Ketogenic Diet and Black Seed Oil (Nigella sativa) Combination Overcomes Doxorubicin Resistance in Breast Cancer: An In Vitro and In Vivo Study
7 affiliations
- 1Department of Clinical Nutrition and Dietetics, Applied Science Private University, Amman 11931, Jordan
- 2College of Medicine, King Saud bin Abdulaziz University for Health Sciences (KSAU-HS), Jeddah 21589, Saudi Arabia
- 3King Abdullah International Medical Research Centre (KAIMRC), Jeddah 21589, Saudi Arabia
- 4Ministry of National Guard Health Affairs (MNGHA), Jeddah 21589, Saudi Arabia
- 5Department of Clinical Pharmacology, Faculty of Medicine, King Abdulaziz University, Rabigh 21911, Saudi Arabia
- 6Faculty of Allied Medical Sciences, Applied Science Private University, Amman 11931, Jordan
- 7Institute of Basics of Health Sciences, Midwifery and Health Visiting, Faculty of Health Sciences, University of Pécs, 7621 Pécs, Hungary
Abstract
Background: This study evaluated the effects of black seed oil (BSO), a simulated ketogenic metabolic condition consisting of a low-glucose medium supplemented with 5 mM β-hydroxybutyrate (BHB), and their combination in EMT6/P and EMT6/DOX cells. The in vivo effects of a ketogenic diet, BSO, and their combination were also investigated in tumor-bearing mice. Methods: Cytotoxicity of black seed oil (BSO), a simulated ketogenic metabolic condition consisting of low-glucose medium supplemented with 5 mM β-hydroxybutyrate (BHB), their combination, and doxorubicin (DOX) was evaluated in EMT6/P and EMT6/DOX cells using the MTT assay. Combination index (CI) analysis was used to determine the interaction between BSO and the simulated ketogenic metabolic condition. Caspase-3 activity, vascular endothelial growth factor VEGF expression, and P-glycoprotein (P-gp) expression were evaluated in EMT6/DOX cells. In vivo, mice bearing EMT6/P and EMT6/DOX tumors received a ketogenic diet, BSO, their combination, DOX, and control treatment, and tumor growth, tumor weight, tumor-free rate, and serum biochemical parameters were assessed. Results: BSO half-maximal inhibitory concentration IC₅₀ was 14.5-fold higher in EMT6/DOX than in EMT6/P, and DOX IC50 was 39.5-fold higher. The simulated ketogenic metabolic condition consisting of a low-glucose medium supplemented with 5 mM BHB + BSO combination gave a CI of 0.1244 in EMT6/P and 0.0239 in EMT6/DOX, indicating synergism in both lines. The combination produced the highest caspase-3 activity (1.54-fold vs. control), the lowest VEGF level (268.34 pg/mL vs. 953.34 pg/mL in control), and the lowest P-gp level (1.6 ng/mL vs. 3.7 ng/mL in control) among the treatments tested. In vivo, the combination reduced tumor volume by 87.15% in EMT6/P and 58.12% in EMT6/DOX, and gave a tumor-free rate of 66.6% in EMT6/P and 50% in EMT6/DOX. No consistent treatment-related change was observed in liver enzymes, renal markers, or lipid profile compared with healthy controls. Conclusions: The combination of a simulated ketogenic metabolic condition and BSO showed synergistic cytotoxic activity in vitro, while the ketogenic diet and BSO combination produced enhanced antitumor activity in vivo. These effects were associated with increased caspase-3 activity and reduced VEGF and P-gp expression in EMT6/DOX cells, without evidence of hepatic, renal, or metabolic toxicity. These findings support further investigation of this combination in doxorubicin-resistant breast cancer.
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BibTeX
@article{hadi2026ketogenic,
title = {{Ketogenic Diet and Black Seed Oil (Nigella sativa) Combination Overcomes Doxorubicin Resistance in Breast Cancer: An In Vitro and In Vivo Study}},
author = {Rawan W. hadi and Majida Al wraikat and Moudi M. Alasmari and Doaa Anbarserry and Wamidh H. Talib and Bence Raposa},
journal = {Nutrients},
year = {2026},
month = oct,
doi = {10.3390/nu18213610},
url = {https://www.mdpi.com/2072-6643/18/21/3610},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Ketogenic Diet and Black Seed Oil (Nigella sativa) Combination Overcomes Doxorubicin Resistance in Breast Cancer: An In Vitro and In Vivo Study
AU - Rawan W. hadi
AU - Majida Al wraikat
AU - Moudi M. Alasmari
AU - Doaa Anbarserry
AU - Wamidh H. Talib
AU - Bence Raposa
PY - 2026
DA - 2026/10/30
JO - Nutrients
DO - 10.3390/nu18213610
UR - https://www.mdpi.com/2072-6643/18/21/3610
AB - Background: This study evaluated the effects of black seed oil (BSO), a simulated ketogenic metabolic condition consisting of a low-glucose medium supplemented with 5 mM β-hydroxybutyrate (BHB), and their combination in EMT6/P and EMT6/DOX cells. The in vivo effects of a ketogenic diet, BSO, and their combination were also investigated in tumor-bearing mice. Methods: Cytotoxicity of black seed oil (BSO), a simulated ketogenic metabolic condition consisting of low-glucose medium supplemented with 5 mM β-hydroxybutyrate (BHB), their combination, and doxorubicin (DOX) was evaluated in EMT6/P and EMT6/DOX cells using the MTT assay. Combination index (CI) analysis was used to determine the interaction between BSO and the simulated ketogenic metabolic condition. Caspase-3 activity, vascular endothelial growth factor VEGF expression, and P-glycoprotein (P-gp) expression were evaluated in EMT6/DOX cells. In vivo, mice bearing EMT6/P and EMT6/DOX tumors received a ketogenic diet, BSO, their combination, DOX, and control treatment, and tumor growth, tumor weight, tumor-free rate, and serum biochemical parameters were assessed. Results: BSO half-maximal inhibitory concentration IC₅₀ was 14.5-fold higher in EMT6/DOX than in EMT6/P, and DOX IC50 was 39.5-fold higher. The simulated ketogenic metabolic condition consisting of a low-glucose medium supplemented with 5 mM BHB + BSO combination gave a CI of 0.1244 in EMT6/P and 0.0239 in EMT6/DOX, indicating synergism in both lines. The combination produced the highest caspase-3 activity (1.54-fold vs. control), the lowest VEGF level (268.34 pg/mL vs. 953.34 pg/mL in control), and the lowest P-gp level (1.6 ng/mL vs. 3.7 ng/mL in control) among the treatments tested. In vivo, the combination reduced tumor volume by 87.15% in EMT6/P and 58.12% in EMT6/DOX, and gave a tumor-free rate of 66.6% in EMT6/P and 50% in EMT6/DOX. No consistent treatment-related change was observed in liver enzymes, renal markers, or lipid profile compared with healthy controls. Conclusions: The combination of a simulated ketogenic metabolic condition and BSO showed synergistic cytotoxic activity in vitro, while the ketogenic diet and BSO combination produced enhanced antitumor activity in vivo. These effects were associated with increased caspase-3 activity and reduced VEGF and P-gp expression in EMT6/DOX cells, without evidence of hepatic, renal, or metabolic toxicity. These findings support further investigation of this combination in doxorubicin-resistant breast cancer.
SN - 2072-6643
ER -