Genetics of HLH (Hemophagocytic Lymphohistiocytosis) Dr Mohammad Keramatipour MD, PhD Tehran University of Medical Sciences

Overview Hemophagocytic Lymphohistiocytosis ?? Genetics of HLH / FHL Genetic testing Role of new sequencing technologies in HLH diagnosis and management

Hemophagocytic Lymphohistiocytosis Hemophagocytic Lymphohistiocytosis (HLH): It is a clinical syndrome of overwhelming immune activation and cytokine production It is caused by failure to clear antigen presenting cells and/or activated T cells

HLH Subtypes Genetic / Familial Acquired Includes Familial Hemophagocytic Lymphohistiocytosis (FHL) as well as certain immunodeficiencies Caused by defects in the cell-mediated cytotoxicity pathways Acquired Various groups of disorders that result in hemophagocytic symptoms Caused by dysregulated immune responses leading to lymphocyte and macrophage activation

Genetic HLH Familial Hemophagocytic Lymphohistiocytosiscan (FHL) can be divided into 5 subtypes: FHL1 – caused by unknown defect on chromosome 9 FHL2 – caused by deficiency of Perforin FHL3 – caused by deficiency of Munc 13-4 FHL4 – caused by deficiency of Syntaxin 11 FHL5 – caused by deficiency of Munc 18-2 Chediak-Higashi & Griscelli II syndromes: characterized by partial albinism and immune deficiency XLP: characterized by massive lymphoproliferation and immune deficiency

FHL Sometimes referred to as Farquhar’s disease after its describer (1952) Autosomal recessive inheritance with estimated incidence of 1:50,000 live births (male > female) Symptoms are usually evident by 1 year (70-80% of case) and can even present at birth or in utero Some forms can present in later childhood or even as adults Overwhelming HLH is the primary symptom, and deficient NK cell-mediated cytotoxicity is characteristic

FHL Diagnosis At least five of the eight following criteria based on the guidelines of the Histiocyte Society [Henter et al 2007]. Biallelic pathogenic variants in any one of PRF1, UNC13D, STX11, or STXBP2 genes

Genetics of FHL Autosomal Recessive Inheritance Biallelic Pedigree patterns Consanguinity Risk assessment

Genetics of FHL Five loci (but 4 genes!!) have been identified: FHL1: locus on 9q21.3-22, no gene identified, homozygosity mapping in 4 consanguineous families of Pakistani descent FHL2: PRF1 gene, 10q22.1 FHL3: UNC13D gene, 17q25.1 FHL4: STX11 gene, Syntaxin, 6q24.2 FHL5: STXBP2 gene, Syntaxin binding protein 2, 19p13.2,

Genetics of FHL Genetic epidemiology: FHL1: 4 families FHL2: 20-30% of all FHL cases (Europe, Japan), >50% of cases with African American descent FHL3: 20-30% of all FHL cases (Europe, Japan) FHL4: 20% in Turks/Kurds, 1% in North American, 5% in Central Europeans FHL5: 20% in North Americans

Genetics of FHL Variants features - testing method: FHL2: PRF1 – coding regions - Sequence analysis FHL3: UNC13D – coding regions, splice sites, deep intronic, large inversion (253 kb) – Sequence analysis, targeted targeted scanning FHL4: STAX11 – 3 variants in Turkish/Kurdish, other variants, deletion/duplication – sequence analysis, targeted scanning FHL5: STAXBP2 – coding regions, splice sites – sequence analysis

Genetics of FHL Important points to consider: More locus heterogeneity: 30% of FHL with no identified mutation in any of 4 genes Negative genetic analysis can not exclude diagnosis Finding one mutation / missing second mutation A very complex genetic workup, sometimes inevitable

Genetics of FHL Genetic workup of a proband Factors to be considered: Clinical presentation, race/ethnicity, immunologic test results General recommended approach: NGS panel of four genes Targeted scanning of complex rare variants Analysis of RAB27A gene for Griscceli syndrome type 2 Male patients: analysis of XLAP and SH2D1A Other differential diagnosis: Chediak-Higashi syndrome

Genetics of FHL Genetic workup of the family / pedigree: Long before any replroductive planning !!! Pre-marriage / preconception counseling Carrier screening of any at-risk member of the pedigree Prenatal diagnosis / preimplantation genetic diagnosis for at-risk pregnancies

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