Administration of rhIL-7 Is Associated with Increase in Precursor T-Cells in Bone Marrows of Patients with Idiopathic CD4 Lymphocytopenia

Introduction

Idiopathic CD4+ Lymphocytopenia (ICL) is a rare, likely heterogeneous syndrome characterized by persistent CD4+ lymphopenia (<300/μl) in the absence of HIV infection or other known immunodeficiency. The diverse clinical presentation may include opportunistic infections, malignancies, and autoimmune diseases. Though the etiology of ICL remains unknown, previous studies have suggested decreased production, proliferation, or survival of CD4+ T-cells. There is currently no FDA-approved therapy for ICL.

Methods

We analyzed bone marrow core biopsies of 12 ICL patients in a phase I/IIA NIH clinical trial designed to evaluate recombinant human Interleukin-7 (rhIL-7) as a potential therapy for ICL . Subjects received 3 injections of rhIL-7 over two 24-week periods. Bone marrow biopsies were performed at week 1 and week 24 to analyze numbers of T-cells and T-cell precursors at baseline and post rhIL-7 therapy. Peripheral blood T-cell counts were assessed in parallel. Double chromogenic immunohistochemical staining using anti-terminal deoxynucleotidyl transferase (TdT) and anti-CD3 antibodies was performed on fixed, paraffin-embedded samples using an automated stainer (Ventana). Single CD3-positive T-cells and double-positive TdT/CD3 precursor T-cells were counted in at least ten consecutive fields under a light microscope and results expressed as mean positive cell number per field before and after treatment with rhIL-7. Similar staining procedure using TdT and CD79a was performed to evaluate precursor B-cells in the same patient cohort. In addition, bone marrow biopsies from 10 healthy control subjects were analyzed in parallel.

Results

Compared to healthy controls, ICL patients showed no significant differences in the number of bone marrow precursor T-cells (p=0.069) but had lower levels of mature T-cells (p<0.0001) in marrow core biopsies. These data correlated with presence of peripheral blood T-lymphopenias. ICL patients had higher levels of precursor B-cells that were double positive for TdT/CD79a (p=0.016) compared to control marrows, but showed no significant differences in the number of mature B-cells (p=0.059). After treatment with rhIL-7, bone marrow precursor T-cells increased significantly between weeks 1-24 (p=0.016). Mature T-cells also increased significantly (p=0.031). Precursor B-cells and mature B-cells did not change significantly (p=0.313 and 0.375, respectively). During the same time, peripheral blood CD3+ T-cell counts and CD4+ T-cell counts also increased.

Conclusion

Novel histological technique for double chromogenic staining allows for the visualization of T-cell precursors in bone marrow biopsies. Our study revealed no evidence that lymphopenia in ICL patients is associated with lack of T-cell precursors in bone marrow, suggesting downstream defects in T-cell differentiation, proliferation or survival. Administration of rhIL-7 was associated with an increase in peripheral blood T-cells and increase in bone marrow precursor T-cells, while B-cell lineage cells remained unchanged.