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

Methods: Genes involved in pHLH were analysed with next generation sequencing (NGS) in patients with MAS in the context of different rheumatic diseases and in secHLH. We performed targeted resequencing on all patients using a panel including the 7 principal HLH-related genes (PRF1, UNC13d, STX11, STXBP2, Rab27a, XIAP, SH2D1A). Sequencing analysis were performed on the MiSeq® platform (Illumina, San Diego, CA); all variants identified were confirmed by Sanger. The possible functional impact of variants was studied by in silico analysis using SIFT and PolyPhen softwares. We took into account only variants with an allelic frequency in the global population <1%, in the dbSNP and Ensembl databases.

Results: We studied 25 patients, 20 MAS, 16 of whom developed this complication in the context of sJIA, and 4, in the context of other rheumatic diseases (vasculitis, Crohn’s disease, systemic lupus erithematosus and Kawasaki disease, respectively) and 5 patients with secHLH. We identified at least 1 heterozygous variant in one of the pHLH associated genes in 14 (56%) patients: 10/20 MAS (50%), 4/5 secHLH (80%), with a detection rate of 56%. Nine patients showed variants in one gene, while variants in two or more genes were found in 5 patients. Five patients with MAS showed an heterozygous mutation in PRF1 gene, the A91V variant was the most frequent (4), and 5 patients showed an heterozygous mutation in UNC13d gene. Three of the 20 MAS patients had mutations in two different genes, two of them had recurrent episodes of MAS with one presenting a severe disease with a prolonged ICU admission. Three (60%) patients with secHLH showed an heterozygous mutation of Rab27a, 2 of UNC13d gene and 1 of PRF1. Two of those patients had mutations in two different genes, one of them presented three episodes of HLH reactivation and the other one presented a severe disease and died. Overall patients carrying mutations in 2 genes showed higher frequency of recurrences (4/5, 80%) compared to patients carrying one mutations or no mutation (6/20, 30%) and higher frequency of severe disease (i.e. needed admission to intensive care unit or died) (4/5, 80% versus 7/20, 35%).

Conclusion: Mutations of PRF1 and UNC13d genes are frequently observed in patients with MAS or secHLH; Rab27a variants may be more frequent in patients with secHLH. Re-occurrence and severe disease tend to be more frequent and more severe in patients who carry mutations in two genes. These data are consistent with a polygenic model of secHLH and MAS. Reference. 1. Sepulveda FE et al. Polygenic mutations in the cytotoxicity pathway increase susceptibility to develop HLH immunopathology in mice. Blood 2016.

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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-sechlh/