Thymoquinone Suppresses Cellular Proliferation, Inhibits VEGF Production and Obstructs Tumor Progression and Invasion in the Rat Model of DMH-Induced Colon Carcinogenesis


A myriad of medicinal effects has been attributed to Thymoquinone (TQ), the major biological-active component of Nigella sativa. TQ has been shown to exhibit potent anti-tumor activities. The present work was undertaken to further explore TQ’s chemopreventive efficacy against 1, 2-dimethylhydrazine (DMH)-induced colon carcinogenesis in the rat model through a two-phase study (initiation and post-initiation) and to evaluate its potential impact on tumor progression and invasion in vivo. TQ treatment in the initiation phase significantly reduced tumor incidence, multiplicity and mean tumor volume. However, although mean tumor volume and multiplicity were decreased upon TQ treatment in the post-initiation phase, TQ did not reduce incidence significantly. Cellular proliferation, as assessed by expression of colonic PCNA, was shown to be inhibited in consequence to TQ treatment in both phases, with a more pronounced reduction in the initiation phase. In addition, our results demonstrated an appreciable negative impact of TQ on vascular endothelial growth factor (VEGF) production in tumor-bearing rats. Furthermore, we provided evidence that TQ-treatment, in both phases, tended to considerably suppress tumor progression and invasion. Taken together, the present study demonstrated that TQ, at an orally daily dose of 10 mg/kg, has a chemopreventive effect in the initiation phase, and has the potential to attenuate tumor burden, suppress progression of pre-neoplastic lesions and to inhibit tumor growth in the post-initiation phase of DMH-induced colon carcinogenesis, We surmise that such effects of TQ may be due to suppression of cellular proliferation and inhibition of VEGF production. The results could provide an effective chemopreventive approach in the primary prevention of colon cancer in humans in the next future, and illuminate a promising horizon to combat progression of benign colonic pre-neoplastic lesions into malignant metastatic tumors and to manage colon cancer.

Share and Cite:

W. Asfour, S. Almadi and L. Haffar, "Thymoquinone Suppresses Cellular Proliferation, Inhibits VEGF Production and Obstructs Tumor Progression and Invasion in the Rat Model of DMH-Induced Colon Carcinogenesis," Pharmacology & Pharmacy, Vol. 4 No. 1, 2013, pp. 7-17. doi: 10.4236/pp.2013.41002.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] J. Ferlay, F. Bray, P. Pisani and D. Parkin, “GLOBOCAN 2002: Cancer Incidence, Mortality and Prevalence Worldwide,” IARC Cancer Base, Vol. 5, IARC Press, Lyon, 2004.
[2] J. Ferlay, H. R. Shin, F. Bray, D. Forman, C. Mathers and D. M. Parkin, “GLOBOCAN 2008 v1.2, Cancer Incidence and Mortality Worldwide,” IARC Cancer Base, Vol. 2, International Agency for Research on Cancer Press, Lyon, 2010.
[3] Z. Msallati, A. El Jord and F. Al-Jerf, “Cancer Incidence in Syria, National,” Cancer Registry, Cancer Prevention Directorate, Ministry of Health, Damascus, 2009.
[4] J. R. Jass, “Pathogenesis of Colorectal Cancer,” Surgical Clinics of North America, Vol. 82, No. 5, 2002, pp. 891-904. doi:10.1016/S0039-6109(02)00047-6
[5] V. Kumar, A. K. Abbas and N. Fausto, “Robbins and Cotran Pathologic Basis of Diseases,” 7th Edition, Saunders, Philadelphia, 2004.
[6] I. Weinstein, “The Origin of Human Cancer: Molecular Mechanisms and Their Implications for Cancer Prevention and Treatment,” Cancer Research, Vol. 48, No. 15, 1988, pp. 4135-4143.
[7] W. C. Willett, “Balancing Life-Style and Genomics Research for Disease Prevention,” Science, Vol. 296, No. 5568, 2002, pp. 695-698. doi:10.1126/science.1071055
[8] Y. S. Kim and J. A. Milner, “Dietary Modulation of Colon Cancer Risk,” Journal of Nutrition, Vol. 137, No. 11, 2007, pp. 2576-2579.
[9] A. E. Tammariello and J. A. Milner, “Mouse Models for Unraveling the Importance of Diet in Colon Cancer Prevention,” Journal of Nutritional Biochemistry, Vol. 21, No. 2, 2010, pp. 77-88. doi:10.1016/j.jnutbio.2009.09.014
[10] D. E. Corpet and F. Pierre, “How Good Are Rodent Models of Carcinogenesis in Predicting Efficacy in Humans? A Systematic Review and Meta-Analysis of Colon Chemoprevention in Rats, Mice and Men,” European Journal of Cancer, Vol. 41, No. 13, 2005, pp. 1911-1922. doi:10.1016/j.ejca.2005.06.006
[11] M. J. Wargovich, A. Jimenez, K. McKee, et al., “Efficacy of Potential Chemopreventive Agents on Rat Colon Aberrant Crypt Formation and Progression,” Carcinogenesis Vol. 21, No. 6, 2000, pp. 1149-1155. doi:10.1093/carcin/21.6.1149
[12] M. Per?e and A. Cerar, “Morphological and Molecular Alterations in 1, 2 Dimethylhydrazine and Azoxymethane Induced Colon Carcinogenesis in Rats,” Journal of Biomedicine and Biotechnology, 2011, Article ID: 473964, 14 p.
[13] M. Lipkin, B. Reddy, H. Newmark and S. A. Lamprecht, “Dietary Factors in Human Colorectal Cancer,” Annual Review of Nutrition, Vol. 19, 1999, pp. 545-586. doi:10.1146/annurev.nutr.19.1.545
[14] Y. J. Surh, “Cancer Chemoprevention with Dietary Phytochemicals,” Nature Reviews Cancer, Vol. 3, No. 10, 2003, pp. 768-780. doi:10.1038/nrc1189
[15] H. Jrah Harzallah, B. Kouidhi, et al., “Chemical Composition, Antimicrobial Potential against Cariogenic Bacteria and Cytotoxic Activity of Tunisian Nigella Sativa Essential Oil and Thymoquinone,” Food Chemistry, Vol. 129, No. 4, 2011, pp. 1469-1474. doi:10.1016/j.foodchem.2011.05.117
[16] H. Gali-Muhtasib, M. Ocker, D. Kuester, et al., “Thymoquinone Reduces Mouse Colon Tumor Cell Invasion and Inhibits Tumor Growth in Murine Colon Cancer Models,” Journal of Cellular and Molecular Medicine, Vol. 12, No. 1, 2008, pp. 330-342. doi:10.1111/j.1582-4934.2007.00095.x
[17] O. A. Badary and A. M. Gamal El-Din, “Inhibitory Effects of Thymoquinone against 20-Methylcholanthrene-Induced Fibrosarcoma Tumorigenesis,” Cancer Detection and Prevention, Vol. 25, No. 4, 2001, pp. 362-368.
[18] O. A. Badary, O. A. Al-Shabanah, M. N. Nagi, et al., “Inhibition of Benzo (a) Pyrene-Induced Forestomach Carcinogenesis in Mice by Thymoquinone,” European Journal of Cancer Prevention, Vol. 8, No. 5, 1999, pp. 435-440. doi:10.1097/00008469-199910000-00009
[19] O. A. Badary, “Thymoquinone Attenuates Ifosfamide-Induced Fanconi syndrome in Rats and Enhances Its Antitumor Activity In mice,” Journal of Ethnopharmacology, Vol. 67, No. 2, 1999, pp. 135-142. doi:10.1016/S0378-8741(98)00242-6
[20] O. A. Badary, M. N. Nagi, O. A. Al-Shabanah, et al., “Thymoquinone Ameliorates the Nephrotoxicity Induced by Cisplatin in Rodents and Potentiates Its Antitumor Activity,” Canadian Journal of Physiology and Pharmacology, Vol. 75, No. 12, 1997, pp. 1356-1361. doi:10.1139/y97-169
[21] N. El-Najjar, M. Chatila, H. Moukadem, et al., “Reactive Oxygen Species Mediate Thymoquinone-Induced Apoptosis and Activate ERK and JNK Signaling,” Apoptosis, Vol. 15, No. 2, 2010, pp. 183-195. doi:10.1007/s10495-009-0421-z
[22] H. Gali-Muhtasib, D. Kuester, C. Mawrin, et al., “Thymoquinone Triggers Inactivation of the Stress Response Pathway Sensor CHEK1 and Contributes to Apoptosis in Colorectal Cancer Cells,” Cancer Research, Vol. 68, No. 14, 2008, pp. 5609-5618.
[23] M. A. El-Mahdy, Q. Zhu, Q. E. Wang, et al., “Thymoquinone Induces Apoptosis through Activation of Caspase-8 and Mitochondrial Events in p53-Null Myeloblastic Leukemia HL-60 Cells,” International Journal of Cancer, Vol. 117, No. 3, 2005, pp. 409-417. doi:10.1002/ijc.21205
[24] M. Alhosin, A. Abusnina, M. Achour, et al., “Induction of Apoptosis by Thymoquinone in Lymphoblastic Leukemia Jurkat Cells Is Mediated by a p73-Dependent Pathway Which Targets the Epigenetic Integrator UHRF1,” Biochemical Pharmacology, Vol. 79, No. 9, 2010, pp. 1251-1260.
[25] K. Womack, M. Anderson, M. Tucci, et al., “Evaluation of Bioflavonoids as Potential Chemotherapeutic Agents,” Biomedical Sciences Instrumentation, Vol. 42, 2006, pp. 464-469.
[26] L. R. Richards, P. Jones, J. Hughes, et al., “The Physiological Effect of Conventional Treatment with Epigallocatechin-3-Gallate, Thymoquinone, and Tannic Acid on the LNCaP Cell Line,” Biomedical Sciences Instrumentation, Vol. 42, 2006, pp. 357-362.
[27] N. Chehl, G. Chipitsyna, Q. Gong, et al., “Anti-Inflammatory Effects of the Nigella Sativa Seed Extract, Thzymoquinone, in Pancreatic Cancer Cells,” HPB (Oxford), Vol. 11, 2009, pp. 373-381. doi:10.1111/j.1477-2574.2009.00059.x
[28] L. R. Richards, P. Jones, H. Benghuzzi, et al., “A Comparison of the Morphological Changes Associated with Conventional and Sustained Treatment with Pigallocatechin3gallate, Thymoquinone, and Tannic Acid on LNCaP Cells,” Biomedical Sciences Instrumentation, Vol. 44, 2008, pp. 465-470.
[29] V. B. Astler and F. A. Coller, “The Prognostic Significance of Direct Extension of Carcinoma of the Colon and Rectum,” Annals of Surgery, Vol. 139, No. 6, 1954, p. 846.
[30] A. M. Shoieb, M. Elgayyar, P. S. Dudrick, et al., “In Vitro Inhibition of Growth and Induction of Apoptosis in Cancer Cell Lines by Thymoquinone,” International Journal of Oncology, Vol. 22, No. 1, 2003, pp. 107-113.
[31] H. Gali-Muhtasib, M. Diab-Assaf, C. Boltze, et al., “Thymoquinone Extracted from Black Seed Triggers Apoptotic Cell Death in Human Colorectal Cancer Cells via a p53-Dependent Mechanism,” International Journal of Oncology, Vol. 25, No. 4, 2004, pp. 857-866.
[32] D. R. Worthen, O. A. Ghosheh and P. A. Crooks, “The in Vitro Anti-Tumor Activity of Some Crude and Purified Components of Blackseed, Nigella sativa L.,” Anticancer Research, Vol. 18, No. 3A, 1998, pp. 1527-1532.
[33] H. U. Gali-Muhtasib, W. G. Abou Kheir, L. A. Kheir, et al., “Molecular Pathway for Thymoquinone-Induced Cell-Cycle Arrest and Apoptosis in Neoplastic Keratinocytes,” Anticancer Drugs, Vol. 15, No. 4, 2004, pp. 389-399. doi:10.1097/00001813-200404000-00012
[34] M. Roepke, A. Diestel, K. Bajbouj, et al., “Lack of p53 Augments Thymoquinone-Induced Apoptosis and Caspase Activation in Human Osteosarcoma Cells,” Cancer Biology & Therapy, Vol. 6, No. 2, 2007, pp. 160-169.
[35] A. O. Kaseb, K. Chinnakannu, D. Chen, A. Sivanandam, S. Tejwani, M. Menon, et al., “Androgen Receptor and E2F-1 Targeted Thymoquinone Therapy for Hormone-Refractory Prostate Cancer,” Cancer Research, Vol. 67, No. 16, 2007, pp. 7782-7788. doi:10.1158/0008-5472.CAN-07-1483
[36] T. Yi, S. G. Cho, Z. Yi, X. Pang, M. Rodriguez, Y. Wang, et al., “Thymoquinone Inhibits Tumor Angiogenesis and Tumor Growth through Suppressing AKT and Extracellular Signal-Regulated Kinase Signaling Pathways,” Molecular Cancer Therapeutics, Vol. 7, No. 7, 2008, pp. 1789-1796.
[37] S. Banerjee, A. O. Kaseb, Z. Wang, D. Kong, M. Mohammad, S. Padhye, et al., “Antitumor Activity of Gemcitabine and Oxaliplatin Is Augmented by Thymoquinone in Pancreatic Cancer,” Cancer Research, Vol. 69, No. 13, 2009, pp. 5575-5583.
[38] S. Attoub, O. Sperandio, H. Raza, K. Arafat, et al., “Thymoquinone as an Anticancer Agent: Evidence from Inhibition of Cancer Cells Viability and Invasion in Vitro and Tumor Growth in Vivo,” Fundamental & Clinical Pharmacology, 2012. doi:10.1111/j.1472-8206.2012.01056.x
[39] T. Yi, S. G. Cho, Z. Yi, X. Pang, et al., “Thymoquinone Inhibits Tumor Angiogenesis and Tumor Growth through Suppressing AKT and ERK Signaling Pathways,” Molecular Cancer Therapeutics, Vol. 7, No. 7, 2008, pp. 1789-1796. doi:10.1158/1535-7163.MCT-08-0124
[40] P. A. Hall, D. A. Levinson and A. L. Woods, “Proliferating Cell Nuclear Antigen (PCNA) Immunolocalization in Paraffin Sections: An Index of Cell Proliferation with Evidence of Deregulated Expression in Some Neoplasm,” Journal of Pathology, Vol. 162, No. 4, 1990, pp. 285-294.
[41] Y. K. Wang, X. L. Ji, Y. G. Gu, et al., “P53 and PCNA Expression in Glandular Dilatation of Gastric Mucosa,” China National Journal of New Gastroenterology, Vol. 2, No. 2, 1996, pp. 106-108.
[42] X. Q. Zhang, “Progress of Study on Suppresser, EGFR and PCNA in Colorectal Cancer,” Chinese Journal of New Gastroenterology, Vol. 4, 1996, pp. 327-328.
[43] Y. Zheng, P. M. Kramer, G. Olson, et al., “Prevention by Retinoids of Azoxymethane Induced Tumors and Aberrant Crypt Foci and Their Modulation of Cell Proliferation in the Colon of Rats,” Carcinogenesis, Vol. 18, No. 11, 1997, pp. 2119-2125.
[44] W. J. Adams, J. A. Lawson, S. E. Nicholson, et al., “The Growth of Carcinogen Induced Colon Cancer in Rats Is Inhibited by Cimetidine,” European Journal of Surgical Oncology, Vol. 19, No. 4, 1993, pp. 332-335.
[45] X. D. Jia and C. Han, “Chemoprevention of Tea on Colorectal Cancer Induced by Dimethylhydrazine in Wistar Rats,” World Journal of Gastroenterology, Vol. 6, No. 5, 2000, pp. 699-703.
[46] B. Shpitz, Y. Bomstein, Y. Mekori, et al., “Proliferating Nuclear Antigen as a Marker of Cell Kinetics in Aberrant Crypt Foci Hyperplastic Polyps, Adenomas, and Adenocarcinomas of the Human Colon,” American Journal of Surgery, Vol. 174, No. 4, 1997, pp. 425-430. doi:10.1016/S0002-9610(97)00122-0
[47] G. Cohen, R. Mustafi, A. Chumsangsri, et al., “Epidermal Growth Factor Receptor Signaling Is Up-Regulated in Human Colonic Aberrant Crypt Foci,” Cancer Research, Vol. 66, No. 11, 2006, pp. 5656-5664. doi:10.1158/0008-5472.CAN-05-0308
[48] D. Longo, “HARRISON’S Hematology and Oncology,” McGraw Hill, New York, 2010, 325 p.
[49] J. Folkman, “What Is the Evidence That Tumors Are Angiogenesis Dependent?” Journal of the National Cancer Institute, Vol. 82, No. 1, 1990, pp. 4-6. doi:10.1093/jnci/82.1.4
[50] L. A. Liotta, J. Kleinerman and G. M. Saidel, “Quantitative Relationships of Intravascular Tumor Cells, Tumor Vessels, and Pulmonary Metastases Following Tumor Implantation,” Cancer Research, Vol. 34, 1974, pp. 997-1004.
[51] G. Gasparini, R. Longo, M. Toi and N. Ferrara, “Angiogenic Inhibitors: A New Therapeutic Strategy in Oncology,” Nature Clinical Practice Oncology, Vol. 2, 2005, pp. 562-577. doi:10.1038/ncponc0342
[52] M. M. Cooney, W. Van Heeckeren, S. Bhakta, et al., “Drug Insight: Vascular Disrupting Agents and Angiogenesis—Novel Approaches for Drug Delivery,” Nature Clinical Practice Oncology, Vol. 3, 2006, pp. 682-692. doi:10.1038/ncponc0663
[53] G. Bergers and L. E. Benjamin, “Tumorigenesis and the Angiogenic Switch,” Nature Reviews Cancer, Vol. 3, No. 6, 2003, pp. 401-410. doi:10.1038/nrc1093
[54] D. J. Kerr, “Targeting Angiogenesis in Cancer: Clinical Development of Bevacizumab,” Nature Clinical Practice Oncology, Vol. 1, 2004, pp. 39-43. doi:10.1038/ncponc0026
[55] H. N. Fernando and H. I. Hurwitz, “Targeted Therapy of Colorectal Cancer: Clinical Experience with Bevacizumab,” Oncologist, Vol. 9, Suppl. 1, 2004, pp. 11-18. doi:10.1634/theoncologist.9-suppl_1-11
[56] G. Des Guetz, B. Uzzan, P. Nicolas, et al., “Microvessel Density and VEGF Expression Are Prognostic Factors in Colorectal Cancer. Meta-Analysis of the Literature,” British Journal of Cancer, Vol. 94, No. 12, 2006, pp. 1823-1832. doi:10.1038/sj.bjc.6603176
[57] N. Ferrara, “Vascular Endothelial Growth Factor: Basic Science and Clinical Progress,” Endocrine Reviews, Vol. 25, No. 4, 2004, pp. 581-611.
[58] N. Ferrara, “Molecular and Biological Properties of Vascular Endothelial Growth Factor,” Journal of Molecular Medicine, Vol. 77, No. 7, 1999, pp. 527-543. doi:10.1007/s001099900019
[59] Y. Takahashi, Y. Kitadai, C. D. Bucana, et al., “Expression of Vascular Endothelial Growth Factor and Its Receptor, KDR, Correlates with Vascularity, Metastasis, and Proliferation of Human Colon Cancer,” Cancer Research, Vol. 55, No. 18, 1995, pp. 3964-3968.
[60] K. Werther, I. J. Christensen, N. Brunner and H. J. Nielsen, “Soluble Vascular Endothelial Growth Factor Levels in Patients with Primary Colorectal Carcinoma, The Danish RANX05 Colorectal Cancer Study Group,” European Journal of Surgical Oncology, Vol. 26, No. 7, 2000, pp. 657-662. doi:10.1053/ejso.2000.0977
[61] K. Werther, I. J. Christensen and H. J. Nielsen, “The Association between Preoperative Concentration of Soluble Vascular Endothelial Growth Factor, Perioperative Blood Transfusion, and Survival in Patients with Primary Colorectal Cancer,” European Journal of Surgery, Vol. 167, No. 4, 2001, pp. 287-292. doi:10.1080/110241501300091480
[62] K. Werther, I. J. Christensen and H. J. Nielsen, “Prognostic Impact of Matched Preoperative Plasma and Serum VEGF in Patients with Primary Colorectal Carcinoma,” British Journal of Cancer, Vol. 86, No. 3, 2002, pp. 417-423. doi:10.1038/sj.bjc.6600075
[63] W. S. Tsai, C. R. Changchien, C. Y. Yeh, et al., “Preoperative Plasma Vascular Endothelial Growth Factor but Not Nitrite Is a Useful Complementary Tumor Marker in Patients with Colorectal Cancer,” Diseases of the Colon & Rectum, Vol. 49, No. 6, 2006, pp. 883-894.
[64] F. De Vita, M. Orditura, E. Lieto, et al., “Elevated Perioperative Serum Vascular Endothelial Growth Factor Levels in Patients with Colon Carcinoma,” Cancer, Vol. 100, No. 2, 2004, pp. 270-278.
[65] G. Galizia, E. Lieto, F. Ferraraccio, et al., “Determination of Molecular Marker Expression Can Predict Outcome in Colon Carcinomas,” Clinical Cancer Research, Vol. 10, 2004, pp. 3490-3499. doi:10.1158/1078-0432.CCR-0960-03
[66] K. F. Chin, J. Greenman, E. Gardiner, et al., “Pre-Operative Serum Vascular Endothelial Growth Factor Can Select Patients for Adjuvant Treatment after Curative Resection in Colorectal Cancer,” British Journal of Cancer, Vol. 83, No. 11, 2000, pp. 1425-1431.
[67] O. Kemik, S. Kemik Ahu, A. Sümer, et al., “Preoperative Vascular Endothelial Growth Factor Levels as a Prognostic Marker for Stage II or III Colorectal Cancer Patients,” Cancer Growth and Metastasis, Vol. 4, 2011, pp. 25-32. doi:10.4137/CGM.S7113
[68] G. L. Semenza, “Targeting HIF-1 for Cancer Therapy,” Nature Reviews Cancer, Vol. 3, 2003, pp. 721-732. doi:10.1038/nrc1187
[69] J. D. Gordan and M. C. Simon, “Hypoxia-Inducible Factors: Central Regulators of the Tumor Phenotype,” Current Opinion in Genetics & Development, Vol. 17, No. 1, 2007, pp. 71-77. doi:10.1016/j.gde.2006.12.006
[70] M. Erdurmus, R. Yagci, B. Yilmaz, et al., “Inhibitory Effects of Topical Thymoquinone on Corneal Neovascularization,” Cornea, Vol. 26, No. 6, 2007, pp. 715-719. doi:10.1097/ICO.0b013e31804f5a45

Copyright © 2023 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.