Antiproliferative Potential of Fruits Extracts on Oral 2D and 3D Culture

Objectives: In this study, we performed an in vitro screening of three waste extracts of Colombian fruits in order to investigate potential antitumor activity against CAL-27 oral cell line in both monolayer and spheroids 3D cultures.
Methods: The cytotoxic and antiproliferative activity of four different fruit extracts (Mangifera indica, Passiflora mollisima, and Physalis peruviana) was evaluated against CAL-27 cell line and primary gingival fibroblasts growing in a monolayer. The antiproliferative effect in the range of 100 to 6,25 μg/mL at 48 and 72 hours was determined using an alamar blue assay. Pro-apoptotic activity and cell cycle distribution were determined by annexin-V/7AAD staining by flow cytometry. The extract with major potential (Passiflora mollisima extract) was evaluated in three concentrations (10, 50,100 μg/mL) on mature spheroids at 72 hours. The extract activity was determined by morphology and size changes using ImageJ software
Results: The primary screen on 2D cultured cells showed an antiproliferative time and dose-dependent and selective effect of Passiflora mollisima extract (IC₅₀ 10μg/mL) on CAL-27 cell line. The extract induced a reduction in cell number, morphological changes, nuclear shrinkage associated with cell death and G0/G1 cell cycle arrest at IC₅₀. The most active extract concentration on spheroids was 50 μg/mL. Passiflora mollisima which reduced (p<0.05) spheroid average area from 198616,5 μm to 164262,3 μm in treated groups, and produced changes in volume and sphericity due to cell detachment.
Conclusions: Extract from Passiflora mollisima showed dose-dependent antiproliferative potential in 2D and 3D cultures related to cell cycle arrest and morphological changes on spheroids. The spheroids model is a promissory tool to select the most active extracts after a cell culture monolayers for screen and preclinical assessment.