Experimental studies have indicated that disulfiram (Antabuse) has antineoplastic effects against melanoma, breast, and prostate cancer. To explore this hypothesis, we examined the association between disulfiram use and these cancers in a nationwide register-based case–control study nested within ever-users (≥one prescription) of disulfiram. Cases were all Danish individuals with a histologically verified first-time diagnosis of malignant melanoma, breast, or prostate cancer during 2000–2009. For each case, we selected four cancer-free controls matched for age, sex, and year of first disulfiram prescription using risk set sampling. Similarly, for secondary analyses, we selected case–control populations for selected tobacco-related and alcohol-related cancer types, that is, cancers of the buccal cavity, liver, lung, and colorectal cancer. Disulfiram use 1 year before cancer diagnosis and the corresponding date for controls were disregarded. We estimated crude and adjusted odds ratios and 95% confidence intervals (CI) for cancer associated with long-term (≥500 daily defined doses) versus one-time (one prescription) use of disulfiram. Among 53 856 disulfiram users, we identified 166, 644, and 464 cases, respectively, of melanoma, breast, or prostate cancer. Adjusted odds ratios for melanoma, breast, or prostate cancer associated with long-term disulfiram use were 1.04 (95% CI: 0.60–1.78), 0.92 (95% CI: 0.70–1.22), and 0.77 (95% CI: 0.56–1.06), respectively. For prostate cancer, dose–response analyses showed a further risk reduction with the highest cumulative dose level of disulfiram; however, the test for trend did not reach statistical significance. Our study provides some epidemiological support for a protective effect of disulfiram against prostate and breast cancer.