Monocyte NOTCH2 expression predicts IFN-β immunogenicity in multiple sclerosis patients
Marsilio Adriani, Petra Nytrova, Cyprien Mbogning, Signe Hässler, Karel Medek, Poul Erik H. Jensen, Paul Creeke, Clemens Warnke, Kathleen Ingenhoven, Bernhard Hemmer, Claudia Sievers, Raija L.P. Lindberg Gasser, Nicolas Fissolo, Florian Deisenhammer, Zsolt Bocskei, Vincent Mikol, Anna Fogdell-Hahn, Eva Kubala Havrdova, Philippe Broët, Pierre Dönnes, Claudia Mauri, Elizabeth C. Jury, The ABIRISK Consortium
Marsilio Adriani, Petra Nytrova, Cyprien Mbogning, Signe Hässler, Karel Medek, Poul Erik H. Jensen, Paul Creeke, Clemens Warnke, Kathleen Ingenhoven, Bernhard Hemmer, Claudia Sievers, Raija L.P. Lindberg Gasser, Nicolas Fissolo, Florian Deisenhammer, Zsolt Bocskei, Vincent Mikol, Anna Fogdell-Hahn, Eva Kubala Havrdova, Philippe Broët, Pierre Dönnes, Claudia Mauri, Elizabeth C. Jury, The ABIRISK Consortium
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Research Article Immunology Neuroscience

Monocyte NOTCH2 expression predicts IFN-β immunogenicity in multiple sclerosis patients

Abstract

Multiple sclerosis (MS) is an autoimmune disease characterized by CNS inflammation leading to demyelination and axonal damage. IFN-β is an established treatment for MS; however, up to 30% of IFN-β–treated MS patients develop neutralizing antidrug antibodies (nADA), leading to reduced drug bioactivity and efficacy. Mechanisms driving antidrug immunogenicity remain uncertain, and reliable biomarkers to predict immunogenicity development are lacking. Using high-throughput flow cytometry, NOTCH2 expression on CD14+ monocytes and increased frequency of proinflammatory monocyte subsets were identified as baseline predictors of nADA development in MS patients treated with IFN-β. The association of this monocyte profile with nADA development was validated in 2 independent cross-sectional MS patient cohorts and a prospective cohort followed before and after IFN-β administration. Reduced monocyte NOTCH2 expression in nADA+ MS patients was associated with NOTCH2 activation measured by increased expression of Notch-responsive genes, polarization of monocytes toward a nonclassical phenotype, and increased proinflammatory IL-6 production. NOTCH2 activation was T cell dependent and was only triggered in the presence of serum from nADA+ patients. Thus, nADA development was driven by a proinflammatory environment that triggered activation of the NOTCH2 signaling pathway prior to first IFN-β administration.

Authors

Marsilio Adriani, Petra Nytrova, Cyprien Mbogning, Signe Hässler, Karel Medek, Poul Erik H. Jensen, Paul Creeke, Clemens Warnke, Kathleen Ingenhoven, Bernhard Hemmer, Claudia Sievers, Raija L.P. Lindberg Gasser, Nicolas Fissolo, Florian Deisenhammer, Zsolt Bocskei, Vincent Mikol, Anna Fogdell-Hahn, Eva Kubala Havrdova, Philippe Broët, Pierre Dönnes, Claudia Mauri, Elizabeth C. Jury, The ABIRISK Consortium

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Figure 2

Using LegendScreen to identify the immune signature associated with neutralizing antidrug antibodies (nADA) in patients with multiple sclerosis (MS).

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Using LegendScreen to identify the immune signature associated with neut...
(A) Peripheral blood mononuclear cells (PBMCs) from MS patients treated with IFN-β for at least 12 months (MS-T; Table 1, discovery cohort) and who were either nADA– (n = 12) or nADA+ (n = 16) were analyzed using LegendScreen. Volcano plots comparing fold change (Log2) in marker expression against the significance of marker expression between nADA– and nADA+ MS patients in monocytes, B cells, and T cells (Log10 P value). Only markers listed in Supplemental Table 1 were included in the analysis. Horizontal dashed line represents P value threshold (P = 0.05), vertical dashed lines represent fold change in marker expression (dashed fold change, 0; dotted blue lines fold change, ±1). Markers significantly different between nADA–/nADA+ after 0.20 FDR and with a Log2 fold change ≥ ±0.4 were selected for further analysis (Supplemental Table 2). (B) Summary flow chart of the screening strategy used to identify the nADA-associated phenotypic signature. (C and D) Correlation of differentially expressed markers (DEMs) selected in A and Supplemental Table 2 with disease progression assessed by MRI measurements T1 (C) and T2 (D). Changes in brain lesions detected by comparing MRI scanning at the time of PBMC isolation to MRI scans performed 12 months before sampling. Spearman’s correlation coefficient (r) (see also Supplemental Table 3). (E) DEM expression in treatment-naive (MS-N; n = 15), nADA– (n = 9), and nADA+ (n = 11) IFN-β–treated (MS-T) patients. Box and whisker plots showing expression of NOTCH2, CD200R, CD169, CD284, CD245, CD262, and CD317 on monocytes that passed inclusion criteria in Table 2. One-way ANOVA with Tukey’s correction for multiple comparisons, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. See Supplemental Figure 3 for analysis of excluded markers. (F) Transcriptional regulation of the selected nADA-associated markers (Supplemental Table 4). mRNA was isolated from FACS-sorted CD14+ monocytes from nADA– (n ≥ 3) and nADA+ (n ≥ 3) MS-T patients and analyzed for CD169, NOTCH2, CD200R, CD284, CD262, CD317, and CD245 and for IFN-β–responsive gene MxA as a control analyzed by qPCR relative to cyclophilin or GAPDH. Fold change compared with HCs shown. Two-tailed t tests, **P = 0.01. Box plots show the median and 25th and 75th percentiles; whiskers represent minimum and maximum values.