A Standing

Deficiency of adenosine deaminase type 2 (DADA2): A description of phenotype and genotype in 15 cases

To describe the clinical features, genotype, and treatment in a series of subjects with confirmed adenosine deaminase 2 (ADA2) deficiency.

Early-Onset Juvenile SLE Associated With a Novel Mutation in Protein Kinase C δ

Juvenile systemic lupus erythematosus (jSLE) is rare before 5 years of age. Monogenic causes are suspected in cases of very early onset jSLE particularly in the context of a family history and/or consanguinity. We performed whole-exome sequencing and homozygosity mapping in the siblings presented with early-onset jSLE. A novel homozygous missense mutation in protein kinase C delta (c.1294G>T; p.Gly432Trp) was identified in both patients. One patient showed a marked clinical response and resolution inflammation with rituximab therapy. This report demonstrates the clinical importance of identifying monogenic causes of rare disease to provide a definitive diagnosis, help rationalize treatment, and facilitate genetic counseling.

SAT0010 A Targeted Next-Generation Sequencing Gene Panel for Autoinflammation

Background Monogenic autoinflammatory diseases (AID) are severe lifelong systemic inflammatory disorders with dysregulated innate immunity, causing significant morbidity, mortality, and economic burden. The number of AID is rapidly expanding. Securing a molecular diagnosis is of major importance for treatment, prognosis, and genetic counselling. Routine genetic screening is time-consuming, costly, and lacks sensitivity since only common disease harbouring exons of a minority of the known AID genes are currently tested using Sanger sequencing. Next-generation sequencing (NGS) offers the ability to rapidly and cost-effectively screen all exons of a gene panel containing hundreds of genes. This approach has not yet been routinely introduced in the UK for AID. Objectives To develop and evaluate the performance of a NGS gene panel for AID. Methods The Agilent SureDesign tool was used to design an NGS panel targeting 113 genes, grouped into 9 broad clinical phenotypes: AID; monogenic vasculitis/vasculopathy; complement defects; monogenic lupus; hemophagocytic lymphohistiocytosis (HLH); early-onset inflammatory bowel disease; autoimmune lymphoproliferative syndromes; monogenic stroke; and hereditary amyloidosis. The targeted region includes coding exons, conserved non-coding exons, upstream promoter regions, and splice sites. Captured and indexed libraries (QXT Target Enrichment System) were sequenced as a multiplex of 16 samples on an Illumina MiSeq sequencer in paired-end mode. Positive controls for panel validation comprised 19 DNA samples from patients with confirmed mutations in a variety of genes on the panel. We then applied the panel to test 29 prospective samples with suspected but unconfirmed monogenic inflammation. Read alignment, variant calling, and annotation were performed using Agilent SureCall v3.0 software. Results In the validation stage, our targeted panel detected all known mutations in the 19 positive control samples with an average read depth of 244X (+/− 29X) across the captured genes. The panel was effective at detecting different types of variants, including rare and common single nucleotide variants (SNVs), insertion/deletions, splice-junction variants, upstream promoter region variants, and somatic mosaicism. Prospective testing of the panel in 29 patients revealed potential pathogenic variants in 14 of the 29 patients giving a detection rate of 48%. These included atypical patients with novel mutations in specific exons/genes, not currently routinely available for testing by Sanger sequencing in the NHS. Conclusions This study demonstrates the clinical utility of a comprehensive NGS-based targeted gene panel which was both highly sensitive and specific in detecting different sequence variants of clinical significance. We are currently integrating this panel into a routine diagnostic laboratory testing strategy for monogenic inflammation. This will facilitate accurate and timely molecular diagnosis, more targeted treatment, and ultimately better outcomes for our patients. Acknowledgement This work was supported by Rosetrees Trust and Swedish Orphan Biovitrum (SOBI). Disclosure of Interest None declared

Monogenic polyarteritis nodosa caused by ADA2 Deficiency: the GOSH experience

Recessive mutations in Cat Eye syndrome Critical Region 1 (CECR1), the gene encoding adenosine deaminase 2 (ADA2) have been recently reported to cause polyarteritis nodosa (PAN) with highly varied clinical expression.

Identification of a novel monogenic autoinflammatory disease due to mutation in a mitochondrial chaperone protein in a single kindred, and cure with allogeneic haematopoietic stem cell transplantation

The monogenic autoinflammatory syndromes are characterised by seemingly unprovoked inflammation which derives from a disruption of innate immunity. Novel as yet undefined autosomal recessive syndromes are increasingly recognised in consanguineous families. This type of family is ideal for genetic mapping.

Severe autoinflammatory disease caused by mutation in a gene controlling actin cytoskeletal dynamics and cure with allogeneic haematopoetic stem cell transplantation

The actin cytoskeleton is crucial at many junctures of normal immune function, and consequently there are many immune specific regulators of actin dynamics. A growing number of primary immunodeficiencies are being defined as caused by mutations in the genes encoding these regulators. In addition to immunodeficiency, immune dysregulation and autoinflammation are increasingly recognised to arise from defects within this pathway.

Loss of function mutation in a mitochondrial chaperone protein leading to dyregulated ROS production, and autoinflammatory disease in a single kindred

Reactive oxygen species (ROS) are known to have many roles in the propagation of the inflammatory process. Indeed increased leucocyte ROS have been implicated in a number of autoinflammatory diseases (TRAPS, FMF, CAPS and FCAS2).

Whole Exome Sequencing reveals a NLRP3 mutation in exon 5 in a patient with CINCA

Cryopyrin-associated periodic Syndromes (CAPS) are caused by heterozygous mutations in the NLRP3 gene. More than 80 disease causing mutations have been identified, mostly clustered in NLRP3 exon 3, but also described in exons 4 and 6. However, up to 50% of clinically diagnosed CAPS patients (with identical clinical features and response to anti-IL-1 treatment) show no mutation in NLRP3 detected by conventional DNA sequencing analysis of exons 3, 4 and 6. Recently somatic NLRP3 mosaicism has been shown to account for up to 70% of these mutation negative CAPS patients.

Late onset of the cryopyrin-associated periodic syndrome (CAPS) associated with low level of somatic mosaicism in six patients

CAPS is caused by mutations in the NLRP3 gene and is inherited in an autosomal dominant fashion. About 40% of children with CINCA are mutation negative by conventional Sanger sequencing, but NLRP3 somatic mosaicism can be identified by sensitive multi-parallel sequencing (MPS) in a significant proportion of such patients.