Late Onset of Primary Hemophagocytic Lymphohistiocytosis (HLH) with a Novel Constellation of Compound Heterozygosity Involving Two Missense Variants in the PRF1 Gene

Abstract

Hemophagocytic lymphohistiocytosis (HLH) is a rare but in most cases life-threatening immune-mediated disease of the hematopoietic system frequently associated with hematologic neoplasms. Here, we report on a case in which we detected a novel constellation of two missense variants affecting the PRF1 gene, leading to de novo primary HLH. Diagnostics included a comprehensive clinical work-up and standard methods of hematopathology as well as extended molecular genomics based on polymerase chain reaction (PCR) reactions and the calculation of three-dimensional molecule reconstructions of PRF1. Subsequently, a comprehensive review of the literature was performed, which showed that this compound heterozygosity has not been previously described. The patient was a 20-year-old female. Molecular diagnostics revealed two heterozygous missense variants in the PRF1 gene (A91V and R104C) on exon 2. Apart from the finding of two inconclusive genetic variants, all clinical criteria defined by the HLH study group of Histiocyte Society were met at initial presentation. The final diagnosis was made in cooperation with the Consortium of German HLH-reference centers. Here, chemotherapy did not lead to sufficient sustained disease control. Therefore, the decision for allogenic hematopoietic stem cell transplantation (alloHSCT) was made. Hitherto, the duration of response was 6 months. Due to severe and unmanageable hepatic graft-versus-host disease (GvHD), the patient died. We report on a novel constellation of a compound heterozygosity containing two missense variants on exon 2 of the PRF1 gene. To the authors' best knowledge, this is the first presentation of a primary HLH case harboring this genomic constellation with late-onset clinical manifestation.

Keywords: PRF1 gene; compound heterozygosity; primary HLH.

Figure 1

(A) The clinical course and treatment sequence. (B) Total body ¹⁸F-FDG PET/MRI in HLH with involvement of several non-enlarged lymph nodes and extranodal organs with increased multifocal FDG uptake, including bone marrow as well as spleen with associated splenomegaly. * During this period, other immunosuppressive agents not listed here in the Swimmer Plot were used (e.g., etanercept).

Figure 2

(A) The normal perforin expression at mean fluorescence intensity (MFI) was >19.6. The cut-off calculation was performed using ROC analysis on 120 healthy persons and 39 patients. Here, we see a significantly reduced but detectable perforin expression in NK cells, which is consistent with the compound heterozygous mutation in PRF1. (B) Lollipop plot of the PRF1 gene with the alterations detected here in exon 2. (C) Alignment of PRF1 gene sequence based on the topology of Mus musculus (PDB ID: 3NSJ) with the corresponding mutations (A91V, R104C) in Homo sapiens as blue spheres. * The 3NSJ PDB contains multiple rotamers of the sidechains at this position and also ‘T’ stands for turn [22]. (D) On the left: cartoon representation of mouse PRF1 in soluble (PDB ID: 3NSJ) and inserted form (PDB ID: 7PAG) showing large conformational changes between the soluble and the extended form (cyan, yellow) of the perforin monomer. On the right: overlay of the soluble and extended form of the perforin monomer highlighted, with the residues involved in the interaction between monomers of the perforin oligomer highlighted as sticks (black, orange), which also includes the mutation ALA90 (blue). (E) The Cryo-EM map (EMD-13269) of oligomerized mouse perforin in its inserted state fitted with five monomers of a C22 symmetry pore complex (left inset). Vertical view of two of the monomers (pink and cyan) showing residues also identical in Homo sapiens were found to be involved in primary interactions (1°, GLU343, ARG213, black sticks, right inset) at the oligomerization interface, and secondary interactions (2°, ARG103, PHE315, orange sticks) were formed as a result of conformational changes from soluble to extended form [23].