Background: The germline TP53 p.R337H variant is a well-established pathogenic founder variant in Brazil, characterized by a elevated population frequency compared with other TP53 pathogenic variants. In the South and Southeast regions of the country, this variant has been estimated to occur in approximately 0.3% of the general population, a frequency that far exceeds germline TP53 mutation rates reported worldwide among Li Fraumeni carriers. Unlike most pathogenic TP53 variants, which affect the DNA-binding domain, the p.R337H substitution involves the oligomerization domain of the p53 protein. Carriers exhibit incomplete and variable penetrance, with an increased risk for a broad spectrum of malignancies consistent with the Li-Fraumeni syndrome tumor spectrum, including childhood adrenocortical carcinoma (ACC), sarcomas, breast cancer, and other solid tumors. Importantly, a substantial proportion of carriers do not fulfill classical clinical criteria for Li-Fraumeni syndrome, indicating that traditional family history–based screening approaches may fail to identify many individuals at increased cancer risk. The high regional frequency and demographic history of this variant reflect a founder effect followed by internal dissemination across Brazilian populations, driven by early population expansion and migratory patterns. Despite significant advances in characterizing p.R337H in high-prevalence Bazilian regions, a critical knowledge gap remains regarding its nationwide distribution and public health implications, particularly in understudied regions where genetic admixture and migration may influence local prevalence. Objectives: To determine the Brazilian prevalence of the pathogenic germline TP53 p.R337H variant using samples from the Brazilian National Neonatal Screening Program. Methods: Anonymized newborn screening samples donated by the Brazilian National Neonatal Screening Program were analyzed. Genomic DNA was extracted from dried blood spots using the MagMAX™ DNA Multi-Sample 2.0 Kit on the KingFisher™ Apex System (Thermo Fisher Scientific). Genotyping was performed by real-time polymerase chain reaction (PCR) using TaqMan® SNP Genotyping Assays, including wild-type and mutant controls. All positive results were confirmed by Sanger sequencing. Results: A total of 12,766 samples were analyzed, comprising 5,000 from the North, 5,000 from the Northeast, and 2,766 from the Central-West regions of Brazil. Three heterozygous carriers of the TP53 p.R337H variant were identified—two from the Central-West region and one from the North region—corresponding to regional prevalences of 0.07% and 0.02%, respectively. In contrast, all samples from the Northeast region were homozygous for the wild-type allele. Conclusions/Implications: The identification of the TP53 p.R337H variant in the North and Central-West regions of Brazil provides direct evidence of its circulation beyond the historically recognized South–Southeast axis. Although the observed prevalence is substantially lower than that reported in Southern and Southeastern Brazil (~0.3%), its detection in a population-based newborn cohort underscores the role of internal migration and population admixture in the dissemination of cancer-predisposing variants. Given the strong association of p.R337H with rare, early-onset, and high-morbidity malignancies, its presence outside traditionally high-risk regions has important clinical and public health implications, supporting the need for expanded genetic surveillance strategies, refined cancer risk assessment, and regionally tailored prevention and early-detection policies aligned with Brazil’s demographic and genetic diversity.