Background/Purpose: Macrophage activation syndrome (MAS), a severe complication of pediatric rheumatic disease, is currently classified among the secondary forms of HLH (sHLH). Primary HLH (pHLH) are caused by mutation of genes coding for proteins involved in cytotoxic functions. Mice carrying heterozygous mutations in more than 1 pHLH gene carry a higher risk to develop HLH following viral infection, suggesting that accumulation of partial genetic defects may be relevant in HLH.

Methods: Genes involved in pHLH in MAS in the context of different rheumatic diseases and in sHLH were analyzed, with next generation sequencing (NGS). A targeted resequencing was performed on all patients using a panel including the 7 principal HLH-related genes (PRF1, UNC13d, STX11, STXBP2, Rab27a, XIAP, SH2D1A) on MiSeq® and NextSeq550® platforms (Illumina, San Diego, CA); all variants identified were confirmed by Sanger. We took into account variants with an allelic frequency in the global population up to 5% in the dbSNP and Ensembl databases.

Results: We analysed 125 patients: 47 MAS, (40 developed this complication in the context of systemic Juvenile Idiopathic Arthritis (sJIA), and 7 in the context of different rheumatic diseases), 32 sHLH, 22 sJIA (without history of MAS) and 24 with different autoinflammatory diseases (AID). sJIA and AID patients were used as control groups. We identified at least 1 heterozygous variant in one of the pHLH-related genes in 41 patients with a detection rate of 52%, 45% of MAS and 62% of sHLH patients. More than one variant was identified in 37% patients from both groups, with 19% of both MAS and sHLH patients carrying polygenic variants. In control groups, 54% of sJIA and 33% of AID patients carry at least a variant in the analysed genes, while polygenic variants have been detected in 14% and 8% of control patients, respectively. The most involved genes in both MAS and sHLH groups were PRF1 and UNC13d, while variants in RAB27a and XIAP have been found only in sHLH patients. The most frequent variants identified in both groups were A91V in PRF1 gene and R928C in UNC13d gene. The A91V variant in PRF1 gene was identified in 19% of both MAS and sHLH patients, while this variant was present, respectively, in only 5% of sJIA and 4% of AID patients. The R928C variant in UNC13d gene was identified in 32% of MAS and 18% of sHLH patients, and in the control group in 9% of sJIA and 17% of AID patients. Variants of PRF1 and UNC13d genes were most frequently observed in patients with both MAS and sHLH, while Rab27a and XIAP variants were more frequent in sHLH. Considering the patients’ clinical characteristics, relapse, CNS involvement, ICU admission and death, in sHLH we observed that three of the 6 patients (50%) carrying multiple variants had recurrent episodes of HLH and that two of them (33%) presented a severe disease with exitus.

Conclusion: Monoallelic variants in pHLH-related genes are more frequent in MAS, sHLH and sJIA and less in AID patients, suggesting different molecular mechanisms involved in the diseases. Re-occurrence and severity of disease seem to be more frequent and more severe in patients who carry mutations in two genes in sHLH group. These data may support a polygenic model of sHLH.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/next-generation-sequencing-analysis-of-familial-haemophagocytic-lymphohistiocytosis-hlh-related-genes-in-macrophage-activation-syndrome-mas-and-secondary-hlh-shlh-2/