İlhan Özdemir

Investigation of the effect of thymoquinone and doxorubicin on the EGFR/FOXP3 signaling pathway in OVCAR3 human ovarian adenocarcinoma cells

ABSTRACT Purpose: To investigate the cytotoxic and apoptotic effects of the combination of doxorubicin (Dox) and thymoquinone (TQ) on ovarian adenocarcinoma cells (OVCAR3) via the EGFR/FOXP3 signaling pathway. Methods: We used human OVCAR3 and human skin keratinocyte cells (HaCaT). Different concentrations of TQ and Dox were applied to the cells for 24, 48, and 72 hours, and the cytotoxicity level was determined via the MTT method. Expression levels of EGFR/FOXP3 for cell proliferation and apoptosis were determined by quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blot analysis. The colony counting was performed after DAPI staining, and the effect on cell proliferation was determined. Results: Cytotoxicity was found to be highest with TQ and Dox treatments, and cell migration was prevented, especially in the group that received combined TQ and Dox treatment. Moreover, using RT-qPCR analysis, activity in the EGFR and FOXP3 pathway was found to be downregulated the most with TQ, and the amount of protein decreased with TQ and Dox. Conclusions: The findings showed that the greatest cytotoxic effect and the most apoptosis occurred during TQ treatment. Additionally, it was determined that a significant decrease in EGFR and FOXP3 levels occurred with the application of TQ and Dox.

Apoptotic effect of thymoquinone on OVCAR3 cells via the P53 and CASP3 activation

The limitations in cancer treatment and the inadequacy of classical methods have made it necessary to discover therapeutics in cancer treatment. The cytotoxicity of thymoquinone, which has quite different properties in terms of biological activities, in ovarian cancer cells, and the changes in the expression levels of apoptotic genes (p53/caspase-3 (casp-3)) were investigated. In the study, thymoquinone (5, 50, 100, 250 and 500 µM and 24, 48, 72 hours) were applied to ovarian adenocarcinoma cancer cell line (OVCAR3), at different concentrations. Cytotoxic effect of thymoquinone on OVCAR-3 cells were analyzed by MTT method, and apoptotic and pro-apoptotic gene expression levels (p53, Casp-3) of thymoquinone in cancer cells were analyzed by quantitative real-time polymerase chain reaction. Thymoquinone, whose effect has been revealed in many types of cancer, was shown to significantly reduce the viability of OVCAR3 cancer cells depending on the dose and time (p < 0.05). It was also determined that Casp-3 and p53 gene expressions increased in OVCAR3 cells. Considering the in-vitro cytotoxic activity and apoptotic gene expressions of thymoquinone, an important treatment agent, since it is a promising agent for the future of cancer treatment, more comprehensive studies may pave the way for its clinical use.

Boswellic Acid Enhances Gemcitabine’s Inhibition of Hypoxia-Driven Angiogenesis in Human Endometrial Cancer

Background and Objectives: Endometrial carcinoma is among the most common gynecological malignancies, with recurrence and chemoresistance remaining major clinical challenges. This study aimed to evaluate the combined effects of Boswellic acid (BA), a natural pentacyclic triterpene, and Gemcitabine (GEM), a nucleoside analog chemotherapeutic, on hypoxia, angiogenesis, and apoptosis in human endometrial cancer cells. Materials and Methods: ECC-1 cells were treated with BA, GEM, or their combination under normoxic and hypoxic conditions. Cell viability (MTT assay); nuclear morphology (NucBlue staining); cell cycle distribution (PI flow cytometry); angiogenesis (VEGF ELISA expression); apoptosis (Caspase-3/7 activity; Bax; Bcl-2 expression); inflammatory cytokines (IL-1β; IL-6; TNF-α); and gene ontology enrichment were analyzed. Results: Both BA and GEM reduced cell viability in a dose- and time-dependent manner, with the combination producing synergistic cytotoxicity and lower IC50 values. Hypoxia enhanced drug sensitivity, particularly in combination therapy. BA and GEM significantly suppressed HIF-1α and VEGF expression, with maximal inhibition observed in the combination group. Apoptotic induction was confirmed by increased Bax and Caspase-3 and decreased Bcl-2 expression, together with elevated Caspase-3/7, -8, and -9 activity. Pro-inflammatory cytokine levels were markedly reduced, and gene ontology analysis revealed enrichment of apoptotic, anti-proliferative, and anti-angiogenic pathways. Conclusions: BA + GEM combination synergistically suppresses hypoxia-driven angiogenesis and promotes apoptosis in endometrial cancer cells. These findings support its potential as an adjuvant therapeutic approach, warranting further preclinical and clinical validation.