Since 1975, the global prevalence of obesity has tripled, with increases occurring across developed and developing countries. According to the latest World Health Organization (WHO) estimates, more than 1.9 billion adults were overweight and 650 million adults were classified as obese (W.H.O, 2021). Overweight and obesity reflect an excessive accumulation or abnormal distribution of fat and are classified according to the body mass index (BMI). BMI between 25 and 30 kg/m² defines overweight whereas BMI ≥ 30kg/m² corresponds to obesity. Over the past decades, trends toward increased intake of calorie-dense foods rich in fat and sugar, and decreased physical activity explain, in part, the obesity pandemic (W.H.O, 2021). Obesity is associated with chronic diseases such as type 2 diabetes, hypertension, dyslipidemia, cardiovascular diseases, non-alcoholic fatty liver disease, and 13 different cancer types, including postmenopausal breast cancer (Avgerinos et al., 2019; Włodarczyk and Nowicka, 2019; Dwivedi et al., 2020; Brown, 2021).

Obesity increases the risk of developing postmenopausal breast cancer by up to 50% (Lauby-Secretan et al., 2016; Chan et al., 2019; Brown, 2021). Further, breast cancer patients (regardless of subtype and menopausal status) with obesity also have poorer overall survival, reduced response to chemotherapy and endocrine-targeted therapies, increased risk of local recurrence and metastasis, and often develop dose-limiting comorbidities (Ewertz et al., 2011; Rock et al., 2015; Iwase et al., 2016; Picon-Ruiz et al., 2017). Nevertheless, the mechanistic links between obesity, lipid metabolism, and breast cancer initiation and progression are still poorly characterized. The obesity-associated increase in both breast cancer risk and progression is thought to be regulated by several systemic and localized factors (Brown, 2021). These mechanisms include inflammation and immunosuppression, hypoxia of breast tissue, increased aromatase-mediated estrogen conversion, perturbations in the adipokines leptin and adiponectin, and changes in insulin signaling (Gunter et al., 2009; Ollberding et al., 2013; Himbert et al., 2017). Intriguingly, all these potential risk factors are also influenced by the gut microbiome ( Figure 1 ), suggesting a critical relationship between microbes in the gut and obesity-driven breast cancer burden, which we will discuss in more detail in this review.

The gut microbiome is a dynamic and functional entity that is vital for the health of its host. It is shifted by host factors such as age, diet, lifestyle, xenobiotic agents, and disease state. Functionally, gut microbiota has an essential role in host physiology, including digestion of plant polysaccharides, biosynthesis of essential vitamins, detoxification of environmental pollutants, maintenance of the intestinal epithelial barrier function, enhancement of the immune function, and out-competition of pathological bacteria (Hansen and Sams, 2018; Heintz-Buschart and Wilmes, 2018; Rowland et al., 2018). The gut microbiome is composed of fungi, archaea, protists, viruses, and bacteria. The latter two account for >99% of the gut microbiome (Liang and Bushman, 2021; Zhang et al., 2022). Bacteroidetes and Firmicutes are the two major phyla of the gut microbiome, accounting for >90% of its composition (Plottel and Blaser, 2011; Tilg and Kaser, 2011). This distribution of bacteria is similar in mice, which are frequently used in gut microbiome studies. Mice and humans also share qualitative similarities in their gut microbiomes at the genus level (Krych et al., 2013). Non-human primates share even more similarities with the human microbiome; especially those in captivity and fed a human-like diet, suggesting that non-human primates may be a superior model to study the microbiome’s impact on health in humans (Nagpal et al., 2018).

Dysbiosis, or perturbations in the gut microbiome, is associated with the development of inflammatory, autoimmune, and malignant diseases, which can occur locally or at distant tissue sites. Many pathophysiologic conditions cause dysbiosis such as inflammatory bowel disease, diabetes mellitus, and obesity (Carding et al., 2015). Obesity alters the diversity and the relative abundance of microbes in the gut. Evidence supporting obesity-associated dysbiosis is from analyses of genetically obese leptin-deficient (ob/ob) mice. 16s ribosomal RNA (rRNA) sequencing revealed a major increase in the Firmicutes to Bacteroidetes (F/B) ratio (a marker of dysbiosis) of the ob/ob mice compared to their lean wild type (WT) siblings fed the same diet (Ley et al., 2005). Similar phyla-level changes were found in mice with diet-induced obesity (DIO) (Turnbaugh et al., 2008), as well as in obese humans (Ley et al., 2006). An unfavorable signature that has been frequently associated with obesity is the expansion in Proteobacteria phyla (Xu et al., 2022). It is imperative to mention that studies on mice with DIO showed a more consistent increase in the F/B ratio than studies on obese humans (Bisanz et al., 2019). This might be attributed to the experimental modeling in mice where the majority of studies have used chow as a control diet which has different dietary composition than the high fat diet (Dalby et al., 2017). Hence, this adds dietary composition differences on top of the adiposity differences between mice groups that could have confounded the microbiome sequencing results. Yet, in addition to the effect of diet, body adiposity within a dietary pattern further modified the gut microbiome in non-human primates, demonstrating that obesity alters the gut microbiome (Newman et al., 2021). Mice with DIO showed a marked decrease in microbial diversity in comparison to their lean counterparts (Turnbaugh et al., 2008). Human studies on obese individuals also revealed a marked decrease in microbial diversity (Le Chatelier et al., 2013).

While adiposity influences the gut microbiome, the inverse is also true. Mice with an intact gut microbiome showed a 42% increase in adiposity compared to germ-free mice, despite a 29% higher food intake by the germ-free mice. Accordingly, gut microbiota transplantation from conventionally-raised (lean) mice to germ-free mice caused a 60% increase in adiposity despite the decreased food intake (Bäckhed et al., 2004). Furthermore, germ-free mice were more resistant to DIO than conventionally raised mice, demonstrating how integral the microbiome is in the obesity pathogenesis process (Bäckhed et al., 2007). Interestingly, transplantation of gut microbiota from obese and lean mice caused differing degrees of adiposity in germ-free mice, with “obese microbiome” transplants causing a greater increase in adiposity than the “lean microbiome” transplants. The “obese microbiome” increased the capacity for energy harvest as demonstrated by gene enrichment for enzymes in pathways involved in galactose metabolism, starch/sucrose metabolism, and butanoate metabolism (Turnbaugh et al., 2006). Hence, obesity and gut microbiome perturbations have a two-way relationship and the perturbations to the gut microbiota caused by obesity are major drivers for obesity pathogenesis.

Mounting evidence suggests a causative role of the gut microbiome in carcinogenesis. Although various microorganisms are associated with different cancer types, none to date were demonstrated to be causative for breast cancer (I.A.R.C., 2023). However, experiments with germ-free mice and rats revealed the tumor-promoting effects of the gut microbiome. Animals with an intact gut microbiome had more spontaneous, genetically-induced, and carcinogen-induced tumors compared to germ-free counterparts in various organs including the lung (Schreiber et al., 1972), liver (Dapito et al., 2012; Yoshimoto et al., 2013), skin (Sacksteder, 1976), colon (Grivennikov et al., 2012), and mammary gland (Mishra et al., 2021). Additionally, gut dysbiosis induced by obesity was associated with enhanced tumor growth and significant loss of gut microbial diversity in a murine triple-negative breast cancer (TNBC) model (Hossain et al., 2021). Moreover, multiple studies have shown that the gut microbiome is different in malignant breast disease, benign breast disease, and control. These observations are summarized in Table 1 ; which include all articles identified by performing a systematic search of PubMed articles published until February 22, 2024, for combinations of search terms: “Obesity”, “Gut microbiome”, and “Breast Cancer”. Mechanisms proposed for dysbiosis-associated breast cancer risk include alteration of normal tissue metabolism, induction of chronic inflammation, direct genotoxicity, and modulation of immune responses (Plottel and Blaser, 2011; Schwabe and Jobin, 2013; Argolo et al., 2018).

Changes in gut microbiome composition and diversity have been documented in breast cancer. A pilot study by Bertazzoni et al. performed using simple culturing techniques (predating current sequencing methods) found significant increases in Bacteroides, Clostridia, and anaerobic Lactobacilli in breast cancer patients compared to healthy controls (Bertazzoni et al., 2006). Later studies of the gut microbiome using 16S rRNA or metagenomic sequencing affirmed the initial findings of dysbiosis in breast cancer patients ( Table 1 ). Most studies show a significant reduction in alpha diversity which represents the compositional complexity of a single sample (intra-sample heterogeneity, Box 1), in breast cancer patients compared to cancer-free controls ( Table 2 ). This is consistent with loss of microbiome richness and evenness, which has been associated with unhealthy gut environments, aging, and disease state (Hou et al., 2021). For beta diversity, which corresponds to the taxonomical or phylogenetic differences between samples (inter-sample heterogeneity, Box 1), an opposite trend is observed where most of the studies showed significant increases in beta diversity ( Table 3 ). This reflects an increased heterogeneity between samples of breast cancer patients compared to healthy women. This may be due to inter-individual differences in breast cancer stages, grades, hormone receptor status, HER2 status, and proliferation levels which are factors that differentially impact the gut microbiome (Luu et al., 2017; Wu et al., 2020; Yang et al., 2021). For instance, clinical stages II and III had significantly higher levels of Bacteroidetes, Blautia, Clostridium coccoides, and Faecalibacterium prausnitzii compared to stages 0 and I (Luu et al., 2017). Also, patients with non-malignant breast disease had an altered microbiome, different from that of breast cancer patients (Luu et al., 2017). Another potential confounder that is often overlooked in these comparisons is dietary differences that might have existed between breast cancer and cancer-free subjects (Ma et al., 2020).

During menopause, the depletion of cycling estrogens can adversely impact physiological systems, including gut health (Peters et al., 2022). A comparison of the microbiome of non-obese pre vs. postmenopausal women (without breast cancer) found significant differences in β-diversity, demonstrating that menopause shifts the gut microbiome. Furthermore, postmenopausal women also have higher Firmicutes proportional abundance, higher F/B ratio, and increased levels of pro-inflammatory cytokines, including interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1). These differences could not be explained by the age difference since they were not observed in male control groups of the same age. Moreover, subjects were matched for their BMI and nutritional background. Hence, this strongly suggests the fundamental interactions between estrogen, microbiota, and inflammation (Santos-Marcos et al., 2018).

Several studies show altered gut microbiota populations in breast cancer patients depend on menopausal status. Bertazzoni et al. (2006) divided the breast cancer study participants according to their menopausal status and found that the genera and species cultured from each group were remarkably different from one another and from the healthy controls. Another study found significant differences in gut microbiota composition and diversity in postmenopausal (vs. matched healthy controls) but not in premenopausal women (Zhu et al., 2018). In particular, this group found a positive correlation between Shewanella putrefaciens and Erwinia amylovora with estradiol (p< 0.05) in postmenopausal patients. This is consistent with gut microbiota interactions with estrogen metabolism, suggesting a potential biomarker for breast cancer. Roseburia inulinivorans, a butyrate-producing bacteria, was found to be lower in postmenopausal breast cancer patients. Butyrate acts as an anti-inflammatory agent, by inhibiting the activation of nuclear factor-κB (NF-κB) in intestinal epithelial cells (Inan et al., 2000). Therefore, this reduction in R. inulinivorans may indicate postmenopausal women are more prone to inflammation and therefore at higher risk of breast carcinogenesis (Zhu et al., 2018). Another study demonstrated a significant difference in β-diversity between breast cancer patients and age-matched controls as well as a significant reduction in α-diversity in the premenopausal breast cancer group compared to controls (Hou et al., 2021). However, while premenopausal and postmenopausal patients had similar BMIs, BMIs were not provided for the age-matched controls. These comparisons may therefore not be controlled for adiposity. Using functional pathways analysis, the gut microbiota of premenopausal breast cancer patients showed enrichment in steroid-related aromatic and androstenedione degradation, which may result in DNA damage induction and, subsequently, breast cancer development (Heikkinen et al., 2015; Hou et al., 2021). Moreover, gut microbes of postmenopausal breast cancer patients showed enrichment in chemical carcinogenesis and aldosterone-related pathways. This could be attributed to the lower estrogen levels post-menopause which has been shown to increase aldosterone levels and, thus, may increase breast cancer risk (Rigiracciolo et al., 2016; Hou et al., 2021). Different gut microbiome compositions in premenopausal women with vs. without breast cancer were confirmed by another study (He et al., 2021). Collectively, these studies show that the gut microbiome is differentially regulated in breast cancer based on menopausal status. Yet, discrepancies in the altered microbes and diversity changes call for further investigations. Differences in other variables such as obesity status, race/ethnicity, age, diet, environmental exposures, sequencing methodology, and sample size are a few of the potential confounders.

Few clinical studies have investigated the role of obesity-modulated gut microbiome in breast cancer. Luu et al. found that the gut microbiome composition in breast cancer patients differs according to BMI (Luu et al., 2017). The bacterial load was lower in obese/overweight patients compared to patients with normal weight. Among other differences, the abundance of Faecalibacterium prausnitzii, was significantly lower in the obese/overweight group. Interestingly, these bacteria produce butyrate, an anti-inflammatory short-chain fatty acid (SCFA), which will be discussed in detail below. In another study, breast cancer patients in the obese/overweight category showed significant enrichment of Clostridiaceae family and Akkermansia genus, and a significant reduction of Lactobacillus and Streptococcus genera (Wu et al., 2020). Total body fat also impacted the microbiome; patients with higher body fat had fewer detectable operational taxonomic units (OTUs, Box 1) and lower alpha diversity. These patients also had significant enrichment in Clostridium genus, and Lachnospira genus, and a significant reduction in Catenbacterium genus. These findings raise the question of whether differences in the gut microbiome between BMI or body fat categories can explain the worse prognosis and lower survival of obese breast cancer patients. A consensus on how obesity may influence microbial populations in the context of breast cancer has not been reached, and further investigations are needed.

The gut microbiomes of obese/overweight breast cancer patients were compared to the microbiomes of BMI-matched cancer-free women by Smith et al. (2021). While there was no significant difference in alpha or beta diversity, differences in gut microbiota were observed. Compared to BMI-matched cancer-free women, bacterial genera such as Phenylobacterium and Balneimonas were significantly reduced in obese/overweight breast cancer patients, while Allobaculum was significantly enriched. Taken together, these findings indicate a two-way relationship between tumorigenesis and gut microbiome composition, where the obesity-modulated microbiome increases breast tumorigenesis and the presence of a tumor imposes a selective pressure on the gut microbiome in obese patients. The latter is corroborated by an animal study on tumor-bearing and tumor-free obese mice, where the presence of the tumor modified the gut microbiome (Hossain et al., 2021). This is true in lean mice as well, where the presence of mammary tumors perturbs microbiome composition, compromises intestinal barrier function, increases translocation of gut bacteria, and induces systemic inflammation (Loman et al., 2022). Additionally, high-risk obese individuals may possess a pro-tumorigenic microbial signature prior to breast cancer development. Using a chemical mammary carcinogenesis model and manipulation of the gut microbiome with fecal microbiota transplants (FMT), we showed that intestinal microbes derived from lard-fed (obese) mice decreased tumor-free survival in lean animals (Soto-Pantoja et al., 2021). Reciprocally, obese mice benefited from FMT from lean animals, with lower tumor burden and increased survival. In a murine TNBC model, FMT from non-tumor bearing mice with DIO increased tumor growth in recipient mice (Bawaneh et al., 2022). These results demonstrate that the pro-tumorigenic effects of the lard diet are, at least in part, caused by shifts in the gut microbiome. Further research is required to determine whether obesity-mediated dysbiosis is a cause and/or a consequence of breast tumorigenesis. Potential mechanisms linking obesity, dysbiosis, and breast carcinogenesis are illustrated in Figure 2 and discussed in the following sections.

In 2014 Urbaniak et al. (2014) established the existence of a breast microbiome and inspired more than a dozen studies demonstrating dysbiosis in breast cancer that are well-summarized in ref (Peters et al., 2023). Differences in the breast microbiome were also found between breast cancer patients and patients with non-malignant breast disease (Hieken et al., 2016; Urbaniak et al., 2016), between breast tumors and paired tumor-adjacent normal tissue (Thompson et al., 2017; Smith et al., 2019; Esposito et al., 2022), and between breast cancer survivors and women who never had breast cancer (Chan et al., 2016; Klann et al., 2020). An interesting comparison of beta diversity was done by Costantini et al. (2018) between three groups: 1) paired tumors and tumor-adjacent normal tissues, 2) tumors of different subjects and 3) tumor-adjacent normal tissues of different subjects. The beta diversity was significantly lower in paired tumors and normal-adjacent tissues within the same subject than in tumors or normal-adjacent tissues of different subjects. This shows that more similarities than differences exist in the microbiome of tumors and normal-adjacent tissues within individuals. Hence, it may indicate that dysbiosis in the breast is antecedent to tumor initiation by establishing a pro-tumorigenic microenvironment.

Obesity induces microbiome perturbations in the breast tissue. Our group has demonstrated that obesity modifies tumoral microbiome populations in the breast (Chiba et al., 2020). We have also shown that an obesogenic Western diet perturbs the breast microbiome in non-human primates. Obesogenic Western diet disturbed non-cancerous breast tissue homeostasis by significantly decreasing bile acid levels and increasing oxidative stress; mechanisms that are associated with increased breast cancer risk (Shively et al., 2018). More studies are needed in the breast cancer initiation front to identify the pre-malignant changes that are likely to occur in obesity due to breast microbiome perturbations.

In conclusion, obesity modulates the gut microbiome in ways that may increase breast cancer risk. Carcinogenic mediators communicate gut microbiome changes in obesity to the breast. These mediators include circulating LPS, SCFAs, estrogens, IGF-1, and DCA which can influence molecular signaling at distant tissue sites such as the breast. However, studying the impact of these mediators on breast cancer risk needs to be carried in the context of obesity to prove causality.

A major challenge in gut microbiome research is the lack of reproducibility between studies. Although some findings align well, there are many differences and contradictions ( Table 1 ). For instance, compositional differences in the gut microbiome between premenopausal breast cancer patients and healthy controls were reported by He et al. (2021), while no differences were found by Zhu et al. (2018). Although both study populations were Chinese; the methodologies used for microbiome identification and inclusion criteria were different. The former used 16s rRNA sequencing while the latter used metagenomic sequencing. Inclusion criteria were also different; the former study excluded patients exposed to antibiotics within one month of fecal sample collection while the latter had a wider exclusion window of three months. Another example is the increase in F/B ratio in breast cancer patients vs. healthy controls observed by Bobin-Dubigeon et al. (2021) but not by Byrd et al. (2021). The discrepancy could be explained by the different populations studied (European vs. African) and the methodologies used. The former study used RT-qPCR to quantify bacteria copy numbers, while the latter used 16s rRNA sequencing.

Overall, several factors may (at least partly) explain discrepancies between gut microbiome studies, including differences in study population (geography, BMI, race/ethnicity, diet, xenobiotic exposure, inclusion/exclusion criteria), sample handling (collection method, storage time, preservatives, external contaminants), experimental protocols (DNA extraction, library preparation, sequencing methodology, reference database), bioinformatics pipelines, and statistical analyses. How variability in these factors leads to different findings and potential solutions for them are comprehensively illustrated in this review (Nearing et al., 2021). Ideally, a universal standardized protocol for microbiome studies should be adopted to eliminate many of the aforementioned variability.

Future research on the interactions between obesity and the gut microbiome on breast cancer development is needed. A better understanding is needed of how an individual’s gut microbiome is influenced by a combination of factors such as obesity, diet, and genetics and how such combinations affect treatment responses. This could lead to personalized treatment strategies that consider the patient’s microbiome. Additionally, further investigation into microbial metabolites specific for obesity, immune system interactions, and hormonal pathways that play a role in breast cancer development and progression are needed. Finally, the determination of obese microbial markers or signatures that can be used for the identification of high-risk individuals and early detection of breast cancer will aid the development of prevention strategies and early interventions. Advancements in these areas will have a positive impact on the breast cancer incidence and mortality of obese populations.

MG: Writing – original draft. AA: Writing – original draft. P-AV: Writing – review & editing. KC: Writing – review & editing.