Bioprospecting is a systematic search for beneficial compounds derived from biological resources like animals, plants, or microorganisms that can be used for societal benefit and commercialization. Bioprospecting of fungi for anticancer compounds has emerged as a promising avenue in the search for novel and efficient cancer therapies. Bioprospecting isolates fungi from various ecological niches (soil, marine, and plant) and focuses on understanding the biosynthetic pathways involved in producing these metabolites. Advanced molecular biology and genomics techniques have enhanced the ability to discover and identify fungal species with potent bioactive compounds. High-throughput screening is employed to evaluate the cytotoxicity of fungal extracts against cancer cell lines. Several compounds, like polyketides, alkaloids, peptides, and terpenoids, are isolated from fungi and demonstrate anticancer activity. They target cellular mechanisms and pathways like cell cycle arrest, apoptosis, and angiogenesis inhibition. Genetic and metabolic engineering approaches enhance the yield and structural diversity of anticancer compounds produced by fungi. This book chapter highlights the importance of exploration and sustainable utilization of fungal biodiversity in the search for new anticancer agents. Integrating advanced biotechnological tools with traditional natural product research aids in developing novel therapies that can improve cancer treatment outcomes and patient quality of life.