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Potential Anticancer Effects of Limonene Associated with Reactive Oxygen Species Generation, Apoptosis Induction, and
Mahshad Ghanbari1, Roham Deyhimfar2, Fatemeh Khatami2
1Students' Scientific Research Center, Tehran University of Medical Education, Tehran, Iran.
Background:
Tumor recurrence and progression occur in many patients with renal cell carcinoma, highlighting a clinical demand for safe and effective treatment options, particularly by employing plant-derived bioactive compounds. Limonene, an aromatic monoterpene commonly found in citrus peels, has been shown to possess anticancer effects in malignancies such as prostate and bladder cancers.
Objectives:
This study aimed to explore the potential anticancer effects of limonene on ACHN papillary renal cancer cells (pRCC).
Methods:
The viability of ACHN, human embryonic kidney (HEK293), and human dermal fibroblast (HDF) cells treated with limonene was assessed using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. Cellular migration, colony formation, 3D spheroid development, morphological alterations, and caspase 3/7 activity were evaluated in vitro. Reactive oxygen species (ROS) levels, apoptosis, and cell cycle alterations were analyzed by flow cytometry. The mRNA levels of BAX, BCL2, E-cadherin, N-cadherin, Caspase-3, Caspase-9, NF-κB, Akt1, MMP-2, MMP-9, mTOR, TNF-α, COX-2, and IL-6 were quantified using quantitative polymerase chain reaction. NF-κB protein levels and IκB phosphorylation were assessed by Western blot. Nuclear NF-κB levels were measured by ELISA.
Results:
The viability of ACHN cells was reduced by limonene in a dose- and time-dependent manner, with comparatively limited effects on HEK293 and HDF cells. Limonene significantly reduced migration, colony formation, and spheroid growth in ACHN cells. Furthermore, limonene elevated ROS levels and increased apoptosis in ACHN cells without affecting cell cycle progression. After limonene exposure, mRNA expression of Akt1, mTOR, NF-κB, MMP-2, N-cadherin, BCL2, TNF-α, COX-2, and IL-6 decreased, whereas BAX and E-cadherin expression increased. Caspase-3 and Caspase-9 mRNA levels were not significantly altered, whereas caspase 3/7 activity increased. Limonene also reduced NF-κB protein levels, IκB phosphorylation, and NF-κB nuclear translocation.
Conclusions:
These preliminary in vitro findings support further investigation of limonene as a therapeutic candidate in additional pRCC models.
Insights
Limonene, a compound from citrus peels, effectively reduced papillary renal cancer cell viability and migration in vitro. Further research is warranted to explore limonene as a potential plant-derived cancer therapy.
Area of Science:
- Oncology
- Natural Products Chemistry
- Molecular Biology
Background:
- Renal cell carcinoma (RCC) recurrence necessitates novel treatments, with plant-derived compounds showing promise.
- Limonene, a citrus monoterpene, exhibits anticancer properties in various malignancies.
Purpose of the Study:
- To investigate the anticancer potential of limonene against human papillary renal cancer cells (ACHN).
Main Methods:
- In vitro assays assessed cell viability, migration, colony formation, and spheroid growth.
- Flow cytometry analyzed reactive oxygen species (ROS), apoptosis, and cell cycle.
- Quantitative PCR and Western blotting evaluated gene and protein expression related to apoptosis, cell signaling, and metastasis.
Main Results:
- Limonene decreased ACHN cell viability, migration, colony formation, and spheroid growth in a dose- and time-dependent manner.
- Limonene induced ROS production and apoptosis in ACHN cells.
- Limonene modulated mRNA and protein expression, including downregulation of NF-κB signaling and key oncogenes, and upregulation of pro-apoptotic markers.
Conclusions:
- Limonene demonstrates significant in vitro anticancer activity against papillary renal cancer cells.
- These findings support limonene as a potential therapeutic candidate for further investigation in renal cell carcinoma.
