Fecal Microbiota Transplantation Effect on Amyotrophic Lateral Sclerosis Patients

Study Overview
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Brief Summary
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Given the role of adaptive immunity in ALS, the pathogenicity of some clostridial strains on motorneurons, the putative role of cyanobacteria in ALS development, and the increasing interest for microbiota in neurodegenerative disorders, the modification of intestinal microbiota might affect ALS at its core. This interventional study aims at evaluating the biological and disease-modifying effects of Fecal Microbiota Transplant (FMT) in patients affected by Amyotrophic Lateral Sclerosis. As a primary aim of the study, the investigators postulate ALS patients treated with FMT compared to the control arm will display increased Tregs number, which is a favourable biomarker of disease activity and progression. Clinical outcomes as disease progression measured by ALS Functional Rating Scale Revised (ALSFRS-R) score, survival, respiratory function and quality of life will be assessed during the whole treatment and follow-up period. Moreover, biological activity of FMT will be evaluated in different biomatrices, together with FMT safety and tolerability in a cohort of ALS patients.

Detailed Description
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The study will include 42 ALS patients with 2:1 allocation in 2 groups of subjects (28 FMT vs 14 placebo); computerized randomization will be stratified by progression rate (ΔFS) </≥0.7. Randomization time will last 18 months. Treatment will be double blinded to patients and physicians, and will be done at baseline and at month 6. FMT is regarded as the active treatment. Post-treatment follow up will be 6 months. ALS patients will undergo upper GI endoscopy with small-intestine biopsies (n° 4 biopsies of small intestine, performed with a standard biopsy forces) at baseline and after 6 months. At baseline patients will be randomized (2:1) to either an allogenic (from donors) infusion of collected feces (fecal microbiota transplantation, FMT) (or no procedure in case of allocation to placebo) in the duodenum-jejunum. The infusion will be performed through a standard nasojejunal tube, that will be placed during endoscopy. Infusion of feces will be performed at time 0 and repeated at month 6. The patients allocated to placebo arm will not receive treatment, but will undergo intestinal biopsy. Upper GI endoscopy 12 months after FMT will be performed only to identify specific microbioma and mucosal immunological evaluation. Fecal samples and small intestine biopsy samples (at baseline, before treatment, and at month 6 and 12) will be obtained from patients to perform the gut microbiota typing. Every endoscopic procedure will be performed with sedation of the patient. Feces for FMT will be obtained by healthy donors for C. difficile infection. Procedures that are usually performed for the selection of donors for transplantation of feces are as follows. Potential donors stool will be chosen in healthy volunteers that will have given a questionnaire with questions about lifestyle, health status, current therapy, etc., significant clinical symptoms of gastrointestinal disease, etc.. Based on this questionnaire, the potential donor will be considered eligible if excluded: I) Habits of life and risk behaviors, II) Concomitant significant known disorders, III) chronic or recent use of concomitant medications that may interfere with the state of the intestinal microflora (eg, antibiotics), IV) Clinical symptoms indicative of gastrointestinal disease or other diseases of importance, V) Personal or family medical history known of neurodegenerative diseases or other autoimmune diseases. Moreover, each suitable potential donor will be subjected to the following screening tests: I) Examination of stool for Clostridium difficile bacterial pathogens and protozoa and helminths of the small intestine and colon, Vancomycin-resistant Enterococci (VRE ), Methicillin-resistant Staphylococcus aureus (MRSA), Gram-negative Multidrug-Resistant Organisms (MDR), II) Serological screening for hepatitis virus A,B and C, HIV 1-2, Treponema pallidum, H. pylori, blood count with differential, dose transaminasemia, creatinine and C-reactive protein. Potential donors negative for this screening will be considered definitively suitable and will be invited to give a stool sample to prepare than for the fecal transplant. The donation will be made in the appropriate circles in the Department of Internal Medicine and Gastroenterology, and the preparation of faeces (manual homogenization in 500 mL of saline solution) for infusion will be performed at the Unit of Analysis 2 ° (Virology and Microbiology). Analysis of T cell sub-populations will be performed both in peripheral blood and gut mucosa: especially the ratio T Regulatory cells (Tregs)/Th17 cells A Contract Research Organization (CRO) will be in charge for study monitoring.

Official Title
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Interplay Between Gut Microbiota and Adaptive Immunity in Amyotrophic Lateral Sclerosis: a Clinical Trial

Conditions
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Amyotrophic Lateral Sclerosis

Intervention / Treatment
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* Biological: Fecal microbiota transplantation
* Biological: Placebo


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Patients diagnosed with a laboratory supported, clinically probable or definite amyotrophic lateral sclerosis according to the Revised El Escorial criteria (Brooks, 2000) Sporadic or familial ALS Female or male patients aged between 18 and 70 years old Disease duration from symptoms onset no longer than 18 months at the screening visit Patients treated with a stable dose of Riluzole (100 mg/day) for at least 30 days prior to screening Patients with a weight > 50 kg and a BMI ≥18 Patients with a FVC (Forced Vital Capacity) equal or more than 70% predicted normal value for gender, height, and age at the screening visit Patients able and willing to comply with study procedures as per protocol Patients able to understand, and capable of providing informed consent at screening visit prior to any protocol-specific procedures Use of effective contraception both for males and females Exclusion Criteria: Known organic gastrointestinal disease History of gastrointestinal malignancy; ongoing malignancies Use of immunosuppressive or chemotherapy within the past 2 years Celiac disease and/or food (e.g.lactose) intolerance Previous gastrointestinal surgery Any condition that would make endoscopic procedures contraindicated Acute infections requiring antibiotics Antimicrobial treatment or probiotics 4 weeks prior to screening Severe comorbidities (heart, renal, liver failure); severe renal (eGFR< 30ml/min/1.73m2), or liver failure or liver aminotransferase (ALT/AST > 2x Upper limit of normal), Autoimmune diseases, inflammatory disorders (SLE, Rheumatoid arthritis, connective tissue disorder) or chronic infections (HIV, hepatitis B or C infection) Abuse of alcohol or drugs HIV, tuberculosis, hepatitis Participation in clinical trials <30 days before screening Existing blood dyscrasia (e.g., myelodysplasia) White blood cells<4,000/mm³, platelets count<100,000/mm³, hematocrit<30% Patients who underwent non-invasive ventilation, tracheotomy and /or gastrostomy Women who are pregnant or breastfeeding

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 70 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Treatment
Allocation: Randomized
Intervention Model: Parallel Assignment
Interventional Model Description: randomized double blind multicentre study on FMT in ALS, with a randomization ratio 2:1 (FMT: placebo).
Masking: Quadruple


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Active Comparator: Fecal microbiota transplantation (FMT)<br>Fecal microbiota transplantation will be performed during two endoscopic procedures (at baseline and at 6 months) by allogenic infusion of collected feces in the duodenum-jejunum. Fecal microbiota will be diluted in saline solution 200 ml and infused at 30 ml/minute speed. Every endoscopic procedure will be performed with sedation of the patient. Feces for FMT will be obtained by known healthy donors for C. difficile infection according to standard selection procedures. | Biological: Fecal microbiota transplantation<br>* Fecal microbiota transplantation will be performed during two endoscopic procedures (baseline and at 6 months) by allogenic infusion of collected feces in the duodenum-jejunum.<br>* Other names: FMT;|
| Placebo Comparator: Placebo<br>ALS patients will undergo upper GI endoscopy with small-intestine biopsies at baseline and after 6 months. Patients in the placebo arm will not receive any treatment during these procedures, but will undergo intestinal biopsy. Every endoscopic procedure will be performed with sedation of the patient. | Biological: Placebo<br>* patients will undergo endoscopic procedure with biopsy during sedation but without any kind of intervention<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change in Tregs number | to assess whether FMT increases Tregs' number in ALS patients treated with FMT compared to the control arm | 6 months | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change in T cell subsets frequency in blood and gut tissue samples | Change from baseline to each time point (month 3, 6, 9, 12) of the T cell distribution especially the ratio Tregs/Th1 or Tregs/Th17comparing FMT arm and placebo arm. | 12 months (at time points: baseline, month 3 - 6 (both arms)- 9 -12 (both arms)) | 
| Change in heavy neurofilaments levels in CSF | assessment of ongoing disease activity by measuring neurofilaments in CSF only after a proper given consent (lumbar puncture will not be mandatory) | 6 months (at baseline and at month 6) | 
| Changes in levels of pro-inflammatory cytokines and cytokines linked to T cell proliferation and differentiation | Changes from baseline to each time point (month 3, 6, 9, 12) in inflammatory status (cytokines profile in CSF) comparing FMT and placebo arm, only after a proper given consent (lumbar puncture will not be mandatory). We will measure: MIP1a, IL-27, IL-1β, IL-2, IL-4, IL-5, IP-10, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-17A, IFNγ, GM-CSF, TNFα, IFNα, MCP-1, IL-9, P-selectin, IL-1α, IL-23, IL-18, IL-21, sICAM-1, IL-22, E-selectin content using specifically assembled kits (Custom Mix&Match panel Human Panel- 27 Plex) for Luminex Screening Assays (Affymetrix, eBioscience). | 6 months (at baseline and at month 6) | 
| Changes in microbiota profile | analysis of fecal, gut and saliva samples to assess whether FMT consistently modifies microbiota in treated patients versus placebo arm | 12 months (at baseline and at month 6 and 12) | 
| Incidence of Adverse Events | Patients will be monitored with particular attention to possible side effects, including but not limited to increased risk of infections, constipation, diarrhea, pain, nausea, headache, fever. Routine blood samples will be performed at each neurological examination including blood cell count, serum cholesterol and triglycerides, liver and renal function, urine examination, fecal calprotectin. | 12 months ( at screening, baseline, month 1-3-6-7-9-12) | 
| Tracheostomy free survival | Overall survival from randomization to date of documented death or tracheostomy | 12 months | 
| Forced vital capacity (FVC) | respiratory function | 12 months (at baseline and month 3, 6, 9, 12) | 
| disease progression | Amyotrophic lateral sclerosis functional rating scale-revised score, a scale which measures individual functioning through questions regarding communication, eating, motricity and respiration (values: maximum 48 corresponding to no disability; minimum 0 corresponding to extreme disability; higher values represent a good outcome) | 12 months (at baseline and month 3, 6, 9, 12) | 
| quality of life: Amyotrophic Lateral Sclerosis Assessment Questionnaire (ALSAQ-40) | measurement of quality of life by changes in Amyotrophic Lateral Sclerosis Assessment Questionnaire (ALSAQ-40) a scale used to measure the subjective well-being of patients with amyotrophic lateral sclerosis; it includes 40 items / questions. Dimension scores are coded on a scale of 0 (perfect health as assessed by the measure) to 100 (worst health as assessed by the measure). | 12 months (at baseline and month 6 and 12) | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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microbiota, fecal transplantation, adaptive immunity, T regulatory lymphocytes