Berry fruits are an important dietary source of health-promoting antioxidant polyphenols. Interestingly, berry leaves of diverse species, including strawberries, have shown higher bioactive phytochemical content in the leaves than in the fruit. Moreover, the vegetative part of the plants is usually discarded, representing a presumably large source of underutilized bioactive biomass. In this investigation, the polyphenol profiles of tropical highland strawberry (Fragaria x ananassa cv. Festival) leaves and fruits were compared by high-performance liquid chromatography coupled with a diode array detector (UHPLC-DAD) and mass spectrometry (HPLC-MS). The total polyphenol strawberry leaf extracts exhibited a 122-fold-higher total polyphenol content and 13-fold higher antioxidant activity (ORAC) than strawberry fruits, and they showed evidence of possible photoprotective effects against UV damage in human melanoma cells (SK-MEL-28) and in murine embryo fibroblasts (NIH/3T3), together with promising anti-proliferative activities against the same melanoma cells. Seven polyphenols were confirmed by HPLC-DAD in the leaf extracts, with differences depending on fraction solubility. Moreover, three substituted quercetin derivatives, three substituted kaempferol derivatives, two anthocyanins, and catechin were confirmed in the soluble fraction by HPLC-MS. Given their higher total polyphenol content and bioactive activities, underutilized strawberry Festival leaves are a potential source of apparently abundant biomass with prospective bioactive applications.