Evaluation of B‐cell intracellular signaling by monitoring the PI3K‐Akt axis in patients with common variable immunodeficiency and activated phosphoinositide 3‐kinase delta syndrome
Lucía del Pino Molina, Juan Manuel Torres, Rebeca Rodríguez‐Pena, Eduardo López‐Granados
Abstract
BACKGROUND: Primary antibody deficiencies (PADs) are characterized by hypogammaglobulinemia and impaired B-cell differentiation. Patients with common variable immunodeficiency (CVID) present severe reductions in at least 2 serum immunoglobulins and impaired terminal differentiation of B cells. Most patients with CVID do not appear to present monogenic defects. Activated phosphoinositide 3-kinase delta syndrome (APDS), caused by gain-of-function mutations in the PIK3CD gene (p110δ), can present in patients with a CVID-like phenotype. Memory B-cell differentiation requires the orchestrated activation of numerous intracellular signaling pathways, which promote transcriptional programs required for long-term B-cell survival. The aim of this study was to develop a flow cytometry assay to trace the PI3K-Akt-mTOR pathway, a critical component of B-cell homeostasis, and analyze its status in PADs. METHODS: We analyzed the intracellular expression of Akt and S6 by flow cytometry and their phosphorylation status in both baseline conditions and upon B-cell receptor activation with anti-IgM in various primary B-cell subsets of patients with CVID and APDS. RESULTS: B cells from CVID patients showed reduced phosphorylation in Akt and S6 proteins after anti-IgM stimulation. Constitutive high baseline B-cell levels of Akt and S6 phosphorylation in a patient with APDS were reduced once m-TOR inhibition therapy was initiated. CONCLUSIONS: Intracellular flow cytometry can be routinely employed to explore alterations in the PI3K-Akt-mTOR pathway in B cells from patients with PADs. AKT and S6 phosphorylation levels are informative biomarkers that could be employed as mTOR inhibitors for monitoring therapies targeting this pathway.