Systemic sclerosis (SSc) is an auto-immune, multi organ disease marked by severe gastrointestinal (GI) involvement and gut dysbiosis. Here, we aimed to determine the safety and efficacy of fecal microbiota transplantation (FMT) using commercially-available anaerobic cultivated human intestinal microbiota (ACHIM) in SSc.
Ten patients with SSc were randomized to ACHIM (n = 5) or placebo (n = 5) in a double-blind, placebo-controlled 16-week pilot. All patients had mild to severe upper and lower GI symptoms including diarrhea, distention/bloating and/or fecal incontinence at baseline. Gastroduodenoscopy transfer of ACHIM or placebo was performed at weeks 0 and 2. Primary endpoints were safety and clinical efficacy on GI symptoms assessed at weeks 4 and 16. Secondary endpoints included changes in relative abundance of total, immunoglobulin (Ig) A- and IgM-coated fecal bacteria measured by 16s rRNA sequencing.
ACHIM side effects were mild and transient. Two placebo controls experienced procedure-related serious adverse events; one developed laryngospasms at week 0 gastroduodenoscopy necessitating study exclusion whilst one encountered duodenal perforation during gastroduodenoscopy at the last study visit (week 16). Decreased bloating, diarrhea and/or fecal incontinence was observed in four of five patients in the FMT group (week 4 or/and 16) and in two of four in the placebo group (week 4 or 16). Relative abundance, richness and diversity of total and IgA-coated and IgM-coated bacteria fluctuated more after FMT, than after placebo.
FMT of commercially-available ACHIM is associated with gastroduodenoscopy complications but reduces lower GI symptoms by possibly altering the gut microbiota in patients with SSc.
All relevant data are within the paper and its Supporting Information files.
Systemic sclerosis (SSc) is a complex, multi-organ disorder characterized by immune-mediated inflammation, progressive organ fibrosis and vascular pathology [
To date, effective treatment alternatives for SSc-related GI disease are lacking and mostly limited to providing partial symptom relief [
Herein, we performed a first-in-man fecal microbiota transplantation (FMT) pilot study with commercially-available anaerobic cultivated human intestinal microbiota (ACHIM) in patients with SSc to determine safety, effects on GI symptoms and on fecal microbiota composition.
This was a single center randomized double-blind placebo controlled pilot trial with active intervention by a standardized FMT culture over 16 weeks with six study visits conducted at Oslo University Hospital between January and May 2018 (See
The trial protocol was approved by the Regional Committees for Medical and Health Research Ethics (REK) on September 8, 2016 (Approval No: 2016/1529) and followed the Helsinki Declaration. All patients gave after verbal information written consent before study start. The study was registered at clinicaltrials.gov (NCT03444220), one month after study start while still all participants and staff was blinded. The authors confirm that all ongoing and related trials for this drug/intervention are registered.
Eligible participants were randomly assigned to treatment. An unblinded staff member (TG) who was not otherwise involved in the clinical aspects of the study generated the random sequence for treatment assignment. All patients and care providers responsible for assessing patients during the study and outcomes were blinded. Active treatment and placebo were packaged, and handled so that participants and study staff were not able to distinguish between treatments. Participants, study staff performing safety and efficacy assessments and analyzing safety and other trial data were blinded to group assignment until the database was locked.
Treatment was installed during gastroduodenoscopy twice with two weeks apart (week 0 and 2). In addition, the patients underwent a third gastroduodenoscopy at the last study visit (week 16) for sampling of biological material (biopsies). The gastroscopy procedure is explained in further detail in
Safety was assessed at each study visit and collected by a standardized safety form and any unfavorable and unintended sign, symptom, or disease temporally associated with the study treatment was registered as an adverse event (AE). A serious AE (SAE) was defined as a state that required hospitalization, death or life threatening situations, or disability or reducing the patient’s capacities.
Efficacy was evaluated by patient-reported outcome using the ULCA GIT 2.0 score questionnaire (UCLA GIT score) and was done at each study visit [
Collection procedures of biological material and details on analysis of short-chain fatty acids (SCFAs), relative abundance of fecal bacteria and Ig-coating is described in
The primary endpoint was safety and clinical efficacy. Safety was collected by a standardized safety form. The primary efficacy endpoint was met if the patients showed improvement of clinical apparent GI involvement at baseline after FMT; measured with the previously validated minimally clinically important difference (MCID) values of the UCLA GIT score (below) [
All analyzes were performed using IBM SPSS Statistics version 25 and STATA version 15. Due to the small sample size, solely descriptive statistics without p-values were applied to clinical data analysis.
The total UCLA GIT score, and the seven single items were assessed at week 0 and the MCID from week 0 to week 4 and week 16. Changes from mild or moderate-severe GI symptoms at week 0 were defined as clinical meaningful if the MCID, the smallest change in score that patients perceive as beneficial, met the “somewhat better” threshold, as previously published [
Thirteen eligible SSc patients were screened and 10 were enrolled in the study; with five patients assigned to active intervention and five to placebo (
Efficacy of FMT measured by the total UCLA GIT score and all seven items of the UCLA GIT score (bloating, diarrhea, reflux, obstipation, fecal soilage, social functioning and emotional well being). Patients receiving FMT (active treatment) marked as A and placebo marked as P. Symptoms at baseline (week 0; w0) are segregated into dark blue (moderate-severe), light blue (mild) and white (no) GI symptoms. Changes to week 4 and week 16 are marked in green (improved), yellow (stable) or red (worsening).
Characteristics at baseline | Total study cohort (n = 9) | FMT intervention (n = 5) | Placebo intervention (n = 4) |
---|---|---|---|
Age, years | 62 (5.7) | 58 (5.6) | 66 (1.5) |
Female gender | 9 (100) | 5 (100) | 4 (100) |
Body Mass Index | 24.0 (2.3) | 23.6 (2.5) | 24.4 (2.1) |
Disease duration, years | 12.0 (10.6) | 7.4 (6.7) | 17.8 (12.6) |
Limited cutaneous SSc | 9 (100) | 5 (100) | 4 (100) |
Anti-centromere antibodies | 8 (89) | 5 (100) | 3 (75) |
mRSS, median (IQR) | 4 (2–5) | 4 (2–8) | 3 (2–4) |
DU | 0 (0) | 0 (0) | 0 (0) |
FVC, % | 97 (11) | 98 (12) | 97 (12) |
DLCO, % | 74 (14) | 76.4 (12) | 72 (18) |
Hemoglobin, g/dL | 13.1 (0.8) | 13.1 (0.8) | 13.1 (1.0) |
CRP, mg/L | 5 (6.7) | 7 (8.8) | 4 (3.1) |
ESR, mm | 13 (6.9) | 14(5.9) | 13 (9.0) |
UCLA GIT score | 0.72 (0.5) | 0.68 (0.4) | 0.77 (0.6) |
Patient global assessment, cm | 3.9 (2.7) | 3.0 (2.8) | 4.8 (2.6) |
Physician global assessment, cm | 2.2 (2.1) | 2.4 (2.8) | 2.1 (1.0) |
Immune modulating drugs | 2 (22) | 1 (20) | 1 (25) |
Proton pump inhibitors | 7 (78) | 3 (60) | 4 (100) |
Calcium channel blockers | 7 (78) | 4 (80) | 3 (75) |
Endothelin receptor antagonist | 2 (22) | 2 (40) | 0 (0) |
Values are n (%) for categorical variables and mean (SD) for continuous variables. FMT: in vitro cultured fecal microbiota transplantation; IQR: interquartile range; n: number; SSc: systemic sclerosis; mRSS: modified Rodnan Skin Score; DU: digital ulcer; FVC: Forced Vital Capacity; DLCO: diffusion capacity of the lung carbon monoxide; CRP: C-reactive protein; ESR: erythrocyte sedimentation rate, UCLA GIT score: University of California Los Angeles Gastrointestinal score; Immune modulating drugs used by the study particitpants included prednisone, methotrexate and rituximab (all medications use was registered)
No patients experienced fever or signs of infection after the interventions. Patients in the FMT group (n = 5) reported more AEs post-intervention than the placebo controls (n = 4), but all the AEs were regarded as mild and transient, including abdominal bloating, diarrhea, nausea and constipation (
By week 4 | At week 16 | |||
---|---|---|---|---|
FMT | Placebo | FMT | Placebo | |
Total adverse events, n (%) | 5 (100) | 3 (75) | 5 (100) | 4 (100) |
Severe adverse events, n (%) | 0 (0) | 0 (0) | 0 (0) | 1 (25) |
Mild adverse events, n (%) | 5 (100) | 3 (75) | 5 (100) | 3 (75) |
Bloating | 2 (40) | 2 (50) | 5 (100) | 1 (25) |
Diarrhea | 2 (40) | 1 (25) | 3 (60) | 2 (50) |
Constipation | 1 (20) | 0 (0) | 5 (100) | 1 (25) |
Nausea | 3 (60) | 1 (25) | 4 (80) | 2 (50) |
Vomiting | 0 (0) | 1 (10) | 1 (0) | 1 (0) |
Abdominal discomfort | 4 (80) | 0 (0) | 5 (100) | 0 (0) |
Fever | 0 (0) | 0 (0) | 0 (0) | 1 (25) |
The patient who experienced laryngospasm during the first gastroduodenoscopy, necessitating exclusion from the rest of the study, is excluded from the table. FMT: in vitro cultured fecal microbiota transplantation, n: number
There were two procedure-related serious adverse events in two patients, both of which had been assigned to placebo. The first SAE was laryngospasms during the baseline gastroduodenoscopy, which necessitated per protocol exclusion from the study (as indicated above). The patient recovered without any sequela. The second SAE was duodenal perforation that occurred during the final gastroduodenoscopy, at the last study visit (week 16) after completing all other study visits. This resulted in hospitalization and treatment with intravenous antibiotics. The patient recovered without any form of sequela.
One of the patients assigned to the placebo group (P2;
At the time of the primary intervention (week 0), all the nine study completers had clinical apparent GI symptoms, consistent with actual presence of SSc-related upper and/or lower GI symptoms (shown as light blue (mild) or dark blue (moderate to severe) in
Effects of the intervention appeared to be most pronounced in patients presenting with lower GI symptoms at baseline such as bloating, diarrhea, and/or fecal incontinence. Specifically, at week 4, four of five patients in the FMT group reported improvement in bloating (
Patient reported fecal incontinence was registered at week 0, 4, 8, 12 and 16 as an exploratory endpoint. Three patients in the FMT group (orange) had fecal incontinence at week 0, with restoration of incontinence at week 4. Two patients in the placebo group (blue) had fecal incontinence at week 0, with restoration of incontinence within week 4 in one patient.
Explorative outcome measure | FMT (n = 5) | Placebo (n = 4) |
---|---|---|
mRSS | -0.8 (-2.4 to 0.8) | -0.8 (-2.3 to 0.8) |
DU, new onset | 0 (0) | 0 (0) |
FVC % | -0.4 (-9.2 to 8.4) | -1.5 (-11.7 to 8.7) |
DLCO % | -2.0 (-6.8 to 2.8) | -6.3 (-11.8 to -0.7) |
CRP, mg/l | -4.6 (-15.0 to 5.8) | -1.5 (-7.4 to 4.4) |
ESR, mm/h | 1.4 (-11.6 to 14.4) | -4.0 (-15.0 to 7.0) |
Patient global assessment, cm | 0.0 (-6.4 to 6.4) | -2.5 (-7.8 to 2.8) |
Physician global assessment, cm | -0.6 (-3.2 to 2.0) | -0.35 (-3.7 to 3.0) |
Results are shown as relative change in the individual measures from week 0 to week 16 with 95% CI. Values are n (%) for categorical variables and mean (95%CI) for continuous variables. CI: confidence interval; FMT: in vitro cultured fecal microbiota transplantation; mRSS: modified Rodnan Skin Score; DU: digital ulcers; FVC: forced vital capacity; DLCO: diffusing lung capacity for carbon monoxide; CRP: C-reactive protein; ESR: Erythrocyte sedimentation rate
We observed relatively stable fecal calprotectin levels in the placebo group at week 0, 4 and 16, but an increase in calprotectin levels in all five FMT treated patients either at week 4 or 16 (
At baseline, there were no significant differences in fecal bacteria composition between the FMT and placebo groups, as measured by intra-individual (alpha) diversity (Shannon index, phylogenetic diversity, number of distinct operational taxonomic units (OTUs)) and global microbiota composition (beta diversity measured by unweighted unifrac analysis)(See
Genera that showed increased relative abundance after FMT were predominantly within the Firmicutes phylum, including genera within the Ruminococcaceae and Lachnospiraceae families. At baseline, relative abundance of IgA and IgM coated fecal bacteria were similar in the FMT and placebo groups. IgA and IgM coating pattern at genera level changed from baseline to weeks 4 and 16 in the FMT group, while the placebo group showed stable relative abundances of these genera. A: Relative abundance of unsorted bacteria. Only bacteria with change (p<0.1) in relative abundance of unsorted bacteria from week 0–4 or week 0–16 are shown. B: Relative abundance of IgA coated bacteria. Only bacteria with changes (p<0.1) in relative abundance of IgA coating from week 0–4 or week 0–16 are shown. C: Relative abundance of IgM coated bacteria. Only bacteria with change (p<0.1) in relative abundance of IgM coating from week 0–4 or week 0–16 are shown.
SSc is marked by high cumulative incidence of severe and hitherto untreatable GI involvement, and alterations of the fecal microbiota composition [
The observed side effects directly related to the FMT by ACHIM were only short lasting and minor, and similar to previously reported FMT side effects [
In this study, the SSc patients mainly reported improvement in lower GI-symptoms. Especially several patients with long standing fecal incontinence had marked improvement of symptoms following FMT. Changes in upper GI-symptoms were less pronounced. Whether this is due to the small sample size, or has other reasons is unclear, but should be addressed in larger trials. As outcome measures we chose the UCLA GIT score because it (i) is the only validated GIT scoring system in SSc and (ii) had readily available definitions of meaningful clinically important difference, expressed as relative changes in the total and individual item scores. We did, however, experience limitations with the UCLA GIT score, particularly regarding identification and longitudinal assessment of fecal incontinence. The UCLA GIT questionnaire only captures symptoms present in the past week. This probably works fine for most items, but from our data set where fecal incontinence was registered as an ever present GI symptom, it seems that the UCLA GIT score underestimated the negative impact of this symptom at baseline, and failed to detect post-intervention changes that were described as important by the patients. Hence, in future studies there seem to be a need for fecal incontinence scores specifically designed to capture clinically meaningful changes. Notably, even though the AEs related to infusion of ACHIM or placebo were short lasting and transient, they were recorded as ever present since last study visit. The UCLA GIT score only covers GI symptoms the previous week. For this reason, there were occasional mismatches between recorded AEs and the UCLA GIT score.
The ideal study duration of FMT trials are unknown. We observed in some patients in this pilot trial that the FMT effects tended to wane, with recurrence of lower GI symptoms towards the end of the study period. The study period of 16 weeks was based on previous FMT trials in inflammatory bowel disease patients. But based on the results of this pilot trial one could argue that endpoints should be estimated at around 4 weeks for short term an at 12 weeks after FMT before recurrence of GI symptoms. The waning effect possibly reflects that gut microbiota in general appears to be resilient to change, and often returns to its pre-intervention state within weeks [
A major advantage of using ACHIM is that it ensures administration of the same bacteria to all study participants, and makes it possible to systemically track the donor-derived microbiome across all the FMT recipients. Given that FMT using ACHIM significantly reduced lower GI symptoms, we sought to determine if specific bugs found in ACHIM were enriched in feces following treatment. We observed post-intervention increase in the relative abundance of three bacterial families (Ruminococcaceae, Lachnospiraceae and Eggerthellaceae) which are dominant in ACHIM (personal communication Tore Midtvedt 2019). Although many of the genera present in ACHIM are known butyrate producers, we did not observe any significant post-intervention changes in SCFA, but this may be due to the small sample size.
Previous studies in murine models suggest activation of adaptive immune responses contributes to the efficacy of FMT [
In this study we did not focus on the mechanistic pathways by which FMT exerts its effects, but such future studies are highly warranted. One could speculate that there is a mechanistic link between dysmotility and dysbiosis in SSc; and that manipulation of gut microbiota with FMT could affect motility patterns, which in turn leads to improvement of GI symptoms.
This study has several limitations including the small number of participants and short study period. Some of the patients included had relatively mild GI symptoms, decreasing the chance of observing a clinical treatment effect. In addition, we only included female patients with longstanding limited cutaneous systemic sclerosis. This may also have led to the rather small changes that were observed in the secondary clinical outcome measures. A larger study is needed to further elucidate the findings from this promising proof-of concept study.
In conclusion, this first in line study indicates that FMT with in vitro standardized cultivated fecal microbiota is safe regarding the manipulation of the gut microbiota, but there are concerns regarding the safety of gastroduodenal administration. We observed alterations of the gut microbiota composition and Ig coating patterns in patients with SSc after FMT. The sample size was too small to make any meaningful conclusion regarding clinical effects of FMT on GI symptoms, but we observed clearly improvement in diarrhea, bloating and fecal incontinence.
Additional information about material and methods.
(PDF)
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(DOC)
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(DOCX)
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Short chain fatty acids analysis at week 0, 4 and 16.
(XLSX)
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(PDF)
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(ZIP)
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Six study visits over a period of 16 weeks. Intervention with ACHIM with two weeks apart (week 0 and 2). Pulmonary function tests performed at week 0 and 12. Fecal samples each week, and collected at all study visits. Clinical exam and UCLA GIT score performed at all study visits (week 0, 2, 4, 8, 12 and 16).
(PDF)
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Individual level of fecal calprotectin (mg/kg) at week 0, week 4 and week 16. A = active treatment group patient. P = placebo group patient.
(PDF)
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Individual beta diversity at week 0 (V1), week 4 (V3) and week 16 (V6).
(PDF)
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The sequencing service was provided by the Norwegian Sequencing Centre, a national technology platform hosted by the University of Oslo and supported by the "Functional Genomics" and "Infrastructure" programs of the Research Council of Norway and the South Eastern Regional Health Authorities. Analyses of fecal SCFA levels were performed at Unger-Vetlesen Institute by bioengineers Gunn Helen Malmstrøm and Jennifer T. Fiennes.