Background: Occupational exposure to hazardous drugs occurs in all aspects of anticancer drug handling. Proper recommendations and guidelines should be applied to control and reduce exposure. Objective: To assess pharmacists' knowledge and practice regarding the safe handling of anticancer drugs. Methods: A cross-sectional study was conducted at seven major hospitals in Baghdad City, Iraq, from December 2023 to February 2024. A pre-designed questionnaire was given to pharmacists who handled anticancer drugs in chemotherapy units. The questionnaire comprises sociodemographic data, knowledge of the safe handling of cytotoxic drugs and thoughts about exposure and risk, practices for safely handling cytotoxic drugs, and challenges for safely handling cytotoxic drugs. Results: A total of 126 pharmacists were enrolled in the study. Less than half of the participants (46%) received training on anticancer drugs. While more than a third (42.9%) of the pharmacists were handling more than 100 cytotoxic preparations weekly, the total knowledge score was 7.14, while the total practice score was 73.36, with less than half of the pharmacists having good knowledge (44.44%) or practice (48.41%). Pharmacists who received training had significantly higher total practice scores than those who did not. Pharmacists with more than 100 weekly preparations were more knowledgeable than those with 50–99 preparations. Conclusions: The majority of participating pharmacists had fair to excellent knowledge and practice regarding the safe handling of cytotoxic medications. However, there were some gaps regarding important knowledge and practice issues. To improve knowledge and practices regarding anticancer handling, training is necessary.