Bidirectional regulation of the gut microbiome-immune axis in the immune microenvironment of colorectal cancer and targeted interventions

Abstract

The initiation and progression of colorectal cancer (CRC) are profoundly influenced by the complex interplay between the gut microbiota and the immune system, underscoring the clinical importance of exploring the bidirectional regulatory mechanisms of the microbiota-immune axis within the CRC immune microenvironment. Emerging evidence indicates that the composition and functional capacity of the gut microbiota play a vital role in modulating the host’s immune responses, while the immune system, in turn, can reciprocally regulate the structure and function of the microbiota. Despite significant insights into the role of the microbiota-immune axis in CRC progression, several critical questions remain unanswered-including how microbial heterogeneity affects therapeutic outcomes and the specific consequences of dysregulated regulatory mechanisms on the immune microenvironment. This review aims to provide a comprehensive analysis of the compositional features of the CRC immune microenvironment, examine the bidirectional molecular mechanisms underpinning the microbiota-immune axis, and evaluate the potential of targeted therapeutic strategies, thereby offering novel research perspectives and clinical applications for CRC treatment.

Key Words: Microbiota-immune axis; Colorectal cancer; Immune microenvironment; Bidirectional regulation; Targeted intervention

Core Tip: This review explores the bidirectional regulation of the microbiota-immune axis in the immune microenvironment of colorectal cancer (CRC). It discusses how gut microbiota influences immune responses and how the immune system modulates microbiota composition. Highlighting the complexity of these interactions, the review addresses unanswered questions about microbiota heterogeneity and its impact on therapeutic outcomes. It also examines targeted interventions and their potential to improve CRC treatment, offering new insights into both the basic mechanisms and clinical applications for effective therapy.

INTRODUCTION

Colorectal cancer (CRC) remains one of the most prevalent and lethal malignancies globally, with the immune microenvironment playing a pivotal role in tumor initiation, progression, and prognosis[1-3]. In recent years, accumulating evidence has underscored the significant involvement of gut microbiome-immune system interactions in the pathogenesis of CRC[4,5]. The gut microbiome not only maintains intestinal barrier integrity but also influences host immune responses, metabolic processes, and neurotransmitter regulation, thereby affecting systemic health[6]. Dysbiosis, characterized by an imbalance in the gut microbiota, has emerged as a key disruptor of homeostasis and is closely associated with the onset and progression of CRC[7].

Gut microbes interact with the immune system through their metabolic byproducts, such as short-chain fatty acids (SCFAs) and bile acids, which modulate immune responses and influence tumor initiation and progression[8-10]. For example, certain gut microbes can either enhance or suppress the activity of specific immune cell subsets, thus shaping the tumor microenvironment. Moreover, alterations in the gut microbial composition can affect hepatic immunity by modulating the gut-liver axis, a critical pathway in CRC metastasis[11].

The bidirectional regulatory mechanisms of the gut microbiome in CRC provide novel insights into this intricate biological process. Beyond immune modulation, gut microbes directly interact with tumor cells, contributing to the creation of a microenvironment conducive to tumor development[12-14]. For instance, Fusobacterium nucleatum (Fn) is believed to play a crucial role in CRC pathogenesis, potentially accelerating disease progression by influencing host immune responses and cellular signaling pathways[15]. Furthermore, the diversity of the gut microbiota has been correlated with CRC prognosis, with dysbiosis potentially causing immune dysfunction and facilitating tumorigenesis and progression[16,17].

Ongoing research into the interplay between the gut microbiome and the immune system expands our understanding of CRC pathogenesis and opens up new avenues for targeted therapeutic interventions. Emerging strategies such as dietary modulation, probiotic supplementation, and fecal microbiota transplantation (FMT) are gaining recognition as promising approaches to restore microbiome balance[18]. These interventions are promising for improving the treatment outcomes and prognosis of patients with CRC.

In conclusion, the gut microbiome-immune axis plays a critical, bidirectional role in CRC development and progression. A deeper understanding of these mechanisms will provide new insights for the development of more effective therapeutic strategies, ultimately improving the clinical outcomes of CRC treatment.

To this end, this review presents an in-depth analysis of the compositional landscape of the immune microenvironment of CRC, explores the bidirectional molecular interactions within the microbiota-immune axis, and assesses emerging targeted therapies, aiming to uncover new directions for research and clinical intervention in CRC treatment.

COMPOSITION AND CHARACTERISTICS OF THE IMMUNE MICROENVIRONMENT IN CRC

The immune microenvironment of CRC is a dynamic and complex ecosystem composed of various immune cells, cytokines, extracellular matrix components, and the gut microbiota[19,20]. It plays a crucial role not only in tumor initiation and progression but also in determining therapeutic responses and clinical outcomes. Recent studies have highlighted a bidirectional regulatory relationship between the gut microbiota and the host immune system, wherein the composition, function, and metabolites of the microbiota are key modulators in CRC development[21,22].

The mucosal layer and intestinal epithelial cells form the first line of defense against microbial invasion. The mucus layer, primarily composed of glycoproteins, acts as a physical barrier against harmful microorganisms and toxins while providing a niche for commensal bacteria. Epithelial cells maintain the integrity of the intestinal barrier through tight junctions and modulate microbial homeostasis by secreting antimicrobial peptides and cytokines. Disruption of this barrier allows pathogenic bacteria to infiltrate the mucosa, trigger chronic inflammation, and promote intestinal lesions, thereby facilitating CRC progression[23]. Alterations in the macroenvironment of the gut often accompany structural and functional changes in the mucosal layer during CRC development. Certain microbial metabolites can activate signaling pathways that influence epithelial cell proliferation and differentiation, thereby indirectly shaping the immune landscape[23].

The spatial distribution and functional status of immune cells are central to the immune microenvironment of CRC. The formation of tertiary lymphoid structures (TLS) is considered a hallmark of active anti-tumor immune responses and is typically associated with improved prognosis. However, under certain conditions, TLS may paradoxically contribute to localized immunosuppression, which can impair effective immune surveillance. This immunosuppressive microenvironment may, in turn, facilitate the development of an “immune desert” phenotype[24,25]. In this context, effector immune cells such as macrophages and T cells are often suppressed or reprogrammed under the influence of tumor-derived cytokines, facilitating immune evasion (Figure 1)[26-28].

Figure 1 Mechanisms of immune evasion in colorectal cancer. The immune evasion of colorectal cancer occurs in four stages: (1) Initiation: Normal cells undergo malignant transformation into cancer cells under the influence of carcinogenic factors; (2) Elimination: Immune cells, such as CD8+ T cells and natural killer cells, recognize and attack transformed cells with reduced major histocompatibility complex class I (MHC-I) expression; (3) Equilibrium: Tumor cells enter a dormant state, during which the immune system-mediated by CD4+ T cells, interleukin-12, and interferon-gamma-exerts selective pressure to control tumor growth; and (4) Escape: Cancer cells further downregulate MHC-I expression and exploit immunosuppressive cells such as regulatory T cells and myeloid-derived suppressor cells to promote tumor progression and achieve immune evasion. MHC-I: Major histocompatibility complex class I; IL-12: Interleukin-12; NK: Natural killer cells; Treg: Regulatory T cell; MDSCs: Myeloid-derived suppressor cells.

Among the microbial metabolites involved in immune modulation, SCFAs and secondary bile acids are of particular interest. SCFAs, including acetate, propionate, and butyrate (NaB), regulate epithelial cell function and enhance barrier integrity via G protein-coupled receptor signaling, while also exhibiting anti-inflammatory effects that may reduce CRC risk. Secondary bile acids influence microbiota composition and immune responses through pathways such as the farnesoid X receptor, and their dysregulation has been closely linked to CRC pathogenesis[23].

Overall, the CRC immune microenvironment is shaped by complex interactions among gut microbiota, epithelial barriers, immune cells, and their molecular mediators. A deeper understanding of these components and their regulatory mechanisms may provide new insights and therapeutic targets for advancing cancer immunotherapy.

BIDIRECTIONAL REGULATION OF THE GUT MICROBIOME-IMMUNE AXIS: MOLECULAR MECHANISMS

The microbiota-immune axis shapes the CRC immune microenvironment through multiple molecular pathways. Gut microbes activate immunity via pattern recognition receptors (PRRs) such as Toll-like receptor 4 (TLR4) and NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3). Their metabolites, such as SCFAs, maintain immune balance by modulating regulatory T cell (Treg)/T helper 17 cell (Th17) dynamics. Certain microbial antigens also enhance cluster of differentiation (CD)8+ T cell-mediated antitumor responses. Conversely, the host immune system regulates microbiota composition through immunoglobulin A (IgA) and cytokines such as interleukin-17 (IL-17) and interleukin-22 (IL-22), influencing inflammation and barrier function. Dysregulation of the programmed cell death protein 1 (PD-1) pathway further disrupts microbial balance and immune surveillance. This bidirectional regulation plays a key role in CRC progression and offers potential therapeutic targets.

Microbiome: Immune system

The gut microbiome plays a crucial role in modulating the immune system through the activation of PRRs, particularly TLR4 and NLRP3[29,30]. TLR4 activation promotes myeloid cell maturation and the release of inflammatory mediators, amplifying both local and systemic immune responses[8]. Beilmann-Lehtonen et al[31] performed immunohistochemical analysis of TLR4 and T cells (CD3+, CD8+) expression in tumor tissues from 549 patients with CRC and found that high TLR4 expression was significantly associated with increased intratumoral T cell infiltration, suggesting that TLR4 participates in the CRC immune microenvironment by regulating immune cells. However, Lee et al[32], using wild-type and TLR4 knockout germ-free mice mono-colonized with Bacteroides fragilis (B. fragilis) and induced colitis-associated CRC, found that B. fragilis significantly inhibited tumor formation by regulating immune responses through activation of the TLR4 signaling pathway. Feng et al[33], using an MC38 xenograft mouse model, demonstrated that polysaccharides from Atractylodes macrocephala (PAM) promoted macrophage immune responses via activation of the Myeloid differentiation primary response 88/TLR4 signaling pathway, significantly inhibiting CRC tumor growth and prolonging mouse survival, indicating that the TLR4-mediated immune mechanism plays a key role in PAM’s anti-CRC effects. The NLRP3 pathway is associated with the activation of intracellular inflammasomes, which induce the secretion of various proinflammatory cytokines by myeloid cells. This process is intricately regulated by the gut microbiome[34]. Serrya et al[35] established an acetic acid-induced ulcerative colitis rat model and found that mycophenolate mofetil significantly alleviated intestinal inflammation and oxidative stress by inhibiting NLRP3 inflammasome and pro-inflammatory cytokine expression, activating Nuclear factor erythroid 2-related factor 2 signaling, and modulating interferon expression, suggesting that NLRP3 inflammasome-mediated immune-inflammatory responses play a critical role in the inflammatory microenvironment associated with CRC.

Metabolites produced by the gut microbiome, specifically SCFAs such as NaB, play a critical role in regulating the immune system. NaB exerts its effects by inhibiting the activity of histone deacetylase, thereby modulating the balance between Treg and Th17 cells, an essential mechanism for maintaining intestinal immune homeostasis. Reportedly, NaB promotes the proliferation of Treg cells while suppressing the differentiation of Th17 cells, effectively reducing intestinal inflammation, which is vital for the prevention and treatment of CRC[36-38]. Alrafas et al[39] used an Azoxymethane (AOM)/Dextran sulfate sodium (DSS)-induced CRC mouse model combined with resveratrol treatment to regulate the gut microbiota and SCFAs NaB levels, finding that increased NaB led to a reduction in pro-inflammatory Th17 cells, thereby inhibiting CRC development. NaB enhances the function of intestinal epithelial cells, reinforces the intestinal barrier, and alleviates local inflammation, contributing to the maintenance of gut health.

The gut microbiome also enhances immune system efficacy by generating specific antigens that elicit targeted immune responses from CD8+ T cells. Certain microbiota-derived antigens have been shown to activate CD8+ T cells through cross-presentation mechanisms, stimulating their proliferation and effector functions. This process is critical for the immune surveillance of intestinal tumors, as CD8+ T cells can directly recognize and eliminate tumor cells (Figure 2)[40,41]. Xu et al[42] employed an MC38 CRC mouse model and demonstrated that the combined therapy of an anti-angiogenic adenovirus encoding endostatin and PD-1 blockade significantly suppressed tumor growth, with the gut microbe B. fragilis enhancing the activation of CD8+ T cells within the tumor microenvironment. In addition to strengthening anti-tumor immune responses in CRC, this mechanism may influence the overall immune state, offering potential targets for the development of novel immunotherapeutic strategies. Liao et al[43] further revealed that the gut microbial metabolite 4-Hydroxyphenylacetic acid promotes CRC progression by activating the Janus kinase 2/Signal transducer and activator of transcription 3 pathway to upregulate C-X-C motif chemokine ligand 3 (CXCL3) expression, thereby recruiting immunosuppressive Polymorphonuclear myeloid-derived suppressor cells that inhibit CD8+ T cell antitumor activity; notably, microbial metabolic intervention or CXCL3 knockdown reversed immunosuppression and enhanced anti-PD-1 efficacy, suggesting the gut microbiota metabolite-CD8+ T cell axis as a novel therapeutic target for CRC.

Figure 2 Mechanism of antigen presentation-induced cluster of differentiation 8+ T cell-specific immune response. The mechanism involves five steps: (1) Tumor antigen release: Tumor cells shed or release antigens; (2) Antigen presentation by dendritic cells (DC): DC, a type of antigen-presenting cells (APCs), capture and process these tumor antigens; (3) Activation of cluster of differentiation (CD)8+ T cells by APCs: The APCs then activate CD8+ T cells through antigen presentation; (4) Migration of activated CD8+ T cells to the tumor site: Activated CD8+ T cells migrate toward the tumor location; and (5) CD8+ T cell-mediated tumor cell killing: Finally, CD8+ T cells exert cytotoxic effects and eliminate tumor cells. APCs: Antigen-presenting cells; DC: Dendritic cells; CD: Cluster of differentiation.

Immune system: Microbiome

The immune system regulates the composition and function of the gut microbiome by secreting IgA, which neutralizes pathogens in the gut. IgA selectively binds to specific bacteria, influencing their colonization and proliferation within the intestine. The presence of the J-chain is essential for the formation and secretion of IgA, facilitating its aggregation in the intestinal mucosa and enhancing immune surveillance of the gut microbiota[15]. This IgA-mediated selective action not only plays a pivotal role in maintaining intestinal microbiota balance but also prevents pathogenic microbial invasion and the development of intestinal diseases. Magat et al[44] detected IgA antibodies against Clostridioides difficile toxin B (tcdB) in plasma from patients with CRC and healthy controls using Enzyme-Linked Immunosorbent Assay, and found that tcdB-specific IgA levels were significantly elevated in patients with CRC, suggesting that gut microbiota-related IgA immune responses play an important role in CRC development.

Cytokines play a crucial role in the interaction between the immune system and the gut microbiome. Cytokines such as IL-17 and IL-22 regulate the function of intestinal epithelial cells and microbial metabolic pathways, influencing the composition and activity of the gut microbiota. Upregulation of IL-17 is closely associated with intestinal inflammation, whereas IL-22 contributes to maintaining the integrity of the intestinal barrier[45-47]. Han et al[48] conducted clinical retrospective analyses and mouse model studies, discovering that statins significantly inhibit CRC development by increasing the gut microbiota Lactobacillus reuteri and its metabolite indole-3-lactic acid, which suppresses the IL-17 signaling pathway, highlighting the gut microbiota-IL-17 axis as a novel target for CRC prevention. Yu et al[49] conducted bioinformatics analysis of clinical fecal samples and FMT experiments in mice, finding that Lactobacillus species significantly alleviated acute chemoradiotherapy-induced intestinal injury by regulating IL-22 expression in gut lymphoid tissue inducer cells, suggesting a protective role of the gut microbiome-IL-22 axis in CRC treatment. The dynamic regulation of this cytokine network through the modulation of microbial metabolic products impacts host immune responses. This feedback mechanism may play a crucial role in the development of CRC, influencing the tumor microenvironment and the host’s immune status.

Within the context of immune system regulation, the upregulation of PD-1 may lead to immune dysregulation, affecting the composition of the gut microbiome. As an inhibitory immune checkpoint, PD-1 dysfunction can promote the colonization and overgrowth of specific pathogenic bacteria, which is closely associated with the development of CRC. Studies have shown that high PD-1 expression correlates with reduced microbial diversity in the gut and an increased abundance of pathogenic bacteria, contributing to dysbiosis. This microbial imbalance can, in turn, disrupt the immune microenvironment and promote tumor progression[50,51]. Gopalakrishnan et al[52] analyzed the oral and gut microbiomes of patients with melanoma (n = 112) undergoing anti-PD-1 immunotherapy and found significant differences in gut microbial diversity and composition between responders and non-responders. Gao et al[53], through multi-omics analysis and mouse model experiments, found that the probiotic Lacticaseibacillus rhamnosus Probio-M9 enhanced the inhibitory effect of anti-PD-1 immunotherapy on CRC by modulating the gut microbiota and its metabolites, promoting cytotoxic T cell activation while suppressing Treg function. Wen et al[54] also found that Chang-Wei-Qing combined with PD-1 inhibitors effectively suppressed AOM/DSS-induced colitis-associated CRC by regulating the gut microbiota and improving intestinal barrier function. Therefore, targeting PD-1 and related pathways may hold promise as novel therapeutic strategies for CRC.

DYSREGULATION OF BIDIRECTIONAL CROSSTALK AND TUMOR MICROENVIRONMENT REMODELING

The bidirectional crosstalk between the gut microbiota and the host immune system plays a central role in shaping the CRC immune microenvironment. Disruption of this balance leads to the remodeling of the tumor microenvironment, thereby promoting tumor initiation and progression. Recent studies have highlighted a strong association between gut microbiota dysbiosis (i.e., "microbial imbalance") and the development of CRC[55,56]. Dysbiosis directly disrupts immune responses within the gut and may indirectly modulate the tumor microenvironment by altering the metabolic and inflammatory states of the host[55,56]. Consequently, elucidating the mechanisms underlying this bidirectional dysregulation is essential for the development of novel therapeutic strategies.

For the pro-tumorigenic mechanisms, chronic inflammation is a key oncogenic mechanism in CRC. Evidence suggests that dysbiosis of the gut microbiota triggers chronic inflammatory responses, which activate the Wnt/β-catenin signaling pathway. Aberrant activation of this pathway is strongly associated with CRC development, as it promotes tumor cell proliferation and survival while inhibiting apoptosis[57]. Additionally, cytokines and chemokines produced during chronic inflammation can exacerbate the formation of the tumor microenvironment, altering the interactions between tumor and immune cells and accelerating tumor progression[58]. Therefore, investigating the role of chronic inflammation, especially its influence on the Wnt/β-catenin signaling pathway, may reveal promising therapeutic targets for CRC.

Concerning the anti-tumorigenic mechanisms, certain gut microbiota can enhance anti-tumor immune responses by activating the stimulator of interferon genes signaling pathway. Studies have shown that specific microbial communities can promote the infiltration of CD103+ dendritic cells (DCs) into tumor tissues, thereby enhancing anti-tumor immune responses. These CD103+ DCs are essential for antigen presentation and T-cell activation, and improve the body's ability to recognize and eliminate tumor cells[59]. This anti-cancer mechanism suggests that modulating the composition of the gut microbiota may help boost the host’s immune defenses, providing new perspectives for immunotherapy in CRC.

Regarding the bidirectional regulatory mechanism, within the gut microbiota-immune axis, the aryl hydrocarbon receptor (AhR) and indoleamine 2,3-dioxygenase 1 (IDO1) pathways represent important molecular nodes. As a transcription factor, AhR can sense microbial metabolites in the gut and modulate the host’s immune response. Specifically, IDO1 catalyzes the metabolism of tryptophan to produce metabolites such as kynurenine (Kyn), which depletes local tryptophan availability and activates AhR signaling to promote the differentiation of T cells into Tregs, thereby suppressing excessive immune responses and inflammation[60]. Fong et al[61] utilized a mouse model of CRC and multi-omics approaches to demonstrate that the probiotic Lactobacillus gallinarum produces indole-3-carboxylic acid, which modulates the IDO1/Kyn/AhR signaling pathway to suppress CD4+ Treg differentiation and enhance CD8+ cytotoxic T cell function, thereby improving the gut microbiota composition and significantly enhancing the efficacy of anti-PD-1 immunotherapy against CRC. This highlights the crucial role of bidirectional regulation between microbial metabolites and the immune system in CRC immunotherapy[61]. Other studies have shown that the IDO1 metabolite Kyn promotes Treg differentiation through AhR activation, suppressing antitumor immunity and contributing to tumor progression. This suggests that microbial metabolite-mediated regulation of the IDO1-Kyn-AhR immune axis plays a key role in CRC development, and targeting this pathway may represent a novel strategy for CRC prevention and therapy[62]. In addition, Shi et al[63] found that Ubiquitin-specific protease 14 (USP14) stabilizes IDO1 protein, thereby promoting tryptophan metabolism and T cell dysfunction, leading to the suppression of antitumor immune responses; inhibition of USP14 reduces IDO1 expression and enhances anti-PD-1 efficacy without affecting AhR activation. Therefore, this bidirectional regulatory network plays a crucial role in the initiation and progression of CRC. Consequently, maintaining the balance between the gut microbiota and the host immune system presents promising opportunities for the development of targeted therapeutic strategies.

Notably, significant spatiotemporal heterogeneity exists between different CRC sites, and this heterogeneity is particularly evident in microbiota-immune interactions. Research has shown that the composition of the microbiota and the degree of immune cell infiltration vary markedly across different regions of the gut, which may influence tumor growth and metastasis[64]. For instance, certain regions may exhibit a robust immune response due to a high abundance of beneficial microbes, whereas other regions may experience immune evasion due to the proliferation of pathogenic bacteria. This spatiotemporal heterogeneity suggests that, when designing individualized treatment strategies for CRC, it is crucial to consider the site-specific microbiota and immune environment to achieve a more precise and effective intervention.

ASSOCIATION BETWEEN MICROBIAL HETEROGENEITY AND TREATMENT RESPONSE

The treatment of CRC involves complex interactions between the gut microbiota and the tumor immune microenvironment. Studies have shown that the gut microbiome directly influences the host’s immune response and holds potential as a predictive biomarker for treatment outcomes[65,66]. In recent years, accumulating evidence has shown that the abundance of specific gut microbial taxa is closely associated with treatment response, prognosis, and resistance to immunotherapy, suggesting that the gut microbiota may serve as a predictive biomarker for therapeutic outcomes and provide novel targets for personalized precision interventions (Table 1).

Table 1 Comparative impact of gut microbiota interventions on immune regulation and colorectal cancer therapy outcomes.

Intervention	Key microbes	Mechanism of action	Impact on CRC treatment
Prognostic Biomarker	Fn/Fp ratio	Fn promotes inflammation and tumorigenesis, while Fp has anti-inflammatory and protective effects; Fn/Fp ratio reflects tumor microenvironment status	Higher Fn abundance correlates with poor prognosis, while higher Fp levels associate with better outcomes; Fn/Fp ratio aids in early screening and prognosis assessment
Enhance Chemosensitivity	NaB-producing bacteria (e.g., Faecalibacterium) and NaB	NaB strengthens gut barrier function, modulates immune activity, and induces tumor cell apoptosis while inhibiting proliferation, migration, and invasion	Improves efficacy of OXA and other chemotherapies while reducing side effects
Modulate Immunotherapy Response	B. fragilis (polysaccharides), Fn (succinic acid)	(1) B. fragilis polysaccharides synergize with CTLA-4 inhibitors to activate T cells; and (2) Fn-derived succinate inhibits the cGAS-interferon-β pathway, reducing CD8+ Tcells infiltration	(1) Enhances immune checkpoint inhibitor efficacy; and (2) High Fn abundance causes anti-PD-1 resistance, reversible via antibiotics or microbiota modulation

CRC: Colorectal cancer; Fn: Fusobacterium nucleatum; Fp: Faecalibacterium; OXA: Oxaliplatin; NaB: Butyrate; CD: Cluster of differentiation; CTLA-4: Cytotoxic T-lymphocyte-associated protein 4; cGAS Cyclic: GMP-AMP synthase; PD-1: Programmed cell death protein 1; B. fragilis: Bacteroides fragilis.

Gut microbiota composition as a prognostic biomarker

Research has identified the ratio of Fn to Faecalibacterium as a key prognostic marker in patients with CRC. Fn promotes gut inflammation and tumorigenesis, whereas Faecalibacterium has anti-inflammatory and protective properties. Variations in the relative abundance of these microbes may reflect the tumor microenvironment, thereby influencing patient survival and treatment response. Evidence suggests that a higher abundance of Fn is associated with poorer clinical outcomes, whereas increased levels of Faecalibacterium correlate with improved prognosis[67,68]. Guo et al[69] assessed the relative abundance of Fn and Faecalibacterium prausnitzii (Fp) in 903 fecal samples using quantitative PCR, revealing a negative correlation between Fn and Fp, and demonstrating that the Fn/Fp ratio exhibited high sensitivity and specificity for early screening of CRC. These findings provide new biomarkers for evaluating the prognosis of patients with CRC.

NaB-producing bacteria enhance chemotherapy sensitivity

Certain gut microbiota, particularly NaB-producing bacteria, can enhance the sensitivity of CRC to the chemotherapeutic agent oxaliplatin (OXA). NaB, an SCFA, modulates the composition and function of the gut microbiome, promoting the health and immune activity of intestinal epithelial cells. In CRC treatment, the presence of NaB-producing bacteria enhances the efficacy of chemotherapy and alleviates chemotherapy-related side effects. Research indicates that these bacteria improve drug bioavailability by enhancing the gut barrier function while simultaneously promoting anti-tumor immune responses, thereby boosting the effectiveness of chemotherapy[70]. Shuwen et al[71] conducted metabolomic and 16S rRNA microbiota analyses in patients with CRC and mouse models, and found that NaB-producing bacteria and their metabolite sodium NaB synergistically enhanced the efficacy of OXA by inhibiting tumor cell proliferation, migration, and invasion, and promoting apoptosis. These findings suggest that NaB-producing bacteria play a crucial role in improving chemosensitivity and therapeutic outcomes in CRC, opening new avenues for optimizing chemotherapy protocols through the modulation of the gut microbiome.

Gut microbiota modulates immunotherapy response

In the context of immunotherapy, patients with CRC often encounter issues related to immune resistance. Reportedly, the modulation of the gut microbiota can effectively reverse this resistance. Particularly, polysaccharides produced by B. fragilis can synergize with the immune checkpoint inhibitor Cytotoxic T-lymphocyte-associated protein 4 to enhance T-cell activation, thereby improving the efficacy of immunotherapy[72]. This mechanism underscores the pivotal role of the gut microbiome in chemotherapy and immunotherapy. Zhang et al[73] integrated multi-omics data analysis and validated their findings in humanized mouse models, revealing that the gut microbiota composition and its metabolites in patients with CRC significantly influence the tumor immune microenvironment. Moreover, microbiota transplantation from these patients reduces the efficacy of immunotherapy, suggesting that modulation of the gut microbiota may enhance the effectiveness of CRC immunotherapy. Jiang et al[74], through FMT and metabolite analysis in metastatic patients with CRC, found that Fn and its metabolite succinic acid reduce CD8+ T cell infiltration in tumors by inhibiting the Cyclic GMP-AMP synthase-interferon-β pathway, leading to resistance to anti-PD-1 immunotherapy, while antibiotics that reduce the abundance of this bacterium can restore immunotherapy efficacy. By adjusting the abundance of specific microbial populations, patient responses to immunotherapy can be improved, further advancing the development of personalized treatment strategies for CRC.

TARGETING THE MICROBIOTA-IMMUNE AXIS: INTERVENTION STRATEGIES

In CRC treatment, intervention strategies targeting the microbiota-immune axis have increasingly garnered attention. These approaches extend beyond direct tumor targeting to address the complex interactions between the gut microbiota and the host immune system, offering novel perspectives for anti-tumor immunity. Recent studies have emphasized the significant role of microbial composition and its metabolic products in the tumor microenvironment, profoundly influencing immune responses and tumor progression. Consequently, targeting the microbiota-immune axis could pave new pathways for immunotherapy in CRC.

Engineered probiotics

Engineered probiotics, designed to secrete specific immune factors such as interleukin-15 (IL-15) and Fms-like tyrosine kinase 3 Ligand (FLT3 L), have demonstrated considerable potential in remodeling the tumor microenvironment. IL-15 is a key cytokine that promotes the proliferation and activation of cytotoxic T and natural killer cells, whereas FLT3 L aids in the development and functional enhancement of DCs[75]. By leveraging engineered microbiota strategies, precise modulation of these immune factors can be achieved, thereby enhancing the anti-tumor immune responses of the host. This approach boosts the activity of immune cells and improves the immune adaptability of the tumor microenvironment, opening new therapeutic possibilities for CRC. Li et al[76] employed single-cell RNA sequencing and a humanized mouse model to demonstrate that delivering interleukin-32 (IL-32) and selenium nanoparticles via engineered probiotic Escherichia coli Nissle 1917 significantly enhanced CD8+ T cell immune activity and alleviated immune tolerance, thereby effectively suppressing CRC progression. Another research team constructed an engineered probiotic strain, Ep-AH, expressing the tumor-adhesive protein Histone-like protein A and the tumoricidal protein azurin. In a CRC mouse model, Ep-AH markedly inhibited tumorigenesis, improved inflammatory responses, and modulated gut microbiota composition[77]. Culpepper et al[78] used the AOM/Il10^-/^- mouse model to evaluate the anti-tumor efficacy of engineered Saccharomyces boulardi and found that it effectively reduced intestinal inflammation and significantly decreased tumor burden, suggesting that engineered probiotics hold promising therapeutic potential for CRC by exerting local anti-inflammatory effects and regulating the intestinal microenvironment. However, despite their potential, engineered probiotics still face several major challenges in CRC treatment, including low colonization efficiency, unclear biosafety and immunological risks, difficulty in precisely regulating functional protein expression, lack of clinical validation, potential disruption of native microbiota homeostasis, and unresolved issues regarding regulatory oversight and ethical review.

Indole propionic acid and nanoparticle delivery systems

Indole propionic acid (IPA), a significant metabolic product, is essential for regulating gut immunity and maintaining intestinal barrier function. Jia et al[79] discovered through microbial supplementation and metabolite intervention experiments that Lactobacillus johnsonii cooperates with Clostridium sporogenes to produce IPA, which promotes the formation of progenitor exhausted CD8+ T cells by enhancing Histone 3 Lysine 27 acetylation at the Transcription factor 7 super-enhancer region, thereby significantly improving the response rate to PD-1 immunotherapy in CRC and other tumor models. Zhou et al[80] found that the probiotic Limosilactobacillus fermentum GR-3 can significantly alleviate inflammation, oxidative stress, and intestinal barrier damage by producing metabolites such as IPA. This probiotic also induces tumor cell apoptosis, reduces tumor incidence, and highlights the important role of IPA in CRC treatment through modulation of the gut microbiota and metabolic pathways. Engineered nanoparticle delivery systems can enable targeted delivery of IPA, thereby enhancing its bioavailability and efficacy within the gut[79,81]. This strategy amplifies the regulatory effects of microbiota-derived metabolites on the immune system as well as inhibits tumor initiation and progression by improving the gut microenvironment. This approach harnesses the potential of gut microbiota in the prevention and treatment of CRC more effectively. Although IPA and nanoparticle delivery systems have shown promising effects in CRC treatment, their clinical application is still limited by IPA’s low stability and bioavailability in vivo, as well as the unclear safety and metabolic mechanisms of the nanoparticle carriers. In addition, individual differences in gut microbiota may affect therapeutic efficacy. Currently, most related studies remain at the animal and in vitro stages, lacking clinical validation, thus requiring further optimization and clinical trial support.

FMT

FMT is an emerging therapeutic strategy, with its success heavily dependent on the selection and matching of donors. Studies have shown that the compatibility between the donor's immune phenotype and the recipient’s microbiome is a crucial determinant of FMT success[82,83]. By establishing new standards for donor-immune phenotype matching, FMT success rates in patients with CRC can be improved, facilitating microbiome restoration and enhancing the host's immune response. Gou et al[84], using AOM/DSS-induced and Apc^Min/+ mouse models combined with metagenomic sequencing and metabolomic analysis, found that Pien Tze Huang improved gut barrier function and inhibited oncogenic and inflammatory pathways by modulating the gut microbiota and metabolites; FMT partially replicated its anti-CRC effects, suggesting that FMT has potential chemo preventive and immunomodulatory roles in CRC treatment. Zhang et al[85] applied FMT intervention in a mouse model of ulcerative colitis and found that FMT restored gut microbiota balance, increased SCFA levels, inhibited inflammation and the Nuclear Factor kappa B pathway, thereby improving gut barrier function and alleviating disease symptoms. Meanwhile, a phase II clinical trial evaluating FMT combined with tislelizumab and fruquintinib for refractory microsatellite stable metastatic CRC showed that this combination significantly prolonged progression-free survival and overall survival with manageable safety, indicating that FMT plays an important role in enhancing immunotherapy efficacy and CRC treatment[82]. The implementation of this strategy requires extensive clinical research and data analysis to optimize donor selection and recipient matching processes.

Combined immunotherapy strategy

Combined immunotherapy holds great promise in cancer treatment, particularly the strategy of combining PD-1 inhibitors with microbiota-derived metabolic vaccines. PD-1 inhibitors can reverse the suppression of the immune system by tumor cells, while microbiota-derived metabolic vaccines promote enhanced immune responses by modulating the microbiome[86]. Ni et al[87] developed a bi-adjuvant neoantigen nanovaccine that enhances immune responses through efficient co-delivery of neoantigens and synergistic adjuvants; combined with anti-PD-1 therapy, it significantly promoted complete regression of CRC tumors, demonstrating the strong potential of combination immunotherapy in CRC treatment. Liu et al[88] analyzed the co-expression and functional status of Tim-3 and PD-1 on tumor-infiltrating CD8+ T cells in patients with CRC and found that combined blockade of the Tim-3 and PD-1 pathways, compared to single blockade, significantly enhanced the function and proliferation of tumor antigen-specific T cells, reduced the number of Tregs, and increased cytotoxic activity, indicating that combination immunotherapy can effectively reverse T cell exhaustion and improve the immune response against CRC[88]. This combination treatment model provides a new avenue for CRC immunotherapy, aiming to improve therapeutic outcomes through dual mechanisms and overcome the limitations of monotherapy. Au et al[89] analyzed cases of cytokine release syndrome (CRS) occurring after mRNA coronavirus disease 2019 vaccination in patients undergoing anti-PD-1 therapy, suggesting that combined immunotherapy may increase the risk of immune-related adverse events and highlighting the need for further monitoring and safety evaluation.

FUTURE RESEARCH

The microbiome-immune axis has garnered increasing attention for its role in remodeling the CRC immune microenvironment. As investigations into this complex bidirectional regulatory mechanism advance, its substantial influence on the tumor microenvironment is becoming increasingly apparent. The onset and progression of CRC are closely linked to the intrinsic characteristics of tumor cells and intricately connected to the composition and function of the gut microbiota. The microbiome, through its modulation of host immune responses, influences tumor immune evasion and potentially contributes to tumor growth and metastasis. Studies have revealed the bidirectional regulatory effects of the microbiome on immune cell functions, which can enhance antitumor immunity and promote immunosuppressive microenvironments, highlighting the importance of a multifaceted approach that considers individual genetic backgrounds, dietary habits, and environmental factors (Table 2).

Table 2 Conceptual framework of the microbiota-immune axis in colorectal cancer.

Level	Key elements	Regulatory mechanisms	Intervention strategies	Translational indicators
Input layer	Microbiome	Pro-carcinogenic bacteria (e.g., Fusobacterium) activate TLR4/β-catenin - Protective bacteria (e.g., Faecalibacterium) produce NaB to strengthen barrier	Microbiome profiling (qPCR/metagenomics)	Fecal α-diversity; Fn/Fp ratio
	Host factors	Genetic mutations (e.g., APC), diet (high-fiber/high-fat), antibiotic exposure history	Risk stratification questionnaire + genetic testing	Patient subtype classification (inflammatory/metabolic)
Core interaction layer	Metabolite-immune crosstalk	SCFAs → HDAC inhibition → Treg induction - Secondary bile acids → FXR → IL-23/Th17 activation	NaB formulations; FXR antagonists	Serum NaB levels; IL-17A in colon tissue
	Cellular network	Microbial antigens → CD103+ DCs → CD8+ T cell activation - PD-1↑ → dysbiosis → MDSC recruitment	Engineered bacteria delivering DC activators (e.g., FLT3 L)	Tumor-infiltrating CD8+ T cell density; circulating MDSC levels
Effector layer	Immune phenotype	Inflamed type (high CD8+, TLS+) - Immune-desert (fibrosis, Treg-dominant)	Spatial multi-omics (CODEX/mIHC)	Immunoscore®, CT-based immune features
	Therapeutic response	Fusobacterium → oxaliplatin resistance via ABCB1 upregulation - B. fragilis → anti-PD-1 sensitization via polysaccharide-CTLA-4 interaction	Microbiome-guided personalized therapy	PFS, ORR
Intervention layer	Microbiome modulation	FMT for microbiota rebalancing - Phage therapy targeting pathogens	Donor-recipient matching; phage cocktail therapy	Post-FMT colonization rate; pathogen load reduction
	Immuno-metabolic combo	AhR inhibitors (e.g., IK-175) block IDO1-Kyn - STING agonists (e.g., ADU-S100) activate CD103+ DCs	Optimization of combination regimens (timing/dosing)	IFN-γ in tumor tissue; CXCL10 in blood
Output layer	Clinical benefit	Improved OS; reduced chemo toxicity (e.g., diarrhea)	QoL scales; CTC monitoring	5-year survival rate; grade ≥ 3 adverse event rate
	Biomarkers	Microbial signature (e.g., Clostridium cluster XIVa abundance); immune dynamics (CD8+/FoxP3+ ratio)	Liquid biopsy (ctDNA + microbial DNA)	AUC of predictive models; longitudinal consistency

Fn: Fusobacterium nucleatum; Fp: Faecalibacterium; TLR4: Toll-like receptor 4; SCFAs: Short-chain fatty acids; HDAC: Histone deacetylase; Treg: Regulatory T cells; FXR: Farnesoid X receptor; IL-23: Interleukin 23; Th17: T helper 17 cells; DCs: Dendritic cells; CD: Cluster of differentiation; PD-1: Programmed cell death protein 1; MDSCs: Myeloid-derived suppressor cells; FLT3 L: Fms-like tyrosine kinase 3 Ligand; TLS: Tertiary lymphoid structures; ABCB1: ATP-binding cassette sub-family B member 1; CTLA-4: Cytotoxic T-lymphocyte-associated protein 4; FMT: Fecal microbiota transplantation; AhR: Aryl hydrocarbon receptor; IDO1: Indoleamine 2:3-dioxygenase 1; Kyn: Kynurenine; STING: Stimulator of interferon genes; IFN-γ: Interferon gamma; CXCL10: C-X-C motif chemokine ligand 10; OS: Overall survival; QoL: Quality of life; CTC: Circulating tumor cells; ctDNA: Circulating tumor DNA; AUC: Area under the curve; NaB: Butyrate.

Future research should focus on several key areas: First, standardizing and harmonizing microbiome analysis methods to improve data comparability and reliability; second, conducting more multicenter, large-scale clinical trials to evaluate the safety and efficacy of microbiome-based interventions such as FMT and probiotic formulations in CRC prevention and treatment; third, emphasizing clinical translation by systematically validating biomarkers and stratifying patients based on immune phenotypes to achieve precision and personalized therapy; and finally, establishing more comprehensive animal models and clinical research platforms to elucidate the specific roles of microbial communities and their metabolites in CRC.

CONCLUSION

In conclusion, the microbiome-immune axis represents a promising frontier in CRC research. A deeper understanding of its complex bidirectional regulatory mechanisms and the promotion of clinical translation will provide a solid scientific foundation and innovative strategies for early diagnosis, prevention, and personalized treatment of CRC.