Citral induced apoptosis in MDA-MB-231 spheroid cells

Abstract

BackgroundBreast cancer remains a leading cause of death in women worldwide. Although breast cancer therapies have greatly advanced in recent years, many patients still develop tumour recurrence and metastasis, and eventually succumb to the disease due to chemoresistance. Citral has been reported to show cytotoxic effect on various cancer cell lines. However, the potential of citral to specifically target the drug resistant breast cancer cells has not yet been tested, which was the focus of our current study.MethodsThe cytotoxic activity of citral was first tested on MDA-MB-231 cells in vitro by MTT assay. Subsequently, spheroids of MDA-MB-231 breast cancer cells were developed and treated with citral at different concentrations. Doxorubicin, cisplatin and tamoxifen were used as positive controls to evaluate the drug resistance phenotype of MDA-MB-231 spheroids. In addition, apoptosis study was performed using AnnexinV/7AAD flowcytometry. Aldefluor assay was also carried out to examine whether citral could inhibit the ALDH-positive population, while the potential mechanism of the effect of citral was carried out by using quantitative real time- PCR followed by western blotting analysis.ResultsCitral was able to inhibit the growth of the MDA-MB-231 spheroids when compared to a monolayer culture of MDA-MB-231 cells at a lower IC50 value. To confirm the inhibition of spheroid self-renewal capacity, the primary spheroids were then cultured to additional passages in the absence of citral. A significant reduction in the number of secondary spheroids were formed, suggesting the reduction of self-renewal capacity of these aldehyde dehydrogenase positive (ALDH+) drug resistant spheroids. Moreover, the AnnexinV/7AAD results demonstrated that citral induced both early and late apoptotic changes in a dose-dependent manner compared to the vehicle control. Furthermore, citral treated spheroids showed lower cell renewal capacity compared to the vehicle control spheroids in the mammosphere formation assay. Gene expression studies using quantitative real time PCR and Western blotting assays showed that citral was able to suppress the self-renewal capacity of spheroids and downregulate the Wnt/β-catenin pathway.ConclusionThe results suggest that citral could be a potential new agent which can eliminate drug-resistant breast cancer cells in a spheroid model via inducing apoptosis.

The Essential Oil of Cymbopogon citratus Stapt and Carvacrol: An Approach of the Antitumor Effect on 7,12-Dimethylbenz-[α]-anthracene (DMBA)-Induced Breast Cancer in Female Rats

“…Regarding its antineoplastic potential, the EOCc showed an anti-cancer effect on cervical cancer cell lines and breast cancer cell line, MCF-7 [ 21 , 22 , 23 ] Likewise, citral induced apoptosis in several hematopoietic cancer cell lines, accompanied by DNA fragmentation and induction of the catalytic activity of caspase-3 [ 24 ], and in ovarian cancer cell lines [ 25 , 26 ]. Additionally, carvacrol, a monoterpenes-phenol is found in the essential oil of many aromatic plants of the Lamiaceae family, including thyme and oregano, showed its anti-cancer potential on the human metastatic breast cancer cell line [ 27 ].…”

Citral Induced Apoptosis through Modulation of Key Genes Involved in Fatty Acid Biosynthesis in Human Prostate Cancer Cells: In Silico and In Vitro Study

“…e therapeutic use of this oil was due to the presence of monoterpenes and myrcene. Few studies demonstrated the anticancer properties of citral in various cancer cell lines including breast (MDI-MB-231), small-cell lung cancer, colorectal cancer (HCT116 and HT29), and cervical cancer (HeLa and ME-180) [12,[29][30][31]. Although various studies in a preliminary level of citral have been studied against various cancer cell lines, the effect of cis-and trans-citral on aggressive prostate cancer (PC3) has not been elucidated so far.…”

TMPE Derived from Saffron Natural Monoterpene as Cytotoxic and Multidrug Resistance Reversing Agent in Colon Cancer Cells

“…Obtained IC 50 values were dependent on structure of monoterpene molecule as well as on cancer cell line used. For linearly-shaped citral, IC 50 values ranged from 10–15 µM in melanoma cell lines [ 25 ] to 60–80 µM in breast [ 26 ] and small cell lung cancer [ 27 ]. On the other hand, cyclic monoterpenes were usually characterized by IC 50 values close to several hundred µM (e.g., β-elemene [ 28 ], safranal [ 29 ], carvacrol, and thymol [ 30 ]); however, one study reported IC 50 values below 1 µM [ 31 ].…”