Rapid Detection of SAP Deficiency in Cytotoxic Lymphocytes from Patients with X-Linked Lymphoproliferative Disease and Their Family Members.

Background: X-linked lymphoproliferative disease (XLP) is a primary immunodeficiency, which is characterized by an extreme susceptibility to Epstein-Barr virus (EBV). In half of the XLP patients, primary EBV infection can be fatal with explosive activation and proliferation of lymphocytes in many organs, which leads to fluminant hepatitis and bone marrow failure with hemophagocytosis. Mutations in the SH2D1A gene, which encodes the SAP protein, have been described in a proportion of patients with the clinical syndrome of XLP. The diagnosis of XLP is still difficult given its clinical heterogeneity, and the lack of a readily available rapid diagnostic laboratory test, particularly in cases without a family history of XLP. XLP should always be a consideration in males with EBV-associated hemophagocytic lymphohistiocytosis (EBV-HLH).

Methods: Four-color flow cytometric analysis was used to establish normal patterns of SAP expression for control subjects, and patterns of SAP staining in cytotoxic lymphocytes (CD8+ T cells, CD56+ T cells and natural killer [NK] cells). This assay was used to study patients with clinical syndromes consistent with XLP, and in some cases; their family members.

Results: 6 patients with XLP confirmed by detection of mutations in the coding regions of SH2D1A showed lack of intracellular SAP in all cytotoxic cell types; one was tested presymptomatically. Carriers of SH2D1A mutations tested (mother and sister) had decreased SAP staining patterns below normal levels. Eleven males with clinical syndromes consistent with XLP, predominantly EBV-HLH who were demonstrated to have normal or increased perforin expression were also studied. All patients showed normal SAP expression including one case with a convincing X-linked family history; all were subsequently shown to have no mutations in SH2D1A. Additionally, one asymptomatic adult male with an intronic mutation previously published as causing XLP was found to have normal SAP expression.

Conclusions: Four-color flow cytometry provides diagnostic information that may speed the identification of this fatal disease, differentiating it from other EBV-HLH. We also suspect it will prove useful in substantiating or excluding disease-causing genetic variants in the SH2D1A gene.