Role of prebiotics in melanoma

Emerging evidence suggests that prebiotics may support melanoma therapy as they have the capacity to modulate the gut microbiome and metabolome. To understand the potential of prebiotics in melanoma, it is necessary to review the studies conducted across various models, ranging from in vitro systems to animal studies and clinical trials. Recent findings suggest that a high-fiber, prebiotic-rich diet enhances progression-free survival and immunotherapy response in melanoma patients, likely by promoting beneficial gut microbiota. In contrast, the use of certain commercial probiotic supplements, often consumed for their anti-inflammatory properties in health conditions, has been associated with impaired immune responses and reduced treatment efficacy, highlighting the importance of supporting the native healthy microbiome rather than introducing specific external strains during cancer therapy, as this may lead to further complications [20]. Moreover, the authors have shown probiotic use to be associated with impaired response to immune checkpoint blockade, larger tumors, lower gut microbiome diversity, and fewer cytotoxic T cells in the tumor microenvironment. A possible reason for impaired immunity could be that probiotic use disrupts the gut microbial diversity. Spencer et al. [8] have shown that microbial alpha diversity and Ruminococcaceae family and Faecalibacterium genus abundances were higher in patients without probiotic use, and that the microbiome and antitumor immunity were impaired in mice receiving probiotics. FMT, in contrast, is a holistic intervention that introduces the stool donors’ entire stool ecosystem for restoration of the recipient gut microbiome rather than specifically targeting bacterial strains. However, variable efficacy of donor stool, unclear mechanistic pathways, and potential transmission of pathogens or opportunistic microbes, and the need to assess safety in a disease-specific context, limit its application [21].

Given these limitations, prebiotics are particularly important to investigate further as they offer a promising alternative to probiotics and FMT. Furthermore, mechanistic studies reveal that combining probiotics with immunotherapy impairs T-helper cell activity, reducing antigen presentation and decreasing dendritic cell function in mouse models, ultimately limiting immune responses and microbial diversity [22]. This reinforces the potential value of prebiotics, which can strengthen immune modulation through endogenous microbial pathways rather than external supplementation and unclear mechanistic pathways for FMT.

Mechanisms of action of prebiotics

As illustrated in Figure 1, prebiotics achieve health benefits by exerting their effects through several key mechanisms, including the following:

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Figure 1. Overview of the prebiotic potential in modulating gut and immune system function. Created in BioRender. Parvathy, S. (2025) https://BioRender.com/rfh1d8k.

SCFAs and immune pathways

SCFAs, such as acetate, propionate, and butyrate, are key metabolites generated through prebiotic fermentation by gut microbiota that influence host innate and adaptive immunity and metabolic regulation. As shown in Figure 2, SCFAs act through a variety of mechanisms, exerting a downstream effect on the immune system. Chiefly, SCFAs inhibit the histone deacetylases (HDACs), consequently regulating mTOR and MAPK pathways, which are involved in chemokine and cytokine production [24, 25]. Second, SCFA binds to the G-protein-coupled receptors (GPCRs) leading to cascade of downstream effects. The entry of intracellular calcium ions into the cytoplasm increases due to reduced intracellular cAMP levels, thereby regulating the gene transcription and translation in immune cells [26]. This initiates anti-inflammatory signaling cascades and strengthens gut barrier integrity [27, 28]. SCFAs also act via increasing acetyl-CoA production and metabolic integration [29]. In fact, SCFAs are shown to have proinflammatory effects as well as anti-inflammatory effects on different cell types, such as macrophages and microglia [30, 31]. These mechanisms collectively bridge microbial metabolism with immune modulation, providing a mechanistic basis for how prebiotic-derived metabolites can enhance antitumor immunity in melanoma [32].

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Figure 2. Downstream effects of SCFAs produced by gut microbiota. SCFAs regulate immune responses through multiple pathways. They inhibit histone deacetylases (HDACs), influencing mTOR/MAPK signalling and cytokine production, and act as ligands for G-protein-coupled receptors (GPCRs), altering intracellular calcium and cAMP to modulate immune cell gene expression. SCFAs also contribute to cellular metabolism by increasing acetyl-CoA availability. Depending on the context and cell type, SCFAs can exert anti-inflammatory or pro-inflammatory effects. Together, these mechanisms link microbial fermentation of dietary prebiotics with host immune regulation. SCFAs: short-chain fatty acids. Created in BioRender. Parvathy, S. (2025) https://BioRender.com/vlza4p9.

Modulating systemic and gut-associated immune responses

Prebiotics enhance gut microbiota that influences innate and adaptive immunity through various mechanisms, as can be demonstrated from Figure 3. The anti-inflammatory response is stimulated through pattern recognition receptors or GPCRs, promoting the increased production of inflammatory regulators, including dendritic cells, lymphocytes, monocytes, macrophages, and various innate immune factors, thereby enhancing immune surveillance and strengthening disease resistance [35]. Different genera of lactic acid bacteria (Lactobacillus casei, L. acidophilus, L. rhamnosus, L. delbrueckii subsp. bulgaricus, L. plantarum, L. lactis, and Streptococcus thermophilus) have immunomodulatory effects through various approaches. They have been shown to promote IL-6 release, which stimulates B cell clonal expansion leading to IgA release [36, 37]. Also, Bifidobacterium and Lactobacillus stimulate differentiation of T regulatory cells (Treg cells), inhibit the expression of JAK and NF-κB genes, increase the total helper T cells (CD4⁺) and activate T lymphocytes (CD25⁺) and natural killer (NK) cells [38, 39]. Li et al. [40] found that Bifidobacteria, Bacteroides, and Akkermansia muciniphila lead to an elevated expression of chemokines (CCL4 and CCL8), inflammasome-related genes (TLR3 and TLR7), and antigen presentation-related genes (CD40, Stat1, and ICOS). This indicates a mechanism by which the beneficial gut bacteria enhanced the recruitment and activation of immune cells in the tumor microenvironment, thereby imparting anti-tumor immunity. Furthermore, Akkermansia, Bifidobacterium, and Faecalibacterium are shown to be associated with programmed death 1 (PD-1) blockade in cancer patients, which is usually expressed by tumor cells to prevent the immune system response and suppress anti-tumor immunity [41–44].

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Figure 3. Effects of different gut microbiota on innate and adaptive immunity. Prebiotics promote the growth of beneficial gut bacteria that influence immune signalling pathways. These microbes stimulate anti-inflammatory responses through GPCR and pattern-recognition receptor activation, supporting dendritic cell maturation and the recruitment of macrophages, lymphocytes, and other innate immune cells. Beneficial taxa enhance T regulatory and effector T cell activity, increase IgA production, and improve antigen presentation within the tumour microenvironment. Together, these interactions strengthen immune surveillance and may enhance antitumor responses to therapies such as PD-1 blockade. GPCRs: G-protein-coupled receptors. Created in BioRender. Parvathy, S. (2025) https://BioRender.com/qed8cnw.

Castalagin

Castalagin is a naturally occurring polyphenol that demonstrates promising prebiotic properties by promoting the growth of immunostimulatory gut bacteria [45]. This microbial modulation enhances immune activation by increasing key immune cell populations such as CD8⁺, CD4⁺, and FOXP3⁺ T-cell populations, which are critical for effective anti-tumor immunity. Notably, castalagin has been shown to enhance the efficacy of anti-PD-1 immunotherapy and may help overcome treatment resistance as observed in melanoma models [46]. These findings highlight castalagin’s potential as a natural prebiotic that not only supports gut health but also contributes to immune-mediated melanoma control.

Oral supplementation of camu camu (Myrciaria dubia), a polyphenol-rich berry containing castalagin, has demonstrated the ability to beneficially alter gut microbial composition, increasing the abundance of bacterial taxa associated with improved immunotherapeutic responses in melanoma [45]. In both mice and humans, castalagin supplementation induced a shift in the gut microbiota towards an increase in beneficial bacteria such as Ruminococcaceae and Alistipes, which are linked to favourable ICI outcomes [47].

Castalagin’s antitumour activity appears to result from the modulation of gut microbiota, enhancing the efficacy of anti-PD-1 antibodies even in antibiotic-treated mice reconstituted with fecal microbiota from ICI-resistant melanoma patients. To enhance the stability and bioavailability, camu camu is often processed through freeze-drying or spray-drying, which allows for its incorporation into foods such as yogurt, grape juice, candies, and bakery products [48]. Notably, camu camu seeds and peels, which are typically discarded as waste products, contain the highest concentrations of polyphenols (182.56 ± 1.76 mg/100 mL), offering an opportunity for sustainable repurposing within the camu camu industry [49].

However, despite its demonstrated immunomodulatory potential, the optimal dosage and human bioavailability of castalagin remain largely undefined. Current evidence is limited to preclinical and small translational studies, with no standardized intake guidelines or pharmaceutical knowledge available. Given castalagin’s polyphenolic nature and susceptibility to extensive first-pass metabolism, factors such as food matrix composition and formulation method are likely to influence its bioavailability and therapeutic potency. Further research optimizing delivery formulations and assessing variability will be critical to advance castalagin toward clinical application.

Inulin

Inulin shows significant promise in the context of melanoma by enriching antitumor microbial populations and enhancing the therapeutic efficacy of MEK inhibitors, particularly in BRAF-mutant melanoma, a subtype characterized by aggressive cell growth and poor prognosis [1, 40]. SCFAs produced during inulin fermentation play a pivotal role in maintaining immune tolerance and preventing excessive inflammatory responses. These metabolites modulate immune homeostasis and enhance targeted antitumor activity [1].

Additionally, molecular dihydrogen (H₂) produced during inulin fermentation can diffuse across the intestinal barrier, contributing to immune regulation by increasing CD4⁺ and CD8⁺ T-cell activity and inhibiting melanoma tumor growth [50].

Natural sources of inulin are the roots of chicory (Cichorium intybus) and yacon (Polymnia sonchifolia), as well as the tuber of dahlia (Dahlia pinnata) and Jerusalem artichoke (Helianthus tuberosus). Inulin with a higher degree of polymerization resists digestion longer, reaching the lower colon where it is fermented by gut microbes [51]. Regular intake of inulin (approximately 5 g per serving, three times daily) can significantly reshape gut microbial diversity within weeks, though research continues to focus on minimizing gastrointestinal discomfort at higher doses [51].

Galactooligosaccharides (GOS)

GOS exerts its prebiotic effects by activating mucin-glycan pathways and providing β-(1→4)-galactan as a nutrient source for beneficial bacteria. This supports microbial growth, enhances gut barrier integrity, promotes SCFA production, and strengthens mucosal immunity [52]. In cancer models, GOS-induced mucin activation has been linked to antitumor immune responses, particularly through increased efficacy of MEK inhibitors and delayed resistance. These effects are supported by upregulation of MHC class I/II molecules, chemokines (CCL4, CCL8), and immune-activating genes (CD40, Stat1, ICOS) [40]. β-GOS is commonly added to dairy products such as infant formula and yoghurt to enhance prebiotic content and support gut health, and is produced by industrial processes of modifying lactose [53]. However, non-fructosylated α-GOS derived from peas (Pisum sativum) represents a promising plant-based alternative, offering similar benefits with fewer gastrointestinal side effects, particularly by reduced gas production during colonic fermentation [54, 55]. Preclinical studies have reported beneficial effects at doses of approximately 5 g/kg body weight in animal models [56], while recent human supplementation trials indicate that GOS can exhibit prebiotic efficacy at doses as low as 1.3–2.0 g/day in healthy adults [57]. However, tolerance and clinical outcomes vary among individuals, as higher intakes may cause gastrointestinal discomfort, and differences in gut microbial composition that can alter metabolic response.

Fructooligosaccharides (FOS)

FOS are typically of shorter chain length than related sugars, such as inulin. FOS contributes to intestinal health and immune regulation, making it a promising adjunct for enhancing anti-melanoma therapies. By modulating cellular metabolism, reducing inflammation, and reinforcing tight junctions within the intestinal epithelium, FOS maintains gut integrity and supports immune activity [34]. Kinome analyses and calcium signaling studies suggest that FOS may boost kinase activity and intracellular calcium levels, which are essential for immune cell activation and antitumor responses [58].

Soybeans are a rich natural source of FOS, highlighting the potential of plant-based foods in promoting gut and immune health. FOS have also been incorporated into soy-kombucha fermentations to produce nutrient-enhanced, dairy-free alternatives with improved flavour and reduced environmental impact compared to animal-based dairy products [59]. Additionally, chamomile tea has been identified as a rich natural source of FOS, containing up to 20% of FOS by dry leaf weight [60], and the roots of Polymnia sonchifolia (yacon) provide another sustainable source for functional food applications [61].

While FOS exhibits prebiotic, gut-barrier, and immune-supporting activities, standardized human dosing and formulation bioavailability remain ongoing. Clinical studies have shown that supplementation at doses of approximately 7.5–15 g/day for four weeks or longer significantly increases Bifidobacterium abundance compared to lower doses [62]. Additionally, phase I trials have also demonstrated that FOS is well tolerated at up to 10 g/day, with only mild gastrointestinal symptoms reported [62].

Lyophilized mushroom extract (LME) and kale extract (KE)

Mushrooms, particularly Pleurotus ostreatus, serve as rich natural sources of prebiotics due to their bioactive polysaccharides like chitin, hemicellulose, α- and β-glucans, mannans, xylans, and galactans. These compounds resist acid hydrolysis and digestion in the upper gastrointestinal tract, which enables LME to promote SCFA production and beneficial bacterial growth, mirroring similar effects observed with inulin [63, 64].

When combined with KE, these natural prebiotics synergistically enhance the production of SCFAs such as succinic acid and butyric acid, which are compounds that are well known for their immunomodulatory and antitumor properties. KE contributes additional minerals and beneficial prebiotic carbohydrates such as FOS, amplifying the overall gut-supportive and immune-enhancing activity [65].

Human trials report immune-stimulatory effects at oral doses of 100–500 mg/day of β-glucans derived from LME, though absorption and efficacy are variable as they depend on molecular weight and branching structure [66]. These findings underscore its prebiotic and immunological potential in the context of melanoma. Human dosage data for KE in prebiotic and immunomodulatory applications remain unavailable, as most findings are derived from pre-clinical or dietary studies. Therefore, further investigations into formulation, bioavailability, and dose-response optimization are needed to confirm the synergistic effects of KE and LME in clinical settings. Overall, these extracts and their biological mechanisms demonstrate strong potential to reinforce gut-mediated immune responses relevant to melanoma therapy [67].

Konjac glucomannan (KGM)

Hydrolyzed KGM, particularly in the form of KGM oligosaccharides (KGOS), demonstrates more substantial prebiotic effects compared to its unhydrolyzed counterpart. Produced through enzymatic hydrolysis using Bacillus amyloliquefaciens WX-1 [68], KGOS is more readily fermented by anaerobic gut bacteria, generating SCFAs that promote gut and immune health. These SCFAs have been shown to improve spleen and thymus function, stimulate T and B lymphocyte proliferation, elevate serum hemolysin levels, increase phagocytic and NK cell activity, along with enhancing the secretion of immune-regulatory cytokines such as TNF-α, IL-2, and IgG [69]. Given melanoma’s reliance on immune activation for effective therapy, KGOS’s ability to activate systemic immunity underscores its therapeutic potential.

KGM is extracted from the tubers of Amorphophallus konjac, a plant valued for its high polysaccharide content and established use as a functional food. KGM is extracted through precipitation from dried konjac tubers, which are the primary edible and economically valuable part of the plant [70]. It is composed mainly of glucose and mannose units arranged in long chains. Given the widespread inclusion of probiotics in commercial dairy products, incorporating prebiotic compounds like KGM is a common symbiotic strategy for improving colonic and overall health [68].

According to the European Food Safety Authority, daily intake of approximately 3 g of KGM, typically divided into three 1 g doses, is generally well tolerated, although higher amounts may lead to mild gastrointestinal discomfort [71]. While these findings support its use as a safe, fermentable dietary fibre, clinical studies optimizing dose-response relationships and bioavailability in immunological or oncological contexts remain limited, urging further investigation in this area.

When comparing the prebiotics, notable differences in their mechanisms, levels of supporting evidence, and translational potential are observed. Oligosaccharides such as inulin, GOS, and FOS possess substantial in vivo and clinical results for modulating the gut microbiota and metabolic function, yet their influence on tumor immunity remains indirect. In contrast, castalagin and KGM demonstrate promising in vivo immunomodulatory effects, primarily through SCFA-mediated enhancement of gut-immune signaling. LME and KE further contribute synergistic prebiotic and immunological effects in preclinical studies, though standardized dosing and bioavailability data remain limited. Collectively, the comparison of these compounds underscores that while classical oligosaccharides provide strong clinical safety and efficacy profiles, novel polyphenol and polysaccharide-based prebiotics such as castalagin and KGM represent emerging candidates with distinct structural complexity and target immunomodulatory potential but require further mechanistic and clinical investigation before translation to oncology settings.