key: cord-0924699-7isjcasa authors: Balmant, Bianca D.; Torrinhas, Raquel S.; Rocha, Ilanna M.; Fonseca, Danielle C.; Formiga, Francisco F.C.; Bonfá, Eloisa S.D.O.; Borba, Eduardo F.; Waitzberg, Dan L. title: SARS-CoV-2 infection, gut dysbiosis and heterogeneous clinical results of hydroxychloroquine on COVID-19 therapy – Is there a link? date: 2020-12-14 journal: Nutrition DOI: 10.1016/j.nut.2020.111115 sha: 56baab612423c1b850a7476c4e050cfb1b4d6e3a doc_id: 924699 cord_uid: 7isjcasa • Growing evidence suggest a link between gut dysbiosis and poor COVID-19 outcomes; • Impact on gut microbiota seems relevant to select candidate drugs for COVID-19 treatment; • Candidate drugs include hidroxychloroquine (HCQ), but available clinical data question its efficacy/safety; • HCQ efficacy/safety may be influenced by the dose provided and its impact on gut dysbiosis; • This suggestion is based on preliminary findings in systemic lupus erythematosus patients. Clinical manifestations of the new severe acute respiratory syndrome coronavirus (SARS-CoV-2) infection can include gastrointestinal signals and symptoms. Individuals with previous clinical conditions that usually enroll gut dysbiosis have been identified at a high-risk to develop more severe infectious phenotypes. Actually, intestinal dysbiosis was observed in infected patients and potentially linked to systemic hyperinflammation. These observations suggest that a previous gut dysbiosis may be aggravated by the SARS-CoV-2 infection and related to the coronavirus disease-19 (COVID-19) progression into more severe stages. While the COVID-19’s pathophysiology is not fully understood it seems relevant to consider the interactions of candidate therapeutic drugs with host, gut microbiota, and SARS-CoV-2. Here we summarize scientific evidence supporting the potential relevance of these interactions and suggest that disputing clinical data on hydroxychloroquine (HCQ) administration in COVID-19 may have been influenced by the dose provided and its impact on gut dysbiosis. The proposition is based on preliminary data of gut microbiota composition in patients with inactive systemic lupus erythematosus under exclusive continuous HCQ treatment, displaying a direct correlation between the drug doses and markers typically associated with gut dysbiosis. 1 Highlights  Growing evidence suggest a link between gut dysbiosis and poor COVID-19 outcomes  Impact on gut microbiota seems relevant to select candidate drugs for COVID- The new severe acute respiratory syndrome coronavirus (SARS-CoV) 2 is responsible for the coronavirus disease 2019 (COVID-19) pandemic, which emerged in Wuhan, China, in December 2019. 1 Beyond the classical respiratory symptoms, a significant proportion of patients with COVID-19 (50.5%) can also have gastrointestinal symptomatology such as diarrhea, nausea and vomiting. 2 These gastrointestinal features have been associated to increased disease susceptibility 3 and poor prognosis. 2, 4 In fact, some respiratory infections are often associated with intestinal dysfunction or secondary to complications, suggesting a lung-intestine crosstalk. 5 Despite that the respiratory tract harbors its own microbial ecology, the gut microbiota has shown to impact the host's systemic immune response, by influencing distant mucous membranes, such as the lungs. 6, 7 Importantly, an healthy gut microbiota profile can contribute to protect the host against respiratory infections 8, 9 while an unbalanced gut microbiota profile (dysbiosis) is associated to several infectious diseases and chronic inflammation. 10, 11 Likewise, gut microbiota also can be sensitive to respiratory viral infections, along with other stressors (exposure to antibiotics and chronic inflammation). 12, 13 Particularly, SARS-CoV-2 binds its spike protein to the surface membrane angiotensin-converting enzyme 2 (ACE2) receptor to gain access and infect human cells. Similarly to the lung, intestinal epithelium also harbors ACE2 receptors, where it seems to influence the composition of resident microbiota. 14 High risk populations, such as elderly, obese, diabetic, hypertensive and metabolic syndrome patients, are prone to have COVID-19 most severe forms, and often present gut dysbiosis. [15] [16] [17] [18] In these patients, dysbiosis is associated to systemic low grade inflammation, with increased intestinal bacterial translocation . 15, 16 Disturbances in gut microbiota composition can contribute to epithelial barrier damage by mechanisms that may include an abnormal hyperactivation of intestinal toll-like receptors (TLR). 18 Therefore, low-grade inflammation, due to increased bacteria translocation, may be a predisposing factor for unbalanced inflammatory responses to SARS-CoV-2 observed in more severe COVID-19 phenotypes. 19 Gou et al. have identified 20 unbalanced gut microbiota components that were predictive for COVID-19 patients to progress to a severe phase compared to non-infected healthy subjects. 20 SARS-CoV-2 translocation contributing to systemic reinfection is a possibility that cannot be ruled out since this virus can be found in feces even when it is no longer detectable in the respiratory tract. 21 Of note, gut microbiota stabilization was one of the six components of a new therapy that effectively increased the rate of cure and reduced mortality of patients with COVID- 19. 22 Inclusion of this intervention was based on a reduction in commensal Lactobacillus and Bifidobacterium, observed in some afflicted patients. 22 Nutritional support and administration of prebiotics and probiotics to balance gut microbiota also has been suggested for COVID-19 patients to reduce the risk of secondary infection due bacterial translocation. 20, 21 While a specific and effective therapy for COVID-19 is lacking, alternative treatments have been proposed. One of the first therapy suggested to manage this new disease included the acute use of the antimalarials drugs chloroquine diphosphate (CQ) or hydroxychloroquine (HCQ). The rational to test CQ/HCQ for SARS-COV-2 infection was based on positive previous experience with these drugs against SARS-CoV-1 (shares 79% genome with the new coronavirus), during the severe acute respiratory syndrome (SARS) outbreak in 2003. 23 Low doses of CQ/HCQ have been used to treat other diseases, such as systemic lupus, with relevant immunomodulatory effects with very low side / adverse effects rate. 24 For some of these diseases, the CQ/HCQ therapeutic effect requires at least three months of continuous use to reach the steady state. 25 Nevertheless, there is a robust discussion on the safety and efficacy in using CQ/HCQ to treat patients with COVID-19, even for short periods. Preliminary data for COVID-19 infection using CQ/HCQ came from non-randomized controlled trials or small sample size trials with controversial results. Some benefits were reported by using low daily doses ( 600 mg/d), 26, 27 but increased toxicity was observed, particularly in the context of high doses ( 1200 mg/d). 28, 29 Importantly, in the Solidarity triala clinical study conducted with the participation of the World Health Organization (WHO), awaiting for peer-reviewed publication -HCQ produced little or no reduction in the mortality of hospitalized COVID-19 patients, when compared to standard of care. 30 Although evidence on increased mortality was not found, some associated safety signals in isolated clinical laboratory findings resulted in the exclusion of HCQ treatment arm from the study. 30 According to WHO, observations from the Solidarity study do not preclude to assess the HCQ effect in other studies in non-hospitalized patients or as pre-or post-exposure prophylaxis for COVID- 19. 30 Considering that gut microbiota might play a role in the COVID-19 pathogenesis, therapies for its treatment or prophylaxis should avoid dysbiosis. As far as we could ascertain, the influence of HCQ use on human gut microbiota composition is unknown. Our research group is investigating dysbiosis signatures (fecal samples 16S genetic sequencing) associated to the inactive (SLEDAI-2k = 0) systemic lupus erythematosus (SLE). In a preliminary analysis, it caught our attention that the abundance of some pro-inflammatory and commensal phyla and genera of bacteria occurred in a heterogeneous grade across the patients. Once they were at same SLE activity grade and under exclusive HCQ treatment at a stable dose, ranging from 2.33 to 5.58 mg/kg/d for 2.62  1.47 years, we searched for potential correlations of HCQ treatment with our negative findings on gut microbiota. In our analysis, only SLE patients (n=12) with C-reactive protein (CRP)  0.3 mg/dL (mean = 0.138  0.097 mg/dL) were considered, to avoid the influence of inflammation bias on gut microbiota composition. 31 With this rigorous parameter, we have calculated the sum of markers typically associated with gut dysbiosis [32] [33] [34] [35] , as an indirect measure of dysbiosis degree (see the Figure legend), and tested it against different doses of HCQ. As observed in the figure, 7 our data suggest that the HCQ dosage is directly correlated with the dysbiosis degree in SLE patients (r = 0.61, CI = 0.04, p = 0.03). This correlation was observed despite the narrow dose range tested and the small sample size, reinforcing the relevance to further evaluate a larger number of patients. Although our observations are very preliminary and were obtained from a different clinical population than COVID-19, they raise the possibility that HCQ administration may influence previous gut dysbiosis in a dose-dependent fashion. We cannot determine whether low HCQ doses may alleviate gut dysbiosis, because data on gut microbiota composition without HCQ intervention is lacking. In a high fat diet-induced mice model of rheumatoid arthritis-associated atherosclerosis, a better gut microbiota profile was observed after HCQ administration and this change was associated to clinical benefits of the treatment. 36 On the other hand, our data in lupus patients suggest that high HCQ doses may contribute to gut dysbiosis. In healthy C57BL/6J mice, despite intestinal integrity and immunological responses were maintained, short-term high dose HCQ affected gut microbiota composition and lactate dehydrogenase (LDH) activity, an indicator for myocardial injury. 37 Different effects of HCQ doses on gut microbiota could explain, at least partially, why available clinical data in COVID-19 are quite heterogeneous on HCQ benefits and side effects according to the low and high doses provided. [26] [27] [28] In looking after a definitive treatment for COVID-19, researchers are concentrating efforts to understand mechanisms or biological predictors associated with prognosis. While we do not fully understand the pathophysiology of this new disease, our data draws attention to the potential relevancy in considering the interactions of candidate drugs with host, gut microbiota, and SARS-CoV-2. The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. 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Clinical efficacy and side effects of antimalarials in systemic lupus erythematosus: a systematic review Pharmacology of Chloroquine and Hydroxychloroquine Efficacy of hydroxychloroquine in patients with COVID-19: results of a randomized clinical trial Hydroxychloroquine and azithromycin as a treatment of COVID-19: results of an open-label non-randomized clinical trial Hydroxychloroquine in patients with mainly mild to moderate coronavirus disease 2019: open label, randomised controlled trial Effect of high vs low doses of chloroquine diphosphate as adjunctive therapy for patients hospitalized with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection: a randomized clinical trial C Reactive Protein (CRP) Preserving microbial diversity Richness and ecosystem development across faecal snapshots of the gut microbiota Akkermansia muciniphila is a promising probiotic Protective effect of hydroxychloroquine on rheumatoid arthritis-associated atherosclerosis Short-term high-dose gavage of hydroxychloroquine changes gut microbiota but not the intestinal integrity and immunological responses in mice Legend. Doses of hydroxychloroquine (HCQ, x axis) ranged from 2.33 to 5.58 mg/kg/d. Gut dysbiosis degree (DYS, y axis) ranged from 0 to 6 points and was determined based on a empirical systematic criteria, where 1 point was attributed for the presence of each of the following items: low Firmicutes / Bacteroidetes ratio (< 0.7); low Firmicutes + Bacteroidetes index (< 85%); low richness (< 360); low diversity (< 5.5); presence of at least one pathological bacterium in abundance (B. fragilis > 0.5%; others > 0.1%); low Akkermansia muciniphila (< 1.0%).