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Extended report
A new autoinflammatory and autoimmune syndrome associated with NLRP1 mutations: NAIAD (NLRP1-associated autoinflammation with arthritis and dyskeratosis)
  1. Sylvie Grandemange1,2,
  2. Elodie Sanchez2,3,
  3. Pascale Louis-Plence2,4,
  4. Frédéric Tran Mau-Them2,3,
  5. Didier Bessis4,5,
  6. Christine Coubes3,
  7. Eric Frouin6,
  8. Marieke Seyger7,
  9. Manon Girard3,
  10. Jacques Puechberty3,
  11. Valérie Costes4,6,
  12. Michel Rodière8,
  13. Aurélia Carbasse8,
  14. Eric Jeziorski4,8,
  15. Pierre Portales9,
  16. Guillaume Sarrabay1,2,4,
  17. Michel Mondain4,10,
  18. Christian Jorgensen2,4,11,
  19. Florence Apparailly2,4,11,
  20. Esther Hoppenreijs12,
  21. Isabelle Touitou1,2,4,
  22. David Geneviève2,3,4
  1. 1 Centre de référence des maladies autoinflammatoires, CeRéMAI, CHRU de Montpellier, Montpellier, France
  2. 2 INSERM, U1183, IRMB, Hôpital Saint Eloi, CHU Montpellier, Montpellier, France
  3. 3 Département de Génétique Médicale, maladies rares et médecine personnalisée, centre de référence anomalies du développement et syndromes malformatifs, plateforme recherche de remaniements chromosomiques par puces à ADN, Hôpital Arnaud de Villeneuve, CHRU Montpellier, Montpellier, France
  4. 4 Université de Montpellier, Montpellier, France
  5. 5 Service de dermatologie, Hôpital Saint Eloi, CHRU Montpellier, Montpellier, France
  6. 6 Laboratoire d'Anatomie et Cytologie Pathologiques, Hôpital Gui de Chauliac, CHRU Montpellier, Montpellier, France
  7. 7 Dermatology Department, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands
  8. 8 Pédiatrie, Hôpital Arnaud de Villeneuve, CHRU Montpellier, Montpellier, France
  9. 9 Laboratoire d'Immunologie, CHRU Montpellier, Montpellier, France
  10. 10 Service d'Oto-Rhino-Laryngologie, CHRU Montpellier, Montpellier, France
  11. 11 Department for Osteoarticular Diseases, University Hospital Lapeyronie, Montpellier, France
  12. 12 Department of Pediatrics, Pediatric Rheumatology, Radboud University Nijmegen Medical Centre and Sint Maartenskliniek, Nijmegen, The Netherlands
  1. Correspondence to Professor David Genevieve, Département de Génétique Médicale, maladies rares et médecine personnalisée, CHRU Arnaud de Villeneuve, 371 avenue du doyen Gaston Giraud, Montpellier 34000, France; d-genevieve{at}chu-montpellier.fr

Abstract

Objectives Inflammasomes are multiprotein complexes that sense pathogens and trigger biological mechanisms to control infection. Nucleotide-binding oligomerisation domain-like receptor (NLR) containing a PYRIN domain 1 (NLRP1), NLRP3 and NLRC4 plays a key role in this innate immune system by directly assembling in inflammasomes and regulating inflammation. Mutations in NLRP3 and NLRC4 are linked to hereditary autoinflammatory diseases, whereas polymorphisms in NLRP1 are associated with autoimmune disorders such as vitiligo and rheumatoid arthritis. Whether human NLRP1 mutation is associated with autoinflammation remains to be determined.

Methods To search for novel genes involved in systemic juvenile idiopathic arthritis, we performed homozygosity mapping and exome sequencing to identify causative genes. Immunoassays were performed with blood samples from patients.

Results We identified a novel disease in three patients from two unrelated families presenting diffuse skin dyskeratosis, autoinflammation, autoimmunity, arthritis and high transitional B-cell level. Molecular screening revealed a non-synonymous homozygous mutation in NLRP1 (c.2176C>T; p.Arg726Trp) in two cousins born of related parents originating from Algeria and a de novo heterozygous mutation (c.3641C>G, p.Pro1214Arg) in a girl of Dutch origin. The three patients showed elevated systemic levels of caspase-1 and interleukin 18, which suggested involvement of NLRP1 inflammasome.

Conclusions We demonstrate the responsibility of human NLRP1 in a novel autoinflammatory disorder that we propose to call NAIAD for NLRP1-associated autoinflammation with arthritis and dyskeratosis. This disease could be a novel autoimmuno-inflammatory disease combining autoinflammatory and autoimmune features. Our data, combined with that in the literature, highlight the pleomorphic role of NLRP1 in inflammation and immunity.

Trial registration number NCT02067962; Results.

  • Arthritis
  • Fever Syndromes
  • Inflammation
  • Juvenile Idiopathic Arthritis

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Introduction

Inflammasomes are large multiprotein complexes that are key modulators of immune and inflammatory responses. Members of the nucleotide-binding oligomerisation domain-like receptor (NLR) family are the major components of inflammasomes. On infection, they play a critical role by sensing microbial structures called pathogen-associated molecular patterns or endogenous danger molecules released by stressed cells, damage-associated molecular patterns. On cell injury, NLRs trigger the inflammasome assembly, which activates caspase-1 and promotes the processing and release of interleukin 1β (IL-1β) and IL-18, two potent proinflammatory cytokines.1 ,2

In the NLR family, only NLR containing a PYRIN domain 1 (NLRP1), NLRP3 and NLRC4 (NLR containing a caspase recruitment domain) are clearly identified to form an inflammasome complex. Because of the key role of inflammasome in inducing the immune response and inflammation, these genes have been associated with autoinflammatory or autoimmune disorders.3–5 Indeed, mutations in NLRP3 and NLRC4 are linked to a spectrum of dominant autoinflammatory diseases (OMIM 607115, 191900, 1201006 ,7 and OMIM 616050 and 61115,8 ,9 respectively). Moreover, polymorphisms in NLRP1 are associated with autoimmune disorders such as vitiligo, type 1 diabetes, systemic lupus erythematosus and rheumatoid arthritis.3 ,4 ,10–12

To search for novel genes involved in a family exhibiting systemic juvenile idiopathic arthritis (sJIA), we used a next-generation sequencing (NGS) approach and identified NLRP1 mutations in a novel monogenic disorder associating autoinflammation, autoimmunity, dyskeratosis and arthritis.

Methods

Healthy controls and patients

All subjects or the subject's legally authorised representative provided informed, signed consent to be in the study. We obtained approval for the study from the Comité de Protection des Personnes sud méditerranée I (no 13 88—reference approval: 1262, ID of the study 2013-A01516–39 and EFS Convention EFS-PM no. 21PLER2015-0013).

We used histopathology, flow cytometry immunophenotyping, genomic studies and quantification of inflammasome components to better characterise this novel disorder (see online supplementary methods).

Results

Clinical features of the patients

Initially, we studied a multiplex family (family 1) with two affected patients (1 and 2) who are double first cousins born of healthy consanguineous parents originating from Algeria. The pregnancy course and birth of both patients were unremarkable. Both patients presented dermatological anomalies as well as early-onset (age 4.5 and 5 months old, respectively) and recurrent episodes of unprovoked fever lasting 3–4 days. Recurrent elevated C reactive protein level was observed (figure 1A) during febrile and afebrile periods (data not shown). Both patients received a diagnosis of atypical sJIA. Clinical and biological features of patients are in table 1.

Table 1

Summary of the main clinical and biological features of patients

Figure 1

Clinical features of patients. (A) C reactive protein (CRP) level over time in the three patients. CRP level in mg/L (Y-axis) was determined at different times (X-axis) during fever episodes or afebrile periods. The dotted horizontal line indicates the normal range (CRP <5 mg/L). (B) Close-up of the skin manifestations. (a, e, k) Skin dyskeratosis with phrynoderma in patients 1, 2 and 3, respectively. Filliform hyperkeratosis, palmoplantar symmetrical hyperkeratosis and papules with pseudo comedones observed in patients 1 (b), 2 (f) and 3 (h). Skin lesions on feet (c) and left hand (d) of patient 1 suggesting human papillomavirus infection. (g, k) Arthritis presented by enlarged left knee in patients 2 and 3. Hyperkeratosis on soles (i) and ungueal dyskeratosis (j) in patient 3. (C) Histopathological lesions of patients. Vocal cord biopsy for patient 1 (a) shows parakeratosis (vertical bold line) with numerous dense eosinophilic dyskeratosic cells (insert). Skin biopsy of the left leg for patient 2 (b) shows hyperkeratosis (vertical bold line) with mild papillomatosis and acanthosis. The granular cell layer is prominent (arrow). Dyskeratosic eosinophilic cells are numerous, dispersed through all layers including the stratum corneum. Skin biopsy of the arm of patient 3 (c) showing mild epithelial hyperplasia with squamous pearls (arrow) and rare dyskeratosic eosinophilic cells (insert). Original magnification ×20 and ×40 for insert.

Patient 1, a 16-year-old boy, was previously reported as having phrynoderma, an alleged acquired form of follicular hyperkeratosis (figure 1Ba, b) caused by vitamin A deficiency.13 Since he was 2 years old, he had had skin lesions (figure 1Ba–d) and chronic lesions including human papillomavirus (HPV)-like lesions on the larynx (see below) that suggested HPV infections but without any serological conversion. In addition, he had recurrent Giardia intestinalis infection. High and sustained levels of IgG and IgA were reported, which suggested a chronic inflammatory/autoimmune immune response (see online supplementary figure S1). The patient also presented uveitis without signs of corneal dyskeratosis. Oligoarticular arthritis had appeared at the age of 12 years, and at the last examination polyarticular arthritis was present (knees, ankles and wrists). The patient was negative for antinuclear antibodies. Growth values were within the normal range.

Patient 2, an 8-year-old girl, presented congenital and progressive disseminated follicular hyperkeratosis (figure 1Be, f). She had a diagnosis of oligoarticular arthritis of the knees from age 5 (figure 1Bg). At age 6, she presented recurrent inflammation and polyarticular arthritis involving the knees, ankles, wrists and hands. Arthritis was first treated by local arthrocentesis with intra-articular knee corticosteroid injections and thereafter relieved with etanercept, 25 mg orally, twice a month. Values for birth variables were within the normal range, whereas those for growth at age 8 were at −2 SDs for length, weight and occipitofrontal circumference (data not shown). The patient was slightly positive for antinuclear antibodies.

Patient 3, a 10-year-old girl, was referred by a physician from the EUROFEVER consortium14 because of a phenotype strikingly similar to that of the patients from family 1. She was the second child of unrelated healthy Dutch parents (family 2). Pregnancy and birth were unremarkable. At age 6 months, the patient showed failure to thrive, hepatosplenomegaly and filiform hyperkeratosis (figure 1B h–j). At age 19 months, autoimmune haemolytic anaemia and thyreoiditis developed. The patient was positive for antinuclear and antiparietal antibodies and at times was positive for M-protein. Four months later, daily high-spiking fever and inflammation developed (figure 1A). Furthermore, the patient showed chronic candidiasis in the mouth and on buttocks and high levels of IgG sustained over time (see online supplementary figure S1). At the age 4 years, respiratory insufficiency developed due to subglottic oedema, and feeding problems ensued. A massive hepatosplenomegaly was observed and splenectomy was performed. Histopathology revealed extensive extra medullar haematopoiesis, some siderosis and sparse white pulp with very few B cells. At the age of 5 years, arthritis developed in both knees. Methotrexate was started, but the fever and inflammation persisted. Anakinra (50 mg per day) was started, which led to total remission of the arthritis and inflammation and discontinuation of corticosteroids. Growth values were improved; growth is currently at −2 SDs for length. Progressive photophobia developed, and ophtalmological evaluation showed corneal dyskeratosis and neovascularisation. Because of the occurrence of amelogenesis imperfecta, several teeth were removed. Anakinra was recently replaced by canakinumab. Currently, the patient has photophobia.

For the three patients, vitamin A levels were measured over time and found persistently below the normal range (see online supplementary figure S2). Vitamin A supplementation for patient 2 was quickly stopped because it did not improve but rather worsened the disease. Patient 3 has received supplementation since 2010, with no modification of disease.

Skeletal X-rays for patients 2 and 3 showed abnormal lower femoral epiphysis and abnormal metaphyses of knees (see online supplementary figure S3). Patient 3 also presented bilateral abnormal striation of the lower metaphyses. These radiographic features were not described in patients with sJIA.15 ,16

Skin disorders were treated with acitretin (0.2–0.5 mg/kg for patients 1 and 2 and 10 mg once daily for patient 3), which led to markedly reduced skin lesions.

Histopathology

Histology of vocal cords (figure 1Ca, patient 1) and skin biopsy (figure 1Cb, c, for patients 2 and 3) showed ancanthosis, with scattered to confluent dyskeratotic keratinocytes. Dyskeratotic cells were characterised by a pyknotic dark staining nuclei and a dense eosinophilic cytoplasm. This feature affected all epidermal layers except the basal layers and predominated at the superficial layer of the epidermis. Lesions were covered by a dense orthokeratosis with focal parakeratosis and elimination of dyskeratotic cells. In some biopsies, the granular layer was completely replaced by dyskeratotic keratinocytes (data not shown). The vocal cords and interarythenoid space had similar epithelial features, demonstrating epidermal hyperplasia, scattered dyskeratotic keratinocytes and hyperkeratosis. In addition, some skin and laryngeal lesions in patient 1 had a more papillomatous and verrucous architecture with hypergranulosis and elongated dermal papillae similar to HPV-induced papillomas (negative HPV staining by in situ hybridisation; data not shown).

Flow cytometry immunophenotyping

Several populations of immune cells were monitored to evaluate the immunological status of patients 1 and 2 from family 1 as compared with their healthy relatives (data not shown) and age-matched controls.17 The patients showed a high number of neutrophils and low number of T lymphocytes, with minor deviations from age-matched control values (table 2). Patient 3 also showed a low number of T lymphocytes (table 2), and recent blood analyses revealed a high number of neutrophils (>8000 cells/mm3; data not shown). For patients 1 and 2, polyclonal T-cell proliferation and vaccine immune responses were normal (data not shown).

Table 2

Percentage and absolute counts of circulating immune cells for the three patients

For all three patients, circulating B cells were analysed as an indicator of B-cell production and maturation steps. Transitional B cells were the earliest B-cell stage detected in peripheral circulation. These cells can be distinguished from other B cells by concomitant high expression of CD38 and CD24 and lack of CD27 expression.18 The three patients showed abnormal B-cell distribution as compared with their healthy relatives (data not shown) and age-matched controls. Indeed, patients 1 and 3 showed high absolute number of circulating transitional B cells (table 2). Patients 2 and 3 showed a reduced number of circulating CD27+ B cells (marginal zone or memory B cells), which suggested low circulation of antibody-secreting cells and confined to lymphoid tissues such as bone marrow.

Molecular screening

For patient 1, first-line screening revealed no mutations in MEFV, TNFRSF1A and NLRP3, responsible for three rare autoinflammatory diseases: (http://fmf.igh.cnrs.fr/ISSAID/infevers/),19 familial Mediterranean fever OMIM 249100;20 ,21 tumour necrosis factor (TNF) receptor-associated periodic syndrome, OMIM 14268022 and cryopyrin-associated periodic syndrome (CAPS) OMIM 607115, 191900, 120100,6 ,7 respectively (figure 2A).

Figure 2

NLRP1 mutations in the two families. (A) Genealogic trees for families 1 and 2 and segregation of both NLRP1 mutations. Pedigrees of families 1 (left) and 2 (right) and segregation of the mutations c.2176C>T, (p.Arg726Trp) and c.3641C>G (p.Pro1214Arg). Males and females are denoted by squares and circles, respectively. Black symbols represent affected patients. Deceased individuals are denoted by squares and circles with diagonal line. The chromatograms show the homozygous autosomal NLRP1 mutation in patients 1 and 2 from family 1 and de novo mutation in patient 3 from family 2. * and ** (in red) represent heterozygous and homozygous mutant alleles, respectively. (B) Schematic representation of the NLRP1 protein. The domain structure of the NLRP1 protein is represented (NM_033004.3; NP_12749.1). The figure shows the location of the mutations identified in this manuscript (p.Arg726Trp and p.Pro1214Arg) and in isolated corneal intraepithelial dyskeratosis (p.Met77Thr)23 (red arrows). CARD, caspase recruitment domain; FIIND, function to find domain; LRR, leucine-rich repeat domain; PYD, pyrin domain; NACHT domain (neuronal apoptosis inhibitor protein, major histocompatibility class 2 transcription activator, incompatibility locus protein from Podospora anserine and telomerase-associated protein).

By homozygosity mapping in patients 1 and 2, their parents and three unaffected relatives of the family 1, we identified two homozygous regions for the two probands, with a pLod score >2 on chromosomes 8 (pLod score 2.18) and 17 (pLod score 2.77) (see online supplementary figure S4). To search for candidate genes, we performed exome sequencing for both patients from family 1 and one unaffected relative (figure 2A). After filtering rare variants and recessive inheritance, we identified only one candidate gene, NLRP1, with a homozygous substitution (c.2176C>T; p.Arg726Trp; NM_033004.3; 17p13.2), in the homozygous region of chromosome 17. The mutation predicted potentially damaged protein function, segregated with the disease in family 1 (figure 2A). The mutation was localised between the NACHT domain (neuronal apoptosis inhibitor protein, major histocompatibility complex class II transcription activator, incompatibility locus protein from Podospora anserine and telomerase-associated protein) and the leucine-rich repeat (LRR) domain (figure 2B). The mutation was not found in any of the 95 healthy volunteers from Algeria or in web databases (1000 genomes and exome variant server). This genetic variation was reported in five individuals (four Asians and one European; allele frequency 0.00004119) with a heterozygous state in the Exac database (rs776245016). The p.R726W mutation of NLRP1 is located in a conserved region, where several mutations associated with CAPS (NLRP3) and recurrent fever (NLRP12) have been described (see online supplementary figure S5).

Because patient 3 (family 2) presented a phenotype strictly similar to patients 1 and 2, we performed Sanger sequencing of NLRP1 as a candidate gene and found a de novo c.3641C>G; p.Pro1214Arg mutation (figure 2A). This mutation was located in the function to find domain (FIIND) (figure 2B). The variation was never reported in 110 chromosome controls, 1000 genomes, the exome variant server or Exac databases. To exclude possible pathogenic mutation(s) in other genes, we extended the sequencing analysis for this patient with an NGS approach including all known and candidate autoinflammatory genes and two Sanger-sequenced genes associated with rare autoimmune diseases (AIRE and CASP10). We found no clearly pathogenic mutation segregating with the phenotype, which further supports NLRP1 as the causative gene.

In an attempt to identify additional patients with NLRP1 mutations, we also screened 17 additional patients with comparable clinical features but failed to identify other NLRP1 mutations.

Serum levels of inflammasome components

Because NLRP1 was implicated in the inflammasome signalling pathway, we hypothesised that this disease could be an inflammasomopathy and measured the main biomarkers of inflammasome activation. The three patients presented a higher level of caspase-1 and IL-18 in serum than healthy controls and the two heterozygous parents of family 1 (figure 3). We detected high levels of IL-1β in serum from patient 3, but this cytokine was not detectable in serum from healthy controls, the two heterozygous parents of family 1 or patients 1 and 2.

Figure 3

Elevated levels of inflammasome components in serum samples of the three patients. Blood samples from healthy controls (n=11), two heterozygous parents (Heteroz.) and the three patients were obtained in Vacutainer tubes containing silicone coated for the isolating serum. The systemic production of caspase-1, interleukin 18 (IL-18) and IL-1β (detection limit of the assays 7, 15 and 4 pg/mL, respectively) in serum was measured by commercially available ELISA. Samples from each individual were analysed in duplicate, and dots represent the mean of the two calculated values. IL-1β and IL-18 were expressed in ng/mL and caspase-1 in pg/mL.

Discussion

Here we report, in two unrelated families, mutations in NLRP1 associated with a novel Mendelian autoinflammatory disease characterised by recurrent fever, arthritis, dyskeratosis and slight autoimmunity. We call this disease NAIAD for NLRP1-associated autoinflammation with arthritis and dyskeratosis.

To date, only one non-synonymous mutation in the Pyrin domain of NLRP1 (p.Met77Thr, figure 2B) has been identified. The mutation is associated with a dominant disorder with isolated corneal intraepithelial hyperplasia with dyskeratosis and parakeratosis.23 Despite strikingly similar clinical aspects of the skin and the corneal dyskeratosis and dental problems for patient 3, the patients we describe presented a more complex phenotype characterised by a unique combination of features, namely recurrent fever with elevated biological markers of inflammation, arthritis initially classified as sJIA in family 1, intraepithelial dyskeratosis, vitamin A deficiency, slight autoimmunity and high transitional B-cell count in two of the three patients. The similar dermatological manifestations between our patients and those described by Soler et al,23 as well as the genetic susceptibility of NLRP1 in skin disorders such as vitiligo10 and psoriasis,24 suggest that NLRP1 may have a unknown role in skin innate immune responses.

Combining linkage and exome sequencing strategies, we pinpointed NLRP1 as the gene responsible for this novel autoinflammatory syndrome. We identified two distinct mutations in NLRP1: a homozygous (p.Arg726Trp) mutation in family 1 and a de novo (p.Pro1214Arg) mutation in family 2. These mutations are located in two different regions of the NLRP1 protein: between the NACHT and LRR domains and in the FIIND, respectively. The p.Arg726Trp mutation is located in the same linker region as the mutation found in the mouse counterpart, NLRP1a, reported to cause a severe autoinflammatory disease driven by the NLRP1a inflammasome.25

In mice, Nlrp1a was highly expressed in haematopoietic stem cells and progenitor cells of both myeloid and lymphoid origin.25 In addition, Nlrp1a activation induced progenitor cell death. The authors speculated that cell-intrinsic deficiency of progenitor cells in Nlrp1a Q593P/Q593P mice leads to pyroptosis due to constitutive activation of the NLRP1a inflammasome. Nlrp1a-mutated mice showed neutrophilia, lymphopenia and splenomegaly. Interestingly, immune cell phenotyping of our three patients revealed neutrophilia, T-cell lymphopenia and abnormal B-cell distribution. We also found a high number of transitional B cells and low number of circulating CD27+ B-cell subsets. A similar decrease in circulating memory B cells was previously reported for early rheumatoid arthritis,26 systemic lupus erythematosus,27 primary Sjogren's syndrome28 and systemic sclerosis.29

Although the sequence homology between the mouse Nlrp1a and human NLRP1 genes is weak,30 data from the Nlrp1a mice model may support the human NLRP1 as the cause of the novel NAIAD syndrome.

Recently, several genes have been found responsible for complex diseases with coexistence of autoinflammatory and autoimmune features:31 IFIH1 responsible for AGS7 (Aicardi-Goutières syndrome 7, OMIM 615846)32 and CECR1 responsible for adenosine deaminase 2 deficiency (OMIM 615688).33 We believe that autoimmunity is a central feature of the NAIAD syndrome, and this result, together with the known association of polymorphisms in NLRP1 with several autoimmune disorders,3 ,4 suggests that NLRP1 plays a role in the development of autoimmunity. We propose that NLRP1 belongs to a novel group of genes responsible for Mendelian disorders combining autoinflammatory and autoimmune features, recently termed autoimmuno-inflammatory disease.34

Among the NLR family, two genes directly assembling an inflammasome have been associated with human autoinflammatory inflammasomopathies: NLRP3 and NLRC4 in CAPS and AIEFC/FCAS4, respectively.35–37 Mutations in NLRP3 and NLRC4 both lead to the constitutive activation of the corresponding inflammasomes, inducing caspase-1 signalling and release of bioactive IL-1β and/or IL-18. Serum IL-18 and caspase-1 levels were elevated in our three patients, which suggests a hyperactivation of the NLRP1 inflammasome. Anakinra therapy, an IL-1 receptor antagonist, ameliorated the fever in patient 3. These data suggest that NAIAD may be a third NLR inflammasome involved in Mendelian disorders. Comparing levels of IL-1β, IL-18 and caspase-1 between our patients and those with classical sJIA would be of interest because the literature suggests that IL-1β and IL-18 are also involved in the pathogenesis of sJIA.38–40

The precise mechanisms of activation of the NLRP1 inflammasome leading to the common features observed in our three patients were not determined and could be totally different. The role of the human NLRP1 during infection remains unknown;30 however, the mutated NLRP1 inflammasome may be responsible for the recurrent infections observed in our patients.

As reported here in NAIAD, both autosomal recessive and autosomal dominant inheritance have been described in autoinflammatory diseases.41 For example, MEFV mutations (in Mediterranean fever) were observed in both recessive20 ,21 and dominant42–45 modes of inheritance. How both dominant and recessive mutations in MEFV lead to excessive processing of IL-1β and/or IL-18 is still unclear, but some hypotheses are emerging. The MEFV recessive mutations may be responsible for loss of function of its negative regulation on the NLRP3 inflammasome,46–48 whereas dominant MEFV mutations may act as gain-of-function mutations leading to a constitutive activation of the Pyrin inflammasome.44 ,45 ,49 ,50 As in Pyrin, recessive mutation in the linker region may affect an inhibitory function of NLRP1 and a dominant mutation in the FIIND may act directly on the NLRP1 inflammasome. Further experiments are needed to elucidate the pathogenesis of NAIAD.

In conclusion, our three patients provide evidence of the involvement of NLRP1 in NAIAD, a novel Mendelian autoinflammatory disorder. Our data, combined with those from the literature, highlight the pleiomorphic roles of NLRP1 and reveal the association of this gene with several autoinflammatory and autoimmune diseases and modes of inheritance, which suggests complex mechanisms of regulation of NLRP1.

Web resources accession numbers and the URL for data are as follows:

Online Mendelian Inheritance in Man (OMIM), http://www.omim.org

dbSNP, http://www.ncbi.nlm.nih.gov/projects/SNP

1000 Genomes Project variant database, http://www.1000genomes.org/

Entrez Gene, http://www.ncbi.nlm.nih.gov/gene/

Exome variant server, NHLBI GO Exome Sequencing Project (ESP), Seattle, Washington, USA, http://evs.gs.washington.edu

Exome Aggregation Consortium (ExAC), Cambridge, Massachusetts, USA, http://exac.broadinstitute.org/

PolyPhen-2,http://genetics.bwh.harvard.edu/pph2/index.shtml

Acknowledgments

The authors thank the patients and the family members who contributed to this study. Immunophenotyping of patient 3 was kindly provided by University Medical Center St Radboud, Nijmegen. The authors thank Maïlys Cren for the immunomonitoring analyses of patients and healthy relatives from family 1, which involved the eCellFrance platform (ANR-11-INSB-005). The authors thank Cécile Rittore who performed the NGS experiments. Affymetrix microarrays were processed in the Microarray Core Facility of the Institute of Research on Biotherapy, CHRU-INSERM-UM1 Montpellier, http://www.irmb-inserm.fr/. The authors also thank Valérie Macioce for editing the English language.

References

Footnotes

  • Handling editor Tore K Kvien

  • SG and ES contributed equally.

  • Contributors DG, ES, SG, and FTM-T analysed exome data and performed molecular analyses. PP and PL-P performed and analysed immunophenotyping. MG and JP performed the linkage analysis. DB, CC, MS, MM, EH, MR, EJ and AC provided clinical information from the patients. EF and VC performed the histology. GS designed the NGS panel and analysed the results. CJ and FA helped conduct the study. DG and IT conducted the study. SG, PL-P, FA, DG and IT wrote the manuscript.

  • Funding Part of this work was funded by the Arthritis Foundation http://www.fondation-arthritis.org/ and the Direction Générale de l'Organisation des Soins of the inter-regional Programme Hospitalier de Recherche Clinique, named GENEinJIA, ClinicalTrials.gov Identifier: NCT02067962.

  • Competing interests None.

  • Patient consent Obtained.

  • Ethics approval Comité de Protection des Personnes sud méditerranée I (committee number 13 88—reference approval: 1262, ID number of the study 2013-A01516-39) and Etablissement du sang français (EFS; EFS-PM no21PLER2015-0013).

  • Provenance and peer review Not commissioned; externally peer reviewed.

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