Clinical Information

The differential diagnosis for patients with primary hypogammaglobulinemia of unclear etiology (after other secondary causes of hypogammaglobulinemia have been ruled out) includes common variable immunodeficiency (CVID) and X-linked agammaglobulinemia (XLA). CVID is the most common diagnosis of humoral immunodeficiency, particularly in adults, but also in children over 4 years of age. However, adult male patients with XLA may be misdiagnosed with CVID. XLA is an independent humoral immunodeficiency and should not be regarded as a subset of CVID.

The BTK gene is present on the long arm of the X-chromosome and encodes for a cytoplasmic tyrosine kinase with 5 distinct structural domains. While BTK gene sequencing is the gold standard for definitively identifying mutations and confirming a diagnosis of XLA, it is labor intensive and expensive. Flow cytometry is a screening test for XLA and should be included in the evaluation of patients with possible CVID, particularly in male patients with <1% B cells. Bruton tyrosine kinase (Btk) is an intracellular protein and absence of the Btk protein by flow cytometry provides a strong rationale for performing a BTK gene-sequencing test. However, 20% to 30% of XLA patients may have intact or truncated Btk protein with abnormal function; therefore, genetic analysis remains the more definitive test for diagnosing XLA (besides other clinical and immunological parameters).

XLA is a prototypical humoral immunodeficiency caused by mutations in the BTK gene, which encodes Btk, a hematopoietic-specific tyrosine kinase. XLA is characterized by normal, reduced, or absent Btk expression in monocytes and platelets, a significant reduction or absence of circulating B cells in blood, and profound hypogammaglobulinemia of all isotypes (IgG, IgA, IgM, and IgE). The clinical presentation includes early onset of recurrent bacterial infections, and absent lymph nodes and tonsils. Btk plays a critical role in B-cell differentiation. The defect in Btk may be "leaky" in some patients (ie, a consequence of mutations in the gene that result in a milder clinical and laboratory phenotype), such that these patients may have some levels of IgG and/or IgM and a small number of B cells in blood.(1)

The vast majority of XLA patients are diagnosed in childhood (median age of diagnosis in patients with sporadic XLA is 26 months), although some patients are recognized in early adulthood or later in life. The diagnosis of XLA in both children and adults indicates that the disorder demonstrates considerable clinical phenotypic heterogeneity, depending on the position of the mutations within the gene. Females are typically carriers and asymptomatic. Testing in adult females should be limited to those in their child-bearing years (<45 years). Carrier testing ideally should be confirmed by genetic testing since it is possible to have a normal flow cytometry test for protein expression in the presence of heterozygous (carrier) BTK gene mutations.

Flow cytometry is a preliminary screening test for XLA. It is important to keep in mind that this flow cytometry test is only a screening tool and approximately 20% to 30% of patients who have a mutation within the BTK gene have normal protein expression (again related to the position of the mutation in the gene and the antibody used for flow cytometric analysis). Therefore, in addition to clinical correlation, genetic testing is recommended to confirm a diagnosis of XLA. Furthermore, it is helpful to correlate gene and protein data with clinical history (genotype-phenotype correlation) in making a final diagnosis of XLA. Consequently, the preferred test for XLA is BTKFP / Bruton's Tyrosine Kinase (BTK) Genotype and Protein Analysis, Full Gene Sequence and Flow Cytometry, which includes both flow cytometry and gene sequencing to confirm the presence of a BTK mutation. If a familial mutation has already been identified, then BTKMP / Bruton's Tyrosine Kinase (BTK) Genotype and Protein Analysis, Known Mutation Sequencing and Flow Cytometry should be ordered.