The identification of microbial networks that are predictive of disease progression and response to therapy will not only increase our understanding of the connection between microbiota and breast cancer, but also pave the way for the development of novel microbiota-based therapeutic interventions. The study by Di Modica and colleagues points to the existence of specific microbiota in patients with HER2+ breast cancer that can influence their response to trastuzumab. This information can potentially be used to develop novel therapeutic regimens combining fecal microbiota transplant with standard cancer therapy.

See related article by Di Modica et al., p. 2195

The quest to better comprehend the factors associated with cancer initiation, development, and response to therapy has led to the development of various predictive tools to guide cancer prevention and treatment strategies. Despite these advancements, development of cancer in people harboring no known risk factors, and disparate responses to cancer therapy in patients with similar clinical profile, are just a few questions that intrigue cancer researchers. Data from the NIH Human Microbiome Project and the Integrative Human Microbiome Project confirm the human microbiome as an important modifier of health and disease, and thus pave the way for studies investigating the connection between the microbiome and cancer. Distinct communities of microbes inhabit all the anatomical sites of the body, including the ones previously considered “sterile,” such as prostate, pancreas, and breast, and influence host cells by affecting various biological processes. Dysbiosis can particularly impact carcinogenesis by altering cellular metabolism, causing sustained inflammation and modulating the immune landscape. Multiple studies have shown that microbes can promote local as well as distant effects via the release of microbial toxins, and distinct microbial features can predict disease severity and clinical response in various cancers, including colon, oral, pancreatic, lung, liver, prostate, and breast. Analyses of whole-genome sequencing and whole-transcriptome sequencing from The Cancer Genome Atlas consortium encompassing 33 cancer types revealed unique microbial signatures in tissue and blood that are predictive of cancer stage. In addition, examination of plasma-derived cell-free microbial nucleic acids can distinguish cancer tissue from healthy tissue for many cancer types (1). Distinct sets of microbiota have been identified in tumor tissues compared with adjacent normal in several cancers and, interestingly, breast cancers harbor a remarkably rich and diverse microbiota (2). As breast cancer is a heterogeneous disease encompassing multiple subtypes, bulk analyses of breast tumors may provide an oversimplified picture, whereas comprehensive analyses of microbial communities associated with different breast cancer subtypes will uncover further complexities.

Another important aspect of the microbiota–cancer connection is the effect of microbiota on drug metabolism, pharmacokinetics, antitumor effects, and toxicity, ultimately altering the response to therapy. Most cancer drugs require enzymatic modification in the gut before entering the circulation for optimum absorption and bioavailability. Gut microbes are enzyme factories that can modulate the efficacy and toxicity of various chemotherapy drugs, though the exact mechanisms remain undetermined. Of interest, resistance and response to immune checkpoint inhibitors (ICI) can also be manipulated by gut microbiota. Distinct differences such as an abundance of Akkermansia muciniphila in the gut microbiota have been noted in responders compared with nonresponder patients. Fecal microbiota transplant (FMT) from responders or supplementation with Akkermansia muciniphila sensitizes nonresponder mice to ICI, exhibiting the functional impact of this microbe (3). With growing evidence pertaining to the impact of gut microbiota on cancer therapy, questions regarding the connections between breast cancer subtypes, targeted therapies, and gut microbiota are becoming more important. Can we define microbial features whose modulation can circumvent drug resistance and enhance drug efficacy?

In this issue, Di Modica and colleagues have taken a significant step toward addressing the impact of gut microbiota on the efficacy of trastuzumab in HER2-positive breast cancer, an aggressive subtype that constitutes approximately 15% to 20% of all breast cancer (4). Though trastuzumab has proven very effective in inhibiting HER2-driven tumors and improving overall survival, intrinsic and acquired resistance to trastuzumab in patients is still a major clinical challenge, especially at advanced stage. Various tumor-centric features such as dysregulation of downstream signaling cascades (e.g., PI3K-AKT), bidirectional cross-talk with transcription factors, and activation of escape networks (e.g., MET, SRC, FAK) have been identified as mediators of HER2-targeted therapy resistance (5). Taking a host-centric approach, Di Modica and colleagues focused their attention on the gut microbiota and asked whether its composition can act as an extrinsic tumor feature capable of modulating trastuzumab efficacy. Indeed, tumor-bearing mice whose gut microbiota was altered with vancomycin or streptomycin exhibited poor response to trastuzumab in comparison with mice harboring native microbiota. As expected, gut microbiota was significantly altered by antibiotic administration, resulting in reduced bacterial taxonomic richness. Reduction in the relative abundance of Actinobacteria and Firmicutes and an increase in the relative abundance of Proteobacteria and Verrucomicrobia were noted in the antibiotic-treated mice. As some of these depleted microbes produce short-chain fatty acids, their reduced abundance decreased the fecal levels of butyrate, propionate, and acetate. Intriguingly, gut dysbiosis owing to antibiotic administration considerably modulated the tumor immune microenvironment with increased CD45+ positive cells and Gr-1+ myeloid cells, and a concurrent reduction of CD3+ lymphocytes in tumor tissue. Perturbations of gut microbiota also altered the immune landscape known to associate with antitumor activity of trastuzumab. Further exploration of intestinal mucosal immunity revealed a large number of differentially expressed genes among the groups and enrichment of pathways related to antigen presentation via MHC class II, response to IFNγ, and IgGA production in mice with native gut microbiota. Investigating the changes in systemic cytokine and chemokine levels, the research team uncovered higher levels of CCL11 and CCL7, and lower levels of CCL5 and IL12p70, in the antibiotic-treated mice compared with control mice. Interestingly, it was noted that neutralization of IL12p70 through an anti-IL12p70 mAb reduced the efficacy of trastuzumab in control mice. Conversely, recombinant IL12p70 (rIL-12p70) administration improved antitumor activity of trastuzumab in antibiotic-treated mice, underlining IL12p70 as the key functional node. Together, these data strongly suggest that the gut microbiota–dendritic cell activation axis modulates the effectiveness of trastuzumab by controlling natural killer cell activation and recruitment via an IL12p70-dependent pathway. These studies represent an important step toward establishing gut microbiota as a potential modifier of trastuzumab efficacy in HER2+ breast cancer.

Furthermore, patients who responded to trastuzumab and experienced pathological complete response exhibited higher gut microbiota diversity compared with patients with residual disease. Gut microbiota in responders was enriched with Bifidobacteriaceae, Clostridiales, Turicibacteraceae, and Bacteroidales, whereas nonresponders exhibited an abundance of Bacteroidetes. It will be interesting to know whether microbiota profiling can help predict response to therapy and distinguish between responders and non-responders in future. Distinct features of gut and breast microbiota pertaining to breast cancer subtype and response to therapy have been reported (6, 7), but effectiveness of FMT as an adjuvant to anti-breast cancer therapy has never been examined. Another major finding of this study was that FMT from responders enhanced the effectiveness of trastuzumab in recipient mice bearing HER2+ breast tumors, further highlighting the connection between gut microbiota and trastuzumab efficacy against HER2+ breast tumors. Although this is the first study to investigate the effect of FMT with trastuzumab in breast cancer, FMT is being actively investigated for other cancers. Increased efficacy of immunotherapy and significant tumor inhibition in recipient mice was observed in response to FMT from patients with cancer that responded to PD-1/PD-L1 therapy in contrast with nonresponders, emphasizing the functional significance of gut microbiota (3, 8). Some preclinical studies have reported that FMT from healthy donors offers protection against chemotherapy-induced toxicity and modulates anti-inflammatory function in colon cancer (9); contrastingly, FMT from patients with colon cancer activates pro-oncogenic signaling pathways, elevates the levels of proinflammatory cytokines, and increases the tumor burden in recipient mice (10, 11). Multiple clinical trials are being conducted to evaluate the effectiveness of FMT in circumventing therapy-related side effects as well as improving response to anticancer therapy. Although therapeutic importance of FMT has been established for inflammatory bowel diseases, Clostridium difficile infections, and intractable functional constipation, development of FMT as an adjuvant to standard anticancer therapy is still in its infancy.

Many important questions remain regarding the clinical use of FMT as adjuvant therapy in cancer, such as how to identify healthy donors, how to ensure the reproducibility of donor–microbiome, and, most importantly, what should be the frequency and timing of FMT. Response to therapy is guided by a combination of tumor characteristics and host-centric features, and it is therefore important to decipher whether FMT from a single responder would be optimum or if a “fecal cocktail” from multiple responders might be more useful. Furthermore, careful analysis of donor microbiomes is essential to prevent accidental introduction of pathogens. Several routes of FMT administration are being developed, including capsule, nasogastric tube, nasoduodenal tube, enema, and colonoscopy. In addition, guidelines pertaining to FMT are progressively being fine-tuned by the FDA and other institutions. There are several unique advantages to this approach, as it may help improve the efficacy of cancer drugs as well as reduce therapy-related side effects.

Our understanding of the connections between microbiota and cancer has improved significantly in recent years and has helped build up our appreciation of the host microbiota and its direct/indirect effects on clinical outcomes. New studies have revealed key microbes with harmful or beneficial roles in cancer progression and response to therapy. Multiple strategies—including prebiotics, probiotics, postbiotics, synbiotics, and dietary modulations, in addition to FMT—are under development to fine-tune the microbiome. With the development of novel strategies to characterize native gut and target-tissue microbiota, to decipher its functional interactions with gene-networks, immune-landscape and circulating metabolome, and to successfully modulate one's microbiota, a picture emerges in which microbiota could be used as a unique tool to better personalize cancer treatment.

No disclosures were reported.

This work was supported by grants NCINIHR01CA204555, Breast Cancer Research Foundation (BCRF) 90047965, CDMRP BCRPW81XWH-20-1-0626 (to D. Sharma).

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