Residual nonmalignant T cells in the bone marrow of patients with acute leukemias may be involved in active immune responses to leukemic cells. Here, we investigated the phenotypic signature of T cells present at diagnosis in 39 pediatric patients with acute lymphoblastic leukemia (ALL) treated within standardized ALL-BFM study protocols. Previously described age associations of lymphocyte subpopulations in the peripheral blood of healthy children were reproduced in leukemic bone marrow. Analysis of individual lymphocyte parameters and risk-associated variables using univariate linear regression models revealed a correlation of higher CD4/CD8 ratios at diagnosis with a favorable bone marrow response on day 15. Separate analysis of CD4+ cells with the CD4+CD25^hiFoxP3+ T_reg cell phenotype showed that the association was caused by non-T_reg CD4+ cells. The association of higher CD4/CD8 ratios with a favorable bone marrow response on day 15 of treatment persisted in a cohort extended to 69 patients. We conclude that CD4+ non-T_reg cells in leukemic bone marrow at diagnosis may have a role in early response to treatment. Prospective analysis of the CD4/CD8 ratio in a large cohort of pediatric patients is now needed. Moreover, future experiments will establish the functional role of the individual T cell subsets in immune control in pediatric ALL.

Residual non-malignant T cells in the bone marrow of patients with acute leukemias may be involved in active immune responses to leukemic cells or contribute to immune evasion. Here, we investigated the phenotypic signature of T cells present at diagnosis in 39 pediatric patients with acute lymphoblastic leukemia (ALL) treated at our institution within standardized ALL-BFM study protocols. All patients are alive in first (n=35) or second (n=4) remission with a median follow-up of 79 months. Mononuclear cells were isolated from bone marrow, stained with fluorescein-conjugated monoclonal antibodies and analyzed by 6-color flow cytometry. Median CD4/CD8 ratios in our patient cohort were 1.22 (range 0.42 to 2.63). A median of 4.8% (0.6 to 24.0%) of T cells coexpressed HLA-DR, and a median of 4.4% (range 0.8 to 10.7%) of CD4+ T cells had the Treg cell phenotype CD25hiFoxP3+. A median of 6.5% (range 0.4 to 15.5%) of T cells were TCRγδ+. NK cell/T cell ratios were highly variable (range 2.5 to 40.2). Previously described age associations of lymphocyte subpopulations in peripheral blood of healthy children were also found in leukemic bone marrow. Specifically, among CD4+ T cells, an increase of antigen-experienced T cells (CD45RA-CCR7-) was found in older children (p=0.03). Analysis of individual lymphocyte parameters and patient related variables by univariate linear regression models revealed noticeable differences between patients with T and B lineage ALL: Patients with T-ALL had lower CD4/CD8 ratios (0.05) and higher proportions of CD4+CD25hiFoxP3+ Treg cells (p=0.03) and HLA-DR+ T cells (p=0.008) than patients with B lineage ALL. We further investigated potential associations with leukemia response to treatment. Higher CD4/CD8 ratio at diagnosis correlated with favorable bone marrow response (≤5% blasts) on day 15 (p=0.012). Separate analysis of CD4+ cells with CD4+CD25hiFoxP3+ Treg cell phenotype revealed that the association was caused by non-Treg CD4+ cells. No correlation of the CD4/CD8 ratio with other response parameters or with relapse was found. To further validate this finding, we retrospectively analyzed CD4/CD8 cell ratios in routine diagnostic immunophenotypings of 32 further pediatric patients with ALL, all treated according to ALL-BFM 2000 protocol in our institution. An association of higher CD4/CD8 ratios with favorable bone marrow response on day 15 of treatment persisted in the total cohort of 71 patients (p=0.05). We conclude that CD4+ nonTreg cells in leukemic bone marrow at diagnosis may have a role in early response to treatment. Prospective analysis of the CD4/CD8 ratio in a large cohort of pediatric patients is now needed. Moreover, future experiments will establish the functional role of the individual T cell subsets in immune control or escape in pediatric ALL. Citation Format: Imke Lustfeld, Bianca Altvater, Martina Ahlmann, Sandra Ligges, Peter Brinkrolf, Christel Katerkamp, Annegret Rosemann, Anja Moericke, Claudia Rossig. High proportions of CD4+ T cells among residual bone marrow T cells in childhood acute lymphoblastic leukemia are associated with favorable early responses. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 458. doi:10.1158/1538-7445.AM2013-458