New or vanishing frontiers: LACC1-associated juvenile arthritis

Abstract

Background

The classification and pathogenic basis of juvenile idiopathic arthritis (JIA) are a subject of some controversy. Essentially, JIA is an exclusion diagnosis that represents a phenotypically heterogeneous group of arthritis of unknown origin. Familial aggregation of JIA supports the concept of genetic influence in the pathogenesis of JIA.

Objective

To present the spectrum of laccase domain-containing 1 (LACC1)-associated juvenile arthritis with clinical, biochemical, and molecular genetic data of a cohort of 43 patients, including 11 previously unpublished cases.

Methods

We studied 11 patients with different categories of juvenile idiopathic arthritis from 5 consanguineous families, all from Saudi Arabia, except 2 patients who were of Jordanian ethnicity. Whole-exome sequencing was used to identify the disease-causing variant of LACC1. We also reviewed the clinical spectrum and molecular genetic data of previously published cases of LACC1-associated juvenile arthritis.

Results

This study describes 43 (29 females, 14 males) patients from consanguineous multiplex families. Most of the included patients were of Arab origin with 86% having early onset disease. The most frequent categories were systemic (19 patients) and rheumatoid factor-negative polyarticular (19 patients). Thirty-seven (86%) had progressive erosive arthritis and 10 (23.3%) had persistent limb lymphedema. None of the patients had features of macrophage activation syndrome. Genetic analysis confirmed LACC1 variant in all patients; 22 patients had common founder mutation (LACC1: c.850T > C,p.C284R), while the others showed different LACC1 variants. All patients were treated aggressively with methotrexate and sequential biologic agents. Most of them showed a poor response to treatment.

Conclusion

This report expands the pathogenic variants of LACC1 and the clinical spectrum associated with this genetic subset of juvenile arthritis. The predominance of autosomal-recessive inheritance and strong genetic evidence allowed us to propose LACC1-associated juvenile arthritis as a distinct disorder.

1. Introduction

To date, juvenile idiopathic arthritis (JIA) has largely been regarded as a polygenic disorder. The International League of Association for Rheumatology (ILAR) classification criteria for JIA is widely accepted. However, it cannot classify all children with chronic arthritis. It is worth mentioning that the ILAR criteria for JIA is mainly based on clinical features for simplicity and easy applicability [[1], [2], [3]]. The exact etiology and pathogenesis of JIA are not well defined. However, advances in the last few years have enabled to identify underlying genes and related proteins and to characterize new pathogenic mechanisms [4,5]. Recently, laccase domain-containing 1 (LACC1) variants have gained more attention in familial JIA. Originally, a single LACC1 variant was identified in monogenic systemic JIA, which differs greatly from other JIA categories regarding genetic and immunologic findings [6]. Since its first description in 2015, several clinical reports of siblings from consanguineous families with different subtypes of JIA have been linked to LACC1 variants, but there has been no comprehensive overview of the clinical spectrum and molecular genetic data [[7], [8], [9], [10]].

Here, we present characteristics, including new clinical phenotypic features, laboratory, and molecular genetic findings of a large new cohort of patients with LACC1-associated juvenile arthritis. We also review all published cases of LACC1-associated juvenile arthritis.

2. Methods

The study cohort comprised patients with familial juvenile arthritis who underwent treatment and regular follow-up in the pediatric rheumatology clinics at King Faisal Specialist Hospital and Research Center (KFSH-RC), King Fahad Medical City, Riyadh, Saudi Arabia and Queen Rania Children Hospital, Amman, Jordan. Patients were from consanguineous families. All patients met the ILAR classification criteria for JIA. All patients were assessed for demographic features, age at first disease manifestation, clinical findings, and diagnostic evaluation. All patients underwent genetic testing.

2.1. Ethical considerations

The ethics committee of the Research Affairs Council at KFSH-RC approved the study protocol (RAC# 2020023). Further ethics approval for enrolling patients in the study was obtained from the participating centers as required by their institutional regulations. Full informed consent for genetic testing was obtained from parents at the time of blood extraction. The patient’s enrollment was documented in the medical file according to institutional regulations.

2.2. Collection of data

We comprehensively reviewed all published cases of LACC1-associated juvenile arthritis focusing on the spectrum of phenotypic characteristics and underlying genetic variants. All previously reported cases since 2015 were included and referenced [[6], [7], [8], [9], [10]].

3. Molecular genetic studies

3.1. Participants

A total of 5 unstudied consanguineous multiplex families with 11 patients were included in this study. All were collected through the participating centers between 2015 and 2019. All patients presented with juvenile arthritis although clinical presentation included a wide range of symptoms. DNA was extracted from peripheral blood samples using standard procedures (Flexi Gene DNA Handbook, Qiagen). Samples were quantitated spectrophotometrically and stored at −20 °C.

3.2. Whole exome sequencing and analysis

Briefly, 100 ng of each DNA was amplified in 12 separate wells using Exome Primer Pools, AmpliSeq HiFi mix (Thermo Fisher, Carlsbad, CA, USA) and 10 amplification cycles. All 12 PCR pools were combined in one well and subjected to primer digestion by incubation with FuPa reagent (Thermo Fisher, Carlsbad, CA, USA). Amplified exome targets were ligated with Ion P1 and Ion Xpress Barcode adapters. After purification, libraries were quantified using qPCR with the Ion Library Quantification Kit (Thermo Fisher, Carlsbad, CA, USA). The prepared exome library was further used for emulsion PCR on an Ion OneTouch System, and templated Ion Sphere particles were enriched using Ion OneTouch ES, both procedures following the manufacturer’s instructions (Thermo Fisher, Carlsbad, CA, USA). The template-positive Ion PI Ion Sphere particles were processed for sequencing on the Ion Proton instrument (Thermo Fisher, Carlsbad, CA, USA). Reads were mapped to UCSC hg19 (http://genome.ucsc.edu/), and variants were identified using the Saudi Human Genome Program (SHGP) pipeline [11].

4. Results

Collectively, including this study, only 43 patients (29 females, 14 males) with seven mutations of LACC1 were identified. The mean age of disease onset was 3.2 (SD± 1.8) years. Most of the patients were of Arab origin (22 Saudi, 3 Moroccan, 2 Jordanian, 2 Lebanese), 11 Turkish, and 3 were Indian. According to the ILAR classification criteria, the most frequent categories were systemic (19 patients) and rheumatoid factor (RF)-negative polyarticular (19 patients). There were two patients with RF-positive polyarticular, two patients with oligoarticular subtype, and one patient with enthesitis-related arthritis (ERA) [1]. The most common founder mutation (LACC1:NM_001128303.2:c.850T > C:p.Cys284Arg) was identified in 51.2% (22/43) of patients. The clinical spectrum is shown in Table 1. Members of the same families generally shared an identical clinical spectrum related to a common LACC1 variants. However, high variability in the age of disease onset, clinical presentation, and disease category of arthritis was observed among siblings of 5 families (Family 8, 9, 11, 15, and 17).

Table 1.

Demographics of patients with LACC1-associated juvenile arthritis.

Referencea	Patient no.	Gender	Disease type	Gene mutations
Family 1	4	F	Systemic	LACC1: c.850T > C, p.C284R
Family 2	3	F	Systemic	LACC1: c.850T > C, p.C284R
Family 3	1 1	M F	Systemic Systemic	LACC1: c.850T > C, p.C284R LACC1: c.850T > C, p.C284R
Family 4	2	F	Systemic	LACC1: c.850T > C, p.C284R
Family 5	2	F	Systemic	LACC1: c.850T > C, p.C284R
Family 6	1 1	M F	Poly (RF-) Poly (RF-)	LACC1: c.850T > C, p.C284R LACC1: c.850T > C, p.C284R
Family 7	2	F	Poly (RF-)	LACC1: c.850T > C, p.C284R
Family 8	1 1 1	M F F	Poly (RF-) Poly (RF-) Poly (RF+)	LACC1: c.850T > C, p.C284R LACC1: c.850T > C, p.C284R LACC1: c.850T > C, p.C284R
Family 9	1 1	F F	Systemic Poly (RF-)	LACC1: c.850T > C, p.C284R LACC1: c.850T > C, p.C284R
Family 10	2	M	Systemic	LACC1: c.827delC, p. T276Kfsa2
Family 11	1 1	F F	Systemic Extended Oligo	LACC1: c.827delC, p. T276fsa2 LACC1: c.827delC, p. T276fsa2
Family 12	2 2	F M	Poly (RF-) Poly (RF-)	LACC1: c.3G > A, p.0 LACC1: c.3G > A, p.0
Family 13	3	M	Poly (RF-)	LACC1: c.1240C > T, p. A414T
Family 14	2	M	Systemic	LACC1: c.988del, p. Ile330del
Family 15	1 1	M F	ERA Oligo	LACC1: Homo c.760A > G, p. Ile254Val/Hetero c.1109G > A, p. C370T LACC1: Homo c.760A > G, p. Ile254Val/Hetero c.1109G > A, p. C370T
Family 16	1 2	M F	Poly (RF-) Poly (RF-)	LACC1: c.128delG, p. C43Tfsa6 LACC1: c.128delG, p. C43Tfsa6
Family 17	2 1	F F	Poly (RF-) Poly (RF+)	LACC1: c.832G > C; p.(Ala278Pro) LACC1: c.832G > C; p.(Ala278Pro)

^aFamily 1–5 (Ref No. 6), family 11 (Ref No. 7), family 12–15 (Ref No. 8), family 16 (Ref No. 9), family 17 (Ref No. 10) M = male, F = female, Poly = polyarticular, RF = rheumatoid factor, Oligo = oligoarticular, ERA = enthesitis-related arthritis.

The frequency of clinical and laboratory features, treatment administered, and outcomes are summarized in Table 2. Thirty-seven (86%) patients developed first disease manifestation prior to their second year, and most of the patients (93%) experienced a chronic disease course. No patient had features suggesting inflammatory bowel disease. All patients in the systemic category had fever and associated skin rash. Patients in systemic and polyarticular categories had symmetrical polyarthritis involving small and large joints. Ten (23.3%) patients with polyarticular subtype developed persistent lymphedema involving at least one extremity. Two patients (family 6) had recurrent perioral and periorbital angioedema; all work-up including C1-estrase were unremarkable. In family 7, one patient had bilateral developmental dysplasia of the hip, single kidney, with refractory hypertension but no vascular anomalies, while the other sibling had pulmonary involvement in form of plastic bronchitis. There was no clear phenotype- genotype correlation. Dual pathologies are not infrequent in highly consanguineous populations and may explain phenotypic heterogeneity observed in this family. Of note, patients with the same disease subtypes shared similar clinical and laboratory features regardless of their ethnicity.

Table 2.

The frequency of clinical and laboratory features and treatment.

Features	Frequency	Percentage (%)
Consanguinity	40/43	93
Age at onset (≤2 years)	37/43	86
Fever	22/43	51.2
Skin rash	16/43	37.2
Joint involvement: Large Small	43/43 31/43	100 72.1
Erosive arthritis	37/43	86
Hepatosplenomegaly	8/43	18.6
Lymphadenopathy	7/43	16.3
Disease course (Chronic)	40/43	93
Pericarditis	3/43	6.9
Limb lymphedema	10/43	23.3
Leukocytosis	23/43	53.5
ESR (normal < 15 mm/h)	37/43	86
RF	7/30	23.3
ANA	8/27	29.6
Treatment: Methotrexate Biologic agents (multiple, sequential)	43/43 40/43	100 93
Response: Poor Partial Complete	26/43 13/43 4/43	60.5 30.2 9.3

ESR = erythrocyte sedimentation rate, RF = rheumatoid factor, ANA = antinuclear antibody.

All patients in the systemic category had leukocytosis, thrombocytosis, and elevated inflammatory markers, while patients with other subtypes had normal leukocyte and platelet counts with elevated inflammatory markers. None of the patients had features suggesting macrophage activation syndrome. Imaging studies revealed erosive arthritic changes. All patients had progressive moderate-to-severe disease and required treatment with a systemic corticosteroid and methotrexate. Forty patients received sequential treatment with multiple biologic agents, including anti-tumor necrosis factor (anti-TNF), anti-interleukin-1 (anti-IL1) (anti-IL-6), rituximab, and abatacept. Few patients required treatment with more than three sequential biologic agents. Most of the patients had a poor to partial response, but four patients showed a good response to the corticosteroid, methotrexate, and anti-TNF and anti-IL-6 agents.

5. Discussion

The classification and pathogenic basis of JIA are a subject of controversy. Essentially, JIA is an exclusion diagnosis that represents a phenotypically heterogeneous group of arthritis of unknown origin [12]. ILAR criteria for JIA is mainly based on clinical features. Remarkably, family history of JIA was not considered in the classification criteria [13]. Like other autoimmune and autoinflammatory disorders, the interactions of epigenetic and environmental factors contribute to and influence the susceptibility to disease and its manifestation [4,14]. Familial aggregation of JIA supports the concept of genetic influence in the pathogenesis of JIA and is probably associated with refractory disease [15,16]. Interestingly, an increased risk for JIA was observed among first-degree relatives of patients with JIA [17]. LACC1-associated juvenile arthritis is an autosomal recessive disorder, characterized by inflammatory arthritis. LACC1-associated juvenile arthritis was identified by a combination of homozygosity mapping and whole exome sequencing [6]. LACC1-associated juvenile arthritis is a rare condition. Unfortunately, the frequency of this disorder worldwide is unknown. This report represents the largest published series of LACC1-associated juvenile arthritis. Consanguineous marriage represents an increased risk for rare diseases such as LACC1-associated juvenile arthritis. This study included 40 affected individuals from unrelated consanguineous Arab and Turkish families, which makes it relatively frequent in these ethnicities.

Our new patients and the previously published case series expand the clinical spectrum and the pathogenic variants of LACC1-associated juvenile arthritis. Obviously, it is not limited to systemic subtype as recognized previously, but it is in line with JIA categories. Furthermore, Rabionet et al. performed targeted LACC1 gene sequencing studies in patients with sporadic systemic and RF-negative polyarticular JIA, which revealed no pathogenic LACC1 gene variants [9]. These findings support the causative role of LACC1 gene variants in familial juvenile arthritis. Typically, patients with LACC1-associated juvenile arthritis presented with early onset disease, chronic disease course, and high inflammatory markers with variable phenotypes. Interestingly, a significant number of patients with LACC1-associated juvenile arthritis have been identified with new disease causing variants. However, on the basis of the available data, we could not find clear genotype-phenotype correlations. Our data showed guarded therapeutic responses to intensive treatment.

Our findings have limitations and accordingly results should be interpreted carefully. Part of the data was retrospectively collected. There are variations in management modalities based on the availability of medications and resources.

In summary, LACC1-associated juvenile arthritis is a rare disorder with a wide clinical spectrum. This report is intended to increase the awareness of health care providers about these diseases. Our data suggest that patients with early onset of chronic inflammatory arthritis and a positive family history should be genetically screened for pathogenic LACC1 variants. We believe that conducting the test might help in diagnosis and counselling. It might also help in predicting the treatment response and outcome. Furthermore, the predominance of autosomal-recessive inheritance and strong genetic evidence allowed us to propose LACC1-associated juvenile arthritis as a distinct disorder to differentiate it from non-Mendelian juvenile arthritis of unknown cause “juvenile idiopathic arthritis”.

Ethical statement

All authors have contributed significantly and are in agreement with the content of the manuscript. Furthermore, Ethics committee of our institution has approved the protocol for the research project. Please, note that this work did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Declaration of competing interest

We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no financial support for this work that could have influenced its outcome.We confirm that the manuscript has been read and approved by all named authors and that there are no other persons who satisfied the criteria for authorship but are not listed. We further confirm that the order of authors listed in the manuscript has been approved by all of us.

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