biomarkers and clinical responses.

Complexity and controversy of mechanism research

Most clinical trials of IDO inhibitors have been unsuccessful to date, partly due to the overlooked compensatory redundancy between IDO1 and TDO2, as well as an incomplete understanding of systemic versus local Trp buffering capacities. Furthermore, the role of IDO in macrophage polarization remains controversial. While some studies suggest that IDO induces M2 polarization, others have observed the opposite effect (M1 polarization) in different disease contexts, indicating that its mechanisms vary significantly across different tumor types and non-malignant diseases. Additionally, although IDO2 is overexpressed in certain cancers, its precise role in mediating immunosuppression and driving tumor progression remains to be fully elucidated.

Challenges in translational research

Although associations between Trp metabolism and immunosuppression have been observed in other malignancies (e.g., colorectal cancer, renal cell carcinoma, and non-small cell lung cancer), studies specifically investigating the correlation between Trp metabolism-mediated Treg infiltration and immunotherapy efficacy in breast cancer remain scarce. Furthermore, current therapeutic approaches predominantly rely on a "one-size-fits-all" paradigm, lacking precise patient stratification based on specific Trp catabolic profiles. More importantly, successful clinical translation necessitates addressing complex confounding factors, such as circadian rhythm disruptions (which can upregulate immunosuppressive Trp pathways), and requires more rigorous evaluation alongside robust biomarker development.

Urgent needs for future research

Firstly, given the limitations of single-enzyme inhibition, the development of dual-target IDO1/TDO2 antagonists is imperative for future interventions. Moreover, there is a critical need to design nanodrug delivery systems with enhanced tumor-targeting capabilities and biocompatibility to overcome systemic toxicity. Notably, it remains essential to further map the dynamic interactions between Trp metabolites and immune cell plasticity utilizing spatial metabolomics and high-resolution multi-omics technologies.

Conclusion and future perspectives

Fig.1 illustrates the roles of IDO1 and TDO2 in Trp metabolism and tumor immune suppression. The metabolic landscape of breast cancer, particularly in aggressive TNBC and HER2-positive subtypes, is increasingly recognized as a sophisticated molecular rheostat governed by the Trp-Kyn shunt. Beyond mere nutrient depletion, the IDO1/TDO2-Kyn-AhR signaling axis is a critical metabolic checkpoint that orchestrates a hostile, immune-excluded TME. By coupling biochemical flux with transcriptional reprogramming—exemplified by the GCN2-mediated arrest of T-cell translation and AhR-driven Treg differentiation—this pathway provides a robust non-genetic foundation for immune evasion and therapeutic resistance.

Fig. 1.

Schematic illustration of the roles of IDO1 and TDO2 in Trp metabolism and tumor immune suppression. IDO1 and TDO2 catalyze the conversion of Trp into Kyn, leading to the suppression of effector T cells (Teff), the induction of regulatory T cells (Treg), and the polarization of macrophages toward an M2-like phenotype, thereby promoting an immunosuppressive tumor microenvironment. Combination therapies targeting IDO1 or TDO2—such as enzyme inhibitors, photodynamic therapy (PDT), and drug delivery systems—can effectively enhance antitumor immunity.

Despite the initial pharmacological enthusiasm, the clinical trajectory of first-generation IDO1 inhibitors has underscored the formidable metabolic plasticity of breast cancer. Clinical trials’ failures (e.g., ECHO-301) highlight a critical oversights: the compensatory redundancy between IDO1 and TDO2, as well as the systemic-to-local Trp buffering capacity. Future breakthroughs require shifting from global enzymatic inhibition toward precision microenvironmental recalibration. This includes the development of dual-target antagonists, stimuli-responsive nanomedicines designed to bypass systemic toxicities, and "metabolic signatures" for patients stratification based on specific Trp catabolic profiles.

Looking ahead, integrating spatial metabolomics and high-resolution multi-omics is crucial to map the dynamic crosstalk between Trp metabolites and immune cell plasticity. Furthermore, the gut-breast axis—where the microbiome modulates circulating Trp metabolites—offers novel dietary or microbial interventions to sensitize "cold" breast tumors to immunotherapy. Ultimately, transitioning from a "one-size-fits-all" approach to a biomarker-driven, rationally integrated metabolic-immunotherapy paradigm holds the potential to fundamentally improve the survival and quality of life for patients facing this recalcitrant disease.

CRediT authorship contribution statement

Guangliang Li: Writing – original draft, Funding acquisition, Data curation. Huanhuan Zhou: Investigation, Data curation. Xiabo Shen: Methodology, Formal analysis, Data curation. Pingting Ying: Writing – review & editing, Supervision, Formal analysis. Haiqi Lu: Writing – review & editing, Supervision, Investigation, Funding acquisition, Conceptualization.

Declaration of competing interest

No conflict of interest exits in the preparation for this manuscript and manuscript is approved by all authors listed. Neither the entire paper nor any part of its content has been accepted or is under consideration for publication elsewhere.