Recent research has increasingly focused on phytochemicals as promising anticancer agents, with glucosinolates (GSLs) and their hydrolytic derivatives playing a central role. These sulfur-containing compounds, found in plants of the Brassicales order, are converted by myrosinase enzymes into biologically active products, primarily isothiocyanates (ITCs) and indoles, which exhibit significant anticancer properties. Indole-3-carbinol, diindolylmethane, sulforaphane (SFN), phenethyl isothiocyanate (PEITC), benzyl isothiocyanate, and allyl isothiocyanate have shown potent anticancer effects in animal models, particularly in breast, prostate, lung, melanoma, bladder, hepatoma, and gastrointestinal cancers. Clinical studies further support the chemopreventive effects of SFN and PEITC, particularly in detoxifying carcinogens and altering biochemical markers in cancer patients. These compounds have demonstrated good bioavailability, low toxicity, and minimal adverse effects, supporting their potential therapeutic application. Their anticancer mechanisms include the modulation of reactive oxygen species, suppression of cancer-related signaling pathways, and direct interaction with tumor cell proteins. Additionally, semi-synthetic derivatives of GSLs have been developed to enhance anticancer efficacy. In conclusion, GSLs and their derivatives offer significant potential as both chemopreventive and therapeutic agents, warranting further clinical investigation to optimize their application in cancer treatment.