Severe Combined Immunodeficiency (SCID), Wiskott-Aldrich Syndrome (WAS), and X-Linked Agammaglobulinemia (XLA) are three primary immune deficiency disorders associated with early lethality if not diagnosed and treated appropriately. Early recognition, diagnosis, and preemptive treatment are critical for improving outcomes in these disorders however;there is a fundamental gad in our ability to perform population-based screening for more than just SCID. The long-term goal is to develop a viable multiplex testing methodology that will allow simultaneous identification of multiple congenital disorders characterized by absence of specific proteins using small sample volumes of blood dried onto filter paper. The objective of this particular application is to identify and characterize unique """"""""fingerprints"""""""" peptides that can be used to rapidly identify patients with three severe immunodeficiency syndromes that will be used as model disorders: Severe Combined Immunodeficiency (SCID), Wiskott-Aldrich Syndrome (WAS), and X-Linked Agammaglobulinemia (XLA). Dried blood spots will be used as the substrate and the testing approach will be tandem mass spectrometry. The central hypothesis of this application is that tandem mass spectrometry can used to detect the presence or absence of key """"""""fingerprint"""""""" peptides derived from CD3, WASP and BTK (markers for SCID, WAS, and XLA respectively). This hypothesis has been formulated based on our preliminary data (deWilde, 2008). The rationale for the proposed research is that rapid, early detection of severe primary immunodeficiency disorder such as SCID, WAS, and XLA will allow for preemptive treatment that will prevent death and morbidity thereby improving quality of life and long-term productivity. The central hypothesis will be tested by pursuing three specific aims: 1.) Identify key """"""""fingerprint"""""""" peptides for CD3, WASP, and BTK;marke proteins for SCID, WAS. And XLA respectively. 2.) Define the suitability of each predicted """"""""fingerprint"""""""" peptide for use in tandem mass spectrometry (MS/MS) and optimize its detection. 3.) Validate the ability of each """"""""fingerprint"""""""" peptide to identify the absence of CD3, WASP, and BTK in dried blood spots from patients with SDIC, WAS, and XLA as well as controls. The approach is innovative because of the new methods we have developed, expanding the use of an existing technology to diagnose congenital disorders that result from absence of a specific protein. These tools have the potential to increase our capacity for early and rapid identification of several severe primary immunodeficiency disorders. The proposed research is significant, because it will provide a testing methodology that is viable both for newborn screening and for rapid diagnostics for these and other severe immunodeficiency and congenital disorders. This will provide opportunities to identify patients early in life and provide life-saving therapies. In addition, this is expected to fundamentally advance the use of proteomic methodologies for diagnostic testing, an area that holds great promise but is largely untapped.

Public Health Relevance

The proposed studies focus on early and rapid diagnosis of the severe, life-threatening immunodeficiency syndromes Severe Combined Immunodeficiency (SCID), Wiskott-Aldrich Syndrome (WAS), and X-Linked Agammaglobulinemia (XLA). This application is designed to utilize an innovative tandem mass spectroscopy approach to quickly diagnose these disorders using blood spots dried onto filter paper, because of the high-throughput, multiplex potential of this method, it will dramatically expand our ability to perform newborn screening and rapid diagnostic testing for these and other disorders using very small blood volumes. This will directly impact patient care by providing early identification and the opportunity for preemptive treatment for life threatening immunodeficiency disorders.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (ZRG1-IMM-C (52))
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Johnson, David R
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Seattle Children's Hospital
United States
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Kerfoot, Sandra A; Jung, Sunhee; Golob, Karin et al. (2012) Tryptic peptide screening for primary immunodeficiency disease by LC/MS-MS. Proteomics Clin Appl 6:394-402