Cytokines, autoantibodies and astrocytes in the pathology of Aicardi-Goutieres syndrome (AGS)

Aicardi-Goutières syndrome (AGS) is a genetically determined encephalopathy
whose importance from a clinical viewpoint is magnified because of the risk of
misdiagnosis as the sequelae of congenital infection. Moreover, from a
biological perspective it is remarkable that some children with AGS develop an
early-onset form of systemic lupus erythematosus (SLE).

AGS and congenital infection are associated with an increased production of
interferon alpha (IFN-*), and a disturbance of IFN-* homeostasis is considered
central to the pathogenesis of SLE. These observations led us to predict that
elucidation of the pathogenesis of AGS would inform our understanding of
autoimmunity.

We have shown that AGS results from mutations in genes encoding the exonuclease
TREX1 (AGS1) and subunits of the RNASEH2 endonuclease protein complex (AGS2,
AGS3 and AGS4), and, the recently identified AGS5 gene and its protein.
Although AGS is normally inherited as a recessive trait, we have described
heterozygous TREX1 mutations in dominantly inherited AGS, in a cutaneous form
of SLE called familial chilblain lupus (FCL), and in patients with an
adult-onset cerebro-retinal microangiopathy associated with Raynaud*s
phenomenon (retinal vasculopathy with cerebral leukodystrophy: RVCL).
Considering the phenotypic and biochemical overlap of AGS with lupus, it is
notable that heterozygous TREX1 mutations were also present in 2% of a cohort
of individuals with SLE. We group AGS, FCL, RVCL and (some cases of) lupus as
Nuclease Immune Mediated Brain and Lupus-like (NIMBL) phenotypes.

This project will focus on AGS (and RVCL) as paradigm for other disorders
sharing the common pathogenesis.

The objectives of the complete NIMBL study are as follows:

Objective 1. Gain insight into the natural history of AGS and RVCL
* Create a registry of paediatric and adult AGS and RVCL patients, and develop
a network of clinicians caring for these patients
* Comprehensively define the clinical, radiological and laboratory features of
AGS and RVCL
* Evaluate disease features which may pertain to heterozygous carriers of
TREX1, RNASEH2A/B/C and AGS5 mutations in AGS families

Objective 2. Acquire knowledge for the development of diagnostic and
therapeutic modalities
* Diagnostics
* Define the molecular spectrum of mutations in TREX1, RNASEH2A/B/C and AGS5 in
AGS and RVCL and correlate this with disease phenotype
* Establish a protocol and quality assurance for the molecular diagnosis of
NIMBL phenotypes
* Produce validated diagnostic criteria
* Identify non-TREX1 / RNASEH2A/B/C / AGS5 mutation-associated AGS patients and
define novel genes causing this disease
* Therapeutics
* Collect data on the efficacy of immunomodulators in AGS
* Collect data on the effectiveness of current treatments for specific features
(e.g. chilblains / Raynaud*s / thrombocytopenia / seizures / headaches) of AGS
and RVCL
* Identify disease features and biomarkers for monitoring future therapeutic
trials
* Use our new model systems to test therapeutic strategies identified by the
NIMBL project

Objective 3. Develop four new animal models relevant to human NIMBL phenotypes
* Ags5 conditional knock-out mouse
* Rnaseh2c knock-in mouse
* Trex1 conditional knock-out mouse
* RVCL Trex1 knock-in mouse

Objective 4. Explain the pathophysiology of NIMBL phenotypes
* Dissect out the intracellular mechanism signalling retained nucleic acids to
immune activation
* Characterise the source, abundance, diversity, size, and dynamics of Trex1
DNA substrates
* Define key aspects of the cell biology of TREX1 and the AGS5 protein
* Determine if / how the pathogenesis of AGS and RVCL differ
* Interrogate cytokine, autoantibody and astrocyte function in AGS

Our local study effort is to measure cytokines and other biomarkers in paired
CSF and serum samples for AGS-related neuroinflammatory reactions (*cytokine
storm*).
In view of the apparent *burning out* of the disease process with time, it will
be of particular importance to measure outputs in early and later stages of the
disease.

Serum samples of AGS patients will be screened for autoantibodies against a
panel of usual autoantigens as well as donor tissues (including skin, heart,
liver) for potentially AGS-specific autoantibodies.

We will characterise the time-course of these parameters (cytokines and
autoantibodies) to determine associations with phenotypic features (including
age at onset and rate of disease progression), and with genotype.
Significant results will then be interrogated further by: 1) examining
production and release of these factors upon cellular activation of fibroblast
and PBMC cultures derived from AGS patients and controls; 2)
immunohistochemistry of AGS brain sections with relevant antibodies and
appropriate cell-specific counterstaining to characterise the cellular origin
of the marker of interest , and 3) staining of patient chilblain lesions upon
any diagnostic skin biopsies for the selected cytokines and for cell-specific
markers including plasmacytoid dendritic cells and T cells.


Materials to be studied will consist of:
- Blood and CSF upon diagnostic sampling
- Monitoring blood samples (every 3-12 months) and score disease-related
features of AGS
- Monitoring CSF samples upon follow-up during clincial disease progression and
MRI (under anesthetics)
- Fibroblast cultures from diagnostic skin biopsies (outpatient care unit)
- Skin biopsies when available for diagnostics in case of chilblains or other
disease-related tissue sampling
- Autopsy material when consented to by parents or caregivers

The burden and risks for the patient will not be increased by participation to
the study.

Local care upon admission for diagnostics and/or monitoring of AGS at the early
stage of the disease will be provided by colleague pediatric neurologists and
pediatricians at the local academic and non-academic hospitals in The
Netherlands.

Once the diagnosis is made, we will ask for consent to visit our clinical
center once every 3-12 months, depending on the contact with local phycisians,
the clinical condition of the patients and the consent of the parents or
caregivers of the affected patients to travel to Amsterdam.