This study investigates the clinical and molecular characteristics of TP53-mutated acute myeloid leukemia (AML), associated with a poor prognosis. A retrospective analysis was conducted on 336 AML patients, focusing on the distinctions between TP53-mutated and non-mutated cases. Our findings reveal that TP53 mutations are linked to a higher proportion of leukemic blasts positive for the primitive stem cell markers CD34 and CD41 (with dual positive: 12.5% vs. 1.3%, p < 0.001), suggesting a more stem/progenitor cell-like nature. Patients with TP53-mutated AML exhibited lower white blood cell counts in peripheral blood (p = 0.016) and reduced blast proportion in bone marrow (with a median of 39.6% in TP53-mutated AML and 56.9% in TP53 wild-type AML, p = 0.01) at diagnosis. TP53-mutated AML was frequently found in therapy-related AML cases and occurred with fewer concurrent genetic mutations compared to non-mutated AML. In terms of outcomes, TP53 mutations corresponded with significantly lower composite complete response rates and shorter overall and progression-free survival. This study highlights the distinct clinical, molecular, and immunophenotypic features of TP53-mutated AML and emphasizes the need for improved therapeutic strategies for this high-risk subtype.