Background: The NAD (P) H: quinone oxidoreductase (NQO1) C609T polymorphism has been widely thought to be associated with the risk of acute leukemia. Objective: This case-control study aimed to assess the impact of NQO1 C609T gene polymorphism in childhood acute lymphoblastic leukemia (ALL). Patients and Method: The study was carried out on one hundred de novo ALL children and one hundred apparently healthy children. Routine genotyping of NQO1C609T gene polymorphism by PCR-RFLP was done for all subjects. Results: No statistically significant difference was observed between the patient group and control group as regards wild and polymorphic genotypes. However, there was a significant difference between ALL patients with wild and polymorphic genotypes regarding their immunophenotyping diagnosis (P=0.02) and FAB classification (P=0.01). There was also a significant difference between ALL patients with wild and polymorphic genotypes regarding their response to treatment. The complete remission in the wild genotype (CC) was 69.2% while in polymorphic genotypes (TT & CT) was 29.4% (P<0.05). Conclusion: The polymorphic genotype forms of the NQO1 C609T (CT and TT) are associated with decreased response to treatment.