Immunologic monitoring during a phase 2a trial of the GNbAC1 antibody in patients with MS

Abstract

Objective: To monitor the systemic immune responses of patients with multiple sclerosis (MS) under treatment with GNbAC1, a monoclonal antibody against the envelope protein of the MS- associated retrovirus, during a phase 2a trial.

Methods: We analyzed the composition of immune cell subsets and the activation level of monocytes by flow cytometry and the response against viral and vaccine antigens by ELISpot.

Results: None of the endpoints measured revealed any immunosuppressive effect of the therapeutic antibody. Activation of monocytes slightly decreased during treatment as predicted by the hypothesized mechanism of action of GNbAC1.

Conclusion: These results support the conclusion that the antibody is safe for use in patients with MS.

Classification of evidence: This study provides Class III evidence that in patients with MS GNbAC1 does not significantly affect several biomarkers of systemic immune response.

Figure 1. Comparison of immune phenotype, antigen reactivity, and monocyte activation in blood of patients and controls

(A) Cellular composition of peripheral blood mononuclear cells (PBMC) from 5 patients and 4 controls over time (immediately before first infusion: D1, the following day: D2, 2 weeks later: D15, and the conclusion of the study: End). Percentages of CD8⁺ T cells, CD4⁺ T cells, CD33⁺ monocytes, and CD19⁺ B cells in patients (black circles and lines) and controls (purple squares and purple lines) at the 4 time points. Results are only reported from Basel participants. Missing data points are indicated by omitted symbols. (B) Interferon (IFN)-γ response to a vaccine (Tetanol) and a viral antigen (varicella-zoster virus [VZV], Varilrix) throughout the trial in 5 patients and 4 controls. Each condition was assayed in triplicate wells with PBMC exposed to the stimulus using the human IFN-γ ELISpot Ready-SET-Go! (eBioscience, San Diego, CA), and this figure displays the medians from these triplicates at each time point for each of 5 patients and 4 controls. Results are only reported from Basel participants. Missing data points are indicated by omitted symbols. (C) Phosphorylation of p38 mitogen-activated protein kinase (MAPK) in monocytes from 10 patients before and during treatment and from 5 healthy controls at the same time points. Cryopreserved PBMC were permeabilized in methanol and then immunolabeled with anti-CD33 antibodies (eBioscience) to identify monocytes and anti–phospho-p38 MAPK antibodies (BD Biosciences, San Jose, CA) to measure phosphorylation. Vertical axis shows relative phospho-p38 immunofluorescence (arbitrary units). When values from more than 1 time point were available before or during the treatment period, these were averaged, producing 1 before-treatment and 1 during-treatment value for each patient. Time points at which patients were given placebo were included in the pretreatment condition. Error bars show SEM. Averaging for controls followed the patients to whom they were yoked, and the data were analyzed by repeated-measures 2-way analysis of variance comparing effects of group (i.e., patients vs controls) and time point (i.e., before vs during treatment). Neither the main effects nor the interaction were significant. Since the parameter of most interest was the comparison between before and during treatment within the patients, these data were also compared using a paired 2-tailed t test (p = 0.0671). (D) Phosphorylation of p38 MAPK in monocytes of 10 patients and 5 controls across the trial in response to lipopolysaccharide (LPS). Vertical axis is the median immunofluorescence in arbitrary units. Purple lines and squares represent healthy untreated controls. Black lines and circles represent patients. Each of the 4 panels shows results from 1 time point from blood exposed to a range of concentrations of LPS, as indicated. The effect of dose was analyzed by linear regression and found to be significant (p < 0.0005 for both patients and controls at each time point). This effect of dose (i.e., the slope of the dose-response regression lines) was compared between patients and controls and between time points, but neither the main effects nor the interaction were significant.