Integrated Genomic Profiling of Pediatric Acute Lymphoblastic Leukemia: Genomic Landscape, Risk Stratification and Association of RAS Pathway Mutations with Early Treatment Response

Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy and genomic profiling has become increasingly vital for risk stratification. We conducted a retrospective single-center study including 96 newly diagnosed pediatric patients with ALL and mixed phenotype acute leukemia to characterize the genomic landscape using conventional and molecular cytogenetics, multiplex ligation-dependent probe amplification and targeted next-generation sequencing and to evaluate associations with treatment response and survival. Mutation co-occurrence analysis revealed a NRAS/KRAS/JAK cluster and an ETV6::RUNX1 cluster characterized by relative mutational exclusivity, with additional associations between NOTCH1–CDK4 and CDK4–CCND3 alterations. DelCDKN2A/CDKN2B alterations showed a trend toward higher day-15 minimal residual disease (MRD) levels and inferior 2-year event free survival rate (EFS). DelIKZF1-positive cases showed lower 2-year EFS. In contrast, ETV6::RUNX1 and high hyperdiploidy were associated with favorable early response and EFS. DelPAX5 did not independently influence outcome. ZNF384 positive cases showed higher early MRD levels despite excellent survival outcomes. NRAS/KRAS mutations were significantly associated with higher positive day-15 MRD (Wilcoxon, p = 0.0067) and remained independently associated after adjustment for white blood cell count and cytogenetic subgroup. Intermediate risk (IR) and high-risk groups showed comparable 2-year EFS, indicating limited discrimination by conventional risk stratification. The IR group displayed a heterogeneous genomic profile, with NRAS/KRAS, Ph-like mutations and delCDKN2A/CDKN2B among the most frequent alterations. These observations highlight the potential of integrated genomic profiling to refine risk stratification, particularly by identifying clinically relevant subgroups within the IR category.