Background: Psidium guajava L. (Psidium guajava) is an edible plant; its parts are widely used to cure and prevent many health disorders. Psidium guajava leaves contain a wide array of polyphenols that inhibit peroxidation and may play a role in the prevention and treatment of common, degenerative chronic disorders such as diabetes, cardiovascular disease, and cancer. Colon cancer is the third most common type of cancer diagnosed and the second most common cause of cancer-related deaths globally. In contrast, prostate cancer is the second most diagnosed cancer and the fifth leading cause of cancer-related death worldwide. This study aims to evaluate the pharmacological potential of the Psidium guajava plants cultivated in Romania on colon and prostate cancer cell lines. Methods: Phenolic compounds extraction was made using an average sample of all nine Psidium guajava varieties. Analyses were carried out using a HP-1200 liquid chromatograph. The effect of the alcoholic extract of Psidium guajava leaves was tested on two colon cancer and one prostate cancer cell line as in vitro models. Results: The Psidium guajava leaf extract exhibited anticancer activity against the tested cell lines, with decreased proliferation, increased apoptosis, and cell cycle arrest. The extract reduced the cancer cell line’s migration and invasion capacity, demonstrating greater selectivity for the colon cancer cell line than for the prostate cancer cell lines. Conclusions: This study provides further proof of the Psidium guajava plant’s anticancer properties against colon cancer cell lines. Further studies are needed to confirm its use either alone or in conjunction with conventional cancer treatments as an alternate treatment for certain kinds of malignancies.