Editorial: Solving doubts about L-Ornithine L-Aspartate for overt HE: whom and how to treat Manuscript: L-Ornithine L-Aspartate in bouts of overt hepatic encephalopathy Abstract: (words 103) Randomized controlled trials about the treatment of bouts of overt hepatic encephalopathy (OHE)

(words 103) Randomized controlled trials about the treatment of bouts of overt hepatic encephalopathy (OHE) are difficult to design and interpret, since bouts of OHE i) are relatively short episodes in which the dynamic of the response to treatment is relevant, ii) their occurrence and treatment is frequently linked to precipitating events. The study by Sidhu et al. in this issue of Hepatology is a model of proper design and provides evidence that L-ornithine-L-aspartate infusion for 5 days is an effective add on treatment to speed up recovery from bouts of OHE in comparison with a placebo treatment that does not contain amino acid nutrients. Main document (Words 1474 with references) Bouts of overt hepatic encephalopathy (OHE) i.e., hepatic encephalopathy causing at least clear disorienting in time or asterixis (1)severely hamper the autonomy and quality of life, causing a heavy burden on caregivers and health systems (2). Hyperammonemia plays a crucial role in the pathophysiology of HE, so that reducing ammonia levels is currently the primary therapeutic target. However, the link between HE and ammonia is not strict, since other factors intervene to alter brain function, in the first place infection (3). Thus, to select the appropriate therapy could be challenging, because of the multiple factors involved (1). A notable increase in the secondary prevention of bouts of OHE has been reached in the last years. It was proved that the risk for episodes of OHE increases after a previous one (4) and evidence exists that individuals who underwent more than one bout in the last six months have a higher risk than those who had only one bout (5, 6). Disaccharides, particularly lactulose, were proven to reduce the risk for episodes of OHE of about 50% in a year (6). Rifaximin in addition to lactulose further halves the risk for bouts of OHE in the following six months in the patients who had This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1002/hep.29530 This article is protected by copyright. All rights reserved. 2 suffered from more than one bout of OHE in the previous six months (5). With lower accuracy than the one reached for lactulose and rifaximin, also oral branched chain amino acids supplementation have been proved to be effective (7). Proper nutrition might also be useful in HE: some evidence of beneficial effects of a diet intervention (about halving the risk for OHE in the follow up of subjects with minimal HE) was recently published (8). In contrast to the studies about OHE prophylaxis, evidence about proper management of a single bout of OHE is poor. This depends mainly on two factors: i) the variety of the events that can precipitate OHE and ii) outcomes that should be taken into account. There is consensus that the treatment of the precipitant event is mandatory and prior (1). On the contrary, the ways to measure the outcome is confused. A too short interval between the beginning of treatment and the evaluation of outcome may bias the results towards the ineffectiveness of any therapy; in contrast, a too much long interval may prove the identity of the efficacy of any treatment, since finally a bout of OHE resolves, as per definition of the term 'bout'. In addition, pooling together OHE of any degree from time disorientation (i.e., the so called grade 2) to coma (i.e., the so called grade 4) might be misleading. The article by Sidhu et al. (9) is an example of a good approach to the problem: it takes into account the daily dynamic of HE in 5 days, which represents a reasonable time-interval to assess the treatment. Further, the outcome of OHE may depend on its severity, so categorizing the OHE grade provides not only information about the clinical status, but might also help to select the most appropriate therapy (10). Of note, Sidhu et al. (9) assessed the global effect of LOLA taking into account the OHE grade. LOLA was found more effective in lower (particularly grade II) than higher OHE grades (III/IV) in comparison with placebo, confirming previous results (5). Second, the dose (usually 20 g.) and the length (3-8 days) of the intravenous LOLA treatment are not homogeneous between RCT (5). This study made a day-a-day evaluation for HE improvement up to Page 2 of 5 Hepatology Hepatology This article is protected by copyright. All rights reserved. 3 5 days, founding that the first 4 days of treatment were enough to observe the beneficial effects of 30 g of i.v. LOLA. HE occurring in acute-on-chronic liver failure was not considered in this RCT, a condition where probably the treatment would be unsuccessful, because of the simultaneous presence of inflammatory-related triggers, very poor liver function and the occurrence of other complications. Notably, some precipitating factors for OHE, including oxidative stress or inflammation, are not covered by the usual treatments. In fact, LOLA did not reduce proinflammatory cytokines compared to placebo in this (9) study (in contrast to the reduction of ammonia plasma levels). Consequently, this drug could be suboptimal in some scenarios where inflammation is the main driver for the bout of OHE, which makes recommendable identifying the main trigger before selecting and initiating the therapy. Another interesting point is whether LOLA could be used in monotherapy or together with other drugs, such as non-absorbable disaccharides or antibiotics. Currently, the AASLD and EASL guidelines indicate that intravenous LOLA can be utilized as an alternative or additional agent to treat patients nonresponsive to conventional therapy (1). Sidhu et al. (9) considered as basal treatment the association of LOLA with non-absorbable disaccharides plus ceftriaxone. This can provide a reasonable baseline treatment in a study evaluating a metabolic drug, such as LOLA, since this kind of treatment does not interact with LOLA and can homogenize bouts of OHE that might require mainly disaccharides (e.g., OHE precipitated by constipation) or antibiotics (e.g., OHE precipitated by infection). Therefore, the results of this study reinforce the message that LOLA should be used as an add-on therapy rather than in monotherapy. A limit of the study is the significant number of subjects who were excluded. While this is reasonable in a trial, the efficacy of LOLA is doubtful in real practice in about half of the patients with OHE. Also, the impact of LOLA probably should have been stratified according to the trigger, Page 3 of 5 Hepatology Hepatology This article is protected by copyright. All rights reserved. 4 depending on the presence of ammonia(e.g., constipation, bleeding) or inflammatory-related (e.g., infections) factors that can influence on its efficacy. Another severe limitation is the selection of placebo. Indeed, 30 g of LOLA is a 30 g mixture of two amino acids. They might also reduce the ammonia output from muscles reducing amino acid muscle catabolism because of an unspecific nutritional effect. Would the effect be found if placebo had had the same nitrogen content than LOLA? In other words, is the effect of LOLA specific or is it a class effect of e.v. amino acids? The absence of any effect of previous studies on e.v. BCAA (7) cannot be invoked as proof that the infusion of an amino acid mixture per se is not effective as an add on treatment for OHE, since all these studies were groundlessly designed using BCAA as an alternative treatment in comparison with standard treatment. In the era of personalized medicine, both known drugs for OHE must be adapted, and selective therapies developed taking into account at least the severity of the bout and the main trigger. Despite its limits, the study performed by Sidhu et al. (9) clarifies the role of LOLA for a bout of OHE especially in patients with hyperammonemia regarding the adequate dose (30 g) and length of the therapy (up to 4 days), as well as the target population (lower HE grades) and the concomitant treatment (at least, lactulose). In addition, this study provides new opportunities for treatment and opens to further studies with an add-on design to improve the speed of recovery for bouts of OHE.