Abstract
Cyclooxygenase-2 (COX-2) is expressed in a variety of human colorectal cancer cells and can contribute to carcinogenesis. This study aimed to investigate the effect of diclofenac (DCF), a selective COX-2 inhibitor, on cell adhesion molecules and apoptosis in human colon adenocarcinoma cells. Levels of homing cell adhesion molecule (H-CAM, CD44), intercellular adhesion molecule-1 (ICAM-1, CD54), vascular cell adhesion molecule-1 (VCAM-1, CD106), and epithelial cell adhesion molecule (EpCAM, CD326) were evaluated in cancer cells overexpressing (HT29) or not expressing (HCT116) COX-2. Cell viability was determined by MTT assay, COX-2 protein levels and activity were assessed by immunofluorescence and fluorometric analysis, respectively. Endogenous levels of polyunsaturated fatty acids (PUFAs) were measured by liquid chromatography-tandem mass spectrometry (LC–MS/MS) while expression of cell adhesion molecules was analyzed by flow cytometry. Annexin V–FITC/propidium iodide-labelling and fluorometric caspase-3 activity measurements were carried out to determine apoptosis. Flow cytometry analysis revealed that the percentage of CD44 and ICAM-1 staining in HCT116 cells was significantly lower compared to HT29 cells. In HT29 cells, phorbol 12-myristate 13-acetate (PMA) induced COX-2 expression and increased CD44 and ICAM-1 levels were down-regulated by diclofenac. Stimulation of COX-2 activity in HT29 cells via PMA significantly decreased diclofenac associated increase in PUFA levels. Treatment with both diclofenac and PMA significantly increased the number of apoptotic cells and caspase-3 activity in colon adenocarcinoma cells compared to control groups. In conclusion, diclofenac’s effect to retard colorectal tumor growth and metastasis occurs in COX-2 overexpressing colon cancer cells by increased apoptosis and decreased expression of CD44 and ICAM-1.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
The authors confirm that the data supporting the findings of this study are available within the article.
Code availability
Not applicable.
Abbreviations
- AA:
-
: Arachidonic acid
- AAD8:
-
: Deuterium-labeled AA
- COX-2:
-
: Cyclooxygenase-2
- COTCF:
-
: Cyclooxgenase-2
- DCF:
-
: Diclofenac
- DGLA:
-
: Dihomogamma-linolenic acid
- DHA:
-
: Docosahexaenoic acid
- DMSO:
-
: Dimethyl sulfoxide
- DSCF:
-
: Dwass-Steel-Critchlow-Fligner
- EPA:
-
: Eicosopentaenoic acid
- EpCAM:
-
: Epithelial cell adhesion molecule
- GSH:
-
: Glutathione
- ICAM-1:
-
: Intercellular adhesion molecule -1
- LC-MS/MS:
-
: Liquid chomatography-tandem mass spectrometry
- MPT:
-
: Mitochondrial permeability transition
- MRM:
-
: Multiple reaction monitoring
- MS/MS:
-
: Tandem mass spectrometry
- OPN:
-
: Osteopontin
- PGG2:
-
: Hydroperoxy endoperoxide
- PMA:
-
: Phorbol 12-myristate 13-acetate
- PUFAs:
-
: Polyunsaturated fatty acids
- UFLC:
-
: Ultrafast liquid chromatography
- VCAM-1:
-
: Vascular cell adhesion molecule-1
References
Agrez MV (1996) Cell adhesion molecules and colon cancer. Aust N Z J Surg 66:791–798. https://doi.org/10.1111/j.1445-2197.1996.tb00752.x
Alexiou D, Karayiannakis AJ, Syrigos KN, Zbar A, Kremmyda A, Bramis I, Tsigris C (2001) Serum levels of E-selectin, ICAM-1 and VCAM-1 in colorectal cancer patients: correlations with clinicopathological features, patient survival and tumour surgery. Eur J Cancer 37:2392–2397. https://doi.org/10.1016/s0959-8049(01)00318-5
Arisan ED, Ergül Z, Bozdağ G, Rencüzoğulları Ö, Çoker-Gürkan A, Obakan-Yerlikaya P, Coşkun D, Palavan-Ünsal N (2018) Diclofenac induced apoptosis via altering PI3K/Akt/MAPK signaling axis in HCT 116 more efficiently compared to SW480 colon cancer cells. Mol Biol Rep 45:2175–2184. https://doi.org/10.1007/s11033-018-4378-2
Aslan M, Kırımlıoğlu E, Afşar E, Çeker T, Yılmaz Ç (2020) Increased PUFA levels in kidney epithelial cells in the course of diclofenac toxicity. Toxicol in Vitro 66:104836. https://doi.org/10.1016/j.tiv.2020.104836
Aslan M, Özcan F, Aslan I, Yücel G (2013) LC-MS/MS analysis of plasma polyunsaturated fatty acids in type 2 diabetic patients after insulin analog initiation therapy. Lipids Health Dis 12:169. https://doi.org/10.1186/1476-511x-12-169
Bendardaf R, Algars A, Elzagheid A, Korkeila E, Ristamäki R, Lamlum H, Collan Y, Syrjänen K, Pyrhönen S (2006) Comparison of CD44 expression in primary tumours and metastases of colorectal cancer. Oncol Rep 16:741–746
Chen WS, Liu JH, Wei SJ, Liu JM, Hong CY, Yang WK (2003) Colon cancer cells with high invasive potential are susceptible to induction of apoptosis by a selective COX-2 inhibitor. Cancer Sci 94:253–258. https://doi.org/10.1111/j.1349-7006.2003.tb01429.x
Chia WK, Ali R, Toh HC (2012) Aspirin as adjuvant therapy for colorectal cancer—reinterpreting paradigms. Nat Rev Clin Oncol 9:561–570. https://doi.org/10.1038/nrclinonc.2012.137
Das UN (2018) Arachidonic acid and other unsaturated fatty acids and some of their metabolites function as endogenous antimicrobial molecules: a review. J Adv Res 11:57–66. https://doi.org/10.1016/j.jare.2018.01.001
Davis JS, Kanikarla-Marie P, Gagea M, Yu PL, Fang D, Sebastian M, Yang P, Hawk E, Dashwood R, Lichtenberger LM, Menter D, Kopetz S (2020) Sulindac plus a phospholipid is effective for polyp reduction and safer than sulindac alone in a mouse model of colorectal cancer development. BMC Cancer 20:871. https://doi.org/10.1186/s12885-020-07311-4
Ding L, Gu H, Lan Z, Lei Q, Wang W, Ruan J, Yu M, Lin J, Cui Q (2019) Downregulation of cyclooxygenase-1 stimulates mitochondrial apoptosis through the NF-κB signaling pathway in colorectal cancer cells. Oncol Rep 41:559–569. https://doi.org/10.3892/or.2018.6785
Dohadwala M, Luo J, Zhu L, Lin Y, Dougherty GJ, Sharma S, Huang M, Pold M, Batra RK, Dubinett SM (2001) Non-small cell lung cancer cyclooxygenase-2-dependent invasion is mediated by CD44. J Biol Chem 276:20809–20812. https://doi.org/10.1074/jbc.C100140200
Gallicchio M, Rosa AC, Dianzani C, Brucato L, Benetti E, Collino M, Fantozzi R (2008) Celecoxib decreases expression of the adhesion molecules ICAM-1 and VCAM-1 in a colon cancer cell line (HT29). Br J Pharmacol 153:870–878. https://doi.org/10.1038/sj.bjp.0707634
Ghanghas P, Jain S, Rana C, Sanyal SN (2016) Chemoprevention of colon cancer through inhibition of angiogenesis and induction of apoptosis by nonsteroidal anti-inflammatory drugs. J Environ Pathol Toxicol Oncol 35:273–289. https://doi.org/10.1615/JEnvironPatholToxicolOncol.2016015704
Haier J, Nasralla M, Nicolson GL (2000) Cell surface molecules and their prognostic values in assessing colorectal carcinomas. Ann Surg 231:11–24. https://doi.org/10.1097/00000658-200001000-00003
Hamada T, Cao Y, Qian ZR, Masugi Y, Nowak JA, Yang J, Song M, Mima K, Kosumi K, Liu L, Shi Y, da Silva A, Gu M, Li W, Keum N, Zhang X, Wu K, Meyerhardt JA, Giovannucci EL, Giannakis M, Rodig SJ, Freeman GJ, Nevo D, Wang M, Chan AT, Fuchs CS, Nishihara R, Ogino S (2017) Aspirin use and colorectal cancer survival according to tumor CD274 (programmed cell death 1 ligand 1) expression status. J Clin Oncol 35:1836–1844. https://doi.org/10.1200/jco.2016.70.7547
Hanahan D, Weinberg RA (2011) Hallmarks of cancer: the next generation. Cell 144:646–674. https://doi.org/10.1016/j.cell.2011.02.013
Hixson LJ, Alberts DS, Krutzsch M, Einsphar J, Brendel K, Gross PH, Paranka NS, Baier M, Emerson S, Pamukcu R et al (1994) Antiproliferative effect of nonsteroidal antiinflammatory drugs against human colon cancer cells. Cancer Epidemiol Biomarkers Prev 3:433–438
Huh JW, Kim HR, Kim YJ, Lee JH, Park YS, Cho SH, Joo JK (2009) Expression of standard CD44 in human colorectal carcinoma: association with prognosis. Pathol Int 59:241–246. https://doi.org/10.1111/j.1440-1827.2009.02357.x
Mt K, Hofland LJ, van Grevenstein WM, van Koetsveld PV, Jeekel J, van Eijck CH (2004) Influence of proinflammatory cytokines on the adhesion of human colon carcinoma cells to lung microvascular endothelium. Int J Cancer 112:943–950. https://doi.org/10.1002/ijc.20506
Kelek SE, Afşar E, Akçay G, Danışman B, Aslan M (2019) Effect of chronic L-carnitine supplementation on carnitine levels, oxidative stress and apoptotic markers in peripheral organs of adult Wistar rats. Food Chem Toxicol 134:110851. https://doi.org/10.1016/j.fct.2019.110851
Kim YM, Pyo H (2013) Different cell cycle modulation by celecoxib at different concentrations. Cancer Biother Radiopharm 28:138–145. https://doi.org/10.1089/cbr.2012.1264
Kohno H, Suzuki R, Sugie S, Tanaka T (2005) Suppression of colitis-related mouse colon carcinogenesis by a COX-2 inhibitor and PPAR ligands. BMC Cancer 5:46. https://doi.org/10.1186/1471-2407-5-46
Lichtenberger LM, Phan T, Fang D, Dial EJ (2018) Chemoprevention with phosphatidylcholine non-steroidal anti-inflammatory drugs in vivo and in vitro. Oncol Lett 15:6688–6694. https://doi.org/10.3892/ol.2018.8098
Liu Q, Chan ST, Mahendran R (2003) Nitric oxide induces cyclooxygenase expression and inhibits cell growth in colon cancer cell lines. Carcinogenesis 24:637–642. https://doi.org/10.1093/carcin/bgg014
Lu X, Yu H, Ma Q, Shen S, Das UN (2010) Linoleic acid suppresses colorectal cancer cell growth by inducing oxidant stress and mitochondrial dysfunction. Lipids Health Dis 9:106. https://doi.org/10.1186/1476-511x-9-106
Menter DG, Schilsky RL, DuBois RN (2010) Cyclooxygenase-2 and cancer treatment: understanding the risk should be worth the reward. Clin Cancer Res 16:1384–1390. https://doi.org/10.1158/1078-0432.Ccr-09-0788
Meyerhardt JA, Shi Q, Fuchs CS, Meyer J, Niedzwiecki D, Zemla T, Kumthekar P, Guthrie KA, Couture F, Kuebler P, Bendell JC, Kumar P, Lewis D, Tan B, Bertagnolli M, Grothey A, Hochster HS, Goldberg RM, Venook A, Blanke C, O’Reilly EM, Shields AF (2021) Effect of celecoxib vs placebo added to standard adjuvant therapy on disease-free survival among patients with stage III colon cancer: the CALGB/SWOG 80702 (Alliance) randomized clinical trial. JAMA 325:1277–1286
Misra S, Obeid LM, Hannun YA, Minamisawa S, Berger FG, Markwald RR, Toole BP, Ghatak S (2008) Hyaluronan constitutively regulates activation of COX-2-mediated cell survival activity in intestinal epithelial and colon carcinoma cells. J Biol Chem 283:14335–14344. https://doi.org/10.1074/jbc.M703811200
Mohammadi A, Yaghoobi MM, Gholamhoseinian Najar A, Kalantari-Khandani B, Sharifi H, Saravani M (2016) HSP90 inhibition suppresses PGE2 production via modulating COX-2 and 15-PGDH expression in HT-29 colorectal cancer cells. Inflammation 39:1116–1123. https://doi.org/10.1007/s10753-016-0343-1
Paschos KA, Canovas D, Bird NC (2009) The role of cell adhesion molecules in the progression of colorectal cancer and the development of liver metastasis. Cell Signal 21:665–674. https://doi.org/10.1016/j.cellsig.2009.01.006
Rawla P, Sunkara T, Barsouk A (2019) Epidemiology of colorectal cancer: incidence, mortality, survival, and risk factors. Prz Gastroenterol 14:89–103. https://doi.org/10.5114/pg.2018.81072
Reina M, Espel E (2017) Role of LFA-1 and ICAM-1 in cancer. Cancers (basel) 9(11):153. https://doi.org/10.3390/cancers9110153
Sanlioglu AD, Dirice E, Aydin C, Erin N, Koksoy S, Sanlioglu S (2005) Surface TRAIL decoy receptor-4 expression is correlated with TRAIL resistance in MCF7 breast cancer cells. BMC Cancer 5:54. https://doi.org/10.1186/1471-2407-5-54
Schack A, Fransgaard T, Klein MF, Gögenur I (2019) Perioperative use of nonsteroidal anti-inflammatory drugs decreases the risk of recurrence of cancer after colorectal resection: a cohort study based on prospective data. Ann Surg Oncol 26:3826–3837. https://doi.org/10.1245/s10434-019-07600-8
Senbanjo LT, Chellaiah MA (2017) CD44: a multifunctional cell surface adhesion receptor is a regulator of progression and metastasis of cancer cells. Front Cell Dev Biol 5:18. https://doi.org/10.3389/fcell.2017.00018
Smakman N, Kranenburg O, Vogten JM, Bloemendaal AL, van Diest P, Borel Rinkes IH (2005) Cyclooxygenase-2 is a target of KRASD12, which facilitates the outgrowth of murine C26 colorectal liver metastases. Clin Cancer Res 11:41–48
Tang W (2003) The metabolism of diclofenac—enzymology and toxicology perspectives. Curr Drug Metab 4:319–329. https://doi.org/10.2174/1389200033489398
Wang D, DuBois RN (2018) Role of prostanoids in gastrointestinal cancer. J Clin Invest 128:2732–2742. https://doi.org/10.1172/JCI97953
Yaghobi Z, Movassaghpour A, Talebi M, Abdoli Shadbad M, Hajiasgharzadeh K, Pourvahdani S, Baradaran B (2021) The role of CD44 in cancer chemoresistance: a concise review. Eur J Pharmacol 903:174147. https://doi.org/10.1016/j.ejphar.2021.174147
Zhang H, Sun XF (2002) Overexpression of cyclooxygenase-2 correlates with advanced stages of colorectal cancer. Am J Gastroenterol 97:1037–1041. https://doi.org/10.1111/j.1572-0241.2002.05625.x
Acknowledgements
The authors thank Dr. Sreeparna Banerjee for providing the cell lines.
Funding
This work was supported by a grant from Akdeniz University Research Foundation (TYL-2020–5435).
Author information
Authors and Affiliations
Contributions
The authors declare that all data were generated in-house and that no paper mill was used.
Tuğçe Çeker: investigation, formal analysis, validation
Çağatay Yilmaz: investigation, formal analysis, validation
Sadi Koksoy: conceptualization, methodology
Mutay Aslan: conceptualization, methodology, writing—reviewing and editing
Corresponding author
Ethics declarations
Ethics approval (include appropriate approvals or waivers)
Not applicable.
Consent to participate (include appropriate statements)
Not applicable.
Consent for publication
The authors give consent for the publication of the article and identifiable details (photographs, graphics, tables, and text) to be published in the Journal of Naunyn–Schmiedeberg’s Archives of Pharmacology.
Conflict of interest
The authors declare no competing interests.
Additional information
Publisher's note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Yilmaz, Ç., Köksoy, S., Çeker, T. et al. Diclofenac down-regulates COX-2 induced expression of CD44 and ICAM-1 in human HT29 colorectal cancer cells. Naunyn-Schmiedeberg's Arch Pharmacol 394, 2259–2272 (2021). https://doi.org/10.1007/s00210-021-02139-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00210-021-02139-6