Characterization of cerebrospinal fluid pleocytosis and cell subsets during blinatumomab therapy for pediatric B-ALL: A real-life study from China

Abstract Introduction: The central nervous system (CNS) is recognized as a key “sanctuary site” in B-cell acute lymphoblastic leukemia (B-ALL). Although blinatumomab, a bispecific T-cell engager antibody targeting CD19 on B cells and CD3 on T cells, has significantly improved outcomes for patients with B-ALL, the risk of CNS relapses (either isolated or concurrent with other sites) remains a clinical concern. Previous research indicates that blinatumomab can enhance T-lymphocyte migration into the CNS by upregulating adhesion molecules and activating T cells peripherally. It mediates anti-tumor effects by promoting the differentiation of CD8+ T cells into potent effector cells locally. A recent study confirms that over half of adult B-ALL patients treated with blinatumomab develop reactive cerebrospinal fluid (CSF) pleocytosis, predominantly lymphocytosis, which is not linked to increased CNS relapse, neurotoxicity, or worse outcomes. However, this specific pattern has not yet been systematically characterized in pediatric B-ALL patients. Methods: This retrospective analysis involved pediatric B-ALL patients who received blinatumomab at Beijing Children's Hospital, Capital Medical University, between March 2023 and July 2023. Inclusion criteria included: 1) age 1-18 years; 2) no CNS leukemia before blinatumomab therapy; 3) no evidence of active CNS infection or major neurological disorders. All patients received a standard dose of blinatumomab for 14 or 28 days. CSF cytospin and CSF lymphocyte subsets were analyzed on day 14 after receiving blinatumomab. Peripheral blood (PB) lymphocyte subsets were evaluated during the same timeframe. Serum cytokine levels, including interleukin-6 (IL-6), interleukin-10 (IL-10), and interferon-gamma (IFN-γ), were measured before blinatumomab treatment, on day 2, day 7, day 14, and day 28 during blinatumomab treatment. Results: Among 17 evaluable patients (mean age 6.0±3.2 years), 3 were low risk, 8 were intermediate risk, and 6 were high risk. The application time of blinatumomab was during consolidation therapy (n=5) and before maintenance therapy (n=12). Neurological adverse events included seizure (n=1, day 5) and limb tremors (n=1, day 3).CSF cytospin (nucleated cells ≥ 100) was observed in 64.7% of cases, with all instances classified as lymphocyte-predominant. Within this cohort, the mean lymphocyte count reached 96.0±3.8 cells. Flow cytometry revealed that CSF lymphocytes were overwhelmingly CD3+ T-cells (mean 97.3%±2.9%), with minimal CD19+ B-cells (median 0.30%[IQR ± 0.90%]). Compared to PB, CSF exhibited a significantly higher proportion of CD4+ T-cells (49.5% vs. 32.5%, P<0.01) and a significantly lower proportion of CD8+ T-cells (38.8% vs. 48.1%, P=0.022). CD19+ B-cell proportions did not differ significantly (0.30% vs. 0.10%, P=0.24). CD4+ T lymphocytes in CSF were predominantly of a central memory phenotype, constituting 61.6% of the CD4+ subset. Conversely, central memory CD8+ T cells in CSF represented the largest proportion of cytotoxic T lymphocytes at 49.6%. In the subgroup with evaluable serum cytokine data (n=9), median levels of IL-6 (1.73 vs. 18.64 pg/mL), IL-10 (1.73 vs. 23.17 pg/mL), and IFN-γ (1.73 vs. 23.05 pg/mL) measured within 48 hours post-infusion initiation were significantly elevated compared to pre-treatment levels (all P<0.05). Conclusions: The study demonstrated that CSF pleocytosis, predominantly lymphocytosis, is a common phenomenon in pediatric B-ALL patients without prior CNS involvement receiving blinatumomab therapy. This phenomenon of reactive CSF lymphocytosis observed during blinatumomab therapy is more likely due to the migration of peripherally activated T cells across the blood-brain barrier (BBB), rather than infiltration by leukemic cells. These findings provide novel insights into CSF lymphocytosis during blinatumomab therapy in pediatric B-ALL patients, potentially alleviating concerns regarding CNS relapse risk after blinatumomab therapy.