Development of Screening for Primary Immunodeficiency

Primary immunodeficiencies are inherited disorders of the immune system, with an estimated prevalence of 1:500 in the USA. Yet, a majority of the patients are still undiagnosed. Most patients with a diagnosis come in contact with the healthcare due to complications caused by the immunological defects. In many cases, it is crucial that the patients receive treatment before their health is seriously compromised. There is therefore a great need for more efficient ways of identifying the patients. As a lack of serum IgA is included in the phenotype of many of the defects, a population-based screening for IgA deficiency would identify patients with various forms of primary immunodeficiencies. The main benefit of such screening would be the early detection of the disorders followed by appropriate treatment, to decrease the risk of severe infections. We investigated the possibility of a screening for primary immunodeficiencies based on the use of serum microarrays, a high-throughput platform suitable for large-scale analysis of dried blood spot samples collected from newborns. Serum samples from adult individuals with known levels of serum IgA were used to investigate the potential of serum microarrays in detection of serum IgA. A high level of correlation was observed throughout the material, compared to a conventional method (paper I). The serum microarrays were then used to establish the prevalence of IgA deficiency in serum samples of Swedish children at the age of four and the IgA was correlated to various health outcomes that are associated to allergy and infections. The prevalence of IgA deficiency was found to be 1:173, as compared to 1:600 in the adult population. IgA deficient children seem to have an increased risk of pseudocroup and parentally reported food hypersensitivity during childhood (paper II). As the screening should be based on the use of dried blood spots samples, collected within days after birth, the suitability of serum microarrays in analysis of these samples was evaluated in paper III. The levels of complement protein C3 were measured in samples from a C3 deficient patients and controls. The levels of C3 in the control group was significantly higher than in the Swedish C3 deficient patient sample, indicating that serum microarrays are suitable for identification of C3 deficiency using the dried blood spot samples. The next step in our evaluation was to increase the sensitivity of the serum microarrays/DBSS approach (paper IV). The C2 protein was used, which is less abundant in newborns than C3. None of the tested anti-C2 antibodies was suitable for discrimination between patients and controls using serum microarrays. However, when using ELISA, the levels of C2 in DBSS eluates of C2 deficient patients were significantly lower than in control samples. Thus, the C2 deficiency is present already at birth. In paper V, serum IgA levels were determined in dried blood spot samples from patients with various forms of primary immunodeficiencies. Only patients born to mothers with selective IgA deficiency lacked IgA at birth. In the remaining patients, the IgA levels were comparable to those of the controls. The results suggest that the serum IgA detected in newborns is to some extent of maternal origin. Thus, a neonatal screening for primary immunodeficiencies based on the determination of IgA levels is not possible due o the presence of maternal IgA in the samples. LIST OF PUBLICATIONS I. Serum microarrays for large scale screening of protein levels. Janzi M., Odling J., Pan-Hammarström Q., Sundberg M., Lundeberg J., Uhlén M., Hammarström L., Nilsson P. Mol Cell Proteomics. 2005, 4(12):1942-7. II. Selective IgA deficiency in early life: Association to infections and allergic diseases during childhood. Janzi M., Kull I., Sjöberg R., Wan J., Melén E., Bayat N., Östblom E., PanHammarström Q., Nilsson P., Hammarström L. Clin Immunol. 2009, Jun 19. [Epub ahead of print] III. Screening for C3 deficiency in newborns using microarrays. Janzi M., Sjöberg R., Wan J., Fischler B., von Döbeln U., Isaac L., Nilsson P., Hammarström L. PLoS One. 2009, 4(4):e5321. IV. Screening for C2 deficiency in newborns. Janzi M., von Döbeln U., Jönsson G., Hammarström L. Manuscript. V. Evaluation of lack of IgA as a diagnostic marker for primary immunodeficiency disorders using Guthrie card eluates. Janzi M., Pan-Hammarström Q., von Döbeln U., Nordvall L., Fasth A., Hammarström L. Manuscript.