The marine environment is a promising source of natural products with possible pharmacological applications. In this sense, marine microorganisms, especially marine fungi, can produce bioactive compounds with various therapeutic properties. Colorectal cancer (CRC) represents a major health problem worldwide, since the treatments used to date are not capable of improving patient survival; that is why natural compounds from marine fungi offer a promising alternative. This study focused on evaluating the antitumor and antioxidant activity of fractions derived from the marine fungi E. maritima and P. lilacinum in two CRC cell lines T84 and SW480. Fractions Fr-EM6, Fr-EM7, Fr-EM8 and Fr-PLMOH-3 demonstrated potent cytotoxic activity in tested CRC cell lines with no activity in the non-tumor line. In particular, the Fr-PLMOH-3 fraction from P. lilacinum showed significant antiproliferative effects on T84 and SW480 cell lines and exhibited a greater cytotoxic effect on cancer stem cells compared to tumor cells. Furthermore, the Fr-EM8 fraction from E. maritima demonstrated a strong antioxidant capacity. These findings highlight the potential of compounds of marine origin as effective and selective antitumor agents for the treatment of CRC. Further studies are needed to explore the underlying mechanisms and potential clinical applications of these bioactive fractions and compounds.