Circulating B-Cell Precursor Cells As Potential Diagnostic and Therapeutic Biomarker in Patients with W­H­I­M Syndrome

Background

Warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome is a combined immunodeficiency caused in most known cases by gain-of-function mutations in the CXCR4 gene. The wide spectrum of multisystem clinical presentations, often with incomplete penetrance and expressivity, can pose a diagnostic challenge. WHIM syndrome has historically been a clinicopathologic diagnosis, with myelokathexis being the pathognomonic feature of WHIM. As genetic testing is becoming readily available, sequencing of the CXCR4 gene is often used to confirm a clinical suspicion of WHIM syndrome by finding genetic variants. There is a need for additional easily accessible and noninvasive biomarkers for diagnosing WHIM syndrome. We hypothesize that B-cell compartment might also show pathognomonic features of WHIM syndrome.

Patients and Methods

We analyzed peripheral blood B-cell subsets in 13 patients diagnosed with WHIM syndrome from 8 families with pathogenic CXCR4 mutations using 9-multicolor flow cytometry after informed consent approval. Samples from 2 patients who received plerixafor were collected at baseline before and after treatment. The duration of treatment was approximately 6 months.

Results

Using multicolor flow cytometry, we analyzed circulating B-cell subsets (CD10,CD19,CD21,CD27,CD38,IgA,IgD,IgG and IgM) in 13 patients (pediatric, n=7; adult, n=6) with WHIM syndrome (9 patients with p.Arg334Stop mutation, 2 patients with p.Val320Glufs*23 mutation, 1 patient with p.Ser338Stop mutation, and 1 patient with p.Ser346Stop mutation). We observed severe peripheral B-cell lymphopenia, accompanied by the presence of a consistent and specific scarce CD19⁺CD10⁺CD38⁺IgM^–IgD^–CD21^–CD27^– B-cell subpopulation. A B-cell subpopulation with an equivalent marker expression pattern was not found in healthy controls (n=50) or in non-WHIM disease controls (n=20). The rare B-cell population found in WHIM syndrome was further characterized as belonging mostly to the pre–B-cell stage revealed by intracellular expression of IgM, CD179a (VpreB chain), and CD179b (lambda 5). The presence of these early B-cell precursors was independent of age and was stable over time in patients with WHIM syndrome. The number of these peripheral B-cell precursors found specifically in patients with WHIM syndrome was greatly reduced when the patients were treated with granulocyte colony-stimulating factor (G-CSF) or plerixafor; counts of the precursor cells returned to baseline after the patients discontinued plerixafor treatment.

Conclusions

Taken together, we identify a unique CD19⁺CD10⁺CD38⁺IgM^–IgD^–CD21^–CD27^– B-cell subpopulation in patients with WHIM syndrome that might be used as a noninvasive biomarker to help identify patients with this rare primary combined immunodeficiency. In addition, these circulating B-cell precursors may be used to monitor response to therapy in patients with WHIM syndrome. The observed impact of G-CSF on the B-cell precursor population may be attributed to downregulation of CXCL12/CXCR4, which influences myelokathexis and impairs egression of the precursor population from the bone marrow but needs further experimental validation.