Outcomes
death, ALS
DISEASE PROGRESSION, APPENDIX
The primary efficacy outcome was the proportion of positive response (Tregs number increase of at least 30%) at treatment end (18 weeks) with respect to baseline, in patients treated with rapamycin compared to the placebo arm. This difference was established based on a previous study demonstrating that slowly progressing ALS patients presented a number of Tregs that was equal to healthy controls, whereas fast progressors had 31% fewer TregsSecondary outcomes were:Assessment of rapamycin safety and tolerability through documentation of the occurrence of any AEs, changes on clinical examination including vital signs and weight, and laboratory examinations (biochemistry, hematology and urinalysis) that were registered throughout the study duration. Symptoms consistent with disease progression, were recorded as AEs.Biological outcomes, assessed as the change from baseline to week 8, 18, 30, 54, comparing rapamycin and placebo arms, of the following biological variables: a) activation and homing capabilities of different T, B, NK cell subpopulations; b) relative expression of inflammasome genes and its activation status; c) phosphorylation of the S6 ribosomal protein (S6RP); d) plasma/CSF neurofilament heavy/light chain protein; e) creatinine and albumin, CK, vitamin D; f) the assessment of rapamycin in CSF samples at week 18 by LC-MS/MS.Laboratory methods are explained in Supplementary Appendix, Section Clinical outcomes through comparison between placebo and treatment arms of: a) the changes in ALSFRS-R from baseline to weeks 4, 8, 12, 18, 30, 42, 54; b) overall survival from randomization to date of documented death or tracheostomy; c) survival rate at weeks 18, 30, 42, and 54; d) respiratory muscle function as assessed by FVC score from baseline to weeks 4, 8, 12, 18, 30, 42, 54 (Supplementary Appendix, Section Quality of life, measured through absolute and relative change from baseline in ALSAQ-40 at week 4, 8, 12, 18, 30, 42 and 54 comparing treatment and placebo arms.