Objective NLRP3 mosaicism is a well-established mechanism causing cryopyrin-associated periodic syndromes (CAPS). The number of reported patients with mosaicism is small, and the knowledge about the long-term disease behavior is limited. Herein we have assembled the largest cohort of individuals with NLRP3 mosaicism reported to date to obtain additional evidence that strengthens the understanding of this disease.

Methods Patients’ data were collected from their medical charts. Genetic analyses were performed using Sanger and next-generation sequencing. In vitro analyses determined the functional consequences of detected variants.

Results Seventeen individuals with NLRP3 mosaicism were enrolled, with 16/17 experiencing different CAPS phenotypes. An overrepresentation of late-onset forms was detected (37.5%). Overall, clinical manifestations, analytical results, and outcomes of treatments were markedly similar to those detected in patients with germline variants. A large mutational diversity was identified, with 16 different variants among 17 individuals. Two main patterns of mosaicism (extended vs myeloid-restricted) were detected, with the last one overrepresented in the late-onset group. The evaluation of mosaicism over time identified three different patterns, being the group with stable mosaicism the largest one.

Conclusions Collected evidence supports the marked similarities among patients carrying somatic or germline NLRP3 variants. The overrepresentation of NLRP3 mosaicism in late-onset forms should be considered in patients with inflammatory manifestations starting in adulthood. Analysis of mosaicism at the biological level confirms the two known patterns of corporal distribution and reveals that mosaicism remains stable over time in most patients, but it may also vary during the course of the disease.

Key Messages What is already known about this subject?

Cryopyrin-associated periodic syndromes (CAPS) is a dominantly-inherited autoinflammatory disease with three different phenotypes of increasing severity across a continuous spectrum.

The disease is a consequence of monoallelic, gain-of-function NLRP3 variants leading to a constitutive hyperactivation of the NLRP3-inflammasome, with the subsequent IL-1β overproduction.

Recent investigations have established the key role of NLRP3 mosaicism in the pathogenesis of CAPS. However, the number of individuals with NLRP3 mosaicism reported to date is small, and the data on the long-term behavior of the disease at clinical and biological levels are scarce.

What does this study add?

We have assembled the largest cohort of individuals with NLRP3 mosaicism reported to date to obtain novel evidence that strengthens the understanding of this syndrome.

The clinical manifestations, the results of analytical tests, and the outcomes of treatments in patients with NLRP3 mosaicism were in line with previous results collected from patients with germline NLRP3 variants.

We have detected a previously unidentified overrepresentation of late-onset forms among patients with NLRP3 mosaicism.

Genetic evidence supports for a large mutational diversity among individuals with NLRP3 mosaicism. Despite the fact that, theoretically, any amino acid residue in the protein may be involved in mosaicism, the regions spanning the amino acid residues 300-310 and 560-570 concentrate the larger proportion of post-zygotic variants.

Additional experiments confirmed the two main patterns of mosaicism distribution (myeloid-restricted vs extended) and that the degree of mosaicism remains stable in most patients but can also change over time.

How might this impact on clinical practice?

Considering that genetic analyses represent the unique way to establish a CAPS diagnosis, the data presented here strongly suggest for an adequate analysis of the sequence reads of the NLRP3 gene in candidate patients, which should include differential analyses for either germline and post-zygotic variants.

The overrepresentation of late-onset forms among patients with NLRP3 mosaicism should be taken into consideration by clinicians treating adult patients in the differential diagnosis of inflammatory diseases starting during adulthood.

The evidence presented here support periodic evaluation of circulating NLRP3 mosaicism to evaluate the disease behavior at biological level over time.

The authors have declared no competing interest.

Funding This work has been partially funded by: PID2021-125106OB-C31 (JIA), PID2020-116709RB-I00 (PP), CNS2022-135101 (PP), PID2023-147531OB-I00 (PP), PID2021-125106OB-C32 (FC), RED2022-134511-T (PP, JIA) and PID2021-125106OB-C33 (OF) grants from the Ministerio de Ciencia e Innovacion (MCIN); Agencia Estatal de Investigacion (AEI); 10.13039/501100011033; Fondo Europeo de Desarrollo Regional (FEDER), UE; European Union Next Generation EU/PRTR. AC21_2/00042 (JIA) and IFI22/00031 (JG-V) grants from the Instituto de Salud Carlos III co-funded by Union Europea Next Generation EU; Mecanismo para la Recuperacion y la Resiliencia (MRR); Plan de Recuperacion, Transformacion y Resiliencia (PRTR). 2021SGR01093; Agencia de Gestio Ajuts Universitaris i de Recerca; Generalitat de Catalunya (FC). 21897/PI/22 (PP) and 21214/FPI/19 (LH-N); Fundacion Seneca, Region de Murcia, Spain.

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The Ethical Review Board of Hospital Clinic approved the study (code HCB/2022/0855).

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Data sharing statement The data included in the present work are, by definition, deidentified, and are available upon reasonable request to the corresponding author. Data shall only be made available after approval by the study contributors of a submitted research proposal, with investigator support and after a signed data access agreement.