P09.01 Blinatumumab resistance in paediatric relapsed acute lymphoblastic leukaemia is intrinsically driven byleukaemia aggressiveness especially resistance to apoptosis

Background Blinatumomab is a bispecific T-cell engager that links CD3 + T cells with CD19+ B-cell precursor acute lymphoblastic leukaemia (BCP-ALL) blasts. It engages patient T cells in vivo without prior genetic modification or lymphodepletion. In children with relapsed BCP-ALL, blinatumomab achieves response rates of 38–60% (Cheng & Liu, 2025), while exhibiting lower toxicity and greater availability than CAR-T therapy. Several resistance mechanisms have been proposed and investigated in vitro or in small cohorts. These mechanisms may be intrinsic to the leukaemia or driven by extrinsic factors such as T-cell dysfunction or the tumor microenvironment (Zhao et al., 2020). In this study, we investigate potential intrinsic mechanisms of blinatumomab resistance in a paediatric cohort with relapsed of BCP ALL.Methods We performed a retrospective analysis of RNA sequencing data obtained at relapse diagnosis, together with minimal residual disease (MRD) monitoring, from bone marrow samples of 44 paediatric patients diagnosed with relapsed BCP-ALL between 2019 and 2023. All patients received blinatumomab either as part of consolidation therapy or as a bridge to hematopoietic stem cell transplantation (HSCT) and had a detectable MRD prior to blinatumomab. Response to blinatumomab was defined by MRD clearance (< 10e-4 or negative) at the end of the treatment cycle. Patients were stratified into good responders (GR, n = 27) and non-good responders (NGR, n = 17). RNA sequencing and MRD monitoring were performed as part of routine diagnostics. Data were analysed integratively using appropriate bioinformatic and statistical methods.Results Differential gene expression, gene set enrichment, and gene ontology analyses identified a transcriptional profile enriched for genes involved in cell cycle regulation and DNA repair. Adjustment for blast percentage derived from flow cytometry demonstrated that these signatures were associated with leukaemic blasts. In addition, a mitochondria complex I NDUFS6–CD147 interaction was found to be differentially elevated in NGR patients, consistent with enhanced apoptosis resistance and cell survival. Genes belonging to the inhibitor of apoptosis protein (IAP) family, as well as SERPINB9 and its associated genes, were upregulated in NGR compared to GR. Analysis of CD19 alternative splicing at relapse diagnosis, including intron 2 retention and intraexonic splicing of exon 2 (CD19 ex2part), revealed no significant differences between the two response groups.Conclusion Leukemia-intrinsic biological features, particularly increased proliferative capacity and apoptosis resistance at relapse diagnosis, are associated with poor response to blinatumomab. In contrast, target-specific adaptations such as CD19 downregulation and alternative splicing appear to emerge during therapy, reflecting treatment-driven tumour evolution as seen in other studies (Zhao et al., 2020). These findings highlight potential diagnostic markers for early prediction of blinatumomab response and support the rationale for adjuvant therapeutic strategies targeting leukaemia biology to overcome resistance.