Combined Regulatory T-Lymphocyte and IL-2 Treatment Is Safe, Tolerable, and Biologically Active for 1 Year in Persons With Amyotrophic Lateral Sclerosis

Abstract

Background and objectives: In a phase 1 amyotrophic lateral sclerosis (ALS) study, autologous infusions of expanded regulatory T-lymphocytes (Tregs) combined with subcutaneous interleukin (IL)-2 were safe and well tolerated. Treg suppressive function increased and disease progression stabilized during the study. The present study was conducted to confirm the reliability of these results.

Methods: Participants with ALS underwent leukapheresis, and their Tregs were isolated and expanded in a current Good Manufacturing Practice facility. Seven participants were randomly assigned in a 1:1 ratio to receive Treg infusions (1 × 106 cells/kg) IV every 4 weeks and IL-2 (2 × 105 IU/m2) injections 3 times/wk or matching placebo in a 24-week randomized controlled trial (RCT). Six participants proceeded into a 24-week dose-escalation open-label extension (OLE). Two additional participants entered directly into the OLE. The OLE included dose escalation of Treg infusions to 2 × 106 cells/kg and 3 × 106 cells/kg at 4-week intervals.

Results: The Treg/IL-2 treatments were safe and well tolerated, and Treg suppressive function was higher in the active group of the RCT. A meaningful evaluation of progression rates in the RCT between the placebo and active groups was not possible due to the limited number of enrolled participants aggravated by the COVID-19 pandemic. In the 24-week OLE, the Treg/IL-2 treatments were also safe and well tolerated in 8 participants who completed the escalating doses. Treg suppressive function and numbers were increased compared with baseline. Six of 8 participants changed by an average of -2.7 points per the ALS Functional Rating Scale-Revised, whereas the other 2 changed by an average of -10.5 points. Elevated levels of 2 markers of peripheral inflammation (IL-17C and IL-17F) and 2 markers of oxidative stress (oxidized low-density lipoprotein receptor 1 and oxidized LDL) were present in the 2 rapidly progressing participants but not in the slower progressing group.

Discussion: Treg/IL-2 treatments were safe and well tolerated in the RCT and OLE with higher Treg suppressive function. During the OLE, 6 of 8 participants showed slow to no progression. The 2 of 8 rapid progressors had elevated markers of oxidative stress and inflammation, which may help delineate responsiveness to therapy. Whether Treg/IL-2 treatments can slow disease progression requires a larger clinical study (ClinicalTrials.gov number, NCT04055623).

Classification of evidence: This study provides Class IV evidence that Treg infusions and IL-2 injections are safe and effective for patients with ALS.

Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.

Figures

Figure 1. Study Design

Seven participants with rapidly progressing ALS were enrolled and underwent leukapheresis. The participants were randomized at a 1:1 ratio to active drug or placebo. Participants received IL-2 or placebo subcutaneous injections between 2 and 4 weeks before week 0. In the RCT, placebo or Treg infusions were administered every 4 weeks beginning week 0 for a total of 6 infusions. The final assessment of the RCT was performed on week 24. Six participants from the RCT entered the open-label extension (OLE). Two additional participants were directly enrolled into the OLE. All 8 participants received IL-2 beginning 2–4 weeks before the first Treg infusion. Treg infusions were administered every 4 weeks in the OLE beginning week 26 (week 0 for 2 participants who entered directly into the OLE) for a total of 6 infusions. The first 2 infusions were a 1× dose of Tregs, the next 2 infusions were a 2× dose, and the final 2 infusions were a 3× dose. The final assessment of the OLE was performed on week 50, and the final study visit occurred on week 54. IL = interleukin; RCT = randomized controlled trial; Treg = regulatory T-lymphocyte.

Figure 2. Treg Suppressive Function and Numbers in the Randomized Controlled Trial

(A) Treg suppressive function was assessed in each participant in the placebo and Treg/IL-2 treatment groups at screening and week 24 (4 weeks after the last infusion of the RCT). (B) Treg numbers were assessed in each participant in the placebo and Treg/IL-2 treatment groups at screening and week 24 (4 weeks after the last infusion of the RCT). Data were depicted as visit-specific estimates ±standard error and were compared for progression of continuous end points using a shared-baseline, linear mixed model. A p < 0.05 was considered significant. IL = interleukin; RCT = randomized controlled trial; Treg = regulatory T-lymphocyte.

Figure 3. Disease Progression During the RCT and OLE

(A) Four participants were randomized to the placebo group and received infusions every 4 weeks as depicted by the vertical dotted lines. Infusions #1–6 were placebo infusions in the RCT, and #7–12 were Treg infusions in the OLE. (B) Three participants were randomized to the active group (Treg/IL-2 treatment) and received infusions every 4 weeks. Infusions #1–6 were Treg infusions in the RCT, and #7–12 were Treg infusions in the OLE. Disease progression was documented in each participant by the ALSFRS-R over the duration of the 54-week study including the screening period. Placebo or IL-2 subcutaneous injections were administered beginning 2–4 weeks before the first infusion at week 0 for the RCT and 2 weeks before the 7th infusion at week 26 for the OLE. (C) Two participants were directly enrolled into the OLE and received Treg infusions every 4 weeks as depicted by the vertical dotted lines. Disease progression was documented in each participant by the ALSFRS-R over the duration of the 28-week study including the screening period. IL-2 subcutaneous injections were administered beginning 4 weeks before the first infusion at week 0. ALSFRS-R = ALS Functional Rating Scale–Revised; IL = interleukin; OLE = open-label extension; RCT = randomized controlled trial; Treg = regulatory T-lymphocyte.

Figure 4. Treg Suppressive Function and Numbers in the Open-Label Extension (OLE)

(A) Treg suppressive function was assessed in each participant at screening, 4 weeks after the second infusion of a 1× dose of Tregs (week 34), 4 weeks after the second infusion of a 2× dose (week 42), and 4 weeks after the second infusion of a 3× dose (week 50). (B) Treg numbers were assessed in each participant at screening, 4 weeks after the second infusion of a 1× dose of Tregs (week 34), 4 weeks after the second infusion of a 2× dose (week 42), and 4 weeks after the second infusion of the 3× dose (week 50). Data were depicted as visit-specific estimates ±standard error and were compared for progression of continuous end points using a shared-baseline, linear mixed model. A p < 0.05 was considered significant. Treg = regulatory T-lymphocyte.

Figure 5. Disease Progression of Participants During the Open-Label Extension (OLE)

Disease progression was documented in each participant by the ALSFRS-R over the duration of the 24-week OLE. The timing of the Treg infusions was depicted by the vertical dotted lines with the corresponding Treg dosages. The baseline ALSFRS-R value for the OLE was taken at week 26 for the 6 participants who went through the RCT and week 0 for the 2 participants who entered directly into the OLE. The total change in the ALSFRS-R was calculated after 24 weeks. Six of the 8 participants showed intermediate to no progression of the ALSFRS-R (average of −2.7 points). Two of the 8 participants showed rapid progression (average of −10.5 points). ALSFRS-R = ALS Functional Rating Scale–Revised; Treg = regulatory T-lymphocyte.

Figure 6. Peripheral Markers of Inflammation and Oxidative Stress During the Open-Label Extension (OLE)

Sera from all 8 participants were collected during the OLE and assayed through an Olink proteomic study for markers of inflammation and oxidative stress. Two markers of inflammation, (A) IL-17F and (B) IL-17C, were higher in the 2 participants who progressed rapidly during the OLE compared with the 6 participants who showed intermediate to no progression. One marker of oxidative stress, (C) oxidized low-density lipoprotein receptor 1 (OLR1), was elevated in the 2 rapidly progressing participants compared with the other 6 participants. An ELISA for another marker of oxidative stress, (D) oxidized low-density lipoprotein (ox-LDL), showed higher levels in the 2 rapidly progressing participants compared with the other 6 participants. Levels of IL-17F, IL-17C, and OLR1 in sera from 9 healthy controls were depicted as shaded areas of the mean ± SD. Levels of ox-LDL in sera from 26 healthy controls were depicted as a shaded area of the mean ± SD. The levels of ox-LDL in the participants were standardized to the control levels. These data were not statistically analyzed as there were only 2 participants in the rapidly progressive group. IL = interleukin.