The gut microbiome modulates response to cancer immunotherapy

Abstract

With the advent of next generation sequencing approaches, there has been a renaissance in the microbiome field. Microbial taxonomy and function can now be characterized relatively easily and rapidly - no longer mandating complex culturing approaches. With this renaissance, there is now a strong and growing appreciation for the role of the microbiome (referring to microbes and their genomes) in modulating many facets of physiology - including overall immunity. This is particularly true of the gut microbiome, and there is now an evolving body of literature demonstrating a role for gut microbes in modulating responses to cancer treatment - particularly immunotherapy. Gut microbes can modulate immunity and anti-tumor responses via a number of different interactions, and these will be discussed herein. Additionally, data regarding the impact of gut microbes on cancer immunotherapy response will be discussed, as will strategies to manipulate the microbiome to enhance therapeutic responses. These efforts to date are not completely optimized, however there is evidence of efficacy though much additional work is needed in this space. Nonetheless it is clear that the microbiome plays a central role in health and disease, and strategies to manipulate it in cancer and overall precision health are being explored.

Gut microbes and their impact on immunity

Direct molecular and cellular interactions

With the introduction and colonization of the microbial species in the gut ecosystem, the host immune system has evolved to maintain a mutualistic relationship between microbes and host immunity [1]. Gut microbiota themselves have a tremendous impact on host physiology, and play a central role in the education and function of the host immune system. In turn, our immune system may shape the composition and function of the microbiota [2].

Gut microbes may interact with the host via a number of different mechanisms (Figure 1). This includes through pathogen-associated molecular patterns (PAMPs) such as lipopolysaccharide and flagellin, which are recognized by pattern recognition receptors (PRRs; such as Toll-like receptors) located on the surface of intestinal epithelial cells [3]. Commensal bacteria can prime DCs which in turn can signal Toll-like receptors (TLRs) to educate the immune system regarding differential recognition of pathogenic vs non-pathogenic microbes. PAMPs from the gut may also interact directly with immune cells, inducing activation and maturation of antigen-presenting cells (APCs) including dendritic cells (DCs). These APCs may then translocate into mesenteric lymph nodes and mediate priming of B and T cells, which then can disseminate systemically and induce the differentiation of naïve T cells into CD4+ T cells [4] which function at distant sites [5].

Figure 1: Gut microbes and host immunity.

Bacterial metabolites, anti-microbial peptides and bacteria can prime dendritic cells (DCs) which migrate to the lymph nodes to stimulate T and B cells. PAMPs from the gut may also interact directly with immune cells, inducing activation and maturation of antigen-presenting cells (APCs) including DCs. These APCs may then translocate into mesenteric lymph nodes and mediate priming of lymphocytes, and disseminate systemically and induce the differentiation of naïve T cells. These specific microbes or metabolites can prime DCs that in turn skew the response towards immune suppression or immune activation

Additionally, gut microbes modulate systemic immune responses via soluble immunomodulatory factors, circulating cytokines that can alter the activity of the immune cells. Preclinical data suggests segmented filamentous bacteria (SFB) can induce secretion of IL-22 from ILC3 cells which drive TH17 polarization in lamina propria DCs via production of serum amyloid A [6,7].

Furthermore, commensal bacteria provides a stimuli for the maturation of the Gut-associated lymphoid tissue (GALT), inducing B cell differentiation, maturation and activation [8–11] and IgA production [12]. The importance of this gut microbiota exposure to mucosal immunity is highlighted by the substantially defective gut mucosal immune systems seen in gnotobiotic mice characterized by a thin gastrointestinal mucus layer and aberrant IgA release in the gut [13,14].

There is also extensive data substantiating the role of gut microbe metabolites in modulating immune responses. Gut microbes produce a complex array of metabolites via utilization of the nutrients derived from the diet and other sources, and these can in turn influence the host immune system by receptor-specific binding to host cells. Examples of metabolites that may impact immunity include short chain fatty acids (SCFAs) and polyamines. SCFAs including acetate, propionate and butyrate and are produced by bacterial fermentation in the gut from dietary fibers that remain undigested or partially digested by the host. As the most abundant microbial metabolites in the intestine, SCFAs plays several crucial roles in host immunity [15]. First, these serve as the major energy sources of intestinal epithelial cells and maintain gut barrier integrity [16]. Prior work has demonstrated that association of acetate producing bacterium Bifidobacterium longum prevents the translocation of toxins from enteropathogenic Escherichia coli O157:H7 into the systemic compartment [17] SCFA may bind to G-coupled protein receptors (GPR43 and GPR41) of intestinal epithelial cells altering host gene expression and inducing autophagy and stimulating production of anti-inflammatory cytokines.. SCFAs can also influence innate immune cells. In the peripheral circulation, SCFAs deactivates nuclear factor-κB (NF-κB) and abrogate the expression of the pro-inflammatory cytokine tumor necrosis factor in mononuclear cells and neutrophils [18,19]. They also mediate immune cell phenotype through epigenetic mechanism, by inhibiting histone deacetylases (HDACs), with recent work demonstrating that this mechanism can promote CD8 memory cell differentiation [20]. Finally, SCFAs also influence antigen-specific adaptive immunity by stimulating the synthesis of IgA by B cells [21,22]. Furthermore, compounds such as tryptophan, indole, and bile acids may interact with aryl hydrocarbon receptor (AhR) and pregnane X receptor (PXR), thus modulating expression of inflammatory genes, upregulainge the expression of tight junctions, and inducing cytokine production by AhR expression lymphoid cells. In contrast, AhR expression on Tregs affects their homing and suppressive activity [23].

Polyamines which can be of dietary, host or microbial sources, are also important players in host immunity. Like SCFAs, polyamines such as putrescine, spermidine and spermine are important in mucosal immunity [16], inducing the secretion of IgA in the gut and maintaining barrier integrity [24,25]. Preclinical data showed that elevated level of polyamine in breastmilk promoted the maturation of gut CD8+ and CD4+ T cells [26]. Oral administration of the polyamine-producing bacterial strain Bifidobacterium animalis subsp. lactis LKM512 along with diet supplemented with the substrate (arginine) has been shown to decrease levels of intestinal TNF and IL-6 [27].

A specific group of gut microbiota, primarily Lactobacillus spp., produces metabolites by metabolizing dietary tryptophan [28]. These metabolites bind to the aryl hydrocarbon receptor (AHR) molecules expressed on the immune, epithelial and some tumor cells, demonstrating important role in regulating colonic inflammation [29]. In addition, the recognition of AHR by microbial metabolites was demonstrated to activate IL-22-producing ILC3 cells [30].

This body of literature demonstrating the profound effects that the gut microbiota exert on both mucosal and systemic immunity thus lays the foundation for investigations into the effects of the gut microbiota on anti-tumor immunity. Dedicated studies assessing metabolic profiles in the gut microbiome and peripheral circulation of patients on immunotherapy are currently underway, and are likely to reveal important insights into functional mechanisms through which gut microbes impact overall immunity and therapeutic response.

Pre-clinical studies substantiating gut microbes in response to immunotherapy

The earliest evidence regarding the role of the gut microbiome on response to immunotherapy was conducted in pre-clinical models. As noted above, there is a vast body of preclinical literature which has established the key role the gut microbiome plays in shaping mucosal as well as systemic immunity. Some of the earliest work indicating that the gut microbiome can also play a role in cancer immunology came from work with the chemotherapeutic cyclophosphamide[31]. Modulating the immune response is a key mechanism of action of cyclophosphamide. One mechanism through which this immune induction occurs is via translation of gram-positive bacteria into secondary lymphoid tissues which then stimulate Th17 and Th1 immune response [31]. Work in preclinical models of adoptive cellular therapy (ACT) has further demonstrated that gut microbiota modulation can help sustain transferred T cells in an IL-12 dependent manner, further supporting a role for the gut microbiota in adaptive anti-tumor immune responses [32].

Two ground-breaking preclinical studies published in Science in 2015, showed that mice with a favorable microbiota had enhanced response to immune checkpoint blockade (ICB) therapy compared to mice with an unfavorable microbiome - and furthermore that the beneficial phenotype was transferable by fecal transplant or cohousing [33,34]. More recent work has demonstrated that fecal microbiota transplant (FMT) from humans who responded versus did not respond to ICB can induce differential response to immunotherapy in mice [35,36]. In these studies, mice who received FMT from complete responder to ICB in turn had tumor response to ICB as well as increased tumor immune infiltrates whereas anti-tumor immunity was not induced in mice who received non-responder donor FMT. The precise mechanisms by which anti-tumor immunity is induced by the gut microbiota remains an area of active investigation. Beyond FMT, other work has focused on trying to define the specific bacterial consortia critical to the induction of anti-tumor immunity, with this work now being translated into clinical trials testing these consortia [37]. Other provocative work has demonstrated that the intratumoral microbiome can also influence response to therapy and can in turn be modulated via the gut microbiome[38,39]. Far from being sterile, pancreatic cancer has a distinct microbiome, apparently as a result of gut translocation[40]. FMT from long-term human survivors of pancreatic cancer into mice was shown to modulate not just the gut microbiome but also the tumor microbiome, as well as shape anti-tumor immunity[41]. Intratumoral bacteria may also induce chemotherapy resistance via in situ drug metabolism[42].

Clinical cohorts and observational studies on gut microbes and response to immunotherapy

These pre-clinical studies laid the foundation for subsequent observational studies in clinical cohorts. The initial evidence that the gut microbiome plays a role in treatment response in cancer were from studies on allogenic hematopoietic stem cell transplant (SCT) where significant association were observed between diversity of the gut microbiome and long-term outcomes in the setting of treatment of acute myelogenous leukemia (AML) with SCT [43–45]. Improved survival was demonstrated in patients with a high diversity of the gut microbiome compared to those with an intermediate or low diversity. Subsequent studies focused on the risk of GVHD in the setting of SCT, showing differences in rates of GVHD depending on the composition of the gut microbiome, and also depending on antibiotic regimens used for potential infectious complications in the context of therapy[46].

More recently, several studies demonstrating strong associations between the gut microbiome and response to ICB in human cohorts across a variety of cancer types have been published[35,36,47–49], building upon the prior foundational preclinical work.

One of the first of these studies was published by Dubin and colleagues in 2016, and reported results from a cohort study of patients with metastatic melanoma undergoing ICB treatment, and focused on the associations of signatures in the gut microbiome with toxicity to therapy - namely immune checkpoint-associated colitis. In this cohort, the investigators reported that an increased abundance of bacteria belonging to the Bacteriodetes phylum was correlated with a decreased risk of ICB-induced colitis [50] (Table 1). This work was followed by another study authored by Chaput et al in 2017, which demonstrated that the gut microbiome of responders to ICB was distinct from that of non-responders; however, the cohort was somewhat small (n=26). The group found associations of baseline gut microbiota composition with both clinical response as well as colitis; whereby patients whose baseline microbiota was enriched with the Faecalibacterium genus and other Firmicutes, had longer progression free survival (P=0.0039) and overall survival (P=0.051) compared to those dominated by Bacteroides [48].

Table 1.

Observational studies of role of gut microbiome in modulating response to immunotherapy

Reference	Population	Study Model	n	Bacteria	Influence of Gut Microbiome in ICB Therapy	Outcome	Mechanism of Immune Modulation Proposed	Geography
Gopalakrishnan, V., et al., 2018	Melanoma	Patients	112	Ruminococcaceae	Enhanced efficacy of PD-1 blockade therapy	Higher abundance in responders	Increased antigen presentation Improved effector T cell function	Texas, USA
						Elevating levels of effector T cells in peripheral blood and TILs
						Increasing densities of CD8+ T cells in tumor microenvironment
Routy, B., et al., 2018	NSCLC and RCC	Patients	100	Akkermansia muciniphila	Enhanced efficacy of PD-1 blockade	Higher abundance in responders	Induce dendritic cell secretion of IL-12	Villejuif, France
						Prolonging progression free survival
Matson, V., et al., 2018	Melanoma	Patients/Mouse	--	Bifidobacterium adolescents	Enhanced efficacy of PD-1 blockade therapy	Higher abundance in responders	Enhance the activation of dendritic cells Increase CD8+ T cells	Chicago, USA
						Decreasing peripherally derived Tregs
				Bifidobacterium longum		Higher abundance in responders
				Veillo nella parvula		Higher abundance in responders	Increased frequency of dendritic cells and greater T helper cell responses Decreased frequency of regulatory T cells
				Klebsiella pneumonia
				Enterococcus faecium		Decreasing peripherally derived Tregs
				Collinsella aerofaciens
				Parabacteroides merdae
				Roseburia intestinalis		Higher abundance in non-responders	Increase in CD8+ tumor-infiltrating lymphocytes
				Ruminococcus obeum
Chaput, N., et al., 2017	Melanoma	Patients	26	Butyrate producing bacterium	Enhanced CTLA-4 blockade efficacy therapy	Higher abundance in responders	--	Villejuif, France
						Inducing activation of Treg
						Promoting development of tolerogenic macrophages and dendritic cells
						Prolonging progression free survival/overall survival
				Faecalibacterium prausnitzii	Enhanced CTLA-4 blockade efficacy therapy	Prolonging overall survival
						Elevating colitis risk
				Gemmiger formicilis	Enhanced efficacy of CTLA-4 blockade therapy	-Prolonging progression free survival/overall survival
						-Elevating colitis risk
Frankel, A., et al., 2017	Melanoma	Patients	39	Bacteroides caccae	Enhanced efficacy of ICB blockade therapy	Higher abundance in responders	--	Texas, USA
				Dorea formicogenerans	Enhanced efficacy of PD-1 blockade therapy (pembrolizumab)
				Bacteroides thetaiotamicron	Enhanced efficacy of CTLA-4 plus PD-1 blockade therapy
				Faecalibacterium prausnitzii	Enhanced efficacy of CTLA-4 plus PD-1 blockade therapy
				Holdemania filiformis	Enhanced efficacy of CTLA-4 plus PD-1 blockade therapy
Dubin, K., et al., 2016	Melanoma	Patients	34	Bacteroidetes	--	Decreased colitis secondary to anti- CTLA-4	Stimulate Tregulatory cell differentiation	New York, USA
Sivan, A., et al., 2015	Melanoma	Mouse	--	Bifidobacterium	Enhanced efficacy of PD-1 and PDL1 blockade therapy	--	Enhance the activation of dendritic cells Increase CD8+ T cells	Chicago, USA
Vetizou, M., et al., 2015	Sarcoma, Melanoma and Colon cancer	Mouse	--	Bacteroides	Enhanced efficacy of CTLA-4 blockade therapy	--	Induce T helper 1 immune responses in tumor-draining lymph nodes Promote maturation of intratumoral dendritic cells	Villejuif, France

MSS=metagenomic shotgun sequencing; rRNA=ribosomal ribonucleic acid; gDNA=genomic deoxyribonucleic acid; PD-1=programmed cell death protein 1;CTLA-4=cytotoxic T-lymphocyte-associated protein 4; NSCLC = non-small cell lung cancer; RCC=renal cell cancer; Treg=regulatory T-cell

Another study published by Frankel and colleague in 2017 demonstrated an association between human gut microbes and metabolites on ICB response in a cohort of patients with metastatic melanoma. In their cohort, responders to treatment with combination anti-CTLA4 and anti-PD1 immune checkpoint blockade had an enrichment of taxa such as Faecalibacterium prausnitznii, Bacteroides thetaiotamicron, and Holdemania filiformis (Table 1) whereas an enrichment in Dorea formicogenerans was found in responders to anti-PD-1 monotherapy [49].

Following this, several groups co-published papers in Science in 2018 reporting a strong association between the diversity and composition of the gut microbiome and response to ICB in patients with melanoma and other cancers[35,36,47]. In these studies, distinct microbial signatures were observed in the gut microbiota of responders vs non-responders to ICB across a range of cancer types, including melanoma, non-small cell lung cancer, and kidney cancer. Interestingly, taxa associated with response differed among the cohorts with modest to little overlap (Table 1) - thus highlighting the complexity of identifying specific taxa to include in a bacterial consortia to enhance responses. However there are some functional redundancies in taxa identified across these cohorts [51] thus supporting the concept that we should be focusing more on bacterial function rather than phylogeny. This notion is strengthened by the observation regarding a higher diversity of gut microbes in responders to ICB, as this might suggest that functional redundancy in the gut microbiome is critically important to fill potential functional “holes” in the repertoire of the gut microbiome.

Further clinical evidence regarding the role of gut microbes in immunotherapy response

An additional piece of evidence substantiating the role of gut microbes in promoting immunotherapy response lies in the growing body of literature suggesting reduced response rates and impaired survival in patients on ICB who receive antibiotics during treatment [52]. One of the first studies to demonstrate this provocative finding was by Routy et al in 2018 [47]. This manuscript showed that if patients received antibiotics before or early after start of treatment with ICB, they had significantly worse survival. However, there were potential confounders in the study limiting conclusions based on these results. Nonetheless, since that time numerous studies have been published demonstrating a negative association between the use of antibiotics and response to immune checkpoint blockade across several cancer types [53–55] with a recent meta-analysis summarizing these studies [52].

Together, these studies illuminate the potential deleterious impact of antibiotics on response to immune checkpoint blockade; however, important questions remain. These studies did not include longitudinal microbiome characterization before and during therapy (both ICB and antibiotics) - thus the direct relationship between antibiotic use and impact on the microbiome and associated immunity / anti-tumor immunity is unknown. Nonetheless the data in aggregate is quite compelling, and warrant (and mandate) additional studies to further define the impact of specific classes of anti-microbials on the gut microbiome and therapy response. Such studies will allow us to guide care and will also allow for more targeted approaches and engagement within and outside our respective disciplines regarding antibiotic stewardship.

Strategies targeting gut microbes in patients on immunotherapy

Based on this compelling preclinical and observational data supporting a role for the gut microbiome in cancer therapeutic outcomes, there are now multiple interventional studies examining the ability of gut microbiota modulation to enhance therapeutic efficacy and abrogate treatment-associated toxicity. Several strategies are being employed, building off of prior work on microbiome modulation in other diseases. (Figure 2).

Figure 2: Modulation of gut microbiome.

Different strategies /approaches that can be employed to modulate the gut microbiome and can be used as interventions to improve immunotherapy treatments and abrogate the treatment associated toxicity i.e. Fecal microbiota transplant (FMT), administration of microbial consortia or probiotics, prebiotics that includes die and supplements, or use of personalized antibiotic therapy or phages.

Fecal microbiota transplant

Fecal microbiota transplant (FMT) is the most well-established strategy to modulate the microbiome. In FMT, the gut microbiome in its entirety is transferred from a donor to a recipient as a stool suspension, administered either endoscopically on in the form of pills. FMT is a proven therapy for recurrent and refractory Clostridium difficile infection (CDI) [56,57] and has also been tested as a treatment modality for many other dysbiotic diseases and is now being investigated in the context of cancer therapy. This includes studies of FMT in SCT patients such as a trial of patients with acute myeloid leukemia, with FMT targeting the prevention of dysbiosis and eradication of multi-drug resistant bacteria (NCT02928523).

There are also a growing number of trials now investigating FMT in cancer patients undergoing immunotherapy (NCT03353402, NCT03341143, NCT03772899, NCT03817125, NCT04130763, NCT04116775, NCT04056026). This field is still at its beginning, and thus these are small pilot Phase I/II studies with primarily safety endpoints, though of course the ultimate strategy is to enhance therapeutic response. These studies vary in selection, delivery method, and FMT duration reflecting that the optimal FMT regimen in this population is still being defined (further discussed below). Very early data from one of these studies being conducted in Israel (NCT03353402) was presented at the 2019 AACR meeting with exciting early safety and efficacy data. This study is enrolling patients with anti-PD1 refractory melanoma who received FMT from a CR donor + anti-PD1, and reported responses in 3 of 10 patients with no adverse safety signals.

FMT is also being studied as a way to abrogate treatment related toxicities. A provocative case study of FMT for steroid-refractory ICB-induced colitis showed remarkable efficacy in two patients, with both histologic and clinical remission [58]. Based on this experience, at least two prospective clinical trials of FMT for ICB-induced toxicity are now underway (NCT03772899 and NCT03819296).

Defined bacterial consortia

Another method of modulating the gut microbiota is through the oral administration of live bacterial consortia or even a single putatively beneficial bacterium as pills (i.e., probiotics). The advantage to this approach is ease and simplicity. One such ongoing trial (NCT03817125) in metastatic melanoma patients undergoing immunotherapy (anti-PD1) employing a defined orally administered microbial formulation (SER-401) is intended to assess safety, clinical response, as well as change in gut microbiome and immunity. Another ongoing clinical trial of a proprietary bacterial strain (MRx0518) in combination with anti-PD-1 therapy is enrolling patients with melanoma, bladder cancer, non-small cell lung cancer and renal cell carcinoma (NCT03637803). While the administration of probiotics along with immunotherapy in cancer is relatively new, it has been employed in combination with other forms of cancer therapies such as chemotherapy and SCT [59–62]. However, significant caution should be taken until preclinical and clinical studies fully evaluate the safety of using commercially available probiotics, especially given significant variability and lack of regulation of over the counter probiotics [63].

Diet and prebiotics

Gut microbiota play a major role in harvesting important nutrients for the host by digestion of the food and, as noted above, resultant metabolites play a substantial role in host immunity [64–66]. For example, the plant polysaccharide inulin can stimulate an outgrowth of the fiber-fermenting Faecalibacterium and Bifidobacterium species [67], two taxa recently associated with enhanced response to immune checkpoint blockade [36,68] whereas supplementation with inulin and fructo-oligosaccharide may select for such beneficial bacteria as Lactobacillus and Bifidobacterium, [69]. Thus, there is a strong rationale for investigating the effects of dietary interventions on the gut microbiota and immunity in the context of ICB. However, while diet is a convenient and accessible strategy, behavior change is notoriously difficult and defining what diet components or patterns are optimal in this context remains an unmet need.

Fecal transplant and unique issues with regard to patients with cancer

Thus, while gut microbiota can be modulated by a variety of methods, the most established strategy is through the use of FMT. However, there are several important considerations in designing and implementing the FMT trials with cancer patients [63]. These include donor selection, patient population, pre-conditioning, FMT modality, duration of treatment, and endpoints. In regards to donor selection, the primary considerations are healthy donor vs ICB complete responder, and microbiome profiling. Though FMT from healthy donors has proven to be effective in treating non-malignant diseases such as CDI [70], whether FMT from healthy donor is sufficient to induce response to ICB or whether a responder donor is needed has not been established nor has the practice of microbiome profiling to select optimal donors.

Another area of controversy is the use of pre-conditioning antibiotics. Antibiotics are routinely used prior to FMT to ablate the native microbiome and facilitate engraftment. However, as has been noted above, antibiotic use has been associated with worse outcomes with ICB[71,72]. A preclinical study reported that pre-treatment with broad-spectrum antibiotics did not affect the overall engraftment of donor microbiota in the recipients but improved the engraftment of beneficial bacterial taxa such as Bifidobacterium [73]. The effects of the specific antibiotic regimen typically used for FMT preconditioning (oral Vancomcyin) in the context of ICB is also an unanswered question.

The optimal route of administration of FMT in the setting of immunotherapy also needs to be defined. For CDI, a single endoscopically delivered FMT is sufficient for durable engraftment. However, this is not the case for other disease states which are less profoundly dysbiotic and thus may be more resistant to colonization[74]. For example, results of trials of FMT for inflammatory bowel disease have been decidedly mixed, but a much more significant benefit was seen in a recent study that employed ongoing FMT dosing x 8 weeks[75]. Though this study utilize repeated enemas, other studies have demonstrated frozen stool pills to be a convenient and effective alternative to traditional endoscopic approaches and this is an approach also being tested in the setting of immunotherapy[76].

Irrespective of the source for the FMT, the fecal samples may be considered as an imprecise source of immune-potentiating microbial community that may potentially contain harmful bacteria including pathogens, drug resistant bacteria and microbial groups associated with metabolic disorders or even carcinogenesis. A study led by DeFilipp et al. demonstrated that two patients following FMT developed bacteremia with extended-spectrum beta-lactamase (ESBL)–producing Escherichia coli and one of them who underwent hematopoietic cell transplantation died [77]. This warrant updating the donor screening strategy routinely, considering any potential emerging bacteria or viruses that may cause adverse events in the recipients while donors remain asymptomatic. However, overall vast experience with FMT for CDI and other indications support that this is generally a safe and well-tolerated approach.

Conclusions and future directions

Through multiple preclinical and human observational studies, it is now well-established that the gut microbiome is associated with response to cancer immunotherapy. However, there remains considerable work to be done in establishing mechanism as well as the microbial features causally linked to response. Preclinical models further suggest microbiome modulation as a therapeutic strategy to enhance response to immunotherapy and this approach is now being tested in multiple ongoing clinical trials. Modalities being tested include FMT, probiotics and bacterial consortia, and prebiotics and diet. Each of these strategies has important variables to be considered, and these early studies with integrated biospecimen collection will yield important data to help us further optimize these approaches and continue to move this field further.

Key Findings.

• The gut microbiome plays a key role in shaping mucosal and systemic immunity

• Work in preclinical models first demonstrated that the gut microbiome can also impact anti-tumor immunity

• Data from multiple human cohorts now supports that there is a strong association between gut microbiome profile and response to immune checkpoint blockade (ICB)

• Microbiome modulation can impact response to ICB in mice and various approaches of microbiome modulation are now being tested as a strategy to enhance response and ameliorate toxicity to ICB in multiple human interventional studies

• The precise mechanisms by which the gut microbiome impacts anti-tumor immunity remains an area of active investigation

Biographies