Development of flow cytometry assay for Wiskott–Aldrich syndrome diagnosis by WASP protein evaluation

Abstract

Wiskott–Aldrich syndrome (WAS) is a rare X-linked primary immunodeficiency characterized by microplatelet thrombocytopenia, eczema, frequent infections and an increased risk of autoimmune disorders and malignant neoplasms. Mutation detection in WAS gene is the gold standard for diagnosis of this disorder. This gene encodes a WASP protein, which works as regulator of cell cytoskeleton and is involved in the transmission of many intracellular signals. Nowadays there is no rapid and reliable method that allows to confirm WAS in a short period of time. Early detection of WAS in patients enables initiation of a donor search and preparation for the HSCT procedure. It also helps to avoid the development of severe and life-threatening conditions during waiting for genetic confirmation of the diagnosis by using pathogenetic therapy. Currently flow cytometry is one of the leading laboratory methods that permits to get the information about the expression of a protein in several hours. The study below describes rapid and reliable based on flow cytometry assay for WAS diagnosis. The study was approved by the Independent Ethics Committee and the Scientific Council of the Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology, and Immunology. The study included 46 patients with suspected WAS from 2 months to 17 years old. Patients were examined from January 2018 to January 2020. WAS gene defect was confirmed in 35 patients. It was calculated that normal threshold value for WASP expression is 7.07 with sensitivity and specificity 100% and 93.1% respectively. Besides negative correlation between WASP expression index and WAS clinical severity was shown (r = –0.63). This flow cytometry assay can be used for chimerism detection in WAS patients after HSCT. The flow cytometry assay for WASP protein evaluation is rapid, highly sensitive and highly specific. It allows to speed up diagnosis of this disorder.