Randomized, double-blind, placebo-controlled trial of rapamycin in amyotrophic lateral sclerosis

Abstract

In preclinical studies rapamycin was found to target neuroinflammation, by expanding regulatory T cells, and affecting autophagy, two pillars of amyotrophic lateral sclerosis (ALS) pathogenesis. Herein we report a multicenter, randomized, double-blind trial, in 63 ALS patients who were randomly assigned in a 1:1:1 ratio to receive rapamycin 2 mg/m²/day,1 mg/m²/day or placebo (EUDRACT 2016-002399-28; NCT03359538). The primary outcome, the number of patients exhibiting an increase >30% in regulatory T cells from baseline to treatment end, was not attained. Secondary outcomes were changes from baseline of T, B, NK cell subpopulations, inflammasome mRNA expression and activation status, S6-ribosomal protein phosphorylation, neurofilaments; clinical outcome measures of disease progression; survival; safety and quality of life. Of the secondary outcomes, rapamycin decreased mRNA relative expression of the pro-inflammatory cytokine IL-18, reduced plasmatic IL-18 protein, and increased the percentage of classical monocytes and memory switched B cells, although no corrections were applied for multiple tests. In conclusion, we show that rapamycin treatment is well tolerated and provides reassuring safety findings in ALS patients, but further trials are necessary to understand the biological and clinical effects of this drug in ALS.

Fig. 1. CONSORT diagram of the study reporting screening, randomization and follow-up of ALS patients enrolled in the trial.

Source data are provided as a Source Data file.

Fig. 2. changes from baseline in blood cells population and inflammasome across treatment arms (P = placebo, R1 = rapamycin 1 mg/m²/d, R2 = rapamycin 2 mg/m²/d, R = combined rapamycin arm).

The figure displays only a selection of the most interesting outcomes (55 cell subpopulation were examined and 11 inflammasome/cytokines, without accounting for multiple outcomes). In detail from left to right: changes from baseline to week 18 (a, n = 32 patients) and 30 (g n = 26 patients) in activated (CD38+, HLA-DR+) CD4 + T cells; changes from baseline to week 18 (b, n = 29 patients) and 30 (h, n = 21 patients) in activated (CD38 + , HLA-DR+) CD8 + T cells; changes from baseline to week 18 (c n = 22 patients) and 30 (i, n = 19 patients) in memory switched B cells (B.C. Mem. Sw.); changes from baseline to week 18 (d, n = 19 patients) and 30 (j, n = 19 patients) in classical monocytes (Class. Mono.); changes from baseline to week 18 (e, n = 27 patients) and 30 (k, n = 20 patients) in IL18 mRNA level (IL18 mRNA); changes from baseline to week 18 (f, n = 49 patients) and 30 (l, n = 39 patients) of plasmatic IL18. Comparison were performed using linear regression models that include indicator variables for treatment arms as the independent variables. For the comparison of Rapamycin and placebo arms, a P value of 0.05 or less was considered to indicate statistical significance and uncertainty in results was expressed with the 95% confidence interval (CI). For the comparisons between Rapamycin 1 mg/m²/d or 2 mg/m²/d arms and the placebo arm, a P value of 0.025 or less was considered to indicate statistical significance and uncertainty in results was expressed with the 97.5% CI, to account for multiple arms comparison with the Bonferroni method. CIs were calculated based on the exact t distribution. All statistical tests were two-tailed. * means p < 0.05 for R and p < 0.025 for R1 and R2; ** means p < 0.01 for R and p < 0.005 for R1 and R2 arms. Error bars represent ± standard deviation. Source data are provided as a Source Data file.

Fig. 3. Mean rates of decline in Amyotrophic Lateral Sclerosis Functional Rating Scale Revised (ALSFRS-R) total score (Intention to Treat population) of patients enrolled in RAP-ALS over the study (baseline to week 54) based on treatment arm allocation (red = R1, rapamycin 1 mg/m²/d, violet = R2, rapamycin 2 mg/m²/d, blue = P, placebo).

Source data are provided as a Source Data file.

Fig. 4. Changes from week 18 to baseline in serum Neurofilament Light (NfL) and phosphorylated Neurofilament Heavy (pNfH) in relation to progression rate across treatment arms.

In detail from left to right, upper panels: changes from week 18 to baseline in serum pNfH in rapamycin (a) and placebo arm (b), in relation to progression rate calculated as the monthly decline in the ALSFRS-R from baseline to week 18. From left to right, lower panels: changes from week 18 to baseline in serum NfL in rapamycin (c) and placebo arm (d), in relation to progression rate calculated as the monthly decline in the ALSFRS-R from baseline to week 18. Individual differences in neurofilament concentration between week 18 and baseline are plotted as colored symbols (Rapamycin arms in red; placebo arm in blue). The shaded areas represent the 95% confidence intervals around the model estimates. The lines and confidence intervals are drawn from the actual distributions of linear model fits. All statistical tests were two-tailed. Source data are provided as a Source Data file.

Fig. 5. Mean scores of ALS Assessment Questionnaire (ALSAQ40) from baseline to study end across treatment arms.

A–E These show the treatment-dependent mean scores of ALSAQ40 physical mobility, Activity Daily Living (ADL) and independence, eating and drinking, communication, and emotional functioning main questions, respectively, from baseline to study end. Panel F shows ALSAQ40 mean total score from baseline to study end (red = R1, rapamycin 1 mg/m²/d, violet = R2, rapamycin 2 mg/m²/d, blue = P, placebo; Intention to treat population). Source data are provided as a Source Data file.