Two papers, from different groups and published back-to-back in the journal Nature Genetics, report on the independent discovery of distinct gain-of-function NLRC4 mutations that cause an autoinflammatory syndrome pathologically distinct from NLRP3 cryopyrinopathies.

Scott Canna, first author of one of the papers, says NLRC4 c.1009A>T “causes a recurrent fever syndrome that phenotypically looks like macrophage activation syndrome ... It drives not just IL-1β overproduction, which is also seen with NLRP3 mutations, but also IL-18.”

In the other paper, Neil Romberg et al. discovered NLRC4 c.1022T>C, which encodes a p.Val341Ala substitution just four amino acids from the p.Thr337Ser mutation discovered by Canna et al. Both mutations affect the nucleotide binding (also known as NACHT) domain of NLRC4 (NLR family CARD domain-containing protein 4).

Romberg adds that the disease caused by NLRC4 c.1022T>C “is distinct from previously described autoinflammatory diseases in having pronounced inflammation of the gastrointestinal tract”. He and his colleagues at Yale University, USA, first discovered the mutation after a phone call from an intensive care unit physician to the other corresponding author of their study, Richard Lifton.

Lifton explains, “The physician related the history of a 17-day-old boy who had been transferred from an outside hospital with a critical illness involving fever, secretory diarrhoea, anaemia, thrombocytopenia and coagulopathy. Despite an extensive evaluation, no diagnosis had been made and it was recognized that his clinical condition was deteriorating. It was suspected that he might have an undescribed genetic disease or an unusual presentation of a known syndrome. The physician asked whether we might be able to use DNA sequencing to make a diagnosis.” Lifton adds, “We agreed to try.”

Unfortunately, the infant died at 23 days of age as a result of this illness, but, by exome sequencing to identify all variants in the protein-coding regions of his genome, the researchers were able to identify the rare mutation (c.1022T>C) in NLRC4, which encodes a known bacteria-recognition receptor.

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Initially, the importance of this finding was unclear as the mutation was shared by the infant's healthy father but, Lifton explains, “We were surprised when ... informed that the infant's father was intubated in the intensive care unit with acute respiratory distress syndrome and coagulopathy without an obvious cause.”

The researchers suspected that a bout of an underlying disease related to the expression of the mutant NLRC4 had been triggered. “The father was found to have elevated markers of systemic inflammation,” adds Romberg, “and in the absence of identification of an infectious, or other, cause, was given high-dose immunosuppression, ultimately recovering and leaving the hospital after 9 weeks.”

Romberg and Lifton then contacted an expert in mouse models of NLRC4 function, Barbara Kazmierczak, also at Yale University. Kazmierczak says of her contribution, “We were able to use some of the techniques initially developed for studying inflammasome function in mouse cells to evaluate NLRC4 activity in cells collected from the infant's father and half-brother, who also have this mutation.”

With similar findings to Canna et al., Kazmierczak notes, “We showed that the NLRC4 mutation caused the patient's macrophages to produce IL-1β and IL-18 in the absence of infection, and we observed spontaneous formation of the macromolecular inflammasome complex. This abnormal behavior in the absence of infection was also accompanied by an abnormal response to bacterial infection. The macrophages secreted lower levels of IL-1β and IL-18 than healthy macrophages after bacterial infection, but underwent pyroptotic cell death at higher rates.”

“After presenting the results of this work at a meeting last fall,” recalls Lifton, “I received a call the next day from Dr Goldbach-Mansky at the NIH, who had a patient with both a similar syndrome and a de novo mutation in NLRC4.”

Raphaela Goldbach-Mansky's team, who authored the Canna et al. study, successfully treated their patient with an IL-1 receptor antagonist. She says, “The papers are quite complementary. Having two independent studies that confirm each other's phenotypic, genotypic and functional work tells researchers that they can and should extrapolate on these findings.”

Goldbach-Mansky notes that this work is a success of “the ability to conduct genetic and functional studies with much greater ease in single patients using 'next gen' sequencing and bioinformatics tools to analyse the results.” She concludes, “We hope that our approaches will become part of the routine clinical workup of patients with autoinflammatory diseases in the near future.”