The “gut-brain-axis” is the focus of growing numbers of researchers, biotech companies and investors. They are uncovering multiple two-way communication pathways between belly and brain, spanning neural, hormonal and immune-system signalling.
- New research into the “brain-immune-gut-axis” also challenges prior assumptions of how isolated the brain is from the rest of the body and, in particular, from the immune system.
- This gut-mediated back-door route to the brain offers several potential advantages for drug developers.
Hippocrates (460-370BC) believed that all diseases begin in the gut. He was closer to the truth than he has been given credit for. It is reasonable to assume that gastro-intestinal disorders or metabolic diseases may start in the gut. But research over the last 15 years into the nature and role of the gut microbiome – the trillions of microorganisms that reside in our intestine – has uncovered its far wider influence, including across many auto-immune conditions, cardiovascular disease and even cancer.
Even then, few would have linked gut health to Parkinson’s, autism or Alzheimer’s. Yet gut and brain health are far more intimately and intricately linked than previously imagined. The “gut-brain-axis” is the focus of growing numbers of researchers, biotech companies and investors. They are uncovering multiple two-way communication pathways between belly and brain, spanning neural, hormonal and immune-system signalling (see Exhibit 1). The mechanisms, players and causalities within these pathways are still being elucidated. But thanks to more powerful genome sequencing and editing tools, and more sophisticated imaging and data analytics, evidence of the gut’s influence on behavior, mood and disease is accumulating fast.
This work is opening up a treasure chest of new drug targets and potential interventions that may help address neurodegenerative and other cognitive diseases that continue to elude traditional drug development. Preclinical and early clinical work suggests that metabolites generated by gut microbes can influence stress, depression and anxiety. Some gut microbes secrete neuroactive substances like serotonin and dopamine that directly stimulate the nervous system – though it is still unclear how, and to what degree.
THE VAGUS NERVE – THE LONGEST CRANIAL NERVE IN THE BODY – FANS OUT FROM THE BRAIN STEM TO THE COLON. IT PROVIDES THE BRAIN WITH SENSORY INFORMATION FROM IMPORTANT ORGANS INCLUDING THE LUNGS, HEART AND INTESTINE, AND, IN THE OTHER DIRECTION, CARRIES VITAL MOTOR SIGNALS INCLUDING TELLING YOUR LUNGS TO BREATHE AND YOUR HEART TO PUMP. THE VAGUS NERVE PROBABLY FERRIES MESSAGES FROM MANY OF THE NEWLY-DISCOVERED NEURO-IMMUNE AND NEURO-ENDOCRINE SIGNALLING MOLECULES, TOO. THE INTESTINAL BARRIER (AROUND OUR GUT WALL) AND THE BLOOD BRAIN BARRIER (PROTECTING THE BRAIN FROM THE SYSTEMIC CIRCULATION) ARE NATURAL BLOCKS TO GUTBRAIN SIGNALLING. BUT THEIR PERMEABILITY IS INFLUENCED BY GUT MICROBES, STRESS AND INFLAMMATION. SO THE AMOUNT (AND KINDS) OF INFORMATION REACHING THE BRAIN FROM THE GUT, AND VICE VERSA, MAY VARY WIDELY. - Source: Nature Reviews Neuroscience (2012)
Axial Biotherapeutics Inc.’s lead program involves mopping up a microbial metabolite which may be implicated in autism spectrum disorder. The gut is also an important modulator of various hormones used to signal satiety – telling us to stop eating. Companies like New York-based Kallyope Inc. and France’s TargEDysare using that information to develop new kinds of obesity treatments, tackling the behavioural aspect of the disease. “We now have very provocative evidence that we can influence various central nervous system disorders through the gut-brain axis,” summed up Nancy Thornberry, Kallyope’s CEO.
Related work into the “brain-immune-gut” axis also challenges prior assumptions of how isolated the brain is from the rest of the body and, in particular, from the immune system (much of which resides in the gut). This is uncovering further potential therapeutic avenues for brain diseases, and is the focus of discovery work at Boston-based PureTech Health PLC and California-based Alector Inc. Meantime, the gut microbiome’s effect on cancer – mediated by the immune system – is another booming R&D sub-sector. “The gut-brain axis and oncology are the hottest areas, with the biggest market potential, in the [gut] microbiome space,” opined Isabelle de Cremoux, managing director at investment firm Seventure Partners, whose Health for Life Capital II is the company’s second gut microbiome-focused fund, on its way to raising €200m
More, Safer Routes To The Brain?
This gut-mediated “back-door” route to the brain offers several potential advantages for drug developers. The first is that therapies do not have to cross the blood-brain-barrier (BBB). The BBB is a tightly-built wall that prevents the majority of pathogens (as well as large immune-system molecules like antibodies) from crossing into the brain from the systemic circulation. The BBB has a vital protective role. But it is also one reason many CNS drug candidates have failed: they cannot get to where they are needed.
The second advantage of the gut-mediate route to the brain is safety: therapies whose actions are restricted to the gut – or which are based on naturally-occurring, human commensal microbes – are less likely to trigger unwanted or dangerous side-effects than systemically-delivered drugs. That will matter to regulators, particularly in the context of highly prevalent conditions like obesity and neuro-degenerative diseases.
Finally, scientists’ greater understanding of the gut-route to the brain may help clarify the benefits (or otherwise) of alternative, non-drug modalities such as medical foods, nutritional supplements or probiotics in influencing health and behavior. These are subject to different regulatory and marketing requirements to drugs; traditional clinical evidence of their efficacy in humans has, thus far, been patchy. This may change as more precise maps are created of the molecular mechanisms and pathways between gut, brain and rest-of-body, helping us better appreciate the full impact of what we swallow.
The flourishing work into gut microbiome has already created a pipeline of new modalities, including carefully-chosen strains of bacteria packaged into capsules (“bugs-as-drugs”), or selected bacterial metabolites. For Seventure’s de Cremoux, the modality is irrelevant; it’s about the quality of the science and of the clinical evidence. “We invest in all these companies. You never know in advance which regulatory pathway is most suitable,” she said.
Axial – Mopping Up Gut Metabolites For Autism
Jim Blair, partner at US-based VC firm Domain Associates, would prefer the end product to be a classic small molecule – though being able to test bugs-as-drugs initially, to elucidate mechanism, was very helpful, he noted. Domain is an investor in Axial Biotherapeutics, which is developing a gut-mediated small molecules for autism spectrum disorder and Parkinson’s disease. The company raised $35m in a series B round this year, including Domain, Longwood Fund, Seventure and Taiho Ventures.Scientists have known for some years that as many as half of children with autism spectrum disorder also suffer from gastro-intestinal upset. GI symptoms such as constipation are observed in many Parkinson’s patients over a decade before cognitive and motor problems show up. Until recently, no-one understood whether or how the GI issues linked to the conditions’ better-recognized behavioral and cognitive symptoms.
Axial’s scientific co-founder Sarkis Mazmanian, professor of microbiology at the California Institute of Technology (Caltech), and his team used these observations to design mouse models of both diseases. These have generated compelling preclinical evidence of a gut-brain link in these conditions. Mice bred to display the behavioral features of autism generated offspring that also displayed GI symptoms; specifically, ‘leaky gut’, in which the intestinal barrier (or gut wall – a crucial barrier protecting us from outside world) becomes more porous than usual. Significantly, treating those mice with Bacteriodes fragilis, a human commensal bacterium, repaired the gut wall as well as correcting some behavioral abnormalities. Treating mice with microbiomes from autistic children induced similar behaviors in the mice.
Axial’s team has used metabolite profiling and other tools to investigate more deeply the molecular mechanisms behind these findings. It discovered that autistic mice had particularly high levels of a microbial metabolite, 4-ethylphenylsulfate (4-EPS), and that reducing 4-EPS levels improves behavior. It also found that feeding animals this metabolite led to behaviour abnormalities. That amounts not to definitive evidence of causality, but something close. “We think we’ve found something that is much stronger than correlation,” said David Donabedian, Axial co-founder and CEO.
So the company’s polymer chemists designed a small, non-biotherapeutic molecule, AB-2004, which mops up that metabolite before passing out of the body in the feces. Axial published preclinical data in May 2019 showing that AB-2004 restored GI integrity and reduced 4-EPS levels in mouse models of ASD. It also has early evidence of a link, in humans, between 4-EPS and two measures of social function. A study of 130 autistic children and 101 children from the general population found statistically significant differences in the mean levels of the metabolite between the groups. Starting levels of 4-EPS varied very widely among the subjects, however, making it tricky to establish a baseline.
“It’s early days, and 'normal’ levels of 4-EPS in the general population aren’t well understood,” conceded Donabedian. But 4-EPS was clearly elevated in a large subset of autistic patients, and “these elevated levels appear to be correlating with certain aspects of the ASD profile,” he said. It may be that the pathological threshold for particular microbial metabolites is lower in some autistic patients, as a function of both genetic predisposition and underlying metabolite profile.
Axial is currently screening adolescent patients with autism and GI symptoms for inclusion in a Phase Ib/IIa clinical study that will further elucidate the roles and relevance of these metabolites. The company’s work may also help to more accurately, and helpfully, classify a very heterogenous condition.
Mazmanian and colleagues’ work with mouse models of Parkinson’s is also uncovering potentially causal mechanisms within the gut. Transplanting fecal samples from human Parkinson’s patients into germ-free mice led to parkinsonian behaviour in the mice, while healthy samples did not. Mice designed to express alpha-synuclein, a protein that mis-folds in Parkinson’s patients to form damaging clumps in parts of the brain, did not show symptoms so long as they lacked gut microbes. Motor deficits and alpha-synuclein aggregation in the brain both appeared in such mice, though, if fed with short-chain-fatty acids – an important category of microbial metabolites.
Axial hopes eventual treatments that emerge from these insights may improve both the behavioral and GI symptoms of Parkinson’s and autism.
PureTech: Draining The Brain To Cure Diseases Of Aging
PureTech Health is also going after new ways to treat brain diseases. One of its scientific collaborators in 2015 published a paper in Nature describing an entirely new component of the brain-immune-gut axis. Jonathan Kipnis, professor and chair of neuroscience and director of the Center for Brain Immunology and Glia at the University of Virginia School of Medicine and his team had uncovered a network of lymphatic vessels within the meninges – the multi-layered membrane covering the brain and spinal cord (see Exhibit 2).
The brain was previously believed to be insulated from the body’s lymphatic and immune systems by the BBB. (The brain itself does not contain lymphatic vessels.) Yet the lymph vasculature serves two very important roles for the rest of the body: removing unwanted fluid and protein, and immune surveillance – picking up warning signals from tissues that are used to mobilize an immune response. The healthy brain needs both those services too. Kipnis' work helped elucidate precisely how they are delivered. (It also complemented the discovery, by Maiken Nedergaard at the University of Rochester Medical Center, of another part of the brain's waste-disposal system. The so-called 'glymphatic system' flushes unwanted molecules out of the fluid in between brain cells far more efficiently than previously thought, by way of the CSF and blood vasculature.)
Importantly, it also found “that the performance of the meningeal lymphatic system declines significantly with age,” said Joe Bolen, PureTech’s chief scientific officer. Bolen likened this decline to a “blocked drain,” causing protein waste and other materials to accumulate and cause damage.
Kipnis showed that blocking meningeal lymphatics in mice models of Alzheimer’s accelerated disease progression. Other researchers have since highlighted the system’s role in clearing disease-associated proteins including tau and alpha-synuclein from the brain. Together, these findings have opened up promising avenues for drug discovery, most obviously in neuro-degenerative disorders associated with aging.
PureTech, with Kipnis’ team (and technology licenses from the University of Virginia), is avidly combing those avenues for new medicines that can help clear potentially pathogenic macromolecules from the CNS. Encouragingly, Kipnis’ 2015 paper suggested that age-related functional deterioration of lymphatic system could be reversed by supplying vascular endothelial growth factor C (VEGFC), which affects the health and permeability of the lymphatic endothelium. “It was a great experiment. Jonathan [Kipnis] injected a VEGFC-expressing virus into the cerebrospinal fluid of old animals, and after a couple of weeks, the lymphatic vessels opened and started draining macro-molecules,” recounted Bolen.
The work is still at a very early stage: much remains to be uncovered about the detailed architecture of meningeal lymphatics and its regulatory pathways. But Bolen was excited: “We’ve isolated lymphatic endothelial cells from mice, sequenced them, and are starting to understand their biology,” he said – a feat that would have been impossible without today’s tools and knowledge. PureTech is also working on delivery technologies that leverage its growing knowledge of lymphatics – including delivering small molecule pro-drugs to mesenteric lymph nodes to modulate the immune system.
PureTech’s most advanced internal drug candidate is an immune-modulating anti-galectin-9 antibody, expected to reach IND stage in 2020. The company also invests in others working along various parts of the brain-gut-immune axis. Among its external affiliates is Vedanta Biosciences, whose therapeutic candidates are carefully chosen gut microbiome-derived live bacteria. With what it described as one of the largest collections of commensal gut bacteria in the world, Vedanta has identified strains whose gut-protective and immune-modulatory properties provide potential applications across infectious and auto-immune diseases, allergy and immune-oncology. The company raised an $18.5m extension to its series C financing in May 2019.
Outside of the PureTech community, recently listed Alector is focusing on genetic mutations that disrupt the brain's immune system. It believes these disruptions may be behind neuro-degenerative diseases including Alzheimer's. Lead antibody candidates targeting those mutations are in early clinical trials.
Kallyope: Telling The Brain You Are Full
Kallyope is harnessing the gut-brain axis to tackle another chronic disease that remains under-served: obesity.
Among the company’s lead programs is a gut-restricted small molecule that stimulates multiple satiety hormones. Some of these hormones are well-known, explained CEO Thornberry. But Kallyope’s scientists have found new, better-defined routes through which to modulate their release.
Thanks to extensive mapping of the brain-gut circuitry using imaging and sequencing technologies, including tracking cell-specific proteins using light (optogenetics) or small molecules (chemogenetics), Kallyope has found ways to “elicit hormone release in a very rational way.” It has also come across cell types and hormones not previously identified in the gut, according to Thornberry.
Tackling obesity through behavior-change, rather than by restricting food intake or blocking fat absorption, is not a new idea. But past attempts to do so with drugs that interfered with neurotransmitters in the brain’s pleasure centers mostly failed. Sanofi’s Acomplia (rimonabant), which blocks cannabinoid receptors to dull appetite, was rejected by FDA in 2007 due to neurologic and psychiatric side effects. Where the drug is available, it carries cardiovascular risk warnings. Abbott Laboratories Inc.’s Meridia (sibutramine), a serotonin, dopamine and noradrenaline re-uptake inhibitor, was withdrawn from the US and European markets in 2010.
The idea is that a gut-mediated passage to the brain’s behavior and appetite-regulation centers will prove safer and more effective. Thornberry hopes that Kallyope’s multi-faceted gut-hormone approach will lead to weight loss “substantially greater” than the single-digit percentage levels achieved, often only temporarily, by existing pharmaceuticals.
Novo Nordisk AS, which sells a leading share of insulins and GLP-1 agonists for diabetes and metabolic disease, signed a research agreement with Kallyope in 2018. The partners will identify peptide-based candidates using Kallyope’s platform; if successful, Novo will carry out further preclinical and clinical development.
You Are Not Hungry, Your Gut Bacteria Are
TargEdys also believes that gut-mediated behaviour change is key to tackling obesity. Its scientific co-founders believe that appetite regulation may involve not only host gut-to-brain signalling (including hormonal signalling), but also the energy status and feeding behaviour of the gut bacteria community itself. In obese people, they speculate, bacterial-derived hunger signals may compete with – and ultimately dominate – those from other hormones like satiety-inducing leptin, for example.
There is already plenty of evidence supporting the microbiome’s role in energy balance. Giving obese mice pre-biotics (to feed their gut microbes) increases leptin sensitivity. On the other hand, kwashiorkor, a severe form of malnutrition, can be transferred from people to mice by fecal transplantation. Studies have shown that both anorexic and obese people have an imbalance in their gut microbiota. These observations, coupled with more recent data around the dynamics of bacterial growth, may support “a theory of genetic competition between the host and its microbiota for food sources,” wrote Serguei Fetissov, professor of physiology at the University of Rouen, France, in a 2017 article in Nature Reviews Endocrinology. Fetissov and Pierre Déchelotte, also a professor at the University of Rouen, are TargEDys’ scientific co-founders.
Bacterial proteins and metabolites (such as short-chain-fatty acids) involved in breaking down our food act on specialist cells in the gut called enteroendocrine cells, which in turn trigger the release of appetite-regulating hormones like PYY (pancreatic peptide YY), ghrelin or leptin. But what if other kinds of bacterial proteins, more closely linked to the bacteria’s own feeding pattern, could mimic these hormones? TargEDys’ scientists believe that a bacterial protein called caseinolytic peptidase B protein homolog (ClpB) may do just that, copying the actions of a satiety hormone called alpha-melanocyte-stimulating hormone (α-MSH). (In the context of broader work on the brain-immune system-gut axis, Fetissov also speculated that the immune system may be involved in long-term appetite regulation by gut bacteria. Antibodies, released in response to changes in diet or gut microbiota, may cross-react with appetite hormones like α-MSH.)
People with anorexia and related eating disorders are found to have higher than normal levels of ClpB, while the reverse is true for many obese or overweight patients. So TargEDys has developed a pro-biotic designed to restore ClpB levels in individuals who are overweight or obese. EnteroSatys, a food supplement available in France, contains a bacterial strain called Hafnia alvei 4597, originally isolated from raw milk cheese, along with bacterial-derived protein. “In our opinion, the problem [of excess weight and obesity] is eating behavior,” said TargEDys CEO Grégory Lambert. EnteroSatys works by “re-educating people on their choice of food and life-style,” and “helping them learn new behaviors.” It does not suppress appetite, Lambert emphasizes; instead it “normalizes it, restoring the natural [ClpB-mediated] mechanism” of appetite regulation.
TargEDys cannot make any health claims about EnteroSatys; no pro-biotic has been approved to do so in Europe (See Box: Pro-biotics, Food or Drug?). But TargEDys can show doctors and nutritionists preclinical, clinical and consumer data supporting the product. A “big” clinical trial is ongoing, according to Lambert, with results expected early in 2020. Preclinical work suggests that the bacteria does reduce food intake and weight, though not as much as Roche’s fat-blocking drug Xenical (orlistat). A consumer study of 45 people showed that 70% felt fuller after taking EnteroSatys, and felt that they had better control over their eating.
There is more to do to prove the product works: scientists do not yet understand precisely how important ClpB is in host appetite control, nor do they have conclusive evidence of causality between changes in microbiota composition and host feeding behavior. If such evidence accumulates, TargEDys may find the partners it needs to distribute its product beyond France; Lambert added that the company might also consider developing the protein as an active pharmaceutical ingredient, or as a live bio-therapeutic. “We’re waiting for the regulation to be a bit clearer” around live-bugs-as-drugs, he said.
Using The Gut To Enhance Immuno-Attacks On Cancer
Several companies are using gut-derived microbes and metabolites to re-direct the immune-system to fight cancer – potentially enhancing the actions of existing checkpoint inhibitors and other marketed immune-oncology drugs. Definitive evidence of such enhancement would take microbiome-derived drugs into the big league.
Such evidence is not yet available. Clinical trials have begun, though, of various microbial mixtures chosen for their immuno-genic actions. Evelo Biosciences, based in Cambridge, MA, has selected single microbe strains that are thought to modulate various immune pathways on their passage through the gut. Its orally-delivered, cancer-focused “monoclonal microbial,” EDP1503, in December 2018 began a Phase I/II trial in combination with Merck & Co. Inc.’s market-leading checkpoint inhibitor Keytruda (pembrolizumab). The trial will span several cancer types, including colorectal, triple-negative breast cancer. Evelo has other programs in inflammatory diseases.
Seres Therapeutics Inc.’s SER-401 is a mix of donor-derived commensal microbial strains chosen to improve patients’ response to checkpoint inhibitor treatment. It contains a bacterial signature found in melanoma patients that respond well to immunotherapy; the idea is to transfer that gut-microbiome advantage to others. The candidate is in a Phase Ib trial in metastatic melanoma, in combination with Bristol-Myers Squibb Co.’s nivolumab; preliminary read-out is expected in 2020. AstraZeneca PLC’s MedImmune signed a research agreement with Seres in March 2019, hoping to glean more about the mechanics of microbiome-enhanced cancer immune-therapy. The big pharma will pay $20m in fees and gets an option to negotiate rights to SER-401 and other candidates.Meanwhile, BMS itself signed up in December 2018 to collaborate on a clinical trial of Vedanta Biosciences Inc.’s bacteria-based mixture VE800, combined with Opdivo (nivolumab), in advanced and metastatic cancers. The study is planned for 2019. VE800 contains 11 clonal strains of human commensal bacteria selected, like Seres’, to amplify the effects of checkpoint inhibitors. BMS is testing its drug with multiple different therapy-partners that might enhance Opdivo’s effectiveness, and thus its competitive standing, in the red-hot immune-oncology field.
Paris, France-headquartered Enterome Bioscience SA has made its drug candidates out of gut microbe metabolites, chosen for their high degree of similarity to various solid tumor antigens. EO2401 contains three such “onco-mimics,” matched to tumor antigens that are highly expressed in some kinds of brain tumor.
Also, investors see potential for Axial Biotherapeutics’ gut-selective technology in oncology: Taiho Ventures’ $10m contribution to the company’s B round in June 2019 is earmarked for that purpose. “There’s lots of enthusiasm around the [gut] microbiome and its effect on cancer,” remarked Domain’s Jim Blair. “It may be that elements of the microbiome can trigger some of the epigenetic processes” now known to be involved in cancer, he said.
ProBiotics: Food or Drug?
Increasing numbers of companies outside the regulated pharmaceuticals industry are capitalizing on scientists’ growing understanding of the gut microbiome’s role in health and disease. They are variously developing and marketing pre- and probiotics, food supplements and indeed food products, all of which offer lower-cost routes to market than conventional drugs, but which are limited in the health claims they can make. Several are working at the boundaries of probiotics and pharmaceuticals, for instance seeking to engineer live micro-organisms specifically to alleviate particular CNS symptoms. Emergent categories include “psychobiotics” – probiotics used to treat psychiatric disorders.
As the boundaries blur between drugs and foods, the need for clearer definitions and regulations is increasing. In Europe and the US, food or food ingredients cannot be associated with the treatment of a disease, only with maintenance of normal physiological functions. But are probiotics food? The European Food Safety Authority shies away from such questions, pointing instead to the European Commission and member states.
The US FDA might categorize probiotics as dietary supplement, food ingredient, medical food or drug, depending on the intended use. But even then, neither the category limits nor the precise evidence requirements are entirely clear. Makers of medical foods, for instance, can communicate benefits in the “dietary management of a disease,” but only if said disease has “distinctive nutritional requirements,” as established by “medical evaluation.” (In Canada, regulators in 2016 authorized a probiotic claiming to moderate anxiety and promote a healthy mood balance – a product to enhance the brain-gut-axis.)
As for bugs-as-drugs (carefully-selected strains of bacteria packaged up as prescription biopharmaceuticals), the FDA has not yet approved such a therapy, though several live gut microbiome-based products are in late-stage clinical trials. There is no firm regulatory framework, but draft guidance exists, and FDA is engaged in discussions with industry on how to treat this emerging category.
A Neuro-Immune Super System?
Scientists now understand far more about the gut microbiome and its links to the brain than even just five years ago. But we are still at the early stages of mapping out the multiple mechanisms by which our resident bacterial communities influence our mental and physical health. For now, “there’s a lot of cataloguing going on,” said PureTech’s Bolen.
Researchers have found compelling correlations between levels of specific gut metabolites and certain disease phenotypes, but only rarely has a causal link been established. There is also more to learn about redundancies – what kinds of back-up mechanisms may come into play when a given pathway is interfered with? There remain knowledge gaps around the gut microbiome itself: scientists still do not know the function of many of the genes within gut microbial genomes, even among well-studied species. And the parameters of a “normal” gut microbiome remain to be determined: we understand that species diversity is important, but details of what comprises a “healthy” or “disease” state for a given individual are elusive.
Yet even as the contours of the gut-brain-immune axis are being uncovered, its relevance to disease is unquestioned. The top dozen or more chronic illnesses can be divided into either diseases of the immune system or of the nervous system, said Bolen. In the face of changing diet, life-styles and environmental stressors, these are “the only two adaptive systems” individuals have. Given the cross-talk between them, “they might more realistically be thought of as one super-system.”