Ammonia as a critical metabolic modulator of anti-tumor immunity

The landscape of the tumor microenvironment (TME) shapes immune responses through complex metabolic crosstalk. Ammonia is a waste byproduct of cellular metabolism, and recent work has demonstrated that ammonia concentrations can reach millimolar levels in the TME. Ammonia creates concentration gradients similar to other immunomodulatory metabolites such as lactate and kynurenine.1,2 This metabolic pattern suggests orchestrated immune suppression, where the balance between ammonia production and detoxification impacts cancer progression and treatment outcomes. Despite its emerging significance, the mechanisms driving ammonia accumulation, reutilization, and downstream effects are not fully mapped (Figure 1).Figure 1: Ammonia accumulation in the tumor microenvironment drives immunometabolic dysfunction and tumor progression.Schematic representation of ammonia (NH3) accumulation in the tumor microenvironment (TME) and its multifaceted effects on tumor progression and immune suppression. Elevated ammonia levels arise from increased glutamine metabolism through the tricarboxylic acid cycle and dysbiosis. This metabolic alteration leads to chronic inflammation, T cell exhaustion, and reduced effector functions. Ammonia accumulation also promotes cancer stemness through pH modulation and causes mitochondrial dysfunction. The dysregulated urea cycle further contributes to ammonia retention, resulting in impaired antigen presentation and reduced T cell proliferation. Additionally, the TME shows a shift towards M2-like macrophage polarization, collectively creating an immunosuppressive environment that favors tumor growth. Created with BioRender.com. αKG: Alpha-ketoglutaric acid; ROS: reactive oxygen species.Sources of ammonia in the tumor microenvironment: In colorectal cancer, ammonia metabolism is particularly complex due to contributions from both cellular metabolism and the gut microbiota. The colonic microenvironment features bacterial urease activity, which significantly elevates local ammonia levels. This bacterial-derived ammonia, combined with tumor cell-generated ammonia via glutamine metabolism, creates uniquely high concentrations in the colorectal TME compared to other cancer types. The sources of ammonia in the TME primarily include the catabolism of glutamine and aspartate, nucleic acid metabolism, and additional inputs resulting from systemic metabolic dysregulation. To meet the demands of rapid proliferation, tumor cells heavily rely on glutamine metabolism as a source of energy, carbon skeletons, and nitrogen donors. During this process, glutamine is converted into glutamate by glutaminase, with ammonia released as a byproduct. This pathway is particularly prominent in most tumors due to the dependency on glutamine.3,4 Furthermore, the synthesis and degradation of purines and pyrimidines also contribute to ammonia production, with the elevated metabolic activity of tumor cells exacerbating its accumulation. In some cases, systemic conditions such as liver dysfunction further amplify ammonia influx. Tumor cells can reutilize ammonia through glutamine synthetase, converting it back into glutamine to support protein synthesis and metabolic adaptation.5 However, dysregulation of the urea cycle impairs efficient ammonia clearance, leading to significant accumulation within the TME.6 Ammonia-mediated metabolic adaptation and tumor progression: Ammonia plays a central role in metabolic adaptation within the TME. Its reutilization enhances carbon and nitrogen metabolism, enabling tumor cells to maintain metabolic homeostasis and survive under conditions of glutamine deprivation. Recent evidence suggests that the role of ammonia role extends beyond basic metabolic adaptation to increase cancer stemness.7 High ammonia levels may be selected for cancer stem cell populations by activating stemness-associated pathways through metabolic reprogramming. These cancer stem cells exhibit an enhanced ability to utilize ammonia for amino acid synthesis and display increased resistance to metabolic stress. This mechanism is particularly relevant in colorectal cancer, where cancer stem cells occupy metabolically distinct niches along the crypt-villus axis. In addition to its effects on cancer stem cells, ammonia accumulation alters the local pH of the TME, creating a favorable environment for tumor cell survival. Elevated pH levels have been linked to increased tumor invasiveness and metastatic potential.8 Emerging evidence suggests that ammonia interacts with other metabolic byproducts, such as lactate and pyruvate, to establish complex pH and signaling gradients within the TME.9 Investigating these interactions may reveal new metabolic niches that promote tumor progression and therapeutic resistance. Ammonia-mediated immune suppression: Ammonia also profoundly influences immune cell function. By altering the metabolic environment, ammonia suppresses anti-tumor immune responses through altering CD8 T cell function.2 Additionally, ammonia may regulate key immunosuppressive pathways, such as the tryptophan-kynurenine axis. Kynurenine accumulation inhibits T cell function through activation of the aryl hydrocarbon receptor, and via competing metabolic fluxes. Emerging evidence suggests that ammonia synergizes with other immunosuppressive metabolites and immune cells. Ammonia alters lactate levels, which can disrupt mitochondrial function and promote T cell exhaustion. Furthermore, ammonia may impair antigen presentation in dendritic cells through pH-dependent mechanisms that disrupt protein folding and peptide loading in the endoplasmic reticulum. This interference could compromise T cell priming and anti-tumor immune responses. Ammonia has also been linked to the regulation of regulatory T cell differentiation and suppressive activity. By driving a nutrient-deprived and high-pH microenvironment, ammonia may preferentially support Treg survival while impairing effector T cell proliferation. Emerging technologies and future directions: Cutting-edge technologies are essential for unraveling the dynamic roles of ammonia in the TME. Spatial single-cell transcriptomics and metabolic imaging offer opportunities to visualize ammonia gradients and map their spatiotemporal distribution within tumor tissues. These tools are particularly relevant in colorectal cancer, where tracking the spatial relationship between bacterial communities and immune cell positioning could provide critical insights. CRISPR-based functional genomics screening offers a powerful approach to identifying genes and pathways that regulate ammonia production, reutilization, and clearance in tumor and immune cells. Genome-wide CRISPR knockout screens may uncover novel metabolic enzymes or transporters critical for ammonia homeostasis, revealing actionable therapeutic targets. Integrating CRISPR screens with single-cell multi-omics technologies will help resolve ammonia-driven heterogeneity and identify vulnerabilities unique to specific tumor or immune cell subpopulations. Therapeutic strategies targeting ammonia metabolism: Novel therapeutic approaches could exploit the unique features of ammonia metabolism in colorectal cancer. For instance, targeting bacterial urease activity in combination with inhibitors of cellular ammonia production may be particularly effective in this context. Additionally, timing therapeutic interventions to align with circadian patterns of ammonia metabolism could enhance efficacy while reducing systemic toxicity. Emerging approaches also include engineered probiotics designed to consume excess ammonia in the TME, potentially offering a tissue-specific approach to ammonia reduction. Given that ammonia accumulation in tumors is often detrimental, reducing its levels has emerged as one of the key research focuses. One promising strategy involves inhibiting ammonia production at its source. For example, activating urea cycle enzymes such as arginase or using clinically approved ammonia scavengers can effectively lower ammonia concentrations within the TME.2,10 In addition, blocking ammonia reutilization has shown efficacy. Inhibitors targeting the glutaminolysis pathway can significantly impair the metabolic adaptability of tumor cells, reducing their survival advantage. Combining ammonia metabolism regulation with other therapeutic approaches represents another promising combinatorial therapy.11 For instance, integrating ammonia inhibition strategies with immunotherapy or radiotherapy may enhance treatment outcomes. Studies have demonstrated that inhibiting ammonia reutilization while simultaneously activating T-cell functions can potentiate immune responses and significantly suppress tumor progression. However, tumor metabolic heterogeneity poses a significant challenge. Different tumor types exhibit variable dependencies on ammonia and differing sensitivities to its accumulation. Thus, further preclinical studies are essential to stratify patients based on these variations and optimize therapeutic strategies accordingly. Although recent studies have provided conceptual insights into ammonia metabolism, the underlying molecular mechanisms remain poorly understood. Evidence suggests that ammonia sensing may involve pH buffering mechanisms or activation of the mTOR signaling pathway. Intriguingly, hypoxia-inducible factor-1α may indirectly respond to metabolic stress caused by ammonia accumulation. Moreover, the effects of ammonia within cells extend beyond its metabolic functions. Ammonia can influence tumor and immune cell behavior through ion gradient changes and downstream signaling pathways. Targeting these signaling pathways, rather than solely focusing on ammonia metabolism, represents a critical and yet unresolved challenge in the field. Conclusion: Ammonia metabolism in the TME plays a dual role in promoting tumor progression and suppressing anti-tumor immunity. Emerging insights into its metabolic and immunosuppressive functions highlight ammonia as a promising therapeutic target, particularly in colorectal cancer where both microbial and cellular sources contribute to its accumulation. The complex interplay between ammonia metabolism and cancer stem cell maintenance offers new opportunities for therapeutic intervention. Leveraging advanced technologies, including CRISPR-based screens, spatial single-cell approaches, and computational modeling, will be critical to uncovering the full functional spectrum of ammonia in the TME.12 These efforts will pave the way for precision therapies targeting ammonia metabolism, creating new opportunities to overcome metabolic resistance and improve patient outcomes. This work was funded by NIH grants R01CA148828, R01CA245546, and R01DK095201 (to YMS), UMCCC Core grant P30CA046592 (to YMS). 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