Carbamazepine, a Histone Deacetylase Inhibitor Induces Apoptosis in Human Colon Adenocarcinoma Cell Line HT-29


Background and Aim

Colon cancer ranks fourth and is responsible for causing 10% cancer-related mortality in western countries. Its incidence is rising in many countries due to widespread adoption of the Western diet and lifestyle. Carbamazepine (CBZ) is a FDA-approved antiepileptic drug and a histone deacetylase inhibitor. The aim of this study is to evaluate the cytotoxic potentials of CBZ in human colon cancer cells (HT-29 cells).


HT-29 cells were treated with 36 and 76 μg/ml of CBZ for 24 h. The cytotoxic effect was evaluated by MTT assay. The intracellular reactive oxygen species (ROS) expression was evaluated through dichloro-dihydro-fluorescein diacetate staining. Morphological changes related to apoptosis were evaluated by dual staining with acridine orange and ethidium bromide. Mitochondrial membrane potential was evaluated by rhodamine 123 staining. Immunofluorescence analysis of caspase 3 was done with confocal microscopy.


CBZ caused significant cytotoxicity in HT-29 cells and the effect was concentration dependent. CBZ treatments also caused significant expression of ROS in HT-29 cells. Dual staining showed early and late apoptotic cells and morphological alterations induced by the CBZ. Confocal microscopic studies confirmed the increased caspase 3 expression in CBZ-treated cells.


CBZ induced apoptosis in HT-29 cell through ROS generation and caspase 3 expression and these results pave the way for further in vivo studies.

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  1. 1.

    Gierbolini J, Giarratano M, Benbadis SR. Carbamazepine-related antiepileptic drugs for the treatment of epilepsy - a comparative review. Expert Opin Pharmacother. 2016;17:885–8.

    Article  Google Scholar 

  2. 2.

    Meador KJ, Seliger J, Boyd A, Razavi B, Falco-Walter J, Le S, et al. Comparative neuropsychological effects of carbamazepine and eslicarbazepine acetate. Epilepsy Behav. 2019;94:151–7.

    Article  Google Scholar 

  3. 3.

    Ceccacci E, Minucci S. Inhibition of histone deacetylases in cancer therapy: lessons from leukaemia. Br J Cancer. 2016;114:605–11.

    CAS  Article  Google Scholar 

  4. 4.

    Perri F, Longo F, Giuliano M, Sabbatino F, Favia G, Ionna F, et al. Epigenetic control of gene expression: potential implications for cancer treatment. Crit Rev Oncol Hematol. 2017;111:166–72.

    CAS  Article  Google Scholar 

  5. 5.

    Miller CP, Singh MM, Rivera-Del Valle N, Manton CA, Chandra J. Therapeutic strategies to enhance the anticancer efficacy of histone deacetylase inhibitors. J Biomed Biotechnol 2011;2011:514261, 1, 17.

  6. 6.

    Eckschlager T, Plch J, Stiborova M, Hrabeta J. Histone deacetylase inhibitors as anticancer drugs. Int J Mol Sci. 2017;18:E1414.

    Article  Google Scholar 

  7. 7.

    Beutler AS, Li S, Nicol R, Walsh MJ. Carbamazepine is an inhibitor of histone deacetylases. Life Sci. 2005;76:3107–15.

    CAS  Article  Google Scholar 

  8. 8.

    Salminen JK, Tammela TL, Auvinen A, Murtola TJ. Antiepileptic drugs with histone deacetylase inhibition activity and prostate cancer risk: a population-based case-control study. Cancer Causes Control. 2016;27:637–45.

    Article  Google Scholar 

  9. 9.

    Kuipers EJ, Grady WM, Lieberman D, Seufferlein T, Sung JJ, Boelens PG, et al. Colorectal cancer. Nat Rev Dis Primers. 2015;1:15065.

    Article  Google Scholar 

  10. 10.

    Hatano Y, Fukuda S, Hisamatsu K, Hirata A, Hara A, Tomita H. Multifaceted interpretation of colon cancer stem cells. Int J Mol Sci. 2017;18:E1446.

    Article  Google Scholar 

  11. 11.

    Wei J, Ma L, Lai YH, Zhang R, Li H, Li C, et al. Bazedoxifene as a novel GP130 inhibitor for colon cancer therapy. J Exp Clin Cancer Res. 2019;38:63.

    Article  Google Scholar 

  12. 12.

    Akbarzadeh L, MoiniZanjani T, Sabetkasaei M. Comparison of anticancer effects of carbamazepine and valproic acid. Iran Red Crescent Med J. 2016;18:e37230.

    PubMed  PubMed Central  Google Scholar 

  13. 13.

    PonselviInduja M, Ezhilarasan D, Ashok Vardhan N. Evolvulus alsinoides methanolic extract triggers apoptosis in HepG2 cells. Avicenna J Phytomed. 2018;8:504–12.

    Google Scholar 

  14. 14.

    Gheena S, Ezhilarasan D. Syringic acid triggers reactive oxygen species-mediated cytotoxicity in HepG2 cells. Hum Exp Toxicol. 2019;38:694–702.

    CAS  Article  Google Scholar 

  15. 15.

    Gohel A, McCarthy MB, Gronowicz G. Estrogen prevents glucocorticoid-induced apoptosis in osteoblasts in vivo and in vitro. Endocrinology. 1999;140:5339–47.

    CAS  Article  Google Scholar 

  16. 16.

    Fan J, Wang P, Wang X, Tang W, Liu C, Wang Y, et al. Induction of mitochondrial dependent apoptosis in human leukemia K562 cells by Meconopsis integrifolia: a species from traditional Tibetan medicine. Molecules. 2015;20:11981–93.

    CAS  Article  Google Scholar 

  17. 17.

    Abaza MS, Bahman AM, Al-Attiyah RJ. Valproic acid, an anti-epileptic drug and a histone deacetylase inhibitor, in combination with proteasome inhibitors exerts antiproliferative, pro-apoptotic and chemosensitizing effects in human colorectal cancer cells: underlying molecular mechanisms. Int J Mol Med. 2014;34:513–32.

    CAS  Article  Google Scholar 

  18. 18.

    Cornago M, Garcia-Alberich C, Blasco-Angulo N, Vall-Llaura N, Nager M, Herreros J, et al. Histone deacetylase inhibitors promote glioma cell death by G2 checkpoint abrogation leading to mitotic catastrophe. Cell Death Dis. 2014;5:e1435.

    CAS  Article  Google Scholar 

  19. 19.

    Tseng JH, Chen CY, Chen PC, Hsiao SH, Fan CC, Liang YC, et al. Valproic acid inhibits glioblastoma multiforme cell growth via paraoxonase 2 expression. Oncotarget. 2017;8:14666–79.

    PubMed  PubMed Central  Google Scholar 

  20. 20.

    Ezhilarasan D, Apoorva VS, Ashok Vardhan N. Syzygium cumini extract induced reactive oxygen species-mediated apoptosis in human oral squamous carcinoma cells. J Oral Pathol Med. 2019;48:115–21.

    CAS  PubMed  Google Scholar 

  21. 21.

    Liu K, Liu PC, Liu R, Wu X. Dual AO/EB staining to detect apoptosis in osteosarcoma cells compared with flow cytometry. Med Sci Monit Basic Res. 2015;21:15–20.

    Article  Google Scholar 

  22. 22.

    Grilo AL, Mantalaris A. Apoptosis: a mammalian cell bioprocessing perspective. Biotechnol Adv. 2019;37:459–75.

    CAS  Article  Google Scholar 

  23. 23.

    Zhang BB, Wang DG, Guo FF, Xuan C. Mitochondrial membrane potential and reactive oxygen species in cancer stem cells. Familial Cancer. 2015;14:19–23.

    CAS  Article  Google Scholar 

  24. 24.

    Gillissen B, Richter A, Richter A, Preissner R, Schulze-Osthoff K, Essmann F, et al. Bax/Bak-independent mitochondrial depolarization and reactive oxygen species induction by sorafenib overcome resistance to apoptosis in renal cell carcinoma. J Biol Chem. 2017;292:6478–92.

    CAS  Article  Google Scholar 

  25. 25.

    Arumugam A, Subramani R, Nandy S, Powell S, Velazquez M, Orozco A, et al. Desacetyl nimbinene inhibits breast cancer growth and metastasis through reactive oxygen species mediated mechanisms. Tumour Biol. 2016;37:6527–37.

    CAS  Article  Google Scholar 

  26. 26.

    Yan J, Pang Y, Sheng J, Wang Y, Chen J, Hu J, et al. A novel synthetic compound exerts effective anti-tumour activity in vivo via the inhibition of tubulin polymerisation in A549 cells. Biochem Pharmacol. 2015;97:51–61.

    CAS  Article  Google Scholar 

  27. 27.

    McIlwain DR, Berger T, Mak TW. Caspase functions in cell death and disease. Cold Spring Harb Perspect Biol. 2013;5:a008656.

    Article  Google Scholar 

  28. 28.

    Choudhary GS, Al-Harbi S, Almasan A. Caspase-3 activation is a critical determinant of genotoxic stress-induced apoptosis. Methods Mol Biol. 2015;1219:1–9.

    CAS  Article  Google Scholar 

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We thank Dr. R. Parameshwari, Researcher, Sri Ramachandra Institute of Higher Education and Research, Chennai, India for her scientific contribution to this work.

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MS performed the research and written the manuscript; DE conceived the ideas, performed the research, and analyzed the data and corrected the manuscript.

Corresponding author

Correspondence to Ezhilarasan D.

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Sohaib M, Ezhilarasan D Carbamazepine, a Histone Deacetylase Inhibitor Induces Apoptosis in Human Colon Adenocarcinoma Cell Line HT-29. J Gastrointest Canc 51, 564–570 (2020).

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  • Antiepileptics
  • Apoptosis
  • Colon cancer
  • Caspase
  • Cytotoxicity