Assessment of interferon-related biomarkers in Aicardi-Goutières syndrome associated with mutations in TREX1, RNASEH2A, RNASEH2B, RNASEH2C, SAMHD1, and ADAR: a case-control study

Summary

Background

Aicardi-Goutières syndrome (AGS) is an inflammatory disorder caused by mutations in any of six genes (TREX1, RNASEH2A, RNASEH2B, RNASEH2C, SAMHD1, and ADAR). The disease is severe and effective treatments are urgently needed. We investigated the status of interferon-related biomarkers in patients with AGS with a view to future use in diagnosis and clinical trials.

Methods

In this case-control study, samples were collected prospectively from patients with mutation-proven AGS. The expression of six interferon-stimulated genes (ISGs) was measured by quantitative PCR, and the median fold change, when compared with the median of healthy controls, was used to create an interferon score for each patient. Scores higher than the mean of controls plus two SD (>2·466) were designated as positive. Additionally, we collated historical data for interferon activity, measured with a viral cytopathic assay, in CSF and serum from mutation-positive patients with AGS. We also undertook neutralisation assays of interferon activity in serum, and looked for the presence of autoantibodies against a panel of interferon proteins.

Findings

74 (90%) of 82 patients had a positive interferon score (median 12·90, IQR 6·14–20·41) compared with two (7%) of 29 controls (median 0·93, IQR 0·57–1·30). Of the eight patients with a negative interferon score, seven had mutations in RNASEH2B (seven [27%] of all 26 patients with mutations in this gene). Repeat sampling in 16 patients was consistent for the presence or absence of an interferon signature on 39 of 41 occasions. Interferon activity (tested in 147 patients) was negatively correlated with age (CSF, r=−0·604; serum, r=−0·289), and was higher in CSF than in serum in 104 of 136 paired samples. Neutralisation assays suggested that measurable antiviral activity was related to interferon α production. We did not record significantly increased concentrations of autoantibodies to interferon subtypes in patients with AGS, or an association between the presence of autoantibodies and interferon score or serum interferon activity.

Interpretation

AGS is consistently associated with an interferon signature, which is apparently sustained over time and can thus be used to differentiate patients with AGS from controls. If future studies show that interferon status is a reactive biomarker, the measurement of an interferon score might prove useful in the assessment of treatment efficacy in clinical trials.

Funding

European Union's Seventh Framework Programme; European Research Council.

Introduction

First described in 1984,¹ Aicardi-Goutières syndrome (AGS) is a mendelian inflammatory disease most typically characterised by microcephaly, spasticity, dystonia, psychomotor retardation, and, in about 35% of cases, childhood death.² We have shown that AGS is genetically heterogeneous, occurring due to mutations in any one of the genes encoding a 3′ repair exonuclease with preferential activity on single-stranded DNA (TREX1),³ the three non-allelic components of the RNASEH2 endonuclease complex acting on ribonucleotides in RNA:DNA hybrids (RNASEH2A, RNASEH2B, RNASEH2C),⁴ a Sam domain and HD domain containing protein that functions as a deoxynucleoside triphosphate triphosphohydrolase (SAMHD1),⁵ and an enzyme catalysing the hydrolytic deamination of adenosine to inosine in double-stranded RNA (ADAR).⁶

Although some children are affected in utero and display features of illness at birth, most infants with AGS appear to experience the onset of disease in the first 12 months after birth.² Moreover, clinical observation suggests that there is frequently an early period of active regression, seemingly occurring over several months, after which the disease apparently stabilises. For these reasons, and because of the occurrence of later-onset features—particularly vasculitic lesions of the skin (chilblains),⁷ and an intracranial vasculitis seen in a subset of patients8, 9—the development of therapies for AGS is fully warranted.

As part of our ongoing efforts to define rational approaches to the treatment of AGS,¹⁰ we wished to identify a reactive biomarker that could be used to monitor treatment efficacy in future clinical trials. Raised concentrations of interferon α in CSF and serum of patients with AGS were first reported by Lebon and colleagues in 1988.¹¹ This finding not only led to a highly consistent diagnostic marker of early disease, but also presaged fundamental insights into the pathogenesis of AGS, where evidence now suggests that the syndrome results from an accumulation of immunostimulatory nucleic acid species, leading to the induction of an inappropriate type I interferon response.12, 13

Type I interferon is also thought to be central to the development of the autoimmune disease systemic lupus erythematosus, in which affected patients frequently have increased expression of type I interferon-stimulated genes (ISGs) in peripheral blood—a so-called interferon signature.14, 15 In keeping with this observation, some children with AGS develop an early-onset form of lupus.16, 17 Importantly, a combinatorial panel of ISGs is already being used for patient stratification and treatment monitoring in a phase 1 clinical trial approved by the US Food and Drug Administration in patients with systemic lupus erythematosus.18, 19

For these reasons, we aimed to assess interferon-related biomarkers in a large cohort of patients with molecularly defined AGS.