Allergy Asthma Immunol Res.  2020 Mar;12(2):292-305. 10.4168/aair.2020.12.2.292.

Flow Cytometry for the Diagnosis of Primary Immunodeficiency Diseases: A Single Center Experience

Affiliations
  • 1Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea. eskang@skku.edu
  • 2Department of Laboratory Medicine, Soonchunhyang University Hospital, Cheonan, Korea.
  • 3Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
  • 4Department of Pediatrics, Severance Children's Hospital, Yonsei University College of Medicine, Seoul, Korea.
  • 5Department of Pediatrics, Gachon University Gil Medical Center, Incheon, Korea.
  • 6Department of Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
  • 7Green Cross Genome, Yongin, Korea.

Abstract

PURPOSE
While there is an urgent need for diagnosis and therapeutic intervention in patients with primary immunodeficiency diseases (PIDs), current genetic tests have drawbacks. We retrospectively reviewed the usefulness of flow cytometry (FCM) as a quick tool for immunophenotyping and functional assays in patients suspected to have PIDs at a single tertiary care institute.
METHODS
Between January 2001 and June 2018, patients suspected of having PIDs were subjected to FCM tests, including lymphocyte subset analysis, detection of surface- or intracellular-target proteins, and functional analysis of immune cells, at Samsung Medical Center, Seoul, Korea. The genetic diagnosis was performed using Sanger or diagnostic exome sequencing.
RESULTS
Of 60 patients diagnosed with definite or probable PID according to the European Society of Immune Deficiencies criteria, 24 patients were provided with useful information about immunological dysfunction after initial FCM testing. In 10 patients, the PID diagnosis was based on abnormal findings in FCM testing without genetic tests. The FCM findings provided strong evidence for the diagnosis of severe combined immunodeficiency (n = 6), X-linked chronic granulomatous diseases (CGD) (n = 6), leukocyte adhesion deficiency type 1 (n = 3), X-linked agammaglobulinemia (n = 11), autoimmune lymphoproliferative syndrome-FASLG (n = 1), and familial hemophagocytic lymphohistiocytosis type 2 (n = 1), and probable evidence for autosomal recessive-CGD (n = 2), autosomal dominant-hyper-immunoglobulin E (IgE)-syndrome (n = 1), and STAT1 gain-of-function mutation (n = 1). In PIDs derived from PIK3CD (n = 2), LRBA (n = 2), and CTLA4 mutations (n = 3), the FCM test provided useful evidence of immune abnormalities and a tool for treatment monitoring.
CONCLUSIONS
The initial application of FCM, particularly with known protein targets on immune cells, would facilitate the timely diagnosis of PIDs and thus would support clinical decisions and improve the clinical outcome.

Keyword

Primary immunodeficiency; diagnosis; phenotype; flow cytometry; genetic testing

MeSH Terms

Agammaglobulinemia
Diagnosis*
Exome
Flow Cytometry*
Genetic Testing
Granulomatous Disease, Chronic
Humans
Immunophenotyping
Korea
Leukocytes
Lymphocyte Subsets
Lymphohistiocytosis, Hemophagocytic
Phenotype
Retrospective Studies
Seoul
Severe Combined Immunodeficiency
Tertiary Healthcare

Figure

  • Fig. 1 Enrollment and classification of patients. PID, primary immunodeficiency disease; HLH, hemophagocytic lymphohistiocytosis; ESID, European Society of Immune Deficiencies; IUIS, International Union of Immunological Societies; WAS, Wiskott-Aldrich syndrome; AD-HIES, autosomal dominant-hyper-IgE syndrome; HIES, hyper-IgE syndrome; BTK, Bruton's tyrosine kinase; CVID, common variable immune deficiency; IgA, immunoglobulin A; CTLA4, cytotoxic T-lymphocyte associated protein 4; ALPS, autoimmune lymphoproliferative syndrome; AR-CGD, autosomal recessive chronic granulomatous diseases; FHL, familial hemophagocytic lymphohistiocytosis; LAD, leukocyte adhesion deficiency; SCID, severe combined immunodeficiency; IPEX-like SD, immunodysregulation polyendocrinopathy enteropathy X-linked syndrome; XL-CGD, X-linked-chronic granulomatous diseases; STAT1, signal transducer and activator of transcription 1; GOF, gain-of-function.

  • Fig. 2 Categorization of PIDs based on the usefulness of Flow cytometry for diagnosis. PID, primary immunodeficiency disease; FCM, flow cytometry; AD-HIES, autosomal dominant-hyper-IgE syndrome; ALPS, autoimmune lymphoproliferative syndrome; AR-CGD, autosomal recessive-chronic granulomatous diseases; CTLA4, cytotoxic T-lymphocyte associated protein 4; CVID, common variable immune deficiency; FHL, familial hemophagocytic lymphohistiocytosis; IPEX-like SD, Immunodysregulation polyendocrinopathy enteropathy X-linked syndrome; LAD, leukocyte adhesion deficiency; SCID, Severe combined immunodeficiency; WAS, Wiskott-Aldrich syndrome; XIAP, X-linked inhibitor of apoptosis protein; XLA, X-linked agammaglobulinemia; XL-CGD, X-linked-chronic granulomatous diseases. *FCM was not performed for these diseases in this study; †We suggest that WAS could be divided into 2 categories based on the variants that influence protein expression.


Reference

1. Picard C, Al-Herz W, Bousfiha A, Casanova JL, Chatila T, Conley ME, et al. Primary immunodeficiency diseases: an update on the classification from the international union of immunological societies expert committee for primary immunodeficiency 2015. J Clin Immunol. 2015; 35:696–726.
2. Ishimura M, Takada H, Doi T, Imai K, Sasahara Y, Kanegane H, et al. Nationwide survey of patients with primary immunodeficiency diseases in Japan. J Clin Immunol. 2011; 31:968–976.
Article
3. Rhim JW, Kim KH, Kim DS, Kim BS, Kim JS, Kim CH, et al. Prevalence of primary immunodeficiency in Korea. J Korean Med Sci. 2012; 27:788–793.
Article
4. Kobrynski L, Powell RW, Bowen S. Prevalence and morbidity of primary immunodeficiency diseases, United States 2001–2007. J Clin Immunol. 2014; 34:954–961.
Article
5. Grimbacher B. ESID Registry Working Party. The European Society for Immunodeficiencies (ESID) registry 2014. Clin Exp Immunol. 2014; 178:Suppl 1. 18–20.
Article
6. European Society for Immunodeficiencies. Registry working party diagnosis criteria. ESID registry - working definitions for clinical diagnosis of PID [Internet]. Amsterdam: European Society for Immunodeficiencies;2019. cited 2019 Jan 17. Available from: https://esid.org/Working-Parties/Registry-Working-Party/Diagnosis-criteria.
7. Rae W, Ward D, Mattocks C, Pengelly RJ, Eren E, Patel SV, et al. Clinical efficacy of a next-generation sequencing gene panel for primary immunodeficiency diagnostics. Clin Genet. 2018; 93:647–655.
Article
8. Bisgin A, Boga I, Yilmaz M, Bingol G, Altintas D. The utility of next-generation sequencing for primary immunodeficiency disorders: experience from a clinical diagnostic laboratory. BioMed Res Int. 2018; 2018:9647253.
Article
9. Stray-Pedersen A, Sorte HS, Samarakoon P, Gambin T, Chinn IK, Coban Akdemir ZH, et al. Primary immunodeficiency diseases: genomic approaches delineate heterogeneous Mendelian disorders. J Allergy Clin Immunol. 2017; 139:232–245.
10. Maffucci P, Filion CA, Boisson B, Itan Y, Shang L, Casanova JL, et al. Genetic diagnosis using whole exome sequencing in common variable immunodeficiency. Front Immunol. 2016; 7:220.
Article
11. Fleisher TA, Madkaikar M, Rosenzweig SD. Application of flow cytometry in the evaluation of primary immunodeficiencies. Indian J Pediatr. 2016; 83:444–449.
Article
12. Song SM, Park MR, Kim DS, Kim J, Kim YJ, Ki CS, et al. Identification of a novel mutation in the CYBB gene, p.Asp378Gly, in a patient with X-linked chronic granulomatous disease. Allergy Asthma Immunol Res. 2014; 6:366–369.
Article
13. Huh HJ, Jhun BW, Choi SR, Kim YJ, Yun SA, Nham E, et al. Bronchiectasis and recurrent respiratory infections with a de novo STAT1 gain-of-function variant: first case in Korea. Yonsei Med J. 2018; 59:1004–1007.
14. Morbach H, Eichhorn EM, Liese JG, Girschick HJ. Reference values for B cell subpopulations from infancy to adulthood. Clin Exp Immunol. 2010; 162:271–279.
Article
15. Kanegane H, Hoshino A, Okano T, Yasumi T, Wada T, Takada H, et al. Flow cytometry-based diagnosis of primary immunodeficiency diseases. Allergol Int. 2018; 67:43–54.
Article
16. Fischer A, Le Deist F, Hacein-Bey-Abina S, André-Schmutz I, Basile Gde S, de Villartay JP, et al. Severe combined immunodeficiency. A model disease for molecular immunology and therapy. Immunol Rev. 2005; 203:98–109.
Article
17. Luk AD, Lee PP, Mao H, Chan KW, Chen XY, Chen TX, et al. Family history of early infant death correlates with earlier age at diagnosis but not shorter time to diagnosis for severe combined immunodeficiency. Front Immunol. 2017; 8:808.
Article
18. Gathmann B, Grimbacher B, Beauté J, Dudoit Y, Mahlaoui N, Fischer A, et al. The European internet-based patient and research database for primary immunodeficiencies: results 2006–2008. Clin Exp Immunol. 2009; 157:Suppl 1. 3–11.
Article
19. Fischer A. Severe combined immunodeficiencies (SCID). Clin Exp Immunol. 2000; 122:143–149.
Article
20. Yu GP, Nadeau KC, Berk DR, de Saint Basile G, Lambert N, Knapnougel P, et al. Genotype, phenotype, and outcomes of nine patients with T-B+NK+ SCID. Pediatr Transplant. 2011; 15:733–741.
Article
21. Yi ES, Choi YB, Lee NH, Lee JW, Sung KW, Koo HH, et al. Allogeneic hematopoietic cell transplantation in patients with primary immunodeficiencies in Korea: eleven-year experience in a single center. J Clin Immunol. 2018; 38:757–766.
Article
22. Abraham RS, Aubert G. Flow cytometry, a versatile tool for diagnosis and monitoring of primary immunodeficiencies. Clin Vaccine Immunol. 2016; 23:254–271.
Article
23. Marcondes NA, Fernandes FB, Spindler BM, Faulhaber GA. Flow cytometry assessment of intracellular BTK expression. Cytometry B Clin Cytom. 2018; 94:568.
Article
24. Kim HY, Kim HJ, Ki CS, Kim DW, Yoo KH, Kang ES. Rapid determination of chimerism status using dihydrorhodamine assay in a patient with X-linked chronic granulomatous disease following hematopoietic stem cell transplantation. Ann Lab Med. 2013; 33:288–292.
Article
25. Yu JE, Azar AE, Chong HJ, Jongco AM 3rd, Prince BT. Considerations in the diagnosis of chronic granulomatous disease. J Pediatric Infect Dis Soc. 2018; 7:S6–11.
Article
26. Puel A, Cypowyj S, Maródi L, Abel L, Picard C, Casanova JL. Inborn errors of human IL-17 immunity underlie chronic mucocutaneous candidiasis. Curr Opin Allergy Clin Immunol. 2012; 12:616–622.
Article
27. Li P, Huang P, Yang Y, Hao M, Peng H, Li F. Updated understanding of autoimmune lymphoproliferative syndrome (ALPS). Clin Rev Allergy Immunol. 2016; 50:55–63.
Article
28. Mizoguchi Y, Tsumura M, Okada S, Hirata O, Minegishi S, Imai K, et al. Simple diagnosis of STAT1 gain-of-function alleles in patients with chronic mucocutaneous candidiasis. J Leukoc Biol. 2014; 95:667–676.
Article
29. Avery DT, Kane A, Nguyen T, Lau A, Nguyen A, Lenthall H, et al. Germline-activating mutations in PIK3CD compromise B cell development and function. J Exp Med. 2018; 215:2073–2095.
30. Oliveira JB, Bleesing JJ, Dianzani U, Fleisher TA, Jaffe ES, Lenardo MJ, et al. Revised diagnostic criteria and classification for the autoimmune lymphoproliferative syndrome (ALPS): report from the 2009 NIH International Workshop. Blood. 2010; 116:e35–40.
Article
31. Bernardo I, Mancebo E, Aguiló I, Anel A, Allende LM, Guerra-Vales JM, et al. Phenotypic and functional evaluation of CD3+CD4-CD8- T cells in human CD8 immunodeficiency. Haematologica. 2011; 96:1195–1203.
Article
32. Ogonek J, Kralj Juric M, Ghimire S, Varanasi PR, Holler E, Greinix H, et al. Immune reconstitution after allogeneic hematopoietic stem cell transplantation. Front Immunol. 2016; 7:507.
Article
33. Rao VK. Approaches to managing autoimmune cytopenias in novel immunological disorders with genetic underpinnings like autoimmune lymphoproliferative syndrome. Front Pediatr. 2015; 3:65.
Article
34. Lo B, Zhang K, Lu W, Zheng L, Zhang Q, Kanellopoulou C, et al. Autoimmune disease. Patients with LRBA deficiency show CTLA4 loss and immune dysregulation responsive to abatacept therapy. Science. 2015; 349:436–440.
35. Lee S, Moon JS, Lee CR, Kim HE, Baek SM, Hwang S, et al. Abatacept alleviates severe autoimmune symptoms in a patient carrying a de novo variant in CTLA-4. J Allergy Clin Immunol. 2016; 137:327–330.
36. Bousfiha AA, Jeddane L, Ailal F, Benhsaien I, Mahlaoui N, Casanova JL, et al. Primary immunodeficiency diseases worldwide: more common than generally thought. J Clin Immunol. 2013; 33:1–7.
Article
37. Kirkpatrick P, Riminton S. Primary immunodeficiency diseases in Australia and New Zealand. J Clin Immunol. 2007; 27:517–524.
Article
38. Lim DL, Thong BY, Ho SY, Shek LP, Lou J, Leong KP, et al. Primary immunodeficiency diseases in Singapore--the last 11 years. Singapore Med J. 2003; 44:579–586.
Full Text Links
  • AAIR
Actions
Cited
CITED
export Copy
Close
Share
  • Twitter
  • Facebook
Similar articles
Copyright © 2024 by Korean Association of Medical Journal Editors. All rights reserved.     E-mail: koreamed@kamje.or.kr