Comprehensive Analysis of the Chemical Composition and In Vitro Cytotoxic Mechanisms of Pallines Spinosa Flower and Leaf Essential Oils Against Breast Cancer Cells

Abstract

Background/aims: In our quest for new natural anticancer agents, we studied the cytotoxicity of the essential oils extracted from flowers and leaves of Pallines spinosa.

Methods: The essential oils were extracted by hydrodistillation and solid phase microextraction (SPME) from flowers and leaves of the plant and their composition was determined by GC/GC-MS. The cytotoxicity of the oils was evaluated against MCF-7 and MDA-MB-231 breast adenocarcinomas, and the non-cancerous MCF-10-2A cells, using a flow cytometry-based assay Apoptosis was evaluated by flow cytometry, nuclear staining, caspases activation, and Western blotting techniques, and cell cycle by measuring DNA contents.

Results: The hydrodistilled flower oil contained mainly sesquiterpenes (96.39%), while the leaf sample was dominated by oxygenated-sesquiterpenes (51.60%) and sesquiterpene-hydrocarbons (34.06%). In contrast, the SPME oil contained mainly monoterpene-hydrocarbons (44.09%) and sesquiterpene-hydrocarbons (34.15%) in the flower and leaf samples, respectively. The cytotoxicity of the flower oil against MCF-7 (IC50 0.25 ± 0.03 µg/mL) and MDA-MB-231 (IC50 0.21 ± 0.03 µg/mL) was much stronger than the leaf oil (IC50 2.4 ± 0.5 µg/mL and 1.5 ± 0.1 µg/mL, respectively). The toxicity of the flower oil was ∼5 to 8-times less in normal MCF-10-2A (IC50 1.3 ± 0.2 µg/mL) and blood mononuclear cells (2.80 ± 0.45 µg/mL) as compared to breast and hematological cancer cells, respectively. Both oils induced a caspase-dependent and -independent apoptosis in MCF-7 and MDA-MB-231 cells, and altered the levels of Bcl-2 and Bax proteins. In addition, the oils arrested cell cycle in both cancer cell lines at G0/G1 phase by modulating the expression of cyclin D1, CDK4 and p21 proteins.

Conclusion: The cytotoxicity of P. spinosa oils were mediated by apoptosis and cell cycle arrest, suggesting the potential use of their bioactive compounds as natural anticancer compounds.

Keywords: Anticancer natural compounds; Apoptosis; Breast cancer cell lines; CDK4; Cell cycle; Cyclin-D; Essential oils; G0/ G1 phase; Pallines spinosa; p21.