Introduction: PEG-Asparaginase (PEG-ASNase) is critical in treating pediatric Acute Lymphoblastic Leukemia (ALL). However, hypersensitivity reactions and inactivations associated with ASNase are major patient challenges. Genetic variability plays a role in these disparities in drug response, presenting Single Nucleotide Variants (SNVs). Pharmacogenetic research aims to identify these variants early in treatment to effectively predict specific drug responses, such as those that may lead to hypersensitivity or pancreatitis in patients who are treated with ASNase, thereby helping to optimize treatment. Objectives: To describe pharmacogenetic variations influencing hypersensitivity and pancreatitis to PEG-ASNase and their correlation with our population. Methods: We conducted a prospective multicenter study involving ALL patients under 18-years old receiving PEG-ASNase. Genotyping was carried out for 8 variants of interest, 6 of which are frequently associated with hypersensitivity in other populations, including GRIA1 (rs4958351), NFACT2 (rs6021191), CNOT3 (rs73062673), GR1A1 (rs6890057), ARGHAP28 (rs9958628), and MYBBP1A (rs3809849); and 3 related to pancreatitis RGS6 (rs17179470), ULK2 (rs281366), and MYBBP1A (rs3809849). The MYBBP1A gene was used to evaluate both hypersensitivity and pancreatitis. The tests were performed using custom TaqMan® genotyping assays. Results: A total of 441 patients were included, with 9.7% presenting clinical allergic reactions, 3.4% presenting pancreatitis, and 10.1% with silent inactivation (ASNase activity < 0.1 IU/mL). Allergic reaction was associated with silent inactivation (p < 0.05). For all genes, the population is in Hardy-Weinberg equilibrium. The variants of each gene showed no association with either allergic reaction or pancreatitis (p > 0.05). Discussion and conclusion: No statistically significant associations were observed between the selected variants and PEG-ASNase-related hypersensitivity or pancreatitis in this cohort. However, findings contribute to characterizing genetic profiles in the Brazilian pediatric population and support future pharmacogenomic approaches. These findings lay the groundwork for future research to uncover new genetic predictors of hypersensitivity, which could contribute to the development of personalized treatment approaches and the prevention of allergic reactions.