Alimentary Pharmacology and Therapeutics Review article: inhibition of methanogenic archaea by statins as a targeted management strategy for constipation and related disorders K. Gottlieb*, V. Wacher*, J. Sliman* & M. Pimentel† *Synthetic Biologics, Inc., Rockville, SUMMARY MD, USA. † Gastroenterology, Cedars-Sinai Background Medical Center, Los Angeles, CA, USA. Observational studies show a strong association between delayed intestinal transit and the production of methane. Experimental data suggest a direct inhibitory activity of methane on the colonic and ileal smooth muscle and Correspondence to: a possible role for methane as a gasotransmitter. Archaea are the only con- Dr K. Gottlieb, Synthetic Biologics, fi Inc., 9605 Medical Center Drive, rmed biological sources of methane in nature and Methanobrevibacter Rockville, MD 20850, USA. smithii is the predominant methanogen in the human intestine. E-mail: [email protected] Aim To review the biosynthesis and composition of archaeal cell membranes, Publication data archaeal methanogenesis and the mechanism of action of statins in this context. Submitted 8 September 2015 First decision 29 September 2015 Methods Resubmitted 7 October 2015 Narrative review of the literature. Resubmitted 20 October 2015 Accepted 20 October 2015 Results EV Pub Online 11 November 2015 Statins can inhibit archaeal cell membrane biosynthesis without affecting This uncommissioned review article was bacterial numbers as demonstrated in livestock and humans. This opens subject to full peer-review. the possibility of a therapeutic intervention that targets a specific aetiologi- cal factor of constipation while protecting the intestinal microbiome. While it is generally believed that statins inhibit methane production via their effect on cell membrane biosynthesis, mediated by inhibition of the HMG- CoA reductase, there is accumulating evidence for an alternative or addi- tional mechanism of action where statins inhibit methanogenesis directly. It appears that this other mechanism may predominate when the lactone form of statins, particularly lovastatin lactone, is administered. Conclusions Clinical development appears promising. A phase 2 clinical trial is currently in progress that evaluates the effect of lovastatin lactone on methanogenesis and symptoms in patients with irritable bowel syndrome with constipation. The review concludes with an outlook for the future and subsequent work that needs to be done. Aliment Pharmacol Ther 2016; 43: 197–212 ª 2015 The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd. 197 This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes. doi:10.1111/apt.13469 K. Gottlieb et al. – INTRODUCTION constipation6 8 and one study that concludes that colonic Methane, which is now of great interest to climate methane production is not associated with clinical pre- researchers, plays an important if somewhat underappre- sentation in IBS patients.9 ciated role in gastrointestinal disorders. The link between Two microbiome studies correlating methanogens and this odourless gas and the intestine was first made in the stool frequency have recently become available. One early days of colonoscopy, when sparks from electrosur- study evaluated the correlation between gut microbiota gical devices sometimes led to intracolonic explosions.1 variation and stool consistency using the Bristol Stool Microbial metabolism of mannitol, a sugar alcohol that Scale (BSS) classification. Enterotypes were distinctly dis- had been commonly used for colon lavage, led to tributed over the BSS scores: Within the RB enterotype, increased concentrations of the flammable gases hydro- found in harder stool samples, the abundance of gen and methane. Modern bowel preps do not carry that Methanobrevibacter and Akkermansia was positively cor- risk. related with colon transit time.10 The results of another Methane can have inorganic (e.g. geothermal-volcanic microbiome study analysing 273 faecal samples showed methane emissions) and organic sources. The only that only patients with a constipation phenotype pre- organisms known to produce methane are methanogenic sented higher abundance of methanogenic archaea, con- archaea. Archaea are fascinating prokaryotes that were firming a link between low transit time and methane previously considered to be bacteria. However, these production capacity.11 prokaryotes, while resembling bacteria on a superficial Association does not prove causation, and it has long level, were found to have so many unique characteristics been known that intestinal dysmotility, as seen in sclero- that they were given their own name, archaea, and were derma, leads to secondary small intestinal bacterial (and, elevated to occupy a position as one of the three presumably, archaeal) overgrowth. It could be argued domains of life: archaea, bacteria, and eucarya.2, 3 that increased methane production is therefore not the Less spectacular than its explosive capacity but more cause but an effect of delayed intestinal transit. Fourteen important is recent evidence that links intestinal healthy volunteers underwent four interventions: placebo, methane production to slowed intestinal transit.4 In this sulphate supplements, or sulphate supplements with review, we will describe how large quantities of methane either senna or loperamide. With faster intestinal transit are produced in the colon and how statins may have a reduction in faecal methanogens and methane produc- therapeutic importance by inhibiting methane produc- tion was seen. The reverse effects were true with lop- tion through the selective suppression of the growth of eramide.12 However, there is experimental evidence that methanogens. methane directly affects intestinal transit and contractil- ity (methane-first hypothesis). METHANE AND CONSTIPATION-ASSOCIATED DISORDERS Experimental evidence While methane is predominantly produced in the colon, Methane and intestinal transit methanogens can also be demonstrated in the small Although 50–80% of methane in humans is passed as intestine, especially when small bowel bacterial over- flatus, the presence of methane production can be accu- growth exists.13 Pimentel and colleagues conducted a rately assessed by breath tests. A comprehensive review three-part study consisting of two experiments – one recently concluded that data from breath testing gener- in vivo (dogs) and one ex vivo (guinea pig ileum) – as ally support the association between delayed intestinal well as a retrospective review of data in irritable bowel transit and the production of methane.5 Methodological syndrome (IBS) patients who had previously undergone differences in study design precluded a formal meta-ana- breath testing and antroduodenal motility studies.4 In lysis approach for studies that correlated diagnosis of dogs, small intestinal fistulae were created and transit constipation-predominant IBS (IBS-C) or chronic idio- of a radiotracer was measured during infusion of ambi- pathic constipation with methane production, neverthe- ent air, followed by methane at a rate that was physio- less, the same report5 lists 14 studies supporting the logical (i.e. resulting in 50 ppm in exhaled air), a association between methane production and constipa- concentration that is typical for patients with irritable tion related disorders in adults. There are a few reports bowel syndrome with constipation (IBS-C). The that primarily evaluated other hypotheses that did not methane infusion resulted in decreased radiotracer show a link between methane on breath testing and recovery in each of five individual dogs compared to 198 Aliment Pharmacol Ther 2016; 43: 197–212 ª 2015 The Authors. Alimentary Pharmacology & Therapeutics published by John Wiley & Sons Ltd. Review: methane, archaea, statins and the gut baseline. The average reduction was 59%, which was survival conditions for archaea – more methane is pro- highly significant (P < 0.001). duced further slowing down intestinal transit leading to In the second experiment, 3-cm-long terminal ileum symptomatic constipation – archaea thrive and multiply preparations from sacrificed guinea pigs were oriented in until a steady state is achieved. an oral-aboral fashion in a physiological buffer solution and stimulated before and after methane was bubbled THE ORIGIN AND PROPERTIES OF METHANE through the tissue bath. Methane exposure at a concen- PRODUCTION tration of 1000 ppm led to a significant augmentation of The five major gases in the colon are: nitrogen (23– the contractile force in response to the applied brush 80%), hydrogen (0.06–47%), carbon dioxide (5.1–29%), stroke, consistent with hyperactivity. oxygen (0.1–2.3%) and methane (0–26%).1 Finally, after retrospective review, 11 methane-produ- In addition, nitric oxide and hydrogen sulphide are cing and 12 sex-matched hydrogen-producing subjects found as well. These gases have great importance were identified who met Rome I criteria for IBS and, in through their function as gasotransmitters and compo- addition, had antroduodenal manometry data available. nents of a complex microbial ecosystem.12, 15 All the The fasting motility index, as a gross measure of forces methane in the intestine is produced by methanogenic (resistive and propulsive) in the small intestine, in bacteria of which Methanobrevibacter smithii,