Early changes in gene expression and inflammatory proteins in systemic juvenile idiopathic arthritis patients on canakinumab therapy

Abstract

Background: Canakinumab is a human anti-interleukin-1β (IL-1β) monoclonal antibody neutralizing IL-1β-mediated pathways. We sought to characterize the molecular response to canakinumab and evaluate potential markers of response using samples from two pivotal trials in systemic juvenile idiopathic arthritis (SJIA).

Methods: Gene expression was measured in patients with febrile SJIA and in matched healthy controls by Affymetrix DNA microarrays. Transcriptional response was assessed by gene expression changes from baseline to day 3 using adapted JIA American College of Rheumatology (aACR) response criteria (50 aACR JIA). Changes in pro-inflammatory cytokines IL-6 and IL-18 were assessed up to day 197.

Results: Microarray analysis identified 984 probe sets differentially expressed (≥2-fold difference; P < 0.05) in patients versus controls. Over 50% of patients with ≥50 aACR JIA were recognizable by baseline expression values. Analysis of gene expression profiles from patients achieving ≥50 aACR JIA response at day 15 identified 102 probe sets differentially expressed upon treatment (≥2-fold difference; P < 0.05) on day 3 versus baseline, including IL-1β, IL-1 receptors (IL1-R1 and IL1-R2), IL-1 receptor accessory protein (IL1-RAP), and IL-6. The strongest clinical response was observed in patients with higher baseline expression of dysregulated genes and a strong transcriptional response on day 3. IL-6 declined by day 3 (≥8-fold decline; P < 0.0001) and remained suppressed. IL-18 declined on day 57 (≥1.5-fold decline, P ≤ 0.002).

Conclusions: Treatment with canakinumab in SJIA patients resulted in downregulation of innate immune response genes and reductions in IL-6 and clinical symptoms. Additional research is needed to investigate potential differences in the disease mechanisms in patients with heterogeneous gene transcription profiles.

Trial registration: Clinicaltrials.gov: NCT00886769 (trial 1). Registered on 22 April 2009; NCT00889863 (trial 2). Registered on 21 April 2009.

Keywords: Biomarkers; Canakinumab; Gene expression; Interleukin-1β; Juvenile idiopathic arthritis; SJIA.

Fig. 1

Hierarchical clustering of genes that were upregulated (a) and downregulated (b) at least twofold in baseline systemic juvenile idiopathic arthritis (SJIA) samples compared with healthy control samples. Subjects are shown in columns and genes (probe sets) are shown in rows. Relative gene expression values are color-coded. With upregulated probe sets the hierarchical clustering of subjects perfectly segregated patients with SJIA from healthy controls. Using downregulated genes, the majority of patients with SJIA clustered together; however, a subset of patients with SJIA was indistinguishable from healthy controls

Fig. 2

Pathway map for selected proteins encoded by genes that were upregulated in baseline systemic juvenile idiopathic arthritis (SJIA) samples. Protein interaction networks for selected proteins that were upregulated in baseline SJIA samples compared with healthy control samples. Proteins were mapped using Metacore™ pathway maps (Thompson Reuters, New York, NY, USA). The network displayed here represents a combination of maps for the signaling pathways for IL-1β, IL-6, and the toll-like receptors TL2 and TL4. Small red arrows adjacent to gene/protein names indicate upregulation in SJIA. ADAM10 disintegrin and metalloproteinase domain-containing protein 10, AP-1 activator protein 1, CARD caspase recruitment domain, CASP caspase, C/EBP CCAAT/enhancer binding protein, COX2 cyclooxlygenase-2, CREB cyclic AMP responsive element binding protein, GRB growth factor receptor-bound protein, IL-1beta interleukin-1 beta, IL-R1 interleukin-1 receptor, IL1RN interleukin-1 receptor antagonist, IL-6 interleukin-6, IL-6R interleukin-6 receptor, sIL-6RA soluble interleukin-6 receptor antagonist, IL6ST interleukin-6 signal transducer, IRAK interleukin receptor-associated kinase, IRF interferon regulatory factor, JAK Janus kinase, MCPIP monocyte chemotactic protein-induced protein, MEK mitogen activated kinase kinase, MMP matrix metalloproteinase, NFκB nuclear factor-κB, SOS son of sevenless protein, STAT signal transducer and activator of transduction, TAB TGFβ-activated kinase binding protein, TAK TGFβ-activated kinase, TLR toll-like receptor

Fig. 3

Early transcriptional response to canakinumab treatment of dysregulated genes in systemic juvenile idiopathic arthritis (SJIA). Genes (probe sets) are shown in rows and subjects are shown in columns, ordered according to disease status (SJIA versus healthy), assessment time point (baseline versus day 3), and by response according to American College of Rheumatology (ACR) criteria at day 15. Relative gene expression values are color-coded. a Genes that were upregulated ≥2-fold in baseline SJIA samples compared with healthy control sample. b Genes that were downregulated ≥2-fold in baseline SJIA samples compared with healthy control samples

Fig. 4

Longitudinal change in serum concentrations of IL-6 (a) and IL-18 (b) after treatment with canakinumab. Data are shown for trial 2. Similar results were observed for plasma samples in trial 1 (data available until day 29; not shown). The left panel (b) shows all data; the right panel represents a magnification of the lower portion of the y-axis. IL-6 interleukin-6, IL-18 interleukin 18